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Salman M, Shahzad H, Gangaraju R, Ishrat T. Fasudil mitigates diabetes-associated cognitive decline and enhances neuroprotection by suppressing NLRP3/Caspase-1/GSDMD signaling in a stroke mouse model. Exp Neurol 2025; 389:115268. [PMID: 40250699 DOI: 10.1016/j.expneurol.2025.115268] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/06/2025] [Revised: 04/12/2025] [Accepted: 04/15/2025] [Indexed: 04/20/2025]
Abstract
Type 2 diabetes mellitus and obesity are progressive metabolic disorders that heighten the risk of negative outcomes and cognitive decline after an ischemic stroke with limited treatment options. Previous research has shown that Fasudil, a RhoA kinase inhibitor, has therapeutic benefits in various neurological diseases; however, it is unknown if Fasudil provides neuroprotection in diabetic encephalopathy after ischemic stroke. This study aimed to explore the protective effects of Fasudil in an experimental model of diabetic encephalopathy following a photothrombotic stroke using high-fat diet-streptozotocin (HFD/STZ) mice to assess behavioral outcomes and molecular analysis. The experimental mice underwent photothrombotic stroke (pt-MCAO) surgery by retro-orbital injection of Rose Bengal (15 mg/kg), followed by 4 min exposure of the proximal-middle cerebral artery to a 532 nm laser exposure. The results indicated that Fasudil treatment provided potential neuronal protection and improved behavioral outcomes in post-stroke HFD/STZ mice. Additionally, Fasudil inhibited NOD-like receptor protein 3 (NLRP3) inflammasomes and their components, enhanced cognitive function by regulating synaptic markers, and significantly reduced neuroinflammation in post-stroke HFD/STZ mice. Fasudil also notably decreased oxidative stress and apoptosis by modulating Bax and cleaved PARP-1 protein expression and reducing the number of TUNEL-positive cells. In summary, Fasudil treatment offers neuroprotection and enhances cognitive function by preventing oxidative damage and NLRP3 inflammasome activation in post-stroke HFD/STZ mice. These results suggest that Fasudil may serve as a promising alternative therapeutic candidate for improving stroke outcomes and addressing the limitations of current treatment options.
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Affiliation(s)
- Mohd Salman
- Department of Anatomy and Neurobiology, University of Tennessee Health Science Center, Memphis, TN, USA.
| | - Hiba Shahzad
- Department of Anatomy and Neurobiology, University of Tennessee Health Science Center, Memphis, TN, USA
| | - Rajashekhar Gangaraju
- Department of Ophthalmology, Anatomy & Neurobiology, University of Tennessee Health Science Center, Memphis, TN, USA
| | - Tauheed Ishrat
- Department of Anatomy and Neurobiology, University of Tennessee Health Science Center, Memphis, TN, USA; Department of Pharmaceutical Sciences, College of Pharmacy, The University of Tennessee Health Science Center, Memphis, TN, United States; Neuroscience Institute, University of Tennessee Health Science Center, Memphis, TN, USA.
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Ahi EP, Verta JP, Kurko J, Ruokolainen A, Debes PV, Primmer CR. Hippo-vgll3 signaling may contribute to sex differences in Atlantic salmon maturation age via contrasting adipose dynamics. Biol Sex Differ 2025; 16:23. [PMID: 40176157 PMCID: PMC11966934 DOI: 10.1186/s13293-025-00705-8] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/30/2024] [Accepted: 03/20/2025] [Indexed: 04/04/2025] Open
Abstract
BACKGROUND Sexual maturation in Atlantic salmon entails a transition in energy utilization, regulated by genes and environmental stimuli in sex-specific manner. Males require less energy, in the form of adiposity, to mature and typically mature younger than females. Maturation age is also influenced in a sex-dependent fashion by the vgll3 genotype (vestigial-like 3), a co-factor in the Hippo pathway. The underlying molecular processes of sex-dependent maturation age, and their interplay with adiposity and vgll3 genotypes, remain unclear. METHODS To elucidate the mechanisms underlying sex- and genotype-specific maturation differences, we investigated the association of early (E) and late (L) maturation vgll3 alleles with the transcription of > 330 genes involved in the regulation of the Hippo pathway and sexual maturation, and related molecular signals in brain, adipose, and gonads. RESULTS The strongest effect of vgll3 genotype was observed in adipose for females and in brain for males, highlighting sex-specific expression differences in association with vgll3 genotype. Genes related to ovarian development showed increased expression in vgll3*EE compared to vgll3*LL females. Moreover, vgll3*EE females compared to vgll3*EE males exhibited reduced markers of pre-adipocyte differentiation and lipolysis yet enhanced expression of genes related to adipocyte maturation and lipid storage. Brain gene expression further showed sex-specific expression signals for genes related to hormones and lipids, as well as tight junction assembly. CONCLUSIONS Overall, these sex-specific patterns point towards a greater lipid storage and slower energy utilization in females compared to males. These results suggest Hippo-dependent mechanisms may be important mediators of sex differences in maturation age in salmon.
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Affiliation(s)
- Ehsan Pashay Ahi
- Organismal and Evolutionary Biology Research Programme, Faculty of Biological and Environmental Sciences, University of Helsinki, Viikinkaari 1, 00014, Helsinki, Finland.
| | - Jukka-Pekka Verta
- Organismal and Evolutionary Biology Research Programme, Faculty of Biological and Environmental Sciences, University of Helsinki, Viikinkaari 1, 00014, Helsinki, Finland
- Faculty of Biosciences and Aquaculture, Nord University, Bodø, Norway
| | - Johanna Kurko
- Organismal and Evolutionary Biology Research Programme, Faculty of Biological and Environmental Sciences, University of Helsinki, Viikinkaari 1, 00014, Helsinki, Finland
| | - Annukka Ruokolainen
- Organismal and Evolutionary Biology Research Programme, Faculty of Biological and Environmental Sciences, University of Helsinki, Viikinkaari 1, 00014, Helsinki, Finland
| | - Paul Vincent Debes
- Organismal and Evolutionary Biology Research Programme, Faculty of Biological and Environmental Sciences, University of Helsinki, Viikinkaari 1, 00014, Helsinki, Finland
- Department of Aquaculture and Fish Biology, Hólar University, Hólar, Iceland
| | - Craig R Primmer
- Organismal and Evolutionary Biology Research Programme, Faculty of Biological and Environmental Sciences, University of Helsinki, Viikinkaari 1, 00014, Helsinki, Finland
- Institute of Biotechnology, Helsinki Institute of Life Science (HiLIFE), University of Helsinki, Helsinki, Finland
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Mukkala AN, Maksimoska V, Noble E, Ailenberg M, Petrut R, Goldfarb R, Kapus A, Szaszi K, Rotstein OD. SR3677 IS HEPATOPROTECTIVE IN MURINE LIVER ISCHEMIA/REPERFUSION INJURY: POTENTIAL ROLE OF BNIP3L/NIX (BCL2/ADENOVIRUS E1B 19 KDA PROTEIN-INTERACTING PROTEIN 3-LIKE). Shock 2025; 63:499-502. [PMID: 39617401 DOI: 10.1097/shk.0000000000002527] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 03/01/2025]
Abstract
ABSTRACT SR3677, a highly selective rho-associated protein kinase 2 inhibitor, administered prior to liver ischemia/reperfusion injury, induced hepatoprotection in both wild-type and Parkin2-/- mice. Experiments in hepatocytes identified BNIP3L/NIX, as a potential mediator of the hepatoprotective effects of SR3677.
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Affiliation(s)
| | | | - Emma Noble
- Keenan Research Centre for Biomedical Science, St. Michael's Hospital, Unity Health Toronto; Toronto, Ontario, Canada
| | - Menachem Ailenberg
- Keenan Research Centre for Biomedical Science, St. Michael's Hospital, Unity Health Toronto; Toronto, Ontario, Canada
| | - Raluca Petrut
- Keenan Research Centre for Biomedical Science, St. Michael's Hospital, Unity Health Toronto; Toronto, Ontario, Canada
| | - Rachel Goldfarb
- Keenan Research Centre for Biomedical Science, St. Michael's Hospital, Unity Health Toronto; Toronto, Ontario, Canada
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Priante G, Ceol M, Gianesello L, Radu CM, Mantese R, Stefanelli LF, Cacciapuoti M, Martino FK, Calò LA, Anglani F, Nalesso F, Del Prete D. Human parietal epithelial cells as Trojan horses in albumin overload. Sci Rep 2025; 15:1761. [PMID: 39800742 PMCID: PMC11725586 DOI: 10.1038/s41598-024-84972-2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/18/2024] [Accepted: 12/30/2024] [Indexed: 01/16/2025] Open
Abstract
Parietal Epithelial Cells (PECs) activation and proliferation are common to several distinct forms of glomerulopathies. Due to several stimuli, PECs can change to a progenitor (CD24+ and CD133/2+) or a pro-sclerotic (CD44+) phenotype. In addition, PECs, which are constantly exposed to filtered albumin, are known to be involved in albumin internalization, but how this mechanism occurs is unknown. We hypothesized that PECs can transport albumin via receptor-mediated endocytosis and that albumin overload may affect the state of PECs. Conditionally immortalized human PECs (hPECs) were incubated with different albumin concentrations at different times. Albumin internalization studies were performed. Protein expression was assessed using In-Cell Western and immunofluorescence. Cell morphology was analyzed by phase-contrast microscopy and F-actin staining. We demonstrate that hPECs internalize albumin via receptor-mediated mechanisms. Under albumin stimulation, megalin, cubilin, ClC-5, CD133/2, CD24, and CD44 were upregulated. The increase of pERK1/2, the upregulation of ROCK1, ROCK2, caspase -3, -6, and -7, and the morphological changes associated with loss of F-actin fibers indicated that inflammation, proliferation and apoptosis mechanisms had been activated. Our results demonstrate that long-term exposure to high doses of albumin induces up-regulation of molecules involved in the tubular protein uptake machinery and suggest that albumin overload is able to trigger a regenerative process as well as an activation state which might lead in vivo to glomerular crescent formation.
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Affiliation(s)
- Giovanna Priante
- Kidney Histomorphology and Molecular Biology Laboratory, Nephrology Unit, Department of Medicine - DIMED, University of Padua, Via Giustiniani 2, 35128, Padua, Italy.
| | - Monica Ceol
- Kidney Histomorphology and Molecular Biology Laboratory, Nephrology Unit, Department of Medicine - DIMED, University of Padua, Via Giustiniani 2, 35128, Padua, Italy
| | - Lisa Gianesello
- Kidney Histomorphology and Molecular Biology Laboratory, Nephrology Unit, Department of Medicine - DIMED, University of Padua, Via Giustiniani 2, 35128, Padua, Italy
| | - Claudia Maria Radu
- Thrombotic and Haemorrhagic Diseases Unit, Department of Medicine-DIMED, University of Padua, Padua, Italy
| | - Rachele Mantese
- Kidney Histomorphology and Molecular Biology Laboratory, Nephrology Unit, Department of Medicine - DIMED, University of Padua, Via Giustiniani 2, 35128, Padua, Italy
| | - Lucia Federica Stefanelli
- Kidney Histomorphology and Molecular Biology Laboratory, Nephrology Unit, Department of Medicine - DIMED, University of Padua, Via Giustiniani 2, 35128, Padua, Italy
| | - Martina Cacciapuoti
- Kidney Histomorphology and Molecular Biology Laboratory, Nephrology Unit, Department of Medicine - DIMED, University of Padua, Via Giustiniani 2, 35128, Padua, Italy
| | - Francesca K Martino
- Kidney Histomorphology and Molecular Biology Laboratory, Nephrology Unit, Department of Medicine - DIMED, University of Padua, Via Giustiniani 2, 35128, Padua, Italy
| | - Lorenzo Arcangelo Calò
- Kidney Histomorphology and Molecular Biology Laboratory, Nephrology Unit, Department of Medicine - DIMED, University of Padua, Via Giustiniani 2, 35128, Padua, Italy
| | - Franca Anglani
- Kidney Histomorphology and Molecular Biology Laboratory, Nephrology Unit, Department of Medicine - DIMED, University of Padua, Via Giustiniani 2, 35128, Padua, Italy
| | - Federico Nalesso
- Kidney Histomorphology and Molecular Biology Laboratory, Nephrology Unit, Department of Medicine - DIMED, University of Padua, Via Giustiniani 2, 35128, Padua, Italy
| | - Dorella Del Prete
- Kidney Histomorphology and Molecular Biology Laboratory, Nephrology Unit, Department of Medicine - DIMED, University of Padua, Via Giustiniani 2, 35128, Padua, Italy
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Lim KH, Park S, Han E, Baek HW, Hyun K, Hong S, Kim HJ, Lee Y, Rah YC, Choi J. Protective Effects of Fasudil Against Cisplatin-Induced Ototoxicity in Zebrafish: An In Vivo Study. Int J Mol Sci 2024; 25:13363. [PMID: 39769128 PMCID: PMC11678128 DOI: 10.3390/ijms252413363] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/22/2024] [Revised: 12/07/2024] [Accepted: 12/10/2024] [Indexed: 01/11/2025] Open
Abstract
While cisplatin is an effective anti-tumor treatment, it induces ototoxicity through mechanisms involving DNA damage, oxidative stress, and programmed cell death. Rho-associated coiled-coil-containing protein kinase (ROCK) is essential for numerous cellular processes, including apoptosis regulation. Studies have suggested that ROCK inhibitors could prevent apoptosis and promote regeneration. We aimed to investigate the protective effects of the ROCK inhibitor fasudil against cisplatin-induced ototoxicity in a zebrafish model. The zebrafish larvae were exposed to 1 mM cisplatin alone or 1 mM cisplatin co-administered with varying concentrations of fasudil for 4 h. The surviving hair cell counts, apoptosis, reactive oxygen species (ROS) levels, mitochondrial membrane potential (ΔΨm), caspase 3 activity, and autophagy activation were assessed. Rheotaxis behavior was also examined. Cisplatin reduced hair cell counts; increased apoptosis, ROS production, and ΔΨm loss; and activated caspase 3 and autophagy. Fasudil (100 and 500 µM) mitigated cisplatin-induced hair cell loss, reduced apoptosis, and inhibited caspase 3 and autophagy activation. Rheotaxis in zebrafish was preserved by the co-administration of fasudil with cisplatin. Cisplatin induces hair cell apoptosis in zebrafish, whereas fasudil is a promising protective agent against cisplatin-induced ototoxicity.
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Affiliation(s)
- Kang Hyeon Lim
- Department of Otorhinolaryngology-Head and Neck Surgery, Korea University College of Medicine, Ansan Hospital, Ansan 15355, Republic of Korea; (K.H.L.); (S.P.); (E.H.); (H.w.B.); (K.H.); (S.H.); (H.-J.K.); (Y.L.); (Y.C.R.)
| | - Saemi Park
- Department of Otorhinolaryngology-Head and Neck Surgery, Korea University College of Medicine, Ansan Hospital, Ansan 15355, Republic of Korea; (K.H.L.); (S.P.); (E.H.); (H.w.B.); (K.H.); (S.H.); (H.-J.K.); (Y.L.); (Y.C.R.)
| | - Eunjung Han
- Department of Otorhinolaryngology-Head and Neck Surgery, Korea University College of Medicine, Ansan Hospital, Ansan 15355, Republic of Korea; (K.H.L.); (S.P.); (E.H.); (H.w.B.); (K.H.); (S.H.); (H.-J.K.); (Y.L.); (Y.C.R.)
| | - Hyun woo Baek
- Department of Otorhinolaryngology-Head and Neck Surgery, Korea University College of Medicine, Ansan Hospital, Ansan 15355, Republic of Korea; (K.H.L.); (S.P.); (E.H.); (H.w.B.); (K.H.); (S.H.); (H.-J.K.); (Y.L.); (Y.C.R.)
| | - Kyungtae Hyun
- Department of Otorhinolaryngology-Head and Neck Surgery, Korea University College of Medicine, Ansan Hospital, Ansan 15355, Republic of Korea; (K.H.L.); (S.P.); (E.H.); (H.w.B.); (K.H.); (S.H.); (H.-J.K.); (Y.L.); (Y.C.R.)
| | - Sumin Hong
- Department of Otorhinolaryngology-Head and Neck Surgery, Korea University College of Medicine, Ansan Hospital, Ansan 15355, Republic of Korea; (K.H.L.); (S.P.); (E.H.); (H.w.B.); (K.H.); (S.H.); (H.-J.K.); (Y.L.); (Y.C.R.)
| | - Hwee-Jin Kim
- Department of Otorhinolaryngology-Head and Neck Surgery, Korea University College of Medicine, Ansan Hospital, Ansan 15355, Republic of Korea; (K.H.L.); (S.P.); (E.H.); (H.w.B.); (K.H.); (S.H.); (H.-J.K.); (Y.L.); (Y.C.R.)
- Zebrafish Translational Medical Research Center, Korea University, Ansan 15355, Republic of Korea
| | - Yunkyoung Lee
- Department of Otorhinolaryngology-Head and Neck Surgery, Korea University College of Medicine, Ansan Hospital, Ansan 15355, Republic of Korea; (K.H.L.); (S.P.); (E.H.); (H.w.B.); (K.H.); (S.H.); (H.-J.K.); (Y.L.); (Y.C.R.)
- Zebrafish Translational Medical Research Center, Korea University, Ansan 15355, Republic of Korea
| | - Yoon Chan Rah
- Department of Otorhinolaryngology-Head and Neck Surgery, Korea University College of Medicine, Ansan Hospital, Ansan 15355, Republic of Korea; (K.H.L.); (S.P.); (E.H.); (H.w.B.); (K.H.); (S.H.); (H.-J.K.); (Y.L.); (Y.C.R.)
| | - June Choi
- Department of Otorhinolaryngology-Head and Neck Surgery, Korea University College of Medicine, Ansan Hospital, Ansan 15355, Republic of Korea; (K.H.L.); (S.P.); (E.H.); (H.w.B.); (K.H.); (S.H.); (H.-J.K.); (Y.L.); (Y.C.R.)
- Zebrafish Translational Medical Research Center, Korea University, Ansan 15355, Republic of Korea
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Huete-Acevedo J, Mas-Bargues C, Arnal-Forné M, Atencia-Rabadán S, Sanz-Ros J, Borrás C. Role of Redox Homeostasis in the Communication Between Brain and Liver Through Extracellular Vesicles. Antioxidants (Basel) 2024; 13:1493. [PMID: 39765821 PMCID: PMC11672896 DOI: 10.3390/antiox13121493] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2024] [Revised: 11/21/2024] [Accepted: 11/29/2024] [Indexed: 01/11/2025] Open
Abstract
Extracellular vesicles (EVs) are small, membrane-bound particles secreted by cells into the extracellular environment, playing an increasingly recognized role in inter-organ communication and the regulation of various physiological processes. Regarding the redox homeostasis context, EVs play a pivotal role in propagating and mitigating oxidative stress signals across different organs. Cells under oxidative stress release EVs containing signaling molecules that can influence the redox status of distant cells and tissues. EVs are starting to be recognized as contributors to brain-liver communication. Therefore, in this review, we show how redox imbalance can affect the release of EVs in the brain and liver. We propose EVs as mediators of redox homeostasis in the brain-liver axis.
