Search Results (823)

Background: Diabetes mellitus is characterized by chronic hyperglycemia, which promotes oxidative stress and endothelial dysfunction. MicroRNAs (miRNAs) have emerged as important regulators of diabetic vascular complications. Our previous RNA-sequencing study identified putative novel miR-1133 as an upregulated miRNA in hyperglycemia-induced human umbilical vein endothelial cells (HUVECs). This study aimed to investigate the potential involvement of putative novel miR-1133 in oxidative stress in hyperglycemia-induced HUVECs. Methods: Functional enrichment and protein–protein interaction (PPI) network analyses were performed to identify biologically relevant predicted target genes of putative novel miR-1133. Phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) was selected for further investigation because it was identified as a hub gene in the PPI network and was involved in the enriched PI3K/Akt signaling pathway. HUVECs were exposed to high glucose (33.3 mM) to establish an in vitro hyperglycemic model, and the effects of transfection with putative novel miR-1133 inhibitor on PIK3CA expression and oxidative stress markers were evaluated. Results: Bioinformatic analyses identified PIK3CA as a hub gene among the predicted targets of putative novel miR-1133 and identified the PI3K/Akt signaling pathway as significantly enriched. Experimentally, hyperglycemia significantly reduced PIK3CA expression, whereas putative novel miR-1133 inhibitor transfection increased PIK3CA expression. Furthermore, putative novel miR-1133 inhibitor transfection was associated with reduced reactive oxygen species and 8-hydroxy-2′-deoxyguanosine levels, together with increased superoxide dismutase 1 (SOD1) mRNA expression and total SOD activity. Conclusions: Transfection with putative novel miR-1133 inhibitor was associated with increased PIK3CA expression and reduced oxidative stress markers in hyperglycemia-induced endothelial cells. Together, bioinformatic and experimental findings support an association between putative novel miR-1133 inhibitor transfection, PIK3CA expression, and oxidative stress under hyperglycemic conditions. Further studies are required to validate the direct interaction between putative novel miR-1133 and PIK3CA and to determine the relevance of these findings in vivo. Full article

Agarose is a promising biopolymer for wound healing due to its biocompatibility and ability to form stable macroporous cryogels. However, its bioinert nature limits its antibacterial functionality, while conventional chemical modification can disrupt its macroporous architecture and mechanical integrity. Here, we present a versatile post-synthetic heterogeneous surface quaternization strategy to transform prefabricated agarose cryogels into surface-modified agarose cryogels (SMACs) with contact-active antibacterial functionality while preserving their interconnected supermacroporous architecture. Using 3-chloro-2-hydroxypropyltrimethylammonium chloride (CHPTAC) in an alkaline environment, quaternary ammonium moieties were covalently immobilized onto the agarose surface. A multivariable optimization approach examining reaction time, temperature, and scaffold concentration yielded surface quaternization values (DS_EA) of 0.05 and 0.11 for SMAC-4 and SMAC-24, respectively, with successful modification confirmed via SEM-EDX and FT-IR analyses. Quaternization altered the physicochemical and mechanical properties of the scaffolds; notably, the average porosity increased from 66.7% in AC-0 to 78.7% in SMAC-24 while maintaining the integrity of the porous structure. Biological evaluation against Escherichia coli and Staphylococcus aureus identified SMAC-24 as the most favorable formulation, exhibiting the strongest antibacterial activity and maintaining 88.1 ± 1.6% human umbilical vein endothelial cell (HUVEC) viability after 24 h of exposure to scaffold extracts. These findings demonstrate that post-synthetic surface quaternization provides agarose cryogels with contact-active antibacterial functionality and favorable cytocompatibility, highlighting their potential as non-leaching antibacterial agarose scaffolds for chronic wound care. Full article

