SREBP-1-mediated lipogenesis confers resistance to ferroptosis and improves endothelial injury.

Atherosclerosis refers to a disease characterized by the formation of lipid plaque deposits within arterial walls, leading to reduced blood flow or blockage of blood outflow. The process of endothelial injury induced by oxidized low-density lipoprotein (ox-LDL) is considered the initial stage of atherosclerosis. Ferroptosis is a form of iron-dependent, non-apoptotic cell death, and current research suggests its association with coronary artery disease (CAD). In this study, we observed a correlation between reduced expression of SREBP-1 and the occurrence of stable CAD. Additionally, during the process of endothelial injury induced by ox-LDL, we also noted decreased expression of the SREBP-1/SCD1/FADS2 and involvement in the ferroptosis process. Mechanistically, ox-LDL induced endothelial injury by inhibiting the lipid biosynthesis process mediated by the SREBP-1/SCD1/FADS2, thereby inducing lipid peroxidation and ferroptosis. On the contrary, overexpression of SREBP-1 or supplementation with monounsaturated fatty acids counteracted iron accumulation, mitochondrial damage, and lipid peroxidation-induced ferroptosis, thereby improving endothelial injury. Our study indicated that the decreased expression of peripheral blood SREBP-1 mRNA is an independent risk factor for stable CAD. Furthermore, in endothelial cells, the lipid biosynthesis process mediated by SREBP-1 could ameliorate endothelial injury by resisting ferroptosis. The study has been registered with the Chinese Clinical Trial Registry, which serves as a primary registry in the World Health Organization International Clinical Trials Registry Platform (ChiCTR2300074315, August 3rd, 2023).

Relationship of human cytomegalovirus-infected endothelial cells and circulating leukocytes in the pathogenesis of disseminated human cytomegalovirus infection: A narrative review.

Among the leucocyte subpopulations circulating in peripheral blood of immune-compromised patients with disseminated Human cytomegalovirus (HCMV) infection, polymorphonuclear leuckocytes (PMNL) and M/M may carry infectious virus. While only in PMNL early HCMV replicative events do occur, monocytes are susceptible to complete virus replication when they enter human organs, where as macrophages become a site of active complete virus replication. In vivo leucocytes and endothelial cells interact continuously, as suggested by several in vitro experimental findings showing the bidirectional HCMV transmission from leucocytes to and from endothelial cells with the critical aid of adhesion molecules. Recently, the neutralising antibody response in sera from subjects with primary HCMV infection was reported to be much higher and earlier than in human embryonic lung fibroblasts (HELF) cells when measured in endothelial cells and epithelial cells, where virus entry is mediated mostly by the pentamer complex gH/gL/pUL128/pUL130/pUL131, whereas it was much lower and delayed when determined in HELF, where virus entry is mediated mostly by the trimer complex gH/gL/gO. Thus, these results suggested that products of UL128L were the molecules primary responsible for the differential neutralising antibody response. This conclusion was confirmed by a series of polyclonal and monoclonal antibodies directed to the components of pUL128L. Very recently, based on two sets of experiments including inhibition and immunoblotting assays, the pentamer complex/trimer complex ratio has been finally identified as the main factor of the neutralising antibody response. This ratio may change with the virus suspension producer and target cell system as well as number of cell culture passages.

Differential endothelial cell cycle status in postnatal retinal vessels revealed using a novel PIP-FUCCI reporter and zonation analysis.