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Affiliation(s)
- Javier Huete-Acevedo
- MiniAging Research Group, Department of Physiology, Faculty of Medicine, University of Valencia, CIBERFES, INCLIVA, Avenida Blasco Ibáñez, 15, 46010 Valencia, Spain; (J.H.-A.); (C.M.-B.); (M.A.-F.); (S.A.-R.)
| | - Cristina Mas-Bargues
- MiniAging Research Group, Department of Physiology, Faculty of Medicine, University of Valencia, CIBERFES, INCLIVA, Avenida Blasco Ibáñez, 15, 46010 Valencia, Spain; (J.H.-A.); (C.M.-B.); (M.A.-F.); (S.A.-R.)
| | - Marta Arnal-Forné
- MiniAging Research Group, Department of Physiology, Faculty of Medicine, University of Valencia, CIBERFES, INCLIVA, Avenida Blasco Ibáñez, 15, 46010 Valencia, Spain; (J.H.-A.); (C.M.-B.); (M.A.-F.); (S.A.-R.)
| | - Sandra Atencia-Rabadán
- MiniAging Research Group, Department of Physiology, Faculty of Medicine, University of Valencia, CIBERFES, INCLIVA, Avenida Blasco Ibáñez, 15, 46010 Valencia, Spain; (J.H.-A.); (C.M.-B.); (M.A.-F.); (S.A.-R.)
| | - Jorge Sanz-Ros
- Department of Pathology, Stanford University School of Medicine, Stanford, CA 94305, USA;
| | - Consuelo Borrás
- MiniAging Research Group, Department of Physiology, Faculty of Medicine, University of Valencia, CIBERFES, INCLIVA, Avenida Blasco Ibáñez, 15, 46010 Valencia, Spain; (J.H.-A.); (C.M.-B.); (M.A.-F.); (S.A.-R.)
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Lange M, Francis C, Furtado J, Kim YB, Liao JK, Eichmann A. Endothelial Rho kinase controls blood vessel integrity and angiogenesis. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.11.19.624343. [PMID: 39605538 PMCID: PMC11601598 DOI: 10.1101/2024.11.19.624343] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Indexed: 11/29/2024]
Abstract
Background The Rho kinases 1 and 2 (ROCK1/2) are serine-threonine specific protein kinases that control actin cytoskeleton dynamics. They are expressed in all cells throughout the body, including cardiomyocytes, smooth muscle cells and endothelial cells, and intimately involved in cardiovascular health and disease. Pharmacological ROCK inhibition is beneficial in mouse models of hypertension, atherosclerosis, and neointimal thickening that display overactivated ROCK. However, the consequences of endothelial ROCK signaling deficiency in vivo remain unknown. To address this issue, we analyzed endothelial cell (EC) specific ROCK1 and 2 deletions. Methods We generated Cdh5-CreERT2 driven, tamoxifen inducible loss of function alleles of ROCK1 and ROCK2 and analyzed mouse survival and vascular defects through cellular, biochemical, and molecular biology approaches. Results We observed that postnatal or adult loss of endothelial ROCK1 and 2 was lethal within a week. Mice succumbed to multi-organ hemorrhage that occurred because of loss of vascular integrity. ECs displayed deficient cytoskeletal actin polymerization that prevented focal adhesion formation and disrupted junctional integrity. Retinal sprouting angiogenesis was also perturbed, as sprouting vessels exhibited lack of polymerized actin and defective lumen formation. In a three-dimensional endothelial sprouting assay, combined knockdown of ROCK1/2 or knockdown or ROCK2 but not ROCK1 led to reduced sprouting, lumenization and cell polarization defects caused by defective actin and altered VE-cadherin dynamics. The isoform specific role of endothelial ROCK2 correlated with ROCK2 substrate specificity for FAK and LIMK. By analyzing single and three allele mutants we show that one intact allele of ROCK2 is sufficient to maintain vascular integrity in vivo. Conclusion Endothelial ROCK1 and 2 maintain junctional integrity and ensure proper angiogenesis and lumen formation. The presence of one allele of ROCK2 is sufficient to maintain vascular growth and integrity. These data indicate the need of careful consideration for the use of ROCK inhibitors in disease settings.
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Affiliation(s)
- Martin Lange
- Cardiovascular Research Center, Department of Internal Medicine, Yale University School of Medicine, New Haven, CT, USA
- Department of Molecular and Cellular Physiology, Yale University School of Medicine, New Haven, CT, USA
| | - Caitlin Francis
- Cardiovascular Research Center, Department of Internal Medicine, Yale University School of Medicine, New Haven, CT, USA
- Department of Molecular and Cellular Physiology, Yale University School of Medicine, New Haven, CT, USA
| | - Jessica Furtado
- Cardiovascular Research Center, Department of Internal Medicine, Yale University School of Medicine, New Haven, CT, USA
- Department of Molecular and Cellular Physiology, Yale University School of Medicine, New Haven, CT, USA
| | - Young-Bum Kim
- Division of Endocrinology, Diabetes, and Metabolism, Department of Medicine, Beth Israel Deaconess Medical Center, and Harvard Medical School, Boston, Mass, USA
- Department of Molecular Medicine and Biopharmaceutical Sciences, Graduate School of Convergence Science and Technology, Seoul National University, Seoul, Korea
| | - James K Liao
- Division of Cardiology/Sarver Heart Center, University of Arizona College of Medicine, Tucson, AZ, USA
| | - Anne Eichmann
- Cardiovascular Research Center, Department of Internal Medicine, Yale University School of Medicine, New Haven, CT, USA
- Department of Molecular and Cellular Physiology, Yale University School of Medicine, New Haven, CT, USA
- Université de Paris, INSERM, PARCC, F-75015, Paris, France
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de Freitas AC, Reolon HG, Abduch NG, Baldi F, Silva RMO, Lourenco D, Fragomeni BO, Paz CCP, Stafuzza NB. Proteomic identification of potential biomarkers for heat tolerance in Caracu beef cattle using high and low thermotolerant groups. BMC Genomics 2024; 25:1079. [PMID: 39538142 PMCID: PMC11562314 DOI: 10.1186/s12864-024-11021-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/01/2024] [Accepted: 11/08/2024] [Indexed: 11/16/2024] Open
Abstract
BACKGROUND Heat stress has deleterious effects on physiological and performance traits in livestock. Within this context, using tropically adapted cattle breeds in pure herds or terminal crossbreeding schemes to explore heterosis is attractive for increasing animal production in warmer climate regions. This study aimed to identify biological processes, pathways, and potential biomarkers related to thermotolerance in Caracu, a tropically adapted beef cattle breed, by proteomic analysis of blood plasma. To achieve this goal, 61 bulls had their thermotolerance evaluated through a heat tolerance index. A subset of 14 extreme animals, including the seven most thermotolerant (HIGH group) and the seven least thermotolerant (LOW group), had their blood plasma samples used for proteomic analysis by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). The differentially regulated proteins detected between HIGH and LOW groups were used to perform functional enrichment analysis and a protein-protein interaction network analysis. RESULTS A total of 217 proteins were detected only in the HIGH thermotolerant group and 51 only in the LOW thermotolerant group. In addition, 81 and 87 proteins had significantly higher and lower abundancies in the HIGH group, respectively. Regarding proteins with the highest absolute log-fold change values, we highlighted those encoded by DUSP5, IGFALS, ROCK2, RTN4, IRAG1, and NNT genes based on their functions. The functional enrichment analysis detected several biological processes, molecular functions, and pathways related to cellular responses to stress, immune system, complement system, and hemostasis in both HIGH and LOW groups, in addition to terms and pathways related to lipids and calcium only in the HIGH group. Protein-protein interaction (PPI) network revealed as important nodes many proteins with roles in response to stress, hemostasis, immune system, inflammation, and homeostasis. Additionally, proteins with high absolute log-fold change values and proteins detected as essential nodes by PPI analysis highlighted herein are potential biomarkers for thermotolerance, such as ADRA1A, APOA1, APOB, APOC3, C4BPA, CAT, CFB, CFH, CLU, CXADR, DNAJB1, DNAJC13, DUSP5, FGA, FGB, FGG, HBA, HBB, HP, HSPD1, IGFALS, IRAG1, KNG1, NNT, OSGIN1, PROC, PROS1, ROCK2, RTN4, RYR1, TGFB2, VLDLR, VTN, and VWF. CONCLUSIONS Identifying potential biomarkers, molecular mechanisms and pathways that act in response to heat stress in tropically adapted beef cattle contributes to developing strategies to improve performance and welfare traits in livestock under tropical climates.
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Affiliation(s)
- Ana Claudia de Freitas
- Beef Cattle Research Center, Animal Science Institute, Sertãozinho, SP, 14160-900, Brazil
- Agricultural Research Agency of the State of Minas Gerais (EPAMIG), Patos de Minas, MG, 38709-899, Brazil
| | - Henrique G Reolon
- Beef Cattle Research Center, Animal Science Institute, Sertãozinho, SP, 14160-900, Brazil
- Department of Animal Science, School of Agricultural and Veterinary Sciences, São Paulo State University (UNESP), Jaboticabal, SP, 14884-900, Brazil
| | - Natalya G Abduch
- Beef Cattle Research Center, Animal Science Institute, Sertãozinho, SP, 14160-900, Brazil
| | - Fernando Baldi
- Department of Animal Science, School of Agricultural and Veterinary Sciences, São Paulo State University (UNESP), Jaboticabal, SP, 14884-900, Brazil
| | | | - Daniela Lourenco
- Department of Animal and Dairy Science, University of Georgia, Athens, GA, 30602, USA
| | | | - Claudia C P Paz
- Sustainable Livestock Research Center, Animal Science Institute, São José do Rio Preto, SP, 15130-000, Brazil
| | - Nedenia B Stafuzza
- Beef Cattle Research Center, Animal Science Institute, Sertãozinho, SP, 14160-900, Brazil.
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Attia H, Badr A, Alshehri O, Alsulaiman W, Alshanwani A, Alshehri S, Arafa M, Hasan I, Ali R. The Protective Effects of Vitamin B Complex on Diclofenac Sodium-Induced Nephrotoxicity: The Role of NOX4/RhoA/ROCK. Inflammation 2024; 47:1600-1615. [PMID: 38413451 DOI: 10.1007/s10753-024-01996-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/02/2023] [Revised: 02/04/2024] [Accepted: 02/19/2024] [Indexed: 02/29/2024]
Abstract
Diclofenac sodium (DIC) is a widely used non-steroidal anti-inflammatory drug. Unfortunately, its prolonged use is associated with nephrotoxicity due to oxidative stress, inflammation, and fibrosis. We aimed to investigate the nephroprotective effects of vitamin B complex (B1, B6, B12) against DIC-induced nephrotoxicity and its impact on NOX4/RhoA/ROCK, a pathway that plays a vital role in renal pathophysiology. Thirty-two Wistar rats were divided into four groups: (1) normal control; (2) vitamin B complex (16 mg/kg B1, 16 mg/kg B6, 0.16 mg/kg B12, intraperitoneal); (3) DIC (10 mg/kg, intramuscular); and (4) DIC plus vitamin B complex group. After 14 days, the following were assayed: serum renal biomarkers (creatinine, blood urea nitrogen, kidney injury molecule-1), oxidative stress, inflammatory (tumor necrosis factor-α, interleukin-6), and fibrotic (transforming growth factor-β) markers as well as the protein levels of NOX4, RhoA, and ROCK. Structural changes, inflammatory cell infiltration, and fibrosis were detected using hematoxylin and eosin and Masson trichrome stains. Compared to DIC, vitamin B complex significantly decreased the renal function biomarkers, markers of oxidative stress and inflammation, and fibrotic cytokines. Glomerular and tubular damage, inflammatory infiltration, and excessive collagen accumulation were also reduced. Protein levels of NOX4, RhoA, and ROCK were significantly elevated by DIC, and this elevation was ameliorated by vitamin B complex. In conclusion, vitamin B complex administration could be a renoprotective approach during treatment with DIC via, at least in part, suppressing the NOX4/RhoA/ROCK pathway.
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Affiliation(s)
- Hala Attia
- Department of Pharmacology and Toxicology, College of Pharmacy, King Saud University, P. O. Box: 2454, Riyadh, 11495, Saudi Arabia.
| | - Amira Badr
- Department of Pharmacology and Toxicology, College of Pharmacy, King Saud University, P. O. Box: 2454, Riyadh, 11495, Saudi Arabia
| | - Orjuwan Alshehri
- College of Pharmacy, King Saud University, Riyadh, 11495, Saudi Arabia
| | - Waad Alsulaiman
- College of Pharmacy, King Saud University, Riyadh, 11495, Saudi Arabia
| | - Aliah Alshanwani
- Department of Physiology, College of Medicine, King Saud University, Riyadh, 11495, Saudi Arabia
| | - Samiyah Alshehri
- Department of Pharmacology and Toxicology, College of Pharmacy, King Saud University, P. O. Box: 2454, Riyadh, 11495, Saudi Arabia
| | - Maha Arafa
- Pathology Department, College of Medicine, King Saud University, Riyadh, 11495, Saudi Arabia
| | - Iman Hasan
- Department of Pharmacology and Toxicology, College of Pharmacy, King Saud University, P. O. Box: 2454, Riyadh, 11495, Saudi Arabia
| | - Rehab Ali
- Department of Pharmacology and Toxicology, College of Pharmacy, King Saud University, P. O. Box: 2454, Riyadh, 11495, Saudi Arabia
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10
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Zhang C, Zheng J, Yu X, Kuang B, Dai X, Zheng L, Yu W, Teng W, Cao H, Li M, Yao J, Liu X, Zou W. "Baihui" (DU20)-penetrating "Qubin" (GB7) acupuncture on blood-brain barrier integrity in rat intracerebral hemorrhage models via the RhoA/ROCK II/MLC 2 signaling pathway. Animal Model Exp Med 2024; 7:740-757. [PMID: 38379356 PMCID: PMC11528382 DOI: 10.1002/ame2.12374] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/28/2023] [Accepted: 11/21/2023] [Indexed: 02/22/2024] Open
Abstract
BACKGROUND Blocking the RhoA/ROCK II/MLC 2 (Ras homolog gene family member A/Rho kinase II/myosin light chain 2) signaling pathway can initiate neuroprotective mechanisms against neurological diseases such as stroke, cerebral ischemia, and subarachnoid hemorrhage. Nevertheless, it is not clear whether and how disrupting the RhoA/ROCK II/MLC 2 signaling pathway changes the pathogenic processes of the blood-brain barrier (BBB) after intracerebral hemorrhage (ICH). The present investigation included the injection of rat caudal vein blood into the basal ganglia area to replicate the pathophysiological conditions caused by ICH. METHODS Scalp acupuncture (SA) therapy was performed on rats with ICH at the acupuncture point "Baihui"-penetrating "Qubin," and the ROCK selective inhibitor fasudil was used as a positive control to evaluate the inhibitory effect of acupuncture on the RhoA/ROCK II/MLC 2 signaling pathway. Post-assessments included neurological deficits, brain edema, Evans blue extravasation, Western blot, quantitative polymerase chain reaction, and transmission electron microscope imaging. RESULTS We found that ROCK II acts as a promoter of the RhoA/ROCK II/MLC 2 signaling pathway, and its expression increased at 6 h after ICH, peaked at 3 days, and then decreased at 7 days after ICH, but was still higher than the pre-intervention level. According to some experimental results, although 3 days is the peak, 7 days is the best time point for acupuncture treatment. Starting from 6 h after ICH, the neurovascular structure and endothelial cell morphology around the hematoma began to change. Based on the changes in the promoter ROCK II, a 7-day time point was selected as the breakthrough point for treating ICH model rats in the main experiment. The results of this experiment showed that both SA at "Baihui"-penetrating "Qubin" and treatment with fasudil could improve the expression of endothelial-related proteins by inhibiting the RhoA/ROCK II/MLC 2 signaling pathway and reduce neurological dysfunction, brain edema, and BBB permeability in rats. CONCLUSION This study found that these experimental data indicated that SA at "Baihui"-penetrating "Qubin" could preserve BBB integrity and neurological function recovery after ICH by inhibiting RhoA/ROCK II/MLC 2 signaling pathway activation and by regulating endothelial cell-related proteins.
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Affiliation(s)
- Ce Zhang
- Heilongjiang University of Chinese MedicineHarbinChina
| | - Jia Zheng
- Heilongjiang University of Chinese MedicineHarbinChina
| | - Xueping Yu
- First Affiliated Hospital of Heilongjiang University of Chinese MedicineHarbinChina
| | - Binglin Kuang
- Heilongjiang University of Chinese MedicineHarbinChina
| | - Xiaohong Dai
- First Affiliated Hospital of Heilongjiang University of Chinese MedicineHarbinChina
| | - Lei Zheng
- Clinical Key Laboratory of Integrated Traditional Chinese and Western Medicine of Heilongjiang University of Chinese MedicineHarbinChina
| | - Weiwei Yu
- First Affiliated Hospital of Heilongjiang University of Chinese MedicineHarbinChina
| | - Wei Teng
- First Affiliated Hospital of Heilongjiang University of Chinese MedicineHarbinChina
| | - Hongtao Cao
- First Affiliated Hospital of Heilongjiang University of Chinese MedicineHarbinChina
| | - Mingyue Li
- First Affiliated Hospital of Heilongjiang University of Chinese MedicineHarbinChina
| | - Jiayong Yao
- First Affiliated Hospital of Heilongjiang University of Chinese MedicineHarbinChina
| | - Xiaoying Liu
- First Affiliated Hospital of Heilongjiang University of Chinese MedicineHarbinChina
| | - Wei Zou
- First Affiliated Hospital of Heilongjiang University of Chinese MedicineHarbinChina
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Rich J, Bennaroch M, Notel L, Patalakh P, Alberola J, Issa F, Opolon P, Bawa O, Rondof W, Marchais A, Dessen P, Meurice G, Le-Gall M, Polrot M, Ser-Le Roux K, Mamchaoui K, Droin N, Raslova H, Maire P, Geoerger B, Pirozhkova I. DiPRO1 distinctly reprograms muscle and mesenchymal cancer cells. EMBO Mol Med 2024; 16:1840-1885. [PMID: 39009887 PMCID: PMC11319797 DOI: 10.1038/s44321-024-00097-z] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/10/2023] [Revised: 06/13/2024] [Accepted: 06/18/2024] [Indexed: 07/17/2024] Open
Abstract
We have recently identified the uncharacterized ZNF555 protein as a component of a productive complex involved in the morbid function of the 4qA locus in facioscapulohumeral dystrophy. Subsequently named DiPRO1 (Death, Differentiation, and PROliferation related PROtein 1), our study provides substantial evidence of its role in the differentiation and proliferation of human myoblasts. DiPRO1 operates through the regulatory binding regions of SIX1, a master regulator of myogenesis. Its relevance extends to mesenchymal tumors, such as rhabdomyosarcoma (RMS) and Ewing sarcoma, where DiPRO1 acts as a repressor via the epigenetic regulators TIF1B and UHRF1, maintaining methylation of cis-regulatory elements and gene promoters. Loss of DiPRO1 mimics the host defense response to virus, awakening retrotransposable repeats and the ZNF/KZFP gene family. This enables the eradication of cancer cells, reprogramming the cellular decision balance towards inflammation and/or apoptosis by controlling TNF-α via NF-kappaB signaling. Finally, our results highlight the vulnerability of mesenchymal cancer tumors to si/shDiPRO1-based nanomedicines, positioning DiPRO1 as a potential therapeutic target.
