Effects of dipeptidyl peptidase 4 inhibition on the endothelial control of the vascular tone.

Dipeptidyl peptidase 4 (DPP4) is a serine protease known to cleave incretin hormones, which stimulate insulin secretion after food intake, a fact that supported the development of its inhibitors (DPP4i or gliptins) for the treatment of type 2 diabetes mellitus. In addition to their glucose-lowering effects, DPP4i show benefits for the cardiovascular system that could be related, at least in part, to their protective action on vascular endothelium. DPP4i have been associated with the reversal of endothelial dysfunction, an important predictor of cardiovascular events and a hallmark of diseases such as atherosclerosis, diabetes mellitus, hypertension, and heart failure. In animal models of these diseases, DPP4i increase nitric oxide bioavailability and limits oxidative stress, thereby improving the endothelium-dependent relaxation. Similar effects on flow-mediated dilation and attenuation of endothelial dysfunction have also been noted in human studies, suggesting a value for gliptins in the clinical scenario, despite the variability of the results regarding the DPP4i used, treatment duration, and presence of comorbidities. In this mini-review, we discuss the advances in our comprehension of the DPP4i effects on endothelial regulation of vascular tone. Understanding the role of DPP4 and its involvement in the signaling mechanisms leading to endothelial dysfunction will pave the way for a broader use of DPP4i in conditions that endothelial dysfunction is a pivotal pathophysiological player.

γ-Bungarotoxin impairs the vascular endothelial barrier function by inhibiting integrin α5.

γ-bungarotoxin (γ-BGT) is an RGD motif-containing protein, derived from the venom of Bungarus multicinctus, leading to acute death in mice. These RGD motif-containing proteins from snake venom belonging to the disintegrin family can interfere with vascular endothelial homeostasis by directly binding cell surface integrins. Targeting integrins that generate vascular endothelial dysfunction may contribute to γ-BGT poisoning, however, the underlying mechanisms have not been investigated in detail. In this study, the results showed that γ-BGT played a role in -promoting the permeability of the vascular endothelial barrier. Depending on its selective binding to integrin α5 in vascular endothelium (VE), γ-BGT initiated downstream events, including focal adhesion kinase dephosphorylation and cytoskeleton remodeling, resulting in the intercellular junction interruption. Those alternations facilitated paracellular permeability of VE and barrier dysfunction. Proteomics profiling identified that as a downstream effector of the integrin α5 / FAK signaling pathway cyclin D1 partially mediated the cellular structural changes and barrier dysfunction. Furthermore, VE-released plasminogen activator urokinase and platelet-derived growth factor D could serve as potential diagnostic biomarkers for γ-BGT-induced vascular endothelial dysfunction. Our results indicate the mechanisms through which γ-BGT as a novel disintegrin directly interacts with the VE, with consequences for barrier dysfunction.

Endothelium-Dependent Induction of Vasculogenic Mimicry in Human Triple-Negative Breast Cancer Cells Is Inhibited by Calcitriol and Curcumin.

In highly aggressive tumors, cancer cells may form channel-like structures through a process known as vasculogenic mimicry (VM). VM is generally associated with metastasis, mesenchymal phenotype, and treatment resistance. VM can be driven by antiangiogenic treatments and/or tumor microenvironment-derived factors, including those from the endothelium. Curcumin, a turmeric product, inhibits VM in some tumors, while calcitriol, the most active vitamin D metabolite, exerts potent antineoplastic effects. However, the effect of these natural products on VM in breast cancer remains unknown. Herein, we studied the effect of both compounds on triple-negative breast cancer (TNBC) VM-capacity in a co-culture model. The process of endothelial cell-induced VM in two human TNBC cell lines was robustly inhibited by calcitriol and partially by curcumin. Calcitriol promoted TNBC cells' morphological change from spindle-like to cobblestone-shape, while curcumin diminished VM 3D-structure. Notably, the treatments dephosphorylated several active kinases, especially those involved in the PI3K/Akt pathway. In summary, calcitriol and curcumin disrupted endothelium-induced VM in TNBC cells partially by PI3K/Akt inactivation and mesenchymal phenotype inhibition. Our results support the possible use of these natural compounds as adjuvants for VM inactivation in patients with malignant tumors inherently capable of forming VM, or those with antiangiogenic therapy, warranting further in vivo studies.

