|
1
|
Li S, Ma T, An Y, Zhang Y, Yang X, Gao A, Wang H. The Impact of Different Dietary Ratios of Soluble Carbohydrate-to-Neutral Detergent Fiber on Rumen Barrier Function and Inflammation in Dumont Lambs. Animals (Basel) 2024; 14:1666. [PMID: 38891713 PMCID: PMC11171165 DOI: 10.3390/ani14111666] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/20/2024] [Revised: 05/27/2024] [Accepted: 05/30/2024] [Indexed: 06/21/2024] Open
Abstract
Appropriate soluble carbohydrate (SCHO)-to-NDF ratios in the diet are essential for rumen health. The effects of different SCHO-to-NDF ratios (1.0, 1.5, and 2.0) on rumen barrier function and inflammation in Dumont lambs (n = 18, 6 replicates per treatment) was investigated. The SCHO:NDF ratio was altered by replacing the forage (Leynus chinensis) with corn grain. With an increase in the proportion of SCHO, the final body weight (FBW), average daily gain (ADG), soluble carbohydrate intake (SCHOI), and LPS level increased; and the neutral detergent fiber intake (NDFI), ruminal papillae height, papillae area, and pH decreased (p < 0.05, plin < 0.05). The medium CHO:NDF group had increased claudin-1 mRNA (p < 0.05, plin = 0.005, pquad = 0.003) and protein (p < 0.05, pquad < 0.001) levels; the high CHO:NDF group had increased occludin mRNA and protein (p < 0.05, plin = 0.001) levels. The level of the anti-inflammatory cytokine IL-10 was significantly greater in the medium CHO:NDF group than in the high CHO:NDF group (p < 0.05, pquad < 0.001). With an increase in the ratio of SCHO, the mRNA level and concentration of the proinflammatory cytokines IL-1β, IL-6, and TNF-α linearly increased (p < 0.05, plin < 0.05), and those in the high CHO:NDF group were significantly greater than those in the low CHO:NDF group. The levels of phosphorylated p65 (plin = 0.003), IκB-α (plin < 0.001), and JNK (plin = 0.001) increased linearly, and those in the high CHO:NDF group were significantly greater than those in the other two groups (p < 0.05). Therefore, when the SCHO-to-NDF ratio was increased to 1.5, the rumen epithelium was not affected, but when the ratio was increased to 2.0, NF-κB and MAPK were activated in the rumen epithelium, leading to impaired barrier function and inflammation. The suitable NFC:NDF ratio for the short-term fattening of Dumont lambs was found to be 1.50.
Collapse
Affiliation(s)
- Shufang Li
- Animal Nutrition and Feed Science, Inner Mongolia Agricultural University, Hohhot 010018, China; (S.L.)
| | - Tian Ma
- Animal Nutrition and Feed Science, Inner Mongolia Agricultural University, Hohhot 010018, China; (S.L.)
| | - Yawen An
- Animal Nutrition and Feed Science, Inner Mongolia Agricultural University, Hohhot 010018, China; (S.L.)
| | - Yu Zhang
- Animal Nutrition and Feed Science, Inner Mongolia Agricultural University, Hohhot 010018, China; (S.L.)
| | - Xiaodong Yang
- Animal Nutrition and Feed Science, Inner Mongolia Agricultural University, Hohhot 010018, China; (S.L.)
| | - Aiwu Gao
- Food Science, Inner Mongolia Agricultural University, Hohhot 010018, China
| | - Hairong Wang
- Animal Nutrition and Feed Science, Inner Mongolia Agricultural University, Hohhot 010018, China; (S.L.)
| |
Collapse
|
|
2
|
Meyer F, Wendling D, Demougeot C, Prati C, Verhoeven F. Cytokines and intestinal epithelial permeability: A systematic review. Autoimmun Rev 2023; 22:103331. [PMID: 37030338 DOI: 10.1016/j.autrev.2023.103331] [Citation(s) in RCA: 31] [Impact Index Per Article: 15.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2023] [Accepted: 04/03/2023] [Indexed: 04/10/2023]
Abstract
BACKGROUND The intestinal mucosa is composed of a well-organized epithelium, acting as a physical barrier to harmful luminal contents, while simultaneously ensuring absorption of physiological nutrients and solutes. Increased intestinal permeability has been described in various chronic diseases, leading to abnormal activation of subepithelial immune cells and overproduction of inflammatory mediators. This review aimed to summarize and evaluate the effects of cytokines on intestinal permeability. METHODS A systematic review of the literature was performed in the Medline, Cochrane and Embase databases, up to 01/04/2022, to identify published studies assessing the direct effect of cytokines on intestinal permeability. We collected data on the study design, the method of assessment of intestinal permeability, the type of intervention and the subsequent effect on gut permeability. RESULTS A total of 120 publications were included, describing a total of 89 in vitro and 44 in vivo studies. TNFα, IFNγ or IL-1β were the most frequently studied cytokines, inducing an increase in intestinal permeability through a myosin light-chain-mediated mechanism. In situations associated with intestinal barrier disruption, such as inflammatory bowel diseases, in vivo studies showed that anti-TNFα treatment decreased intestinal permeability while achieving clinical recovery. In contrast to TNFα, IL-10 decreased permeability in conditions associated with intestinal hyperpermeability. For some cytokines (e.g. IL-17, IL-23), results are conflicting, with both an increase and a decrease in gut permeability reported, depending on the study model, methodology, or the studied conditions (e.g. burn injury, colitis, ischemia, sepsis). CONCLUSION This systematic review provides evidence that intestinal permeability can be directly influenced by cytokines in numerous conditions. The immune environment probably plays an important role, given the variability of their effect, according to different conditions. A better understanding of these mechanisms could open new therapeutic perspectives for disorders associated with gut barrier dysfunction.
Collapse
Affiliation(s)
- Frédéric Meyer
- PEPITE EA4267, Université de Franche-Comté, F-25000 Besançon, France; Department of rheumatology, University Hospital Besançon, F-25000 Besançon, France
| | - Daniel Wendling
- Department of rheumatology, University Hospital Besançon, F-25000 Besançon, France; EA 4266, EPILAB, Université de Franche-Comté, F-25000 Besançon, France
| | - Céline Demougeot
- PEPITE EA4267, Université de Franche-Comté, F-25000 Besançon, France
| | - Clément Prati
- PEPITE EA4267, Université de Franche-Comté, F-25000 Besançon, France; Department of rheumatology, University Hospital Besançon, F-25000 Besançon, France
| | - Frank Verhoeven
- PEPITE EA4267, Université de Franche-Comté, F-25000 Besançon, France; Department of rheumatology, University Hospital Besançon, F-25000 Besançon, France.
| |
Collapse
|
|
3
|
Jacques C, Floris I. Special Focus on the Cellular Anti-Inflammatory Effects of Several Micro-Immunotherapy Formulations: Considerations Regarding Intestinal-, Immune-Axis-Related- and Neuronal-Inflammation Contexts. J Inflamm Res 2022; 15:6695-6717. [PMID: 36536643 PMCID: PMC9759027 DOI: 10.2147/jir.s389614] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/19/2022] [Accepted: 11/24/2022] [Indexed: 04/11/2024] Open
Abstract
INTRODUCTION Chronic inflammation is a pernicious underlying status, well-known for its contribution to the progressive development of various diseases. In this regard, Micro-immunotherapy (MI) might be a promising therapeutic strategy. MI employs low doses (LD) and ultra-low doses (ULD) of immune regulators in their formulations. In particular, as both IL-1β and TNF-α are often used at ULD in MI medicines (MIM), a special emphasis has been made on formulations that include these factors in their compositions. METHODS Several in vitro models have been employed in order to assess the effects of two unitary MIM consisting of ULD of IL-1β and TNF-α (u-MIM-1 and u-MIM-2, respectively), and four complex MIM (c-MIM-1, -2, -3 and -4) characterized by the presence of ULD of IL-1β and TNF-α amongst other factors. Thus, we first investigated the anti-inflammatory effects of u-MIM-1 and u-MIM-2 in a model of inflamed colon carcinoma cells. In addition, the anti-inflammatory potential of c-MIM-1, -2, -3 and -4, was assessed in in vitro models of intestinal and neuronal inflammation. RESULTS The results revealed that u-MIM-1 and u-MIM-2 both induced a slight decrease in the levels of IL-1β and TNF-α transcripts. Regarding the c-MIMs' effects, c-MIM-1 displayed the capability to restore the altered transepithelial electrical resistance in inflamed-HCoEpiC cells. Moreover, c-MIM-1 also slightly increased the expression of the junction-related protein claudin-1, both at the mRNA and protein levels. In addition, our in vitro investigations on c-MIM-2 and c-MIM-3 revealed their immune-modulatory effects in LPS-inflamed human monocytes, macrophages, and granulocytes, on the secretion of cytokines such as TNF-α, PGE2, and IL-6. Finally, c-MIM-4 restored the cell viability of LPS/IFN-γ-inflamed rat cortical neurons, while reducing the secretion of TNF-α in rat glial cells. DISCUSSION Our results shed the light on the potential role of these MIM formulations in managing several chronic inflammation-related conditions.
Collapse
Affiliation(s)
- Camille Jacques
- Preclinical Research Department, Labo’Life France, Nantes, France
| | - Ilaria Floris
- Preclinical Research Department, Labo’Life France, Nantes, France
| |
Collapse
|
|
4
|
Inflammatory cytokines directly disrupt the bovine intestinal epithelial barrier. Sci Rep 2022; 12:14578. [PMID: 36028741 PMCID: PMC9418144 DOI: 10.1038/s41598-022-18771-y] [Citation(s) in RCA: 17] [Impact Index Per Article: 5.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/01/2022] [Accepted: 08/18/2022] [Indexed: 11/08/2022] Open
Abstract
The small intestinal mucosa constitutes a physical barrier separating the gut lumen from sterile internal tissues. Junctional complexes between cells regulate transport across the barrier, preventing water loss and the entry of noxious molecules or pathogens. Inflammatory diseases in cattle disrupt this barrier; nonetheless, mechanisms of barrier disruption in cattle are poorly understood. We investigated the direct effects of three inflammatory cytokines, TNFα, IFNγ, and IL-18, on the bovine intestinal barrier utilizing intestinal organoids. Flux of fluorescein isothiocyanate (FITC)-labeled dextran was used to investigate barrier permeability. Immunocytochemistry and transmission electron microscopy were used to investigate junctional morphology, specifically tortuosity and length/width, respectively. Immunocytochemistry and flow cytometry was used to investigate cellular turnover via proliferation and apoptosis. Our study shows that 24-h cytokine treatment with TNFα or IFNγ significantly increased dextran permeability and tight junctional tortuosity, and reduced cellular proliferation. TNFα reduced the percentage of G2/M phase cells, and IFNγ treatment increased cell apoptotic rate. IL-18 did not directly induce significant changes to barrier permeability or cellular turnover. Our study concludes that the inflammatory cytokines, TNFα and IFNγ, directly induce intestinal epithelial barrier dysfunction and alter the tight junctional morphology and rate of cellular turnover in bovine intestinal epithelial cells.
Collapse
|
|
5
|
Tumor Necrosis Factor Alpha Effects on the Porcine Intestinal Epithelial Barrier Include Enhanced Expression of TNF Receptor 1. Int J Mol Sci 2021; 22:ijms22168746. [PMID: 34445450 PMCID: PMC8395858 DOI: 10.3390/ijms22168746] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/06/2021] [Revised: 07/30/2021] [Accepted: 08/11/2021] [Indexed: 12/15/2022] Open
Abstract
Tumor necrosis factor alpha (TNFα) has been shown to impair the intestinal barrier, inducing and maintaining inflammatory states of the intestine. The aim of the current study was to analyze functional, molecular and regulatory effects of TNFα in a newly established non-transformed jejunal enterocyte model, namely IPEC-J2 monolayers. Incubation with 1000 U/mL TNFα induced a marked decrease in transepithelial electrical resistance (TEER), and an increase in permeability for the paracellular flux marker [3H]-D-mannitol compared to controls. Immunoblots revealed a significant decrease in tight junction (TJ) proteins occludin, claudin-1 and claudin-3. Moreover, a dose-dependent increase in the TNF receptor (TNFR)-1 was detected, explaining the exponential nature of pro-inflammatory effects, while TNFR-2 remained unchanged. Recovery experiments revealed reversible effects after the removal of the cytokine, excluding apoptosis as a reason for the observed changes. Furthermore, TNFα signaling could be inhibited by the specific myosin light chain kinase (MLCK) blocker ML-7. Results of confocal laser scanning immunofluorescence microscopy were in accordance with all quantitative changes. This study explains the self-enhancing effects of TNFα mediated by MLCK, leading to a differential regulation of TJ proteins resulting in barrier impairment in the intestinal epithelium.
Collapse
|
|
6
|
Nedzvetsky VS, Masiuk DM, Gasso VY, Yermolenko SV, Huslystyi AO, Spirina VA. Low doses of imidacloprid induce disruption of intercellular adhesion and initiate proinflammatory changes in Caco-2 cells. REGULATORY MECHANISMS IN BIOSYSTEMS 2021. [DOI: 10.15421/022159] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/17/2022] Open
Abstract
Imidacloprid is the most widely used pesticide of the neonicotinoid class. Neonicotinoid toxicities against various insects are well known. Nevertheless, there are rising evidences that neonicotinoids exert cytotoxic effects on different non-target organisms including mammals, fish, birds etc. Besides, depending on pesticide application, the exposed plants absorb some part of used neonicotinoids and their residues are detected in agricultural products worldwide. Thus, the continuous consumption of fruits and vegetables contaminated with neonicotinoids is a high risk factor for humans despite the low doses. Intestine epithelial cells are the first targets of the neonicotinoid cytotoxicity in humans because of its direct way of administration. The epithelial cells provide the barrier function of the intestinal system via specialized intercellular adhesion. The effects of imidacloprid on the intestine barrier function and inflammatory cytokines production are still unknown. In the present study, we exposed the human Caucasian colon adenocarcinoma (Caco-2) epithelial cells to low doses (0.10–0.75 µg/mL) of imidacloprid in order to assess the expression of tight and adherens junctions proteins, occludin and E-cadherin, and production of proinflammatory cytokine TNF α and iNOS. Imidacloprid induced dose-dependent decline in both occludin and E-cadherin levels. By contrast, TNF-α and iNOS contents were upregulated in imidacloprid-exposed Caco-2 cells. Decrease in tight and adherens junctions proteins indicates that the barrier function of intestine epithelial cells could be damaged by imidacloprid administration. In addition, TNF-α and iNOS upregulation indicates that imidacloprid is potent to activate proinflammatory response in enterocytes. Thus, imidacloprid can affect intestine barrier function through the increase of proinflammatory cytokine production and decrease in adhesiveness of enterocytes. The further assessment of the role of adhesion proteins and inflammatory cytokines in neonicotinoid pesticide cytotoxicity as it affects enterocyte barrier function is required to highlight the risk factor of use of neonicotinoids.