Cell cycle regulation is critical to blood vessel formation and function, but how the endothelial cell cycle integrates with vascular regulation is not well-understood, and available dynamic cell cycle reporters do not precisely distinguish all cell cycle stage transitions in vivo. Here we characterized a recently developed improved cell cycle reporter (PIP-FUCCI) that precisely delineates S phase and the S/G2 transition. Live image analysis of primary endothelial cells revealed predicted temporal changes and well-defined stage transitions. A new inducible mouse cell cycle reporter allele was selectively expressed in postnatal retinal endothelial cells upon Cre-mediated activation and predicted endothelial cell cycle status. We developed a semi-automated zonation program to define endothelial cell cycle status in spatially defined and developmentally distinct retinal areas and found predicted cell cycle stage differences in arteries, veins, and remodeled and angiogenic capillaries. Surprisingly, the predicted dearth of S-phase proliferative tip cells relative to stalk cells at the vascular front was accompanied by an unexpected enrichment for endothelial tip and stalk cells in G2, suggesting G2 stalling as a contribution to tip-cell arrest and dynamics at the front. Thus, this improved reporter precisely defines endothelial cell cycle status in vivo and reveals novel G2 regulation that may contribute to unique aspects of blood vessel network expansion.

CTRP4 ameliorates inflammation, thereby attenuating the interaction between HUVECs and THP-1 monocytes through SIRT6/Nrf2 signaling.

CTRP4, identified as an adipokine, has demonstrated notable anti-inflammatory and anti-obesity effects in various disease models. Consequently, our research sought to explore the impact of CTRP4 on inflammation and the interaction between endothelial cells and monocytes in hyperlipidemic conditions. Using Western blotting, we assessed the expression levels of various proteins in HUVECs and THP-1 monocytes. Our study findings indicate that treatment with CTRP4 effectively mitigated the attachment of THP-1 monocytes to HUVECs. Furthermore, it reduced the expression of adhesion molecules and inflammation indicators in experimental cells exposed to hyperlipidemic conditions. Notably, CTRP4 treatment led to an increase in SIRT6 expression and the nuclear translocation of Nrf2. Interestingly, when SIRT6 or Nrf2 was silenced using siRNA, the positive effects of CTRP4 in HUVECs and THP-1 cells were nullified. Our results suggest that CTRP4 exhibits anti-inflammatory properties, thereby improving the interaction between endothelial cells and monocytes through the SIRT6/Nrf2-dependent pathway. This study provides insights into CTRP4 as a potential therapeutic target for mitigating obesity-related atherosclerosis.

IL-38 alleviates atherogenic responses via SIRT6/HO-1 signaling: A promising strategy against obesity-related atherosclerosis.

Interleukin-38 (IL-38), a member of the IL-1 family, is known for its anti-inflammatory properties mediated through ligand signaling in various disease models. It plays a significant role in atherosclerosis development, forming a theoretical basis for therapeutic strategies. However, the direct effects of IL-38 on atherogenic responses in the vascular endothelium and monocytes remain unclear. In this investigation, IL-38 treatment reduced THP-1 monocyte adhesion to HUVECs, decreased the expression of vascular adhesion molecules, and mitigated inflammation in the presence of palmitate. IL-38 treatment upregulated SIRT6 expression and enhanced autophagy markers such as LC3 conversion and p62 degradation. The effects of IL-38 were nullified by siRNA-mediated suppression of SIRT6 or heme oxygenase-1 (HO-1) in HUVECs and palmitate-treated THP-1 cells. These findings reveal that IL-38 mitigates inflammation through the SIRT6/HO-1 pathway, offering a potential therapeutic approach for addressing obesity-related atherosclerosis.

Long-term abuse of caffeine sodium benzoate induces endothelial cells injury and leads to coagulation dysfunction.

Our hospital admitted a patient who had difficulty in coagulation even after blood replacement, and the patient had abused caffeine sodium benzoate (CSB) for more than 20 years. Hence, we aimed to explore whether CSB may cause dysfunction in vascular endothelial cells and its possible mechanism. Low, medium, and high concentrations of serum of long-term CSB intake patients were used to treat HUVECs, with LPS as the positive control. MTT and CCK8 were performed to verify CSB's damaging effect on HUVECs. The expression of ET-1, ICAM-1, VCAM-1, and E-selectin were measured by ELISA. TUNEL assay and Matrigel tube formation assay were carried out to detect apoptosis and angiogenesis of HUVECs. Flow cytometry was applied to analyze cell cycles and expression of CD11b, PDGF, and ICAM-1. Expression of PDGF-BB and PCNA were examined by western blot. The activation of MAPK signaling pathway was detected by qRT-PCR and western blot. Intracellular Ca2+ density was detected by fluorescent probes. CCK8 assay showed high concentration of CSB inhibited cell viability. Cell proliferation and angiogenesis were inhibited by CSB. ET-1, ICAM-1, VCAM-1, and E-selectin upregulated in CSB groups. CSB enhanced apoptosis of HUVECs. CD11b, ICAM-1 increased and PDGF reduced in CSB groups. The expression level and phosphorylation level of MEK, ERK, JUN, and p38 in MAPK pathway elevated in CSB groups. The expression of PCNA and PDGF-BB was suppressed by CSB. Intracellular Ca2+ intensity was increased by CSB. Abuse of CSB injured HUVECs and caused coagulation disorders.

