Acute necrotizing encephalopathy (ANE) secondary to influenza infection is characterized by fulminant neurological deterioration and a high mortality rate. The underlying mechanisms remain unclear, and specific treatments are currently lacking. Therefore, understanding the pathogenesis and identifying diagnostic and therapeutic targets for influenza-induced ANE are crucial. Peripheral blood samples were collected from two groups: influenza-infected patients without ANE (mild) and influenza infection with ANE patients (severe). Differentially expressed genes (DEG) were identified through microRNA sequencing analysis, followed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. The expression levels of the four specific miRNAs were validated using qRT-PCR. In the severe group, 24 genes were up-regulated, and 67 genes were down-regulated compared to the mild group. The expression levels of hsa-miR-1290, hsa-miR-4657, has-miR-1231, and hsa-miR-342-3p were validated by qRT-PCR, and the levels of has-miR-4657 and hsamiR- 342-3p showed significant differences between severe and mild groups. GO analysis demonstrated that the DEGs were predominantly involved in the positive regulation of cellular processes, intracellular anatomical structure, and protein binding. KEGG pathway analysis revealed that DEGs were mainly enriched in calcium signaling pathway and axon guidance. The down-regulated hsa-miR-4657 and hsa-miR-342-3p might be associated with the development of ANE in pediatric patients with influenza by regulation of calcium pathways and axon guidance.

Endothelial-to-mesenchymal transition (EndMT) is a biological process that converts endothelial cells to mesenchymal cells with increased proliferative and migrative abilities. EndMT has been implicated in the development of pulmonary vascular remodeling in pulmonary arterial hypertension (PAH), a fatal and progressive lung vascular disease. Transforming growth factor β1 (TGF-β1), an inflammatory cytokine, is known to induce EndMT in many types of endothelial cells including lung vascular endothelial cells (LVECs). An increase in cytosolic free Ca2+ concentration ([Ca2+]cyt) is a major stimulus for cellular proliferation and phenotypic transition, but it is unknown whether Ca2+ signaling is involved in EndMT. In this study, we tested the hypothesis that TGF-β1-induced EndMT in human LVEC is Ca2+-dependent. Treatment of LVEC with TGF-β1 for 5-7 days resulted in increase in SNAI1/2 expression, induction of EndMT, upregulation of STIM/Orai1, and enhancement of store-operated Ca2+ entry (SOCE). Removal (or chelation) of extracellular or intracellular Ca2+ with EGTA or BAPTA-AM, respectively, abolished EndMT in response to TGF-β1. Moreover, EGTA diminished TGF-β1-induced increase in SNAI in a dose-dependent manner. Knockdown of either STIM1 or Orai1 was sufficient to prevent TGF-β-mediated increase in SNAI1/2 and EndMT but did not rescue the continuous adherent junctions. Blockade of Orai1 channels by AnCoA4 inhibited TGF-β-mediated EndMT and restored PECAM1-positive continuous adherent junctions. In conclusion, intracellular Ca2+ signaling plays a critical role in TGF-β-associated EndMT through enhanced SOCE and STIM1-Orai1 interaction. Thus, targeting Ca2+ signaling pathways regulating EndMT may be a novel therapeutic approach to treat PAH and other forms of precapillary pulmonary hypertension.NEW & NOTEWORTHY EndMT has been reported to contribute to the pathogenesis of PAH. In this study, we aimed to determine the role of Ca2+ signaling in the development of EndMT in human lung vascular endothelial cells. Our data suggest that TGF-β1 requires store-operated Ca2+ entry through STIM1/Orai channels to induce SNAI-mediated EndMT. For the first time, we demonstrated that TGF-β1-induced EndMT is a Ca2+-dependent event, whereas inhibition of STIM1/Orai interaction attenuated EndMT in response to TGF-β1.

Disseminated Intravascular Coagulopathy (DIC) and Persistent Inflammation, Immunosuppression, and Catabolism Syndrome (PICS) are critical care syndromes that frequently coexist in critically ill patients, but mechanisms underlying their shared pathways are not well understood.

This review discusses the pathophysiology of DIC and PICS and explores the shared mechanisms behind DIC and PICS and their implications for clinical management.

DIC and PICS share a common pathophysiological foundation of endothelial dysfunction, coagulation dysregulation, and inflammation, leading to a vicious cycle of microvascular injury and systemic inflammation, culminating in organ dysfunction. DIC has also been identified as an independent risk factor for PICS. Anticoagulation therapies such as antithrombin, recombinant human soluble thrombomodulin (rhTM), and heparin attenuates inflammation, a mechanism underlying both syndromes, thereby improving outcomes in PICS.

DIC and PICS share critical pathophysiological pathways that exacerbate outcomes in critically ill patients. Recognizing these interconnections is essential for developing targeted therapies. Standardizing PICS definitions and advancing research to clarify mechanisms, interplay, and causality between DIC and PICS are crucial next steps.

Heart failure (HF) is a life-threatening cardiovascular disease associated with high mortality, diminished quality of life, and a significant economic burden on both patients and society. The pathogenesis of HF is closely related to the endothelium, where endothelial ion channels play an important role in regulating intracellular Ca2+ signals. These ion channels are essential to maintain vascular function, including endothelium-dependent vascular tone, inflammation response, and oxidative stress. Sodium-glucose cotransporter 2 inhibitors (SGLT2i) have shown promising cardiovascular benefits in HF patients, reducing mortality risk and hospitalization in several large clinical trials. Clinical and preclinical studies indicate that the cardioprotective effects of SGLT2i in HF are mediated by endothelial nitric oxide (NO) pathways, as well as by reducing inflammation and reactive oxygen species in cardiac endothelial cells. Additionally, SGLT2i may confer endothelial protection by lowering intracellular Ca2+ level through the inhibition of sodium-hydrogen exchanger 1 (NHE1) and sodium-calcium exchanger (NCX) in endothelial cells. In this review, we discuss present knowledge regarding the expression and role of Ca2+-related ion channels in endothelial cells in HF, focusing on the effects of SGLT2i on endothelial NHE1, NCX as well as on vascular tone.

While increased intracellular calcium (Ca2+) has been identified as a key effect of nanoparticles on endothelial cells, the mechanism has not been fully elucidated or examined under shear stress. Here, we show the effect of several types of 20 nm particles on Ca2+ in the presence of shear stress in human umbilical vein endothelial cells (HUVECs), human coronary artery endothelial cells (HCAECs), and human cardiac microvascular endothelial cells (HMVEC-Cs). Intracellular Ca2+ levels increased by nearly three-fold in these cell types upon exposure to 100 μg mL-1 20 nm Au particles, which was not seen in response to larger or smaller particles. An antagonist to the calcium channel - transient receptor potential vanilloid-type 4 (TRPV4) - drastically reduced the amount of calcium by 9.3-fold in HUVECs exposed to 0.6 Pa shear stress and 100 μg mL-1 20 nm gold particles, a trend upheld in both HCAECs and HMVEC-Cs. Cell alignment in the direction of fluid flow is a well-known phenomenon in endothelial cells, and interestingly, cells in the presence of 20 nm particles with fluid flow had a higher alignment index than cells in the fluid flow alone. When compared with previous works, these results indicated that 20 nm particles may be inducing endothelial permeability by activating the TRPV4 channel in vitro. The potential of nanoparticle delivery technologies hinges on an improved understanding of this effect toward improved delivery with limited toxicity.