Eukaryotic elongation factor 2 kinase inhibitor, A484954 inhibits perivascular sympathetic nerve stimulation-induced vasoconstriction in isolated renal artery.

Eukaryotic elongation factor 2 (eEF2) kinase (eEF2K) repressively regulates protein translation through phosphorylating eEF2. We previously showed that expression and activity of eEF2K are increased in isolated mesenteric arteries from spontaneously hypertensive rats (SHR) contributing to development of essential hypertension. Furthermore, we have recently shown that 7-Amino-1-cyclopropyl-3-ethyl-1,2,3,4-tetrahydro-2,4-dioxopyrido[2,3-d]pyrimidine-6-carboxamide (A484954), a selective eEF2K inhibitor, induces endothelium-dependent relaxation in isolated mesenteric arteries from SHR inducing an antihypertensive effect. In order to test the hypothesis that inhibition of eEF2K activity induces vasodilatation by suppressing sympathetic nerve activity, we examined the effects of A484954 on perivascular sympathetic nerve stimulation-induced contraction in isolated renal artery from normotensive and hypertensive rats. Electrodes were placed near the isolated renal arteries that were applied with transmural nerve stimulation (TNS). Then, contraction of the arteries was isometrically measured. A484954 inhibited TNS-induced contraction. The A484954-mediated inhibition of TNS-induced contraction was significantly prevented by NG-nitro-L-arginine methyl ester. In SHR isolated renal artery, TNS-induced contraction was enhanced compared with normotensive Wistar rats. Furthermore, A484954-mediated inhibition of TNS-induced contraction in SHR was enhanced compared with Wistar rats. In conclusion, this study demonstrates for the first time that A484954 inhibits perivascular sympathetic nerve stimulation-induced vasoconstriction at least in part perhaps through nitric oxide (NO) release from NO-operating nerve.

Far-infrared radiation alleviates cisplatin-induced vascular damage and impaired circulation via activation of HIF-1α.

Severe vascular damage and complications are often observed in cancer patients during treatment with chemotherapeutic drugs such as cisplatin. Thus, development of potential options to ameliorate the vascular side effects is urgently needed. In this study, the effects and the underlying mechanisms of far-infrared radiation (FIR) on cisplatin-induced vascular injury and endothelial cytotoxicity/dysfunction in mice and human umbilical vein endothelial cells (HUVECs) were investigated. An important finding is that the severe vascular stenosis and poor blood flow seen in cisplatin-treated mice were greatly mitigated by FIR irradiation (30 minutes/day) for 1-3 days. Moreover, FIR markedly increased the levels of phosphorylation of PI3K and Akt, and VEGF secretion, as well as the expression and the activity of hypoxia-inducible factor 1α (HIF-1α) in cisplatin-treated HUVECs in a promyelocytic leukemia zinc finger protein (PLZF)-dependent manner. However, FIR-stimulated endothelial angiogenesis and VEGF release were significantly diminished by transfection with HIF-1α siRNA. We also confirmed that HIF-1α, PI3K, and PLZF contribute to the inhibitory effect of FIR on cisplatin-induced apoptosis in HUVECs. Notably, FIR did not affect the anticancer activity and the HIF-1α/VEGF cascade in cisplatin-treated cancer cells under normoxic or hypoxic condition, indicating that the actions of FIR may specifically target endothelial cells. It is the first study to demonstrate that FIR effectively attenuates cisplatin-induced vascular damage and impaired angiogenesis through activation of HIF-1α-dependent processes via regulation of PLZF and PI3K/Akt. Taken together, cotreatment with the noninvasive and easily performed FIR has a therapeutic potential to prevent the pathogenesis of vascular complications in cancer patients during cisplatin treatment.

Comparison of Three Species of Rhubarb in Inhibiting Vascular Endothelial Injury via Regulation of PI3K/AKT/NF-κB Signaling Pathway.

