VCAM-1 Expression Research Articles

BAP31 Promotes Adhesion Between Endothelial Cells and Macrophages Through the NF-κB Signaling Pathway in Sepsis.

To investigate the role of B cell receptor associated protein 31 (BAP31) in the pathogenesis of sepsis. Cecal ligation and puncture (CLP)-induced C57BL/6J mice, and LPS-challenged endothelial cells (HUVECs) were established to mimic a sepsis animal model and a sepsis cell model, respectively. Cre/LoxP and shRNA methods were used for BAP31 knockdown in vivo and in vitro respectively. Neutrophils/macrophages-endothelial cocultures were used to evaluate neutrophils or macrophages infiltration and adhesion to endothelial cells. Cox proportional hazards model was used to evaluate the survival time of mice. Western blotting (WB) and Quantitative real-time polymerase chain reaction (qRT-PCR) were used to detect toll-like receptor (TLR) signaling pathway, transforming growth factor β activated kinase 1 (TAK1) signaling pathway and phosphoinositide-3 kinases-protein kinase B (PI3K/AKT) signaling pathway. Deletion of BAP31 reduced CLP-induced mortality of mice, histological damage with less interstitial edema, and neutrophils and macrophages infiltration. IHC and IF showed that BAP31 knockdown significantly decreases the expressions of ICAM1 and VCAM1 both in vivo and in vitro. Coculture showed that LPS-induced neutrophils or macrophages adhesion to endothelial cells was significantly weakened in BAP31 knockdown cells. In addition, BAP31 knockdown of endothelial cells decreased the expression of CD80 and CD86 on the surface of macrophages as well as interleukin 1β (IL-1β) and tumor necrosis factor α (TNF-α) during sepsis. Mechanistically, LPS-induced the activation of TLR4, MyD88 and TRAF6, and the phosphorylation of TAK1, PI3K, AKT, IκBα and IKKα/β, resulting in activation of nuclear factor kappa B (NF-κB) p65 in endothelial cells. However, BAP31 knockdown significantly reversed the expressions of associated proteins. BAP31 up-regulated the expressions of ICAM1 and VCAM1 in endothelial cells leading to sepsis-associated organ injury. This may be involved in activation of TLR signaling pathway, TAK1 pathway, and PI3K-AKT signaling pathway.

A terminal metabolite of niacin promotes vascular inflammation and contributes to cardiovascular disease risk.

Despite intensive preventive cardiovascular disease (CVD) efforts, substantial residual CVD risk remains even for individuals receiving all guideline-recommended interventions. Niacin is an essential micronutrient fortified in food staples, but its role in CVD is not well understood. In this study, untargeted metabolomics analysis of fasting plasma from stable cardiac patients in a prospective discovery cohort (n = 1,162 total, n = 422 females) suggested that niacin metabolism was associated with incident major adverse cardiovascular events (MACE). Serum levels of the terminal metabolites of excess niacin, N1-methyl-2-pyridone-5-carboxamide (2PY) and N1-methyl-4-pyridone-3-carboxamide (4PY), were associated with increased 3-year MACE risk in two validation cohorts (US n = 2,331 total, n = 774 females; European n = 832 total, n = 249 females) (adjusted hazard ratio (HR) (95% confidence interval) for 2PY: 1.64 (1.10-2.42) and 2.02 (1.29-3.18), respectively; for 4PY: 1.89 (1.26-2.84) and 1.99 (1.26-3.14), respectively). Phenome-wide association analysis of the genetic variant rs10496731, which was significantly associated with both 2PY and 4PY levels, revealed an association of this variant with levels of soluble vascular adhesion molecule 1 (sVCAM-1). Further meta-analysis confirmed association of rs10496731 with sVCAM-1 (n = 106,000 total, n = 53,075 females, P = 3.6 × 10-18). Moreover, sVCAM-1 levels were significantly correlated with both 2PY and 4PY in a validation cohort (n = 974 total, n = 333 females) (2PY: rho = 0.13, P = 7.7 × 10-5; 4PY: rho = 0.18, P = 1.1 × 10-8). Lastly, treatment with physiological levels of 4PY, but not its structural isomer 2PY, induced expression of VCAM-1 and leukocyte adherence to vascular endothelium in mice. Collectively, these results indicate that the terminal breakdown products of excess niacin, 2PY and 4PY, are both associated with residual CVD risk. They also suggest an inflammation-dependent mechanism underlying the clinical association between 4PY and MACE.

Long non-coding RNA H19X as a regulator of mononuclear cell adhesion to the endothelium in systemic sclerosis.