Radio-immunotherapy has antitumor activity but also causes toxicity, which limits its clinical application. JS-201 is a dual antibody targeting PD-1 and TGF-β signaling. We investigated the antitumor effect of JS-201 combined with radiotherapy (RT) and the effect on radiation-induced lung injury (RILI). Different tumor models were established to detect the antitumor effects of the combination of JS-201 and RT, and RILI models were established to observe the effects of JS-201. Transcriptome sequencing showed that JS-201 optimized the tumor microenvironment by inhibiting extracellular matrix formation and angiogenesis. Combining JS-201 with RT further increased the inflammatory response and immune infiltration and showed great abscopal effects in Lewis lung cancer luciferase-positive models. Single-cell sequencing demonstrated that JS-201 reduced fibroblast proliferation by inhibiting the TGF-β/Smad pathway and the release of neutrophil extracellular traps mediated by ROS, thereby relieving radiation-induced pulmonary fibrosis. In conclusion, the JS-201 and RT combination enhances antitumor effects while mitigating acute and chronic RILI, and it may have potential for translational investigation as a cancer treatment strategy.

Endometritis poses a significant challenge to the dairy industry, impairing bovine reproductive performance and causing substantial economic losses. Although Morinda officinalis oligosaccharides (MOO) exhibit anti-inflammatory properties, their therapeutic potential against endometritis remains unclear. This study investigated MOO's protective effects against LPS-induced uterine injury in mice and inflammation in bovine endometrial epithelial cells (BENDs), and explored the underlying mechanisms. In mice, MOO attenuated uterine inflammation by improving histopathology, reducing pro-inflammatory cytokines and decreasing oxidative stress. In BEND cells, MOO alleviated LPS-induced inflammation, oxidative stress, and apoptosis via downregulating pro-inflammatory mediators (IL-1β, IL-6, TNF-α, IL-8, TLR4, RELA), restoring antioxidant enzymes (HMOX1, NQO1, Nrf2, NOX4), and modulating apoptosis markers (BAX, cleaved CASP3, CASP9, BCL2). MOO reduced ROS accumulation, preserved mitochondrial membrane potential, and inhibited calcium influx. Critically, the calcium channel agonist Bay K 8644 reversed MOO's protective effects, confirming calcium signaling modulation as a key mechanism. This study provides the first evidence that MOO mitigates LPS-induced uterine damage and BENDs inflammation through calcium signaling regulation, suggesting its potential for treating inflammation-related reproductive disorders in livestock.

SARS-CoV-2 variants present diverse clinical manifestations, necessitating deeper insights into their pathogenic effects. This study employs multi-omics approaches to investigate the molecular mechanisms underlying SARS-CoV-2 infection, focusing on vascular damage. Plasma proteomic analysis of unvaccinated participants infected with Omicron BA.2.76 or ancestral variants identifies key signaling pathways associated with endothelial dysfunction, with the vinculin (VCL) pathway emerging as a hallmark of Omicron infections, contributing to lung exudation. Metabolomic analysis of plasma samples from the same cohort reveals disruptions in immune function, cell membrane integrity, and metabolic processes, including altered tricarboxylic acid cycle and glycolysis pathways. An integrated analysis of proteomic and metabolomic data underscores the role of VCL in inflammation and extravasation, highlighting its interactions with adhesion molecules and inflammatory metabolites. A validation cohort of plasma samples from Omicron-infected participants confirms this association by replicating proteomic analysis, showing elevated VCL levels correlated with inflammatory markers. Functional studies in a male rat model of lung injury demonstrate that anti-VCL intervention reduces plasma VCL levels, mitigates alveolar edema, and restores alveolar-capillary barrier integrity, as assessed by histological staining and electron microscopy, thereby illustrating VCL modulation's impact on vascular leakage and extravasation. These findings establish VCL as a potential therapeutic target for mitigating vascular complications in SARS-CoV-2 infections.

While the relationship between albumin corrected calcium (ACC) levels and unfavourable outcomes has been previously investigated, existing studies have been limited in their specific application to patients undergoing maintenance hemodialysis (MHD) in intensive care unit (ICU). This retrospective cohort study aimed to explore the association between baseline ACC and 28-day in-hospital mortality in ICU patients undergoing MHD. Logistic regression model, smooth curve fitting, piecewise linear regression, subgroup analyses, and a series of sensitivity analyses were employed. Of the 2114 patients with a median age of 64 years, 290 (13.72%) died within 28 days after ICU admission. Multivariate regression analysis revealed that, in comparison with the lowest tertile, the highest tertile of ACC was associated with a higher mortality rate (OR 1.69, 95% CI 1.09-1.53, P = 0.0032). When the ACC levels were < 8.04 mg/dL, the mortality rate decreased with an adjusted OR of 0.44 (95% CI 0.20-0.98, P = 0.0438) for every 1 mg/dL increase in the ACC levels. When the ACC levels were ≥ 8.04 mg/dl, the mortality rate increased with an adjusted OR of 1.36 (95% CI 1.13-1.64, P = 0.0011) for every 1 mg/dl increase in the ACC levels. Non-linear relationship between ACC and 28-day in-hospital mortality were identified in patients undergoing MHD in the ICU. However, the findings of this study need to be confirmed through prospective studies.

To investigate whether ex vivo plasma from injured patients causes endothelial calcium (Ca 2+ ) influx as a mechanism of trauma-induced endothelial permeability.

Endothelial permeability after trauma contributes to postinjury organ dysfunction. While the mechanisms remain unclear, emerging evidence suggests intracellular Ca 2+ signaling may play a role.

Ex vivo plasma from injured patients with "low injury/low shock" (injury severity score <15, base excess ≥-6 mEq/L) and "high injury/high shock" (injury severity score ≥15, base excess <-6 mEq/L) were used to treat endothelial cells. Experimental conditions included Ca 2+ removal from the extracellular buffer, cyclopiazonic acid pretreatment to deplete intracellular Ca 2+ stores, and GSK2193874 pretreatment to block the transient receptor potential vanilloid 4 (TRPV4) Ca 2+ channel. Live cell fluorescence microscopy and electrical cell-substrate impedance sensing were used to assess cytosolic Ca 2+ increases and permeability, respectively. Western blot and live cell actin staining were used to assess myosin light chain phosphorylation and actomyosin contraction.