Proinflammatory signaling mechanism of endocan in macrophages: Involvement of TLR2 mediated MAPK-NFkB pathways.

Endocan is an endothelial cell-specific proteoglycan that contributes to vascular dysfunction by impairing endothelial function and inducing vascular smooth muscle cell migration. However, its role in regulating macrophage inflammation, a key pathological feature of vascular dysfunction, is not well understood. In this study, we investigated the effect of endocan on macrophage inflammation to better understand its contribution to vascular dysfunction. We found that endocan upregulated pro-inflammatory cytokines including IL-1ß, IL-6 and TNF-α in RAW 264.7 cells and activated MAPK/NFkB signaling pathways. Inhibiting these pathways reduced endocan-induced cytokine levels, while inhibiting TLR2 compromised the MAPK/NFkB regulation. Additionally, LPS-induced HUVEC conditioned medium stimulated cytokine levels in RAW 264.7 cells, which were reduced by endocan siRNA treatment in HUVEC. These results suggest that endocan positively regulates pro-inflammation in macrophages through the TLR2-MAPK-NFkB axis, highlighting the potential of targeting endocan to reduce inflammation in vascular dysfunction.

Umbilical cord mesenchymal stem cells from gestational diabetes show impaired ability to up-regulate paracellular permeability from sub-endothelial niche.

In vitro studies have shown that Wharton's jelly mesenchymal stem cells (WJ-MSCs) can cross umbilical and uterine endothelial barriers and up-regulate endothelial junctional integrity from sub-endothelial niches. This pericytic behaviour may be lost in pregnancies complicated by gestational diabetes (GDM), where increased vascular permeability and junctional disruption are reported. The aim of the present study was to investigate whether WJ-MSCs isolated from GDM pregnancies displayed any changes in morphology, proliferation, VEGF-A secretion, and their ability to influence paracellular junctional composition and permeability. WJ-MSCs were isolated from human umbilical cords from normal pregnancies (nWJ-MSCs, n=13) and those complicated by GDM (gWJ-MSCs), either diet-controlled (d-GDM, n=13) or metformin-treated (m-GDM, n=9). We recorded that 4-fold more WJ-MSCs migrated from m-GDM, and 2.5-fold from d-GDM cord samples compared with the normal pregnancy. gWJ-MSCs showed a less predominance of spindle-shaped morphology and secreted 3.8-fold more VEGF-A compared with nWJ-MSCs. The number of cells expressing CD105 (Endoglin) was higher in gWJ-MSCs compared with nWJ-MSCs (17%) at P-2. The tracer leakage after 24 h across the HUVEC + gWJ-MSCs bilayer was 22.13% and 11.2% higher in the m-GDM and d-GDM, respectively, HUVEC + nWJ-MSCs. Transfection studies with siRNAs that target Endoglin were performed in n-WJ-MSCs; transfected cells were co-cultured with HUVEC followed by permeability studies and VE-cadherin analyses. Loss of Endoglin also led to increased VEGF-A secretion, increased permeability and affected endothelial stabilization. These results reinforce the pericytic role of nWJ-MSCs to promote vascular repair and the deficient ability of gWJ-MSCs to maintain endothelial barrier integrity.

Transcriptome wide functional analysis of HBx expressing human hepatocytes stimulated with endothelial cell cross-talk.