Collapse
|
|
7
|
Gustin A, Cromarty R, Schifanella L, Klatt NR. Microbial mismanagement: how inadequate treatments for vaginal dysbiosis drive the HIV epidemic in women. Semin Immunol 2021; 51:101482. [PMID: 34120819 DOI: 10.1016/j.smim.2021.101482] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/22/2021] [Accepted: 05/24/2021] [Indexed: 12/18/2022]
Abstract
Women and girls represent a key population driving new HIV infections and persistence of the HIV pandemic. A key determinant of HIV susceptibility is the composition of the vaginal microbiome, which can influence the local immune cell population, inflammation status, and HIV prevention drug levels. While a low-diversity composition dominated by Lactobacillus crispatus is associated with a decreased risk of HIV acquisition, high diversity environments associated with bacterial vaginosis increase risk of HIV. Given the important role of the vaginal microbiome in determining HIV susceptibility, altering the microbiome towards a Lactobacillus-dominated state is an attractive complementary strategy to reduce HIV incidence rates. Here, we provide an overview of the mechanisms by which the vaginal microbiome may contribute to HIV acquisition risk. Furthermore, we address the advantages and limitations of historical treatments and emerging technologies under investigation to modify the vaginal microbiome, including: antibiotics, bacteriophages, probiotics, topicals, and engineered bacteria. By addressing the current state of vaginal microbiome knowledge and strategies for manipulation, we hope to amplify the growing calls for increased resources and research into vaginal microbial health, which will be essential to accelerating preventative efforts amongst the world's most vulnerable populations.
Collapse
Affiliation(s)
- Andrew Gustin
- Department of Global Health, University of Washington, Seattle, WA, USA
| | - Ross Cromarty
- Department of Surgery, Division of Surgical Outcomes and Precision Medicine Research, University of Minnesota, Minneapolis, MN, USA; Department of Medicine, University of Minnesota, Minneapolis, MN, USA
| | - Luca Schifanella
- Department of Surgery, Division of Surgical Outcomes and Precision Medicine Research, University of Minnesota, Minneapolis, MN, USA
| | - Nichole R Klatt
- Department of Surgery, Division of Surgical Outcomes and Precision Medicine Research, University of Minnesota, Minneapolis, MN, USA.
| |
Collapse
|
|
8
|
Al-Sadi R, Engers J, Haque M, King S, Al-Omari D, Ma TY. Matrix Metalloproteinase-9 (MMP-9) induced disruption of intestinal epithelial tight junction barrier is mediated by NF-κB activation. PLoS One 2021; 16:e0249544. [PMID: 33826658 PMCID: PMC8026081 DOI: 10.1371/journal.pone.0249544] [Citation(s) in RCA: 42] [Impact Index Per Article: 10.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/06/2020] [Accepted: 03/20/2021] [Indexed: 12/13/2022] Open
Abstract
BACKGROUND Matrix Metalloproteinase-9 (MMP-9) has been shown to play a key role in mediating inflammation and tissue damage in inflammatory bowel disease (IBD). In patients with IBD, the intestinal tight junction (TJ) barrier is compromised as characterized by an increase in intestinal permeability. MMP-9 is elevated in intestinal tissue, serum and stool of patients with IBD. Previous studies from our laboratory showed that MMP-9 causes an increase in intestinal epithelial TJ permeability and that the MMP-9 induced increase in intestinal permeability is an important pathogenic factor contributing to the development of intestinal inflammation in IBD. However, the intracellular mechanisms that mediate the MMP-9 modulation of intestinal barrier function remain unclear. AIMS The main aim of this study was to further elucidate the molecular mechanisms involved in MMP-9 induced increase in intestinal epithelial TJ permeability using Caco-2 monolayers as an in-vitro model system. RESULTS MMP-9 induced increase in Caco-2 TJ permeability was associated with activation and cytoplasmic-to-nuclear translocation of NF-κB p65. Knocking-down NF-κB p65 by siRNA transfection prevented the MMP-9 induced expression of the NF-κB target gene IL-8, myosin light chain kinase (MLCK) protein expression, and subsequently prevented the increase in Caco-2 TJ permeability. In addition, the effect of MMP-9 on Caco-2 intestinal epithelial TJ barrier function was not mediated by apoptosis or necrosis. CONCLUSION Our data show that the MMP-9 induced disruption of Caco-2 intestinal epithelial TJ barrier function is regulated by NF-κB pathway activation of MLCK.
Collapse
Affiliation(s)
- Rana Al-Sadi
- Department of Medicine, Penn State University College of Medicine, Milton S. Hershey Medical Center, Hershey, Pennsylvania, United States of America
- * E-mail:
| | - Jessica Engers
- Department of Medicine, Penn State University College of Medicine, Milton S. Hershey Medical Center, Hershey, Pennsylvania, United States of America
| | - Mohammad Haque
- Department of Medicine, Penn State University College of Medicine, Milton S. Hershey Medical Center, Hershey, Pennsylvania, United States of America
| | - Steven King
- Department of Medicine, Penn State University College of Medicine, Milton S. Hershey Medical Center, Hershey, Pennsylvania, United States of America
| | - Deemah Al-Omari
- Department of Biology, University of New Mexico, Albuquerque, New Mexico, United States of America
| | - Thomas Y. Ma
- Department of Medicine, Penn State University College of Medicine, Milton S. Hershey Medical Center, Hershey, Pennsylvania, United States of America
| |
Collapse
|
|
9
|
Lobo de Sá FD, Schulzke JD, Bücker R. Diarrheal Mechanisms and the Role of Intestinal Barrier Dysfunction in Campylobacter Infections. Curr Top Microbiol Immunol 2021; 431:203-231. [PMID: 33620653 DOI: 10.1007/978-3-030-65481-8_8] [Citation(s) in RCA: 12] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022]
Abstract
Campylobacter enteritis is the most common cause of foodborne bacterial diarrhea in humans. Although various studies have been performed to clarify the pathomechanism in Campylobacter infection, the mechanism itself and bacterial virulence factors are yet not completely understood. The purpose of this chapter is to (i) give an overview on Campylobacter-induced diarrheal mechanisms, (ii) illustrate underlying barrier defects, (iii) explain the role of the mucosal immune response and (iv) weigh preventive and therapeutic approaches. Our present knowledge of pathogenetic and diarrheal mechanisms of Campylobacter jejuni is explained in the first part of this chapter. In the second part, the molecular basis for the Campylobacter-induced barrier dysfunction is compared with that of other species in the Campylobacter genus. The bacteria are capable of overcoming the intestinal epithelial barrier. The invasion into the intestinal mucosa is the initial step of the infection, followed by a second step, the epithelial barrier impairment. The extent of the impairment depends on various factors, including tight junction dysregulation and epithelial apoptosis. The disturbed intestinal epithelium leads to a loss of water and solutes, the leak flux type of diarrhea, and facilitates the uptake of harmful antigens, the leaky gut phenomenon. The barrier dysfunction is accompanied by increased pro-inflammatory cytokine secretion, which is partially responsible for the dysfunction. Moreover, cytokines also mediate ion channel dysregulation (e.g., epithelial sodium channel, ENaC), leading to another diarrheal mechanism, which is sodium malabsorption. Future perspectives of Campylobacter research are the clarification of molecular pathomechanisms and the characterization of therapeutic and preventive compounds to combat and prevent Campylobacter infections.
Collapse
Affiliation(s)
- Fábia Daniela Lobo de Sá
- Institute of Clinical Physiology/Nutritional Medicine, Medical Department, Division of Gastroenterology, Infectiology, Rheumatology, Charité - University Medicine Berlin, Campus Benjamin Franklin, Hindenburgdamm 30, 12203, Berlin, Germany
| | - Jörg-Dieter Schulzke
- Institute of Clinical Physiology/Nutritional Medicine, Medical Department, Division of Gastroenterology, Infectiology, Rheumatology, Charité - University Medicine Berlin, Campus Benjamin Franklin, Hindenburgdamm 30, 12203, Berlin, Germany
| | - Roland Bücker
- Institute of Clinical Physiology/Nutritional Medicine, Medical Department, Division of Gastroenterology, Infectiology, Rheumatology, Charité - University Medicine Berlin, Campus Benjamin Franklin, Hindenburgdamm 30, 12203, Berlin, Germany.
| |
Collapse
|
|
10
|
Inflammation, HIV, and Immune Quiescence: Leveraging on Immunomodulatory Products to Reduce HIV Susceptibility. AIDS Res Treat 2020; 2020:8672850. [PMID: 33178456 PMCID: PMC7609152 DOI: 10.1155/2020/8672850] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2020] [Revised: 09/23/2020] [Accepted: 10/15/2020] [Indexed: 12/29/2022] Open
Abstract
The relationship between inflammation and HIV has been a focus of research over the last decade. In HIV-infected individuals, increased HIV-associated immune activation significantly correlated to disease progression. While genital inflammation (GI) has been shown to significantly increase the risk of HIV acquisition and transmission, immune correlates for reduced risk remain limited. In certain HIV-exposed seronegative individuals, an immune quiescent phenotype characterized reduced risk. Immune quiescence is defined by specific, targeted, highly regulated immune responses that hinder overt inflammation or immune activation. Targeted management of inflammation, therefore, is a plausible strategy to mitigate HIV risk and slow disease progression. Nonsteroidal anti-inflammatory drugs (NSAIDs) such as hydroxychloroquine and aspirin have shown encouraging preliminary results in low-risk women by reducing systemic and genital immune activation. A topical NSAID, containing ibuprofen, is effective in treating vulvovaginal inflammation. Additionally, the glucocorticoids (GCs), prednisolone, and dexamethasone are used to treat HIV-associated immune activation. Collectively, these data inform on immune-modulating drugs to reduce HIV risk. However, the prolonged use of these pharmaceutical drugs is associated with adverse effects, both systemically and to a lesser extent topically. Natural products with their reduced side effects coupled with anti-inflammatory properties render them viable options. Lactic acid (LA) has immunomodulatory properties. LA regulates the genital microbiome by facilitating the growth of Lactobacillus species, while simultaneously limiting bacterial species that cause microbial dysbiosis and GI. Glycerol monolaurate, besides being anti-inflammatory, also inhibited SIV infections in rhesus macaques. The proposed pharmaceutical and natural products could be used in combination with either antiretrovirals for treatment or preexposure prophylaxis for HIV prevention. This review provides a summary on the associations between inflammation, HIV risk, and disease progression. Furthermore, we use the knowledge from immune quiescence to exploit the use of pharmaceutical and natural products as strategic interventions to manage inflammation, toward mitigating HIV infections.
Collapse
|
|
11
|
Cromarty R, Sigal A, Liebenberg LJ, Mckinnon LR, Abdool Karim SS, Passmore JAS, Archary D. Betamethasone induces potent immunosuppression and reduces HIV infection in a PBMC in vitro model. J Investig Med 2020; 69:28-40. [PMID: 33004468 PMCID: PMC7803916 DOI: 10.1136/jim-2020-001424] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 08/25/2020] [Indexed: 01/15/2023]
Abstract
Genital inflammation is an established risk factor for increased HIV acquisition risk. Certain HIV-exposed seronegative populations, who are naturally resistant to HIV infection, have an immune quiescent phenotype defined by reduced immune activation and inflammatory cytokines at the genital tract. Therefore, the aim of this study was to create an immune quiescent environment using immunomodulatory drugs to mitigate HIV infection. Using an in vitro peripheral blood mononuclear cell (PBMC) model, we found that inflammation was induced using phytohemagglutinin and Toll-like receptor (TLR) agonists Pam3CSK4 (TLR1/2), lipopolysaccharide (LPS) (TLR4) and R848 (TLR7/8). After treatment with anti-inflammatory drugs, ibuprofen (IBF) and betamethasone (BMS), PBMCs were exposed to HIV NL4-3 AD8. Multiplexed ELISA was used to measure 28 cytokines to assess inflammation. Flow cytometry was used to measure immune activation (CD38, HLA-DR and CCR5) and HIV infection (p24 production) of CD4+ T cells. BMS potently suppressed inflammation (soluble cytokines, p<0.05) and immune activation (CD4+ T cells, p<0.05). BMS significantly reduced HIV infection of CD4+ T cells only in the LPS (0.98%) and unstimulated (1.7%) conditions (p<0.02). In contrast, IBF had minimal anti-inflammatory and immunosuppressive but no anti-HIV effects. BMS demonstrated potent anti-inflammatory effects, regardless of stimulation condition. Despite uniform immunosuppression, BMS differentially affected HIV infection according to the stimulation conditions, highlighting the complex nature of these interactions. Together, these data underscore the importance of interrogating inflammatory signaling pathways to identify novel drug targets to mitigate HIV infection.
