Search Results (120)

Peripheral edema is a pathological condition caused by abnormal fluid accumulation in the interstitial space due to elevated vascular permeability and inflammation. This study evaluated the therapeutic efficacy of Terminalia chebula fruit extract (TCE) in inflammation-induced peripheral edema and clarified its molecular mechanisms. Using hydrogen peroxide (H₂O₂)-stimulated human umbilical vein endothelial cells (HUVECs), TCE was tested for effects on cell viability, inflammatory gene expression, intracellular reactive oxygen species, endothelial barrier integrity, and vascular endothelial growth factor (VEGF)-induced migration. Its influence on nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and mitogen-activated protein kinase (MAPK) signaling was examined. In vivo, TCE was assessed in acetic acid-induced peritoneal vascular permeability and carrageenan-induced paw edema models, followed by histological analysis and serum tumor necrosis factor-α (TNF-α) measurement. TCE restored cell viability (76.2% to 94.8%), reduced TNF, IL6, and PTGS2 mRNA expression, and decreased reactive oxygen species by 27.2%. It enhanced barrier integrity, increased transendothelial electrical resistance, and inhibited VEGF-induced migration. TCE suppressed NF-κB and MAPK activation. In vivo, TCE reduced Evans blue extravasation by 41.6% and paw edema by 67.5%. Histology showed reduced dermal thickening and inflammatory infiltration, and serum TNF-α levels were lowered. TCE attenuates peripheral edema by preserving endothelial barrier function and suppressing inflammatory signaling, supporting its potential as a therapeutic agent for inflammation-associated vascular dysfunction and edema. Full article

Background: Type 2 diabetes mellitus (DM) is a major cardiovascular risk factor that induces monocyte dysfunction and contributes to their accumulation in atherosclerotic lesions. Monocyte recruitment and accumulation in the tissues contribute to chronic inflammation and are essential to the pathobiology of diabetes-induced atherosclerosis. However, the mechanisms that drive the accumulation of monocytes in the diabetic environment are not clearly understood. Methods: Primary monocytes from type 2 (T2) DM and non-T2DM individuals were isolated using magnet-assisted cell sorting. To examine the influence of a diabetic milieu on monocyte function, monocytes from T2DM patients, db/db mice, or human monocytes subjected to hyperglycaemia were analysed for their responses to pro-atherogenic cytokines using Boyden chamber assays. Furthermore, the interactions of non-diabetic and diabetic monocytes with TNFα-inflamed endothelium were studied using live-cell imaging under physiological flow conditions. RT-qPCR and FACS were used to study the expression of relevant molecules involved in monocyte-endothelium interaction. Results: CD14⁺CD16⁻ monocytes isolated from T2DM patients or monocytes exposed to hyperglycaemic conditions showed reduced chemotactic responses towards atherosclerosis-promoting cytokines, CCL2 and CX3CL1, indicating monocyte dysfunction. Under flow conditions, the transendothelial migration (TEM) capacity of T2DM monocytes was significantly reduced. Even though these monocytes adhered to the endothelial monolayer, only a few transmigrated. Interestingly, the T2DM monocytes and monocytes exposed to hyperglycaemic conditions accumulated in the ablumen following transendothelial migration. The time period in the ablumen of T2DM cells was prolonged, as there was a significant impairment of the reverse transendothelial migration (rTEM). Mechanistically, the T2DM milieu specifically induced the activation of monocyte integrins, Macrophage-1 antigen (Mac-1; integrin αMβ2 consisting of CD11b and CD18), and Lymphocyte function-associated antigen 1 (LFA-1; αLβ2 consisting of CD11a and CD18). Furthermore, elevated levels of CD18 transcripts were detected in T2DM monocytes. Junctional Adhesion Molecule 3 (JAM-3)–MAC-1 interactions are known to impede rTEM and T2DM milieu-potentiated JAM-3 expression in human coronary artery endothelial cells (HCAEC). Finally, the overexpression of JAM-3 on HCAEC was sufficient to completely recapitulate the impaired rTEM phenotype. Conclusions: Our results revealed for the first time that the enhanced T2DM monocyte accumulation in the ablumen is not secondary to the elevated transmigration through the endothelium. Instead, the accumulation of monocytes is due to the direct consequence of a dysfunctional rTEM, potentially due to enhanced JAM3-MAC1 engagement. Our results highlight the importance of restoring the rTEM capacity of monocytes to reduce monocyte accumulation-dependent inflammation induction and atherogenesis in the T2DM environment. Full article