Identification of genes dysregulated during the hepatitis B virus (HBV)-host cell interaction adds to the understanding of underlying molecular mechanisms and aids in discovering effective therapies to improve prognosis in hepatitis B virus (HBV)-infected individuals. Through bioinformatics analyses of transcriptomics data, this study aimed to identify potential genes involved in the cross-talk of human hepatocytes expressing the HBV viral protein HBx with endothelial cells. Transient transfection of HBV viral gene X (HBx) was performed in THLE2 cells using pcDNA3 constructs. Through mRNA Sequencing (RNA Seq) analysis, differentially expressed genes (DEGs) were identified. THLE2 cells transfected with HBx (THLE2x) were further treated with conditioned medium from cultured human umbilical vein derived endothelial cells (HUVEC-CM). Gene Ontology (GO) enrichment analysis revealed that interferon and cytokine signaling pathways were primarily enriched for the downregulated DEGs in THLE2x cells treated with HUVEC-CM. One significant module was selected following protein-protein interaction (PPI) network generation, and thirteen hub genes were identified from the module. The prognostic values of the hub genes were evaluated using Kaplan-Meier (KM) plotter, and three genes (IRF7, IFIT1, and IFITM1) correlated with poor disease specific survival (DSS) in HCC patients with chronic hepatitis. A comparison of the DEGs identified in HUVEC-stimulated THLE2x cells with four publicly available HBV-related HCC microarray datasets revealed that PLAC8 was consistently downregulated in all four HCC datasets as well as in HUVEC-CM treated THLE2x cells. KM plots revealed that PLAC8 correlated with worse relapse free survival and progression free survival in HCC patients with hepatitis B virus infection. This study provided molecular insights which may help develop a deeper understanding of HBV-host stromal cell interaction and open avenues for future research.

Impact of Methylated Cyclodextrin KLEPTOSE® CRYSMEB on Inflammatory Responses in Human In Vitro Models.

KLEPTOSE® CRYSMEB methylated cyclodextrin derivative displays less methylated group substitution than randomly methylated cyclodextrin. It has demonstrated an impact on atherosclerosis and neurological diseases, linked in part to cholesterol complexation and immune response, however, its impact on inflammatory cascade pathways is not clear. Thus, the impact of KLEPTOSE® CRYSMEB on various pharmacological targets was assessed using human umbilical vein endothelial cells under physiological and inflammatory conditions, followed by screening against twelve human primary cell-based systems designed to model complex human tissue and disease biology of the vasculature, skin, lung, and inflammatory tissues using the BioMAP® Diversity PLUS® panel. Finally, its anti-inflammatory mechanism was investigated on peripheral blood mononuclear cells to evaluate anti-inflammatory or pro-resolving properties. The results showed that KLEPTOSE® CRYSMEB can modulate the immune system in vitro and potentially manage vascular issues by stimulating the expression of molecules involved in the crosstalk between immune cells and other cell types. It showed anti-inflammatory effects that were driven by the inhibition of pro-inflammatory cytokine secretion and could have different impacts on different tissue types. Moreover, this cyclodextrin showed no clear impact on pro-resolving lipid mediators. Additionally, it appeared that the mechanism of action of KLEPTOSE® CRYSMEB seems to not be shared by other well-known anti-inflammatory molecules. Finally, KLEPTOSE® CRYSMEB may have an anti-inflammatory impact, which could be due to its effect on receptors such as TLR or direct complexation with LPS or PGE2, and conversely, this methylated cyclodextrin could stimulate a pro-inflammatory response involving lipid mediators and on proteins involved in communication with immune cells, probably via interaction with membrane cholesterol.

Platelet Microparticle-Derived MiR-320b Inhibits Hypertension with Atherosclerosis Development by Targeting ETFA.