To define the functional relevance of H19 X-linked (H19X) co-expressed long non-coding RNA (lncRNA) in endothelial cell (EC) activation as a key process in SSc vasculopathy. H19X expression in SSc skin biopsies was analysed from single-cell RNA sequencing (scRNA-seq) data. Differential expression and pathway enrichment analysis between cells expressing (H19Xpos) and non-expressing H19X (H19Xneg) cells was performed. H19X function was investigated in human dermal microvascular ECs (HDMECs) by silencing. H19X and EC adhesion molecule levels were analysed by real-time quantitative PCR and western blot after stimulation with pro-inflammatory cytokines. Cytoskeletal rearrangements were analysed by fluorescent staining. Endothelial adhesion was evaluated by co-culture of HDMECs and fluorescent-labelled peripheral blood mononuclear cells (PBMCs). Shedding vascular cell adhesion protein 1 (VCAM1) was evaluated by ELISA on HDMEC supernatant. The scRNA-seq data showed significant upregulation of H19X in SSc compared with healthy ECs. In HDMECs, H19X was consistently induced by IFN type I and II. H19X knockdown lead to a significant decrease in the mRNA of several adhesion molecules. In particular, VCAM1 was significantly reduced at the protein and mRNA levels. Co-expression analysis of the scRNA-seq data confirmed higher expression of VCAM1 in H19Xpos ECs. ECs were also strongly associated with the 'cell adhesion molecule' pathway. Moreover, the VCAM1 downstream pathway displayed less activation following H19X knockdown. Contractility of HDMECs, PBMC adhesion to HDMECs and VCAM1 shedding were also reduced following H19X knockdown. lncRNA H19X may contribute to EC activation in SSc vasculopathy, acting as a regulator of expression of adhesion molecules in ECs.

Mosaic loss of Y chromosome is associated with aging and epithelial injury in chronic kidney disease

BackgroundMosaic loss of Y chromosome (LOY) is the most common chromosomal alteration in aging men. Here, we use single-cell RNA and ATAC sequencing to show that LOY is present in the kidney and increases with age and chronic kidney disease.ResultsThe likelihood of a cell having LOY varies depending on its location in the nephron. Cortical epithelial cell types have a greater proportion of LOY than medullary or glomerular cell types, which may reflect their proliferative history. Proximal tubule cells are the most abundant cell type in the cortex and are susceptible to hypoxic injury. A subset of these cells acquires a pro-inflammatory transcription and chromatin accessibility profile associated with expression of HAVCR1, VCAM1, and PROM1. These injured epithelial cells have the greatest proportion of LOY and their presence predicts future kidney function decline. Moreover, proximal tubule cells with LOY are more likely to harbor additional large chromosomal gains and express pro-survival pathways. Spatial transcriptomics localizes injured proximal tubule cells to a pro-fibrotic microenvironment where they adopt a secretory phenotype and likely communicate with infiltrating immune cells.ConclusionsWe hypothesize that LOY is an indicator of increased DNA damage and potential marker of cellular senescence that can be applied to single-cell datasets in other tissues.

Efficacy of a mixed extract of Salvia miltiorrhiza and Paeonia lactiflora in inhibiting the aging of vascular wall through in vitro and in vivo experiments.

Vascular wall aging has been strongly associated with cardiovascular diseases. Thus, this study aimed to investigate the efficacy of USCP-GVH-014, a mixed extract of Salvia miltiorrhiza Bunge and Paeonia lactiflora Pall., in inhibiting vascular wall aging through invitro and in vivo experiments. The results revealed that USCP-GVH-014 inhibited abnormal cell proliferation, collagen overproduction, and MMP-2 and MMP-9 overexpression caused by various stimuli and recovered the antioxidant enzyme superoxide dismutase on human aortic smooth muscle cells. In addition, it inhibited the increase in ICAM-1 and VCAM-1 expression induced by tumor necrosis factor alpha on human aortic endothelial cells and prevented the aging of the vascular wall by regulating related proteins such as epidermal growth factor and interleukin-1ß. Furthermore, it reduced vascular aging in in vivo studies. These results demonstrate that USCP-GVH-014 effectively reduces vascular aging, thereby rendering it a potential therapeutic candidate for cardiovascular diseases.