Compared with low injury/low shock plasma, high injury/high shock induced greater cytosolic Ca 2+ increase. Cytosolic Ca 2+ increase, myosin light chain phosphorylation, and actin cytoskeletal contraction were lower without extracellular Ca 2+ present. High injury/high shock plasma did not induce endothelial permeability without extracellular Ca 2+ present. TRPV4 inhibition lowered trauma plasma-induced endothelial Ca 2+ influx and permeability.

This study illuminates a novel mechanism of postinjury endotheliopathy involving Ca 2+ influx through the TRPV4 channel. TRPV4 inhibition mitigates trauma-induced endothelial permeability. Moreover, widespread endothelial Ca 2+ influx may contribute to trauma-induced hypocalcemia. This study provides the mechanistic basis for the development of Ca 2+ -targeted therapies and interventions in the care of severely injured patients.

Hypertension is the most frequent health-related condition worldwide and is a primary risk factor for renal and cardiovascular diseases. However, the underlying molecular mechanisms are still poorly understood. To uncover these mechanisms, multi-omics studies have significant potential, but such studies are challenged by genetic and environmental confounding - an issue that can be effectively reduced by studying intra-pair differences in twins. Here, we coupled data on hypertension diagnoses from the nationwide Danish Patient Registry to a study population of 740 twins for whom genome-wide DNA methylation and gene expression data were available together with measurements of systolic and diastolic blood pressure. We investigated five phenotypes: incident hypertension cases, systolic blood pressure, diastolic blood pressure, hypertension (140/90 mmHg), and hypertension (130/80 mmHg). Statistical analyses were performed using Cox (incident cases) or linear (remaining) regression analyses at both the individual-level and twin pair-level. Significant genes (p < 0.05) at both levels and in both types of biological data were investigated by bioinformatic analyses, including gene set enrichment analysis and interaction network analysis. Overall, most of the identified pathways related to the immune system, particularly inflammation, and biology of vascular smooth muscle cell. Of specific genes, lysine methyltransferase 2 A (KMT2A) was found to be central for incident hypertension, ataxia-telangiectasia mutated (ATM) for systolic blood pressure, and beta-actin (ACTB) for diastolic blood pressure. Noteworthy, lysine methyltransferase 2A (KMT2A) was also identified in the systolic and diastolic blood pressure analyses. Here, we present novel biomarkers for hypertension. This study design is surprisingly rare in the field of hypertension. We identified biological pathways related to vascular smooth muscle cells and the immune system, particular inflammation, to be associated with hypertension and blood pressure. Of specific genes, we identified KMT2A (lysine methyltransferase 2A) to be central for blood pressure and hypertension development. ACTB beta-actin, ATM ataxiatelangiectasia mutated, BP blood pressure, EWAS epigenome-wide association studies, KMT2A lysine methyltransferase 2A, LMER linear mixed effect regression, LR linear regression, TWAS transcriptome-wide association studies.

Cell Adhesion Molecules (CAMs) play a crucial role in regulating immune responses and repairing damage caused by hypoxia. However, the relationship between the expression characteristics of CAMs in yak lung tissues and their adaptation to the plateau environment remains unclear. To address this question, we compared lung tissues from yaks and cattle at the same altitude. After digesting the lung tissues with trypsin or Type I collagenase for varying durations, we observed that fewer cells were isolated from yak tissues compared to cattle. RNA sequencing (RNA-seq) analysis revealed that the Differentially Expressed Genes (DEGs) in lung tissues of yaks and cattle were significantly enriched in cell adhesion-related pathways. Quantitative real-time PCR (qRT-PCR) further identified changes in the expression levels of five distinct types of CAMs. Among these, the cadherin family (CDH1, CDH2, CDH11, PCDH12, CD34) exhibited significantly higher expression in yaks than in cattle. These cadherins play a critical role in regulating lung inflammation and maintaining the alveolar-capillary barrier, thereby ensuring the structural stability of the lungs. Immunohistochemical staining demonstrated that the expression patterns of cell adhesion-related proteins (CDH1, CDH11, ITGB6, SELP, CD44) were largely consistent with the qRT-PCR results. In conclusion, compared to cattle, the enhanced cell adhesion capacity of yak lung tissues contributes to their superior adaptation to the harsh plateau environment.

Acute lung injury (ALI), which poses a significant public health threat, is commonly caused by sepsis. ALI is associated with permeability and glycolysis changes in pulmonary microvascular endothelial cells. Our study demonstrates that heparin-binding protein (HBP), released from neutrophils during sepsis, exacerbates endothelial permeability and glycolysis, thereby triggering ALI. Through coimmunoprecipitation and mass spectrometry, TRIM21 was identified as a HBP interaction partner. Notably, HBP enhances the protein stability of TRIM21 by inhibiting K48 ubiquitination. TRIM21 binds to and promotes K63-linked ubiquitination of P65, facilitating its nuclear translocation. TRIM21 regulates HPMEC permeability and glycolysis in a manner dependent on P65 nuclear translocation. HBP stabilizes TRIM21 and enhances TRIM21 interactions with P65. Rescue experiments conducted in vivo and in vitro demonstrate that modulation of endothelial permeability and glycolysis by HBP is predominantly mediated through the TRIM21-P65 axis. Our results suggest that targeting the HBP/TRIM21/P65 axis is a novel therapeutic strategy to ameliorate ALI.

The human immune system is closely linked to microbiota such as a complex symbiotic relationship during the coevolution of vertebrates and microorganisms. The transfer of microorganisms from the mother's microbiota to the newborn begins before birth during gestation and is considered the initial phase of the intestinal microbiota (IM). The gut is an important site where microorganisms can establish colonies. The IM contains polymicrobial communities, which show complex interactions with diet and host immunity. The tendency towards dysbiosis of the intestinal microbiota is influenced by local but also extra-intestinal factors such as inflammatory processes, infections, or a septic state that can aggravate it. Pathogens could trigger an immune response, such as proinflammatory responses. In addition, changes in the host immune system also influence the intestinal community and structure with additional translocation of pathogenic and non-pathogenic bacteria. Finally, local intestinal inflammation has been found to be an important factor in the growth of pathogenic microorganisms, particularly in its role in sepsis. The aim of this article is to be able to detect the current knowledge of the mechanisms that can lead to dysbiosis of the intestinal microbiota and that can cause bacterial translocation with a risk of infection or septic state and vice versa.