Collapse
Affiliation(s)
- Ross Cromarty
- Mucosal Immunology Laboratory, CAPRISA, Durban, KwaZulu-Natal, South Africa
| | - Alexander Sigal
- Africa Health Research Institute (AHRI), Durban, KwaZulu-Natal, South Africa
- Max-Planck-Institute for Infection Biology, Berlin, Germany
| | - Lenine Julie Liebenberg
- Mucosal Immunology Laboratory, CAPRISA, Durban, KwaZulu-Natal, South Africa
- Department of Medical Microbiology, University of KwaZulu-Natal, Durban, KwaZulu-Natal, South Africa
| | - Lyle Robert Mckinnon
- Mucosal Immunology Laboratory, CAPRISA, Durban, KwaZulu-Natal, South Africa
- Department of Medical Microbiology and Infectious Diseases, University of Manitoba, Winnipeg, Manitoba, Canada
| | - Salim Safurdeen Abdool Karim
- Mucosal Immunology Laboratory, CAPRISA, Durban, KwaZulu-Natal, South Africa
- Department of Epidemiology, Columbia University Mailman School of Public Health, New York, New York, USA
| | - Jo-Ann Shelly Passmore
- Mucosal Immunology Laboratory, CAPRISA, Durban, KwaZulu-Natal, South Africa
- Institute of Infectious Diseases and Molecular Medicine, University of Cape Town Faculty of Health Sciences, Cape Town, Western Cape, South Africa
| | - Derseree Archary
- Mucosal Immunology Laboratory, CAPRISA, Durban, KwaZulu-Natal, South Africa
- Department of Medical Microbiology, University of KwaZulu-Natal, Durban, KwaZulu-Natal, South Africa
| |
Collapse
|
|
12
|
Ahmadi S, Wang S, Nagpal R, Wang B, Jain S, Razazan A, Mishra SP, Zhu X, Wang Z, Kavanagh K, Yadav H. A human-origin probiotic cocktail ameliorates aging-related leaky gut and inflammation via modulating the microbiota/taurine/tight junction axis. JCI Insight 2020; 5:132055. [PMID: 32302292 DOI: 10.1172/jci.insight.132055] [Citation(s) in RCA: 150] [Impact Index Per Article: 30.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/05/2019] [Accepted: 04/08/2020] [Indexed: 12/25/2022] Open
Abstract
Inflammation is a major risk factor of morbidity and mortality in older adults. Although its precise etiology is unknown, low-grade inflammation in older adults is commonly associated with increased intestinal epithelial permeability (leaky gut) and abnormal (dysbiotic) gut microbiota. The increasing older population and lack of treatments to reduce aging-related microbiota dysbiosis, leaky gut, and inflammation culminates in a rise in aging-related comorbidities, constituting a significant public health concern. Here, we demonstrate that a human-origin probiotic cocktail containing 5 Lactobacillus and 5 Enterococcus strains isolated from healthy infant gut prevented high-fat diet-induced (HFD-induced) microbiota dysbiosis, leaky gut, inflammation, metabolic dysfunctions, and physical function decline in older mice. Probiotic-modulated gut microbiota primarily reduced leaky gut by increasing tight junctions, which in turn reduced inflammation. Mechanistically, probiotics modulated microbiota in a way to increase bile salt hydrolase activity, which in turn increased taurine abundance in the gut that stimulated tight junctions and suppressed gut leakiness. Furthermore, in Caenorhabditis elegans, taurine increased life span, reduced adiposity and leaky gut, and enhanced physical function. The results suggest that such probiotic therapies could prevent or treat aging-related leaky gut and inflammation in the elderly.
Collapse
Affiliation(s)
- Shokouh Ahmadi
- Department of Internal Medicine-Molecular Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA
| | - Shaohua Wang
- Department of Internal Medicine-Molecular Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA
| | - Ravinder Nagpal
- Department of Internal Medicine-Molecular Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA
| | - Bo Wang
- Department of Chemistry, North Carolina A&T State University, Greensboro, North Carolina, USA
| | - Shalini Jain
- Department of Internal Medicine-Endocrinology and Metabolism.,Mouse Metabolic Phenotyping Core
| | - Atefeh Razazan
- Department of Internal Medicine-Molecular Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA
| | - Sidharth P Mishra
- Department of Internal Medicine-Molecular Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA
| | - Xuewei Zhu
- Department of Internal Medicine-Molecular Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA.,Department of Microbiology and Immunology, and
| | - Zhan Wang
- Department of Internal Medicine-Molecular Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA
| | - Kylie Kavanagh
- Department of Pathology-Comparative Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA.,Biomedical Sciences, University of Tasmania, Hobart, Australia
| | - Hariom Yadav
- Department of Internal Medicine-Molecular Medicine, Wake Forest School of Medicine, Winston-Salem, North Carolina, USA.,Department of Microbiology and Immunology, and
| |
Collapse
|
|
13
|
Butkevych E, Lobo de Sá FD, Nattramilarasu PK, Bücker R. Contribution of Epithelial Apoptosis and Subepithelial Immune Responses in Campylobacter jejuni- Induced Barrier Disruption. Front Microbiol 2020; 11:344. [PMID: 32210941 PMCID: PMC7067706 DOI: 10.3389/fmicb.2020.00344] [Citation(s) in RCA: 17] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2019] [Accepted: 02/17/2020] [Indexed: 12/17/2022] Open
Abstract
Campylobacter jejuni is a widespread zoonotic pathogen and the leading bacterial cause of foodborne gastroenteritis in humans. Previous infection studies showed disruption of intercellular contacts, induction of epithelial apoptosis, and immune activation, all three contributing to intestinal barrier dysfunction leading to diarrhea. The present study aims to determine the impact of subepithelial immune cells on intestinal barrier dysfunction during Campylobacter jejuni infection and the underlying pathological mechanisms. Infection was performed in a co-culture of confluent monolayers of the human colon cell line HT-29/B6-GR/MR and THP-1 immune cells. Twenty-two hours after infection, transepithelial electrical resistance (TER) was decreased by 58 ± 6% compared to controls. The infection resulted in an increase in permeability for fluorescein (332 Da; 4.5-fold) and for FITC-dextran (4 kDa; 3.5-fold), respectively. In contrast, incubation of the co-culture with the pan-caspase inhibitor Q-VD-OPh during the infection resulted in a complete recovery of the decrease in TER and a normalization of flux values. Fluorescence microscopy showed apoptotic fragmentation in infected cell monolayers resulting in a 5-fold increase of the apoptotic ratio, accompanied by an increased caspase-3 cleavage and caspase-3/7 activity, which both were not present after Q-VD-OPh treatment. Western blot analysis revealed increased claudin-1 and claudin-2 protein expression. Inhibition of apoptosis induction did not normalize these tight junction changes. TNFα concentration was increased during the infection in the co-culture. In conclusion, Campylobacter jejuni infection and the consequent subepithelial immune activation cause intestinal barrier dysfunction mainly through caspase-3-dependent epithelial apoptosis. Concomitant tight junction changes were caspase-independent. Anti-apoptotic and immune-modulatory substances appear to be promising agents for treatment of campylobacteriosis.
Collapse
Affiliation(s)
- Eduard Butkevych
- Institute of Clinical Physiology/Nutritional Medicine, Medical Department, Division of Gastroenterology, Infectiology and Rheumatology, Charité-Universitätsmedizin Berlin, Berlin, Germany
| | - Fábia Daniela Lobo de Sá
- Institute of Clinical Physiology/Nutritional Medicine, Medical Department, Division of Gastroenterology, Infectiology and Rheumatology, Charité-Universitätsmedizin Berlin, Berlin, Germany
| | - Praveen Kumar Nattramilarasu
- Institute of Clinical Physiology/Nutritional Medicine, Medical Department, Division of Gastroenterology, Infectiology and Rheumatology, Charité-Universitätsmedizin Berlin, Berlin, Germany
| | - Roland Bücker
- Institute of Clinical Physiology/Nutritional Medicine, Medical Department, Division of Gastroenterology, Infectiology and Rheumatology, Charité-Universitätsmedizin Berlin, Berlin, Germany
| |
Collapse
|
|
14
|
Protective effect of Saccharomyces boulardii on intestinal mucosal barrier of dextran sodium sulfate-induced colitis in mice. Chin Med J (Engl) 2020; 132:1951-1958. [PMID: 31335471 PMCID: PMC6708699 DOI: 10.1097/cm9.0000000000000364] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
Background: The effect and mechanism of Saccharomyces boulardii (Sb) in inflammatory bowel disease are unclear. The objective of the study was to evaluate the impact of Sb on intestinal mucosal barrier and intestinal flora in a colitis mouse model. Methods: Forty C57BL/6J male mice were randomly assigned to five groups: normal control group (A), pathologic control group (B), Sb treatment group (C), mesalazine treatment group (D), and Sb combined with mesalazine treatment group (E). Colitis was induced by the addition of 2.5% (wt/vol) dextran sodium sulfate (DSS) in the drinking water ad libitum for 7 days. The general condition, weight change, stool property, and bloody stool level of mice were observed to evaluate the disease activity index. The expression of zona occludens-1 (ZO-1) and occludin in intestinal tissue were measured by immunohistochemistry. The level of tumor necrosis factor-α (TNF-α) and interleukin (IL)-8 in plasma was measured by enzyme linked immunosorbent assay. Inter-cellular tight junctions were observed by transmission electron microscopy. The feces and intestinal contents were collected sterilely, and intestinal flora was analyzed by 16S rRNA sequencing. Results: Compared with group B, Sb reduced the disease activity index and histological score of group C (disease activity index: group B 2.708 ± 0.628, group C 1.542 ± 0.616, PBC = 0.005; histological score: group B 9.875 ± 3.271, group C 4.750 ± 1.832, PBC = 0.005) in DSS-induced colitis in mice. Sb exerted a protect effect on the expression of ZO-1 (group B 2.075 ± 1.176, group C 4.225 ± 1.316, PBC = 0.019) and occludin (group B 2.200 ± 0.968, group C 3.525 ± 1.047, PBC = 0.023). Compared with group B, Sb decreased the level of TNF-α and IL-8 of group C (TNF-α: group B 716.323 ± 44.691 ng/L, group C 521.740 ± 90.121 ng/L, PBC = 0.001; IL-8: group B 128.992 ± 11.475 pg/mL, group C 106.283 ± 15.906 pg/mL, PBC = 0.012). Treatment with Sb preserved the tight junctions and ameliorated microvilli and inter-cellular space. Treatment with Sb also showed its own characteristics: a higher percentage of Bacteroidetes and a lower percentage of Firmicutes, with significant differences or a significant trend. The proportion of the S24-7 family was increased significantly in the Sb treatment group. Conclusions: Sb shows an anti-inflammatory effect and has a protective effect on the intestinal mucosal mechanical barrier. Sb may up-regulate the abundance of family S24-7 specifically, and maybe a mechanism underlying its function.
Collapse
|
|
15
|
Wang X, Wang H, Zhang R, Li D, Gao MQ. LRRC75A antisense lncRNA1 knockout attenuates inflammatory responses of bovine mammary epithelial cells. Int J Biol Sci 2020; 16:251-263. [PMID: 31929753 PMCID: PMC6949150 DOI: 10.7150/ijbs.38214] [Citation(s) in RCA: 17] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/06/2019] [Accepted: 10/14/2019] [Indexed: 12/12/2022] Open
Abstract
Long noncoding RNAs (lncRNAs) play multiple key roles during inflammatory processes. In this study, a novel lncRNA identified by the high-throughput sequencing analysis was found significantly down-regulated in Escherichia coli-introduced cell model of bovine mastitis. Given that this lncRNA consists of the antisense of leucine-rich repeat-containing protein 75A (LRRC75A), it was named LRRC75A antisense lncRNA1 (LRRC75A-AS1). The expression of LRRC75A-AS1 was down-regulated in bovine mammary epithelial cells and mammary tissues under inflammatory condition. Knockout (KO) of LRRC75A-AS1 by CRISPR-Cas9 system in bovine mammary alveolar cell-T (MAC-T) cell line could enhance expressions of tight junction (TJ) proteins Claudin-1, Occludin and ZO-1, reduce cell monolayer permeability, and inhibit Staphylococcus aureus adhesion and invasion. Meanwhile, it also down-regulated expressions of inflammatory factors and attenuated activation of NF-κB pathway. Similarly, knockdown of LRRC75A caused the changes as LRRC75A-AS1 KO did, while overexpression of LRRC75A enabled the opposite effects. TJ of epithelioid cells barriers the pathogenic microorganisms outside during inflammation, in which LRRC75A-AS1 can regulate the expression of TJ proteins through LRRC75A, affecting the development of inflammation.
Collapse
Affiliation(s)
- Xixi Wang
- College of Veterinary Medicine, Northwest A&F University, Yangling 712100, China
| | - Hao Wang
- College of Veterinary Medicine, Northwest A&F University, Yangling 712100, China
| | - Ruiqi Zhang
- College of Veterinary Medicine, Northwest A&F University, Yangling 712100, China
| | - Dan Li
- College of Veterinary Medicine, Northwest A&F University, Yangling 712100, China
| | - Ming-Qing Gao
- College of Veterinary Medicine, Northwest A&F University, Yangling 712100, China
| |
Collapse
|
|
16
|
Xu P, Elamin E, Elizalde M, Bours PPHA, Pierik MJ, Masclee AAM, Jonkers DMAE. Modulation of Intestinal Epithelial Permeability by Plasma from Patients with Crohn's Disease in a Three-dimensional Cell Culture Model. Sci Rep 2019; 9:2030. [PMID: 30765731 PMCID: PMC6375954 DOI: 10.1038/s41598-018-38322-8] [Citation(s) in RCA: 40] [Impact Index Per Article: 6.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/23/2018] [Accepted: 12/10/2018] [Indexed: 12/11/2022] Open
Abstract
Intestinal epithelial barrier is affected by multiple factors, such as tumour necrosis factor-α (TNF-α). Plasma concentration of TNF-α is higher in patients with Crohn’s disease (CD) than healthy controls (HC) and correlates positively with disease activity. This study aimed to determine the effect of plasma from active, inactive CD patients on intestinal barrier function and to investigate the underlying mechanism. Plasma samples were collected from CD patients and HC. 3D Caco-2 cysts were treated with plasma or TNF-α, with or without pre-incubation of adalimumab (a monoclonal antibody that antagonizes TNF-α) or JNK inhibitor SP600125. The results demonstrated that exposure of the cysts to plasma from CD patients resulted in enhanced paracellular permeability in a disease activity-dependent manner. Compared to HC, active CD plasma decreased ZO-1 and OCCLUDIN expression on mRNA and protein levels, and led to an increased JNK phosphorylation. Pre-incubation with adalimumab or SP600125 ameliorated TJ disruption and barrier dysfunction induced by plasma from CD patients. These results indicate that plasma from CD patients is able to induce epithelial barrier disruption, in part through TNF-α induced TJs modulation. The data also demonstrate an involvement of MAPK pathway, in particular the JNK isoform, in CD patient plasma-induced barrier dysfunction.