Tripterygium Wilfordii Reduces the Inflammatory Response in Transplant Vasculopathy

Background: Transplant vasculopathy (TV) is the most challenging threat during organ transplantation. The pathological induction of TV results from vascular endothelial cell injury. This study established the in vitro model of TV and investigated the attenuating effect of triptolide (TPL) on TV through multiple in vitro and in vivo assays. Methods: The inflammation model was established by stimulating rat vascular endothelial cells with lipopolysaccharides (LPS). Rat vascular endothelial cells were isolated, cultured, and characterized as CD31-positive. For in vivo assays, Wistar and Sprague–Dawley rats were divided into S-S, W-S and W-S-TPL groups. The transplanted arteries were harvested at 1,4, and 8 weeks post-operation. Results: After LPS treatment, the cell viability was significantly reduced, and the IL-1β and TNF-α levels were significantly elevated, indicating that the LPS-induced in vitro inflammation model was successfully constructed. Nevertheless, the changes in the expression of inflammatory factors and vascular adhesion molecules, as well as the p-p65 levels, which were triggered by LPS, were significantly mitigated upon the stimulation of TPL. Results from rat models showed that intimal thickening was observed in the W-S group and significantly reduced in the TPL-treated group a week after transplantation. Consistent with the in vitro data, VCAM-1, ICAM-1, p-p38, and p-p65 expression was significantly attenuated in the TPL-treated group compared with the W-S group. Conclusion: This work confirmed that TPL may alleviate the progression of allograft vasculopathy and reduce the inflammation process by inhibiting the phosphorylation of p65 and p38, which are the key protein of the NF-κB and MAPK pathway, respectively.

Endothelial IL17RD promotes Western diet-induced aortic myeloid cell infiltration

The Interleukin-17 (IL17) family is a group of cytokines implicated in the etiology of several inflammatory diseases. Interleukin-17 receptor D (IL17RD), also known as Sef (similar expression to fibroblast growth factor) belonging to the family of IL17 receptors, has been shown to modulate IL17A-associated inflammatory phenotypes. The objective of this study was to test the hypothesis that IL17RD promotes endothelial cell activation and consequent leukocyte adhesion. We utilized primary human aortic endothelial cells and demonstrated that RNAi targeting of IL17RD suppressed transcript levels by 83 % compared to non-targeted controls. Further, RNAi knockdown of IL17RD decreased the adhesion of THP-1 monocytic cells onto a monolayer of aortic endothelial cells in response to IL17A. Additionally, we determined that IL17A did not significantly enhance the activation of canonical MAPK and NFκB pathways in endothelial cells, and further did not significantly affect the expression of VCAM-1 and ICAM-1 in aortic endothelial cells, which is contrary to previous findings. We also determined the functional relevance of our findings in vivo by comparing the expression of endothelial VCAM-1 and ICAM-1 and leukocyte infiltration in the aorta in Western diet-fed Il17rd null versus wild-type mice. Our results showed that although Il17rd null mice do not have significant alteration in aortic expression of VCAM-1 and ICAM-1 in endothelial cells, they exhibit decreased accumulation of proinflammatory monocytes and neutrophils, suggesting that endothelial IL17RD induced in vivo myeloid cell accumulation is not dependent on upregulation of VCAM-1 and ICAM-1 expression. We further performed proteomics analysis to identify potential molecular mediators of the IL17A/IL17RD signaling axis. Collectively, our results underscore a critical role for Il17rd in the regulation of aortic myeloid cell infiltration in the context of Western diet feeding.

Interferon Beta Modulation of Brain Endothelial Cell Activation in Ischemic Stroke

Background and Hypothesis:Every year, more than 690,000 people in the United States suffer an ischemic stroke. Many survivors are left with long-term disability. While the initial insult to the brain is caused by hypoxia resulting from cerebral artery occlusion, a secondary insult is caused by peripheral immune cell infiltration across the blood brain barrier (BBB) and subsequent cytotoxic insult. Previous studies have demonstrated that interferon beta (IFNβ) limits peripheral immune cell infiltration across the BBB and reduces brain infarction volume. We hypothesize that during ischemic stroke IFNβ suppresses brain endothelial cells (bECs) activation to reduce their expression of adhesion molecules as one of the mechanisms by which it decreases peripheral immune cell infiltration across the BBB.
 Experimental Design:In this project, bEnd.3 cells, a cell line of bECs, were activated by TNF-α, a pro-inflammatory cytokine. Tissue plasminogen activator (tPA), an FDA-approved thrombolytic for ischemic stroke, was included in the study. bEnd.3 cells were treated with IFNβ at 1.5 hours prior TNF-α or TNF-α + tPA stimulation to evaluate its modulation of adhesion cell expression. The adhesion molecule expression was determined by flow cytometry. Results were further confirmed by in vivo studies in which stroke animals were subjected to tPA treatment in the presence or absence of IFNβ.
 Results:Our results showed that TNF-α induced ICAM-1, VCAM-1, E-selectin, and P-selectin expression. Importantly, we found IFNβ suppressed the expression of aforementioned adhesion molecules in bEnd.3 cells treated with TNF-α or TNF-α+tPA. Our in vivo results demonstrated that IFNβ treatment reduced ICAM-1 and E-selectin, but not VCAM-1 or P-selectin expression in the ischemic brain.
 Conclusion and Potential Impact:Our study demonstrates that IFNβ modulates bEC expression of adhesion molecules in vitro and in vivo of ischemic stroke, suggesting IFNβ, an FDA-approved drug for Multiple Sclerosis, shows potential to improve ischemic stroke outcomes.