Metastatic dissemination is driven by genetic, biochemical, and biophysical cues that favor the distant colonization of organs and the formation of life-threatening secondary tumors. We have previously demonstrated that endothelial cells (ECs) actively remodel during extravasation by enwrapping arrested tumor cells (TCs) and extruding them from the vascular lumen while maintaining perfusion. In this work, we dissect the cellular and molecular mechanisms driving endothelial remodeling. Using high-resolution intravital imaging in zebrafish embryos, we demonstrate that the actomyosin network of ECs controls tissue remodeling and subsequent TC extravasation. Furthermore, we uncovered that this cytoskeletal remodeling is driven by altered endothelial-calcium (Ca2+) signaling caused by arrested TCs. Accordingly, we demonstrated that the inhibition of voltage-dependent calcium L-type channels impairs extravasation. Lastly, we identified P2X4, TRP, and Piezo1 mechano-gated Ca2+ channels as key mediators of the process. These results further highlight the central role of endothelial remodeling during the extravasation of TCs and open avenues for successful therapeutic targeting.

Myocardial infarction (MI), a severe cardiovascular condition, is typically triggered by coronary artery disease, resulting in ischemic damage and the subsequent necrosis of the myocardium. Macrophages, known for their remarkable plasticity, are capable of exhibiting a range of phenotypes and functions as they react to diverse stimuli within their local microenvironment. In recent years, there has been an increasing number of studies on the regulation of macrophage behavior based on tissue engineering strategies, and its regulatory mechanisms deserve further investigation. This review first summarizes the effects of key regulatory factors of engineered biomaterials (including bioactive molecules, conductivity, and some microenvironmental factors) on macrophage behavior, then explores specific methods for inducing macrophage behavior through tissue engineering materials to promote myocardial repair, and summarizes the role of macrophage-host cell crosstalk in regulating inflammation, vascularization, and tissue remodeling. Finally, we propose some future challenges in regulating macrophage-material interactions and tailoring personalized biomaterials to guide macrophage phenotypes.

Coagulopathy in sepsis is common and is associated with high mortality. Although immunothrombosis is necessary for infection control, excessive thrombus formation can trigger a systemic thrombo-inflammatory response. Immunothrombosis plays a core role in sepsis-induced coagulopathy, and research has revealed a complex interplay between inflammation and coagulation. Different mechanisms underlying sepsis-related coagulopathy are discussed, including factors contributing to the imbalance of pro- and anticoagulation relevant to endothelial cells. The potential therapeutic implications of anticoagulants on these mechanisms are discussed. This review contributes to our understanding of the pathogenesis of coagulopathy in patients with sepsis. Recent studies suggest that endothelial cells play an important role in immunoregulation and hemostasis. Meanwhile, the non-anticoagulation effects of anticoagulants, especially heparin, which act in the pathogenesis of coagulopathy in septic patients, have been partially revealed. We believe that further insights into the pathogenesis of sepsis-induced coagulopathy will help physicians evaluate patient conditions effectively, leading to advanced early recognition and better decision-making in the treatment of sepsis.

Diarrheas are an important adverse effect of afatinib, a tyrosine kinase inhibitor (TKI) anti-cancer drug, leading to mortality and morbidity in cancer patients with their pathophysiological mechanisms related to intestinal barrier dysfunctions being poorly understood. This study aimed to investigate the effect of afatinib on intestinal epithelial barrier integrity using a human colon-derived organoid model (colonoids). Afatinib (0.5 μM) significantly decreased the transepithelial electrical resistance (TEER) by ∼60% and increased apical-to-basolateral dextran flux by > 20 folds without causing apparent cytotoxicity in human colonoids. The delocalization of zonula occludens-1 (ZO-1) and a decrease in mRNA and protein expression of claudin-4 and ZO-1 were also observed in the afatinib-treated human colonoids. Afatinib induced nuclear translocation of nuclear factor kappa B (NF-κB) as well as mRNA and protein expression of NF-κB targets including tumor necrosis factor (TNF)-alpha, interleukin-8 (IL-8), and inducible nitric oxide synthase (iNOS) indicating the initiation of the NF-κB-mediated epithelial inflammatory responses. Interestingly, afatinib induced mRNA and protein expression of myosin light chain (MLC) kinase (MLCK) and MLC phosphorylation, a known inducer of intestinal epithelial barrier disruption. Treatment with iNOS inhibitor (1400W) or MLCK inhibitor (ML-7) reversed the effect of afatinib on mRNA expressions of ZO-1 and claudin-4, and TEER. Collectively, our results indicate that afatinib induces intestinal epithelial barrier dysfunction via mechanisms involving NF-κB-iNOS-MLCK pathways. This finding may pave the way for developing therapeutic strategies to reduce adverse effects and enhance efficacy of TKI in cancer patients.

Ultrasound and microbubble mediated blood brain barrier opening is a non-invasive and effective technique for drug delivery to targeted brain region. However, the exact mechanisms are not fully resolved. The influences of Ca2+ signaling on sonoporation and endothelial tight junctional regulation affect the efficiency and biosafety of the technique. Therefore, an improved understanding of how ultrasound evokes Ca2+ signaling in the brain endothelial monolayer, and its correlation to endothelial permeability change is necessary. Here, we examined the effects of SonoVue microbubbles or integrin-targeted microbeads on ultrasound induced bioeffects in brain microvascular endothelial monolayer using an acoustically-coupled microscopy system, where focused ultrasound exposure and real-time recording of Ca2+ signaling and membrane perforation were performed. Microbubbles induced robust Ca2+ responses, often accompanied by cell poration, while ultrasound with microbeads elicited reversible Ca2+ response without membrane poration. At the conditions evoking reversible Ca2+ signaling, intracellular Ca2+ release and reactive oxygen species played key roles for microbubbles induced Ca2+ signaling while activation of mechanosensitive ion channels was essential for the case of microbeads. Trans-well diffusion analysis revealed significantly higher trans-endothelial transport of 70 kDa FITC-dextran for both integrin-targeted microbeads and microbubbles compared to the control group. Further immunofluorescence staining showed disruption of cell junctions with microbubble stimulation and reversible remodeling of many cell junctions by ultrasound with integrin-targeted microbeads. This investigation provides new insights for ultrasound induced Ca2+ signaling and its influence on endothelial permeability, which may help develop new strategies for safe and efficient drug/gene delivery in the vascular system.

The endothelial microvasculature is essential for the regulation of vasodilation and vasoconstriction, and improved functioning of the endothelium is linked to improved outcomes for individuals with coronary artery disease (CAD). People with endothelial dysfunction exhibit a loss of nitric oxide (NO)-mediated vasodilation, achieving vasodilation instead through mitochondria-derived H2O2. Mitochondrial dynamics is an important autoregulatory mechanism that contributes to mitochondrial and endothelial homeostasis and plays a role in the formation of reactive oxygen species (ROS), including H2O2. Dysregulation of mitochondrial dynamics leads to increased ROS production, decreased ATP production, impaired metabolism, activation of pathological signal transduction, impaired calcium sensing, and inflammation. We hypothesize that dysregulation of endothelial mitochondrial dynamics plays a crucial role in the endothelial microvascular dysfunction seen in individuals with CAD. Therefore, proper regulation of endothelial mitochondrial dynamics may be a suitable treatment for individuals with endothelial microvascular dysfunction, and we furthermore postulate that improving this microvascular dysfunction will directly improve outcomes for those with CAD.

