How high-salt intake leads to the occurrence of many cardiovascular diseases such as atherosclerosis is a fundamental question in pathology. Here we postulated that high-salt-induced NFAT5 controls the inflammasome activation by directly regulating NLRP3, which mediates the expression of inflammatory- and adhesion-related genes in vascular endothelium, resulting in the formation of atherosclerosis.

Inflammation-induced activation of endothelium constitutes one of the earliest changes during atherogenesis. New imaging techniques that allow detecting activated endothelial cells can improve the identification of persons at high cardiovascular risk in early stages. Quantum dots (QDs) have attractive optical properties such as bright fluorescence and high photostability, and have been increasingly studied and developed for bio-imaging and bio-targeting applications. We report here the development of vascular cell adhesion molecule-1 binding peptide (VCAM-1 binding peptide) functionalized QDs (VQDs) from amino QDs. It was found that the QD fluorescence signal in tumor necrosis factor [Formula: see text] (TNF-[Formula: see text]) treated endothelial cells in vitro was significantly higher when these cells were labeled with VQDs than amino QDs. The VQD labeling of TNF-[Formula: see text]-treated endothelial cells was VCAM-1 specific since pre-incubation with recombinant VCAM-1 blocked cells' uptake of VQDs. Our ex vivo and in vivo experiments showed that in the inflamed endothelium, QD fluorescence signal from VQDs was also much stronger than that of amino QDs. Moreover, we observed that the QD fluorescence peak was significantly blue-shifted after VQDs interacted with aortic endothelial cells in vivo and in vitro. A similar blue-shift was observed after VQDs were incubated with recombinant VCAM-1 in tube. We anticipate that the specific interaction between VQDs and VCAM-1 and the blue-shift of the QD fluorescence peak can be very useful for VCAM-1 detection in vivo.

Objective: AIP1 expression is downregulated in human atherosclerotic plaques and global deletion of AIP1 in mice exacerbates atherosclerosis in ApoE-KO mouse models. However, the direct role of AIP1 in endothelium, vascular remodeling and associated vascular diseases has not been determined. Approach and Results: We used endothelial cell (EC)-specific AIP1-deficient (AIP1-ECKO) mice to define the role of AIP1 in vascular remodeling and intima-media thickening in a mouse carotid artery ligation model characterized by both neointimal hyperplasia and inward vessel remodeling. Compared to WT littermates, AIP1-ECKO mice had 2.2-fold larger intima area and 4.4-fold thicker intima as measured by intima/media ratio in arteries with more proliferating vascular smooth muscle cells (VSMCs) at week 2-4 post-injury. Increased reactive oxygen species (ROS) in endothelium at early time points induced inflammation and vessel dysfunction in AIP1-ECKO prior to VSMC accumulations. Moreover, knockdown of AIP1 in human EC enhanced ROS generation which was attenuated by co-silencing of NOX2. Mechanistically, AIP1 via its proline-rich region binds to the SH3 domain of cytosolic subunit p47phox to disrupt formation of an active NOX2 complex, attenuating ROS production. Conclusion: Our study supports that AIP1 regulates vascular remodeling with intima-media thickening by suppressing endothelial NOX2-dependent oxidative stress. Highlights: •In a carotid ligation model, endothelial cell (EC)-specific AIP1-deficient (AIP1-ECKO) mice had much larger media area, thicker vessel wall and augmented neointima formation.•Increased production of reactive oxygen species in vascular EC at early time points concomitant with vessel dysfunction in AIP1-ECKO.•AIP1 via its proline-rich region binds to the SH3 domain of cytosolic subunit p47phox to disrupt formation of an active NOX2 complex, attenuating ROS production.

