Expression Of Adhesion Molecules Research Articles

Angiogenic potential in periodontal stem cells from upper and lower jaw: A pilot study.

Teeth and supporting oral tissues are attractive and accessible sources of stem cells. Periodontal ligament stem cells (PDLSC) are readily isolated from extracted third molars, and exhibit the ability to self-renew and differentiate into multiple mesodermal cell fates. Clinical experience suggests that the exact location of periodontal defects affects the oral bone remodeling and wound healing. Compared to the mandible, the maxilla heals quicker and more efficiently. Angiogenesis is key in tissue regeneration including dental tissues, yet few studies focus on the angiogenic potential of PDLSC, none of which considered the differences between upper and lower jaw PDLSC (u-PDLSC and l-PDLSC, respectively). Here we studied the angiogenic potential of u-PDLSC and l-PDLSC and compared the results to well-established mesenchymal stem cells (MSC). Cells were characterized in terms of surface markers, proliferation, and vascular endothelial growth factor (VEGF) secretion, and angiogenic assays were performed. Newly formed capillaries were stained with CD31, and their expression of platelet endothelial cell adhesion molecule (PECAM-1), angiopoietin 2 (ANGPT2), and vascular endothelial growth factor receptor 1 and 2 (VEGFR-1, VEGFR-2) were measured. Periodontal stem cells from the upper jaw showed a higher proliferation capacity, secreted more VEGF, and formed capillary networks faster and denser than l-PDLSC. Gene expression of angiogenesis-related genes was significantly higher in u-PDLSC than in l-PDLSC or MSC, given that culture conditions were suitable. The oral cavity is a valuable source of stem cells, particularly PDLSC, which are promising for oral tissue engineering due to their robust growth, lifelong accessibility, low immunogenicity, and strong differentiation potential. Notably, u-PDLSC exhibit higher VEGF secretion and accelerate capillary formation compared to l-PDLSC or MSC. This study suggests a potential molecular mechanism in capillary formation, emphasizing the significance of precise location isolation of PDLSC.

Content and tissue expression of Collagen I, Collagen IV, and Laminin in the Extracellular Matrix in Prostate Adenocarcinoma.

Prostate cancer is one of the most common neoplasm in the male population. It is not known why some tumors become more aggressive than others. Although most studies show changes in the expression of cell adhesion molecules and the extracellular matrix correlated with the Gleason score, no study has objectively measured the tissue content of these molecules. This study aims to measure the content and tissue expression of collagen type I and IV and laminin in the extracellular matrix of patients with prostate adenocarcinoma and correlate these findings with the Gleason score and clinical characteristics. Forty-one patients who underwent radical prostate surgery at the Urology Department of a reference Hospital in Brazil between January 2015 and December 2020 were studied. The tissue protein content was estimated under light microscopy at a final magnification of 200 × . The mean collagen I score in prostate adenocarcinoma tissue samples was 7.16 ± 1.03 pixels/field. The mean type IV collagen score was 3.44 ± 0.61 pixels/field. The mean laminin score was 5.19 ± 0.79 pixels/field. The total Gleason score was correlated with both collagen and laminin. All the correlations were negative, which shows that the higher the collagen/laminin expression was, the lower the total Gleason score (p-value < 0,05). According to the Pearson correlation analysis, age has no statistical relationship with collagen and laminin content. PSA, in turn, showed a correlation only with laminin, but r = -0.378 (p = 0.015). Among the associated diseases and lifestyle habits, there is only statistical significance in the comparison of alcoholism for collagen I. For collagen IV and laminin, no statistical significance was obtained with the clinical variables analyzed.

17β-estradiol and methylprednisolone association as a therapeutic option to modulate lung inflammation in brain-dead female rats.

Brain death (BD) is known to compromise graft quality by causing hemodynamic, metabolic, and hormonal changes. The abrupt reduction of female sex hormones after BD was associated with increased lung inflammation. The use of both corticoids and estradiol independently has presented positive results in modulating BD-induced inflammatory response. However, studies have shown that for females the presence of both estrogen and corticoids is necessary to ensure adequate immune response. In that sense, this study aims to investigate how the association of methylprednisolone (MP) and estradiol (E2) could modulate the lung inflammation triggered by BD in female rats. Female Wistar rats (8 weeks) were divided into four groups: sham (animals submitted to the surgical process, without induction of BD), BD (animals submitted to BD), MP/E2 (animals submitted to BD that received MP and E2 treatment 3h after BD induction) and MP (animals submitted to BD that received MP treatment 3h after BD induction). Hemodynamics, systemic and local quantification of IL-6, IL-1β, VEGF, and TNF-α, leukocyte infiltration to the lung parenchyma and airways, and adhesion molecule expression were analyzed. After treatment, MP/E2 association was able to reinstate mean arterial pressure to levels close to Sham animals (p<0.05). BD increased leukocyte infiltration to the airways and MP/E2 was able to reduce the number of cells (p=0.0139). Also, the associated treatment modulated the vasculature by reducing the expression of VEGF (p=0.0616) and maintaining eNOS levels (p=0.004) in lung tissue. Data presented in this study show that the association between corticoids and estradiol could represent a better treatment strategy for lung inflammation in the female BD donor by presenting a positive effect in the hemodynamic management of the donor, as well as by reducing infiltrated leukocyte to the airways and release of inflammatory markers in the short and long term.