Collapse
Affiliation(s)
- Pan Xu
- Division of Gastroenterology-Hepatology, Department of Internal Medicine, Maastricht University Medical Centre, Maastricht, The Netherlands.,School of Nutrition and Translational Research in Metabolism, Maastricht University Medical Center, Maastricht, The Netherlands
| | - Elhaseen Elamin
- Division of Gastroenterology-Hepatology, Department of Internal Medicine, Maastricht University Medical Centre, Maastricht, The Netherlands.,School of Nutrition and Translational Research in Metabolism, Maastricht University Medical Center, Maastricht, The Netherlands
| | - Montserrat Elizalde
- Division of Gastroenterology-Hepatology, Department of Internal Medicine, Maastricht University Medical Centre, Maastricht, The Netherlands.,School of Nutrition and Translational Research in Metabolism, Maastricht University Medical Center, Maastricht, The Netherlands
| | - Paul P H A Bours
- Division of Gastroenterology-Hepatology, Department of Internal Medicine, Maastricht University Medical Centre, Maastricht, The Netherlands
| | - Marieke J Pierik
- Division of Gastroenterology-Hepatology, Department of Internal Medicine, Maastricht University Medical Centre, Maastricht, The Netherlands.,School of Nutrition and Translational Research in Metabolism, Maastricht University Medical Center, Maastricht, The Netherlands
| | - Ad A M Masclee
- Division of Gastroenterology-Hepatology, Department of Internal Medicine, Maastricht University Medical Centre, Maastricht, The Netherlands.,School of Nutrition and Translational Research in Metabolism, Maastricht University Medical Center, Maastricht, The Netherlands
| | - Daisy M A E Jonkers
- Division of Gastroenterology-Hepatology, Department of Internal Medicine, Maastricht University Medical Centre, Maastricht, The Netherlands. .,School of Nutrition and Translational Research in Metabolism, Maastricht University Medical Center, Maastricht, The Netherlands.
| |
Collapse
|
|
17
|
Hering NA, Luettig J, Krug SM, Wiegand S, Gross G, van Tol EA, Schulzke JD, Rosenthal R. Lactoferrin protects against intestinal inflammation and bacteria-induced barrier dysfunction in vitro. Ann N Y Acad Sci 2017; 1405:177-188. [PMID: 28614589 DOI: 10.1111/nyas.13405] [Citation(s) in RCA: 63] [Impact Index Per Article: 7.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/27/2017] [Revised: 05/05/2017] [Accepted: 05/10/2017] [Indexed: 12/15/2022]
Abstract
The iron-binding glycoprotein lactoferrin (LF) is naturally present in human breast milk. Several studies suggest that LF contributes to infant health and development owing to a variety of protective effects, including antimicrobial and anti-inflammatory features. Therefore, we aimed to elucidate its protective properties on intestinal epithelial barrier dysfunction induced by infection or inflammation using the human epithelial cell culture models HT-29/B6 and T84. During barrier perturbation induced by the proinflammatory cytokine tumor necrosis factor α (TNF-α), bovine LF restored tight junction (TJ) morphometry and inhibited TNF-α-induced epithelial apoptosis. This resulted in an attenuation of the TNF-α-induced decrease in transepithelial resistance (TER) and increases in permeability of fluorescein and FITC-dextran (4 kDa) and was as effective as the apoptosis inhibitor Q-VD-Oph. The enteropathogenic bacterium Yersinia enterocolitica is a frequent cause of diarrhea in early childhood. This involves focal changes in TJ protein expression and localization. LF diminished the Y. enterocolitica-induced drop in TER in the present in vitro model, which was paralleled by an inhibition of the Yersinia-induced reduction of claudin-8 expression via c-Jun kinase signaling. In conclusion, LF exerts protective effects against inflammation- or infection-induced barrier dysfunction in human intestinal cell lines, supporting its relevance for healthy infant development.
Collapse
Affiliation(s)
- Nina A Hering
- Department of General, Visceral and Vascular Surgery, Campus Benjamin Franklin, Charité - Universitätsmedizin Berlin, Berlin, Germany
| | - Julia Luettig
- Institute of Clinical Physiology, Department of Gastroenterology, Campus Benjamin Franklin, Charité - Universitätsmedizin Berlin, Berlin, Germany
| | - Susanne M Krug
- Institute of Clinical Physiology, Department of Gastroenterology, Campus Benjamin Franklin, Charité - Universitätsmedizin Berlin, Berlin, Germany
| | - Stephanie Wiegand
- Institute of Clinical Physiology, Department of Gastroenterology, Campus Benjamin Franklin, Charité - Universitätsmedizin Berlin, Berlin, Germany
| | - Gabriele Gross
- Mead Johnson Pediatric Nutrition Institute, Nijmegen, the Netherlands
| | - Eric A van Tol
- Mead Johnson Pediatric Nutrition Institute, Nijmegen, the Netherlands
| | - Jörg D Schulzke
- Institute of Clinical Physiology, Department of Gastroenterology, Campus Benjamin Franklin, Charité - Universitätsmedizin Berlin, Berlin, Germany
| | - Rita Rosenthal
- Institute of Clinical Physiology, Department of Gastroenterology, Campus Benjamin Franklin, Charité - Universitätsmedizin Berlin, Berlin, Germany
| |
Collapse
|
|
18
|
Clemente AM, Castronovo G, Antonelli A, D’Andrea MM, Tanturli M, Perissi E, Paccosi S, Parenti A, Cozzolino F, Rossolini GM, Torcia MG. Differential Th17 response induced by the two clades of the pandemic ST258 Klebsiella pneumoniae clonal lineages producing KPC-type carbapenemase. PLoS One 2017; 12:e0178847. [PMID: 28586386 PMCID: PMC5460819 DOI: 10.1371/journal.pone.0178847] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/06/2017] [Accepted: 05/19/2017] [Indexed: 01/29/2023] Open
Abstract
The spread of KPC-type carbapenemases is mainly attributed to the global dissemination of Klebsiella pneumoniae (KP) strains belonging to the clonal group (CG) 258, including sequence type (ST) 258 and other related STs. Two distinct clades of CG258-KP have evolved, which differ mainly for the composition of their capsular polysaccharides, and recent studies indicate that clade 1 evolved from an ancestor of clade 2 by recombination of a genomic fragment carrying the capsular polysaccharide (cps) locus. In this paper, we investigated the ability of two ST258-KP strains, KKBO-1 and KK207-1, selected as representatives of ST258-KP clade 2 and clade 1, respectively, to activate an adaptive immune response using ex vivo-stimulation of PBMC from normal donors as an experimental model. Our data showed that KKBO-1 (clade 2) induces a Th17 response more efficiently than KK207-1 (clade 1): the percentage of CD4+IL17+ cells and the production of IL-17A were significantly higher in cultures with KKBO-1 compared to cultures with KK207-1. While no differences in the rate of bacterial internalization or in the bacteria-induced expression of CD86 and HLA-DR by monocytes and myeloid dendritic cells were revealed, we found that the two strains significantly differ in inducing the production of cytokines involved in the adaptive immune response, as IL-1β, IL-23 and TNF-α, by antigen-presenting cells, with KKBO-1 being a more efficient inducer than KK207-1. The immune responses elicited by KK207-1 were comparable to those elicited by CIP 52.145, a highly virulent K. pneumoniae reference strain known to escape immune-inflammatory responses. Altogether, present results suggest that CG258-KP of the two clades are capable of inducing a different response of adaptive immunity in the human host.
Collapse
Affiliation(s)
- Ann Maria Clemente
- Department of Experimental and Clinical Medicine, University of Firenze, Firenze, Italy
| | - Giuseppe Castronovo
- Department of Experimental and Clinical Medicine, University of Firenze, Firenze, Italy
| | - Alberto Antonelli
- Department of Experimental and Clinical Medicine, University of Firenze, Firenze, Italy
| | - Marco Maria D’Andrea
- Department of Experimental and Clinical Medicine, University of Firenze, Firenze, Italy
- Department of Medical Biotechnologies, University of Siena, Siena, Italy
| | - Michele Tanturli
- Department of Experimental and Clinical Biomedical Sciences, University of Firenze, Firenze, Italy
| | - Eloisa Perissi
- Department of Experimental and Clinical Medicine, University of Firenze, Firenze, Italy
| | - Sara Paccosi
- Department of Health Science, University of Firenze, Firenze, Italy
| | - Astrid Parenti
- Department of Health Science, University of Firenze, Firenze, Italy
| | - Federico Cozzolino
- Department of Experimental and Clinical Biomedical Sciences, University of Firenze, Firenze, Italy
| | - Gian Maria Rossolini
- Department of Experimental and Clinical Medicine, University of Firenze, Firenze, Italy
- Clinical Microbiology and Virology Unit, Careggi University Hospital, Firenze, Italy
| | - Maria Gabriella Torcia
- Department of Experimental and Clinical Medicine, University of Firenze, Firenze, Italy
- * E-mail:
| |
Collapse
|
|
19
|
Myrrh exerts barrier-stabilising and -protective effects in HT-29/B6 and Caco-2 intestinal epithelial cells. Int J Colorectal Dis 2017; 32:623-634. [PMID: 27981377 DOI: 10.1007/s00384-016-2736-x] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Accepted: 12/05/2016] [Indexed: 02/04/2023]
Abstract
PURPOSE Myrrh, the oleo-gum resin of Commiphora molmol, is well known for its anti-inflammatory properties. In different animal models, it protected against DSS-, TNBS- and oxazolone-induced colitis. To date, no information concerning the effect of myrrh on barrier properties are available. Thus, this study investigates the effect of myrrh on paracellular barrier function in the absence or presence of the pro-inflammatory cytokine TNFα. METHODS Monolayers of human colon cell lines HT-29/B6 and Caco-2 were incubated with myrrh under control conditions or after challenge with the pro-inflammatory cytokine TNFα. Barrier function was analysed by electrophysiological and permeability measurements, Western blotting, immunostaining in combination with confocal microscopy, and freeze-fracture electron microscopy. RESULTS In Caco-2 cells, myrrh induced an increase in transepithelial resistance (TER) which was associated with downregulation of the channel-forming tight junction (TJ) protein claudin-2 via inhibition of the PI3 kinase signalling pathway. In HT-29/B6 cells, myrrh had no effect on barrier properties under basic conditions, but protected against barrier damage induced by TNFα, as indicated by a decrease in TER and an increase in fluorescein permeability. The TNFα effect was associated with a redistribution of the sealing TJ protein claudin-1, an increase in the expression of claudin-2 and a change in TJ ultrastructure. Most importantly, all TNFα effects were inhibited by myrrh. The effect of myrrh on claudin-2 expression in this cell line was mediated via inhibition of the STAT6 pathway. CONCLUSIONS This study shows for the first time that myrrh exerts barrier-stabilising and TNFα-antagonising effects in human intestinal epithelial cell models via inhibition of PI3K and STAT6 signalling. This suggests therapeutic application of myrrh in intestinal diseases associated with barrier defects and inflammation.
Collapse
|
|
20
|
Cao Y, Wu BJ, Zheng WP, Yin ML, Liu T, Song HL. Effect of heme oxygenase-1 transduced bone marrow mesenchymal stem cells on damaged intestinal epithelial cells in vitro. Cell Biol Int 2017; 41:726-738. [PMID: 28206713 DOI: 10.1002/cbin.10749] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2016] [Accepted: 02/13/2017] [Indexed: 12/21/2022]
Abstract
In this study, we explored the effects of mesenchymal stem cells (MSCs) from bone marrow overexpressing heme oxygenase-1 (HO-1) on the damaged human intestinal epithelial barrier in vitro. Rat MSCs were isolated from bone marrow and transduced with rat HO-1 recombinant adenovirus (HO-MSCs) for stable expression of HO-1. Colorectal adenocarinoma 2 (Caco2) cells were treated with tumor necrosis factor-α (TNF-α) to establish a damaged colon epithelial model. Damaged Caco2 were cocultured with MSCs, Ad-MSCs, Ad-HO + MSCs or HO-MSCs. mRNA and protein expression of Zona occludens-1 (ZO-1) and human HO-1 and the release of cytokines were measured. ZO-1 and human HO-1 in Caco2 were significantly decreased after treatment with TNF-α; and this effect was reduced when coculture with MSCs from bone marrow. Expression of ZO-1 was not significantly affected by Caco2 treatment with TNF-α, Ad-HO, and MSCs. In contrast, ZO-1 and human HO-1 increased significantly when the damaged Caco2 was treated with HO-MSCs. HO-MSCs showed the strongest effect on the expression of ZO-1 in colon epithelial cells. Coculture with HO-MSCs showed the most significant effects on reducing the expression of IL-2, IL-6, IFN-γ and increasing the expression of IL-10. HO-MSCs protected the intestinal epithelial barrier, in which endogenous HO-1 was involved. HO-MSCs play an important role in the repair process by reducing the release of inflammatory cytokines and increasing the release of anti-inflammatory factors. These results suggested that HO-MSCs from bone marrow were more effective in repairing the damaged intestinal epithelial barrier, and the effectiveness of MSCs was improved by HO-1 gene transduction, which provides favorable support for the application of stem cell therapy in the intestinal diseases.