Pancreatic ductal adenocarcinoma (PDA) exhibits diverse molecular aberrancies that contribute to its aggressive behaviour and poor patient survival. P-21-activated kinase 1 (PAK1) and PAK4 drive the tumorigenesis of PDA. However, their roles in tumour vasculature and the impact on immune response are unclear. This study aims to investigate the effects of PAK1 and PAK4 on tumour vasculature, immune cell infiltration, and the connection between using PAK1-knockout (KO), PAK4 KO, and wild-type (WT) PDA cells in cell-based and mouse experiments. Tumour tissues isolated from a syngeneic mouse model were immuno-stained to determine the changes in tumour vasculature and immune cell infiltration/activation, followed by a proteomic study to assess biological processes involved. PAK1KO or PAK4KO suppressed tumour growth by reducing angiogenesis while enhancing vascular normalisation, enhanced the infiltration/activation of T-cells and dendritic cells associated with upregulation of ICAM-1 and VCAM-1 in the tumour microenvironment, and stimulated vascular immune crosstalk via an ICAM-1-mediated mechanism. This was supported by proteomic profiles indicating the regulation of endothelial cell and leukocyte trans-endothelial migration in PAK1- or PAK4-knockout tumours. In conclusion, PAK1KO or PAK4KO enhanced tumour vascular normalisation while reducing angiogenesis, stimulating immune cell infiltration and activation to suppress tumour growth. Full article

Background/Objectives: Methamphetamine (METH), a potent psychostimulant, exerts harmful effects on the vascular system by promoting oxidative stress, inflammation, and endothelial injury. While its impact on the blood–brain barrier is well documented, its influence on the retinal microvasculature remains less understood. This study investigated the effects of METH on syndecan-1 expression and endothelial function in primary rat retinal microvascular endothelial cells (RRMECs) and isolated ophthalmic arteries. Methods: We assessed METH-induced changes in mRNA and protein expression levels of syndecan-1, matrix metalloproteinase (MMP)-2, and MMP-9. Endothelial function was evaluated using scratch migration assays and trans-endothelial electrical resistance (TEER) measurements. The mechanistic involvement of MMP-9 and trace amine-associated receptor 1 (TAAR-1), a known receptor for METH, was examined using selective pharmacological inhibitors. Results: METH exposure significantly decreased syndecan-1 expression and increased MMP-9 levels. These changes were accompanied by impaired endothelial migration and reduced TEER in RRMECs. Similar findings were confirmed in cultured ophthalmic arteries, reinforcing the translational relevance of our in vitro results. Inhibition of MMPs restored syndecan-1 expression and rescued endothelial function. Furthermore, TAAR-1 antagonism protected against syndecan-1 degradation, reduced MMP-9 upregulation, and improved endothelial migration and barrier resistance. Conclusions: Our findings suggest that METH induces loss of syndecan-1 and retinal vascular integrity by promoting TAAR-1–mediated MMP-9 upregulation. Targeting the TAAR-1/MMP-9 axis may offer a promising therapeutic strategy for preventing METH-induced microvascular damage in the retina. Full article