Endothelial dysfunction in neurodegenerative disease: Is endothelial inflammation an overlooked druggable target?

Neurological diseases with a neurodegenerative component have been associated with alterations in the cerebrovasculature. At the anatomical level, these are centred around changes in cerebral blood flow and vessel organisation. At the molecular level, there is extensive expression of cellular adhesion molecules and increased release of pro-inflammatory mediators. Together, these has been found to negatively impact blood-brain barrier integrity. Systemic inflammation has been found to accelerate and exacerbate endothelial dysfunction, neuroinflammation and degeneration. Here, we review the role of cerebrovasculature dysfunction in neurodegenerative disease and discuss the potential contribution of intermittent pro-inflammatory systemic disease in causing endothelial pathology, highlighting a possible mechanism that may allow broad-spectrum therapeutic targeting in the future.

Regnase-1 D141N mutation induces CD4+ T cell-mediated lung granuloma formation via upregulation of Pim2.

Regnase-1 is an RNase that plays a critical role in negatively regulating immune responses by destabilizing inflammatory messenger RNAs (mRNAs). Dysfunction of Regnase-1 can be a major cause of various inflammatory diseases with tissue injury and immune cell infiltration into organs. This study focuses on the role of the RNase activity of Regnase-1 in developing inflammatory diseases. We have constructed mice with a single point mutation at the catalytic center of the Regnase-1 RNase domain, which lacks endonuclease activity. D141N mutant mice demonstrated systemic inflammation, immune cell infiltration into various organs, and progressive development of lung granuloma. CD4+ T cells, mainly affected by this mutation, upregulated the mTORC1 pathway and facilitated the autoimmune trait in the D141N mutation. Moreover, serine/threonine kinase Pim2 contributed to lung inflammation in this mutation. Inhibition of Pim2 kinase activity ameliorated granulomatous inflammation, immune cell infiltration, and proliferation in the lungs. Additionally, Pim2 inhibition reduced the expression of adhesion molecules on CD4+ T cells, suggesting a role for Pim2 in facilitating leukocyte adhesion and migration to inflamed tissues. Our findings provide new insights into the role of Regnase-1 RNase activity in controlling immune functions and underscore the therapeutic relevance of targeting Pim2 to modulate abnormal immune responses.

The Immunomodulatory Effects and Mechanism of Wogonin for the Treatment of Multiple Sclerosis in a Murine Experimental Autoimmune Encephalomyelitis Model

Abstract Multiple sclerosis (MS) stands out as the most prevalent inflammatory autoimmune disease affecting the central nervous system (CNS). The primary driver of MS involves myelin-specific autoreactive T cells that infiltrate the CNS, initiating attacks on myelinated axons and triggering neuroinflammatory responses. Wogonin, a flavonoid, exhibits pharmacologic properties, including neuroprotective, anticancer, antiviral, anti-inflammatory, and anti-oxyradical effects. Our study demonstrates that wogonin significantly suppresses TLR-dependent DC activation, migration, and the expression of adhesion molecules in LPS-stimulated bone marrow-derived dendritic cells (BMDCs). In the experimental autoimmune encephalomyelitis (EAE) model, preventive treatment with wogonin led to reduced lymphocyte infiltration, prevented demyelination in the CNS, and consequently mitigated disease symptoms. Moreover, wogonin suppressed the release of neuroinflammatory cytokines by activated microglial cells, Th1 cells, Th2 cells, and Th17 cells in vitro and exerted therapeutic effects on attenuating the disease severity in the early disease onset stage and late disease progression stage. Our study suggests that wogonin treatment may not only prevent the onset of MS by inhibiting DC activation and migration but also potentially ameliorate the progression of MS by reducing neuroinflammation. This opens avenues for the development of new approaches in the therapy of autoimmune diseases.