As a Rho kinase (ROCK) inhibitor, fasudil has been used in clinical trials of several cardiovascular diseases. This study was to investigate the vasorelaxant effect of fasudil on resistance arterial rings including mesenteric, renal, ventral tail and basilar artery. We also examined the potential mechanisms of its vasodilatory action using mesenteric artery rings. A DMT multiwire myograph system was used to test the tension of isolated small arteries. K(+) channel blockers, NO-cGMP pathway blockers and Ca(2+)-free physiological salt solution (PSS) were employed to verify the underlying mechanisms. Fasudil (10(-7)-10(-4)M) relaxed four types of small artery rings pre-contracted by 60mmol/l KCl (pEC50: 6.01±0.09, 5.47±0.03, 5.54±0.04, and 5.72±0.10 for mesenteric, renal, ventral tail and basilar artery rings, respectively). Pre-incubation with fasudil (1, 3, or 10μmol/l) attenuated KCl (10-60mmol/l) and angiotensin II (Ang II; 1μmol/l)-induced vasoconstriction in mesenteric artery rings. Fasudil at the concentration of 10(-6)mol/l showed different relaxant potency in endothelium intact (pEC50:6.01±0.09) or denued (5.75±0.06) mesenteric artery. The influx and release of Ca(2+) were inhibited by fasudil. In addition, fasudil could block the increased phosphorylation level of myosin light chain (MLC) and myosin-binding subunit of myosin phosphatase (MYPT1) induced by Ang II. However, pretreatment with various K(+) channel blockers did not affect the relaxant effects of fasudil remarkably. The present results demonstrate that fasudil has a vasorelaxant effect on isolated rat resistance arteries, including mesenteric, renal, ventral tail and basilar artery, and may exert its action through the endothelium, Ca(2+) channels, and the Rho/ROCK pathway.

Epoxyeicosatrienoic acids (EETs), angiogenic mediators degraded by soluble epoxide hydrolase (sEH), have been shown to exert beneficial effects on the cardiovascular system. The current study assessed the impact of increased EETs with an sEH inhibitor, t-AUCB, on two-kidney-one-clip (2K1C)-induced renovascular endothelial dysfunction, associated with hypertension, in rats. The hypertensive rats exhibited increased systolic blood pressure, reduced renal blood flow, impaired endothelium-dependent relaxation and eNOS phosphorylation in the renal arteries, elevated ROS production in the endothelium of the renal arteries, and decreased EET levels in plasma, the renal arteries, and endothelial cells; however, t-AUCB reversed all the deleterious effects. Moreover, we found that the stimulation of AMPK/UCP2 scavenged ROS and restored endothelial function in the renal arteries of hypertensive rats undergoing therapy with t-AUCB. In addition, we were the first to reveal the potential role of miR-155-5p in the occurrence and development of vascular endothelial dysfunction in hypertension. Importantly, t-AUCB recovered NO bioavailability by regulating the NF-κB/miR-155-5p/eNOS/NO/IκB cycle after the activation of AMPK/UCP2 and the subsequent inhibition of ROS in hypertensive rat renal artery endothelial cells. This study will provide evidence for this additional new mechanism, underlying the benefits of EETs and the related agents against hypertensive vasculopathy.

The migration of circulating mesenchymal stem cells (MSCs) to injured tissue is an important step in tissue regeneration and requires adhesion to the microvascular endothelium. The current study investigated the underlying mechanism of MSC adhesion to endothelial cells during inflammation. In in vitro MSC culture, tumor necrosis factor‑α (TNF‑α) increased the level of vascular cell adhesion molecule‑1 (VCAM‑1) expression in a dose‑dependent manner. The nuclear factor-κB (NF-κB), extracellular signal‑regulated kinase (ERK) and c‑Jun N‑terminal kinase (JNK) signaling pathway inhibitors, pyrrolidine dithiocarbamate (PDTC), U0126 and SP600125, respectively, suppressed VCAM‑1 expression induced by TNF‑α at the mRNA and protein levels (P<0.05). TNF‑α augmented the activation of NF‑κB, ERK and JNK, and promoted MSC adhesion to human umbilical vein endothelial cells; however, the inhibitors of NF‑κB, ERK and JNK did not affect this process in these cells. The results of the current study indicate that adhesion of circulating MSCs to the endothelium is regulated by TNF-α-induced VCAM-1 expression, which is potentially mediated by the NF‑κB, ERK and JNK signaling pathways.