Background/Aim: Rhubarb, a traditional Chinese medicine derived from three species, is commonly used in the prescriptions for promoting blood circulation and removing blood stasis based on its traditional effects of removing blood stasis and dredging the meridians. It has been reported that rhubarb can protect blood vessels by reducing inflammation and inhibiting vascular endothelial injury (VEI), but the effective components and mechanism of rhubarb inhibiting VEI are still unclear. This study aimed to compare the differences in chemical compositions of the three species of rhubarb and their inhibitory effect on VEI, so as to explain the material basis and select the dominant species to inhibit VEI, and to elucidate the mechanism of rhubarb's inhibitory effect on VEI. Methods: Plant metabolomics was used to compare the chemical components of three species of rhubarb. The efficacy of three species of rhubarb in inhibiting VEI was compared through cell experiments in vitro. At the same time, combined with network pharmacology and molecular docking, the effective components and pathways of rhubarb involved in inhibiting VEI were screened. The mechanism of rhubarb inhibiting VEI was verified by molecular biology. Results: There were significant differences in the distribution of chemical components among the three species of rhubarb. We identified 36 different chemical components in the positive ion mode and 38 different chemical components in the negative ion mode. Subsequently, the results showed significant differences in inhibiting VEI among the three species of rhubarb based on the contents of inflammatory factors (such as IL-1ß, IL-6, and TNF-α), ROS, and NO and confirmed that R. tanguticum had the best inhibitory effect on VEI in the light of the comprehensive efficacy, compared with R. palmatum and R. officinale. Three species of rhubarb alleviated the inflammatory response in LPS-induced EA.hy926 cells by reducing the contents of inflammatory cytokines IL-6, IL-1ß, and TNF-α and decreasing expressions of PI3K, AKT, NF-κB p65, and STAT3 protein in the PI3K/AKT/NF-κB pathway and the inhibition of proteins phosphorylation. In addition, three species of rhubarb could lessen the contents of ROS and NO in EA.hy926 cells induced by LPS. All results indicated that the process of inflammation-induced cellular oxidative stress, which resulted in VEI, was obviously improved by three species of rhubarb. Conclusion: R. tanguticum was more effective among three species of rhubarb, and it had been proved that gallic acid, gallic-acid-O-galloyl-glucoside, procyanidin B-2,3,3'-di-O-gallatein, and other potential components could reduce the contents of inflammatory factors (such as IL-1ß, IL-6, and TNF-α), ROS, and NO by inhibiting the PI3K/AKT/NF-κB signaling pathway and protected the vascular endothelium and the blood vessels by improving the inflammation and oxidative stress reaction.

Comparative effects of estrogen and silibinin on cardiovascular risk biomarkers in ovariectomized rats.

BACKGROUND: Silibinin is a polyphenolic compound that could modulate estrogen receptor activation. Vascular dysfunction is considered a key initiator in atherosclerosis and may occur in the postmenopausal period. This manuscript compares estrogen and silibinin's impacts on factors that change endothelial function in ovariectomized (OVX) rats. METHODS: 32 female Wistar rats were subdivided into control; OVX; OVX + estrogen (1 mg/kg/day); and OVX + silibinin (50 mg/kg/day) groups. After the experimental period, lipid profile, atherogenic indices, and histopathology of endothelium were monitored. The vascular oxidative stress, adhesion molecules, inflammatory cytokine levels, nitric oxide (NO), angiotensin-II (Ang-II), and endothelin-1 (ET-1) were also analyzed. RESULTS: Silibinin treatment, similar to estrogen, significantly normalized the adverse changes of OVX on vascular function, including improved lipid profile and oxidative stress, increased endothelial nitric oxide synthase (eNOS) expression, diminished inflammatory status, and reduced adhesion molecule levels, ET-1 and Ang-II substances. Our findings also revealed that the administration with estrogen or silibinin resulted in a normal endothelium layer in the aorta tissues of OVX rats. CONCLUSION: Estrogen and silibinin have similar effects in improving vascular function. These treatments' protective impacts on vasculature indicate their potential benefits on the cardiovascular system in the postmenopausal period.

Improve endothelialization of metallic cardiovascular stent via femtosecond laser induced micro/nanostructure dependent cells proliferation and drug delivery control.