Collapse
Affiliation(s)
- Yi Cao
- Tianjin First Central Hospital Clinic Institute, Tianjin Medical University, Tianjin, China
| | - Ben-Juan Wu
- Department of Organ Transplantation, Tianjin First Central Hospital, Tianjin, China
| | - Wei-Ping Zheng
- Department of Organ Transplantation, Tianjin First Central Hospital, Tianjin, China
| | - Ming-Li Yin
- Tianjin First Central Hospital Clinic Institute, Tianjin Medical University, Tianjin, China
| | - Tao Liu
- Key Laboratory of Emergency Care Medicine of Ministry of Health, Tianjin First Central Hospital, Tianjin, China
| | - Hong-Li Song
- Department of Organ Transplantation, Tianjin First Central Hospital, Tianjin, China.,Tianjin Key Laboratory of Organ Transplantation, Tianjin, China
| |
Collapse
|
|
21
|
Luettig J, Rosenthal R, Lee IFM, Krug SM, Schulzke JD. The ginger component 6-shogaol prevents TNF-α-induced barrier loss via inhibition of PI3K/Akt and NF-κB signaling. Mol Nutr Food Res 2016; 60:2576-2586. [PMID: 27487982 DOI: 10.1002/mnfr.201600274] [Citation(s) in RCA: 53] [Impact Index Per Article: 5.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/30/2016] [Revised: 06/20/2016] [Accepted: 07/21/2016] [Indexed: 12/26/2022]
Abstract
SCOPE Anti-inflammatory properties of the ginger-derived pungent component 6-shogaol (6-SG) have been studied intensively in recent years. Purpose of this study was to characterize the influence of 6-SG on inflammation-related intestinal barrier dysfunction, especially its paracellular component. METHODS AND RESULTS The effect of 6-SG was studied in the human intestinal cell models HT-29/B6 and Caco-2 either under control conditions or challenged by the pro-inflammatory cytokine tumor necrosis factor α (TNF-α). Electrophysiological measurements, freeze-fracture electron microscopy, and protein analyses were performed. 6-SG partially prevented both, the TNF-α-induced decrease in transepithelial resistance and the rise in fluorescein permeability. By inhibiting phosphatidylinositol-3-kinase/Akt signaling 6-SG prevented the TNF-α-induced increase in protein expression of claudin-2, a channel-forming tight junction protein. In addition, the TNF-α-induced disassembly of the sealing tight junction protein claudin-1 was attenuated, the latter of which was due to TNF-α-triggered phosphorylation of nuclear factor kappa light chain enhancer of activated B cells (NF-κB). CONCLUSION 6-SG has barrier-protective effects by affecting TNF-α-induced claudin-2 upregulation and claudin-1 disassembly via inhibition of phoshatidylinositol-3-kinase/Akt and nuclear factor kappa light chain enhancer of activated B-cell signaling. Therefore, 6-SG-containing food might be beneficial for barrier preservation during intestinal inflammation.
Collapse
Affiliation(s)
- Julia Luettig
- Nutritional Medicine and Clinical Physiology, Department of Gastroenterology, Rheumatology and Infectious Diseases, Charité-Universitätsmedizin Berlin, Campus Benjamin Franklin, Berlin, Germany
| | - Rita Rosenthal
- Nutritional Medicine and Clinical Physiology, Department of Gastroenterology, Rheumatology and Infectious Diseases, Charité-Universitätsmedizin Berlin, Campus Benjamin Franklin, Berlin, Germany
| | - In-Fah M Lee
- Nutritional Medicine and Clinical Physiology, Department of Gastroenterology, Rheumatology and Infectious Diseases, Charité-Universitätsmedizin Berlin, Campus Benjamin Franklin, Berlin, Germany
| | - Susanne M Krug
- Nutritional Medicine and Clinical Physiology, Department of Gastroenterology, Rheumatology and Infectious Diseases, Charité-Universitätsmedizin Berlin, Campus Benjamin Franklin, Berlin, Germany
| | - Jörg D Schulzke
- Nutritional Medicine and Clinical Physiology, Department of Gastroenterology, Rheumatology and Infectious Diseases, Charité-Universitätsmedizin Berlin, Campus Benjamin Franklin, Berlin, Germany
| |
Collapse
|
|
22
|
Maher S, Mrsny RJ, Brayden DJ. Intestinal permeation enhancers for oral peptide delivery. Adv Drug Deliv Rev 2016; 106:277-319. [PMID: 27320643 DOI: 10.1016/j.addr.2016.06.005] [Citation(s) in RCA: 248] [Impact Index Per Article: 27.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/29/2016] [Revised: 06/07/2016] [Accepted: 06/09/2016] [Indexed: 12/15/2022]
Abstract
Intestinal permeation enhancers (PEs) are one of the most widely tested strategies to improve oral delivery of therapeutic peptides. This article assesses the intestinal permeation enhancement action of over 250 PEs that have been tested in intestinal delivery models. In depth analysis of pre-clinical data is presented for PEs as components of proprietary delivery systems that have progressed to clinical trials. Given the importance of co-presentation of sufficiently high concentrations of PE and peptide at the small intestinal epithelium, there is an emphasis on studies where PEs have been formulated with poorly permeable molecules in solid dosage forms and lipoidal dispersions.
Collapse
|
|
23
|
Seidelin JB. Regulation of antiapoptotic and cytoprotective pathways in colonic epithelial cells in ulcerative colitis. Scand J Gastroenterol 2016; 50 Suppl 1:1-29. [PMID: 26513451 DOI: 10.3109/00365521.2016.1101245] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/04/2023]
Abstract
Ulcerative colitis is an inflammatory bowel disease involving the colon resulting in bloody diarrhea and increased risk of colorectal cancer in certain patient subgroups. Increased apoptosis in the epithelial cell layer causes increased permeability, especially during flares; this leads to translocation of luminal pathogens resulting in a continued inflammatory drive. The present work investigates how epithelial apoptosis is regulated in ulcerative colitis. The main results are that Fas mediated apoptosis is inhibited during flares of ulcerative colitis, probably by an upregulation of cellular inhibitor of apoptosis protein 2 (cIAP2) and cellular FLICE-like inhibitory protein. cIAP2 is upregulated in regenerative epithelial cells both in ulcerative colitis and in experimental intestinal wounds. Inhibition of cIAP2 decreases wound healing in vitro possibly through inhibition of migration. Altogether, it is shown that epithelial cells in ulcerative colitis responds to the hostile microenvironment by activation of cytoprotective pathways that tend to counteract the cytotoxic effects of inflammation. However, the present studies also show that epithelial cells produce increased amounts of reactive oxygen species during stimulation with tumor necrosis factor-α and interferon-γ resulting in DNA instability. The combined effect of increased DNA-instability and decreased apoptosis responses could lead to neoplasia.
Collapse
Affiliation(s)
- Jakob B Seidelin
- a Department of Gastroenterology, Medical Section , Herlev Hospital, University of Copenhagen , Herlev , Denmark
| |
Collapse
|
|
24
|
Tugizov S. Human immunodeficiency virus-associated disruption of mucosal barriers and its role in HIV transmission and pathogenesis of HIV/AIDS disease. Tissue Barriers 2016; 4:e1159276. [PMID: 27583187 DOI: 10.1080/21688370.2016.1159276] [Citation(s) in RCA: 47] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/07/2016] [Revised: 02/22/2016] [Accepted: 02/23/2016] [Indexed: 12/20/2022] Open
Abstract
Oral, intestinal and genital mucosal epithelia have a barrier function to prevent paracellular penetration by viral, bacterial and other pathogens, including human immunodeficiency virus (HIV). HIV can overcome these barriers by disrupting the tight and adherens junctions of mucosal epithelia. HIV-associated disruption of epithelial junctions may also facilitate paracellular penetration and dissemination of other viral pathogens. This review focuses on possible molecular mechanisms of HIV-associated disruption of mucosal epithelial junctions and its role in HIV transmission and pathogenesis of HIV and acquired immune deficiency syndrome (AIDS).
Collapse
Affiliation(s)
- Sharof Tugizov
- Department of Medicine, School of Medicine, University of California, San Francisco, San Francisco, CA, USA; Department of Orofacial Science, School of Dentistry, University of California, San Francisco, San Francisco, CA, USA
| |
Collapse
|
|
25
|
Ribet D, Cossart P. How bacterial pathogens colonize their hosts and invade deeper tissues. Microbes Infect 2015; 17:173-83. [PMID: 25637951 DOI: 10.1016/j.micinf.2015.01.004] [Citation(s) in RCA: 470] [Impact Index Per Article: 47.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/25/2014] [Revised: 01/18/2015] [Accepted: 01/19/2015] [Indexed: 02/06/2023]
Abstract
Bacterial pathogens have evolved a wide range of strategies to colonize and invade human organs, despite the presence of multiple host defense mechanisms. In this review, we will describe how pathogenic bacteria can adhere and multiply at the surface of host cells, how some bacteria can enter and proliferate inside these cells, and finally how pathogens may cross epithelial or endothelial host barriers and get access to internal tissues, leading to severe diseases in humans.
Collapse
Affiliation(s)
- David Ribet
- Institut Pasteur, Unité des Interactions Bactéries-Cellules, Département de Biologie Cellulaire et Infection, F-75015 Paris, France; INSERM, U604, F-75015 Paris, France; INRA, USC2020, F-75015 Paris, France.
| | - Pascale Cossart
- Institut Pasteur, Unité des Interactions Bactéries-Cellules, Département de Biologie Cellulaire et Infection, F-75015 Paris, France; INSERM, U604, F-75015 Paris, France; INRA, USC2020, F-75015 Paris, France.
| |
Collapse
|
|
26
|
Shi S, Wang H, Gao H, Li Z, Chen FX, Zuo XL, Li YQ. Increased gap density predicts weakness of the epithelial barrier in vivo by confocal laser endomicroscopy in indomethacin-induced enteropathy. Dig Dis Sci 2014; 59:1398-405. [PMID: 24573719 DOI: 10.1007/s10620-014-3076-8] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/14/2013] [Accepted: 02/11/2014] [Indexed: 01/26/2023]
Abstract
BACKGROUND AND AIMS The intestinal epithelial barrier plays an important role in the pathogenesis of non-steroidal anti-inflammatory drug-induced enteropathy, and its disruption is often associated with increased cell shedding. The purpose of this report is to observe the gap density in indomethacin-induced small intestinal damage by confocal laser endomicroscopy (CLE) and to investigate the mechanisms involved in this process and how mucosal protectants improve intestinal epithelial barrier dysfunction. CLE is expected to provide a new way for evaluating non-steroidal anti-inflammatory drugs-induced enteropathy in humans and assessing drug efficacy. METHODS Using the new technique of CLE, we established a method to evaluate, in real time, intestinal damage after the administration of indomethacin in Wistar rats by investigating the gap density in the small intestine. The mucosal protectant teprenone and acid-suppressant rabeprazole were then given by gavage before and after the administration of indomethacin, and the mechanisms affecting the intestinal epithelial barrier were investigated. RESULTS Using CLE, gaps could be clearly observed and easily distinguished from goblet cells. Gap density was increased after the administration of indomethacin. During this process, the expression of tumor necrosis factor-α, nuclear factor-κB, and caspase-3 was up-regulated and the expression of tight junctions was down-regulated, which led to the damage of the epithelial barrier. Teprenone and rabeprazole could intervene in this pathway and protect the integrity of the epithelial barrier. CONCLUSIONS CLE can be objective, accurate, and real time in investigating gap density. Teprenone and rabeprazole can prevent indomethacin-induced intestinal lesions and protect the epithelial barrier by intervening in the tumor necrosis factor-α pathway. Gap density was expected to be an indicator of evaluating intestinal inflammation and drug efficacy.
Collapse
Affiliation(s)
- Sha Shi
- Department of Gastroenterology, Qilu Hospital, Shandong University, No. 107, Wenhuaxi Road, Jinan, 250012, People's Republic of China,
| | | | | | | | | | | | | |
Collapse
|
|
27
|
Piegholdt S, Pallauf K, Esatbeyoglu T, Speck N, Reiss K, Ruddigkeit L, Stocker A, Huebbe P, Rimbach G. Biochanin A and prunetin improve epithelial barrier function in intestinal CaCo-2 cells via downregulation of ERK, NF-κB, and tyrosine phosphorylation. Free Radic Biol Med 2014; 70:255-64. [PMID: 24631489 DOI: 10.1016/j.freeradbiomed.2014.02.025] [Citation(s) in RCA: 45] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/23/2013] [Revised: 02/14/2014] [Accepted: 02/25/2014] [Indexed: 01/10/2023]
Abstract
The single-layered gut epithelium represents the primary line of defense against environmental stressors; thereby monolayer integrity and tightness are essentially required to maintain gut health and function. To date only a few plant-derived phytochemicals have been described as affecting intestinal barrier function. We investigated the impact of 28 secondary plant compounds on the barrier function of intestinal epithelial CaCo-2/TC-7 cells via transepithelial electrical resistance (TEER) measurements. Apart from genistein, the compounds that had the biggest effect in the TEER measurements were biochanin A and prunetin. These isoflavones improved barrier tightness by 36 and 60%, respectively, compared to the untreated control. Furthermore, both isoflavones significantly attenuated TNFα-dependent barrier disruption, thereby maintaining a high barrier resistance comparable to nonstressed cells. In docking analyses exploring the putative interaction with the tyrosine kinase EGFR, these novel modulators of barrier tightness showed very similar values compared to the known tyrosine kinase inhibitor genistein. Both biochanin A and prunetin were also identified as potent reducers of NF-κB and ERK activation, zonula occludens 1 tyrosine phosphorylation, and metalloproteinase-mediated shedding activity, which may account for the barrier-improving ability of these isoflavones.