Objective: This study investigated the protective effects of dipeptidyl peptidase-4 inhibitor MK-626 on vascular endothelial function by regulating lncRNAs in hypertensive vasculature. Methods: Angiotensin Ⅱ (Ang Ⅱ)-loaded osmotic pumps were implanted in mice with or without MK-626 administration. GLP-1 levels in plasma were measured by ELISA. Aortic rings were suspended in myograph for tension measurement. Microarray was performed to analyze lncRNA and mRNA expression profiles. Protein expression and phosphorylation were examined by Western blot. The differentially expressed (DE)-genes were validated by qRT-PCR. The intracellular Ca2+ concentration was detected by laser confocal system. Results: MK-626 elevated plasma GLP-1 level, increased eNOS phosphorylation, improved endothelium-dependent relaxations, and reduced systolic blood pressure in Ang Ⅱ-induced hypertensive mice. Microarray revealed the dysregulations of 723 lncRNAs and 742 mRNAs were reversed by MK-626 in hypertensive mouse aortae. qRT-PCR validation showed that 13 DE-lncRNAs and eight dysregulated mRNAs in both hypertensive mouse aortae and mouse aortic endothelial cells (MAECs) were rescued by MK-626. Among them, four mRNAs (Cacna1C, Itgav, Itga8, and Npnt) were co-expressed with lncRNA ENSMUST00000155383. Cacna1C protein expression was reduced in the ECs but was elevated in smooth muscle cells from Ang Ⅱ-infused mice, which were both reversed by MK-626. Knockdown of lncRNA ENSMUST00000155383 suppressed the increased Cacna1c protein and mRNA expression, elevated Ca2+ level, and enhanced eNOS phosphorylation induced by MK-626 in the hypertensive mouse ECs. Conclusion: The dysregulations of lncRNA ENSMUST00000155383-associated genes might play crucial roles in hypertension-induced endothelial dysfunction through affecting calcium pathway. MK-626 might ameliorate endothelial dysfunction by upregulating lncRNA ENSMUST00000155383, enhancing Ca2+ concentration, and subsequently restoring eNOS activity in hypertension.

Organogenesis and regeneration require coordination of cellular proliferation, regulated in part by secreted growth factors and cognate receptors, with tissue nutrient supply provided by expansion and patterning of blood vessels. Here we reveal unexpected combinatorial integration of a growth factor co-receptor with a heterodimeric partner and ligand known to regulate angiogenesis and vascular patterning. We show that ErbB2, which can mediate epidermal growth factor (EGF) and neuregulin signalling in multiple tissues, is unexpectedly expressed by endothelial cells where it partners with neuropilin 1 (Nrp1) to form a functional receptor for the vascular guidance molecule semaphorin 3d (Sema3d). Loss of Sema3d leads to improper patterning of the coronary veins, a phenotype recapitulated by endothelial loss of ErbB2. These findings have implications for possible cardiovascular side-effects of anti-ErbB2 therapies commonly used for cancer, and provide an example of integration at the molecular level of pathways involved in tissue growth and vascular patterning.

Familial hypercholesterolemia (FH) causes elevation of low-density lipoprotein cholesterol (LDL-C) in blood and carries an increased risk of early-onset cardiovascular disease. A caveat for exploration of new therapies for FH is the lack of adequate experimental models. We have created a comprehensive FH stem cell model with differentiated hepatocytes (iHeps) from human induced pluripotent stem cells (iPSCs), including genetically engineered iPSCs, for testing therapies for FH. We used FH iHeps to assess the effect of simvastatin and proprotein convertase subtilisin/kexin type 9 (PCSK9) antibodies on LDL-C uptake and cholesterol lowering in vitro. In addition, we engrafted FH iHeps into the liver of Ldlr-/-/Rag2-/-/Il2rg-/- mice, and assessed the effect of these same medications on LDL-C clearance and endothelium-dependent vasodilation in vivo. Our iHep models recapitulate clinical observations of higher potency of PCSK9 antibodies compared with statins for reversing the consequences of FH, demonstrating the utility for preclinical testing of new therapies for FH patients.