Diabetic kidney disease is a complication in one out of three patients with diabetes. Aberrant glucose metabolism in diabetes leads to structural and functional damage in glomerular tissue and a systemic inflammatory immune response. Complex cellular signaling is at the core of metabolic and functional derangement. Unfortunately, the mechanism underlying the role of inflammation in glomerular endothelial cell dysfunction during diabetic kidney disease is not fully understood. Mathematical models in systems biology allow the integration of experimental evidence and cellular signaling networks to understand mechanisms involved in disease progression. This study developed a logic-based ordinary differential equations model to study inflammatory crosstalk between macrophages and glomerular endothelial cells during diabetic kidney disease progression using a protein signaling network stimulated with glucose and lipopolysaccharide. This modeling approach reduced the biological parameters needed to study signaling networks. The model was fitted to and validated against available biochemical data from \textit{in vitro} experiments. The model identified mechanisms for dysregulated signaling in macrophages and glomerular endothelial cells during diabetic kidney disease. In addition, the influence of signaling interactions on glomerular endothelial cell morphology through selective knockdown and downregulation was investigated. Simulation results showed that partial knockdown of VEGF receptor 1, PLC-γ, adherens junction proteins, and calcium partially recovered the intercellular gap width between glomerular endothelial cells. These findings contribute to understanding signaling and molecular perturbations that affect the glomerular endothelial cells in the early stage of diabetic kidney disease.

Endothelial-to-mesenchymal transition (EndMT) is a biological process that converts endothelial cells to mesenchymal cells with increased proliferative and migrative abilities. EndMT has been implicated in the development of pulmonary vascular remodeling in pulmonary arterial hypertension (PAH), a fatal and progressive lung vascular disease. Transforming growth factor β 1 (TGF-β 1 ), an inflammatory cytokine, is known to induce EndMT in many types of endothelial cells including lung vascular endothelial cells (LVEC). An increase in cytosolic free Ca 2+ concentration ([Ca 2+ ] cyt ) is a major stimulus for cellular proliferation and phenotypic transition, but it is unknown whether Ca 2+ signaling is involved in EndMT. In this study we tested the hypothesis that TGF-β 1 -induced EndMT in human LVEC is Ca 2+ -dependent. Treatment of LVEC with TGF-β 1 for 5-7 days resulted in increase in SNAI1/2 expression, induction of EndMT, upregulation of STIM/Orai1 and enhancement of store-operated Ca 2+ entry (SOCE). Removal (or chelation) of extracellular or intracellular Ca 2+ with EGTA or BAPTA-AM respectively abolished EndMT in response to TGF-β 1 . Moreover, EGTA diminished TGF-β 1 -induced increase in SNAI in a dose-dependent manner. Knockdown of either STIM1 or Orai1 was sufficient to prevent TGF-β-mediated increase in SNAI1/2 and EndMT, but did not rescue the continuous adherent junctions. Blockade of Orai1 channels by AnCoA4 inhibited TGF-β-mediated EndMT and restored PECAM1-positive continuous adherent junctions. In conclusion, intracellular Ca 2+ signaling plays a critical role in TGF-β-associated EndMT through enhanced SOCE and STIM1-Orai1 interaction. Thus, targeting Ca 2+ signaling pathways regulating EndMT may be a novel therapeutic approach to treat PAH and other forms of pre-capillary pulmonary hypertension.

EndMT has been reported to contribute to the pathogenesis of PH. In this study we aimed to determine the role of Ca 2+ signaling in the development of EndMT in human lung vascular endothelial cells. Our data suggest that TGF-β 1 requires store-operated Ca 2+ entry through STIM1/Orai channels to induce SNAI-mediated EndMT. For the first time we demonstrated that TGF-β 1 -induced EndMT is Ca 2+ -dependent event while inhibition of STIM1/Orai interaction attenuated EndMT in response to TGF-β 1 .

Several empirical studies have linked subway and ambient particle exposure to toxicity, pro-inflammatory responses, and vascular dysfunction. However, the health effects of pollutants generated from varying sources, particularly when combined with lipopolysaccharide (LPS), are still unexplored. Therefore, the aim of this study was to investigate the characteristic health effects of iron oxide particles (the main components of subway particles) in comparison with urban aerosols (UA) and vehicle exhaust particles (VEP), alone and in combination with LPS. This study revealed that iron oxides caused a more significant reduction in human umbilical vein endothelial cell viability, increased lactate dehydrogenase release, and decreased the production of plasminogen activator inhibitor-1, a fibrinolytic modulator, and endothelin-1, a vasoconstrictor, compared to those by VEP and UA at marginally toxic and toxic concentrations. While VEP and UA induced an increase in interleukin (IL)-6 production, iron oxides, particularly Fe3O4, increased IL-8 production at slightly toxic and non-cytotoxic concentrations. In addition, co-exposure of all particles and LPS at non-cytotoxic concentrations promoted pro-inflammatory cytokine (IL-6 and IL-8) production relative to exposure to the particles alone. Interestingly, the tendency towards either coagulation or fibrinolytic conditions was dependent on the concentration of exposed particles at the same LPS concentration. Furthermore, increases in inflammation, neutrophil and lymphocyte recruitment around blood vessels, and edema were observed in murine lungs exposed to a combination of iron oxides and LPS compared to those in mice exposed to iron oxide alone. Thus, iron oxide-rich subway particulate poses more health risks than outdoor ambient particles since they can significantly impair endothelial function, particularly through gross cellular and vascular homeostatic protein damage, and induce exacerbated inflammatory responses during co-exposure. These findings provide novel empirical evidence for epidemiological studies seeking mechanisms responsible for the observed health impact of transport- and occupational-related exposures on vascular dysfunction.

Inflammation induced by angiotensin II (Ang II) is a key event in the progression of numerous cardiovascular diseases. Astragaloside IV (AS-IV), a glycoside extracted from Astragalus membranaceus Bunge, has been shown to inhibit Ang II-induced inflammatory responses in vivo. However, the mechanisms underlying the beneficial effects are still unclear. This study investigated whether AS-IV attenuates endothelial inflammation induced by Ang II via the activation of endothelial nitric oxide synthase (eNOS)/nitric oxide (NO) pathway. Human umbilical vein endothelial cells (HUVECs) were cultured in the presence of AS-IV with or without the specific inhibitor of NOS or Ca2+- and phosphatidylinositol 3-kinase (PI3K)/Akt-dependent cascade prior to Ang II exposure. Incubation of HUVECs with AS-IV enhanced NO production and eNOSser1177 phosphorylation. These responses were abrogated by the inhibition of NOS or Ca2+- and PI3K/Akt-dependent pathway. In addition, preincubation of HUVECs with AS-IV inhibited Ang II-induced cytokine and chemokine production, adhesion molecule expression, monocyte adhesion, and nuclear factor kappa B (NF-κB) activation as evidenced by the attenuation of inhibitor of kappa B alpha phosphorylation and subsequent NF-κB DNA binding. These effects of AS-IV were abolished by the suppression of NOS or Ca2+- and PI3K/Akt-dependent cascade. Our findings indicate that AS-IV attenuates inflammatory responses triggered by Ang II possibly via the activation of Ca2+/PI3K/Akt/eNOS/NO pathway in endothelial cells.