Collapse
Affiliation(s)
- Stefanie Piegholdt
- Institute of Human Nutrition and Food Science, Christian-Albrechts-University Kiel, D-24118 Kiel, Germany
| | - Kathrin Pallauf
- Institute of Human Nutrition and Food Science, Christian-Albrechts-University Kiel, D-24118 Kiel, Germany
| | - Tuba Esatbeyoglu
- Institute of Human Nutrition and Food Science, Christian-Albrechts-University Kiel, D-24118 Kiel, Germany
| | - Nancy Speck
- Department of Dermatology and Allergology, University Hospital Schleswig-Holstein, Campus Kiel, D-24105 Kiel, Germany
| | - Karina Reiss
- Department of Dermatology and Allergology, University Hospital Schleswig-Holstein, Campus Kiel, D-24105 Kiel, Germany
| | - Lars Ruddigkeit
- Department of Chemistry and Biochemistry, University of Bern, CH-3012 Bern, Switzerland
| | - Achim Stocker
- Department of Chemistry and Biochemistry, University of Bern, CH-3012 Bern, Switzerland
| | - Patricia Huebbe
- Institute of Human Nutrition and Food Science, Christian-Albrechts-University Kiel, D-24118 Kiel, Germany
| | - Gerald Rimbach
- Institute of Human Nutrition and Food Science, Christian-Albrechts-University Kiel, D-24118 Kiel, Germany.
| |
Collapse
|
|
28
|
Schmid T, Bogdan M, Günzel D. Discerning apical and basolateral properties of HT-29/B6 and IPEC-J2 cell layers by impedance spectroscopy, mathematical modeling and machine learning. PLoS One 2013; 8:e62913. [PMID: 23840862 PMCID: PMC3698131 DOI: 10.1371/journal.pone.0062913] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/15/2012] [Accepted: 03/26/2013] [Indexed: 11/19/2022] Open
Abstract
Quantifying changes in partial resistances of epithelial barriers in vitro is a challenging and time-consuming task in physiology and pathophysiology. Here, we demonstrate that electrical properties of epithelial barriers can be estimated reliably by combining impedance spectroscopy measurements, mathematical modeling and machine learning algorithms. Conventional impedance spectroscopy is often used to estimate epithelial capacitance as well as epithelial and subepithelial resistance. Based on this, the more refined two-path impedance spectroscopy makes it possible to further distinguish transcellular and paracellular resistances. In a next step, transcellular properties may be further divided into their apical and basolateral components. The accuracy of these derived values, however, strongly depends on the accuracy of the initial estimates. To obtain adequate accuracy in estimating subepithelial and epithelial resistance, artificial neural networks were trained to estimate these parameters from model impedance spectra. Spectra that reflect behavior of either HT-29/B6 or IPEC-J2 cells as well as the data scatter intrinsic to the used experimental setup were created computationally. To prove the proposed approach, reliability of the estimations was assessed with both modeled and measured impedance spectra. Transcellular and paracellular resistances obtained by such neural network-enhanced two-path impedance spectroscopy are shown to be sufficiently reliable to derive the underlying apical and basolateral resistances and capacitances. As an exemplary perturbation of pathophysiological importance, the effect of forskolin on the apical resistance of HT-29/B6 cells was quantified.
Collapse
Affiliation(s)
- Thomas Schmid
- Department of Mathematics and Computer Science, Universität Leipzig, Leipzig, Germany
| | - Martin Bogdan
- Department of Mathematics and Computer Science, Universität Leipzig, Leipzig, Germany
| | - Dorothee Günzel
- Institute of Clinical Physiology, Charité Universitätsmedizin Berlin, Berlin, Germany
- * E-mail:
| |
Collapse
|
|
29
|
Vandenbroucke RE, Dejonckheere E, Van Hauwermeiren F, Lodens S, De Rycke R, Van Wonterghem E, Staes A, Gevaert K, López-Otin C, Libert C. Matrix metalloproteinase 13 modulates intestinal epithelial barrier integrity in inflammatory diseases by activating TNF. EMBO Mol Med 2013; 5:1000-16. [PMID: 23723167 PMCID: PMC3721470 DOI: 10.1002/emmm.201202100] [Citation(s) in RCA: 102] [Impact Index Per Article: 8.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/29/2012] [Revised: 04/09/2013] [Accepted: 04/11/2013] [Indexed: 12/19/2022] Open
Abstract
Several pathological processes, such as sepsis and inflammatory bowel disease (IBD), are associated with impairment of intestinal epithelial barrier. Here, we investigated the role of matrix metalloproteinase MMP13 in these diseases. We observed that MMP13−/− mice display a strong protection in LPS- and caecal ligation and puncture-induced sepsis. We could attribute this protection to reduced LPS-induced goblet cell depletion, endoplasmic reticulum stress, permeability and tight junction destabilization in the gut of MMP13−/− mice compared to MMP13+/+ mice. Both in vitro and in vivo, we found that MMP13 is able to cleave pro-TNF into bioactive TNF. By LC-MS/MS, we identified three MMP13 cleavage sites, which proves that MMP13 is an alternative TNF sheddase next to the TNF converting enzyme TACE. Similarly, we found that the same mechanism was responsible for the observed protection of the MMP13−/− mice in a mouse model of DSS-induced colitis. We identified MMP13 as an important mediator in sepsis and IBD via the shedding of TNF. Hence, we propose MMP13 as a novel drug target for diseases in which damage to the gut is essential.
Collapse
|
|
30
|
Xie F, Sakwiwatkul K, Zhang C, Wang Y, Zhai L, Hu S. Atractylodis macrocephalae Koidz. polysaccharides enhance both serum IgG response and gut mucosal immunity. Carbohydr Polym 2013; 91:68-73. [DOI: 10.1016/j.carbpol.2012.07.083] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/07/2012] [Revised: 07/14/2012] [Accepted: 07/31/2012] [Indexed: 10/28/2022]
|
|
31
|
Moran GW, O'Neill C, McLaughlin JT. GLP-2 enhances barrier formation and attenuates TNFα-induced changes in a Caco-2 cell model of the intestinal barrier. ACTA ACUST UNITED AC 2012; 178:95-101. [PMID: 22809889 DOI: 10.1016/j.regpep.2012.07.002] [Citation(s) in RCA: 40] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2011] [Revised: 01/30/2012] [Accepted: 07/05/2012] [Indexed: 11/29/2022]
Abstract
INTRODUCTION Tight junctions are intercellular permeability seals that regulate paracellular transport across epithelia. Tight junction function, expression and localisation of constituent proteins are significantly altered by cytokines such as TNFα. Glucagon-like peptide-2 (GLP-2) is an intestinotrophic enteroendocrine peptide. It is not known whether GLP-2 regulates the barrier or tight junctions. The aim of this study was to investigate whether GLP-2 has an effect on tight junction function or protein expression, alone or in response to TNFα exposure. METHODS Caco-2 cells were grown to confluence on filters in the presence or absence of GLP-2. The time course of transepithelial electrical resistance developing across the monolayer was measured; tight junction protein expression was quantified by immunoblotting. At day 20, TNFα in the presence or absence of GLP-2 was added. Changes in TEER and tight junction proteins expression were quantified. Both TNFα and GLP-2 were added on the basolateral side. RESULTS GLP-2 exposed Caco-2 cell monolayers showed a significant increase in transepithelial electrical resistance compared to that in untreated control cells. At the same time, expression of the tight junction proteins occludin and zona occludens-1 (ZO-1) was increased at day 17 post-seeding (1.6-fold; p=0.037 and 4.7 fold; p=0.039 respectively). Subsequent TNFα exposure induced a significant 9.3-fold (p<0.001) decrease in transepithelial electrical resistance and a corresponding reduction in the expression of ZO-1 (5.3 fold; p<0.01). However, the TNFα-induced reduction in transepithelial electrical resistance in GLP-2-exposed cells was highly attenuated to 1.8-fold (p<0.01). No change in tight junction protein expression was noted in GLP-2 exposed cells after cytokine exposure. CONCLUSION GLP-2 enhances formation of the epithelial barrier and its constituent proteins in Caco-2 cells, and diminishes the effects of TNFα. If these effects are replicated in vivo the GLP-2 receptor may present a therapeutic target in intestinal inflammation.
Collapse
Affiliation(s)
- G W Moran
- Inflammation Sciences Research Group, University of Manchester, Manchester, M13 9PL, UK.
| | | | | |
Collapse
|
|
32
|
SHIN DONGYEOK, KIM GIYOUNG, KIM CHANGIL, KIM WUNJAE, KANG HOSUNG, CHOI YUNGHYUN. Anti-invasive effects of decitabine, a DNA methyltransferase inhibitor, through tightening of tight junctions and inhibition of matrix metalloproteinase activities in AGS human gastric carcinoma cells. Oncol Rep 2012; 28:1043-50. [DOI: 10.3892/or.2012.1858] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/20/2012] [Accepted: 04/27/2012] [Indexed: 11/06/2022] Open
|
|
33
|
Stenman LK, Holma R, Korpela R. High-fat-induced intestinal permeability dysfunction associated with altered fecal bile acids. World J Gastroenterol 2012; 18:923-9. [PMID: 22408351 PMCID: PMC3297051 DOI: 10.3748/wjg.v18.i9.923] [Citation(s) in RCA: 145] [Impact Index Per Article: 11.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/01/2011] [Revised: 09/20/2011] [Accepted: 01/18/2012] [Indexed: 02/06/2023] Open
Abstract
AIM: To investigate whether high-fat-feeding is associated with increased intestinal permeability via alterations in bile acid metabolism.
METHODS: Male C57Bl/6J mice were fed on a high-fat (n = 26) or low-fat diet (n = 24) for 15 wk. Intestinal permeability was measured from duodenum, jejunum, ileum and colon in an Ussing chamber system using 4 kDa FITC-labeled dextran as an indicator. Fecal bile acids were analyzed with gas chromatography. Segments of jejunum and colon were analyzed for the expression of farnesoid X receptor (FXR) and tumor necrosis factor (TNF).
RESULTS: Intestinal permeability was significantly increased by high-fat feeding in jejunum (median 0.334 for control vs 0.393 for high-fat, P = 0.03) and colon (0.335 for control vs 0.433 for high-fat, P = 0.01), but not in duodenum or ileum. The concentration of nearly all identified bile acids was significantly increased by high-fat feeding (P < 0.001). The proportion of ursodeoxycholic acid (UDCA) in all bile acids was decreased (1.4% ± 0.1% in high-fat vs 2.8% ± 0.3% in controls, P < 0.01) and correlated inversely with intestinal permeability (r = -0.72, P = 0.01). High-fat feeding also increased jejunal FXR expression, as well as TNF expression along the intestine, especially in the colon.
CONCLUSION: High-fat-feeding increased intestinal permeability, perhaps by a mechanism related to bile acid metabolism, namely a decreased proportion of fecal UDCA and increased FXR expression.
Collapse
|
|
34
|
Ocampo SM, Romero C, Aviñó A, Burgueño J, Gassull MA, Bermúdez J, Eritja R, Fernandez E, Perales JC. Functionally enhanced siRNA targeting TNFα attenuates DSS-induced colitis and TLR-mediated immunostimulation in mice. Mol Ther 2011; 20:382-90. [PMID: 22044934 DOI: 10.1038/mt.2011.236] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023] Open
Abstract
Tumor necrosis factor (TNFα) is a proinflammatory cytokine involved in the pathogenesis of inflammatory bowel disease (IBD). Although TNFα has been extensively targeted using systemic drugs, the use of antisense and small interfering RNA (siRNA) to drive down its expression at the site of inflammation should provide important advantages. In this study, native and chemically modified siRNA against TNFα was developed and characterized using a murine model of IBD. siRNA with 2'-O-methyl and propanediol modifications (siTNF-OMe-P) were resistant to nuclease degradation and provided better silencing efficacy in vitro as compared to unmodified siRNA. Every modification reduced nonspecific Toll-like receptor (TLR)-mediated immunomodulation in human peripheral blood mononuclear cells (PBMC) cells. Intrarectal administration of siTNF-OMe-P significantly ameliorated the clinical endpoints and histopathological severity in 5% dextran sulphate sodium (DSS)-treated mice as compared to unmodified and other chemically modified siRNAs. Differential gene expression assessed in siTNF-OMe-P-treated animals correlated with improved colon integrity and reduced TLR activation as compared to all treatment groups. All in all, this study demonstrates that propanediol and 2'-O-methyl modifications have profound functional consequences for siRNA efficacy in vivo. Consequently, this strategy has potential implications for therapeutic intervention in IBD and other diseases.
Collapse
Affiliation(s)
- Sandra M Ocampo
- Institute for Advanced Chemistry of Catalonia (IQAC), Spanish Research Council (CSIC), Barcelona, Spain
| | | | | | | | | | | | | | | | | |
Collapse
|
|
35
|
Park HS, Kim GY, Choi IW, Kim ND, Hwang HJ, Choi YW, Choi YH. Inhibition of matrix metalloproteinase activities and tightening of tight junctions by diallyl disulfide in AGS human gastric carcinoma cells. J Food Sci 2011; 76:T105-11. [PMID: 22417372 DOI: 10.1111/j.1750-3841.2011.02114.x] [Citation(s) in RCA: 28] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/14/2023]
Abstract
The effect of diallyl disulfide (DADS), a major component of an oil-soluble allyl sulfide garlic (Allium sativum) derivative, on the correlation between anti-invasive activity and tightening of tight junctions (TJs) was investigated in human gastric adenocarcinoma AGS cells. Our data indicated that the inhibitory effects of DADS on cell motility and invasiveness were found to be associated with increased tightness of the TJs, which was demonstrated by an increase in transepithelial electrical resistance. Activities of matrix metalloprotease (MMP)-2 and -9 in AGS cells were dose-dependently inhibited by treatment with DADS, and this was also correlated with a decrease in expression of their mRNA and proteins; however, tissue inhibitor of metalloproteinase (TIMP)-1 and -2 mRNA levels and proteins were increased. Additionally, immunoblotting results indicated that DADS repressed the levels of claudin proteins (claudin-2, -3, and -4), major components of TJs that play key roles in control and selectivity of paracellular transport. Although further studies are needed, these results suggest that DADS treatment may inhibit tumor cell motility and invasion and, therefore, act as a dietary source to decrease the risk of cancer metastasis.