FSH Is Responsible for Androgen Deprivation Therapy-Associated Atherosclerosis in Mice by Exaggerating Endothelial Inflammation and Monocyte Adhesion.

Androgen deprivation therapy (ADT) is the mainstay treatment for advanced prostate cancer. But ADTs with orchiectomy and gonadotropin-releasing hormone (GnRH) agonist are associated with increased risk of cardiovascular diseases, which appears less significant with GnRH antagonist. The difference of follicle-stimulating hormone (FSH) in ADT modalities is hypothesized to be responsible for ADT-associated cardiovascular diseases. We administered orchiectomy, GnRH agonist, or GnRH antagonist in male ApoE-/- mice fed with Western diet and manipulated FSH levels by testosterone and FSH supplementation or FSH antibody to investigate the role of FSH elevation on atherosclerosis. By combining lipidomics, in vitro study, and intraluminal FSHR (FSH receptor) inhibition, we delineated the effects of FSH on endothelium and monocytes and the underlying mechanisms. Orchiectomy and GnRH agonist, but not GnRH antagonist, induced long- or short-term FSH elevation and significantly accelerated atherogenesis. In orchiectomized and testosterone-supplemented mice, FSH exposure increased atherosclerosis. In GnRH agonist-treated mice, blocking of short FSH surge by anti-FSHβ antibody greatly alleviated endothelial inflammation and delayed atherogenesis. In GnRH antagonist-treated mice, FSH supplementation aggravated atherogenesis. Mechanistically, FSH, synergizing with TNF-α (tumor necrosis factor alpha), exacerbated endothelial inflammation by elevating VCAM-1 (vascular cell adhesion protein 1) expression through the cAMP/PKA (protein kinase A)/CREB (cAMP response element-binding protein)/c-Jun and PI3K (phosphatidylinositol 3 kinase)/AKT (protein kinase B)/GSK-3β (glycogen synthase kinase 3 beta)/GATA-6 (GATA-binding protein 6) pathways. In monocytes, FSH upregulated CD29 (cluster of differentiation 29) expression via the PI3K/AKT/GSK-3β/SP1 (specificity protein 1) pathway and promoted monocyte-endothelial adhesion both in vitro and in vivo. Importantly, FSHR knockdown by shRNA in endothelium of carotid arteries markedly reduced GnRH agonist-induced endothelial inflammation and atherosclerosis in mice. FSH is responsible for ADT-associated atherosclerosis by exaggerating endothelial inflammation and promoting monocyte-endothelial adhesion.

Kupffer Cell Inactivation Alters Endothelial Cell Adhesion Molecules in Cecal Ligation and Puncture-Induced Sepsis.

The activation of Kupffer cells, resident macrophages in the liver, is closely associated with the inflammatory response during sepsis, which leads to multiple-organ failure. However, how Kupffer cell activation affects adhesion molecules (ICAM-1 and VCAM-1) in sepsis has not been determined. This study investigated Kupffer cell inactivation's (by gadolinium chloride; GdCl3) effects on adhesion molecule expression in CLP-induced sepsis. The induction of sepsis resulted in increased expression of liver and lung ICAM-1 and VCAM-1. GdCl3 pretreatment significantly decreased liver ICAM-1 expression but had no effect on VCAM-1 expression. In contrast, GdCl3 pretreatment had no effect on sepsis-induced increased adhesion molecule expression in the lungs. Similarly, the immunoreactivity of ICAM-1 was decreased in liver sinusoidal endothelial cells but increased in pulmonary endothelial cells in septic mice pretreated with GdCl3. Further, GdCl3 pretreatment had no effect on the immunoreactivity of VCAM-1 in endothelial cells of the liver and lungs. Hence, the findings of this study demonstrate the differential effects of Kupffer cell inactivation on liver and lung adhesion molecules and suggest the complexity of their involvement in the pathophysiology of sepsis.

P2Y2-P2X7 receptors cross-talk in primed mesenteric endothelial cells upregulates NF-κB signaling favoring mononuclear cell adhesion in schistosomiasis.