Hypertension and atherosclerosis often occur simultaneously. This study aimed to explore the role and mechanism of platelet microparticle (PMP) -derived microRNA-320b (miR-320b) in patients with hypertension accompanied by atherosclerosis.We collected samples from 13 controls without hypertension and atherosclerosis and 20 patients who had hypertension accompanied by atherosclerosis. In vitro, platelets were activated by Thrombin receptor-activating peptide to produce PMPs. HUVECs were induced by CoCl2 to mimic a hypoxic environment in vitro. RT-qPCR was employed to detect the expression levels of CD61, miR-320b, and ETFA. The protein expression level of ETFA was evaluated via Western blotting. Furthermore, 3- (4,5-dimethylthiazol-2-yl) -2,5-diphenyltetrazolium bromide, 5-ethynyl-2'-deoxyuridine, and wound healing assays were employed to assess the proliferation and migration of HUVECs. Enzyme-linked immunosorbent assay was used to measure the oxidative stress and inflammation-related factor expression.The expression of miR-320b was reduced in both platelets and PMPs but increased in plasma. MiR-320b promoted CoCl2-induced HUVEC viability, proliferation, and migration. The levels of the oxidative stress factors SOD and GSH as well as the inflammatory factor IL-10 were elevated in the CoCl2 + miR-320b mimics group compared with both the CoCl2 + mimics NC and CoCl2 groups. Conversely, the levels of the oxidative stress factors MDA and ROS as well as the inflammatory factors IL-6, TNF-α, and IL-1ß were decreased. These results were regulated by miR-320b targeting ETFA.PMP-derived miR-320b inhibits the development of hypertension accompanied by atherosclerosis by targeting ETFA.

Synthesis and Biological Activity of Homohypotaurine Obtained by the Enzyme-Based Conversion of Homocysteine Sulfinic Acid Using Recombinant Escherichia Coli Glutamate Decarboxylase.

l-Homocysteine, formed from S-adenosyl methionine following demethylation and adenosine release, accumulates when the methionine recycling pathway and other pathways become impaired, thus leading to hyperhomocysteinemia, a biomarker in cardiovascular diseases, neurological/psychiatric disorders, and cancer. The partial oxidation of the l-homocysteine thiol group and its decarboxylation on C-alpha lead to the formation of l-homocysteinesulfinic acid (l-HCSA) and homohypotaurine (HHT), respectively. Both compounds are not readily available from commercial suppliers, which hinders the investigation of their biological activities. Herein, the chemical synthesis of l-HCSA, from l-homocystine, was the starting point for establishing the bio-based synthesis of HHT using recombinant Escherichia coli glutamate decarboxylase (EcGadB), an enzyme already successfully employed for the bio-based synthesis of GABA and its phosphinic analog. Prior to HHT synthesis, kcat (33.92 ± 1.07) and KM (38.24 ± 3.45 mM) kinetic constants were determined for l-HCSA on EcGadB. The results of our study show that the EcGadB-mediated synthesis of HHT can be achieved with good yields (i.e., 40% following enzymatic synthesis and column chromatography). Purified HHT was tested in vitro on primary human umbilical vein endothelial cells and rat cardiomyoblasts and compared to the fully oxidized analog, homotaurine (OT, also known as tramiprosate), in widespread pharmaceutical use. The results show that both cell lines display statistically significant recovery from the cytotoxic effects induced by H2O2 in the presence of HHT.

Identification of potential classes of glycoligands mediating dynamic endothelial adhesion of human tumor cells.