Inflammation contributes to the development of thrombosis, but the mechanistic basis for this association remains poorly understood. Innate immune responses and coagulation pathways are activated in parallel following infection or injury, and represent an important host defense mechanism to limit pathogen spread in the bloodstream. However, dysregulated proinflammatory activity is implicated in the progression of venous thromboembolism and arterial thrombosis. In this review, we focus on the role of myeloid cells in propagating thromboinflammation in acute inflammatory conditions, such as sepsis and coronavirus disease 2019 (COVID-19), and chronic inflammatory conditions, such as obesity, atherosclerosis, and inflammatory bowel disease. Myeloid cells are considered key drivers of thromboinflammation via upregulated tissue factor activity, formation of neutrophil extracellular traps (NETs), contact pathway activation, and aberrant coagulation factor-mediated protease-activated receptor (PAR) signaling. We discuss how strategies to target the intersection between myeloid cell-mediated inflammation and activation of blood coagulation represent an exciting new approach to combat immunothrombosis. Specifically, repurposed anti-inflammatory drugs, immunometabolic regulators, and NETosis inhibitors present opportunities that have the potential to dampen immunothrombotic activity without interfering with hemostasis. Such therapies could have far-reaching benefits for patient care across many thromboinflammatory conditions.

Lipid emulsion (LE), a widely used parenteral nutrition, exhibits a well-documented ability to reverse the vasodilatory effects induced by acetylcholine in blood vessels. However, the specific mechanisms underlying this action are not yet fully understood. This study aimed to elucidate the mechanism by which LE reverses vasodilation in vitro through dose-response curve experiments, calcium imaging, and fluorescence assays. The results revealed a significant attenuation of acetylcholine (Ach)-induced vasodilation in rat thoracic aortic rings following LE exposure. In human aortic endothelial cells, pretreatment with LE significantly suppressed ATP-induced calcium elevation. This suppression persisted even after elimination of extracellular calcium with a calcium chelator. Moreover, LE pre-exposure reduced the intracellular calcium concentration ([Ca2+]i) elevation in endothelial cells following cyclopiazonic acid (CPA) treatment, suggesting enhanced endoplasmic reticulum (ER) calcium reuptake. Additionally, nitric oxide (NO) fluorescence assays showed a decrease in NO production upon ATP stimulation post-LE pretreatment of endothelial cells. Taken together, these results indicate that the reversal of vasodilation by LE may involve enhanced ER calcium uptake, leading to a reduction in intracellular calcium concentration and suppression of NO (key vasodilatory agent) synthesis.

The contribution of Erk1/2 to endothelial barrier regulation is convoluted and differs depending on the vascular bed. We explored the effects of Erk1/2 inhibition on endothelial barrier maintenance and its relationship with cAMP-dependent barrier strengthening. Thus, myocardial endothelial cells (MyEnd) were isolated and protein expression, localization and activity of structural and signaling molecules involved in maintenance of endothelial function were investigated by Western blot, immunostainings and G-LISA, respectively. The transendothelial electrical resistance (TEER) from confluent MyEnd monolayers was measured and used as a direct indicator of barrier integrity in vitro. Miles assay was performed to evaluate vascular permeability in vivo. Erk1/2 inhibition with U0126 affected neither the structural organization of adherens or tight junctions nor the protein level of their components, However, TEER drop significantly upon U0126 application, but the effect was transitory as the barrier function recovered 30 min after treatment. Erk1/2 inhibition delayed cAMP-mediated barrier strengthening but did not prevent barrier fortification despite diminishing Rac1 activation. Moreover, Erk1/2 inhibition, induced vascular leakage that could be prevented by local cAMP elevation in vivo. Our data demonstrate that Erk1/2 is required to prevent vascular permeability but is not critical for cAMP-mediated barrier enhancement.

Pulmonary hypertension (PH) is a fatal disease with no existing curative drugs. NF-E2-related factor 2 (NRF2) a pivotal molecular in cellular protection, was investigated in PH models to elucidate its role in regulating abnormal phenotypes in pulmonary artery cells. We examined the expression of NRF2 in PH models and explored the role of NRF2 in regulating abnormal phenotypes in pulmonary artery cells. We determined the expression level of NRF2 in lung tissues of PH model decreased significantly. We found that NRF2 was reduced in rat pulmonary artery endothelial cells (rPAEC) under hypoxia, while it was overexpressed in rat pulmonary artery smooth muscle cells (rPASMC) under hypoxia. Next, the results showed that knockdown NRF2 in rPAEC promoted endothelial-mesenchymal transformation and upregulated reactive oxygen species level. After the rPASMC was treated with siRNA or activator, we found that NRF2 could accelerate cell migration by affecting MMP2/3/7, and promote cell proliferation by regulating PDGFR/ERK1/2 and mTOR/P70S6K pathways. Therefore, the study has shown that the clinical application of NRF2 activator in the treatment of pulmonary hypertension may cause side effects of promoting the proliferation and migration of rPASMC. Attention should be paid to the combination of NRF2 activators.

The blood-brain barrier (BBB) plays a critical role in maintaining ion and fluid homeostasis, essential for brain metabolism and neuronal function. Regulation of nutrient, water, and ion transport across the BBB is tightly controlled by specialized ion transporters and channels located within its unique cellular components. These dynamic transport processes not only influence the BBB's structure but also impact vital signaling mechanisms, essential for its optimal function. Disruption in ion, pH, and fluid balance at the BBB is associated with brain pathology and has been implicated in various neurological conditions, including stroke, epilepsy, trauma, and neurodegenerative diseases such as Alzheimer's disease (AD). However, knowledge gaps exist regarding the impact of ion transport dysregulation on BBB function in neurodegenerative dementias. Several factors contribute to this gap: the complex nature of these conditions, historical research focus on neuronal mechanisms and technical challenges in studying the ion transport mechanisms in in vivo models and the lack of efficient in vitro BBB dementia models. This review provides an overview of current research on the roles of ion transporters and channels at the BBB and poses specific research questions: 1) How are the expression and activity of key ion transporters altered in AD and vascular dementia (VaD); 2) Do these changes contribute to BBB dysfunction and disease progression; and 3) Can restoring ion transport function mitigate BBB dysfunction and improve clinical outcomes. Addressing these gaps will provide a greater insight into the vascular pathology of neurodegenerative disorders.