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Affiliation(s)
- Jeremy Rich
- UMR8126 CNRS, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
| | - Melanie Bennaroch
- UMR8126 CNRS, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
| | - Laura Notel
- UMR8126 CNRS, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
| | - Polina Patalakh
- UMR8126 CNRS, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
| | - Julien Alberola
- UMR8126 CNRS, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
| | - Fayez Issa
- INSERM U1016, CNRS UMR 8104, Institut Cochin, Université Paris-Cité, Paris, France
| | - Paule Opolon
- Pathology and Cytology Section, UMS AMMICA, CNRS, INSERM, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
| | - Olivia Bawa
- Pathology and Cytology Section, UMS AMMICA, CNRS, INSERM, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
| | - Windy Rondof
- Bioinformatics Platform, UMS AMMICA, CNRS, INSERM, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
- Department of Pediatric and Adolescent Oncology, Gustave Roussy Cancer campus, INSERM U1015, Université Paris-Saclay, Villejuif, France
| | - Antonin Marchais
- Bioinformatics Platform, UMS AMMICA, CNRS, INSERM, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
- Department of Pediatric and Adolescent Oncology, Gustave Roussy Cancer campus, INSERM U1015, Université Paris-Saclay, Villejuif, France
| | - Philippe Dessen
- Bioinformatics Platform, UMS AMMICA, CNRS, INSERM, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
| | - Guillaume Meurice
- Bioinformatics Platform, UMS AMMICA, CNRS, INSERM, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
| | - Morgane Le-Gall
- Proteom'IC facility, Université Paris Cité, CNRS, INSERM, Institut Cochin, F-75014, Paris, France
| | - Melanie Polrot
- Pre-clinical Evaluation Unit (PFEP), INSERM, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
| | - Karine Ser-Le Roux
- Pre-clinical Evaluation Unit (PFEP), INSERM, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
| | - Kamel Mamchaoui
- Sorbonne Université, Inserm, Institut de Myologie, Centre de Recherche en Myologie, F-75013, Paris, France
| | - Nathalie Droin
- Genomic Platform, UMS AMMICA US 23 INSERM UAR 3655 CNRS, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
- UMR1287 INSERM, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
| | - Hana Raslova
- UMR1287 INSERM, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France
| | - Pascal Maire
- INSERM U1016, CNRS UMR 8104, Institut Cochin, Université Paris-Cité, Paris, France
| | - Birgit Geoerger
- Department of Pediatric and Adolescent Oncology, Gustave Roussy Cancer campus, INSERM U1015, Université Paris-Saclay, Villejuif, France
| | - Iryna Pirozhkova
- UMR8126 CNRS, Gustave Roussy Cancer campus, Université Paris-Saclay, Villejuif, France.
- INSERM U1016, CNRS UMR 8104, Institut Cochin, Université Paris-Cité, Paris, France.
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12
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Yang J, Xiao S, Li L, Zhu A, Xiao W, Wang Q. Actin Dysregulation Mediates Nephrotoxicity of Cassiae Semen Aqueous Extracts. TOXICS 2024; 12:556. [PMID: 39195658 PMCID: PMC11360101 DOI: 10.3390/toxics12080556] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/28/2024] [Revised: 07/05/2024] [Accepted: 07/26/2024] [Indexed: 08/29/2024]
Abstract
Cassiae semen, commonly consumed as roasted tea, has been widely used for both medicinal purposes and dietary supplements. In this study, we investigated the nephrotoxic effects and underlying mechanisms of Cassiae semen aqueous extracts (CSAEs) using computational and animal models. Both male and female Sprague Dawley rats were treated with 4.73-47.30 g/kg (body weight) of CSAEs by oral gavage twice a day for 7-28 days. We found that serum and urinary biomarkers of kidney injury and kidney coefficients were increased in a dose-dependent manner, and were accompanied by morphological alterations in the kidneys of CSAEs-treated rats. Computational and molecular docking approaches predicted that the three most abundant components of CSAEs-obtusifolin, aurantio-obtusin, and obtusin-exhibited strong affinity for the binding of F-actin, ROCK1, and Rac1, and the RhoA-ROCK pathway was identified as the most likely regulatory mechanism mediating the nephrotoxicity of CSAEs. Consistently, immunofluorescence staining revealed F-actin and cytoskeleton were frequently disturbed in renal cells and brush borders at high doses of CSAEs. Results from gene expression analyses confirmed that CSAEs suppressed the key proteins in the RhoA-ROCK signaling pathway and consequently the expression of F-actin and its stabilization genes. In summary, our findings suggest that Cassiae semen can depolymerize and destabilize actin cytoskeleton by inhibition of the RhoA-ROCK pathway and/or direct binding to F-actin, leading to nephrotoxicity. The consumption of Cassiae semen as a supplement and medicine warrants attention.
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Affiliation(s)
- Jinlan Yang
- Department of Toxicology, School of Public Health, Peking University, Beijing 100191, China; (J.Y.); (S.X.); (L.L.); (A.Z.); (W.X.)
| | - Sheng Xiao
- Department of Toxicology, School of Public Health, Peking University, Beijing 100191, China; (J.Y.); (S.X.); (L.L.); (A.Z.); (W.X.)
| | - Ludi Li
- Department of Toxicology, School of Public Health, Peking University, Beijing 100191, China; (J.Y.); (S.X.); (L.L.); (A.Z.); (W.X.)
| | - An Zhu
- Department of Toxicology, School of Public Health, Peking University, Beijing 100191, China; (J.Y.); (S.X.); (L.L.); (A.Z.); (W.X.)
| | - Wusheng Xiao
- Department of Toxicology, School of Public Health, Peking University, Beijing 100191, China; (J.Y.); (S.X.); (L.L.); (A.Z.); (W.X.)
| | - Qi Wang
- Department of Toxicology, School of Public Health, Peking University, Beijing 100191, China; (J.Y.); (S.X.); (L.L.); (A.Z.); (W.X.)
- Key Laboratory of State Administration of Traditional Chinese Medicine (TCM) for Compatibility Toxicology, Peking University, Beijing 100191, China
- Beijing Key Laboratory of Toxicological Research and Risk Assessment for Food Safety, Peking University, Beijing 100191, China
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13
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Gonzalez-Martinez N, Gibson MI. Post-thaw application of ROCK-inhibitors increases cryopreserved T-cell yield. RSC Med Chem 2023; 14:2058-2067. [PMID: 37859712 PMCID: PMC10583820 DOI: 10.1039/d3md00378g] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/01/2023] [Accepted: 08/18/2023] [Indexed: 10/21/2023] Open
Abstract
Emerging cell-based therapies such as CAR-T (Chimeric Antigen Receptor T) cells require cryopreservation to store and deliver intact and viable cells. Conventional cryopreservation formulations use DMSO to mitigate cold-induced damage, but do not address all the biochemical damage mechanisms induced by cold stress, such as programmed cell death (apoptosis). Rho-associated protein kinases (ROCK) are a key component of apoptosis, and their activation contributes to apoptotic blebbing. Here we demonstrate that the ROCK inhibitor fasudil hydrochloride, when supplemented into the thawing medium of T-cells increases the overall yield of healthy cells. Cell yield was highest using 5 or 10% DMSO cryopreservation solutions, with lower DMSO concentrations (2.5%) leading to significant physical damage to the cells. After optimisation, the post-thaw yield of T-cells increased by approximately 20% using this inhibitor, a significant increase in the context of a therapy. Flow cytometry analysis did not show a significant reduction in the relative percentage of cell populations undergoing apoptosis, but there was a small reduction in the 8 hours following thawing. Fasudil also led to a reduction in reactive oxygen species. Addition of fasudil into the cryopreservation solution, followed by dilution (rather than washing) upon thaw also gave a 20% increase in cell yield, demonstrating how this could be deployed in a cell-therapy context, without needing to change clinical thawing routines. Overall, this shows that modulation of post-thaw biochemical pathways which lead to apoptosis (or other degradative pathways) can be effectively targeted as a strategy to increase T-cell yield and function post-thaw.
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Affiliation(s)
| | - Matthew I Gibson
- Department of Chemistry, University of Warwick Gibbet Hill Road Coventry CV4 7AL UK
- Division of Biomedical Sciences, Warwick Medical School, University of Warwick Gibbet Hill Road Coventry CV4 7AL UK
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14
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Benarroch E. What Is the Role of the Rho-ROCK Pathway in Neurologic Disorders? Neurology 2023; 101:536-543. [PMID: 37722862 PMCID: PMC10516277 DOI: 10.1212/wnl.0000000000207779] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2023] [Accepted: 06/29/2023] [Indexed: 09/20/2023] Open
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15
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Bhargava S, Jankowski J, Merckelbach E, Roth CE, Craveiro RB, Wolf M. Development, Establishment, and Validation of a Model for the Mineralization of Periodontium Remodelling Cells: Cementoblasts. Int J Mol Sci 2023; 24:13829. [PMID: 37762132 PMCID: PMC10531176 DOI: 10.3390/ijms241813829] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/14/2023] [Revised: 09/04/2023] [Accepted: 09/06/2023] [Indexed: 09/29/2023] Open
Abstract
Chronic kidney disease (CKD) patients undergoing dialysis are at high risk of bone fractures. CKD-induced mineral and bone disorder is extended to periodontal disease due to changes in the ionic composition of saliva in CKD patients, dysregulating mineralization, hindering regeneration and thereby promoting the progression of dental complications. Despite the importance of cementum for overall oral health, the mechanisms that regulate its development and regeneration are not well comprehended, and a lack of sufficient in vitro experimental models has hindered research progress. In this study, the impact of experimental conditions on the calcification of cementoblasts was systematically investigated, aimed at establishing a standardized and validated model for the calcification of cementoblasts. The effects of phosphate, calcium, ascorbic acid, β-glycerolphosphate, dexamethasone, and fetal calf serum on the calcification process of cementoblasts were analyzed over a wide range of concentrations and time points by investigating calcium content, cell viability, gene expression and kinase activity. Cementoblasts calcified in a concentration- and time-dependent manner with higher concentrations of supplements cause a higher degree of calcification but decreased cell viability. Phosphate and calcium have a significantly stronger effect on cementoblast calcification processes compared to osteogenic supplements: ascorbic acid, β-glycerolphosphate, and dexamethasone induce calcification over a wide range of osteogenic signalling pathways, with osteopontin being a central target of gene regulation. Conversely, treatment with ascorbic acid, β-glycerolphosphate, and dexamethasone leads to activating only selected pathways, especially promoting bone sialoprotein expression. The developed and validated cementoblast calcification protocol, incubating up to 60% confluent cementoblasts with 1.9 mmol L-1 of phosphate supplementation for a reasonable, multi-pathway calcification induction and 10 mmol L-1 β-glycerolphosphate, 75 µmol L-1 ascorbic acid and 10 nmol L-1 dexamethasone for a reasonable osteogenic differentiation-based calcification induction, provides standard in vitro experimental models for better understanding cementoblast function and regeneration.
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Affiliation(s)
- Shruti Bhargava
- Institute of Molecular Cardiovascular Research, Medical Faculty, RWTH Aachen University, 52062 Aachen, Germany; (S.B.); (E.M.)
| | - Joachim Jankowski
- Institute of Molecular Cardiovascular Research, Medical Faculty, RWTH Aachen University, 52062 Aachen, Germany; (S.B.); (E.M.)
- Aachen-Maastricht Institute for Cardiorenal Disease (AMICARE), University Hospital RWTH Aachen, 52062 Aachen, Germany
- Experimental Vascular Pathology, Cardiovascular Research Institute Maastricht (CARIM), University of Maastricht, 6211 Maastricht, The Netherlands
| | - Erik Merckelbach
- Institute of Molecular Cardiovascular Research, Medical Faculty, RWTH Aachen University, 52062 Aachen, Germany; (S.B.); (E.M.)
| | - Charlotte Elisa Roth
- Department of Orthodontics, Dental Clinic, University of Aachen, Pauwelsstr. 30, 52074 Aachen, Germany; (C.E.R.); (R.B.C.); (M.W.)
| | - Rogerio Bastos Craveiro
- Department of Orthodontics, Dental Clinic, University of Aachen, Pauwelsstr. 30, 52074 Aachen, Germany; (C.E.R.); (R.B.C.); (M.W.)
| | - Michael Wolf
- Department of Orthodontics, Dental Clinic, University of Aachen, Pauwelsstr. 30, 52074 Aachen, Germany; (C.E.R.); (R.B.C.); (M.W.)
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16
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Gao X, Bayraktutan U. TNF-α evokes blood-brain barrier dysfunction through activation of Rho-kinase and neurokinin 1 receptor. Immunobiology 2023; 228:152706. [PMID: 37454559 DOI: 10.1016/j.imbio.2023.152706] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/22/2023] [Revised: 05/17/2023] [Accepted: 07/04/2023] [Indexed: 07/18/2023]
Abstract
Ischaemic stroke, accompanied by neuroinflammation, impairs blood-brain barrier (BBB) integrity through a complex mechanism involving activation of both RhoA/Rho kinase/myosin light chain-2 and neurokinin 1 receptor (NK1R). Using an in vitro model of human BBB composed of brain microvascular endothelial cells (BMEC), astrocytes and pericytes, this study examined the potential contributions of these elements to BBB damage induced by elevated availability of pro-inflammatory cytokine, TNF-α. Treatment of human BMECs with TNF-α significantly enhanced RhoA activity and the protein expressions of Rho kinase and phosphorylated Ser19MLC-2 while decreasing that of NK1R. Pharmacological inhibition of Rho kinase by Y-27632 and NK1R by CP96345 neutralised the disruptive effects of TNF-α on BBB integrity and function as ascertained by reversal of decreases in transendothelial electrical resistance and increases in paracellular flux of low molecular weight permeability marker, sodium fluorescein, respectively. Suppression of RhoA activation, mitigation of actin stress fibre formation and restoration of plasma membrane localisation of tight junction protein zonula occludens-1 appeared to contribute to the barrier-protective effects of both Y-27632 and CP96345. Attenuation of TNF-α-mediated increases in NK1R protein expression in BMEC by Y-27632 suggests that RhoA/Rho kinase pathway acts upstream to NK1R. In conclusion, specific inhibition of Rho kinase in cerebrovascular conditions, accompanied by excessive release of pro-inflammatory cytokine TNF-α, helps preserve endothelial cell morphology and inter-endothelial cell barrier formation and may serve as an important therapeutic target.
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Affiliation(s)
- Xin Gao
- Academic Unit of Mental Health and Clinical Neurosciences, School of Medicine, University of Nottingham, Nottingham, UK
| | - Ulvi Bayraktutan
- Academic Unit of Mental Health and Clinical Neurosciences, School of Medicine, University of Nottingham, Nottingham, UK.
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Lu W, Chen Z, Wen J. The role of RhoA/ROCK pathway in the ischemic stroke-induced neuroinflammation. Biomed Pharmacother 2023; 165:115141. [PMID: 37437375 DOI: 10.1016/j.biopha.2023.115141] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/20/2023] [Revised: 07/03/2023] [Accepted: 07/07/2023] [Indexed: 07/14/2023] Open
Abstract
It is widely known that ischemic stroke is the prominent cause of death and disability. To date, neuroinflammation following ischemic stroke represents a complex event, which is an essential process and affects the prognosis of both experimental stroke animals and stroke patients. Intense neuroinflammation occurring during the acute phase of stroke contributes to neuronal injury, BBB breakdown, and worse neurological outcomes. Inhibition of neuroinflammation may be a promising target in the development of new therapeutic strategies. RhoA is a small GTPase protein that activates a downstream effector, ROCK. The up-regulation of RhoA/ROCK pathway possesses important roles in promoting the neuroinflammation and mediating brain injury. In addition, nuclear factor-kappa B (NF-κB) is another vital regulator of ischemic stroke-induced neuroinflammation through regulating the functions of microglial cells and astrocytes. After stroke onset, the microglial cells and astrocytes are activated and undergo the morphological and functional changes, thereby deeply participate in a complicated neuroinflammation cascade. In this review, we focused on the relationship among RhoA/ROCK pathway, NF-κB and glial cells in the neuroinflammation following ischemic stroke to reveal new strategies for preventing the intense neuroinflammation.
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Affiliation(s)
- Weizhuo Lu
- Medical Branch, Hefei Technology College, Hefei, China
| | - Zhiwu Chen
- Department of Pharmacology, School of Basic Medical Sciences, Anhui Medical University, Hefei, China.
| | - Jiyue Wen
- Department of Pharmacology, School of Basic Medical Sciences, Anhui Medical University, Hefei, China.
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18
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Behnam M, Asadpour R, Topraggaleh TR, Hamali H. Improvement of post-thaw quality and fertilizing ability of bull spermatozoa using Rho kinase inhibitor in freezing extender. Front Vet Sci 2023; 10:1155048. [PMID: 37483290 PMCID: PMC10359164 DOI: 10.3389/fvets.2023.1155048] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/31/2023] [Accepted: 06/12/2023] [Indexed: 07/25/2023] Open
Abstract
In this study, it was hypothesized that the addition of an appropriate concentration of Y-27632 (a ROCK inhibitor) to the freezing extender prevents cryopreservation-induced apoptosis and improves embryonic development after in vitro fertilization (IVF). Semen samples were collected from five fertile Simmental bulls using an artificial vagina twice a week for 4 weeks. Selected samples were pooled and diluted with Tris-egg-yolk-glycerol (TEYG) extender containing different concentrations of Y-27632 (0, 10, 20, 30, and 40 μM) and then frozen in liquid nitrogen. After thawing, computer-assisted semen analysis (CASA), plasma membrane integrity, and acrosome intactness were evaluated in terms of morphological abnormalities, intracellular generation of reactive oxygen species (ROS), DNA fragmentation, phosphatidylserine (PS) externalization, and apoptotic-related gene expression. Finally, groups of frozen and thawed spermatozoa were used for bovine oocyte IVF. The results show that the semen extender at a concentration of 20 μM Y-27632 effectively improved total motility (TM), curvilinear velocity (VCL), as well as the plasma membrane and acrosome integrity compared to the control group (p < 0.05). Intracellular ROS levels were significantly (p < 0.05) lower in samples treated with 30 μM Y-27632 compared to the control specimen. Furthermore, supplementation of the semen extender with 20 μM Y-27632 resulted in more viable spermatozoa compared with the control group (p < 0.05). According to qRT-PCR results, the expression levels of BAX and CASPASE-9 genes in samples treated with 30 μM Y-27632 were significantly downregulated, while the expression of BCL2 was increased compared to the control (p < 0.05). The results of IVF demonstrated that the treatment of frozen-thawed spermatozoa with 20 μM Y-27632 increased blastocyst rates compared to the control group (p < 0.05). In conclusion, the addition of 20 μM Y-27632 into the freezing extender can improve the functionality and the fertilizing capacity of frozen spermatozoa due to its antioxidative and anti-apoptotic properties.