Schistosomiasis is an intravascular infectious disease that impacts over 200 million people globally. In its chronic stage, it leads to mesenteric inflammation with significant involvement of monocytes/macrophages. Endothelial cells lining the vessel lumens play a crucial role, and mount of evidence links this disease to a downregulation of endoprotective cell signaling favoring a primed and proinflammatory endothelial cell phenotype and therefore the loss of immunovascular homeostasis. One hallmark of infectious and inflammatory conditions is the release of nucleotides into the extracellular milieu, which, in turn, act as innate messengers, activating purinergic receptors and triggering cell-to-cell communication. ATP influences the progression of various diseases through P2X and P2Y purinergic receptor subtypes. Among these receptors, P2Y2 (P2Y2R) and P2X7 (P2X7R) receptors stand out, known for their roles in inflammation. However, their specific role in schistosomiasis has remained largely unexplored. Therefore, we hypothesized that endothelial P2Y2R and P2X7R could contribute to monocyte adhesion to mesenteric endothelial cells in schistosomiasis. Using a preclinical murine model of schistosomiasis associated with endothelial dysfunction and age-matched control mice, we showed that endothelial P2Y2R and P2X7R activation increased monocyte adhesion to cultured primary endothelial cells in both groups. However, a distinct upregulation of endothelial P2Y2R-driven canonical Ca2+ signaling was observed in the infected group, amplifying adhesion. In the control group, the coactivation of endothelial P2Y2R and P2X7R did not alter the maximal monocyte adhesion induced by each receptor individually. However, in the infected group, this coactivation induced a distinct upregulation of P2Y2R-P2X7R-driven canonical signaling, IL-1β release, and VCAM-1 expression, with underlying mechanisms involving inflammasome and NF-κB signaling. Therefore, current data suggest that schistosomiasis alters endothelial cell P2Y2R/P2X7R signaling during inflammation. These discoveries advance our understanding of schistosomiasis. This intricate interplay, driven by PAMP-triggered endothelial P2Y2R/P2X7R cross-talk, emerges as a potential key player in the mesenteric inflammation during schistosomiasis.

Ultrasound molecular imaging for early detection of acute renal ischemia-reperfusion injury.

Microcirculatory perfusion disorder and inflammatory response are critical links in acute kidney injury (AKI). We aim to construct anti-vascular cell adhesion molecule-1(VCAM-1) targeted microbubbles (TM) to monitor renal microcirculatory perfusion and inflammatory response. TM carrying VCAM-1 polypeptide was constructed by biological coupling. The binding ability of TM to human umbilical vein endothelial cells (HUVECs) was detected. Bilateral renal ischemia-reperfusion injury (IRI) models of mice were established to evaluate microcirculatory perfusion and inflammatory response using TM. Thirty-six mice were randomly divided into six groups according to the different reperfusion time (0.5, 2, 6, 12, and 24 h) and sham-operated group (Sham group). The correlation of TM imaging with serum and histopathological biomarkers was investigated. TM has advantages such as uniform distribution, regular shape, high stability, and good biosafety. TM could bind specifically to VCAM-1 molecule expressed by tumor necrosis factor-alpha (TNF-α)-treated HUVECs. In the renal IRI-AKI model, the area under the curve (AUC) of TM significantly decreased both in the renal cortical and medullary after 2 h of reperfusion compared with the Sham group (p < 0.05). Normalized intensity difference (NID) of TM at different reperfusion time was all higher than that of blank microbubbles (BM) and the Sham group (p < 0.05). Ultrasound molecular imaging of TM could detect AKI early before commonly used renal function markers, histopathological biomarkers, and BM imaging. AUC of TM was negatively correlated with serum creatinine (Scr), blood urea nitrogen (BUN), and Cystatin C (Cys-C) levels, and NID of TM was linearly correlated with VCAM-1, TNF-α, and interleukin-6 (IL-6) expression (p < 0.05). Ultrasound molecular imaging based on TM carrying VCAM-1 polypeptide can accurately evaluate the changes in renal microcirculatory perfusion and inflammatory response, which might be a promising modality for early diagnosis of AKI.

MiR-130a-3p inhibits endothelial inflammation by regulating the expression of MAPK8 in endothelial cells