Tight control of cytoplasmic Ca2+ concentration in endothelial cells is essential for the regulation of endothelial barrier function. Here, we investigated the role of Cavβ3, a subunit of voltage-gated Ca2+ (Cav) channels, in modulating Ca2+ signaling in brain microvascular endothelial cells (BMECs) and how this contributes to the integrity of the blood-brain barrier.

We investigated the function of Cavβ3 in BMECs by Ca2+ imaging and Western blot, examined the endothelial barrier function in vitro and the integrity of the blood-brain barrier in vivo, and evaluated disease course after induction of experimental autoimmune encephalomyelitis in mice using Cavβ3-/- (Cavβ3-deficient) mice as controls.

We identified Cavβ3 protein in BMECs, but electrophysiological recordings did not reveal significant Cav channel activity. In vivo, blood-brain barrier integrity was reduced in the absence of Cavβ3. After induction of experimental autoimmune encephalomyelitis, Cavβ3-/- mice showed earlier disease onset with exacerbated clinical disability and increased T-cell infiltration. In vitro, the transendothelial resistance of Cavβ3-/- BMEC monolayers was lower than that of wild-type BMEC monolayers, and the organization of the junctional protein ZO-1 (zona occludens-1) was impaired. Thrombin stimulates inositol 1,4,5-trisphosphate-dependent Ca2+ release, which facilitates cell contraction and enhances endothelial barrier permeability via Ca2+-dependent phosphorylation of MLC (myosin light chain). These effects were more pronounced in Cavβ3-/- than in wild-type BMECs, whereas the differences were abolished in the presence of the MLCK (MLC kinase) inhibitor ML-7. Expression of Cacnb3 cDNA in Cavβ3-/- BMECs restored the wild-type phenotype. Coimmunoprecipitation and mass spectrometry demonstrated the association of Cavβ3 with inositol 1,4,5-trisphosphate receptor proteins.

Independent of its function as a subunit of Cav channels, Cavβ3 interacts with the inositol 1,4,5-trisphosphate receptor and is involved in the tight control of cytoplasmic Ca2+ concentration and Ca2+-dependent MLC phosphorylation in BMECs, and this role of Cavβ3 in BMECs contributes to blood-brain barrier integrity and attenuates the severity of experimental autoimmune encephalomyelitis disease.

With the in-depth investigation of various diseases, angiogenesis has gained increasing attention. Among the contributing factors to angiogenesis research, endothelial epigenetics has emerged as an influential player. Endothelial epigenetic therapy exerts its regulatory effects on endothelial cells by controlling gene expression, RNA, and histone modification within these cells, which subsequently promotes or inhibits angiogenesis. As a result, this therapeutic approach offers potential strategies for disease treatment. The purpose of this review is to outline the pertinent mechanisms of endothelial cell epigenetics, encompassing glycolysis, lactation, amino acid metabolism, non-coding RNA, DNA methylation, histone modification, and their connections to specific diseases and clinical applications. We firmly believe that endothelial cell epigenetics has the potential to become an integral component of precision medicine therapy, unveiling novel therapeutic targets and providing new directions and opportunities for disease treatment.

Aging is a major risk factor for cardiovascular diseases (CVD), and mitochondrial autophagy impairment is considered a significant physiological change associated with aging. Endothelial cells play a crucial role in maintaining vascular homeostasis and function, participating in various physiological processes such as regulating vascular tone, coagulation, angiogenesis, and inflammatory responses. As aging progresses, mitochondrial autophagy impairment in endothelial cells worsens, leading to the development of numerous cardiovascular diseases. Therefore, regulating mitochondrial autophagy in endothelial cells is vital for preventing and treating age-related cardiovascular diseases. However, there is currently a lack of systematic reviews in this area. To address this gap, we have written this review to provide new research and therapeutic strategies for managing aging and age-related cardiovascular diseases.

Caffeine (1,3,7-trimethylxanthine) is a naturally occurring methylxanthine that acts as a potent central nervous system stimulant found in more than 60 different plants and fruits. Although caffeinated beverages are widely and casually consumed, the application of caffeine beyond dietary levels as pharmacologic therapy has been recognized since the beginning of its recorded use. The analgesic and vasoactive properties of caffeine are well known, but the extent of their molecular basis remains an area of active research. There is existing evidence in the literature as to caffeine's effect on TRP channels, the role of caffeine in pain management and analgesia, as well as the role of TRP in pain and analgesia; however, there has yet to be a review focused on the interaction between caffeine and TRP channels. Although the influence of caffeine on TRP has been demonstrated in the lab and in animal models, there is a scarcity of data collected on a large scale as to the clinical utility of caffeine as a regulator of TRP. This review aims to prompt further molecular research to elucidate the specific ligand-host interaction between caffeine and TRP by validating caffeine as a regulator of transient receptor potential (TRP) channels-focusing on the transient receptor potential vanilloid 1 (TRPV1) receptor and transient receptor potential ankyrin 1 (TRPA1) receptor subtypes-and its application in areas of pain.

The endothelium regulates crucial aspects of vascular function, including hemostasis, vasomotor tone, proliferation, immune cell adhesion, and microvascular permeability. Endothelial cells (ECs), especially in arterioles, are pivotal for flow distribution and peripheral resistance regulation. Investigating vascular endothelium physiology, particularly in microvascular ECs, demands precise isolation and culturing techniques.

Freshly isolated ECs are vital for examining protein expression, ion channel behavior, and calcium dynamics. Establishing primary endothelial cell cultures is crucial for unraveling vascular functions and understanding intact microvessel endothelium roles. Despite the significance, detailed protocols and comparisons with intact vessels are scarce in microvascular research. We developed a reproducible method to isolate microvascular ECs, assessing substrate influence by cultivating cells on fibronectin and gelatin matrix gels. This comparative approach enhances our understanding of microvascular endothelial cell biology.

Microvascular mesenteric ECs expressed key markers (VE-cadherin and eNOS) in both matrix gels, confirming cell culture purity. Under uncoated conditions, ECs were undetected, whereas proteins linked to smooth muscle cells and fibroblasts were evident. Examining endothelial cell (EC) physiological dynamics on distinct matrix substrates revealed comparable cell length, shape, and Ca2+ elevations in both male and female ECs on gelatin and fibronectin matrix gels. Gelatin-cultured ECs exhibited analogous membrane potential responses to acetylcholine (ACh) or adenosine triphosphate (ATP), contrasting with their fibronectin-cultured counterparts. In the absence of stimulation, fibronectin-cultured ECs displayed a more depolarized resting membrane potential than gelatin-cultured ECs.

Gelatin-cultured ECs demonstrated electrical behaviors akin to intact endothelium from mouse mesenteric arteries, thus advancing our understanding of endothelial cell behavior within diverse microenvironments.