One critical step of metastasis formation is the extravasation of circulating tumor cells from the bloodstream. This process requires the dynamic interaction of cell adhesion molecules like E-selectin on endothelial cells with carbohydrate ligands on tumor cells. To characterize these glycans in a comprehensible approach, the rolling, tethering, and firm adhesion of nine human tumor cell lines on human umbilical vein endothelial cells was analyzed using laminar flow adhesion assays. The tumor cell lines were grouped into three subsets by their canonical E-selectin ligand status (sialyl-Lewis A and X +/+, -/+, -/-) and their adhesiveness was compared after enzymatic, pharmacologic, chemical treatment or antibody blockade of the tumor cells or endothelial cells, respectively. Tumor cells were also screened regarding their glycosyltransferase expression profile. We found that although E-selectin and terminal α2,3-sialic acid largely determined firm adhesion, adhesive events did not exclusively depend on the presence of sialyl-Lewis A and/or sialyl-Lewis X. Nevertheless, two of the three sialyl-Lewis A/X-/- tumor cells additionally or fully depended on vascular cell adhesion molecule-1 for firm adhesion. The significance of O-GalNAc- and N-glycans for adhesion varied remarkably among the tumor cells. The sialyl-Lewis A/X+/+ subset showed glycoprotein-independent adhesion, suggesting a role of glycolipids as well. All sialyl-Lewis A/X-/- tumor cells lacked FUT3 and FUT7 expression as opposed to sialyl-Lewis A/X+/+ or -/+ cell lines. In summary, the glycans on tumor cells mediating endothelial adhesion are not as much restricted to sialyl-Lewis A /X as previously assumed. The present study specifically suggests α2,3-linked sialic acid, O-GalNAc glycans, glycosphingolipids, and FUT3/FUT7 products as promising targets for future studies.

Protective effects of villi mesenchymal stem cells on human umbilical vein endothelial cells by inducing SPOCD1 expression in cases of gestational diabetes mellitus.

BACKGROUND: Gestational diabetes mellitus (GDM) is characterized by a lack of response to insulin in pregnancies, and often accompanied by severe complications. GDM is associated with structural and functional alterations, particularly endothelial dysfunction, in various tissues. This study is aimed to investigate the effect of placental mesenchymal stem cells (MSCs) on the endothelial biological function of human umbilical vein endothelial cells (HUVECs) and their molecular mechanisms. METHODS: Villi mesenchymal stem cells (VMSCs) were co-cultured with HUVECs, and transcriptomic analysis of differential genes was performed in HUVECs under high-glucose induction. Lentiviral transfection was performed to construct HUVECs with stable knockdown or overexpression of SPOCD1. The immunohistochemical assays were used to detect the expression of SPOCD1 in GDM patients. TUNEL fluorescence staining was applied for detection of the HUVEC apoptosis. ß galactosidase staining assay was performed to detect the cell senescence. Electron microscopy was used to detect the cell pyroptosis. qRT-PCR and western blot assays were conducted for identifying the mRNA & protein expressions of genes. RESULTS: VMSCs, when co-cultured with HUVECs, could inhibit the apoptosis, pyroptosis and senescence induced by high-glucose condition in HUVECs. Transcriptomic results showed an upregulation of SPOCD1 expression induced by VMSCs in HUVECs. Overexpression of SPOCD1 inhibited high-level glucose-induced apoptosis, pyroptosis and senescence in HUVECs via the ß-catenin pathway. CONCLUSION: VMSCs induce ß-catenin activation by upregulating the expression of SPOCD1 in HUVECs, which ultimately inhibits high-level glucose-induced apoptosis, pyroptosis and senescence in HUVECs. This observation provides potential therapeutic insight for future GDM treatment.

Co-culture with chorionic villous mesenchymal stem cells promotes endothelial cell proliferation and angiogenesis via ABCA9-AKT pathway.

Human chorionic villous mesenchymal stem cells (CV-MSCs) are a promising and effective therapeutic option for tissue injury. Vascular dysfunction during pregnancies is significantly involved in the pathogenesis of preeclampsia (PE). This work aims to investigate how CV-MSCs regulate the function of vascular endothelial cells. In this study, RNA-seq analysis was used to examine the changes in HUVECs treated with CV-MSC conditioned medium (CM). We examined the levels of ABCA9 and AKT signaling in human umbilical vein endothelial cells (HUVECs) by immunohistochemistry, western blotting, and qRT-PCR assays. CCK-8, colony formation, and tube formation assays were used to understand the role of ABCA9 in HUVEC proliferation and angiogenesis mediated by CV-MSCs. The CV-MSC treatment significantly enhanced the HUVEC proliferation and angiogenesis. Furthermore, a significant increase in the ABCA9 expression and AKT pathway activation was observed in CV-MSCs -treated HUVECs. Consistent with these findings, ABCA9 overexpression exhibited the same proliferation-and angiogenesis-promoting effect in HUVECs as induced by CV-MSC CM, also accompanied the AKT signaling activation. In addition, inhibition of ABCA9 inactivated the AKT signaling in HUVECs and reduced the HUVEC proliferation and angiogenesis. Importantly, the elevation of proliferation and angiogenesis induced by ABCA9 overexpression in HUVECs could be reversed by AKT pathway inhibition. Our results suggest that ABCA9-dependent AKT signaling activation mediated by CV-MSCs could promote HUVEC proliferation and angiogenesis.