A human pluripotent stem cell-derived in vitro model of the blood–brain barrier in cerebral malaria

BackgroundBlood–brain barrier (BBB) disruption is a central feature of cerebral malaria (CM), a severe complication of Plasmodium falciparum (Pf) infections. In CM, sequestration of Pf-infected red blood cells (Pf-iRBCs) to brain endothelial cells combined with inflammation, hemolysis, microvasculature obstruction and endothelial dysfunction mediates BBB disruption, resulting in severe neurologic symptoms including coma and seizures, potentially leading to death or long-term sequelae. In vitro models have advanced our knowledge of CM-mediated BBB disruption, but their physiological relevance remains uncertain. Using human induced pluripotent stem cell-derived brain microvascular endothelial cells (hiPSC-BMECs), we aimed to develop a novel in vitro model of the BBB in CM, exhibiting enhanced barrier properties.MethodshiPSC-BMECs were co-cultured with HB3var03 strain Pf-iRBCs up to 9 h. Barrier integrity was measured using transendothelial electrical resistance (TEER) and sodium fluorescein permeability assays. Localization and expression of tight junction (TJ) proteins (occludin, zonula occludens-1, claudin-5), cellular adhesion molecules (ICAM-1, VCAM-1), and endothelial surface markers (EPCR) were determined using immunofluorescence imaging (IF) and western blotting (WB). Expression of angiogenic and cell stress markers were measured using multiplex proteome profiler arrays.ResultsAfter 6-h of co-culture with Pf-iRBCs, hiPSC-BMECs showed reduced TEER and increased sodium fluorescein permeability compared to co-culture with uninfected RBCs, indicative of a leaky barrier. We observed disruptions in localization of occludin, zonula occludens-1, and claudin-5 by IF, but no change in protein expression by WB in Pf-iRBC co-cultures. Expression of ICAM-1 and VCAM-1 but not EPCR was elevated in hiPSC-BMECs with Pf-iRBC co-culture compared to uninfected RBC co-culture. In addition, there was an increase in expression of angiogenin, platelet factor-4, and phospho-heat shock protein-27 in the Pf-iRBCs co-culture compared to uninfected RBC co-culture.ConclusionThese findings demonstrate the validity of our hiPSC-BMECs based model of the BBB, that displays enhanced barrier integrity and appropriate TJ protein localization. In the hiPSC-BMEC co-culture with Pf-iRBCs, reduced TEER, increased paracellular permeability, changes in TJ protein localization, increase in expression of adhesion molecules, and markers of angiogenesis and cellular stress all point towards a novel model with enhanced barrier properties, suitable for investigating pathogenic mechanisms underlying BBB disruption in CM.

A novel dual inhibitor of the angiotensin and endothelin systems for the treatment of hypertension

Hypertension, affecting nearly half of the US adult population, is a primary cause of morbidity and mortality worldwide. Despite the availability of several classes of antihypertensive medications, only 25% of patients have their blood pressure under control, highlighting the need for novel antihypertensive therapeutics with better effcacy. Among hypertensive cases, salt-sensitive and drug-resistant hypertension are particularly concerning, as they are unresponsive to a combined regimen of three or more classes of medications, including angiotensin II (Ang II) receptor blockers (ARBs), angiotensin-converting enzyme inhibitors (ACEis), and diuretics. Salt-sensitive, observed in 50% of the general and 73% of black hypertensive patients, disproportionately affects the latter, leading to significantly higher mortality. Salt-sensitive hypertension in humans, as well as in various experimental models of hypertension (e.g., Ang II-induced, Dahl-salt sensitive [DSS] rats, deoxycorticosterone acetate [DOCA]-salt hypertensive rats), is characterized by significantly elevated levels of plasma endothelin-1 (ET-1), oxidative stress, and kidney dysfunction. ET-1 is often considered a final common pathway in orchestrating cardiovascular dysfunction through a range of proinflammatory mediators such as Ang II, cytokines, hypoxia, thrombin, reactive oxygen species (ROS), oxidized LDL, and vasopressin, all of which stimulate ET-1 production. In this capacity, we report the discovery of a gut microbiota-derived metabolite, indole-3-acetic acid (IAA), that simultaneously blocks ET-1 receptors (ETAR and ETBR) and the ET-biosynthetic enzyme ECE-1. Our data demonstrate that IAA inhibits ET-1-induced constriction of resistance mesenteric arteries, ROS production in human aortic smooth muscle cells, activation of cultured endothelial cells, adhesion molecule expression, and leukocyte adhesion. IAA inhibited purified human ECE-1 and suppressed Ang-II-induced and hypoxia-elicited ET-1 production from endothelial cells in vitro. Administration of a once-daily, extended-release formulation of IAA inhibits blood pressure elevation and kidney dysfunction in both DSS and Ang II-induced hypertension. IAA treatment preserved endothelial nitric oxide (NO) production and maintained smooth muscle sensitivity to a NO donor, sodium nitroprusside. Furthermore, IAA inhibited the expression of ROS-associated proteins such as Nox2 and Nox4, reduced oxidative damages, and decreased the production of ET-1 in vivo. Overall, IAA is the first compound that, through novel dual targeting of both the angiotensin and endothelin systems, demonstrates profound antihypertensive action in both salt-sensitive and angiotensin II-induced hypertension. Thus, IAA has the potential to effectively treat hypertension that does not respond to conventional therapies. American Heart Association. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Abstract 121: Ponatinib Promotes Endothelial Cell Inflammation And Leukocyte Trafficking Via Tumor Necrosis Factor Signaling: A Potential Mechanism Of Vascular Toxicity