Regarding restenosis occurrence risk after metallic stent deployment in artery, stents with vascular smooth muscle cells antiproliferative agents sustained released from poly(lactic-co-glycolic acid) (PLGA) coating and endothelial cells proliferation favored surface textures were both attempted for endothelialization enhancement. In order to explore the interaction between the surface texture and performance of drug loaded PLGA coating, femtosecond laser surface treatment was used to change the surface characteristics of 316L stainless steel. Two different surface patterns in form of stripe (FSL100) and isolated island-like structure (FSL800) were firstly generated by femtosecond laser processing with 100 and 800 mW energy, then Rapamycin loaded PLGA coating was further deposited to polished and femtosecond laser processed 316L surfaces via a dip-coating method. The subsequent drug loading capacity and release profile studies confirmed the roles of surface texture. Morphological transition characteristics of the PLGA coating on the FLS100 sample indicate that the coating has integrity during degradation compared to the polished one. Finally, rapamycin eluting FLS100 stent was deployed to iliac arteries of New Zealand White rabbits with vascular plaques to demonstrate its endothelialization potential and resistance to restenosis.

A Central Role for TRPM4 in Ca2+-Signal Amplification and Vasoconstriction.

Transient receptor potential melastatin-4 (TRPM4) is activated by an increase in intracellular Ca2+ concentration and is expressed on smooth muscle cells (SMCs). It is implicated in the myogenic constriction of cerebral arteries. We hypothesized that TRPM4 has a general role in intracellular Ca2+ signal amplification in a wide range of blood vessels. TRPM4 function was tested with the TRPM4 antagonist 9-phenanthrol and the TRPM4 activator A23187 on the cardiovascular responses of the rat, in vivo and in isolated basilar, mesenteric, and skeletal muscle arteries. TRPM4 inhibition by 9-phenanthrol resulted in hypotension and a decreased heart rate in the rat. TRPM4 inhibition completely antagonized myogenic tone development and norepinephrine-evoked vasoconstriction, and depolarization (high extracellular KCl concentration) evoked vasoconstriction in a wide range of peripheral arteries. Vasorelaxation caused by TRPM4 inhibition was accompanied by a significant decrease in intracellular Ca2+ concentration, suggesting an inhibition of Ca2+ signal amplification. Immunohistochemistry confirmed TRPM4 expression in the smooth muscle cells of the peripheral arteries. Finally, TRPM4 activation by the Ca2+ ionophore A23187 was competitively inhibited by 9-phenanthrol. In summary, TRPM4 was identified as an essential Ca2+-amplifying channel in peripheral arteries, contributing to both myogenic tone and agonist responses. These results suggest an important role for TRPM4 in the circulation. The modulation of TRPM4 activity may be a therapeutic target for hypertension. Furthermore, the Ca2+ ionophore A23187 was identified as the first high-affinity (nanomolar) direct activator of TRPM4, acting on the 9-phenanthrol binding site.

γ Peptide Nucleic Acid-Based miR-122 Inhibition Rescues Vascular Endothelial Dysfunction in Mice Fed a High-Fat Diet.

The blood levels of microRNA-122 (miR-122) is associated with the severity of cardiovascular disorders, and targeting it with efficient and safer miR inhibitors could be a promising approach. Here, we report the generation of a γ-peptide nucleic acid (γPNA)-based miR-122 inhibitor (γP-122-I) that rescues vascular endothelial dysfunction in mice fed a high-fat diet. We synthesized diethylene glycol-containing γP-122-I and found that its systemic administration counteracted high-fat diet (HFD)-feeding-associated increase in blood and aortic miR-122 levels, impaired endothelial function, and reduced glycemic control. A comprehensive safety analysis established that γP-122-I affects neither the complete blood count nor biochemical tests of liver and kidney functions during acute exposure. In addition, long-term exposure to γP-122-I did not change the overall adiposity, or histology of the kidney, liver, and heart. Thus, γP-122-I rescues endothelial dysfunction without any evidence of toxicity in vivo and demonstrates the suitability of γPNA technology in generating efficient and safer miR inhibitors.

Infection of Endothelial Cells by Dengue Virus Induces ROS Production by Different Sources Affecting Virus Replication, Cellular Activation, Death and Vascular Permeability.