Hypoxic regulation of ADAMTS-2 and -3 (a disintegrin and matrix metalloproteinase with thrombospondin motifs 2 and 3) procollagen N proteinases by HIF-1α in endothelial cells.

ADAMTS-2 and ADAMTS-3, known as procollagen amino proteases (PNP), are primarily responsible for processing the amino ends of the fibrillar collagen precursors. ADAMTS-2 is a highly expressed gene in type I collagen-rich tissues, such as skin, bones, tendons, and aorta. ADAMTS-3 is mainly expressed in cartilage, where it colocalizes with type II procollagen and in the nervous system. Studies about ADAMTS-2 and ADAMTS-3 enzymes primarily focused on their collagen processing activity. Knowledge about the transcriptional regulations of these genes is rather limited. Here we analyzed the transcriptional regulations of ADAMTS-2 and ADAMTS-3 genes under chemically induced hypoxic conditions in endothelial cell model, HUVECs. We elucidated that hypoxia is the potent positive regulator of ADAMTS-2 and ADAMTS-3 genes. qRT-PCR and western blotting studies revealed that ADAMTS-2 and ADAMTS-3 expressions were increased at mRNA and protein levels under chemically induced hypoxic conditions in HUVECs. In addition, Transient transfection experiments of ADAMTS-2 and ADAMTS-3 promoter-reporter constructs indicated that low oxygen conditions increased ADAMTS-2 and ADAMTS-3 promoter activities. Furthermore, the DNA-protein interaction assay provided evidence of the functional binding of HIF-1α on bioinformatically determined HRE regions on the ADAMTS-2 and ADAMTS-3 promoters.

Long-Term Experimental Hyperglycemia Does Not Impair Macrovascular Endothelial Barrier Integrity and Function in vitro.

Hyperglycemia is a hallmark of type 2 diabetes implicated in vascular endothelial dysfunction and cardiovascular complications. Many in vitro studies identified endothelial apoptosis as an early outcome of experimentally modeled hyperglycemia emphasizing cell demise as a significant factor of vascular injury. However, endothelial apoptosis has not been observed in vivo until the late stages of type 2 diabetes. Here, we studied the long-term (up to 4 weeks) effects of high glucose (HG, 30 mM) on human umbilical vein endothelial cells (HUVEC) in vitro. HG did not alter HUVEC monolayer morphology, ROS levels, NO production, and exerted minor effects on the HUVEC apoptosis markers. The barrier responses to various clues were indistinguishable from those by cells cultured in physiological glucose (5 mM). Tackling the key regulators of cytoskeletal contractility and endothelial barrier revealed no differences in the histamine-induced intracellular Ca2+ responses, nor in phosphorylation of myosin regulatory light chain or myosin light chain phosphatase. Altogether, these findings suggest that vascular endothelial cells may well tolerate HG for relatively long exposures and warrant further studies to explore mechanisms involved in vascular damage in advanced type 2 diabetes.

Dienogest does not augment the gene expression of adhesion molecules, MCP-1, and monocyte adherence in human endothelial cells.