Background: ABL tyrosine kinase inhibitors (TKI) have dramatically improved chronic myeloid leukemia (CML) survival. Due to resistance to the first ABL-TKI imatinib (Ima), most patients are treated with newer agents, including ponatinib (Pon). Although Pon improves cancer remission, the survival benefit is mitigated by a 4-fold increase in arterial thrombosis. The mechanism of Pon-induced arterial thrombosis and the safety of Asciminib (Asc), a newly approved alternative, are not known. CML patients are older and have notably elevated serum TNFa levels. Hypothesis: Since arterial thrombotic events occur when inflamed vascular plaques rupture, we tested the hypothesis that Pon enhances endothelial cell (EC) response to TNFa, promoting EC inflammation and leukocyte trafficking. Methods and Results: In vitro: Human umbilical vein ECs (HUVECs) were treated with Pon, Ima, or Asc at concentrations observed in humans. Adhesion molecules that mediate leukocyte rolling (E- and P-selectin) and adhesion (ICAM1, VCAM1), and TNF receptors (TNFR) 1 and 2 were quantified at 24 hours by qPCR, flow cytometry (FC), and immunoblot. Pon increased TNFR2, P-selectin, VCAM, and ICAM by FC, qPCR, and immunoblot while Ima and Asc had no impact. In primary human coronary artery ECs Pon, but not Asc nor Ima, similarly increased VCAM and P-selectin mRNA. Pretreatment of HUVECs with a nonspecific TNFR inhibitor prevented Pon induction of ICAM1 and P-selectin mRNA. In vivo: Mice were treated with ABL TKIs by oral gavage to achieve drug plasma levels consistent with those in CML patients. After three days, leukocyte trafficking was quantified by intravital microscopy (IVM) in mouse mesenteric vessels. Pon, but not Asc, increased the number of rolling and adherent leukocytes, and increased serum TNFa by ELISA. Treatment with a TNFR inhibitor blocked Pon induced leukocyte trafficking by IVM. Conclusion: Pon, but not Ima nor Asc, induces expression of TNF receptors and adhesion molecules in human ECs in vitro and promotes leukocyte trafficking in vivo. Since inflamed plaques lead to acute arterial thrombosis, this mechanism could contribute to the high risk of arterial thrombosis in Pon-treated cancer patients. If so, Asc may be a safer choice as it does not induce EC inflammation.

PM2.5 Temporally Decreases Human Brain Endothelial Barrier Proteins and Concomitantly Increases Inflammation and Autophagy in a Dose Dependent Manner

Exposure to airborne particulate matter 2.5 (particle size &lt;PM2.5 um) pollution is a significant co-morbidity for stroke and other cardio- and cerebrovascular diseases, causing premature mortality. However, mechanisms underlying PM2.5-mediated endothelial dysfunction and integrity loss are not known. Our goal was to elucidate how PM2.5 alters brain endothelial phenotype causing inflammation and loss of barrier integrity to provide better insight to the potential pathogenesis of PM2.5. We hypothesized that PM2.5 would alter integral endothelial barrier proteins as well as inflammatory mediators in a dose and temporal dependent manner, subsequently exacerbating the impact of acute ischemic-like injury. Primary adult male human brain microvascular endothelial cells (HBMEC) were preconditioned with PM2.5 (300, 75, or 15 μg/m3; 12, 24, or 36h) or vehicle, then exposed to normoxia (21% O2) or ischemic-like injury, hypoxia plus glucose deprivation (HGD; 1% O2), for 3h. HBMEC barrier function was assessed via trans-endothelial electrical resistance (TEER). HBMEC levels or expression of barrier markers (claudin-5, occludin, ZO-1), adhesion molecules (ICAM-1 and PECAM-1), inflammatory proteins (iNOS, COX-2, LOX-1, TNF-α, IL-1β), and autophagic protein (Beclin-1) were examined using qRT-PCR, in cell western, and standard immunoblotting. Ischemia-like injury as well as PM2.5 decreased HBMEC barrier function. PM2.5 also induced a concomitant temporal and dose dependent decrease in claudin-5 and increase in iNOS as well as TNF-α levels. Additionally, PM2.5 increased protein levels of beclin-1 and LOX-1 like that of HGD alone; however, combination of PM2.5 and HGD decreased levels of beclin-1. HGD alone or in combination with PM2.5 decreased levels of PECAM-1, but PM2.5 alone did not alter PECAM-1. PM2.5 in the presence or absence of HGD induced differential alteration in claudin-5 isoform protein levels. Intriguingly, PM2.5 decreased claudin-5 mRNA levels like that observed in HBMECs exposed to HGD alone. PM2.5 plus HGD significantly increased mRNA levels of both ICAM-1 and IL-1β. In conclusion, PM2.5 mediated inflammatory and vascular pathogenesis involves a potential complex intracellular molecular framework associated with the brain endothelium which is compounded by ischemic like injury. These findings have revealed detrimental effects of PM2.5 on the brain vascular endothelium which will help to identify targets in the endothelium to restore or preserve cerebrovascular homeostasis. University of Arizona Valley Research Partnership Grant VRP37 P2 (RJG), American Heart Association 19AIREA34480018 (RJG), UA VRP Grant VRP55 P1a (TSW). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Targeting the High-Density Lipoprotein Proteome for the Treatment of Post-Acute Sequelae of SARS-CoV-2.