Micro- and nanoplastics (MNPs) are increasingly detected in human tissues, yet their polymer-specific effects on endothelial cells remain poorly understood, particularly at the placental and fetal level. We compared cadmium selenide quantum dot-labelled polypropylene (PP) and polyvinyl chloride (PVC) model nanoplastics (NPs) in human umbilical vein endothelial cells (HUVECs) using particle characterization, MTT assays, flow cytometry, confocal microscopy, apoptosis analysis, and synchrotron nano-X-ray fluorescence imaging. PP nanoplastics caused an early reduction in metabolic activity, showed the strongest cell-associated fluorescence, and produced the greatest increase in membrane permeability and late apoptotic/necrotic populations. PVC nanoplastics displayed a more punctate distribution with greater overlap with membrane-associated regions and induced a stronger increase in LC3B-positive vesicular structures. Nano-XRF detected Cd-enriched signals associated with the labelled particles and a polymer-specific Cd–Cl spatial association in PVC-exposed cells. Sulfur mapping further revealed localized sulfur-poor regions along the cell periphery in exposed cells, suggesting localized remodeling of peripheral membranes. These findings indicate that PP and PVC NPs interact differently with endothelial cells and elicit distinct structural and functional responses. Polymer composition should therefore be considered when assessing the vascular and prenatal effects of MNP exposure. Full article

Diabetes mellitus induces endothelial dysfunction and oxidative stress, contributing to the development of vascular complications. Although metformin exerts well-established vasculoprotective and antioxidant effects, the molecular mechanisms underlying these actions remain incompletely understood. Novel miR-1133 was previously identified as a metformin-responsive putative microRNA (miRNA), and bioinformatic analysis identified PIK3CA as a biologically plausible candidate target for further investigation. This study investigated the functional significance of novel miR-1133 in the antioxidant effects of metformin in hyperglycemia-induced human umbilical vein endothelial cells (HUVECs). HUVECs were exposed to normal glucose (5.5 mmol/L), hyperglycemia (33.3 mmol/L), or hyperglycemia supplemented with metformin (10 μM) for 24 h. Metformin-treated hyperglycemic HUVECs were subsequently transfected with a novel miR-1133 mimic during continued hyperglycemic and metformin exposure. PIK3CA expression and oxidative stress markers, including reactive oxygen species (ROS), superoxide dismutase (SOD), and 8-hydroxy-2′-deoxyguanosine (8-OHdG), were evaluated. Hyperglycemia increased novel miR-1133 expression, whereas metformin reduced its expression. Metformin treatment was associated with increased PIK3CA mRNA and protein expression, reduced ROS and 8-OHdG levels, and increased SOD expression and activity. Novel miR-1133 mimic transfection was associated with reduced PIK3CA expression, increased ROS and 8-OHdG levels, and decreased SOD expression and activity compared with the corresponding transfection control. Collectively, these findings demonstrate that metformin attenuates oxidative stress in hyperglycemia-induced endothelial cells and overall provide the first functional characterization of novel miR-1133. The reciprocal changes in PIK3CA expression and oxidative stress observed following novel miR-1133 mimic transfection suggest that novel miR-1133 may represent a candidate mediator of the endothelial antioxidant responses associated with metformin. Further studies are required to validate the direct interaction between novel miR-1133 and PIK3CA and to elucidate the underlying molecular mechanisms. Full article

Background: Thin endometrium is a significant cause of infertility due to impaired regeneration, reduced receptivity, and insufficient angiogenesis. Mesenchymal stem cell-derived secretome (MSCsec) is a promising cell-free therapy because it contains bioactive molecules that promote tissue repair. This study evaluated the effects of Wharton’s jelly-derived MSC secretome (WJ-MSCsec) on human endometrial stromal cells (EnSCs) and endothelial cells in vitro. Methods: WJ-MSCs were isolated from umbilical cord tissue, characterized, and cultured under serum-free conditions. The concentrated secretome was analyzed using a human angiogenesis proteome array. EnSCs were isolated from endometrial biopsies. EnSC proliferation and migration in the presence of WJ-MSCsec were assessed using CCK-8 and scratch wound-healing assays. Pro-angiogenic activity of WJ-MSCsec was evaluated using a Matrigel tube formation assay with human umbilical vein endothelial cells (HUVECs). Results: Proteomic analysis of WJ-MSCsec revealed the presence of angiogenic, mitogenic, immunomodulatory, and chemotactic factors. Treatment with WJ-MSCsec significantly enhanced EnSC proliferation and accelerated wound closure. Furthermore, WJ-MSCsec markedly promoted endothelial tube formation, increasing total tube length, junction number, and mesh formation. Conclusions: WJ-MSCsec stimulates EnSC proliferation and migration while enhancing angiogenesis in vitro. WJ-MSCsec represents a promising cell-free therapeutic strategy for endometrial regeneration and reproductive disorders associated with impaired endometrial function. Full article