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Affiliation(s)
- Mina Behnam
- Department of Clinical Science, Faculty of Veterinary Medicine, University of Tabriz, Tabriz, Iran
| | - Reza Asadpour
- Department of Clinical Science, Faculty of Veterinary Medicine, University of Tabriz, Tabriz, Iran
| | - Tohid Rezaei Topraggaleh
- Reproductive Health Research Center, Clinical Research Institute, Urmia University of Medical Sciences, Urmia, Iran
- Department of Anatomical Sciences, School of Medicine, Urmia University of Medical Sciences, Urmia, Iran
| | - Hossein Hamali
- Department of Clinical Science, Faculty of Veterinary Medicine, University of Tabriz, Tabriz, Iran
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19
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Lahooti B, Akwii RG, Patel D, ShahbaziNia S, Lamprou M, Madadi M, Abbruscato TJ, Astrinidis A, Bickel U, Al-Ahmad A, German NA, Mattheolabakis G, Mikelis CM. Endothelial-Specific Targeting of RhoA Signaling via CD31 Antibody-Conjugated Nanoparticles. J Pharmacol Exp Ther 2023; 385:35-49. [PMID: 36746610 PMCID: PMC10029826 DOI: 10.1124/jpet.122.001384] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/25/2022] [Revised: 01/04/2023] [Accepted: 01/17/2023] [Indexed: 02/08/2023] Open
Abstract
Existing vascular endothelial growth factor-oriented antiangiogenic approaches are known for their high potency. However, significant side effects associated with their use drive the need for novel antiangiogenic strategies. The small GTPase RhoA is an established regulator of actin cytoskeletal dynamics. Previous studies have highlighted the impact of endothelial RhoA pathway on angiogenesis. Rho-associate kinase (ROCK), a direct RhoA effector, is potently inhibited by Fasudil, a clinically relevant ROCK inhibitor. Here, we aimed to target the RhoA signaling in endothelial cells by generating Fasudil-encapsulated CD31-targeting liposomes as a potential antiangiogenic therapy. The liposomes presented desirable characteristics, preferential binding to CD31-expressing HEK293T cells and to endothelial cells, inhibited stress fiber formation and cytoskeletal-related morphometric parameters, and inhibited in vitro angiogenic functions. Overall, this work shows that the nanodelivery-mediated endothelial targeting of RhoA signaling can offer a promising strategy for angiogenesis inhibition in vascular-related diseases. SIGNIFICANCE STATEMENT: Systemic administration of antiangiogenic therapeutics induces side effects to non-targeted tissues. This study, among others, has shown the impact of the RhoA signaling in the endothelial cells and their angiogenic functions. Here, to minimize potential toxicity, this study generated CD31-targeting liposomes with encapsulated Fasudil, a clinically relevant Rho kinase inhibitor, and successfully targeted endothelial cells. In this proof-of-principle study, the efficient Fasudil delivery, its impact on the endothelial signaling, morphometric alterations, and angiogenic functions verify the benefits of site-targeted antiangiogenic therapy.
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Affiliation(s)
- Behnaz Lahooti
- Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, Amarillo, Texas (B.L., R.G.A., D.P., S.S., T.J.A., U.B., A.A.-A., N.A.G., C.M.M.); Laboratory of Molecular Pharmacology, Department of Pharmacy, University of Patras, Patras, Greece(M.L., C.M.M.); Department of Marketing and Business Analytics, Lucas College and Graduate School of Business, San Jose State University, San Jose, California (M.M.); Department of Pediatrics, University of Tennessee Health Sciences Center and Le Bonheur Children's Hospital, Memphis, Tennessee (A.A.); and School of Basic Pharmaceutical and Toxicological Sciences, College of Pharmacy, University of Louisiana Monroe, Monroe, Louisiana (G.M.)
| | - Racheal G Akwii
- Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, Amarillo, Texas (B.L., R.G.A., D.P., S.S., T.J.A., U.B., A.A.-A., N.A.G., C.M.M.); Laboratory of Molecular Pharmacology, Department of Pharmacy, University of Patras, Patras, Greece(M.L., C.M.M.); Department of Marketing and Business Analytics, Lucas College and Graduate School of Business, San Jose State University, San Jose, California (M.M.); Department of Pediatrics, University of Tennessee Health Sciences Center and Le Bonheur Children's Hospital, Memphis, Tennessee (A.A.); and School of Basic Pharmaceutical and Toxicological Sciences, College of Pharmacy, University of Louisiana Monroe, Monroe, Louisiana (G.M.)
| | - Dhavalkumar Patel
- Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, Amarillo, Texas (B.L., R.G.A., D.P., S.S., T.J.A., U.B., A.A.-A., N.A.G., C.M.M.); Laboratory of Molecular Pharmacology, Department of Pharmacy, University of Patras, Patras, Greece(M.L., C.M.M.); Department of Marketing and Business Analytics, Lucas College and Graduate School of Business, San Jose State University, San Jose, California (M.M.); Department of Pediatrics, University of Tennessee Health Sciences Center and Le Bonheur Children's Hospital, Memphis, Tennessee (A.A.); and School of Basic Pharmaceutical and Toxicological Sciences, College of Pharmacy, University of Louisiana Monroe, Monroe, Louisiana (G.M.)
| | - Siavash ShahbaziNia
- Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, Amarillo, Texas (B.L., R.G.A., D.P., S.S., T.J.A., U.B., A.A.-A., N.A.G., C.M.M.); Laboratory of Molecular Pharmacology, Department of Pharmacy, University of Patras, Patras, Greece(M.L., C.M.M.); Department of Marketing and Business Analytics, Lucas College and Graduate School of Business, San Jose State University, San Jose, California (M.M.); Department of Pediatrics, University of Tennessee Health Sciences Center and Le Bonheur Children's Hospital, Memphis, Tennessee (A.A.); and School of Basic Pharmaceutical and Toxicological Sciences, College of Pharmacy, University of Louisiana Monroe, Monroe, Louisiana (G.M.)
| | - Margarita Lamprou
- Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, Amarillo, Texas (B.L., R.G.A., D.P., S.S., T.J.A., U.B., A.A.-A., N.A.G., C.M.M.); Laboratory of Molecular Pharmacology, Department of Pharmacy, University of Patras, Patras, Greece(M.L., C.M.M.); Department of Marketing and Business Analytics, Lucas College and Graduate School of Business, San Jose State University, San Jose, California (M.M.); Department of Pediatrics, University of Tennessee Health Sciences Center and Le Bonheur Children's Hospital, Memphis, Tennessee (A.A.); and School of Basic Pharmaceutical and Toxicological Sciences, College of Pharmacy, University of Louisiana Monroe, Monroe, Louisiana (G.M.)
| | - Mahboubeh Madadi
- Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, Amarillo, Texas (B.L., R.G.A., D.P., S.S., T.J.A., U.B., A.A.-A., N.A.G., C.M.M.); Laboratory of Molecular Pharmacology, Department of Pharmacy, University of Patras, Patras, Greece(M.L., C.M.M.); Department of Marketing and Business Analytics, Lucas College and Graduate School of Business, San Jose State University, San Jose, California (M.M.); Department of Pediatrics, University of Tennessee Health Sciences Center and Le Bonheur Children's Hospital, Memphis, Tennessee (A.A.); and School of Basic Pharmaceutical and Toxicological Sciences, College of Pharmacy, University of Louisiana Monroe, Monroe, Louisiana (G.M.)
| | - Thomas J Abbruscato
- Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, Amarillo, Texas (B.L., R.G.A., D.P., S.S., T.J.A., U.B., A.A.-A., N.A.G., C.M.M.); Laboratory of Molecular Pharmacology, Department of Pharmacy, University of Patras, Patras, Greece(M.L., C.M.M.); Department of Marketing and Business Analytics, Lucas College and Graduate School of Business, San Jose State University, San Jose, California (M.M.); Department of Pediatrics, University of Tennessee Health Sciences Center and Le Bonheur Children's Hospital, Memphis, Tennessee (A.A.); and School of Basic Pharmaceutical and Toxicological Sciences, College of Pharmacy, University of Louisiana Monroe, Monroe, Louisiana (G.M.)
| | - Aristotelis Astrinidis
- Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, Amarillo, Texas (B.L., R.G.A., D.P., S.S., T.J.A., U.B., A.A.-A., N.A.G., C.M.M.); Laboratory of Molecular Pharmacology, Department of Pharmacy, University of Patras, Patras, Greece(M.L., C.M.M.); Department of Marketing and Business Analytics, Lucas College and Graduate School of Business, San Jose State University, San Jose, California (M.M.); Department of Pediatrics, University of Tennessee Health Sciences Center and Le Bonheur Children's Hospital, Memphis, Tennessee (A.A.); and School of Basic Pharmaceutical and Toxicological Sciences, College of Pharmacy, University of Louisiana Monroe, Monroe, Louisiana (G.M.)
| | - Ulrich Bickel
- Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, Amarillo, Texas (B.L., R.G.A., D.P., S.S., T.J.A., U.B., A.A.-A., N.A.G., C.M.M.); Laboratory of Molecular Pharmacology, Department of Pharmacy, University of Patras, Patras, Greece(M.L., C.M.M.); Department of Marketing and Business Analytics, Lucas College and Graduate School of Business, San Jose State University, San Jose, California (M.M.); Department of Pediatrics, University of Tennessee Health Sciences Center and Le Bonheur Children's Hospital, Memphis, Tennessee (A.A.); and School of Basic Pharmaceutical and Toxicological Sciences, College of Pharmacy, University of Louisiana Monroe, Monroe, Louisiana (G.M.)
| | - Abraham Al-Ahmad
- Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, Amarillo, Texas (B.L., R.G.A., D.P., S.S., T.J.A., U.B., A.A.-A., N.A.G., C.M.M.); Laboratory of Molecular Pharmacology, Department of Pharmacy, University of Patras, Patras, Greece(M.L., C.M.M.); Department of Marketing and Business Analytics, Lucas College and Graduate School of Business, San Jose State University, San Jose, California (M.M.); Department of Pediatrics, University of Tennessee Health Sciences Center and Le Bonheur Children's Hospital, Memphis, Tennessee (A.A.); and School of Basic Pharmaceutical and Toxicological Sciences, College of Pharmacy, University of Louisiana Monroe, Monroe, Louisiana (G.M.)
| | - Nadezhda A German
- Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, Amarillo, Texas (B.L., R.G.A., D.P., S.S., T.J.A., U.B., A.A.-A., N.A.G., C.M.M.); Laboratory of Molecular Pharmacology, Department of Pharmacy, University of Patras, Patras, Greece(M.L., C.M.M.); Department of Marketing and Business Analytics, Lucas College and Graduate School of Business, San Jose State University, San Jose, California (M.M.); Department of Pediatrics, University of Tennessee Health Sciences Center and Le Bonheur Children's Hospital, Memphis, Tennessee (A.A.); and School of Basic Pharmaceutical and Toxicological Sciences, College of Pharmacy, University of Louisiana Monroe, Monroe, Louisiana (G.M.)
| | - George Mattheolabakis
- Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, Amarillo, Texas (B.L., R.G.A., D.P., S.S., T.J.A., U.B., A.A.-A., N.A.G., C.M.M.); Laboratory of Molecular Pharmacology, Department of Pharmacy, University of Patras, Patras, Greece(M.L., C.M.M.); Department of Marketing and Business Analytics, Lucas College and Graduate School of Business, San Jose State University, San Jose, California (M.M.); Department of Pediatrics, University of Tennessee Health Sciences Center and Le Bonheur Children's Hospital, Memphis, Tennessee (A.A.); and School of Basic Pharmaceutical and Toxicological Sciences, College of Pharmacy, University of Louisiana Monroe, Monroe, Louisiana (G.M.)
| | - Constantinos M Mikelis
- Department of Pharmaceutical Sciences, School of Pharmacy, Texas Tech University Health Sciences Center, Amarillo, Texas (B.L., R.G.A., D.P., S.S., T.J.A., U.B., A.A.-A., N.A.G., C.M.M.); Laboratory of Molecular Pharmacology, Department of Pharmacy, University of Patras, Patras, Greece(M.L., C.M.M.); Department of Marketing and Business Analytics, Lucas College and Graduate School of Business, San Jose State University, San Jose, California (M.M.); Department of Pediatrics, University of Tennessee Health Sciences Center and Le Bonheur Children's Hospital, Memphis, Tennessee (A.A.); and School of Basic Pharmaceutical and Toxicological Sciences, College of Pharmacy, University of Louisiana Monroe, Monroe, Louisiana (G.M.)
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20
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Chen YC, Chen KF, Andrew Lin KY, Su HP, Wu DN, Lin CH. Evaluation of toxicity of polystyrene microplastics under realistic exposure levels in human vascular endothelial EA.hy926 cells. CHEMOSPHERE 2023; 313:137582. [PMID: 36529175 DOI: 10.1016/j.chemosphere.2022.137582] [Citation(s) in RCA: 15] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/04/2022] [Revised: 12/13/2022] [Accepted: 12/14/2022] [Indexed: 06/17/2023]
Abstract
Microplastics (MPs) have emerged as a global concern, with a recent study being the first to detect them in the bloodstream of healthy people. However, precise information regarding the toxic effects of MPs on the human vascular system is currently lacking. In this study, we used human vascular endothelial EA. hy926 cells to examine the toxic potential of polystyrene MPs (PSMPs) under realistic blood concentrations. Our findings indicated that PSMPs can cause oxidative stress by reducing the expression of antioxidants, thereby leading to apoptotic cytotoxicity in EA. hy926 cells. Furthermore, the protective potential of heat shock proteins can be reduced by PSMPs. PSMP-induced apoptosis might also lower the expression of rho-associated protein kinase-1 and nuclear factor-κB expression, thus dampening LRR- and pyrin domain-containing protein 3 in EA. hy926 cells. Moreover, we observed that PSMPs induce vascular barrier dysfunction via the depletion of zonula occludens-1 protein. However, although protein expression of the nuclear hormone receptor 77 was inhibited, no significant increase in ectin-like oxidized low-density lipoprotein receptor-1 was noted in PSMP-treated EA. hy926 cells. These results demonstrate that exposure to PSMPs may not sufficiently increase the risk of developing atherosclerosis. Overall, our research signifies that exposure to realistic blood concentrations of PSMPs is associated with low atherosclerotic cardiovascular risk in humans.
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Affiliation(s)
- Yi-Chun Chen
- Department of Biotechnology, National Formosa University, Yunlin, 63208, Taiwan; Department of Science and Environment Studies, The Education University of Hong Kong, New Territories, Hong Kong
| | - Ku-Fan Chen
- Department of Civil Engineering, National Chi Nan University, Nantou, 54561, Taiwan
| | - Kun-Yi Andrew Lin
- Department of Environmental Engineering, National Chung Hsing University, Taichung, 40227, Taiwan
| | - Han-Pang Su
- Third Research Division, Taiwan Research Institute, New Taipei City, 251030, Taiwan
| | - Dong-Ni Wu
- Department of Biotechnology, National Formosa University, Yunlin, 63208, Taiwan
| | - Chia-Hua Lin
- Department of Biotechnology, National Formosa University, Yunlin, 63208, Taiwan.
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21
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Kopecny LR, Lee BWH, Coroneo MT. A systematic review on the effects of ROCK inhibitors on proliferation and/or differentiation in human somatic stem cells: A hypothesis that ROCK inhibitors support corneal endothelial healing via acting on the limbal stem cell niche. Ocul Surf 2023; 27:16-29. [PMID: 36586668 DOI: 10.1016/j.jtos.2022.12.008] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/09/2022] [Revised: 12/18/2022] [Accepted: 12/22/2022] [Indexed: 12/29/2022]
Abstract
Rho kinase inhibitors (ROCKi) have attracted growing multidisciplinary interest, particularly in Ophthalmology where the question as to how they promote corneal endothelial healing remains unresolved. Concurrently, stem cell biology has rapidly progressed in unravelling drivers of stem cell (SC) proliferation and differentiation, where mechanical niche factors and the actin cytoskeleton are increasingly recognized as key players. There is mounting evidence from the study of the peripheral corneal endothelium that supports the likelihood of an internal limbal stem cell niche. The possibility that ROCKi stimulate the endothelial SC niche has not been addressed. Furthermore, there is currently a paucity of data that directly evaluates whether ROCKi promotes corneal endothelial healing by acting on this limbal SC niche located near the transition zone. Therefore, we performed a systematic review examining the effects ROCKi on the proliferation and differentiation of human somatic SC, to provide insight into its effects on various human SC populations. An appraisal of electronic searches of four databases identified 1 in vivo and 58 in vitro studies (36 evaluated proliferation while 53 examined differentiation). Types of SC studied included mesenchymal (n = 32), epithelial (n = 11), epidermal (n = 8), hematopoietic and other (n = 8). The ROCK 1/2 selective inhibitor Y-27632 was used in almost all studies (n = 58), while several studies evaluated ≥2 ROCKi (n = 4) including fasudil, H-1152, and KD025. ROCKi significantly influenced human somatic SC proliferation in 81% of studies (29/36) and SC differentiation in 94% of studies (50/53). The present systemic review highlights that ROCKi are influential in regulating human SC proliferation and differentiation, and provides evidence to support the hypothesis that ROCKi promotes corneal endothelial division and maintenance via acting on the inner limbal SC niche.