Here, we target the high-density lipoprotein (HDL) proteome in a case series of 16 patients with post-COVID-19 symptoms treated with HMG-Co-A reductase inhibitors (statin) plus angiotensin II type 1 receptor blockers (ARBs) for 6 weeks. Patients suffering from persistent symptoms (post-acute sequelae) after serologically confirmed SARS-CoV-2 infection (post-COVID-19 syndrome, PCS, n = 8) or following SARS-CoV-2 vaccination (PVS, n = 8) were included. Asymptomatic subjects with corresponding serological findings served as healthy controls (n = 8/8). HDL was isolated using dextran sulfate precipitation and the HDL proteome of all study participants was analyzed quantitatively by mass spectrometry. Clinical symptoms were assessed using questionnaires before and after therapy. The inflammatory potential of the patients' HDL proteome was addressed in human endothelial cells. The HDL proteome of patients with PCS and PVS showed no significant differences; however, compared to controls, the HDL from PVS/PCS patients displayed significant alterations involving hemoglobin, cytoskeletal proteins (MYL6, TLN1, PARVB, TPM4, FLNA), and amyloid precursor protein. Gene Ontology Biological Process (GOBP) enrichment analysis identified hemostasis, peptidase, and lipoprotein regulation pathways to be involved. Treatment of PVS/PCS patients with statins plus ARBs improved the patients' clinical symptoms. After therapy, three proteins were significantly increased (FAM3C, AT6AP2, ADAM10; FDR < 0.05) in the HDL proteome from patients with PVS/PCS. Exposure of human endothelial cells with the HDL proteome from treated PVS/PCS patients revealed reduced inflammatory cytokine and adhesion molecule expression. Thus, HDL proteome analysis from PVS/PCS patients enables a deeper insight into the underlying disease mechanisms, pointing to significant involvement in metabolic and signaling disturbances. Treatment with statins plus ARBs improved clinical symptoms and reduced the inflammatory potential of the HDL proteome. These observations may guide future therapeutic strategies for PVS/PCS patients.

Corn Silk Polysaccharide Reduces the Risk of Kidney Stone Formation by Reducing Oxidative Stress and Inhibiting COM Crystal Adhesion and Aggregation.

HK-2 cells were damaged by 100 nm COM crystals to build an injury model. The cells were protected by CSPs with -COOH contents of 3.92% (CSP0) and 16.38% (CCSP3), respectively. The changes in the biochemical indexes of HK-2 cells and the difference in adhesion amount and aggregation degree of nano-COM on the cell surface before and after CSP protection were detected. CSP0 and CCSP3 protection can obviously inhibit HK-2 cell damage caused by nano-COM crystals, restore cytoskeleton morphology, reduce intracellular ROS level, inhibit phosphoserine eversion, restore the polarity of the mitochondrial membrane potential, normalize the cell cycle process, and reduce the expression of adhesion molecules, OPN, Annexin A1, HSP90, HAS3, and CD44 on the cell surface. Finally, the adhesion and aggregation of nano-COM crystals on the cell surface were effectively inhibited. The carboxymethylated CSP3 exhibited a higher protective effect on cells than the original CSP0, and cell viability was further improved with the increase in polysaccharide concentration. CSPs can protect HK-2 cells from calcium oxalate crystal damage and effectively reduce the adhesion and aggregation of nano-COM crystals on the cell surface, which is conducive to inhibiting the formation of calcium oxalate kidney stones.