UDP-Gal-β-1,4 galactosyltransferase-V (GalT-V) is a member of a large family of galactosyltransferases whose function is to transfer galactose from the nucleotide sugar UDP-galactose to a glycosphingolipid glucosylceramide, to generate lactosylceramide (LacCer). It also causes the N and O glycosylation of proteins in the Trans Golgi area. LacCer is a bioactive lipid second messenger that activates an “oxidative stress pathway”, leading to critical phenotypes, e.g., cell proliferation, migration angiogenesis, autophagy, and apoptosis. It also activates an “inflammatory pathway” that contributes to the progression of disease pathology. β-1,4-GalT-V gene expression is regulated by the binding of the transcription factor Sp-1, one of the most O-GlcNAcylated nuclear factors. This review elaborates the role of the Sp-1/GalT-V axis in disease phenotypes and therapeutic approaches targeting not only Sp-1 but also Notch-1, Wnt-1 frizzled, hedgehog, and β-catenin. Recent evidence suggests that β-1,4GalT-V may glycosylate Notch-1 and, thus, regulate a VEGF-independent angiogenic pathway, promoting glioma-like stem cell differentiation into endothelial cells, thus contributing to angiogenesis. These findings have significant implications for cancer and cardiovascular disease, as tumor vascularization often resumes aggressively following anti-VEGF therapy. Moreover, LacCer can induce angiogenesis independent of VEGF and its level are reported to be high in tumor tissues. Thus, targeting both VEGF-dependent and VEGF-independent pathways may offer novel therapeutic strategies. This review also presents an up-to-date therapeutic approach targeting the β-1,4-GalT-V interactome. In summary, the β-1,4-GalT-V interactome orchestrates a broad network of signaling pathways essential for maintaining cellular homeostasis. Conversely, its dysregulation can promote unchecked proliferation, angiogenesis, and inflammation, contributing to the initiation and progression of multiple diseases. Environmental factors and smoking can influence β-1,4-GalT-V expression and its interactome, whereas elevated β-1,4-GalT-V expression may serve as a diagnostic biomarker of colorectal cancer, inflammation—exacerbated by factors that may worsen pre-existing cancer malignancies, such as smoking and a Western diet—and atherosclerosis, amplifying disease progression. Increased β-1,4-GalT-V expression is frequently associated with tumor aggressiveness and chronic inflammation, underscoring its potential as both a biomarker and therapeutic target in colorectal and other β-1,4-GalT-V-driven cancers, as well as in cardiovascular and inflammatory diseases. Full article

Background. Elevated lipoprotein(a) [Lp(a)] levels are a key factor in the early formation and progression of atherosclerosis. Monocytes in individuals with an elevated Lp(a) level are represented by an activated inflammatory phenotype and have an increased ability for transendothelial migration. This work studies the association between Lp(a), monocytes, and the progression of carotid atherosclerosis in patients with premature coronary heart disease (CHD). Methods. This study included 102 patients with CHD manifested before 55 in men and 60 in women who underwent two carotid duplex scans with an interval of 5 [3; 8] years. The criteria for the progression of carotid atherosclerosis were the appearance of new plaque and an increase in stenosis by >10% in any of the six segments. The lipid profile, Lp(a), and hematology with the calculation of the lymphocyte–monocyte ratio (LMR) were determined in all the patients. Results. The median blood monocyte count was 0.54 × 10⁹/L, and the median LMR was 4.18. In 70 patients, we revealed the criteria for carotid atherosclerosis progression. The groups did not differ by demographics, risk factors, or the blood lipid and lipoprotein levels, except for Lp(a); this concentration was higher in the patients with carotid atherosclerosis progression. The odds of atherosclerosis progression were highest in the patients with an elevated Lp(a) level and a blood monocyte count above the median (16.8, 3.4–83.0, p < 0.001). Carotid atherosclerosis progression was associated with LMR < 4.18 and an elevated Lp(a) level (OR = 4.3, 1.1–17.2, p = 0.04) and not associated with the patients with Lp(a) levels < 30 mg/dL and an LMR above the median. Conclusions. An elevated Lp(a) level and monocyte count provide the highest probability of the progression of carotid atherosclerosis in patients with premature CHD. Full article