Accumulation of senescent endothelial cells (ECs) accelerates atherosclerosis, yet the molecular orchestrators linking metabolic dysregulation to epigenetic reprogramming during EC senescence remain elusive. Here, we investigated the specific role of Myeloid cell leukemia 1 (MCL1) in endothelial senescence and atherogenesis. Through integrative bioinformatics, MCL1 was identified as a candidate senescence regulator. MCL1 expression was validated in human carotid atherosclerotic plaques, ApoE^−/− mice, and multiple EC senescence models, primarily HRAS^G12V-induced senescent human umbilical vein endothelial cells (HUVECs). Loss- and gain-of-function assays were performed in HUVECs. Underlying metabolic and epigenetic mechanisms were explored using Seahorse extracellular flux analysis, lactate measurements, Co-IP and CUT&Tag sequencing. In vivo therapeutic potential was evaluated via AAV-mediated MCL1 knockdown in high-fat diet-fed ApoE^−/− mice. MCL1 expression was significantly upregulated in atherosclerotic plaques and senescent ECs. Functionally, MCL1 knockdown attenuated senescence markers (SA-β-gal, P21), suppressed the senescence-associated secretory phenotype (SASP), and restored cell proliferation, whereas MCL1 overexpression exacerbated EC senescence. Mechanistically, MCL1 promoted glycolytic reprogramming and intracellular lactate accumulation. This metabolic byproduct served as an epigenetic substrate for histone H4K12 lactylation (H4K12la), which specifically enriched at the CDKN1A (P21) promoter, correlating with its transcription. In vivo, AAV-mediated MCL1 silencing effectively reduced vascular H4K12la levels, alleviated vascular senescence, and substantially constrained atherosclerotic lesion areas. Our findings identify MCL1 as a pivotal regulator of endothelial senescence and atherosclerosis, operating through a metabolic-\–epigenetic mechanism involving glycolytic reprogramming, lactate accumulation, and H4K12la-associated P21 upregulation. These findings suggest that targeting the MCL1-associated pathway may represent a potential therapeutic strategy for atherosclerosis. Full article

Mucuna pruriens (Linn.) DC. var. pruriens is used as a traditional medication to enhance male sexual function, yet integrated evidence linking vascular-cell mechanisms to in vivo efficacy and safety of a standardized extract remains limited. This study evaluated the ethanolic seed extract of M. pruriens var. pruriens (MPSE) on erectile function-related mechanisms, addressing the nitric oxide (NO)–cyclic guanosine monophosphate (cGMP)–phosphodiesterase type 5 (PDE5) pathway in vitro, together with mounting frequency, anxiety-like behavior, locomotor activity, and cardiovascular safety in vivo. Soxhlet extraction with 80% ethanol yielded a dark brown, syrupy extract (12.28% w/w), which was standardized by high-performance thin-layer chromatography to contain 278 mg/g extract (27.8% w/w) of L-DOPA (Rf 0.38). Mechanistic effects were assessed in human umbilical vein endothelial cells (HUVECs), pulmonary artery smooth muscle cells (PASMCs), and a PDE5A1 enzyme assay. Male Wistar rats received MPSE (50, 100, and 200 mg/kg) orally for 15 days, with hemodynamic assessment in normotensive and Nω-nitro-L-arginine methyl ester (L-NAME)-induced hypertensive rats. MPSE significantly increased eNOS expression and NO production in HUVECs, elevated intracellular cGMP in PASMCs, and directly inhibited PDE5A1 enzymatic activity. Repeated administration significantly increased mounting frequency relative to baseline without altering serum testosterone levels, anxiety-like behavior, locomotor activity, mean arterial pressure, or heart rate. These findings indicate that MPSE modulates erectile function-related mechanisms, primarily through peripheral vascular effects on the NO–cGMP–PDE5 pathway, and suggest a favorable safety profile. Erectile function itself was not directly assessed and warrants confirmation in models of erectile response. MPSE nevertheless represents a promising plant-derived candidate for erectile dysfunction. Full article