Coal-fired PM2.5 induces endothelial cell injury and the expression of atherosclerosis-related adhesion molecules: Involvement of the p38 and JNK signaling pathways

It has been reported that PM2.5 causes endothelial cell injury and promotes atherosclerosis. However, the exact mechanism through which coal-fired PM2.5 induces endothelial cell injury, and the involvement of the p38 and JNK signaling pathways in this process remain unclear. In this study, EA.hy926 cells were exposed to coal-fired PM2.5 at concentrations of 25, 50, and 100 μg/mL, and the toxic effects were observed. The phosphorylation of the JNK and p38 signaling pathways was investigated through western blot analysis. Additionally, the expression of adhesion molecules (ICAM-1 and E-selectin) was assessed using ELISA and flow cytometry. Changes in cellular toxicity and adhesion molecules were evaluated after pretreatment with the p38 inhibitor SB203580 (10 μM) and the JNK inhibitor SP600125 (25 μM). We observed that exposure to coal-fired PM2.5 led to a decrease in cell survival rate and proliferation while promoting apoptosis. Exposure to coal-fired PM2.5 promoted the expression of ICAM-1 and E-selectin, as well as the phosphorylation of p38 and JNK in EA.hy926 cells. After using p38 and JNK inhibitors, there was an observed increase in cell survival rate and proliferation, accompanied by a decrease in apoptosis. The levels of ICAM-1 and E-selectin showed no significant changes with the addition of p38 and JNK inhibitors. Our results indicated that coal-fired PM2.5 caused cellular toxicity and increased the levels of ICAM-1 and E-selectin. The p38 and JNK signaling pathways might play a role in the reduction of cell viability, while the regulation of ICAM-1 and E-selectin might not be influenced by these pathways.

Transgenic human C-reactive protein affects oxidative stress but not inflammation biomarkers in the aorta of spontaneously hypertensive rats

BackgroundC-reactive protein (CRP) is an acute inflammatory protein detected in obese patients with metabolic syndrome. Moreover, increased CRP levels have been linked with atherosclerotic disease, congestive heart failure, and ischemic heart disease, suggesting that it is not only a biomarker but also plays an active role in the pathophysiology of cardiovascular diseases. Since endothelial dysfunction plays an essential role in various cardiovascular pathologies and is characterized by increased expression of cell adhesion molecules and inflammatory markers, we aimed to detect specific markers of endothelial dysfunction, inflammation, and oxidative stress in spontaneously hypertensive rats (SHR) expressing human CRP. This model is genetically predisposed to the development of the metabolic syndrome.MethodsTransgenic SHR male rats (SHR-CRP) and non-transgenic SHR (SHR) at the age of 8 months were used. Metabolic profile (including serum and tissue triglyceride (TAG), serum insulin concentrations, insulin-stimulated incorporation of glucose, and serum non-esterified fatty acids (NEFA) levels) was measured. In addition, human serum CRP, MCP-1 (monocyte chemoattractant protein-1), and adiponectin were evaluated by means of ELISA, histological analysis was used to study morphological changes in the aorta, and western blot analysis of aortic tissue was performed to detect expression of endothelial, inflammatory, and oxidative stress markers.ResultsThe presence of human CRP was associated with significantly decreased insulin-stimulated glycogenesis in skeletal muscle, increased muscle and hepatic accumulation of TAG and decreased plasmatic cGMP concentrations, reduced adiponectin levels, and increased monocyte chemoattractant protein-1 (MCP-1) levels in the blood, suggesting pro-inflammatory and presence of multiple features of metabolic syndrome in SHR-CRP animals. Histological analysis of aortic sections did not reveal any visible morphological changes in animals from both SHR and SHR-CRP rats. Western blot analysis of the expression of proteins related to the proper function of endothelium demonstrated significant differences in the expression of p-eNOS/eNOS in the aorta, although endoglin (ENG) protein expression remained unaffected. In addition, the presence of human CRP in SHR in this study did not affect the expression of inflammatory markers, namely p-NFkB, P-selectin, and COX2 in the aorta. On the other hand, biomarkers related to oxidative stress, such as HO-1 and SOD3, were significantly changed, indicating the induction of oxidative stress.ConclusionsOur findings demonstrate that CRP alone cannot fully induce the expression of endothelial dysfunction biomarkers, suggesting other risk factors of cardiovascular disorders are necessary to be involved to induce endothelial dysfunction with CRP.

Carboxymethylated Rhizoma alismatis polysaccharides reduces the risk of calcium oxalate stone formation by reducing cellular inflammation and oxidative stress.