Background: Tankyrase (TNKS1) regulates the WNT/β-catenin pathway, while CDK8 is a transcriptional regulator overexpressed in renal cell carcinoma (RCC). This study aims to identify novel dual inhibitors of tankyrase and Cyclin-dependent kinase 8 (CDK8), utilizing bioinformatics and in vitro methods and to assess their efficiency in renal cancer cells. Methods: To identify leads, the ChemBridge library was screening using high-throughput virtual screening (HTVS), which was followed by protein–ligand interaction analysis, Molecular Dynamics (MD) simulation, and Gibbs binding free energy estimation. A-498, Caki-1, and HK-2 cells were employed to validate in vitro efficacy. Results: TCS9725 was discovered by HTVS with binding affinities of −8.1 kcal/mol and −8.2 kcal/mol for TNKS1 and CDK8, respectively. TCS9725 had robust binding interactions with root mean square deviation values of 0.00 nm. The ΔG binding estimate was −27.45 for TNKS1 and −27.88 for CDK8, respectively. ADME predictions favored specific small-molecule inhibition profiles. TCS9725 reduced TNKS1 and CDK8 activities with IC50s of 243 nM and 403.6 nM, respectively. The compound efficiently inhibited the growth of A-498 and Caki-1 cells with GI50 values of 385.9 nM and 243.6 nM, respectively, with high selectivity compared to the non-cancerous kidney cells. TCS9725 decreased STAT1 and β-catenin positivity in A-498 and Caki-1 cells. The compound induced apoptosis and reduced TGFβ-stimulated trans-endothelial migration and p-smad2/3 signaling in both RCC cells. Conclusions: This work provides valuable insights into the therapeutic potential of TCS9725, a dual inhibitor of TNKS1 and CDK8. Further developments of this molecule could lead to new and effective treatments for this devastating disease. Full article

Tick-borne encephalitis remains a significant burden on human health in the endemic areas in Central Europe and Eastern Asia. The causative agent, tick-borne encephalitis virus (TBEV), is a neurotropic virus belonging to the genus of Orthoflavivirus. After TBEV enters the central nervous system (CNS), it mainly targets neurons, causing encephalitis and leading to life-long disabilities, coma and, in rare cases, death. The neuroinvasive mechanisms of orthoflaviviruses are poorly understood. Here we investigate the mechanism of TBEV neuroinvasion, hypothesizing that TBEV influences blood–brain barrier (BBB) properties and uses transcellular routes to cross the endothelial barrier and enter the CNS. To test this hypothesis, we employed an in vitro transwell system consisting of endothelial cell monolayers cultured on insert membranes and studied the barrier properties following inoculation to tick-borne orthoflaviviruses. It was shown that TBEV and closely related but naturally attenuated Langat virus (LGTV) crossed the intact endothelial cell monolayer without altering its barrier properties. Interestingly, transendothelial migration of TBEV was significantly affected when two cellular surface antigens, the laminin-binding protein and vimentin, were blocked with specific antibodies. Taken together, these results indicate that orthoflaviviruses use non-destructive transcellular routes through endothelial cells to establish infection within the CNS. Full article

We previously observed elevated plasma levels of citrulline and arginine in diabetic retinopathy patients compared to diabetic controls. We tested our hypothesis that citrulline plus arginine induces angiogenesis and increases permeability in retinal endothelial cells. Human retinal microvascular endothelial cells (HRMECs) were treated with citrulline, arginine, or citrulline + arginine, and angiogenesis was measured with cell proliferation, migration, and tube formation assays. Permeability was measured in HRMEC monolayers via trans-endothelial electrical resistance (TEER) and FITC-labeled dextran. We also measured arginase activity, arginase-1 and arginase-2 expression, protein expression and phosphorylation of endothelial nitric oxide synthase (eNOS), and nitric oxide (NO) production. Citrulline + arginine induced endothelial cell proliferation (p = 0.018), migration (p = 0.011), and tube formation (p = 0.0042). Citrulline + arginine also increased FITC-dextran flow-through (p = 1.5 × 10⁻⁵) and decreased TEER (p = 0.010). Citrulline + arginine had no effect on arginase activity, but it increased eNOS (p = 6.3 × 10⁻⁴) and phosphorylated eNOS (p = 0.029), as well as NO production (p = 0.025). Inhibiting eNOS prevented the increase in NO (p = 0.0092), inhibited citrulline + arginine-induced cell migration (p = 0.0080) and tube formation (p = 0.0092), and blocked citrulline + arginine-related alterations in FITC-dextran flow-through (p = 3.6 × 10⁻⁴) and TEER (p = 3.9 × 10⁻⁴). These data suggest that citrulline + arginine treatment induces angiogenesis and increases permeability in retinal endothelial cells by activating eNOS and increasing NO production. Full article