Exacerbated inflammatory response and altered vascular function are hallmarks of dengue disease. Reactive oxygen species (ROS) production has been associated to endothelial barrier disturbance and microvascular alteration in distinct pathological conditions. Increased ROS has been reported in in vitro models of dengue virus (DENV) infection, but its impact for endothelial cell physiology had not been fully investigated. Our group had previously demonstrated that infection of human brain microvascular endothelial cells (HBMEC) with DENV results in the activation of RNA sensors and production of proinflammatory cytokines, which culminate in cell death and endothelial permeability. Here, we evaluated the role of mitochondrial function and NADPH oxidase (NOX) activation for ROS generation in HBMEC infected by DENV and investigated whether altered cellular physiology could be a consequence of virus-induced oxidative stress. DENV-infected HBMECs showed a decrease in the maximal respiratory capacity and altered membrane potential, indicating functional mitochondrial alteration, what might be related to mtROS production. Indeed, mtROS was detected at later time points after infection. Specific inhibition of mtROS diminished virus replication, cell death, and endothelial permeability, but did not affect cytokine production. On the other hand, inhibition of NOX-associated ROS production decreased virus replication and cell death, as well as the secretion of inflammatory cytokines, including IL-6, IL-8, and CCL5. These results demonstrated that DENV replication in endothelial cells induces ROS production by different pathways, which impacts biological functions that might be relevant for dengue pathogenesis. Those data also indicate oxidative stress events as relevant therapeutical targets to avoid vascular permeability, inflammation, and neuroinvasion during DENV infection.

Interferon-ß Suppresses Transcriptionally Active Parvovirus B19 Infection in Viral Cardiomyopathy: A Subgroup Analysis of the BICC-Trial.

Human parvovirus B19 (B19V) is the predominant virus currently detected in endomyocardial biopsies (EMBs). Recent findings indicate that, specifically, transcriptionally active B19V with detectable viral RNA is of prognostic relevance in inflammatory viral cardiomyopathy. We aimed to evaluate B19V replicative status (viral RNA) and beneficial effects in a sub-collective of the prospective randomized placebo-controlled phase II multi-center BICC-Trial (Betaferon In Chronic Viral Cardiomyopathy) after interferon beta-1b (IFN-ß) treatment. EMBs of n = 64 patients with B19V mono-infected tissue were retrospectively analyzed. Viral RNA could be detected in n = 18/64 (28.1%) of B19V DNA positive samples (mean age 51.7 years, 12 male), of whom n = 13 had been treated with IFN-ß. Five patients had received placebo. PCR analysis confirmed in follow-up that EMBs significantly reduced viral RNA loads in n = 11/13 (84.6%) of IFN-ß treated patients (p = 0.001), independently from the IFN-ß dose, in contrast to the placebo group, where viral RNA load was not affected or even increased. Consequently, a significant improvement of left ventricular ejection fraction (LVEF) after treatment with IFN-ß was observed (LVEF mean baseline 51.6 ± 14.1% vs. follow-up 61.0 ± 17.5%, p = 0.03). In contrast, in the placebo group, worsening of LVEF was evaluated in n = 4/5 (80.0%) of patients. We could show for the first-time the beneficial effects from treatment with IFN-ß, suppressing B19V viral RNA and improving the hemodynamic course. Our results need further verification in a larger prospective randomized controlled trial.

Simultaneous Determination of Seven Lipophilic and Hydrophilic Components in Salvia miltiorrhiza Bunge by LC-MS/MS Method and Its Application to a Transport Study in a Blood-Brain-Barrier Cell Model.

Salvia miltiorrhiza Bunge (SM) has been extensively used in Alzheimer's disease treatment, the permeability through the blood-brain barrier (BBB) determining its efficacy. However, the transport mechanism of SM components across the BBB remains to be clarified. A simple, precise, and sensitive method using LC-MS/MS was developed for simultaneous quantification of tanshinone I (TS I), dihydrotanshinone I (DTS I), tanshinone IIA (TS IIA), cryptotanshinone (CTS), protocatechuic aldehyde (PAL), protocatechuic acid (PCTA), and caffeic acid (CFA) in transport samples. The analytes were separated on a C18 column by gradient elution. Multiple reaction monitoring mode via electrospray ionization source was used to quantify the analytes in positive mode for TS I, DTS I, TS IIA, CTS, and negative mode for PAL, PCTA, and CFA. The linearity ranges were 0.1-8 ng/mL for TS I and DTS I, 0.2-8 ng/mL for TS IIA, 1-80 ng/mL for CTS, 20-800 ng/mL for PAL and CFA, and 10-4000 ng/mL for PCTA. The developed method was accurate and precise for the compounds. The relative matrix effect was less than 15%, and the analytes were stable for analysis. The established method was successfully applied for transport experiments on a BBB cell model to evaluate the apparent permeability of the seven components.