This study aims to elucidate the mechanism and potential of Rhizoma alismatis polysaccharides (RAPs) in preventing oxidative damage to human renal proximal tubule epithelial cells. The experimental approach involved incubating HK-2 cells with 100nm calcium oxalate monohydrate for 24h to establish a cellular injury model. Protection was provided by RAPs with varying carboxyl group contents: 3.57%, 7.79%, 10.84%, and 15.33%. The safeguarding effect of RAPs was evaluated by analyzing relevant cellular biochemical indicators.Findings demonstrate that RAPs exhibit notable antioxidative properties. They effectively diminish the release of reactive oxygen species, lactate dehydrogenase, and malondialdehyde, a lipid oxidation byproduct. Moreover, RAPs enhance superoxide dismutase activity and mitochondrial membrane potential while attenuating the permeability of the mitochondrial permeability transition pore. Additionally, RAPs significantly reduce levels of inflammatory factors, including NLRP3, TNF-α, IL-6, and NO. This reduction corresponds to the inhibition of overproduced pro-inflammatory mediator nitric oxide and the caspase 3 enzyme, leading to a reduction in cellular apoptosis. RAPs also display the ability to suppress the expression of the HK-2 cell surface adhesion molecule CD44.The observed results collectively underscore the substantial anti-inflammatory and anti-apoptotic potential of all four RAPs. Moreover, their capacity to modulate the expression of cell surface adhesion molecules highlights their potential in inhibiting the formation of kidney stones. Notably, RAP3, boasting the highest carboxyl group content, emerges as the most potent agent in this regard.

Neutrophil extracellular traps induced by IL-1β promote endothelial dysfunction and aggravate limb ischemia.

Vascular inflammation and endothelial dysfunction contribute to vascular diseases. While neutrophil extracellular traps (NETs) participate in some vascular pathologies, their roles in lower limb ischemia remain poorly defined. This study investigated the functional significance of NETs in vascular inflammation and remodeling associated with limb ischemia. Single-cell RNA sequencing (scRNA-seq) and flow cytometry revealed neutrophil activation and upregulated NETs formation in human limb ischemia, with immunofluorescence confirming IL-1β-induced release of NETs for vascular inflammation. Endothelial cell activation was examined via scRNA-seq and western blotting, indicating enhanced proliferation, expression of adhesion molecules (VCAM-1, ICAM-1), inflammatory cytokines (IL-1β, IL-6) and decreased expression of VE-cadherin, that could be mediated by NETs to exacerbate endothelial inflammation. Mechanistically, NETs altered endothelial cell function via increased pSTAT1/STAT1 signaling. Vascular inflammation and subsequent ischemia were alleviated in vivo by NETosis or IL-1β inhibition in ischemic mice. IL-1β-NETs induce endothelial activation and inflammation in limb ischemia by stimulating STAT1 signaling. Targeting NETs may thus represent a novel therapeutic strategy for inflammatory vascular diseases associated with limb ischemia. Graphical abstract of NETs regulation of the development of vascular inflammation in lower limb ischemia via pSTAT1/STAT1 signaling pathway.

Runx3 Regulates CD8+ T Cell Local Expansion and CD43 Glycosylation in Mice by H1N1 Influenza A Virus Infection.

We have recently reported that transcription factor Runx3 is required for pulmonary generation of CD8+ cytotoxic T lymphocytes (CTLs) that play a crucial role in the clearance of influenza A virus (IAV). To understand the underlying mechanisms, we determined the effects of Runx3 knockout (KO) on CD8+ T cell local expansion and phenotypes using an inducible general Runx3 KO mouse model. We found that in contrast to the lungs, Runx3 general KO promoted enlargement of lung-draining mediastinal lymph node (mLN) and enhanced CD8+ and CD4+ T cell expansion during H1N1 IAV infection. We further found that Runx3 deficiency greatly inhibited core 2 O-glycosylation of selectin ligand CD43 on activated CD8+ T cells but minimally affected the cell surface expression of CD43, activation markers (CD44 and CD69) and cell adhesion molecules (CD11a and CD54). Runx3 KO had a minor effect on lung effector CD8+ T cell death by IAV infection. Our findings indicate that Runx3 differently regulates CD8+ T cell expansion in mLNs and lungs by H1N1 IAV infection. Runx3 is required for CD43 core 2 O-glycosylation on activated CD8+ T cells, and the involved Runx3 signal pathway may mediate CD8+ T cell phenotype for pulmonary generation of CTLs.

BjussuMP-II, a venom metalloproteinase, induces the release and cleavage of pro-inflammatory cytokines and disrupts human umbilical vein endothelial cells