MicroRNA-130a-3p (miR-130a-3p) has been reported as closely related to atherosclerosis (AS). This study is to survey the effects of miR-130a-3p in endothelial cells (ECs) treated with oxidized low-density lipoprotein (ox-LDL) and explore underlying mechanisms. The proliferation and apoptosis of ox-LDL-treated HUVEC cells were determined by CCK-8, EdU, and flow cytometry assays. ELISA and Western blot analysis measured the expressions of cytokines and protein levels. Bioinformatics and dual-luciferase reporter assay were performed to predict and confirm that Mitogen-activated protein kinase 8 (MAPK8) was a direct target of miR-130a-3p, and MAPK8 was negatively associated with miR-130a-3p. As expected, miR-130a-3p was down-regulated in ox-LDL-treated HUVEC cells, and up-regulation of miR-130a-3p promoted proliferation and inhibited apoptosis of ox-LDL-treated HUVEC cells. Furthermore, miR-130a-3p mimics suppressed the expressions of TNF-α and IL-6 and decreased the protein levels of VCAM-1, ICAM-1 and E-selectin. MAPK8 was highly expressed in ox-LDL-treated HUVEC cells, and silence of MAPK8 promoted proliferation inhibited apoptosis, suppressed inflammatory responses, and decreased the levels of VCAM-1, ICAM-1, and E-selectin, over-expression of MAPK8 partially restored the functional effects of miR-130a-3p on proliferation, inflammatory responses, and the expressions of VCAM-1, ICAM-1 and E-selectin. This study indicates that miR-130a-3p may emerge as an effective target for treating AS.

Hope for vascular cognitive impairment: Ac-YVAD-cmk as a novel treatment against white matter rarefaction.

Vascular cognitive impairment (VCI) is the second leading cause of dementia with limited treatment options, characterised by cerebral hypoperfusion-induced white matter rarefaction (WMR). Subcortical VCI is the most common form of VCI, but the underlying reasons for region susceptibility remain elusive. Recent studies employing the bilateral cortical artery stenosis (BCAS) method demonstrate that various inflammasomes regulate white matter injury and blood-brain barrier dysfunction but whether caspase-1 inhibition will be beneficial remains unclear. To address this, we performed BCAS on C57/BL6 mice to study the effects of Ac-YVAD-cmk, a caspase-1 inhibitor, on the subcortical and cortical regions. Cerebral blood flow (CBF), WMR, neuroinflammation and the expression of tight junction-related proteins associated with blood-brain barrier integrity were assessed 15 days post BCAS. We observed that Ac-YVAD-cmk restored CBF, attenuated BCAS-induced WMR and restored subcortical myelin expression. Within the subcortical region, BCAS activated the NLRP3/caspase-1/interleukin-1beta axis only within the subcortical region, which was attenuated by Ac-YVAD-cmk. Although we observed that BCAS induced significant increases in VCAM-1 expression in both brain regions that were attenuated with Ac-YVAD-cmk, only ZO-1 and occludin were observed to be significantly altered in the subcortical region. Here we show that caspase-1 may contribute to subcortical regional susceptibility in a mouse model of VCI. In addition, our results support further investigations into the potential of Ac-YVAD-cmk as a novel treatment strategy against subcortical VCI and other conditions exhibiting cerebral hypoperfusion-induced WMR.

Establishment of a novel amyotrophic lateral sclerosis patient (TARDBP N345K/+)-derived brain microvascular endothelial cell model reveals defective Wnt/β-catenin signaling: investigating diffusion barrier dysfunction and immune cell interaction.

Amyotrophic lateral sclerosis (ALS) is a major neurodegenerative disease for which there is currently no curative treatment. The blood-brain barrier (BBB), multiple physiological functions formed by mainly specialized brain microvascular endothelial cells (BMECs), serves as a gatekeeper to protect the central nervous system (CNS) from harmful molecules in the blood and aberrant immune cell infiltration. The accumulation of evidence indicating that alterations in the peripheral milieu can contribute to neurodegeneration within the CNS suggests that the BBB may be a previously overlooked factor in the pathogenesis of ALS. Animal models suggest BBB breakdown may precede neurodegeneration and link BBB alteration to the disease progression or even onset. However, the lack of a useful patient-derived model hampers understanding the pathomechanisms of BBB dysfunction and the development of BBB-targeted therapies. In this study, we differentiated BMEC-like cells from human induced pluripotent stem cells (hiPSCs) derived from ALS patients to investigate BMEC functions in ALS patients. TARDBP N345K/+ carrying patient-derived BMEC-like cells exhibited increased permeability to small molecules due to loss of tight junction in the absence of neurodegeneration or neuroinflammation, highlighting that BMEC abnormalities in ALS are not merely secondary consequences of disease progression. Furthermore, they exhibited increased expression of cell surface adhesion molecules like ICAM-1 and VCAM-1, leading to enhanced immune cell adhesion. BMEC-like cells derived from hiPSCs with other types of TARDBP gene mutations (TARDBP K263E/K263E and TARDBP G295S/G295S) introduced by genome editing technology did not show such BMEC dysfunction compared to the isogenic control. Interestingly, transactive response DNA-binding protein 43 (TDP-43) was mislocalized to cytoplasm in TARDBP N345K/+ carrying model. Wnt/β-catenin signaling was downregulated in the ALS patient (TARDBP N345K/+)-derived BMEC-like cells and its activation rescued the leaky barrier phenotype and settled down VCAM-1 expressions. These results indicate that TARDBP N345K/+ carrying model recapitulated BMEC abnormalities reported in brain samples of ALS patients. This novel patient-derived BMEC-like cell is useful for the further analysis of the involvement of vascular barrier dysfunctions in the pathogenesis of ALS and for promoting therapeutic drug discovery targeting BMEC.