Collapse
Affiliation(s)
- Hyun Soo Park
- Dept. of Pharmacy, Pusan Natl. Univ., Busan 609-735, Republic of Korea
| | | | | | | | | | | | | |
Collapse
|
|
36
|
Shin DY, Kim GY, Kim JI, Yoon MK, Kwon TK, Lee SJ, Choi YW, Kang HS, Yoo YH, Choi YH. Anti-invasive activity of diallyl disulfide through tightening of tight junctions and inhibition of matrix metalloproteinase activities in LNCaP prostate cancer cells. Toxicol In Vitro 2010; 24:1569-76. [PMID: 20600798 DOI: 10.1016/j.tiv.2010.06.014] [Citation(s) in RCA: 33] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/24/2010] [Revised: 06/22/2010] [Accepted: 06/25/2010] [Indexed: 01/25/2023]
Abstract
Diallyl disulfide (DADS) is a major component of an oil-soluble allyl sulfide garlic (Allium sativum) derivative, which has been shown to exert a potential for anti-cancer activity. However, the biochemical mechanisms underlying DADS-induced anti-invasiveness and anti-metastasis have not been thoroughly studied. In this study, we investigated the effect of DADS on the correlation between tightening of tight junctions (TJs) and anti-invasive activity in human prostate carcinoma LNCaP cells. Inhibitory effects of DADS on cell motility and invasiveness were found to be associated with increased tightness of the TJ, which was demonstrated by an increase in transepithelial electrical resistance (TER). Additionally, immunoblotting results indicated that DADS repressed the levels of the claudin proteins, which are major components of TJs that play a key role in control and selectivity of paracellular transport. Furthermore, the activities of matrix metalloproteinase (MMP)-2 and -9 in LNCaP cells were dose-dependently inhibited by treatment with DADS, and this was also correlated with a decrease in expression of their mRNA and proteins. Although further studies are needed, the present study indicates that TJs and MMPs are critical targets of DADS-induced anti-invasiveness in human prostate cancer LNCaP cells.
Collapse
Affiliation(s)
- Dong Yeok Shin
- Department of Biochemistry, Dongeui University College of Oriental Medicine, Busan, Republic of Korea
| | | | | | | | | | | | | | | | | | | |
Collapse
|
|
37
|
Cui W, Li LX, Sun CM, Wen Y, Zhou Y, Dong YL, Liu P. Tumor necrosis factor alpha increases epithelial barrier permeability by disrupting tight junctions in Caco-2 cells. Braz J Med Biol Res 2010; 43:330-7. [PMID: 20445948 DOI: 10.1590/s0100-879x2010007500020] [Citation(s) in RCA: 57] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/17/2009] [Accepted: 03/03/2010] [Indexed: 02/10/2023] Open
Abstract
The objectives of this study were to determine the effect of tumor necrosis factor alpha (TNF-alpha) on intestinal epithelial cell permeability and the expression of tight junction proteins. Caco-2 cells were plated onto Transwell microporous filters and treated with TNF-alpha (10 or 100 ng/mL) for 0, 4, 8, 16, or 24 h. The transepithelial electrical resistance and the mucosal-to-serosal flux rates of the established paracellular marker Lucifer yellow were measured in filter-grown monolayers of Caco-2 intestinal cells. The localization and expression of the tight junction protein occludin were detected by immunofluorescence and Western blot analysis, respectively. SYBR-Green-based real-time PCR was used to measure the expression of occludin mRNA. TNF-alpha treatment produced concentration- and time-dependent decreases in Caco-2 transepithelial resistance and increases in transepithelial permeability to the paracellular marker Lucifer yellow. Western blot results indicated that TNF-alpha decreased the expression of phosphorylated occludin in detergent-insoluble fractions but did not affect the expression of non-phosphorylated occludin protein. Real-time RT-PCR data showed that TNF-alpha did not affect the expression of occludin mRNA. Taken together, our data demonstrate that TNF-alpha increases Caco-2 monolayer permeability, decreases occludin protein expression and disturbs intercellular junctions.
Collapse
Affiliation(s)
- W Cui
- Department of Infectious Diseases, the First Affiliated Hospital, China Medical University, Shenyang, Liaoning, China
| | | | | | | | | | | | | |
Collapse
|
|
38
|
Abstract
OBJECTIVE Epithelial apoptosis rates are increased in ulcerative colitis (UC). The increased apoptosis rate could expose mucosal cells to luminal pathogens and thereby be regarded as a primary pathogenic factor in UC. On the other hand, the local inflammatory reaction could cause epithelial apoptosis secondary to the release of cytotoxic mediators. If apoptosis is a primary defect, apoptosis rates could influence the degree of spreading of inflammation and the clinical course of UC. If apoptosis is a side effect of local inflammation, apoptosis rates would be expected only to correlate with the degree of local inflammation. The aim of the study was to investigate the relationship between epithelial apoptosis and clinical characteristics of UC. MATERIAL AND METHODS Twenty patients with UC (12 with active disease) and 20 control subjects were included. Freshly isolated colonic epithelial cells were cultured. Apoptosis was determined by flow cytometry. Cells were stimulated with Fas ligand. The disease was characterized by endoscopic findings, microscopic inflammation grade, surrogate markers of disease activity (hemoglobin level, white blood cell count, C-reactive protein, or albumin), and the clinical course 6 months after biopsy. RESULTS Epithelial apoptosis correlated with local inflammation, both macroscopic (p< 0.02) and microscopic (p< 0.008). Disease extent, disease course, or surrogate markers of disease activity did not correlate with apoptosis rate. However, increased microscopic inflammation inversely correlated with apoptosis response to the Fas ligand (p< 0.06). CONCLUSIONS The epithelial apoptosis rate is influenced primarily by the local inflammatory response. Colonocytes upregulate cytoprotective mediators that decrease apoptosis susceptibility during active UC.
Collapse
Affiliation(s)
- Jakob Benedict Seidelin
- Department of Gastroenterology, Medical Section, Herlev Hospital, University of Copenhagen, Denmark.
| | | |
Collapse
|
|
39
|
Kim SO, Choi YH. The ethyl alcohol extract of Hizikia fusiforme inhibits matrix metalloproteinase activity and regulates tight junction related protein expression in Hep3B human hepatocarcinoma cells. J Med Food 2010; 13:31-8. [PMID: 20136433 DOI: 10.1089/jmf.2009.1233] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022] Open
Abstract
We tested the correlation between the tightness of tight junctions (TJs) and the anti-invasive activity of the ethyl alcohol extract of Hizikia fusiforme (EHF) in Hep3B human hepatocarcinoma cells. EHF inhibited cell proliferation, motility, and invasiveness, which were associated with increased TJ tightness, as demonstrated by an increase in transepithelial electrical resistance. EHF dose-dependently decreased the secretion of matrix metalloprotease-2 and -9, which correlated with a decrease in mRNA and protein expression, but increased tissue inhibitor of metalloproteinase-1 and -2 mRNA levels. Additionally, immunoblotting results indicated that EHF suppressed the major components of TJ, claudins (-1, -3, and -4), which play a key role in the control and selectivity of paracellular transport. These data indicate that EHF may inhibit cancer cell invasion through the tightening of TJs, which may counteract the up-regulation of claudins. Furthermore, EHF treatment decreased the expression of insulin-like growth factor-1 receptor proteins, while concurrently increasing that of thrombospondin-1 and E-cadherin. In conclusion, these results suggest that EHF treatment may inhibit tumor metastasis and invasion and therefore act as a dietary resource for decreasing the risk of developing cancer.
Collapse
Affiliation(s)
- Sung Ok Kim
- Department of Biomaterial Control (BK21 Program), Dongeui University Graduate School, Busan, Republic of Korea
| | | |
Collapse
|
|
40
|
Kayamuro H, Abe Y, Yoshioka Y, Katayama K, Nomura T, Yoshida T, Yamashita K, Yoshikawa T, Kawai Y, Mayumi T, Hiroi T, Itoh N, Nagano K, Kamada H, Tsunoda SI, Tsutsumi Y. The use of a mutant TNF-α as a vaccine adjuvant for the induction of mucosal immune responses. Biomaterials 2009; 30:5869-76. [DOI: 10.1016/j.biomaterials.2009.07.009] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/19/2009] [Accepted: 07/06/2009] [Indexed: 10/20/2022]
|
|
41
|
Theiss AL, Jenkins AK, Okoro NI, Klapproth JMA, Merlin D, Sitaraman SV. Prohibitin inhibits tumor necrosis factor alpha-induced nuclear factor-kappa B nuclear translocation via the novel mechanism of decreasing importin alpha3 expression. Mol Biol Cell 2009; 20:4412-23. [PMID: 19710421 DOI: 10.1091/mbc.e09-05-0361] [Citation(s) in RCA: 57] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/14/2023] Open
Abstract
Expression of prohibitin 1 (PHB), a multifunctional protein in the cell, is decreased during inflammatory bowel disease (IBD). Little is known regarding the regulation and role of PHB during intestinal inflammation. We examined the effect of tumor necrosis factor alpha (TNF-alpha), a cytokine that plays a central role in the pathogenesis of IBD, on PHB expression and the effect of sustained PHB expression on TNF-alpha activation of nuclear factor-kappa B (NF-kappaB) and epithelial barrier dysfunction, two hallmarks of intestinal inflammation. We show that TNF-alpha decreased PHB protein and mRNA abundance in intestinal epithelial cells in vitro and in colon mucosa in vivo. Sustained expression of prohibitin in intestinal epithelial cells in vitro and in vivo (prohibitin transgenic mice, PHB TG) resulted in a marked decrease in TNF-alpha-induced nuclear translocation of the NF-kappaB protein p65, NF-kappaB/DNA binding, and NF-kappaB-mediated transcriptional activation despite robust IkappaB-alpha phosphorylation and degradation and increased cytosolic p65. Cells overexpressing PHB were protected from TNF-alpha-induced increased epithelial permeability. Expression of importin alpha3, a protein involved in p50/p65 nuclear import, was decreased in cells overexpressing PHB and in colon mucosa of PHB TG mice. Restoration of importin alpha3 levels sustained NF-kappaB activation by TNF-alpha during PHB transfection. These results suggest that PHB inhibits NF-kappaB nuclear translocation via a novel mechanism involving alteration of importin alpha3 levels. TNF-alpha decreases PHB expression in intestinal epithelial cells and restoration of PHB expression in these cells can protect against the deleterious effects of TNF-alpha and NF-kappaB on barrier function.
Collapse
Affiliation(s)
- Arianne L Theiss
- Division of Digestive Diseases, Department of Medicine, Emory University, Atlanta, GA 30322, USA.
| | | | | | | | | | | |
Collapse
|
|
42
|
Choi YH, Choi WY, Hong SH, Kim SO, Kim GY, Lee WH, Yoo YH. Anti-invasive activity of sanguinarine through modulation of tight junctions and matrix metalloproteinase activities in MDA-MB-231 human breast carcinoma cells. Chem Biol Interact 2008; 179:185-91. [PMID: 19063874 DOI: 10.1016/j.cbi.2008.11.009] [Citation(s) in RCA: 28] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/04/2008] [Revised: 11/08/2008] [Accepted: 11/11/2008] [Indexed: 10/21/2022]
Abstract
Tight junctions (TJs) are critical structures for the maintenance of cellular polarity, acting as paracellular permeability barriers and playing an essential role in regulation of the diffusion of fluid, electrolytes and macromolecules through the paracellular pathway. Matrix metalloproteinases (MMPs) have been implicated as possible mediators of invasiveness and metastasis in some cancers. In this study, it was investigated the effect of sanguinarine, a benzophenanthridine alkaloid, on the correlation between the tightening of TJs and the anti-invasive activity in human breast carcinoma MDA-MB-231 cells. The inhibitory effects of sanguinarine on cell proliferation, motility and invasiveness were found to be associated with the increased tightness of the TJ, which was demonstrated by an increase in transepithelial electrical resistance (TER). Additionally, immunoblotting results indicated that sanguinarine repressed the levels of the claudin proteins, major components of TJs that play a key role in the control and selectivity of paracellular transport. Furthermore, the activities of MMP-2 and -9 in MDA-MB-231 cells were dose-dependently inhibited by treatment with sanguinarine, and this was also correlated with a decrease in the expression of their mRNA and proteins.
Collapse
Affiliation(s)
- Yung Hyun Choi
- Department of Biochemistry, Dongeui University College of Oriental Medicine, Busan 614-052, South Korea.
| | | | | | | | | | | | | |
Collapse
|
|
43
|
Wang Q, Guo XL, Wells-Byrum D, Noel G, Pritts TA, Ogle CK. Cytokine-induced epithelial permeability changes are regulated by the activation of the p38 mitogen-activated protein kinase pathway in cultured Caco-2 cells. Shock 2008; 29:531-7. [PMID: 17724435 DOI: 10.1097/shk.0b013e318150737f] [Citation(s) in RCA: 33] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
Increased intestinal/epithelial permeability in sepsis and endotoxemia has been noted to be induced by proinflammatory cytokines such as interferon-gamma, TNF-alpha, and IL-1beta. The p38 mitogen-activated protein kinase (MAPK) signaling pathway plays an important role in regulating the inflammatory response induced by these cytokines. We tested the hypothesis that epithelial permeability changes are regulated through the p38 MAPK signaling pathway. Caco-2 cells were cultured for 21 days and then stimulated with a cytokine mixture (CytoMix: TNF-alpha, interferon-gamma, and IL-1beta). Epithelial barrier function was evaluated by measuring permeability in an Ussing chamber. CytoMix-induced changes of MAPKs (p38, c-Jun amino-terminal kinase, and extracellular-regulated kinase), NO production, and inflammatory responses (IL-6 and IL-8 levels) were also assessed. The signaling pathways were further studied by pretreating cells with SB203580, a specific p38 MAPK inhibitor. CytoMix increased permeability at 24 and 48 h but not at 4 h. This was associated with increased IL-6 and IL-8 production, as well as increases in phosphorylation of all three MAPKs. Treatment with SB203580 completely blocked p38 activity with transient inhibition of p38 phosphorylation. SB203580 also prevented the CytoMix-induced permeability increase and reduced NO, IL-6, and IL-8 levels. The results suggest that p38 MAPK plays an important role in regulating epithelial barrier function during inflammation.