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Affiliation(s)
- Lloyd R Kopecny
- School of Clinical Medicine, University of New South Wales, Sydney, Australia.
| | - Brendon W H Lee
- Department of Ophthalmology, School of Clinical Medicine, University of New South Wales, Level 2 South Wing, Edmund Blacket Building, Prince of Wales Hospital, Randwick, NSW, 2031, Australia
| | - Minas T Coroneo
- Department of Ophthalmology, Prince of Wales Hospital, Sydney, Australia
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22
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Bradic M, Taleb S, Thomas B, Chidiac O, Robay A, Hassan N, Malek J, Ait Hssain A, Abi Khalil C. DNA methylation predicts the outcome of COVID-19 patients with acute respiratory distress syndrome. J Transl Med 2022; 20:526. [PMID: 36371196 PMCID: PMC9652914 DOI: 10.1186/s12967-022-03737-5] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/10/2022] [Accepted: 10/30/2022] [Indexed: 11/15/2022] Open
Abstract
BACKGROUND COVID-19 infections could be complicated by acute respiratory distress syndrome (ARDS), increasing mortality risk. We sought to assess the methylome of peripheral blood mononuclear cells in COVID-19 with ARDS. METHODS We recruited 100 COVID-19 patients with ARDS under mechanical ventilation and 33 non-COVID-19 controls between April and July 2020. COVID-19 patients were followed at four time points for 60 days. DNA methylation and immune cell populations were measured at each time point. A multivariate cox proportional risk regression analysis was conducted to identify predictive signatures according to survival. RESULTS The comparison of COVID-19 to controls at inclusion revealed the presence of a 14.4% difference in promoter-associated CpGs in genes that control immune-related pathways such as interferon-gamma and interferon-alpha responses. On day 60, 24% of patients died. The inter-comparison of baseline DNA methylation to the last recorded time point in both COVID-19 groups or the intra-comparison between inclusion and the end of follow-up in every group showed that most changes occurred as the disease progressed, mainly in the AIM gene, which is associated with an intensified immune response in those who recovered. The multivariate Cox proportional risk regression analysis showed that higher methylation of the "Apoptotic execution Pathway" genes (ROC1, ZNF789, and H1F0) at inclusion increases mortality risk by over twofold. CONCLUSION We observed an epigenetic signature of immune-related genes in COVID-19 patients with ARDS. Further, Hypermethylation of the apoptotic execution pathway genes predicts the outcome. TRIAL REGISTRATION IMRPOVIE study, NCT04473131.
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Affiliation(s)
- Martina Bradic
- grid.5386.8000000041936877XDepartment of Genetic Medicine, Weill Cornell Medicine, New York, USA ,grid.51462.340000 0001 2171 9952Marie-Josee and Henry R. Kravis Center for Molecular Oncology, Memorial Sloan Kettering Cancer Center, New York, NY USA
| | - Sarah Taleb
- grid.452146.00000 0004 1789 3191Division of Genomics and Translational Biomedicine, College of Health and Life Sciences- HBKU, Doha, Qatar
| | - Binitha Thomas
- grid.416973.e0000 0004 0582 4340Epigenetics Cardiovascular Lab, Weill Cornell Medicine-Qatar, Doha, Qatar
| | - Omar Chidiac
- grid.416973.e0000 0004 0582 4340Epigenetics Cardiovascular Lab, Weill Cornell Medicine-Qatar, Doha, Qatar
| | - Amal Robay
- grid.416973.e0000 0004 0582 4340Epigenetics Cardiovascular Lab, Weill Cornell Medicine-Qatar, Doha, Qatar
| | - Nessiya Hassan
- grid.413548.f0000 0004 0571 546XNursery and midwifery research department, Hamad Medical Corporation., Doha, Qatar
| | - Joel Malek
- grid.416973.e0000 0004 0582 4340Genomics Core. Weill Cornell Medicine-Qatar., Doha, Qatar
| | - Ali Ait Hssain
- grid.413548.f0000 0004 0571 546XMedical Intensive Care Unit, Hamad Medical Corporation., Doha, Qatar
| | - Charbel Abi Khalil
- Department of Genetic Medicine, Weill Cornell Medicine, New York, USA. .,Epigenetics Cardiovascular Lab, Weill Cornell Medicine-Qatar, Doha, Qatar. .,Joan and Sanford I. Weill Department of Medicine., Weill Cornell Medicine, New York, USA.
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23
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Nallakumarasamy A, Jeyaraman M, Maffulli N, Jeyaraman N, Suresh V, Ravichandran S, Gupta M, Potty AG, El-Amin SF, Khanna M, Gupta A. Mesenchymal Stromal Cell-Derived Extracellular Vesicles in Wound Healing. Life (Basel) 2022; 12:1733. [PMID: 36362890 PMCID: PMC9699035 DOI: 10.3390/life12111733] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/16/2022] [Accepted: 10/24/2022] [Indexed: 07/26/2023] Open
Abstract
The well-orchestrated process of wound healing may be negatively impacted from interrupted or incomplete tissue regenerative processes. The healing potential is further compromised in patients with diabetes mellitus, chronic venous insufficiency, critical limb ischemia, and immunocompromised conditions, with a high health care burden and expenditure. Stem cell-based therapy has shown promising results in clinical studies. Mesenchymal stem cell-derived exosomes (MSC Exos) may favorably impact intercellular signaling and immunomodulation, promoting neoangiogenesis, collagen synthesis, and neoepithelization. This article gives an outline of the biogenesis and mechanism of extracellular vesicles (EVs), particularly exosomes, in the process of tissue regeneration and discusses the use of preconditioned exosomes, platelet-rich plasma-derived exosomes, and engineered exosomes in three-dimensional bioscaffolds such as hydrogels (collagen and chitosan) to prolong the contact time of exosomes at the recipient site within the target tissue. An appropriate antibiotic therapy based on culture-specific guidance coupled with the knowledge of biopolymers helps to fabricate nanotherapeutic materials loaded with MSC Exos to effectively deliver drugs locally and promote novel approaches for the management of chronic wounds.
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Affiliation(s)
- Arulkumar Nallakumarasamy
- Department of Orthopaedics, All India Institute of Medical Sciences, Bhubaneswar 751019, Odissa, India
- Fellow in Orthopaedic Rheumatology, Dr. RML National Law University, Lucknow 226010, Uttar Pradesh, India
- Indian Stem Cell Study Group (ISCSG) Association, Lucknow 226010, Uttar Pradesh, India
| | - Madhan Jeyaraman
- Indian Stem Cell Study Group (ISCSG) Association, Lucknow 226010, Uttar Pradesh, India
- Department of Orthopaedics, Faculty of Medicine—Sri Lalithambigai Medical College and Hospital, Dr. MGR Educational and Research Institute, Chennai 600095, Tamil Nadu, India
- Department of Medical Research and Translational Medicine, Faculty of Medicine—Sri Lalithambigai Medical College and Hospital, Dr. MGR Educational and Research Institute, Chennai 600095, Tamil Nadu, India
- Department of Biotechnology, School of Engineering and Technology, Sharda University, Greater Noida 201310, Uttar Pradesh, India
- South Texas Orthopaedic Research Institute (STORI Inc.), Laredo, TX 78045, USA
| | - Nicola Maffulli
- Department of Musculoskeletal Disorders, School of Medicine and Surgery, University of Salerno, 84084 Fisciano, Italy
- San Giovanni di Dio e Ruggi D’Aragona Hospital “Clinica Ortopedica” Department, Hospital of Salerno, 84124 Salerno, Italy
- Barts and the London School of Medicine and Dentistry, Centre for Sports and Exercise Medicine, Queen Mary University of London, London E1 4DG, UK
- School of Pharmacy and Bioengineering, Keele University School of Medicine, Stoke on Trent ST5 5BG, UK
| | - Naveen Jeyaraman
- Indian Stem Cell Study Group (ISCSG) Association, Lucknow 226010, Uttar Pradesh, India
- Fellow in Joint Replacement, Department of Orthopaedics, Atlas Hospitals, Tiruchirappalli 620002, Tamil Nadu, India
| | - Veerasivabalan Suresh
- Department of Obstetrics-Gynecology, Madras Medical College and Hospital, Chennai 600003, Tamil Nadu, India
| | - Srinath Ravichandran
- Department of General and GI Surgery, Stepping Hill Hospital, Stockport NHS Foundation Trust, Stockport SK27JE, UK
| | - Manu Gupta
- Polar Aesthetics Dental & Cosmetic Centre, Noida 201301, Uttar Pradesh, India
| | - Anish G. Potty
- South Texas Orthopaedic Research Institute (STORI Inc.), Laredo, TX 78045, USA
| | - Saadiq F. El-Amin
- El-Amin Orthopaedic & Sports Medicine Institute, Lawrenceville, GA 30043, USA
- Regenerative Sports Medicine, Lawrenceville, GA 30043, USA
- BioIntegrate, Lawrenceville, GA 30043, USA
| | - Manish Khanna
- Indian Stem Cell Study Group (ISCSG) Association, Lucknow 226010, Uttar Pradesh, India
- Department of Orthopaedics, Autonomous State Medical College, Ayodhya 224135, Uttar Pradesh, India
| | - Ashim Gupta
- Indian Stem Cell Study Group (ISCSG) Association, Lucknow 226010, Uttar Pradesh, India
- South Texas Orthopaedic Research Institute (STORI Inc.), Laredo, TX 78045, USA
- BioIntegrate, Lawrenceville, GA 30043, USA
- Regenerative Orthopaedics, Noida 201301, Uttar Pradesh, India
- Future Biologics, Lawrenceville, GA 30043, USA
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24
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Saadeldin IM, Tanga BM, Bang S, Seo C, Koo O, Yun SH, Kim SI, Lee S, Cho J. ROCK Inhibitor (Y-27632) Abolishes the Negative Impacts of miR-155 in the Endometrium-Derived Extracellular Vesicles and Supports Embryo Attachment. Cells 2022; 11:cells11193178. [PMID: 36231141 PMCID: PMC9564368 DOI: 10.3390/cells11193178] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/17/2022] [Revised: 10/07/2022] [Accepted: 10/07/2022] [Indexed: 11/16/2022] Open
Abstract
Extracellular vesicles (EVs) are nanosized vesicles that act as snapshots of cellular components and mediate cellular communications, but they may contain cargo contents with undesired effects. We developed a model to improve the effects of endometrium-derived EVs (Endo-EVs) on the porcine embryo attachment in feeder-free culture conditions. Endo-EVs cargo contents were analyzed using conventional and real-time PCR for micro-RNAs, messenger RNAs, and proteomics. Porcine embryos were generated by parthenogenetic electric activation in feeder-free culture conditions supplemented with or without Endo-EVs. The cellular uptake of Endo-EVs was confirmed using the lipophilic dye PKH26. Endo-EVs cargo contained miR-100, miR-132, and miR-155, together with the mRNAs of porcine endogenous retrovirus (PERV) and β-catenin. Targeting PERV with CRISPR/Cas9 resulted in reduced expression of PERV mRNA transcripts and increased miR-155 in the Endo-EVs, and supplementing these in embryos reduced embryo attachment. Supplementing the medium containing Endo-EVs with miR-155 inhibitor significantly improved the embryo attachment with a few outgrowths, while supplementing with Rho-kinase inhibitor (RI, Y-27632) dramatically improved both embryo attachment and outgrowths. Moreover, the expression of miR-100, miR-132, and the mRNA transcripts of BCL2, zinc finger E-box-binding homeobox 1, β-catenin, interferon-γ, protein tyrosine phosphatase non-receptor type 1, PERV, and cyclin-dependent kinase 2 were all increased in embryos supplemented with Endo-EVs + RI compared to those in the control group. Endo-EVs + RI reduced apoptosis and increased the expression of OCT4 and CDX2 and the cell number of embryonic outgrowths. We examined the individual and combined effects of RI compared to those of the miR-155 mimic and found that RI can alleviate the negative effects of the miR-155 mimic on embryo attachment and outgrowths. EVs can improve embryo attachment and the unwanted effects of the de trop cargo contents (miR-155) can be alleviated through anti-apoptotic molecules such as the ROCK inhibitor.
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Affiliation(s)
- Islam M. Saadeldin
- Laboratory of Theriogenology, College of Veterinary Medicine, Chungnam National University, Daejeon 34134, Korea
- Research Institute of Veterinary Medicine, Chungnam National University, Daejeon 34134, Korea
| | - Bereket Molla Tanga
- Laboratory of Theriogenology, College of Veterinary Medicine, Chungnam National University, Daejeon 34134, Korea
| | - Seonggyu Bang
- Laboratory of Theriogenology, College of Veterinary Medicine, Chungnam National University, Daejeon 34134, Korea
| | - Chaerim Seo
- Laboratory of Theriogenology, College of Veterinary Medicine, Chungnam National University, Daejeon 34134, Korea
| | | | - Sung Ho Yun
- Korea Basic Science Institute (KBSI), Ochang 28119, Korea
| | - Seung Il Kim
- Korea Basic Science Institute (KBSI), Ochang 28119, Korea
| | - Sanghoon Lee
- Laboratory of Theriogenology, College of Veterinary Medicine, Chungnam National University, Daejeon 34134, Korea
| | - Jongki Cho
- Laboratory of Theriogenology, College of Veterinary Medicine, Chungnam National University, Daejeon 34134, Korea
- Correspondence: ; Tel.: +82-42-821-6788
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Wang B, Wang Y, Tan Y, Guo J, Chen H, Wu PY, Wang X, Zhang H. Assessment of Fasudil on Contrast-Associated Acute Kidney Injury Using Multiparametric Renal MRI. Front Pharmacol 2022; 13:905547. [PMID: 35784704 PMCID: PMC9242620 DOI: 10.3389/fphar.2022.905547] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/27/2022] [Accepted: 05/31/2022] [Indexed: 11/25/2022] Open
Abstract
Aims: To evaluate the utility of fasudil in a rat model of contrast-associated acute kidney injury (CA-AKI) and explore its underlying mechanism through multiparametric renal magnetic resonance imaging (mpMRI). Methods: Experimental rats (n = 72) were grouped as follows: controls (n = 24), CA-AKI (n = 24), or CA-AKI + Fasudil (n = 24). All animals underwent two mpMRI studies (arterial spin labeling, T1 and T2 mapping) at baseline and post iopromide/fasudil injection (Days 1, 3, 7, and 13 respectively). Relative change in renal blood flow (ΔRBF), T1 (ΔT1) and T2 (ΔT2) values were assessed at specified time points. Serum levels of cystatin C (CysC) and interleukin-1β (IL-1β), and urinary neutrophil gelatinase-associated lipocalin (NGAL) concentrations were tested as laboratory biomarkers, in addition to examining renal histology and expression levels of various proteins (Rho-kinase [ROCK], α-smooth muscle actin [α-SMA]), hypoxia-inducible factor-1α (HIF-1α), and transforming growth factor-β1 (TGF-β1) that regulate renal fibrosis and hypoxia. Results: Compared with the control group, serum levels of CysC and IL-1β, and urinary NGAL concentrations were clearly increased from Day 1 to Day 13 in the CA-AKI group (all p < 0.05). There were significant reductions in ΔT2 values on Days 1 and 3, and ΔT1 reductions were significantly more pronounced at all time points (Days 1–13) in the CA-AKI + Fasudil group (vs. CA-AKI) (all p < 0.05). Fasudil treatment lowered expression levels of ROCK-1, and p-MYPT1/MYPT1 proteins induced by iopromide, decreasing TGF-β1 expression and suppressing both extracellular matrix accumulation and α-SMA expression relative to untreated status (all p < 0.05). Fasudil also enhanced PHD2 transcription and inhibition of HIF-1α expression after CA-AKI. Conclusions: In the context of CA-AKI, fasudil appears to reduce renal hypoxia, fibrosis, and dysfunction by activating (Rho/ROCK) or inhibiting (TGF-β1, HIF-1α) certain signaling pathways and reducing α-SMA expression. Multiparametric MRI may be a viable noninvasive tool for monitoring CA-AKI pathophysiology during fasudil therapy.
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Affiliation(s)
- Bin Wang
- Department of Medical Imaging, First Hospital of Shanxi Medical University, Taiyuan, China
- Department of Medical Imaging, Shanxi Medical University, Taiyuan, China
| | - Yongfang Wang
- Department of Medical Imaging, First Hospital of Shanxi Medical University, Taiyuan, China
| | - Yan Tan
- Department of Medical Imaging, First Hospital of Shanxi Medical University, Taiyuan, China
| | - Jinxia Guo
- GE Healthcare MR Research China, Beijing, China
| | - Haoyuan Chen
- Department of Medical Imaging, Shanxi Medical University, Taiyuan, China
| | - Pu-Yeh Wu
- GE Healthcare MR Research China, Beijing, China
| | - Xiaochun Wang
- Department of Medical Imaging, First Hospital of Shanxi Medical University, Taiyuan, China
- *Correspondence: Xiaochun Wang, ; Hui Zhang,
| | - Hui Zhang
- Department of Medical Imaging, First Hospital of Shanxi Medical University, Taiyuan, China
- *Correspondence: Xiaochun Wang, ; Hui Zhang,
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RhoA Signaling in Neurodegenerative Diseases. Cells 2022; 11:cells11091520. [PMID: 35563826 PMCID: PMC9103838 DOI: 10.3390/cells11091520] [Citation(s) in RCA: 59] [Impact Index Per Article: 19.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/05/2022] [Revised: 04/25/2022] [Accepted: 04/29/2022] [Indexed: 02/04/2023] Open
Abstract
Ras homolog gene family member A (RhoA) is a small GTPase of the Rho family involved in regulating multiple signal transduction pathways that influence a diverse range of cellular functions. RhoA and many of its downstream effector proteins are highly expressed in the nervous system, implying an important role for RhoA signaling in neurons and glial cells. Indeed, emerging evidence points toward a role of aberrant RhoA signaling in neurodegenerative diseases such as Parkinson’s disease, Alzheimer’s disease, Huntington’s disease, and amyotrophic lateral sclerosis. In this review, we summarize the current knowledge of RhoA regulation and downstream cellular functions with an emphasis on the role of RhoA signaling in neurodegenerative diseases and the therapeutic potential of RhoA inhibition in neurodegeneration.
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Wang Z, Li X, Huang L, Liu G, Chen Y, Li B, Zhao X, Xie R, Li Y, Fang W. Long Non-coding RNAs (lncRNAs), A New Target in Stroke. Cell Mol Neurobiol 2022; 42:501-519. [PMID: 32865676 PMCID: PMC11441288 DOI: 10.1007/s10571-020-00954-8] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/22/2020] [Accepted: 08/22/2020] [Indexed: 02/07/2023]
Abstract
Stroke has become the most disabling and the second most fatal disease in the world. It has been a top priority to reveal the pathophysiology of stroke at cellular and molecular levels. A large number of long non-coding RNAs (lncRNAs) are identified to be abnormally expressed after stroke. Here, we summarize 35 lncRNAs associated with stroke, and clarify their functions on the prognosis through signal transduction and predictive values as biomarkers. Changes in the expression of these lncRNAs mediate a wide range of pathological processes in stroke, including apoptosis, inflammation, angiogenesis, and autophagy. Based on the exploration of the functions and mechanisms of lncRNAs in stroke, more timely, accurate predictions and more effective, safer treatments for stroke could be developed.