Anti-Inflammatory Activity of Orally Administered Monostroma nitidum Rhamnan Sulfate against Lipopolysaccharide-Induced Damage to Mouse Organs and Vascular Endothelium.

We previously reported that rhamnan sulfate (RS) purified from Monostroma nitidum significantly suppressed lipopolysaccharide (LPS)-induced inflammation in cultured human vascular endothelial cells. Here, we analyzed the effect of orally administered RS on LPS-induced damage to mouse organs and vascular endothelium. RS (1 mg) was orally administered daily to BALB/c mice, 50 µg of LPS was intraperitoneally administered on day 8, and Evans blue was injected into the tail vein 6 h later. After 30 min, LPS-treated mice showed pulmonary Evans blue leakage and elevated plasma levels of liver damage markers, whereas this reaction was suppressed in LPS + RS-treated mice. Immunohistochemical and Western blot analysis of mouse organs 24 h after LPS treatment showed significant neutrophil infiltration into the lung, liver, and jejunum tissues of LPS-treated mice and high expression levels of inflammation-related factors in these tissues. Expression levels of these factors were significantly suppressed in LPS + RS-treated mice. Analysis of lung glycocalyx showed a significant reduction in glycocalyx in LPS-treated mice but not in LPS + RS-treated mice. Levels of syndecan-4, one of the glycocalyx components, decreased in LPS-treated mice and increased in LPS + RS-treated mice. The current results suggest that orally administered RS protects organs and vascular endothelium from LPS-induced inflammation and maintains blood circulation.

Aryl Hydrocarbon Receptor Inhibition Restores Indoxyl Sulfate-Mediated Endothelial Dysfunction in Rat Aortic Rings.

The uremic toxin indoxyl sulfate (IS), elevated in chronic kidney disease (CKD), is known to contribute towards progressive cardiovascular disease. IS activates the aryl hydrocarbon receptor (AhR) mediating oxidative stress and endothelial dysfunction via activation of the CYP1A1 pathway. The present study examines AhR inhibition with the antagonist, CH223191, on IS-mediated impairment of vascular endothelial function and disruption of redox balance. The acute effects of IS on endothelium-dependent relaxation were assessed in aortic rings from Sprague Dawley rats exposed to the following conditions: (1) control; (2) IS (300 µM); (3) IS + CH223191 (1 µM); (4) IS + CH223191 (10 µM). Thereafter, tissues were assessed for changes in expression of redox markers. IS reduced the maximum level of endothelium-dependent relaxation (Rmax) by 42% (p < 0.001) compared to control, this was restored in the presence of increasing concentrations of CH223191 (p < 0.05). Rings exposed to IS increased expression of CYP1A1, nitro-tyrosine, NADPH oxidase 4 (NOX4), superoxide, and reduced eNOS expression (p < 0.05). CH223191 (10 µM) restored expression of these markers back to control levels (p < 0.05). These findings demonstrate the adverse impact of IS-mediated AhR activation on the vascular endothelium, where oxidative stress may play a critical role in inducing endothelial dysfunction in the vasculature of the heart and kidneys. AhR inhibition could provide an exciting novel therapy for CVD in the CKD setting.

Activation of the aryl hydrocarbon receptor by clozapine induces preadipocyte differentiation and contributes to endothelial dysfunction.