Impaired endothelial function, excessive oxidative stress, and persistent bacterial infection collectively contribute to delayed healing of diabetic chronic wounds. Methylglyoxal (MGO)-induced metabolic stress is a critical driver of endothelial injury; however, effective multifunctional strategies capable of restoring vascular function and maintaining cellular homeostasis remain limited. In this study, the endothelial protective potential of an amphibian-derived Bowman–Birk inhibitor (BBI)-type peptide, OSTI-1872, and its rationally designed structural analogues were investigated using an MGO-induced injury model in human umbilical vein endothelial cells (HUVECs). Among the tested peptides, the optimised analogue OSTI-2337 exhibited superior protective activity. OSTI-2337 markedly attenuated MGO-induced oxidative stress, restored nitric oxide bioavailability, enhanced VEGF expression, promoted endothelial migration and tube formation, and reduced oxidative DNA damage. Mechanistically, these effects were associated with coordinated regulation of MGO detoxification and redox homeostasis, as evidenced by enhanced GLO1 expression and modulation of the PI3K/AKT/GSK3β/Nrf2 axis, accompanied by increased expression of downstream antioxidant proteins HO-1 and NQO1. In addition, OSTI-1872 and OSTI-2337 displayed antibacterial activity against representative bacterial strains, suggesting their potential advantages for complex diabetic wound environments. Collectively, these findings demonstrate that structural optimisation significantly enhances the biological activity of amphibian BBI-type peptides and identify OSTI-2337 as a multifunctional peptide candidate capable of integrating endothelial protection, MGO detoxification, redox regulation, and antibacterial activity for the management of diabetes-associated vascular injury and chronic wound complications. Full article

Stem cell-derived extracellular vesicles (EVs) hold therapeutic potential for hypoxia-associated injury, yet the molecular mechanisms underlying their protective effects remain incompletely defined. Herein, we performed comparative proteomic profiling of extracellular vesicles (EVs) secreted by human placenta-derived perivascular stem cells (hPPSCs) and human umbilical cord mesenchymal stem cells (hUCMSCs). We found that hPPSCs-EVs are enriched in proteasome-related proteins. Enzymatic activity assays further confirmed that proteasome activity in hPPSCs-EVs was significantly higher than that in hUCMSCs-EVs. Additionally, we observed that hPPSCs-EVs were efficiently endocytosed by human umbilical vein endothelial cells (hUVECs), leading to marked downregulation of hypoxia-inducible factor 1-alpha (HIF-1α) and intracellular ubiquitinated proteins. Importantly, this HIF-1α degradation persisted even when the host ubiquitin–proteasome system was blocked, indicating that hPPSCs-EVs can function independently of the host proteasomal pathway. These results reveal that hPPSCs-EVs deliver functional proteasomes to endothelial cells, thereby compensating for impaired protein degradation and protecting cells under hypoxic stress. Collectively, our findings provide evidence for an intercellular transfer of proteolytic capacity via stem cell-derived EVs, a mechanism that preserves endothelial proteostasis and highlights the therapeutic potential of proteasome-rich EVs for hypoxia-associated diseases. Full article

Preeclampsia is a serious pregnancy complication characterised by maternal vascular dysfunction, placental dysfunction, and organ injury, with no effective treatment currently available. Kampo, a system of Japanese traditional medicine comprising standardised herbal formulations, could target pathophysiological pathways driving preeclampsia. We evaluated the effects of select Kampo formulations on markers of preeclampsia using primary human trophoblasts, placental explants, human umbilical vein endothelial cells (HUVECs), and uterine microvascular endothelial cells (UtMVECs). Twelve formulations were initially screened in HUVECs, and six formulations advanced for further study. TNFα was used to induce endothelial dysfunction, and angiogenic, antioxidant, inflammatory, and vascular dysfunction markers were assessed. Overall, Kampo formulations had minimal effect on sFlt-1 expression and only modest effects on sFlt-1 secretion by primary human trophoblast. In contrast, several formulations consistently increased placental growth factor (PlGF) expression and secretion, upregulated HMOX1 in trophoblasts, and enhanced PlGF secretion from placental explants. In endothelial cells, Kampo treatment partially reversed TNFα-induced dysfunction, demonstrated by reduced VCAM1 expression, and additional endothelial cell type-dependent effects on ET-1 and inflammatory pathways. These findings indicate that selected Kampo formulations modulate key pathways involved in the pathophysiology underpinning preeclampsia and warrant further investigation as potential therapeutic candidates. Full article