Snake venom metalloproteases (SVMPs) are hydrolytic enzymes dependent on metal binding, primarily zinc (Zn2+), at their catalytic site. They are classified into three classes (P–I to P-III). BjussuMP-II, a P–I SVMP isolated from Bothrops jararacussu snake venom, has a molecular mass of 24 kDa. It exhibits inhibitory activity on platelet aggregation and hydrolyzes fibrinogen. TNF-α upregulates the expression of adhesion molecules on endothelial cell surfaces, promoting leukocyte adhesion and migration during inflammation. Literature indicates that SVMPs may cleave the TNF-α precursor, possibly due to significant homology between metalloproteases from mammalian extracellular matrix and SVMPs. This study aimed to investigate BjussuMP-II’s effects on human umbilical vein endothelial cells (HUVEC), focusing on viability, detachment, adhesion, release, and cleavage of TNF-α, IL-1β, IL-6, IL-8, and IL-10. HUVEC were incubated with BjussuMP-II (1.5–50 μg/mL) for 3–24 h. Viability was determined using LDH release, MTT metabolization, and 7AAD for membrane integrity. Adhesion and detachment were assessed by incubating cells with BjussuMP-II and staining with Giemsa. Cytokines were quantified in HUVEC supernatants using EIA. TNF-α cleavage was evaluated using supernatants from PMA-stimulated cells or recombinant TNF-α. Results demonstrated BjussuMP-II’s proteolytic activity on casein. It was not toxic to HUVEC at any concentration or duration studied but interfered with adhesion and promoted detachment. PMA induced TNF-α release by HUVEC, but this effect was not observed with BjussuMP-II, which cleaved TNF-α. Additionally, BjussuMP-II cleaved IL-1β, IL-6, and IL-10. These findings suggest that the zinc metalloprotease BjussuMP-II could be a valuable biotechnological tool for treating inflammatory disorders involving cytokine deregulation.

Role of chitosan in intestinal integrity: TLR4 and IFNAR signaling in the induction of E-cadherin and CD103 in mice

Chitin and its derivative chitosan (Q) are abundant structural elements in nature. Q has modulatory and anti-inflammatory effects and also regulates the expression of adhesion molecules. The interaction between cells expressing the αEβ7 integrin and E-cadherin facilitates tolerogenic signal transmission and localization of lymphocytes at the frontline for interaction with luminal antigens. In this study we evaluated the ability of orally administered Q to stimulate E-cadherin and CD103 expression in vitro and in vivo. Our findings show that Q promoted epithelial cell migration, accelerated wound healing and increased E-cadherin expression in IEC-18 cells and isolated intestinal epithelial cells (IECs) after Q feeding. The upregulation of E-cadherin was dependent on TLR4 and IFNAR signaling, triggering CD103 expression in lymphocytes. Q reinforced the E-cadherin-αEβ7 axis, crucial for intestinal barrier integrity and contributed to the localization of lymphocytes on the epithelium.

Endothelial PTP1B Deletion Promotes VWF Exocytosis and Venous Thromboinflammation.

Endothelial activation promotes the release of procoagulant extracellular vesicles and inflammatory mediators from specialized storage granules. Endothelial membrane exocytosis is controlled by phosphorylation. We hypothesized that the absence of PTP1B (protein tyrosine phosphatase 1B) in endothelial cells promotes venous thromboinflammation by triggering endothelial membrane fusion and exocytosis. Mice with inducible endothelial deletion of PTP1B (End.PTP1B-KO) underwent inferior vena cava ligation to induce stenosis and venous thrombosis. Primary endothelial cells from transgenic mice and human umbilical vein endothelial cells were used for mechanistic studies. Vascular ultrasound and histology showed significantly larger venous thrombi containing higher numbers of Ly6G (lymphocyte antigen 6 family member G)-positive neutrophils in mice with endothelial PTP1B deletion, and intravital microscopy confirmed the more pronounced neutrophil recruitment following inferior vena cava ligation. RT2 PCR profiler array and immunocytochemistry analysis revealed increased endothelial activation and adhesion molecule expression in primary End.PTP1B-KO endothelial cells, including CD62P (P-selectin) and VWF (von Willebrand factor). Pretreatment with the NF-κB (nuclear factor kappa B) kinase inhibitor BAY11-7082, antibodies neutralizing CD162 (P-selectin glycoprotein ligand-1) or VWF, or arginylglycylaspartic acid integrin-blocking peptides abolished the neutrophil adhesion to End.PTP1B-KO endothelial cells in vitro. Circulating levels of annexin V+ procoagulant endothelial CD62E+ (E-selectin) and neutrophil (Ly6G+) extracellular vesicles were also elevated in End.PTP1B-KO mice after inferior vena cava ligation. Higher plasma MPO (myeloperoxidase) and Cit-H3 (citrullinated histone-3) levels and neutrophil elastase activity indicated neutrophil activation and extracellular trap formation. Infusion of End.PTP1B-KO extracellular vesicles into C57BL/6J wild-type mice most prominently enhanced the recruitment of endogenous neutrophils, and this response was blunted in VWF-deficient mice or by VWF-blocking antibodies. Reduced PTP1B binding and tyrosine dephosphorylation of SNAP23 (synaptosome-associated protein 23) resulting in increased VWF exocytosis and neutrophil adhesion were identified as mechanisms, all of which could be restored by NF-κB kinase inhibition using BAY11-7082. Our findings show that endothelial PTP1B deletion promotes venous thromboinflammation by enhancing SNAP23 phosphorylation, endothelial VWF exocytosis, and neutrophil recruitment.