BACKGROUND: The superior therapeutic benefit of clozapine is often associated with metabolic disruptions as obesity, insulin resistance, tachycardia, higher blood pressure, and even hypertension. AIMS: These adverse vascular/ metabolic events under clozapine are similar to those caused by polycyclic aromatic hydrocarbons (PAHs), and clozapine shows structural similarity to well-known ligands of the aryl hydrocarbon receptor (AhR). Therefore, we speculated that the side effects caused by clozapine might rely on AhR signaling. METHODS: We examined clozapine-induced AhR activation by luciferase reporter assays in hepatoma HepG2 cells and we proved upregulation of the prototypical AhR target gene Cyp1A1 by realtime-PCR (RT-PCR) analysis and enzyme activity. Next we studied the physiological role of AhR in clozapine's effects on human preadipocyte differentiation and on vasodilatation by myography in wild-type and AhR-/- mice. RESULTS: In contrast to other antipsychotic drugs (APDs), clozapine triggered AhR activation and Cyp1A1 expression in HepG2 cells and adipocytes. Clozapine induced adipogenesis via AhR signaling. After PGF2α-induced constriction of mouse aortic rings, clozapine strongly reduced the maximal vasorelaxation under acetylcholine in rings from wild-type mice, but only slightly in rings from AhR-/- mice. The reduction was also prevented by pretreatment with the AhR antagonist CH-223191. CONCLUSION: Identification of clozapine as a ligand for the AhR opens new perspectives to explain common clozapine therapy-associated adverse effects at the molecular level.

Topical application of TAK1 inhibitor encapsulated by gelatin particle alleviates corneal neovascularization.

Rationale: Corneal neovascularization (CoNV) is a severe complication of various types of corneal diseases, that leads to permanent visual impairment. Current treatments for CoNV, such as steroids or anti-vascular endothelial growth factor agents, are argued over their therapeutic efficacy and adverse effects. Here, we demonstrate that transforming growth factor-ß (TGF-ß)-activated kinase 1 (TAK1) plays an important role in the pathogenesis of CoNV. Methods: Angiogenic activities were assessed in ex vivo and in vitro models subjected to TAK1 inhibition by 5Z-7-oxozeaenol, a selective inhibitor of TAK1. RNA-Seq was used to examine pathways that could be potentially affected by TAK1 inhibition. A gelatin-nanoparticles-encapsulated 5Z-7-oxozeaenol was developed as the eyedrop to treat CoNV in a rodent model. Results: We showed that 5Z-7-oxozeaenol reduced angiogenic processes through impeding cell proliferation. Transcriptome analysis suggested 5Z-7-oxozeaenol principally suppresses cell cycle and DNA replication, thereby restraining cell proliferation. In addition, inhibition of TAK1 by 5Z-7-oxozeaenol blocked TNFα-mediated NFκB signalling, and its downstream genes related to angiogenesis and inflammation. 5Z-7-oxozeaenol also ameliorated pro-angiogenic activity, including endothelial migration and tube formation. Furthermore, topical administration of the gelatin-nanoparticles-encapsulated 5Z-7-oxozeaenol led to significantly greater suppression of CoNV in a mouse model compared to the free form of 5Z-7-oxozeaenol, likely due to extended retention of 5Z-7-oxozeaenol in the cornea. Conclusion: Our study shows the potential of TAK1 as a therapeutic target for pathological angiogenesis, and the gelatin nanoparticle coupled with 5Z-7-oxozeaenol as a promising new eyedrop administration model in treatment of CoNV.

Acute systemic inhibition of inflammation augments endothelium-dependent dilation in women with a history of preeclamptic pregnancy.

Women who have had preeclampsia demonstrate microvascular endothelial-dysfunction, mediated in part by reduced nitric oxide (NO)-dependent dilation. Preeclamptic pregnancies are associated with elevated inflammation, and inhibition of inflammation attenuates endothelial damage in animal models of preeclampsia. However, it is unclear if inhibition of vascular inflammation improves endothelial function in women after a preeclamptic pregnancy. Using the cutaneous microcirculation as a model, we hypothesized that acute systemic inhibition of vascular inflammation (oral salsalate; 1500 mg/twice daily, 4 days) would improve endothelium- and NO-dependent vasodilation in women with a history of preeclampsia (PE) but not in women with a history of uncomplicated pregnancy (HC). Twelve HC (30 ± 1yrs, 10 ± 2 months postpartum) and 10 PE (30 ± 2yrs, 8 ± 2 months postpartum) participated in a double-blind placebo-controlled study. Following each treatment, 2 intradermal microdialysis fibers were placed in the skin of the ventral forearm for graded infusion of acetylcholine (Ach, 10-7-102mM) or Ach + 15 mM L-NAME (NO synthase antagonist). Red blood cell flux was measured over each site by laser-Doppler flowmetry (LDF). Cutaneous vascular conductance was calculated (CVC = LDF/mean arterial pressure) and normalized to maximum (%CVCmax; 28 mM SNP + local heat 43 °C). ACh-induced (77 ± 3 vs. 92 ± 3%CVCmax; p = 0.01) and NO-dependent (20 ± 6 vs. 33 ± 4%; p = 0.02) vasodilation were attenuated in PE compared to HC. Salsalate augmented ACh-induced (95 ± 2%CVCmax; p = 0.002) and NO-dependent (39 ± 3%; p = 0.009) dilation in PE compared to placebo but had no effect in HC (all p > 0.05). Salsalate treatment augmented endothelium-dependent vasodilation via NO-mediated pathways in women who have had preeclampsia, suggesting that inflammatory signaling mediates persistent endothelial dysfunction following preeclampsia.