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Affiliation(s)
- Ziyu Wang
- State Key Laboratory of Natural Medicines, School of Basic Medical Sciences and Clinical Pharmacy, China Pharmaceutical University, Nanjing, 210009, People's Republic of China
| | - Xiang Li
- State Key Laboratory of Natural Medicines, School of Basic Medical Sciences and Clinical Pharmacy, China Pharmaceutical University, Nanjing, 210009, People's Republic of China
| | - Liangliang Huang
- State Key Laboratory of Natural Medicines, School of Basic Medical Sciences and Clinical Pharmacy, China Pharmaceutical University, Nanjing, 210009, People's Republic of China
| | - Ge Liu
- State Key Laboratory of Natural Medicines, School of Basic Medical Sciences and Clinical Pharmacy, China Pharmaceutical University, Nanjing, 210009, People's Republic of China
| | - Yan Chen
- State Key Laboratory of Natural Medicines, School of Basic Medical Sciences and Clinical Pharmacy, China Pharmaceutical University, Nanjing, 210009, People's Republic of China
| | - Binbin Li
- State Key Laboratory of Natural Medicines, School of Basic Medical Sciences and Clinical Pharmacy, China Pharmaceutical University, Nanjing, 210009, People's Republic of China
| | - Xueyan Zhao
- State Key Laboratory of Natural Medicines, School of Basic Medical Sciences and Clinical Pharmacy, China Pharmaceutical University, Nanjing, 210009, People's Republic of China
| | - Rong Xie
- Department of Neurosurgery, Huashan Hospital, Fudan University, Shanghai, 200040, People's Republic of China
| | - Yunman Li
- State Key Laboratory of Natural Medicines, School of Basic Medical Sciences and Clinical Pharmacy, China Pharmaceutical University, Nanjing, 210009, People's Republic of China.
| | - Weirong Fang
- State Key Laboratory of Natural Medicines, School of Basic Medical Sciences and Clinical Pharmacy, China Pharmaceutical University, Nanjing, 210009, People's Republic of China.
- Department of Physiology, School of Basic Medicine and Clinical Pharmacy, China Pharmaceutical University, Tongjiaxiang 24, Mailbox 207, Nanjing, 210009, Jiangsu, People's Republic of China.
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The Role of PKC and HIF-1 and the Effect of Traditional Chinese Medicinal Compounds on Cerebral Ischemia-Reperfusion Injury. EVIDENCE-BASED COMPLEMENTARY AND ALTERNATIVE MEDICINE 2022; 2022:1835898. [PMID: 35265143 PMCID: PMC8898791 DOI: 10.1155/2022/1835898] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 11/03/2021] [Revised: 01/26/2022] [Accepted: 02/07/2022] [Indexed: 11/25/2022]
Abstract
Neuronal death occurs during cerebral ischemia. However, when hemoperfusion and oxygen supply are resumed to the ischemic focus of the brain tissue, the brain tissue damage is further aggravated, resulting in cerebral ischemia-reperfusion injury (CIRI) to the patients. Protein kinase C (PKC) plays an important role in CIRI. Through the IP3/DAG/Ca2+ signaling pathway, it promotes the influx of calcium ions in neurons and causes calcium overload, which aggravates the damage. At the same time, when brain cells are hypoxic, hypoxia-inducible factor-1 (HIF-1) is expressed, which regulates the expression of Bcl-2 and Bax through the PI3K/Akt signaling pathway and reduces nerve cell injury. It also fights hypoxic-ischemic injury by increasing the production of vascular endothelial growth factor (VEGF) to promote blood vessel formation. The PKC and HIF-1 signaling pathways are also linked to CIRI. HIF-1 activates the PKC and ERK pathways via the upregulation of VEGF, leading to increased Cx43 phosphorylation and dysfunction and aggravating CIRI. Existing studies have shown that certain traditional Chinese medicine (TCM) compounds regulate the PKC and HIF-1 signaling pathways and alleviate CIRI. These compounds downregulate the PKC and the activity of the PKC-related signaling pathways to alleviate CIRI. They can also promote the expression of HIF-1, increase the content of VEGF in ischemic tissues to promote the generation of blood vessels, and improve microcirculation. TCM compounds can inhibit the cascade of reactions underlying disease occurrence and development by targeting multiple components using different herbal formulations to improve the structural and material changes in the brain cells, which alleviate CIRI and protect the brain tissue. This study briefly describes the role of PKC and HIF-1, their relationship in CIRI, and the effect of TCM on them.
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Shi J, Wei L. Rho Kinases in Embryonic Development and Stem Cell Research. Arch Immunol Ther Exp (Warsz) 2022; 70:4. [PMID: 35043239 PMCID: PMC8766376 DOI: 10.1007/s00005-022-00642-z] [Citation(s) in RCA: 9] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/14/2021] [Accepted: 12/14/2021] [Indexed: 12/12/2022]
Abstract
The Rho-associated coiled-coil containing kinases (ROCKs or Rho kinases) belong to the AGC (PKA/PKG/PKC) family of serine/threonine kinases and are major downstream effectors of small GTPase RhoA, a key regulator of actin-cytoskeleton reorganization. The ROCK family contains two members, ROCK1 and ROCK2, which share 65% overall identity and 92% identity in kinase domain. ROCK1 and ROCK2 were assumed to be functionally redundant, based largely on their major common activators, their high degree kinase domain homology, and study results from overexpression with kinase constructs or chemical inhibitors. ROCK signaling research has expanded to all areas of biology and medicine since its discovery in 1996. The rapid advance is befitting ROCK’s versatile functions in modulating various cell behavior, such as contraction, adhesion, migration, proliferation, polarity, cytokinesis, and differentiation. The rapid advance is noticeably driven by an extensive linking with clinical medicine, including cardiovascular abnormalities, aberrant immune responsive, and cancer development and metastasis. The rapid advance during the past decade is further powered by novel biotechnologies including CRISPR-Cas and single cell omics. Current consensus, derived mainly from gene targeting and RNA interference approaches, is that the two ROCK isoforms have overlapping and distinct cellular, physiological and pathophysiology roles. In this review, we present an overview of the milestone discoveries in ROCK research. We then focus on the current understanding of ROCK signaling in embryonic development, current research status using knockout and knockin mouse models, and stem cell research.
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Affiliation(s)
- Jianjian Shi
- Herman B Wells Center for Pediatric Research, Department of Pediatrics, School of Medicine, Indiana University, 1044 West Walnut Street, R4-370, Indianapolis, IN, 46202-5225, USA.
| | - Lei Wei
- Herman B Wells Center for Pediatric Research, Department of Pediatrics, School of Medicine, Indiana University, 1044 West Walnut Street, R4-370, Indianapolis, IN, 46202-5225, USA.
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30
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Wei L, Shi J. Insight Into Rho Kinase Isoforms in Obesity and Energy Homeostasis. Front Endocrinol (Lausanne) 2022; 13:886534. [PMID: 35769086 PMCID: PMC9234286 DOI: 10.3389/fendo.2022.886534] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/28/2022] [Accepted: 05/06/2022] [Indexed: 11/13/2022] Open
Abstract
Obesity and associated complications increasingly jeopardize global health and contribute to the rapidly rising prevalence of type 2 diabetes mellitus and obesity-related diseases. Developing novel methods for the prevention and treatment of excess body adipose tissue expansion can make a significant contribution to public health. Rho kinase is a Rho-associated coiled-coil-containing protein kinase (Rho kinase or ROCK). The ROCK family including ROCK1 and ROCK2 has recently emerged as a potential therapeutic target for the treatment of metabolic disorders. Up-regulated ROCK activity has been involved in the pathogenesis of all aspects of metabolic syndrome including obesity, insulin resistance, dyslipidemia and hypertension. The RhoA/ROCK-mediated actin cytoskeleton dynamics have been implicated in both white and beige adipogenesis. Studies using ROCK pan-inhibitors in animal models of obesity, diabetes, and associated complications have demonstrated beneficial outcomes. Studies via genetically modified animal models further established isoform-specific roles of ROCK in the pathogenesis of metabolic disorders including obesity. However, most reported studies have been focused on ROCK1 activity during the past decade. Due to the progress in developing ROCK2-selective inhibitors in recent years, a growing body of evidence indicates more attention should be devoted towards understanding ROCK2 isoform function in metabolism. Hence, studying individual ROCK isoforms to reveal their specific roles and principal mechanisms in white and beige adipogenesis, insulin sensitivity, energy balancing regulation, and obesity development will facilitate significant breakthroughs for systemic treatment with isoform-selective inhibitors. In this review, we give an overview of ROCK functions in the pathogenesis of obesity and insulin resistance with a particular focus on the current understanding of ROCK isoform signaling in white and beige adipogenesis, obesity and thermogenesis in adipose tissue and other major metabolic organs involved in energy homeostasis regulation.
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Affiliation(s)
- Lei Wei
- *Correspondence: Lei Wei, ; Jianjian Shi,
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31
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Mei Y, Wu Y, Ma L, Zhang H, Li L, Wang F. Overexpression of ROCK1 promotes cancer cell proliferation and is associated with poor prognosis in human urothelial bladder cancer. Mamm Genome 2021; 32:466-475. [PMID: 34322718 DOI: 10.1007/s00335-021-09896-y] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/12/2021] [Accepted: 07/14/2021] [Indexed: 11/27/2022]
Abstract
Rho-associated protein kinase 1 (ROCK1) has been reported to be overexpressed in many types of tumors, but its role in urothelial bladder cancer is poorly understood. The study aims to investigate the role of ROCK1 in urothelial bladder cancer and explored the underlying mechanism. Protein and mRNA levels of ROCK1 were detected in 64 urothelial bladder cancer patients using western blot, immunohistochemistry and qRT-PCR. Relationships between ROCK1 expression and clinicopathological factors and survival rate were analyzed. ROCK1 was silenced by shRNA in multiple urothelial bladder cancer cells to explore its function and underlying mechanism. ROCK1 expression was significantly increased in tumor tissues compared with the paired adjacent healthy tissues of patients. Higher ROCK1 expression of tumor tissues positively correlated with poor prognosis of patients (p = 0.0435). ROCK1 silence significantly inhibited cell proliferation and colony formation, and enhanced activation of apoptotic pathway in urothelial bladder cancer cells. High ROCK1 expression predicts poor prognosis of urothelial bladder cancer. ROCK1 silence inhibit cell proliferation and promote apoptosis, which may be of value as a therapeutic target for bladder cancer treatment.
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Affiliation(s)
- Yanhui Mei
- Department of Urology, Binzhou Medical University Hospital, No 661 Huanghe 2nd Road, Binzhou, 256603, Shandong, China
| | - Yuhai Wu
- Department of Urology, Binzhou Medical University Hospital, No 661 Huanghe 2nd Road, Binzhou, 256603, Shandong, China
| | - Longbo Ma
- Department of Oncology, Binzhou Medical University Hospital, No 661 Huanghe 2nd Road, Binzhou, 256603, Shandong, China
| | - Hongli Zhang
- Department of Oncology, Guizhou Provincial People's Hospital, No 83 Zhongshan East Road, Guiyang, 550002, Guizhou, China
| | - Lei Li
- Department of Oncology, Binzhou Medical University Hospital, No 661 Huanghe 2nd Road, Binzhou, 256603, Shandong, China
| | - Feng Wang
- Department of Oncology, Binzhou Medical University Hospital, No 661 Huanghe 2nd Road, Binzhou, 256603, Shandong, China.
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Kim S, Kim SA, Han J, Kim IS. Rho-Kinase as a Target for Cancer Therapy and Its Immunotherapeutic Potential. Int J Mol Sci 2021; 22:ijms222312916. [PMID: 34884721 PMCID: PMC8657458 DOI: 10.3390/ijms222312916] [Citation(s) in RCA: 19] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/20/2021] [Revised: 11/24/2021] [Accepted: 11/24/2021] [Indexed: 02/07/2023] Open
Abstract
Cancer immunotherapy is fast rising as a prominent new pillar of cancer treatment, harnessing the immune system to fight against numerous types of cancer. Rho-kinase (ROCK) pathway is involved in diverse cellular activities, and is therefore the target of interest in various diseases at the cellular level including cancer. Indeed, ROCK is well-known for its involvement in the tumor cell and tumor microenvironment, especially in its ability to enhance tumor cell progression, migration, metastasis, and extracellular matrix remodeling. Importantly, ROCK is also considered to be a novel and effective modulator of immune cells, although further studies are needed. In this review article, we describe the various activities of ROCK and its potential to be utilized in cancer treatment, particularly in cancer immunotherapy, by shining a light on its activities in the immune system.
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Affiliation(s)
- Seohyun Kim
- KU-KIST Graduate School of Converging Science and Technology, Korea University, Seoul 02841, Korea; (S.K.); (S.A.K.); (J.H.)
- Center for Theragnosis, Biomedical Research Institute, Korea Institute of Science and Technology (KIST), Seoul 02792, Korea
| | - Seong A. Kim
- KU-KIST Graduate School of Converging Science and Technology, Korea University, Seoul 02841, Korea; (S.K.); (S.A.K.); (J.H.)
- Center for Theragnosis, Biomedical Research Institute, Korea Institute of Science and Technology (KIST), Seoul 02792, Korea
| | - Jihoon Han
- KU-KIST Graduate School of Converging Science and Technology, Korea University, Seoul 02841, Korea; (S.K.); (S.A.K.); (J.H.)
- Center for Theragnosis, Biomedical Research Institute, Korea Institute of Science and Technology (KIST), Seoul 02792, Korea
| | - In-San Kim
- KU-KIST Graduate School of Converging Science and Technology, Korea University, Seoul 02841, Korea; (S.K.); (S.A.K.); (J.H.)
- Center for Theragnosis, Biomedical Research Institute, Korea Institute of Science and Technology (KIST), Seoul 02792, Korea
- Correspondence:
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Kimura T, Horikoshi Y, Kuriyagawa C, Niiyama Y. Rho/ROCK Pathway and Noncoding RNAs: Implications in Ischemic Stroke and Spinal Cord Injury. Int J Mol Sci 2021; 22:ijms222111573. [PMID: 34769004 PMCID: PMC8584200 DOI: 10.3390/ijms222111573] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/01/2021] [Revised: 10/21/2021] [Accepted: 10/24/2021] [Indexed: 01/18/2023] Open
Abstract
Ischemic strokes (IS) and spinal cord injuries (SCI) are major causes of disability. RhoA is a small GTPase protein that activates a downstream effector, ROCK. The up-regulation of the RhoA/ROCK pathway contributes to neuronal apoptosis, neuroinflammation, blood-brain barrier dysfunction, astrogliosis, and axon growth inhibition in IS and SCI. Noncoding RNAs (ncRNAs), such as microRNAs (miRNAs) and long noncoding RNAs (lncRNAs), were previously considered to be non-functional. However, they have attracted much attention because they play an essential role in regulating gene expression in physiological and pathological conditions. There is growing evidence that ROCK inhibitors, such as fasudil and VX-210, can reduce injury in IS and SCI in animal models and clinical trials. Recently, it has been reported that miRNAs are decreased in IS and SCI, while lncRNAs are increased. Inhibiting the Rho/ROCK pathway with miRNAs alleviates apoptosis, neuroinflammation, oxidative stress, and axon growth inhibition in IS and SCI. Further studies are required to explore the significance of ncRNAs in IS and SCI and to establish new strategies for preventing and treating these devastating diseases.
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Affiliation(s)
- Tetsu Kimura
- Correspondence: ; Tel.: +81-18-884-6175; Fax: +81-18-884-6448
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Peaslee C, Esteva-Font C, Su T, Munoz-Howell A, Duwaerts CC, Liu Z, Rao S, Liu K, Medina M, Sneddon JB, Maher JJ, Mattis AN. Doxycycline Significantly Enhances Induction of Induced Pluripotent Stem Cells to Endoderm by Enhancing Survival Through Protein Kinase B Phosphorylation. Hepatology 2021; 74:2102-2117. [PMID: 33982322 PMCID: PMC8544023 DOI: 10.1002/hep.31898] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/19/2020] [Revised: 03/15/2021] [Accepted: 04/22/2021] [Indexed: 01/10/2023]
Abstract
BACKGROUND AND AIMS Induced pluripotent stem cells (iPSCs) provide an important tool for the generation of patient-derived cells, including hepatocyte-like cells, by developmental cues through an endoderm intermediate. However, most iPSC lines fail to differentiate into endoderm, with induction resulting in apoptosis. APPROACH AND RESULTS To address this issue, we built upon published methods to develop an improved protocol. We discovered that doxycycline dramatically enhances the efficiency of iPSCs to endoderm differentiation by inhibiting apoptosis and promoting proliferation through the protein kinase B pathway. We tested this protocol in >70 iPSC lines, 90% of which consistently formed complete sheets of endoderm. Endoderm generated by our method achieves similar transcriptomic profiles, expression of endoderm protein markers, and the ability to be further differentiated to downstream lineages. CONCLUSIONS Furthermore, this method achieves a 4-fold increase in endoderm cell number and will accelerate studies of human diseases in vitro and facilitate the expansion of iPSC-derived cells for transplantation studies.
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Affiliation(s)
- Caitlin Peaslee
- Department of Pathology, University of California San Francisco, San Francisco, CA
| | - Cristina Esteva-Font
- Department of Pathology, University of California San Francisco, San Francisco, CA
| | - Tao Su
- Department of Pathology, University of California San Francisco, San Francisco, CA
| | - Antonio Munoz-Howell
- Children’s Hospital Oakland Research Institute, University of California San Francisco, San Francisco, CA
| | - Caroline C. Duwaerts
- Department of Medicine, University of California San Francisco, San Francisco, CA
- Liver Center, University of California San Francisco, San Francisco, CA
| | - Zhe Liu
- Department of Cell and Tissue Biology, University of California San Francisco, San Francisco, CA
- Diabetes Center, University of California San Francisco, San Francisco, CA
- Eli and Edythe Broad Center for Regeneration Medicine and Stem Cell Research, University of California San Francisco, San Francisco, CA
| | - Sneha Rao
- Department of Cell and Tissue Biology, University of California San Francisco, San Francisco, CA
- Diabetes Center, University of California San Francisco, San Francisco, CA
- Eli and Edythe Broad Center for Regeneration Medicine and Stem Cell Research, University of California San Francisco, San Francisco, CA
| | - Ke Liu
- Children’s Hospital Oakland Research Institute, University of California San Francisco, San Francisco, CA
| | - Marisa Medina
- Children’s Hospital Oakland Research Institute, University of California San Francisco, San Francisco, CA
- Liver Center, University of California San Francisco, San Francisco, CA
| | - Julie B. Sneddon
- Department of Cell and Tissue Biology, University of California San Francisco, San Francisco, CA
- Diabetes Center, University of California San Francisco, San Francisco, CA
- Eli and Edythe Broad Center for Regeneration Medicine and Stem Cell Research, University of California San Francisco, San Francisco, CA
- Department of Anatomy, University of California San Francisco, San Francisco, CA
| | - Jacquelyn J. Maher
- Department of Medicine, University of California San Francisco, San Francisco, CA
- Liver Center, University of California San Francisco, San Francisco, CA
- Eli and Edythe Broad Center for Regeneration Medicine and Stem Cell Research, University of California San Francisco, San Francisco, CA
| | - Aras N. Mattis
- Department of Pathology, University of California San Francisco, San Francisco, CA
- Liver Center, University of California San Francisco, San Francisco, CA
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Bai L, Kee HJ, Han X, Zhao T, Kee SJ, Jeong MH. Protocatechuic acid attenuates isoproterenol-induced cardiac hypertrophy via downregulation of ROCK1-Sp1-PKCγ axis. Sci Rep 2021; 11:17343. [PMID: 34462460 PMCID: PMC8405624 DOI: 10.1038/s41598-021-96761-2] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/06/2021] [Accepted: 08/12/2021] [Indexed: 12/25/2022] Open
Abstract
Cardiac hypertrophy is an adaptive response of the myocardium to pressure overload or adrenergic agonists. Here, we investigated the protective effects and the regulatory mechanism of protocatechuic acid, a phenolic compound, using a mouse model of isoproterenol-induced cardiac hypertrophy. Our results demonstrated that protocatechuic acid treatment significantly downregulated the expression of cardiac hypertrophic markers (Nppa, Nppb, and Myh7), cardiomyocyte size, heart weight to body weight ratio, cross-sectional area, and thickness of left ventricular septum and posterior wall. This treatment also reduced the expression of isoproterenol-induced ROCK1, Sp1, and PKCγ both in vivo and in vitro. To investigate the mechanism, we performed knockdown and overexpression experiments. The knockdown of ROCK1, Sp1, or PKCγ decreased the isoproterenol-induced cell area and the expression of hypertrophic markers, while the overexpression of Sp1 or PKCγ increased the levels of hypertrophic markers. Protocatechuic acid treatment reversed these effects. Interestingly, the overexpression of Sp1 increased cell area and induced PKCγ expression. Furthermore, experiments using transcription inhibitor actinomycin D showed that ROCK1 and Sp1 suppression by protocatechuic acid was not regulated at the transcriptional level. Our results indicate that protocatechuic acid acts via the ROCK1/Sp1/PKCγ axis and therefore has promising therapeutic potential as a treatment for cardiac hypertrophy.