Intraoperative persistent hypotension (IPH) during pancreaticoduodenectomy (PD) is linked to adverse postoperative outcomes, yet its risk factors remain unclear.

To clarify the risk factors associated with IPH during PD, ensuring patient safety in the perioperative period.

A retrospective analysis of patient records from January 2018 to December 2022 at the First Affiliated Hospital of Nanjing Medical University identified factors associated with IPH in PD. These factors included age, gender, body mass index, American Society of Anesthesiologists classification, comorbidities, medication history, operation duration, fluid balance, blood loss, urine output, and blood gas parameters. IPH was defined as sustained mean arterial pressure < 65 mmHg, requiring prolonged deoxyepinephrine infusion for > 30 min despite additional deoxyepinephrine and fluid treatments.

Among 1596 PD patients, 661 (41.42%) experienced IPH. Multivariate logistic regression identified key risk factors: increased age [odds ratio (OR): 1.20 per decade, 95% confidence interval (CI): 1.08-1.33] (P < 0.001), longer surgery duration (OR: 1.15 per additional hour, 95%CI: 1.05-1.26) (P < 0.01), and greater blood loss (OR: 1.18 per 250-mL increment, 95%CI: 1.06-1.32) (P < 0.01). A novel finding was the association of arterial blood Ca2+ < 1.05 mmol/L with IPH (OR: 2.03, 95%CI: 1.65-2.50) (P < 0.001).

IPH during PD is independently associated with older age, prolonged surgery, increased blood loss, and lower plasma Ca2+.

Calcium plays central roles in numerous biological processes, thereby, its levels in the blood are under strict control to maintain homeostatic balance and enable the proper functioning of living organisms. The regulatory mechanisms ensuring this balance can be affected by pathologies such as cancer, and as a result, hyper- or hypocalcemia can occur. These states, characterized by elevated or decreased calcium blood levels, respectively, have a significant effect on general homeostasis. This article focuses on a particular form of calcium metabolism disorder, which is hypercalcemia in neoplasms. It also constitutes a summary of the current knowledge regarding the diagnosis of hypercalcemia and its management. Hypercalcemia of malignancy is estimated to affect over 40% of cancer patients and can be associated with both solid and blood cancers. Elevated calcium levels can be an indicator of developing cancer. The main mechanism of hypercalcemia development in tumors appears to be excessive production of parathyroid hormone-related peptides. Among the known treatment methods, bisphosphonates, calcitonin, steroids, and denosumab should be mentioned, but ongoing research promotes progress in pharmacotherapy. Given the rising global cancer prevalence, the problem of hypercalcemia is of high importance and requires attention.

Heart failure (HF) is one of the major causes of death among diabetic patients. Although studies have shown that curcumin analog C66 can remarkably relieve diabetes-associated cardiovascular and kidney complications, the role of SJ-12, SJ-12, a novel curcumin analog, in diabetic cardiomyopathy and its molecular targets are unknown. 7-week-old male C57BL/6 mice were intraperitoneally injected with single streptozotocin (STZ) (160 mg/kg) to develop diabetic cardiomyopathy (DCM). The diabetic mice were then treated with SJ-12 via gavage for two months. Body weight, fast blood glucose, cardiac utrasonography, myocardial injury markers, pathological morphology of the heart, hypertrophic and fibrotic markers were assessed. The potential target of SJ-12 was evaluated via RNA-sequencing analysis. The O-GlcNAcylation levels of SP1 were detected via immunoprecipitation. SJ-12 effectively suppressed myocardial hypertrophy and fibrosis, thereby preventing heart dysfunction in mice with STZ-induced heart failure. RNA-sequencing analysis revealed that SJ-12 exerted its therapeutic effects through the modulation of the calcium signaling pathway. Furthermore, SJ-12 reduced the O-GlcNAcylation levels of SP1 by inhibiting O-linked N-acetylglucosamine transferase (OGT). Also, SJ-12 stabilized Sarcoplasmic/Endoplasmic Reticulum Calcium ATPase 2a (SERCA2a), a crucial regulator of calcium homeostasis, thus reducing hypertrophy and fibrosis in mouse hearts and cultured cardiomyocytes. However, the anti-fibrotic effects of SJ-12 were not detected in SERCA2a or OGT-silenced cardiomyocytes, indicating that SJ-12 can prevent DCM by targeting OGT-dependent O-GlcNAcylation of SP1.These findings indicate that SJ-12 can exert cardioprotective effects in STZ-induced mice by reducing the O-GlcNAcylation levels of SP1, thus stabilizing SERCA2a and reducing myocardial fibrosis and hypertrophy. Therefore, SJ-12 can be used for the treatment of diabetic cardiomyopathy.

Nearly half of adults have hypertension, a major risk factor for cardiovascular disease. Mitochondrial hyperacetylation is linked to hypertension, but the role of acetylation of specific proteins is not clear. We hypothesized that acetylation of mitochondrial CypD (cyclophilin D) at K166 contributes to endothelial dysfunction and hypertension.

To test this hypothesis, we studied CypD acetylation in patients with essential hypertension, defined a pathogenic role of CypD acetylation in deacetylation mimetic CypD-K166R mutant mice and endothelial-specific GCN5L1 (general control of amino acid synthesis 5 like 1)-deficient mice using an Ang II (angiotensin II) model of hypertension.

Arterioles from hypertensive patients had 280% higher CypD acetylation coupled with reduced Sirt3 (sirtuin 3) and increased GCN5L1 levels. GCN5L1 regulates mitochondrial protein acetylation and promotes CypD acetylation, which is counteracted by mitochondrial deacetylase Sirt3. In human aortic endothelial cells, GCN5L1 depletion prevents superoxide overproduction. Deacetylation mimetic CypD-K166R mice were protected from vascular oxidative stress, endothelial dysfunction, and Ang II-induced hypertension. Ang II-induced hypertension increased mitochondrial GCN5L1 and reduced Sirt3 levels resulting in a 250% increase in GCN5L1/Sirt3 ratio promoting CypD acetylation. Treatment with mitochondria-targeted scavenger of cytotoxic isolevuglandins (mito2HOBA) normalized GCN5L1/Sirt3 ratio, reduced CypD acetylation, and attenuated hypertension. The role of mitochondrial acetyltransferase GCN5L1 in the endothelial function was tested in endothelial-specific GCN5L1 knockout mice. Depletion of endothelial GCN5L1 prevented Ang II-induced mitochondrial oxidative stress, reduced the maladaptive switch of vascular metabolism to glycolysis, prevented inactivation of endothelial nitric oxide, preserved endothelial-dependent relaxation, and attenuated hypertension.

These data support the pathogenic role of CypD acetylation in endothelial dysfunction and hypertension. We suggest that targeting cytotoxic mitochondrial isolevuglandins and GCN5L1 reduces CypD acetylation, which may be beneficial in cardiovascular disease.