Changes in glycemic variability, gastric emptying and vascular endothelial function after switching from twice-daily to once-weekly exenatide in patients with type 2 diabetes: a subpopulation analysis of the twin-exenatide study.

BACKGROUND: We investigated the changes in blood glucose fluctuation, gastric emptying, and vascular endothelial function by switching from an exenatide twice-daily formulation (BID) to a once-weekly formulation (QW) since the evaluation of postprandial glucose excursion and glycemic variability (GV) by continuous glucose monitoring (CGM) after switching was lacking. METHODS: Twenty-nine patients with type 2 diabetes treated with exenatide BID were included in this study and switched to exenatide QW for 24 weeks. GV assessed by CGM, gastric emptying (by 13 C-acetate breath test) and vascular endothelial function (by reactive hyperemia - peripheral arterial tonometry) were evaluated at baseline and 24 weeks after switching. RESULTS: HbA1c decreased significantly from the baseline to week 24, while postprandial glucose levels after breakfast and dinner significantly increased (both P <0.05). However, the increases in GV indices were modest and not statistically significant at week 24. Vascular endothelial function was also not significantly changed after switching (P >0.05). Gastric emptying was significantly accelerated at week 24 (Tmax 83.4 ± 12.1 min vs. 58.2 ± 16.4 min) (P <0.001) and correlated with increased postprandial glucose levels after breakfast and dinner (both P <0.05). CONCLUSIONS: Despite the increase in postprandial glucose associated with accelerated gastric emptying after switching from exenatide BID to QW, change in GV was modest and no significant deterioration in vascular endothelial function was observed after switching. These results support the superiority of treatment with exenatide QW over exenatide BID in clinical practice; however, attention should be paid to the monitoring and management of postprandial glucose levels when selecting exenatide QW. TRIAL REGISTRATION: Clinical trial registry number; UMIN000016390 and jRCTs031180320 . Approval date of Registry and the Registration: December 12, 2014.

Vascular endotheliitis associated with infections: Its pathogenetic role and therapeutic implication.

Vascular endothelial cells are major participants in and regulators of immune responses and inflammation. Vascular endotheliitis is regarded as a host immune-inflammatory response of the endothelium forming the inner surface of blood vessels in association with a direct consequence of infectious pathogen invasion. Vascular endotheliitis and consequent endothelial dysfunction can be a principle determinant of microvascular failure, which would favor impaired perfusion, tissue hypoxia, and subsequent organ failure. Emerging evidence suggests the role of vascular endotheliitis in the pathogenesis of coronavirus disease 2019 (COVID-19) and its related complications. Thus, once initiated, vascular endotheliitis and resultant cytokine storm cause systemic hyperinflammation and a thrombotic phenomenon in COVID-19, leading to acute respiratory distress syndrome and widespread organ damage. Vascular endotheliitis also appears to be a contributory factor to vasculopathy and coagulopathy in sepsis that is defined as life-threatening organ dysfunction due to a dysregulated response of the host to infection. Therefore, protecting endothelial cells and reversing vascular endotheliitis may be a leading therapeutic goal for these diseases associated with vascular endotheliitis. In this review, we outline the etiological and pathogenic importance of vascular endotheliitis in infection-related inflammatory diseases, including COVID-19, and possible mechanisms leading to vascular endotheliitis. We also discuss pharmacological agents which may be now considered as potential endotheliitis-based treatment modalities for those diseases.