The BET inhibitor apabetalone decreases neuroendothelial proinflammatory activation in vitro and in a mouse model of systemic inflammation.

Brain vascular inflammation is characterized by endothelial activation and immune cell recruitment to the blood vessel wall, potentially causing a breach in the blood - brain barrier, brain parenchyma inflammation, and a decline of cognitive function. The clinical-stage small molecule, apabetalone, reduces circulating vascular endothelial inflammation markers and improves cognitive scores in elderly patients by targeting epigenetic regulators of gene transcription, bromodomain and extraterminal proteins. However, the effect of apabetalone on cytokine-activated brain vascular endothelial cells (BMVECs) is unknown. Here, we show that apabetalone treatment of BMVECs reduces hallmarks of in vitro endothelial activation, including monocyte chemoattractant protein-1 (MCP-1) and RANTES chemokine secretion, cell surface expression of endothelial cell adhesion molecule VCAM-1, as well as endothelial capture of THP-1 monocytes in static and shear stress conditions. Apabetalone pretreatment of THP-1 downregulates cell surface expression of chemokine receptors CCR1, CCR2, and CCR5, and of the VCAM-1 cognate receptor, integrin α4. Consequently, apabetalone reduces THP-1 chemoattraction towards soluble CCR ligands MCP-1 and RANTES, and THP-1 adhesion to activated BMVECs. In a mouse model of brain inflammation, apabetalone counters lipopolysaccharide-induced transcription of endothelial and myeloid cell markers, consistent with decreased neuroendothelial inflammation. In conclusion, apabetalone decreases proinflammatory activation of brain endothelial cells and monocytes in vitro and in the mouse brain during systemic inflammation.

EXPRESSION OF GENES OF BIOMOLECULES ASSOCIATED WITH THE ETIOPATHOGENESIS OF ATHEROSCLEROSIS IN ATHEROSCLEROTIC PLAQUES OF CORONARY ARTERIES

HighlightsThe study showed differences in the expression of a number of genes in atherosclerotic plaques of different types in patients with coronary atherosclerosis. The obtained data can become the basis for the development of test systems in order to determine the dynamics of the atherosclerotic process and detect signs of destabilization of the atherosclerotic plaque as early as possible. Aim. To study the differential expression of genes encoding molecules associated with the etiopathogenesis of atherosclerosis by the method of genome-wide RNA sequencing in stable atherosclerotic plaque of fibrous type and unstable atherosclerotic plaque of dystopic-necrotic type.Methods. The study was performed on samples of atherosclerotic plaques of patients with coronary atherosclerosis without acute coronary syndrome with stable angina pectoris of functional class II–IV (FC) at the age of 45-65 years. Tissue sampling of atherosclerotic plaques was performed intraoperatively in the presence of indications. Genome-wide RNA sequencing was performed using Illumina’s TruSeq RNA Sample Preparation Kit (Illumina, USA).Results. An increase in the level of gene expression in stable atherosclerotic plaques was noted for A2M, ADAMTS13, CSF3, CX3CL1, CXCL1, FGF2, GDF15, ICAM1, IL1A, IL1B, IL6, IL10, PDGFA, PTX3. There was an eightfold statistically significant increase in the level of CFD, CXCL16, FABP4, FLT3, IFNG, IL7, IL15, SELL, TGFA, THBD, TNNT1, VCAM1 and VEGFA gene expression (p&lt;0,001) in unstable atherosclerotic plaques of dystrophic-necrotic type.Conclusion. The study showed differences in the expression of a number of genes in atherosclerotic plaques of different types in patients with coronary atherosclerosis. The obtained data can become the basis for the development of test systems in order to determine the dynamics of the atherosclerotic process and detect signs of destabilization of the atherosclerotic plaque as early as possible.

Tumor-Derived Exosomal LINC01480 Upregulates VCAM1 Expression by Acting as a Competitive Endogenous RNA of miR-204-5p to Promote Gastric Cancer Progression.