Red swamp crayfish (Procambarus clarkii) is one of the most important aquaculture species in China. Frequent outbreaks of diseases seriously threatened the sustainable development of the industry. It is necessary to understand the causes of disease and study the mechanism of disease resistance in P. clarkii. In this paper, the pathogenic bacteria causing head ulcers in juvenile P. clarkii were found and identified as Citrobacter freundii, which can cause severe pathological changes in the hepatopancreas and intestines of juvenile P. clarkii. Detection of humoral immune factors revealed that PO activity and lysozyme activity of infected P. clarkii were significantly enhanced at 15 and 20 dpi, respectively. Transcriptome analysis was conducted of hepatopancreas from normal and diseased P. clarkii after C. freundii injection, as well as bacteria-free control of P. clarkii. It was found that DEGs are rich in NF-κB, oxidative phosphorylation, JAK/STAT, Leukocyte transendothelial migration, MAPK, and PPAR signaling pathway. These pathways are related to immune modulation, metabolism, and pathogen clearance. Meanwhile, immune-related genes such as Gip, nfyA, psmD13, and FGFR were significantly highly expressed in the normal group, which was verified by qRT-PCR results, suggesting that they may be the key regulatory genes for juvenile P. clarkii resistance to C. freundii. This study will help to elucidate the molecular mechanism of the immune response of P. clarkii to C. freundii. The results are instructive for the prevention and treatment of P. clarkii diseases and for further understanding of the invertebrate immune system. Full article

Polychlorinated biphenyls (PCBs) are heterogeneous, synthetic, and widespread organochlorine compounds, and are one of the persistent organic pollutants present in improperly dumped waste and electronic equipment (e-waste), with a high bioaccumulation potential. In this study, the toxicity of Aroclor 1254 (a mixture of commercial PCBs) in human corneal epithelial cells (HCEpiCs), in an in vitro model of an ocular barrier, was evaluated. Aroclor 1254 (0.1–10 μg/mL) reduced cell viability, trans-endothelial electric resistance (TEER) and cell migration. Moreover, it induced an inflammatory response, as indicated by the increase in cPLA₂ activity, PGE2 production, phosphorylation of ERK 1/2 and p-38, and release of inflammatory cytokines. Aroclor 1254 can damage corneal cells, compromising the integrity of the eye’s outermost barrier. This damage may facilitate the occurrence of infectious processes that are physiologically prevented by the corneal barrier. Full article

Objectives: Reduced expression of adhesion molecules in tumor vasculature can limit infiltration of effector T cells. To improve T cell adhesion to tumor endothelial cell (EC) antigens and enhance transendothelial migration, we developed bispecific, T-cell engaging antibodies (bsAb) that activate T cells after cross-linking with EC cell surface antigens. Methods: Recombinant T-cell stimulatory anti-VEGFR2–anti-CD3 and costimulatory anti-TIE2–anti-CD28 or anti-PD-L1–anti-CD28 bsAb were engineered and expressed. Primary lines of human umbilical vein endothelial cells (HUVEC) that constitutively express VEGFR2 and TIE2 growth factor receptors and PD-L1, but very low levels of adhesion molecules, served as models for anergic tumor EC. Results: In cocultures with HUVEC, anti-VEGFR2–anti-CD3 bsAb increased T cell binding and elicited rapid T cell activation. The release of proinflammatory cytokines TNF-α, IFN-γ, and IL-6 was greatly augmented by the addition of anti-TIE2–anti-CD28 or anti-PD-L1–anti-CD28 costimulatory bsAb. Concomitantly, T cell-released cytokines upregulated E-selectin, ICAM1, and VCAM1 adhesion molecules on HUVEC. HUVEC cultured in breast cancer cell-conditioned medium to mimic the influence of tumor-secreted factors were similarly activated by T cell-engaging bsAb. Migration of T cells in transwell assays was significantly increased by anti-VEGFR2–anti-CD3 bsAb. The combination with costimulatory anti-TIE2–anti-CD28 bsAb augmented activation and proliferation of migrated T cells and their cytotoxic capacity against spheroids of the MCF-7 breast cancer cell line seeded in the lower transwell chamber. Conclusions: T cells activated by anti-VEGFR2–anti-CD3 and costimulatory EC-targeting bsAb can reverse the energy of quiescent EC in vitro, resulting in improved T cell migration through an EC layer. Full article