Collapse
Affiliation(s)
- Quan Wang
- Department of Research, Shriners Hospital for Children, Cincinnati OH 45229-3095, USA.
| | | | | | | | | | | |
Collapse
|
|
44
|
Li Q, Zhang Q, Wang M, Zhao S, Ma J, Luo N, Li N, Li Y, Xu G, Li J. Interferon-gamma and tumor necrosis factor-alpha disrupt epithelial barrier function by altering lipid composition in membrane microdomains of tight junction. Clin Immunol 2007; 126:67-80. [PMID: 17964857 DOI: 10.1016/j.clim.2007.08.017] [Citation(s) in RCA: 92] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/27/2006] [Revised: 08/14/2007] [Accepted: 08/20/2007] [Indexed: 11/15/2022]
Abstract
Tight junctions (TJs) are specialized membrane microdomains of plasma membrane and play an important role in barrier function. IFN-gamma and TNF-alpha have been implicated in intestinal barrier dysfunction. In the present study, we analyzed the effect of IFN-gamma and TNF-alpha on epithelial barrier function and determined the contribution of apoptosis to this process using T84 cells, a model intestinal epithelial cell line. We found that TNF-alpha and IFN-gamma synergistically affected the epithelial barrier and disrupted the structure of TJs. We demonstrated for the first time that treatment with TNF-alpha and IFN-gamma changed lipid composition and fatty acyl substitutions of phospholipids in membrane microdomains of TJs. Alterations of lipid environment affected TJs barrier function and partly removed flotillin-1 and displaced occludin from membrane microdomains of TJs to detergent-soluble fractions. The distribution of claudin isoforms was unaffected by TNF-alpha and IFN-gamma treatment. These findings indicated the interaction between inflammatory cytokines and alterations of lipid composition in membrane microdomains of TJs in the inflammatory processes. The apoptosis inhibitor did not prevent cytokine-induced decrease in TER and increase in permeability to FITC-dextran. Our results suggest that the cytokines directly influence TJ function and modulate both the membrane microdomain localization of TJ proteins and lipid composition of TJs. The effects of proinflammatory cytokines on TJ structure and function provide a mechanism in the pathophysiology of Crohn's disease (CD). Understanding the intracellular mechanisms involved could be important in devising future therapeutic strategies to induce retightening of the leaky TJ barrier.
Collapse
Affiliation(s)
- Qiurong Li
- Institute of General Surgery, Jinling Hospital, Nanjing, PR China
| | | | | | | | | | | | | | | | | | | |
Collapse
|
|
45
|
Jørgensen VL, Ibsen M, Andresen L, Schulzke JD, Perner A. Effects of endotoxaemia on markers of permeability, metabolism and inflammation in the large bowel of healthy subjects. Acta Anaesthesiol Scand 2007; 51:1085-92. [PMID: 17697304 DOI: 10.1111/j.1399-6576.2007.01381.x] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
BACKGROUND Increased permeability and increased luminal concentrations of L-lactate have previously been shown in the large bowel in septic patients. To advance these observations, a human model of colorectal barrier failure in sepsis is desirable. Therefore, we assessed the effects of endotoxaemia on markers of permeability, metabolism and inflammation in the large bowel in healthy subjects. METHODS Twelve healthy male subjects received intravenous endotoxin (2 ng/kg body weight) or saline in a paired cross-over design. Colorectal permeability was assessed after 3, 6, 9 and 12 h by the systemic recovery of luminally instilled (99m)Tc-diethylenetriaminepentaacetate. Luminal concentrations of L-lactate were assessed by equilibrium dialysis. Mucosal biopsies from the large bowel were sampled after 6 and 12 h, and the apoptotic ratio of the epithelium was assessed by terminal deoxynucleotidyl transferase-mediated desoxyuridinetriphosphate nick end-labelling (TUNEL) assay and the expression of inducible nitric oxide synthase (iNOS) mRNA by reverse transcriptase-polymerase chain reaction. RESULTS Systemic effects of endotoxaemia were observed, including fever, tachycardia and strongly increased plasma values of tumour necrosis factor-alpha. By contrast, the colorectal permeability, luminal lactate concentrations, mucosal infiltration of inflammatory cells, epithelial apoptotic ratio and expression of iNOS were all unaffected by endotoxin. CONCLUSIONS No effect of a single intravenous dose of endotoxin was observed on markers of large bowel permeability, metabolism and inflammation in healthy subjects. This suggests that this part of the gut is relatively resistant to the systemic inflammation induced by experimental endotoxaemia in humans.
Collapse
Affiliation(s)
- V L Jørgensen
- Department of Anaesthesia and Intensive Care, Herlev Hospital, University of Copenhagen, Copenhagen, Denmark
| | | | | | | | | |
Collapse
|
|
46
|
Al-Sadi RM, Ma TY. IL-1beta causes an increase in intestinal epithelial tight junction permeability. JOURNAL OF IMMUNOLOGY (BALTIMORE, MD. : 1950) 2007; 178:4641-9. [PMID: 17372023 PMCID: PMC3724221 DOI: 10.4049/jimmunol.178.7.4641] [Citation(s) in RCA: 459] [Impact Index Per Article: 25.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
Abstract
IL-1beta is a prototypical proinflammatory cytokine that plays a central role in the intestinal inflammation amplification cascade. Recent studies have indicated that a TNF-alpha- and IFN-gamma-induced increase in intestinal epithelial paracellular permeability may be an important mechanism contributing to intestinal inflammation. Despite its central role in promoting intestinal inflammation, the role of IL-1beta on intestinal epithelial tight junction (TJ) barrier function remains unclear. The major aims of this study were to determine the effect of IL-1beta on intestinal epithelial TJ permeability and to elucidate the mechanisms involved in this process, using a well-established in vitro intestinal epithelial model system consisting of filter-grown Caco-2 intestinal epithelial monolayers. IL-1beta (0-100 ng/ml) produced a concentration- and time-dependent decrease in Caco-2 transepithelial resistance. Conversely, IL-1beta caused a progressive time-dependent increase in transepithelial permeability to paracellular marker inulin. IL-1beta-induced increase in Caco-2 TJ permeability was accompanied by a rapid activation of NF-kappaB. NF-kappaB inhibitors, pyrrolidine dithiocarbamate and curcumin, prevented the IL-1beta-induced increase in Caco-2 TJ permeability. To further confirm the role of NF-kappaB in the IL-1beta-induced increase in Caco-2 TJ permeability, NF-kappaB p65 expression was silenced by small interfering RNA transfection. NF-kappaB p65 depletion completely inhibited the IL-1beta-induced increase in Caco-2 TJ permeability. IL-1beta did not induce apoptosis in the Caco-2 cell. In conclusion, our findings show for the first time that IL-1beta at physiologically relevant concentrations causes an increase in intestinal epithelial TJ permeability. The IL-1beta-induced increase in Caco-2 TJ permeability was mediated in part by the activation of NF-kappaB pathways but not apoptosis.
Collapse
Affiliation(s)
- Rana M. Al-Sadi
- Department of Internal Medicine, University of New Mexico School of Medicine, Albuquerque, NM 87131
| | - Thomas Y. Ma
- Department of Internal Medicine, University of New Mexico School of Medicine, Albuquerque, NM 87131
- Albuquerque Veterans Affairs Medical Center, Albuquerque, NM 87102
| |
Collapse
|
|
47
|
Van Limbergen J, Russell RK, Nimmo ER, Ho GT, Arnott ID, Wilson DC, Satsangi J. Genetics of the innate immune response in inflammatory bowel disease. Inflamm Bowel Dis 2007; 13:338-55. [PMID: 17206667 DOI: 10.1002/ibd.20096] [Citation(s) in RCA: 57] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
The discovery of nucleotide-binding oligomerization domain 2/caspase recruitment domain-containing protein 15 (NOD2/CARD15) as the first susceptibility gene in Crohn's disease (CD) has shifted the focus of research into the pathogenesis of inflammatory bowel disease (IBD) firmly to the innate immune response and the integrity of the epithelial barrier. The subsequent implication in IBD of variant alleles of OCTN, DLG5, MDR1, and TLRs has provided further support for a new, more complex model of innate immunity function in the gastrointestinal tract. In this review, we examine the recent advances in our understanding of the influence of genetics of the innate immune response on IBD. We will focus on germline variation of genes encoding pathogen-recognition receptors, proteins involved in epithelial homeostasis and secreted antimicrobial proteins.
Collapse
Affiliation(s)
- Johan Van Limbergen
- Gastrointestinal Unit, Molecular Medicine Centre, University of Edinburgh, Western General Hospital, Crewe Road South, Edinburgh EH4 2XU, UK.
| | | | | | | | | | | | | |
Collapse
|
|
48
|
Malo MS, Biswas S, Abedrapo MA, Yeh L, Chen A, Hodin RA. The pro-inflammatory cytokines, IL-1beta and TNF-alpha, inhibit intestinal alkaline phosphatase gene expression. DNA Cell Biol 2007; 25:684-95. [PMID: 17233117 DOI: 10.1089/dna.2006.25.684] [Citation(s) in RCA: 39] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/04/2023] Open
Abstract
High levels of the pro-inflammatory cytokines, interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha), are present in the gut mucosa of patients suffering form various diseases, most notably inflammatory bowel diseases (IBD). Since the inflammatory milieu can cause important alterations in epithelial cell function, we examined the cytokine effects on the expression of the enterocyte differentiation marker, intestinal alkaline phosphatase (IAP), a protein that detoxifies bacterial lipopolysaccharides (LPS) and limits fat absorption. Sodium butyrate (NaBu), a short-chain fatty acid and histone deacetylase (HDAC) inhibitor, was used to induce IAP expression in HT-29 cells and the cells were also treated +/- the cytokines. Northern blots confirmed IAP induction by NaBu, however, pretreatment (6 h) with either cytokine showed a dose-dependent inhibition of IAP expression. IAP Western analyses and alkaline phosphatase enzyme assays corroborated the Northern data and confirmed that the cytokines inhibit IAP induction. Transient transfections with a reporter plasmid carrying the human IAP promoter showed significant inhibition of NaBu-induced IAP gene activation by the cytokines (100 and 60% inhibition with IL-1beta and TNF-alpha, respectively). Western analyses showed that NaBu induced H4 and H3 histone acetylation, and pretreatment with IL-1beta or TNF-alpha did not change this global acetylation pattern. In contrast, chromatin immunoprecipitation showed that local histone acetylation of the IAP promoter region was specifically inhibited by either cytokine. We conclude that IL-1beta and TNF-alpha inhibit NaBu-induced IAP gene expression, likely by blocking the histone acetylation within its promoter. Cytokine-mediated IAP gene silencing may have important implications for gut epithelial function in the setting of intestinal inflammatory conditions.
Collapse
Affiliation(s)
- Madhu S Malo
- Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts 02114, USA
| | | | | | | | | | | |
Collapse
|
|
49
|
Abstract
A critical function of the intestinal mucosa is to form a barrier that separates luminal contents from the underlying interstitium. This intestinal barrier is primarily regulated by the apical junctional complex (AJC) consisting of tight junctions (TJs) and adherens junctions (AJs) and is compromised in a number of intestinal diseases, including inflammatory bowel disease (IBD). In vitro studies have demonstrated that proinflammatory cytokines, such as interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha), that are increased in the intestinal mucosa of patients with IBD can induce a leaky mucosal barrier. There is a growing evidence that the increased permeability and altered AJC structure observed in IBD are mediated by internalization of junctional proteins. This review summarizes barrier defects observed in IBD and addresses mechanisms by which proinflammatory cytokines, such as IFN-gamma and TNF-alpha, modulate AJC structure and epithelial barrier function.
Collapse
Affiliation(s)
- Matthias Bruewer
- Department of General Surgery, University of Muenster, Muenster 48149, Germany
| | | | | |
Collapse
|
|
50
|
Guttman JA, Samji FN, Li Y, Vogl AW, Finlay BB. Evidence that tight junctions are disrupted due to intimate bacterial contact and not inflammation during attaching and effacing pathogen infection in vivo. Infect Immun 2006; 74:6075-84. [PMID: 16954399 PMCID: PMC1695516 DOI: 10.1128/iai.00721-06] [Citation(s) in RCA: 70] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
It is widely accepted that tight junctions are altered during infections by attaching and effacing (A/E) pathogens. These disruptions have been demonstrated both in vitro and more recently in vivo. For in vivo experiments, the murine model of A/E infection with Citrobacter rodentium is the animal model of choice. In addition to effects on tight junctions, these bacteria also colonize the colon at high levels, efface colonocyte microvilli, and cause hyperplasia and inflammation. Although we have recently demonstrated that tight junctions are disrupted by C. rodentium, the issue of direct effects of bacteria on epithelial cell junctions versus the indirect effects of inflammation still remains to be clarified. Here, we demonstrate that during the C. rodentium infections, inflammation plays no discernible role in the alteration of tight junctions. The distribution of the tight junction proteins, claudin-1, -3, and -5, are unaffected in inflamed colon, and junctions appear morphologically unaltered when viewed by electron microscopy. Additionally, tracer molecules are not capable of penetrating the inflamed colonic epithelium of infected mice that have cleared the bacteria. Finally, infected colonocytes from mice exposed to C. rodentium for 14 days, which have high levels of bacterial attachment to colonocytes as well as inflammation, have characteristic, altered claudin localization whereas cells adjacent to infected colonocytes retain their normal claudin distribution. We conclude that inflammation plays no discernible role in tight junction alteration during A/E pathogenesis and that tight junction disruption in vivo appears dependent only on the direct intimate attachment of the pathogenic bacteria to the cells.
Collapse
Affiliation(s)
- Julian A Guttman
- The University of British Columbia, Michael Smith Laboratories, and Department of Cellular and Physiological Sciences, 301-2185 East Mall, Vancouver, BC, Canada V6T 1Z4
| | | | | | | | | |
Collapse
|