Exosomes are a type of cell-derived vesicles that range in size from 30 to 100 nm. They are widely present in various organisms and participate in diverse biological processes, playing crucial roles in tumorigenesis and progression. This study aimed to investigate whether LINC01480 in tumor-derived exosomes is involved in the molecular mechanism of gastric cancer by competitively upregulating the VCAM1 expression through binding miR-204-5p. The study analyzed transcriptome data related to gastric cancer from the cancer genome atlas database and constructed a risk-scoring model for epithelial-mesenchymal transition (EMT)-related lncRNAs to identify eight EMT-related lncRNAs associated with prognosis. EMT-related mRNAs positively correlated with LINC01480 were screened in the ExoRBase database. In vitro cell experiments showed that exosomal LINC01480 can promote the proliferation, migration, invasion, and EMT of gastric cancer cells by upregulating VCAM1 expression through competitive binding with miR-204-5p. In vivo experiments on nude mice showed that exosomal LINC01480 promotes the development of gastric cancer. These results suggest that exosomal LINC01480 could be a potential therapeutic target for gastric cancer.

The Protective Role of Mitochondria-Associated Endoplasmic Reticulum Membrane (MAM) Protein Sigma-1 Receptor in Regulating Endothelial Inflammation and Permeability Associated with Acute Lung Injury.

Earlier studies from our lab identified endoplasmic reticulum (ER) chaperone BiP/GRP78, an important component of MAM, to be a novel determinant of endothelial cell (EC) dysfunction associated with acute lung injury (ALI). Sigma1R (Sig1R) is another unique ER receptor chaperone that has been identified to associate with BiP/GRP78 at the MAM and is known to be a pluripotent modulator of cellular homeostasis. However, it is unclear if Sig1R also plays a role in regulating the EC inflammation and permeability associated with ALI. Our data using human pulmonary artery endothelial cells (HPAECs) showed that siRNA-mediated knockdown of Sig1R potentiated LPS-induced the expression of proinflammatory molecules ICAM-1, VCAM-1 and IL-8. Consistent with this, Sig1R agonist, PRE-084, known to activate Sig1R by inducing its dissociation from BiP/GRP78, blunted the above response. Notably, PRE-084 failed to blunt LPS-induced inflammatory responses in Sig1R-depleted cells, confirming that the effect of PRE-084 is driven by Sig1R. Furthermore, Sig1R antagonist, NE-100, known to inactivate Sig1R by blocking its dissociation from BiP/GRP78, failed to block LPS-induced inflammatory responses, establishing that dissociation from BiP/GRP78 is required for Sig1R to exert its anti-inflammatory action. Unlike Sig1R, the siRNA-mediated knockdown or Subtilase AB-mediated inactivation of BiP/GRP78 protected against LPS-induced EC inflammation. Interestingly, the protective effect of BiP/GRP78 knockdown or inactivation was abolished in cells that were depleted of Sig1R, confirming that BiP/GRP78 knockdown/inactivation-mediated suppression of EC inflammation is mediated via Sig1R. In view of these findings, we determined the in vivo relevance of Sig1R in a mouse model of sepsis-induced ALI. The intraperitoneal injection of PRE-084 mitigated sepsis-induced ALI, as evidenced by a decrease in ICAM-1, IL-6 levels, lung PMN infiltration, and lung vascular leakage. Together, these data evidence a protective role of Sig1R against endothelial dysfunction associated with ALI and identify it as a viable target in terms of controlling ALI in sepsis.

Endoglin and soluble endoglin in liver sinusoidal endothelial dysfunction in vivo

Liver sinusoidal endothelial cells (LSECs) play a crucial role in regulating the hepatic function. Endoglin (ENG), a transmembrane glycoprotein, was shown to be related to the development of endothelial dysfunction. In this study, we hypothesized the relationship between changes in ENG expression and markers of liver sinusoidal endothelial dysfunction (LSED) during liver impairment.Male C57BL/6J mice aged 9–12 weeks were fed with 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) diet (intrahepatic cholestasis) or choline-deficient l-amino acid defined high-fat diet (CDAA-HFD) (non-alcoholic steatohepatitis (NASH)).Significant increases in liver enzymes, fibrosis, and inflammation biomarkers were observed in both cholestasis and NASH. Decreased p-eNOS/eNOS and VE-cadherin protein expression and a significant increase in VCAM-1 and ICAM-1 expression were detected, indicating LSED in both mouse models of liver damage. A significant reduction of ENG in the DDC-fed mice, while a significant increase of ENG in the CDAA-HFD group was observed. Both DDC and CDAA-HFD-fed mice showed a significant increase in MMP-14 protein expression, which is related to significantly increased levels of soluble endoglin (sENG) in the plasma.In conclusion, we demonstrated that intrahepatic cholestasis and NASH result in an altered ENG expression, predominantly in LSECs, suggesting a critical role of ENG expression for the proper function of liver sinusoids. Both pathologies resulted in elevated sENG levels, cleaved by MMP-14 expressed predominantly from LSECs, indicating sENG as a liver injury biomarker.