OBJECTIVE: Medroxyprogesterone acetate (MPA) may increase the risk of atherosclerosis during hormone replacement therapy (HRT); therefore, the effect of progestogens other than MPA on atherosclerotic lesions requires evaluation. Adhesion of monocytes to vascular endothelial cells is an important early step in atherosclerosis progression. MCP-1 is a key chemokine that promotes monocyte migration and adhesion to vascular endothelial cells. In this study, we investigated the effects of dienogest (DNG), an alternative progestogen, on monocyte adhesion and cytokine expression in human umbilical vein endothelial cells (HUVECs). STUDY DESIGN: HUVECs were treated with DNG, natural progesterone, or MPA, followed by interleukin (IL)-1ß stimulation. The mRNA expression of adhesion molecules (E-selectin and ICAM-1) and cytokines (MCP-1 and IL-6) was examined using real-time PCR. A flow chamber system was used to examine the effect of DNG on the adhesion of U937 monocytic cells to monolayer HUVECs. RESULTS: Unlike MPA, DNG did not alter the mRNA expression of E-selectin, ICAM-1, MCP-1, and IL-6 in HUVECs. Moreover, it did not increase the number of monocytes adhering to HUVECs in the flow chamber system. However, MPA treatment significantly enhanced monocyte adhesion to HUVECs (p < 0.05). CONCLUSIONS: DNG had no effect on the mRNA expression of adhesion molecules and cytokines in HUVECs, as well as the monocyte adhesion to HUVECs, suggesting that DNG can be explored as an alternative to MPA for HRT.

Investigation of the optimal light parameters for photobiomodulation to induce osteogenic differentiation of the human bone marrow stem cell and human umbilical vein endothelial cell co-culture.

Bones have an important role in the human body with their complex nature. Mesenchymal stem cells and endothelial cells together support their unique and complex nature. Photobiomodulation (PBM) is a promising method that provides cell proliferation, osteogenic differentiation, and bone regeneration. However, there are still unknowns in the mechanism of osteogenic differentiation induced by PBM. The main aim of the study is to understand the molecular mechanism of PBM at 655 and 808 nm of wavelengths and identify the most effective energy densities of both wavelengths for osteogenic differentiation. The effect of PBM on osteogenic differentiation of Human Bone Marrow Stem Cell (hBMSC) and Human Umbilical Vein Endothelial Cell (HUVEC) co-culture was examined at 1, 3, and 5 J/cm2 energy densities of red and near-infrared light through different analysis such as cell viability, scratch assay, intracellular reactive oxygen species production, and ATP synthesis, nitric oxide release, temperature monitoring, and osteogenic differentiation analyses. Even though all PBM-treated groups exhibited better results compared to the control group, 5 J/cm2 energy density induced faster cell proliferation and migration at both wavelengths. The increases in ATP and NO levels as signaling molecules, and the increases in DNA, ALPase, and calcium contents as osteogenic markers were higher in the groups treated with 5 J/cm2 energy density at both wavelengths. Only a slight change was obtained in the level of intracellular ROS after any light applications. It can be concluded that NO release has a very important role together with ATP production in PBM therapy to trigger DNA synthesis, ALPase activity, and mineralization for osteogenic differentiation of the hBMSC and HUVEC co-culture at 655 and 808 nm of wavelengths.

Protective Effect of Cannabidiol on Hydrogen Peroxide-Induced Oxidative Damage in Human Umbilical Vein Endothelial Cells (HUVECs).

Natural antioxidants play an important role in promoting good health because of their prevention for oxidative damage. The work aimed to explore the antioxidant mechanism and activity of cannabidiol (CBD) at the cellular level. The human umbilical vein endothelial cell (HUVEC) with oxidative damage was employed as the model to study the protective capability of CBD. The results showed that CBD pre-treatment before the cells were exposed to hydrogen peroxide (H2 O2 ) resulted in an obvious increase of cell viability (about 100 %) and antioxidant related enzymes activity, and a decline of malondialdehyde (MDA) level. Besides, CBD could alleviate the increase of intracellular reactive oxygen species (ROS) content, the contraction of nucleus, and condensation of chromatin. The changes showed a dose-dependent effect. Additionally, the free radicals scavenging capacity of CBD was comparable to that of typical natural antioxidant, anthocyanidins. In summary, CBD could be employed as a potent antioxidant source for avoiding the oxidative damage. These results could provide the foundation for the development of CBD antioxidant products.