Recent studies have shown that Transmembrane protein 100 (TMEM100) is a gene at locus 17q32 encoding a 134-amino acid protein with two hypothetical transmembrane domainsa, and first identified as a transcript from the mouse genome. As a downstream target gene of bone morphogenetic protein (BMP)-activin receptor-like kinase 1 (ALK1) signaling, it was activated to participate in inducing arterial endothelium differentiation, maintaining vascular integrity, promoting cell apoptosis, inhibiting metastasis and proliferation of cancer cells. However, evidence for the function of TMEM100 in inflammation is still limited. In this study, we explore the role of TMEM100 in inflammatory cytokine secretion and the role of MAPK signaling pathways in tumor necrosis factor-alpha (TNF-α)-induced TMEM100 expression in LX-2 cells. We found that the expression of TMEM100 was decreased markedly in human liver fibrosis tissues, and its expression was also inhibited in LX-2 cells induced by TNF-α, suggesting that it might be associated with the development of inflammation. Therefore, we demonstrated that overexpression of TMEM100 by transfecting pEGFP-C2-TMEM100 could lead to the down-regulation of IL-1β and IL-6 secretion. Moreover, we found that expression changes of TMEM100 could be involved in inhibition or activation of MAPK signaling pathways accompanied with regulating phosphorylation levels of ERK and JNK protein in response to TNF-α. These results suggested that TMEM100 might play an important role in the secretion of inflammatory cytokines (IL-1β and IL-6) of LX-2 cells induced by TNF-α, and MAPK (ERK and JNK) signaling pathways might participate in its induction of expression.

The current study observed the effects and investigated the mechanism of remifentanil (RMF) on the isolated cerebral basilar arteries of spontaneously hypertensive rats (SHR) and Wistar‑Kyoto (WKY) rats. A pressure myograph system was used to observe and compare the effects of different concentrations of RMF (10‑10‑10‑5 mol/l) on the diameter changes of freshly isolated cerebral basilar arteries, which have been pre‑shrunk by phenylephrine (PE), an endothelium‑independent vasoconstrictor. Vascular smooth‑muscle cells of the cerebral basilar artery (BASMCs) were freshly obtained via enzymolysis. BKCa (large‑conductance calcium‑activated potassium channels) current (IBKCa) and Kv (voltage‑gated potassium channels) current (IKv) were recorded using a whole‑cell patch‑clamp technique. The changes in IBKCa and IKv produced by different concentrations of RMF (10‑10 to 10‑5 mol/l) on the two types of rats with the holding potential of ‑40 mV were observed and compared. The cerebral basilar arteries of the SHR and WKY rats were relaxed by RMF in a concentration-dependent manner (P<0.05; n=5). At the same concentration, the diastolic effect of RMF on SHR was weaker than that observed in WKY rats (P<0.05, n=5). When the rats were pre‑perfused with 10‑3 mol/l of the BKCa channel blocker tetraethylammonium (TEA), the diastolic amplitudes of RMF in SHR and WKY rats were decreased, and the fitting curves shifted down (P<0.05; n=7 and 6, respectively). However, no statistically significant difference was observed with 10‑3 mol/l of the Kv channel blocker 4‑aminopyridine (4‑AP; n=6 and 9, respectively; P>0.05). Outward currents were increased by RMF in both BASMCs of SHR and WKY rats in a voltage‑ and dose‑dependent manner (P<0.05; n=6). At the same concentration, the effect of RMF on the outward currents in BASMCs of WKY rats was stronger than that on SHR (P<0.05; n=6). The enhancing effect of RMF can be partially blocked by either 10‑3 mol/l TEA (P<0.05; n=6) or 10‑3 mol/l 4‑AP (P<0.05 or 0.01; n=6 and 9, respectively) however can be totally blocked by the mixture of TEA and 4‑AP (P<0.05, n=7). RMF served a diastolic role in the cerebral basilar arteries of rats in a dose‑dependent manner, likely by activating the BKCa and Kv channels. However, SHR demonstrated a less pronounced diastolic reaction to RMF than that observed in WKY rats.