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Affiliation(s)
- Liyan Bai
- Heart Research Center, Chonnam National University Hospital, 42 Jebong-ro, Dong-gu, Gwangju, 61469, Republic of Korea
- Hypertension Heart Failure Research Center, Chonnam National University Hospital, Gwangju, 61469, Republic of Korea
| | - Hae Jin Kee
- Heart Research Center, Chonnam National University Hospital, 42 Jebong-ro, Dong-gu, Gwangju, 61469, Republic of Korea.
- Hypertension Heart Failure Research Center, Chonnam National University Hospital, Gwangju, 61469, Republic of Korea.
| | - Xiongyi Han
- Heart Research Center, Chonnam National University Hospital, 42 Jebong-ro, Dong-gu, Gwangju, 61469, Republic of Korea
- Hypertension Heart Failure Research Center, Chonnam National University Hospital, Gwangju, 61469, Republic of Korea
| | - Tingwei Zhao
- Heart Research Center, Chonnam National University Hospital, 42 Jebong-ro, Dong-gu, Gwangju, 61469, Republic of Korea
- Hypertension Heart Failure Research Center, Chonnam National University Hospital, Gwangju, 61469, Republic of Korea
| | - Seung-Jung Kee
- Department of Laboratory Medicine, Chonnam National University, Medical School and Hospital, Gwangju, 61469, Republic of Korea
| | - Myung Ho Jeong
- Heart Research Center, Chonnam National University Hospital, 42 Jebong-ro, Dong-gu, Gwangju, 61469, Republic of Korea.
- Hypertension Heart Failure Research Center, Chonnam National University Hospital, Gwangju, 61469, Republic of Korea.
- Department of Cardiology, Chonnam National University Medical School, Gwangju, 61469, Republic of Korea.
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Takechi H, Hakeda-Suzuki S, Nitta Y, Ishiwata Y, Iwanaga R, Sato M, Sugie A, Suzuki T. Glial insulin regulates cooperative or antagonistic Golden goal/Flamingo interactions during photoreceptor axon guidance. eLife 2021; 10:66718. [PMID: 33666170 PMCID: PMC7987344 DOI: 10.7554/elife.66718] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/20/2021] [Accepted: 03/02/2021] [Indexed: 11/29/2022] Open
Abstract
Transmembrane protein Golden goal (Gogo) interacts with atypical cadherin Flamingo (Fmi) to direct R8 photoreceptor axons in the Drosophila visual system. However, the precise mechanisms underlying Gogo regulation during columnar- and layer-specific R8 axon targeting are unknown. Our studies demonstrated that the insulin secreted from surface and cortex glia switches the phosphorylation status of Gogo, thereby regulating its two distinct functions. Non-phosphorylated Gogo mediates the initial recognition of the glial protrusion in the center of the medulla column, whereas phosphorylated Gogo suppresses radial filopodia extension by counteracting Flamingo to maintain a one axon-to-one column ratio. Later, Gogo expression ceases during the midpupal stage, thus allowing R8 filopodia to extend vertically into the M3 layer. These results demonstrate that the long- and short-range signaling between the glia and R8 axon growth cones regulates growth cone dynamics in a stepwise manner, and thus shapes the entire organization of the visual system.
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Affiliation(s)
- Hiroki Takechi
- Graduate School of Life Science and Technology, Tokyo Institute of Technology, Yokohama, Japan
| | - Satoko Hakeda-Suzuki
- Graduate School of Life Science and Technology, Tokyo Institute of Technology, Yokohama, Japan
| | - Yohei Nitta
- Center for Transdisciplinary Research, Niigata University, Niigata, Japan.,Brain Research Institute, Niigata University, Niigata, Japan
| | - Yuichi Ishiwata
- Graduate School of Life Science and Technology, Tokyo Institute of Technology, Yokohama, Japan
| | - Riku Iwanaga
- Graduate School of Life Science and Technology, Tokyo Institute of Technology, Yokohama, Japan
| | - Makoto Sato
- Mathematical Neuroscience Unit, Institute for Frontier Science Initiative, Kanazawa University, Kanazawa, Japan.,Laboratory of Developmental Neurobiology, Graduate School of Medical Sciences, Kanazawa University, Kanazawa, Japan
| | - Atsushi Sugie
- Center for Transdisciplinary Research, Niigata University, Niigata, Japan.,Brain Research Institute, Niigata University, Niigata, Japan
| | - Takashi Suzuki
- Graduate School of Life Science and Technology, Tokyo Institute of Technology, Yokohama, Japan
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Saadeldin IM, Tukur HA, Aljumaah RS, Sindi RA. Rocking the Boat: The Decisive Roles of Rho Kinases During Oocyte, Blastocyst, and Stem Cell Development. Front Cell Dev Biol 2021; 8:616762. [PMID: 33505968 PMCID: PMC7829335 DOI: 10.3389/fcell.2020.616762] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/13/2020] [Accepted: 12/07/2020] [Indexed: 01/09/2023] Open
Abstract
The rho-associated coiled-coil-containing proteins (ROCKs or rho kinase) are effectors of the small rho-GTPase rhoA, which acts as a signaling molecule to regulate a variety of cellular processes, including cell proliferation, adhesion, polarity, cytokinesis, and survival. Owing to the multifunctionality of these kinases, an increasing number of studies focus on understanding the pleiotropic effects of the ROCK signaling pathway in the coordination and control of growth (proliferation, initiation, and progression), development (morphology and differentiation), and survival in many cell types. There is growing evidence that ROCKs actively phosphorylate several actin-binding proteins and intermediate filament proteins during oocyte cytokinesis, the preimplantation embryos as well as the stem cell development and differentiation. In this review, we focus on the participation of ROCK proteins in oocyte maturation, blastocyst formation, and stem cell development with a special focus on the selective targeting of ROCK isoforms, ROCK1, and ROCK2. The selective switching of cell fate through ROCK inhibition would provide a novel paradigm for in vitro oocyte maturation, experimental embryology, and clinical applications.
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Affiliation(s)
- Islam M Saadeldin
- Department of Animal Production, College of Food and Agricultural Sciences, King Saud University, Riyadh, Saudi Arabia.,Department of Comparative Medicine, King Faisal Specialist Hospital & Research Centre, Riyadh, Saudi Arabia
| | - Hammed A Tukur
- Department of Animal Production, College of Food and Agricultural Sciences, King Saud University, Riyadh, Saudi Arabia
| | - Riyadh S Aljumaah
- Department of Animal Production, College of Food and Agricultural Sciences, King Saud University, Riyadh, Saudi Arabia
| | - Ramya A Sindi
- Department of Laboratory Medicine, Faculty of Applied Medical Sciences, Umm Al-Qura University, Makkah, Saudi Arabia
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Abstract
Since the discovery that extracellular vesicles (EVs) mediate intercellular communication, there is an exponential increase in the interest on EVs, especially in pathological settings. EVs are membranous vesicles that are secreted by various cell types and the release of EVs is conserved in every prokaryotic and eukaryotic organism tested to date. These vesicles were initially thought to be garbage disposal vehicles and subsequent studies over the past 4 decades have attributed several functional roles to EVs, some of which are critical for homeostasis. The molecular cargo of nucleic acids, proteins, lipids and metabolites packaged in EVs often mirror the host cells phenotypic status. EVs can be taken up by recipient cells and upon uptake, EVs through its molecular cargo, can induce a cascade of signal transduction events in recipient cells. EVs are categorised into several subtypes depending on their biogenesis and secretion. Due to several subtypes, differing sizes within a subtype and varying cargo, EVs are heterogenous in nature and the biophysical and biochemical properties of EVs often overlap between EV subtypes. Hence, it is important to be cautious when selecting the method of EV isolation and characterisation. This chapter provides a brief introduction to EVs and their subtypes.
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Affiliation(s)
- Pamali Fonseka
- Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, Victoria, Australia.
| | - Akbar L Marzan
- Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, Victoria, Australia
| | - Suresh Mathivanan
- Department of Biochemistry and Genetics, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, Victoria, Australia
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Engin A, Engin AB. N-Methyl-D-Aspartate Receptor Signaling-Protein Kinases Crosstalk in Cerebral Ischemia. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2021; 1275:259-283. [PMID: 33539019 DOI: 10.1007/978-3-030-49844-3_10] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 03/24/2023]
Abstract
Although stroke is very often the cause of death worldwide, the burden of ischemic and hemorrhagic stroke varies between regions and over time regarding differences in prognosis, prevalence of risk factors, and treatment strategies. Excitotoxicity, oxidative stress, dysfunction of the blood-brain barrier, neuroinflammation, and lysosomal membrane permeabilization, sequentially lead to the progressive death of neurons. In this process, protein kinases-related checkpoints tightly regulate N-methyl-D-aspartate (NMDA) receptor signaling pathways. One of the major hallmarks of cerebral ischemia is excitotoxicity, characterized by overactivation of glutamate receptors leading to intracellular Ca2+ overload and ultimately neuronal death. Thus, reduced expression of postsynaptic density-95 protein and increased protein S-nitrosylation in neurons is responsible for neuronal vulnerability in cerebral ischemia. In this chapter death-associated protein kinases, cyclin-dependent kinase 5, endoplasmic reticulum stress-induced protein kinases, hyperhomocysteinemia-related NMDA receptor overactivation, ephrin-B-dependent amplification of NMDA-evoked neuronal excitotoxicity and lysosomocentric hypothesis have been discussed.Consequently, ample evidences have demonstrated that enhancing extrasynaptic NMDA receptor activity triggers cell death after stroke. In this context, considering the dual roles of NMDA receptors in both promoting neuronal survival and mediating neuronal damage, selective augmentation of NR2A-containing NMDA receptor activation in the presence of NR2B antagonist may constitute a promising therapy for stroke.
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Affiliation(s)
- Atilla Engin
- Department of General Surgery, Faculty of Medicine, Gazi University, Ankara, Turkey
| | - Ayse Basak Engin
- Department of Toxicology, Faculty of Pharmacy, Gazi University, Ankara, Turkey.
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40
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Generation and biobanking of patient-derived glioblastoma organoids and their application in CAR T cell testing. Nat Protoc 2020; 15:4000-4033. [PMID: 33169003 DOI: 10.1038/s41596-020-0402-9] [Citation(s) in RCA: 105] [Impact Index Per Article: 21.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/17/2020] [Accepted: 08/20/2020] [Indexed: 12/21/2022]
Abstract
Glioblastoma tumors exhibit extensive inter- and intratumoral heterogeneity, which has contributed to the poor outcomes of numerous clinical trials and continues to complicate the development of effective therapeutic strategies. Most in vitro models do not preserve the cellular and mutational diversity of parent tumors and often require a lengthy generation time with variable efficiency. Here, we describe detailed procedures for generating glioblastoma organoids (GBOs) from surgically resected patient tumor tissue using a chemically defined medium without cell dissociation. By preserving cell-cell interactions and minimizing clonal selection, GBOs maintain the cellular heterogeneity of parent tumors. We include details of how to passage and cryopreserve GBOs for continued use, biobanking and long-term recovery. In addition, we describe procedures for investigating patient-specific responses to immunotherapies by co-culturing GBOs with chimeric antigen receptor (CAR) T cells. It takes ~2-4 weeks to generate GBOs and 5-7 d to perform CAR T cell co-culture using this protocol. Competence with human cell culture, tissue processing, immunohistology and microscopy is required for optimal results.
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Duess JW, Gosemann JH, Puri P, Thompson J. Teratogenesis in the chick embryo following post-gastrulation exposure to Y-27632 -effect of Y-27632 on embryonic development. Toxicol Appl Pharmacol 2020; 409:115277. [PMID: 33049266 DOI: 10.1016/j.taap.2020.115277] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/13/2020] [Revised: 10/04/2020] [Accepted: 10/07/2020] [Indexed: 01/08/2023]
Abstract
The pyridine derivative Y-27632 inhibits Rho-associated coiled-coil-containing protein kinase (ROCK) signaling, which is involved in numerous developmental processes during embryogenesis, primarily by controlling actin-cytoskeleton assembly and cell contractility. Somite formation requires rearrangement of the cytoskeleton and assists in major morphological mechanisms, including ventral body wall formation. Administration of Y-27632 impairs cytoskeletal arrangements in post-gastrulation chick embryos leading to ventral body wall defects (VBWD) at later stages of development. The aim of this study was to investigate the effect of Y-27632 on somite development in post-gastrulation chick embryos during early embryogenesis. After 60 h incubation, embryos in shell-less culture were treated with Y-27632 or vehicle for controls. Following administration, abnormality rates were assessed. In treatment groups, embryos showed a kinked longitudinal body axis. Western blot confirmed impaired ROCK downstream signaling by decreased expression of phosphorylated cofilin-2. Histology, Lysotracker studies and RT-PCR demonstrated increased cell death in somites, the neural tube and the ectoderm. RT-PCR and Western blot of factors known to be involved during somitogenesis revealed reduced expression in the treatment group compared to controls. We hypothesize that administration of Y-27632 disrupts somite development causing axial kinking and embryo malformation, which may lead to VBWD.
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Affiliation(s)
- Johannes W Duess
- Department of Pediatric Surgery, University of Leipzig, Leipzig, Germany; National Children's Research Centre, Our Lady's Children's Hospital, Crumlin, Dublin 12, Ireland; School of Medicine and Medical Science, University College Dublin, Belfield, Dublin 4, Ireland.
| | - Jan-Hendrik Gosemann
- Department of Pediatric Surgery, University of Leipzig, Leipzig, Germany; National Children's Research Centre, Our Lady's Children's Hospital, Crumlin, Dublin 12, Ireland
| | - Prem Puri
- National Children's Research Centre, Our Lady's Children's Hospital, Crumlin, Dublin 12, Ireland; School of Medicine and Medical Science, University College Dublin, Belfield, Dublin 4, Ireland
| | - Jennifer Thompson
- National Children's Research Centre, Our Lady's Children's Hospital, Crumlin, Dublin 12, Ireland; School of Medicine and Medical Science, University College Dublin, Belfield, Dublin 4, Ireland
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Lee JY, Stevens RP, Kash M, Zhou C, Koloteva A, Renema P, Paudel SS, Stevens T. KD025 Shifts Pulmonary Endothelial Cell Bioenergetics and Decreases Baseline Lung Permeability. Am J Respir Cell Mol Biol 2020; 63:519-530. [PMID: 32628869 PMCID: PMC7528923 DOI: 10.1165/rcmb.2019-0435oc] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/09/2019] [Accepted: 07/06/2020] [Indexed: 12/22/2022] Open
Abstract
KD025 is a ROCK2 inhibitor currently being tested in clinical trials for the treatment of fibrotic lung diseases. The therapeutic effects of KD025 are partly due to its inhibition of profibrotic pathways and fat metabolism. However, whether KD025 affects pulmonary microvascular endothelial cell (PMVEC) function is unknown, despite evidence that alveolar-capillary membrane disruption constitutes major causes of death in fibrotic lung diseases. We hypothesized that KD025 regulates PMVEC metabolism, pH, migration, and survival, a series of interrelated functional characteristics that determine pulmonary barrier integrity. We used PMVECs isolated from Sprague Dawley rats. KD025 dose-dependently decreased lactate production and glucose consumption. The inhibitory effect of KD025 was more potent compared with other metabolic modifiers, including 2-deoxy-glucose, extracellular acidosis, dichloroacetate, and remogliflozin. Interestingly, KD025 increased oxidative phosphorylation, whereas 2-deoxy-glucose did not. KD025 also decreased intracellular pH and induced a compensatory increase in anion exchanger 2. KD025 inhibited PMVEC migration, but fasudil (nonspecific ROCK inhibitor) did not. We tested endothelial permeability in vivo using Evans Blue dye in the bleomycin pulmonary fibrosis model. Baseline permeability was decreased in KD025-treated animals independent of bleomycin treatment. Under hypoxia, KD025 increased PMVEC necrosis as indicated by increased lactate dehydrogenase release and propidium iodide uptake and decreased ATP; it did not affect Annexin V binding. ROCK2 knockdown had no effect on PMVEC metabolism, pH, and migration, but it increased nonapoptotic caspase-3 activity. Together, we report that KD025 promotes oxidative phosphorylation; decreases glycolysis, intracellular pH, and migration; and strengthens pulmonary barrier integrity in a ROCK2-independent manner.