Background: Glucose-dependent insulinotropic polypeptide receptor (GIPR) and glucagon-like peptide-1 receptor (GLP-1R) activation exert anti-diabetic and anti-obesity effects. Tirzepatide, a dual GIPR/GLP-1R agonist, has demonstrated cardiovascular benefits in clinical studies. However, the direct vascular actions of tirzepatide and their potential advantages over selective GLP-1 receptor agonists (GLP-1RAs) remain unclear. We investigated the vasoprotective effects of tirzepatide and compared them with those of GLP-1 receptor agonists in vivo and in vitro. Methods: Non-diabetic C57BL/6 and diabetic KK-Ay mice received tirzepatide, semaglutide, or vehicle. Arterial remodeling was induced by femoral artery wire injury. A subset of mice was co-treated with the nitric oxide synthase inhibitor Nω-nitro-L-arginine methyl ester (L-NAME). After 4 weeks, biochemical, morphometric, and immunofluorescence analyses were performed. In vitro, human umbilical vein endothelial cells (HUVECs) were stimulated with tirzepatide or liraglutide to assess nitric oxide (NO) production. Results: In non-diabetic mice, tirzepatide suppressed intimal hyperplasia, including at a low dose that did not affect metabolic parameters, whereas semaglutide had no significant effect on intimal hyperplasia at the same molar dose. The protective effects of tirzepatide were abolished by L-NAME. In diabetic mice, tirzepatide and semaglutide similarly improved metabolic parameters and attenuated intimal hyperplasia. In HUVECs, tirzepatide increased NO production in a dose-dependent manner, and this effect was preserved under hyperglycemic conditions. Tirzepatide and liraglutide induced comparable NO production at equivalent molar concentrations. Conclusions: Tirzepatide, but not semaglutide, exerted vasoprotective effects under non-diabetic conditions in a NO-dependent manner, whereas both agents exhibited comparable vasoprotective effects under diabetic conditions. Full article

Polyhydroxyalkanoates (PHAs) are a family of microbial polyesters distinguished by their excellent biocompatibility and tunable degradation profiles. However, systematic evaluation of the interaction effect between their chemical structure, material properties, and biological performance remains limited, particularly in veterinary medicine applications. In this study, we conducted comprehensive in vitro and in vivo evaluations of five commercially available PHA (co)polymers: poly(3-hydroxybutyrate) (PHB); poly(3-hydroxybutyrate-co-4-hydroxybutyrate) containing 5 mol% or 15 mol% 4-hydroxybutyrate monomer (P34HB 5%, P34HB 15%); and poly(3-hydroxybutyrate-co-3-hydroxyvalerate) containing 5 mol% or 15 mol% 3-hydroxyvalerate monomer (PHBV 5%, PHBV 15%). Material characterization confirmed that the introduction of proper comonomers effectively reduced crystallinity and accelerated the degradation process. Biological assays demonstrated quite excellent cytocompatibility and hemocompatibility among all tested PHA materials. Notably, P34HB 5% nanoparticles promoted human umbilical vein endothelial cell (HUVEC) migration. In a 12-week murine subcutaneous implantation model, the PHBV 15% group exhibited the thinnest fibrous capsule formation and the mildest inflammatory response. Furthermore, intramuscular injection of nanoparticles revealed favorable muscle tissue compatibility in all groups. These results provided a co-polymer ratio-based justification for PHA selection, thus providing support for the application of these biomaterials in veterinary drug delivery platforms, wound dressings, and biodegradable implants. Full article