Background: Chronic chagasic cardiomyopathy is the most severe clinical manifestation of Chagas disease, which affects approximately seven million people worldwide. Latin American countries bear the highest burden, with the greatest morbidity and mortality rates. Currently, diagnostic methods do not provide information on the risk of progression to severe stages of the disease. Recently, microRNAs (miRNAs) have been proposed as promising tools for monitoring the progression of Chagas disease. This study aimed to analyze the expression profiles of the miRNAs miR-1, miR-16, miR-208, and miR-208b in cardiac tissue, plasma, and plasma extracellular vesicles from Ninoa TcI-infected mice during the acute and indeterminate phases of Chagas disease. Methods: The cardiac-specific miRNAs and miR-16 levels were examined in all samples using RT-qPCR. Additionally, pathway analysis was performed to investigate the impact of potential miRNA target genes across various databases. Results: Elevated miR-208b expression was observed in cardiac tissue and plasma during the acute phase. Bioinformatic analysis identified three pathways implicated in disease progression: phosphatidylinositol 3-kinase signaling, Fc gamma receptor-mediated phagocytosis, and leukocyte transendothelial migration, as well as cholinergic synapse pathways. Conclusions: MiR-208b was upregulated during the acute phase and downregulated in the indeterminate phase, suggesting it may play a crucial role in disease progression. Full article

Moringa leaves provide numerous health benefits due to their anti-inflammatory properties. This study presents the first evidence that endothelial cell inflammation can potentially be ameliorated by moringa leaf extract. Here, we established an experimental human blood vessel cell model of inflammation using EA.hy926 cells. TNF-α was added after pre-treating the cells with crude leaf extract from Moringa oleifera Lam., a constituent fraction of the extract, and the bioactive component 3-hydroxy-β-ionone. The extract and the active ingredient significantly decreased the levels of pro-inflammatory mediators such as IL-6, IL-8, and MCP-1; decreased IκB-α and NF-κB p65 phosphorylation; and decreased the expression of VCAM-1, PECAM-1, and ICAM-1, three significant adhesion molecules. Furthermore, they attenuated THP-1 monocyte adhesion to the EA.hy926 monolayer and decreased monocyte transmigration across the monolayer. These findings suggest that 3-hydroxy-β-ionone and moringa leaf extract have anti-inflammatory properties and can be used as therapeutic agents to reduce the progression of diseases involving the inflamed endothelium by decreasing the production of inflammatory cytokines, chemokines, and adhesion molecules. This is promising for conditions such as atherosclerosis and neuroinflammation. Full article

Heart failure is a complex syndrome characterized by cardiac hypertrophy, fibrosis, and diastolic/systolic dysfunction. These changes share many pathological features with significant inflammatory responses in the myocardium. Among the various regulatory systems that impact on these heterogeneous pathological processes, angiotensin II (Ang II)-activated macrophages play a pivotal role in the induction of subcellular defects and cardiac adverse remodeling during the progression of heart failure. Ang II stimulates macrophages via its AT1 receptor to release oxygen-free radicals, cytokines, chemokines, and other inflammatory mediators in the myocardium, and upregulates the expression of integrin adhesion molecules on both monocytes and endothelial cells, leading to monocyte-endothelial cell-cell interactions. The transendothelial migration of monocyte-derived macrophages exerts significant biological effects on the proliferation of fibroblasts, deposition of extracellular matrix proteins, induction of perivascular/interstitial fibrosis, and development of hypertension, cardiac hypertrophy and heart failure. Inhibition of macrophage activation using Ang II AT1 receptor antagonist or depletion of macrophages from the peripheral circulation has shown significant inhibitory effects on Ang II-induced vascular and myocardial injury. The purpose of this review is to discuss the current understanding in Ang II-induced maladaptive cardiac remodeling and dysfunction, particularly focusing on molecular signaling pathways involved in macrophages-mediated hypertension, cardiac hypertrophy, fibrosis, and failure. In addition, the challenges remained in translating these findings to the treatment of heart failure patients are also addressed. Full article