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Affiliation(s)
- Ji Young Lee
- Department of Physiology and Cell Biology
- Department of Internal Medicine
- Division of Pulmonary and Critical Care Medicine
- Center for Lung Biology
- College of Medicine, and
- University of South Alabama, Mobile, Alabama
| | - Reece P. Stevens
- Department of Physiology and Cell Biology
- Center for Lung Biology
- College of Medicine, and
- University of South Alabama, Mobile, Alabama
| | - Mary Kash
- College of Medicine, and
- University of South Alabama, Mobile, Alabama
| | - Chun Zhou
- Department of Physiology and Cell Biology
- Center for Lung Biology
- College of Medicine, and
- University of South Alabama, Mobile, Alabama
| | - Anna Koloteva
- Department of Physiology and Cell Biology
- Center for Lung Biology
- College of Medicine, and
- University of South Alabama, Mobile, Alabama
| | - Phoibe Renema
- Department of Physiology and Cell Biology
- Center for Lung Biology
- College of Medicine, and
- University of South Alabama, Mobile, Alabama
| | - Sunita S. Paudel
- Department of Physiology and Cell Biology
- Center for Lung Biology
- College of Medicine, and
- University of South Alabama, Mobile, Alabama
| | - Troy Stevens
- Department of Physiology and Cell Biology
- Department of Internal Medicine
- Center for Lung Biology
- College of Medicine, and
- University of South Alabama, Mobile, Alabama
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de Sousa GR, Vieira GM, das Chagas PF, Pezuk JA, Brassesco MS. Should we keep rocking? Portraits from targeting Rho kinases in cancer. Pharmacol Res 2020; 160:105093. [PMID: 32726671 DOI: 10.1016/j.phrs.2020.105093] [Citation(s) in RCA: 17] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/30/2020] [Revised: 07/15/2020] [Accepted: 07/19/2020] [Indexed: 12/12/2022]
Abstract
Cancer targeted therapy, either alone or in combination with conventional chemotherapy, could allow the survival of patients with neoplasms currently considered incurable. In recent years, the dysregulation of the Rho-associated coiled-coil kinases (ROCK1 and ROCK2) has been associated with increased metastasis and poorer patient survival in several tumor types, and due to their essential roles in regulating the cytoskeleton, have gained popularity and progressively been researched as targets for the development of novel anti-cancer drugs. Nevertheless, in a pediatric scenario, the influence of both isoforms on prognosis remains a controversial issue. In this review, we summarize the functions of ROCKs, compile their roles in human cancer and their value as prognostic factors in both, adult and pediatric cancer. Moreover, we provide the up-to-date advances on their pharmacological inhibition in pre-clinical models and clinical trials. Alternatively, we highlight and discuss detrimental effects of ROCK inhibition provoked not only by the action on off-targets, but most importantly, by pro-survival effects on cancer stem cells, dormant cells, and circulating tumor cells, along with cell-context or microenvironment-dependent contradictory responses. Together these drawbacks represent a risk for cancer cell dissemination and metastasis after anti-ROCK intervention, a caveat that should concern scientists and clinicians.
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Affiliation(s)
| | | | | | | | - María Sol Brassesco
- Department of Biology, Faculty of Philosophy, Sciences and Letters at Ribeirão Preto, University of São Paulo, Brazil.
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PDPN Is Expressed in Various Types of Canine Tumors and Its Silencing Induces Apoptosis and Cell Cycle Arrest in Canine Malignant Melanoma. Cells 2020; 9:cells9051136. [PMID: 32380790 PMCID: PMC7290317 DOI: 10.3390/cells9051136] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2020] [Revised: 05/02/2020] [Accepted: 05/04/2020] [Indexed: 02/07/2023] Open
Abstract
Podoplanin (PDPN), a small transmembrane mucin-like glycoprotein, is ectopically expressed. It is also known to be linked with several aspects of tumor malignancy in some types of human tumors, including invasion, metastasis, and cancer stemness. However, there are few reports on the expression of dog PDPN (dPDPN) in canine tumors, and the association between dPDPN and tumor malignancy has not been elucidated. We identified that 11 out of 18 types of canine tumors expressed dPDPN. Furthermore, 80% of canine malignant melanoma (MM), squamous cell carcinoma, and meningioma expressed dPDPN. Moreover, the expression density of dPDPN was positively associated with the expression of the Ki67 proliferation marker. The silencing of dPDPN by siRNAs resulted in the suppression of cell migration, invasion, stem cell-like characteristics, and cell viability in canine MM cell lines. The suppression of cell viability was caused by the induction of apoptosis and G2/M phase cell cycle arrest. Overall, this study demonstrates that dPDPN is expressed in various types of canine tumors and that dPDPN silencing suppresses cell viability through apoptosis and cell cycle arrest, thus providing a novel biological role for PDPN in tumor progression.
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Ashrafizadeh M, Javanmardi S, Moradi-Ozarlou M, Mohammadinejad R, Farkhondeh T, Samarghandian S, Garg M. Natural products and phytochemical nanoformulations targeting mitochondria in oncotherapy: an updated review on resveratrol. Biosci Rep 2020; 40:BSR20200257. [PMID: 32163546 PMCID: PMC7133519 DOI: 10.1042/bsr20200257] [Citation(s) in RCA: 30] [Impact Index Per Article: 6.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/04/2020] [Revised: 03/11/2020] [Accepted: 03/12/2020] [Indexed: 12/11/2022] Open
Abstract
Mitochondria are intracellular organelles with two distinct membranes, known as an outer mitochondrial membrane and inner cell membrane. Originally, mitochondria have been derived from bacteria. The main function of mitochondria is the production of ATP. However, this important organelle indirectly protects cells by consuming oxygen in the route of energy generation. It has been found that mitochondria are actively involved in the induction of the intrinsic pathways of apoptosis. So, there have been efforts to sustain mitochondrial homeostasis and inhibit its dysfunction. Notably, due to the potential role of mitochondria in the stimulation of apoptosis, this organelle is a promising target in cancer therapy. Resveratrol is a non-flavonoid polyphenol that exhibits significant pharmacological effects such as antioxidant, anti-diabetic, anti-inflammatory and anti-tumor. The anti-tumor activity of resveratrol may be a consequence of its effect on mitochondria. Multiple studies have investigated the relationship between resveratrol and mitochondria, and it has been demonstrated that resveratrol is able to significantly enhance the concentration of reactive oxygen species, leading to the mitochondrial dysfunction and consequently, apoptosis induction. A number of signaling pathways such as sirtuin and NF-κB may contribute to the mitochondrial-mediated apoptosis by resveratrol. Besides, resveratrol shifts cellular metabolism from glycolysis into mitochondrial respiration to induce cellular death in cancer cells. In the present review, we discuss the possible interactions between resveratrol and mitochondria, and its potential application in cancer therapy.
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Affiliation(s)
- Milad Ashrafizadeh
- Department of Basic Science, Faculty of Veterinary Medicine, University of Tabriz, Tabriz, Iran
| | - Sara Javanmardi
- Department of Clinical Sciences, Faculty of Veterinary Medicine, University of Tabriz, Tabriz, Iran
| | - Masoumeh Moradi-Ozarlou
- Department of Pathobiology, Faculty of Veterinary Medicine, University of Tabriz, Tabriz, Iran
| | - Reza Mohammadinejad
- Neuroscience Research Center, Institute of Neuropharmacology, Kerman University of Medical Sciences, Kerman, Iran
| | - Tahereh Farkhondeh
- Cardiovascular Diseases Research Center, Birjand University of Medical Sciences, Birjand, Iran
| | - Saeed Samarghandian
- Healthy Ageing Research Center, Neyshabur University of Medical Sciences, Neyshabur, Iran
- Department of Basic Medical Sciences, Neyshabur University of Medical Sciences, Neyshabur, Iran
| | - Manoj Garg
- Amity Institute of Molecular Medicine and Stem cell Research (AIMMSCR), Amity University, Noida, Uttar Pradesh 201313, India
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Effects of Short-Term Inhibition of Rho Kinase on Dromedary Camel Oocyte In Vitro Maturation. Animals (Basel) 2020; 10:ani10050750. [PMID: 32344840 PMCID: PMC7277376 DOI: 10.3390/ani10050750] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/30/2020] [Revised: 04/19/2020] [Accepted: 04/22/2020] [Indexed: 12/26/2022] Open
Abstract
Simple Summary Our results revealed, for the first time, that short-term inhibition of Rho-associated protein kinases (ROCK) for 4 h prior to in vitro maturation (IVM) in a biphasic IVM approach improved oocyte nuclear maturation, producing more MII oocyte, through modulating the expression of cytokinesis- and antiapoptosis-related mRNA transcripts. This positive result suggests ROCK inhibitor as a potential candidate molecule to exploit in the control of oocyte meiotic maturation. Abstract This is the first report on a biphasic in vitro maturation (IVM) approach with a meiotic inhibitor to improve dromedary camel IVM. Spontaneous meiotic resumption poses a major setback for in vitro matured oocytes. The overall objective of this study was to improve in vitro maturation of dromedary camel oocytes using ROCK inhibitor (Y-27632) in a biphasic IVM to prevent spontaneous meiotic resumption. In the first experiment, we cultured immature cumulus–oocyte complexes (COCs, n = 375) in a prematuration medium supplemented with ROCK inhibitor (RI) for 2 h, 4 h, 6 h, and 24 h before submission to normal in vitro maturation to complete 28 h. The control was cultured for 28 h in the absence of RI. In the first phase of experiment two, we cultured COCs (n = 480) in the presence or absence (control) of RI for 2 h, 4 h, 6 h, and 24 h, and conducted real-time relative quantitative PCR (qPCR) on selected mRNA transcripts. The same was done in the second phase, but qPCR was done after completion of normal IVM. Assessment of nuclear maturation showed that pre-IVM for 4 h yielded an increase in MII oocyte (54.67% vs. 26.6% of control; p < 0.05). As expected, the same group showed the highest degree (2) of cumulus expansion. In experiment 2, qPCR results showed significantly higher expression of ACTB and BCL2 in the RI group treated for 4 h when compared with the other groups. However, their relative quantification after biphasic IVM did not reveal any significant difference, except for the positive response of BCL2 and BAX/BCL2 ratio after 4 and 6 h biphasic IVM. In conclusion, RI prevents premature oocyte maturation and gave a significantly positive outcome during the 4 h treatment. This finding is a paradigm for future investigation on dromedary camel biphasic IVM and for improving the outcome of IVM in this species.
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Novel Apoptotic Mediators Identified by Conservation of Vertebrate Caspase Targets. Biomolecules 2020; 10:biom10040612. [PMID: 32326640 PMCID: PMC7225963 DOI: 10.3390/biom10040612] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2020] [Revised: 04/08/2020] [Accepted: 04/13/2020] [Indexed: 12/27/2022] Open
Abstract
Caspases are proteases conserved throughout Metazoans and responsible for initiating and executing the apoptotic program. Currently, there are over 1800 known apoptotic caspase substrates, many of them known regulators of cell proliferation and death, which makes them attractive therapeutic targets. However, most caspase substrates are by-standers, and identifying novel apoptotic mediators amongst all caspase substrates remains an unmet need. Here, we conducted an in silico search for significant apoptotic caspase targets across different species within the Vertebrata subphylum, using different criteria of conservation combined with structural features of cleavage sites. We observed that P1 aspartate is highly conserved while the cleavage sites are extensively variable and found that cleavage sites are located primarily in coiled regions composed of hydrophilic amino acids. Using the combination of these criteria, we determined the final list of the 107 most relevant caspase substrates including 30 novel targets previously unknown for their role in apoptosis and cancer. These newly identified substrates can be potential regulators of apoptosis and candidates for anti-tumor therapy.
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Pénzes M, Túrós D, Máthé D, Szigeti K, Hegedűs N, Rauscher AÁ, Tóth P, Ivic I, Padmanabhan P, Pál G, Dobolyi Á, Gyimesi M, Málnási-Csizmadia A. Direct myosin-2 inhibition enhances cerebral perfusion resulting in functional improvement after ischemic stroke. Theranostics 2020; 10:5341-5356. [PMID: 32373216 PMCID: PMC7196296 DOI: 10.7150/thno.42077] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/13/2019] [Accepted: 02/03/2020] [Indexed: 12/29/2022] Open
Abstract
Acute ischemic stroke treatment faces an unresolved obstacle as capillary reperfusion remains insufficient after thrombolysis and thrombectomy causing neuronal damage and poor prognosis. Hypoxia-induced capillary constriction is mediated by actomyosin contraction in precapillary smooth muscle cells (SMCs) therefore smooth muscle myosin-2 could be an ideal target with potentially high impact on reperfusion of capillaries. Methods: The myosin-2 inhibitor para-aminoblebbistatin (AmBleb) was tested on isolated human and rat arterioles to assess the effect of AmBleb on vasodilatation. Transient middle cerebral artery occlusion (MCAO) was performed on 38 male Wistar rats followed by local administration of AmBleb into the ischemic brain area. Development of brain edema and changes in cerebrovascular blood flow were assessed using MRI and SPECT. We also tested the neurological deficit scores and locomotor asymmetry of the animals for 3 weeks after the MCAO operation. Results: Our results demonstrate that AmBleb could achieve full relaxation of isolated cerebral arterioles. In living animals AmBleb recovered cerebral blood flow in 32 out of the 65 affected functional brain areas in MCAO operated rats, whereas only 8 out of the 67 affected areas were recovered in the control animals. Animals treated with AmBleb also showed significantly improved general and focal deficit scores in neurological functional tests and showed significantly ameliorated locomotor asymmetry. Conclusion: Direct inhibition of smooth muscle myosin by AmBleb in pre-capillary SMCs significantly contribute to the improvement of cerebral blood reperfusion and brain functions suggesting that smooth muscle myosin inhibition may have promising potential in stroke therapies as a follow-up treatment of physical or chemical removal of the occluding thrombus.
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Feldman MB, Wood M, Lapey A, Mou H. SMAD Signaling Restricts Mucous Cell Differentiation in Human Airway Epithelium. Am J Respir Cell Mol Biol 2020; 61:322-331. [PMID: 30848657 DOI: 10.1165/rcmb.2018-0326oc] [Citation(s) in RCA: 21] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
Mucin-secreting goblet cell metaplasia and hyperplasia (GCMH) is a common pathological phenotype in many human respiratory diseases, including asthma, chronic obstructive pulmonary disease, cystic fibrosis, primary ciliary dyskinesia, and infections. A better understanding of how goblet cell quantities or proportions in the airway epithelium are regulated may provide novel therapeutic targets to mitigate GCMH in these devastating diseases. We identify canonical SMAD signaling as the principal pathway restricting goblet cell differentiation in human airway epithelium. Differentiated goblet cells express low levels of phosphorylated SMAD. Accordingly, inhibition of SMAD signaling markedly amplifies GCMH induced by mucous mediators. In contrast, SMAD signaling activation impedes goblet cell generation and accelerates the resolution of preexisting GCMH. SMAD signaling inhibition can override the suppressive effects imposed by a GABAergic receptor inhibitor, suggesting the GABAergic pathway likely operates through inhibition of SMAD signaling in regulating mucous differentiation. Collectively, our data demonstrate that SMAD signaling plays a determining role in mucous cell differentiation, and thus raise the possibility that dysregulation of this pathway contributes to respiratory pathophysiology during airway inflammation and pulmonary diseases. In addition, our study also highlights the potential for SMAD modulation as a therapeutic target in mitigating GCMH.
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Affiliation(s)
- Michael B Feldman
- Division of Pulmonary and Critical Care Medicine and.,Harvard Medical School, Boston, Massachusetts
| | - Michael Wood
- the Mucosal Immunology and Biology Research Center, Massachusetts General Hospital, Boston, Massachusetts
| | - Allen Lapey
- Division of Pediatric Pulmonary Medicine, Massachusetts General Hospital for Children, Boston, Massachusetts; and
| | - Hongmei Mou
- the Mucosal Immunology and Biology Research Center, Massachusetts General Hospital, Boston, Massachusetts.,Division of Pediatric Pulmonary Medicine, Massachusetts General Hospital for Children, Boston, Massachusetts; and.,Department of Pediatrics, Harvard Medical School, Boston, Massachusetts
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Hu J, Zhang H, Li J, Jiang X, Zhang Y, Wu Q, Shen L, Shi J, Gao N. ROCK1 activation-mediated mitochondrial translocation of Drp1 and cofilin are required for arnidiol-induced mitochondrial fission and apoptosis. JOURNAL OF EXPERIMENTAL & CLINICAL CANCER RESEARCH : CR 2020; 39:37. [PMID: 32075676 PMCID: PMC7031977 DOI: 10.1186/s13046-020-01545-7] [Citation(s) in RCA: 32] [Impact Index Per Article: 6.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 12/09/2019] [Accepted: 02/14/2020] [Indexed: 01/11/2023]
Abstract
BACKGROUND Arnidiol is a pentacyclic triterpene diol that has multiple pharmacological activities. However, the apoptotic activities of arnidiol in human cancer cells have not yet been explored, nor has the mechanism by which arnidiol induces apoptosis been examined in depth. METHODS MDA-MB-231 cells and xenografted mice were treated with arnidiol. Mitochondrial fission and apoptosis were determined by immunofluorescence, flow cytometry and related molecular biological techniques. The interaction and colocalization of cofilin and Drp1 was determined by immunoprecipitation and immunofluorescence assays. RESULTS Arnidiol induces mitochondrial fission and apoptosis through mitochondrial translocation of Drp1 and cofilin. Importantly, the interaction of Drp1 and cofilin in mitochondria is involved in arnidiol-induced mitochondrial fission and apoptosis. Knockdown of either Drp1 or cofilin abrogated arnidiol-induced mitochondrial translocation, interaction of Drp1 and cofilin, mitochondrial fission and apoptosis. Only dephosphorylated Drp1 (Ser637) and cofilin (Ser3) were translocated to the mitochondria. Mutants of Drp1 S637A and cofilin S3A, which mimic the dephosphorylated forms, enhanced mitochondrial fission and apoptosis induced by arnidiol, whereas mutants of Drp1 S637D and cofilin S3E, which mimic the phosphorylated forms, suppressed mitochondrial fission and apoptosis induced by arnidiol. A mechanistic study revealed that ROCK1 activation plays an important role in the arnidiol-mediated Drp1 and cofilin dephosphorylation and mitochondrial translocation, mitochondrial fission, and apoptosis. CONCLUSIONS Our data reveal a novel role of both Drp1 and cofilin in the regulation of mitochondrial fission and apoptosis and suggest that arnidiol could be developed as a potential agent for the treatment of human cancer.
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Affiliation(s)
- Jinjiao Hu
- College of Pharmacy, Army Medical University, 30 Gaotanyan Street, Shapingba District, Chongqing, 400038, China
| | - Hongwei Zhang
- College of Pharmacy, Army Medical University, 30 Gaotanyan Street, Shapingba District, Chongqing, 400038, China
| | - Jie Li
- College of Pharmacy, Army Medical University, 30 Gaotanyan Street, Shapingba District, Chongqing, 400038, China
| | - Xiuxing Jiang
- College of Pharmacy, Army Medical University, 30 Gaotanyan Street, Shapingba District, Chongqing, 400038, China
| | - Yanhao Zhang
- College of Pharmacy, Army Medical University, 30 Gaotanyan Street, Shapingba District, Chongqing, 400038, China
| | - Qin Wu
- Key Laboratory of Basic Pharmacology of Ministry of Education and Joint International Research Laboratory of Ethnomedicine of Ministry of Education, Zunyi Medical University, Zunyi, China
| | - Liwen Shen
- Key Laboratory of Basic Pharmacology of Ministry of Education and Joint International Research Laboratory of Ethnomedicine of Ministry of Education, Zunyi Medical University, Zunyi, China
| | - Jingshan Shi
- Key Laboratory of Basic Pharmacology of Ministry of Education and Joint International Research Laboratory of Ethnomedicine of Ministry of Education, Zunyi Medical University, Zunyi, China.
| | - Ning Gao
- College of Pharmacy, Army Medical University, 30 Gaotanyan Street, Shapingba District, Chongqing, 400038, China. .,Key Laboratory of Basic Pharmacology of Ministry of Education and Joint International Research Laboratory of Ethnomedicine of Ministry of Education, Zunyi Medical University, Zunyi, China.
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