Endothelial dysfunction is an early event in the development of cardiovascular diseases and is characterized by impaired barrier function, inflammatory activation of endothelial cells (ECs), and alterations in lipid metabolism. In addition to typical (mononuclear) endothelial cells (TECs), multinucleated variant endothelial cells (MVECs) are present within the vascular wall; however, their functional role remains poorly understood. The aim of the present study was to investigate the molecular and functional characteristics of MVECs and their potential contribution to the development of endothelial dysfunction. Primary human umbilical vein endothelial cells (HUVECs) were used, and multinucleated cells were generated by polyethylene glycol-induced fusion. Cells were incubated under control conditions or exposed to low-density lipoproteins (LDL; 100 µg/mL, 24 h). A comprehensive analysis was performed, including transcriptomic and proteomic (secretome) profiling using gene set enrichment analysis (GSEA), as well as functional assays assessing transendothelial LDL transport, intracellular cholesterol accumulation, macrophage migration, and the expression and secretion of pro-inflammatory cytokines (IL-6, IL-8). MVECs exhibited pronounced differences compared to TECs. GSEA revealed reduced enrichment of pathways related to canonical nuclear factor kappa B (NF-κB) signaling and negative regulation of NF-κB transcription factor activity, actin cytoskeleton organization, focal adhesion assembly, basement membrane organization, and vesicle-mediated transport in MVECs relative to TECs, indicating impaired cytoskeletal integrity, altered cell–matrix interactions, dysregulated inflammatory signaling, and reduced vesicular trafficking activity. Functionally, MVECs demonstrated an increased capacity for cholesterol accumulation and enhanced transendothelial migration of macrophages. Notably, transendothelial LDL transport across the MVEC monolayer was not increased, suggesting a predominance of intracellular lipid accumulation. MVECs also exhibited a pronounced pro-inflammatory phenotype, characterized by elevated expression and secretion of IL-6 and IL-8. Taken together, these findings indicate that MVECs represent a functionally altered endothelial phenotype with impaired barrier function, dysregulated lipid metabolism, and enhanced inflammatory activity. Local accumulation of MVECs within the vascular wall may contribute to the formation of pro-atherogenic regions and play a role in the initiation and progression of endothelial dysfunction. Full article

Lactate is increasingly recognized as a signaling molecule capable of modulating gene expression and vascular function. This descriptive pilot study investigated the effects of different lactate concentrations (0 mM, 10 mM, 20 mM, and 30 mM) and exposure times (1 h, 3 h, and 6 h) on the transcriptomic responses of human umbilical vein endothelial cells (HUVECs). Using next-generation RNA sequencing, an unbiased genome-wide analysis was performed, followed by focused examination of genes relevant to endothelial biology, calcium signaling, and glycocalyx integrity. Results showed that there was no statistically significant effect of lactate concentration on the expression of the examined genes. In contrast, prolonged incubation time was associated with differential expression of KLF2, KLF4, FOXO1, CD34, and VCAM1. These findings suggest that exposure time, rather than lactate concentration, may be associated with endothelial gene expression patterns under static conditions. This exploratory pilot study provides preliminary transcriptomic observations and highlights the need for further mechanistic investigations including functional and protein-level analyses. Full article

An undescribed seco-kaurane diterpenoid, benincaside A (BA), was isolated from the seeds of Benincasa hispida. The seeds of B. hispida have been traditionally used in folk medicine and previous studies have reported anti-tumor potential in B. hispida seed extracts. Accordingly, we investigated the cytotoxicity and underlying mechanisms of BA in colorectal cancer cells. BA inhibited growth in HT29, Colo205, HCT116, and CT26 colorectal cancer cells, as determined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, while showing no toxicity toward normal human umbilical vein endothelial cells (HUVEC) and human fibroblast WS-1 cells. In HCT116 cells, BA-induced deoxyribonucleic acid (DNA) damage and apoptosis, as evidenced by morphological changes, 4,6-diamidino-2-phenylindole dihydrochloride (DAPI) staining, and assays of caspase-8 and caspase-3 activities. BA triggered apoptotic cell death via the extrinsic pathway, as indicated by elevated caspase-8 and caspase-3 activities. Intracellular reactive oxygen species (ROS) generation was observed in BA-treated HCT116 cells. The growth-inhibitory effects were significantly attenuated by pretreatment with N-acetylcysteine (NAC, an antioxidant), caffeine (an ATM kinase inhibitor), z-VAD-fmk (pan-caspase inhibitor), or z-IETD-fmk (caspase-8-specific inhibitor). Colorimetric assays confirmed increased caspase-8 and caspase-3 activities in BA-treated cells. This study is the first to report ROS-dependent signaling as a key mechanism underlying BA-induced cell death in HCT116 human colorectal cancer cells. Full article