Catalpol relieved angiotensin II-induced blood-brain barrier destruction via inhibiting the TLR4 pathway in brain endothelial cells.

CONTEXT: Catalpol is a major bioactive constituent of Rehmannia glutinosa Libosch (Scrophulariaceae), a traditional Chinese medicine, which is widely used in multiple diseases, including hypertension. OBJECTIVES: To explore whether catalpol protects against angiotensin II (Ang II)-triggered blood-brain barrier (BBB) leakage. MATERIALS AND METHODS: The bEnd.3 cells and BBB models were pre-treated with or without catalpol (50, 200 and 500 µM) or TAK-242 (1 µM) for 2 h and then with Ang II (0.1 µM) or LPS (1 µg/mL) for 24 h. Cell viability was determined by the MTT assay. The levels of Toll-like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88), inducible nitric oxide synthase (iNOS), tumour necrosis factor-α (TNF-α), caveolin-1 (Cav-1) and p-eNOS/eNOS were tested by western blot. The BBB permeability was evaluated by the flux of bovine serum albumin-fluorescein isothiocyanate (BSA-FITC) across monolayers. nuclear factor kappa-B (NF-κB) p65 nuclear translocation was explored by immunofluorescence staining. RESULTS: Ang II (0.1 µM) decreased the cell viability to 86.52 ± 1.79%, elevated the levels of TLR4, MyD88, iNOS, TNF-α and Cav-1 respectively to 3.7-, 1.5-, 2.3-, 2.2- and 2.7-fold, reduced the level of p-eNOS/eNOS to 1.6-fold in bEnd.3 cells, and eventually increased BBB permeability. Catalpol dose-dependently reversed these changes at 50-500 µM. Meanwhile, catalpol (500 µM) inhibited the upregulated levels of TLR4 pathway-related proteins and NF-κB p65 nuclear translocation, decreased the enhanced transcytosis, and relieved the BBB disruption caused by both LPS (the TLR4 activator) and Ang II. The effects are same as TAK-242 (the TLR4 inhibitor). CONCLUSIONS: Catalpol relieved the Ang II-induced BBB damage, which indicated catalpol has high potential for the treatment of hypertension-induced cerebral small vessel disease (cSVD).

Qingda granule alleviate angiotensin â ±-induced hypertensive renal injury by suppressing oxidative stress and inflammation through NOX1 and NF-κB pathways.

Hypertension has become one of the important diseases harmful to human health. In China, Qingda granule (QDG) has been used to treat hypertension for decades. Previous studies by our team have shown that oxidative stress may be one of the pathways through which QDG inhibits hypertension-induced organs injury. However, the specific molecular mechanism of its anti-hypotension and renal oxidative stress response were unclearly. This study investigated QDG's potential protective mechanism against hypertension-induced renal injury. Mice were infused with Angiotensin Ⅱ (Ang Ⅱ, 500 ng/kg/min) or equivalent saline solution (Control) and administered oral QDG (1.145 g/kg/day) or saline for four weeks. QDG treatment mitigated the elevated blood pressure and reduced renal pathological changes induced by Ang Ⅱ. As per the RNA sequencing results, QDG affects oxidative stress signaling. In agreement with these findings, QDG significantly attenuated the Ang Ⅱ-induced increase in Nitrogen oxides 1 (NOX1) and reactive oxygen species and the decrease in superoxide dismutase in renal tissue. Additionally, QDG significantly inhibited Interleukin 6 (IL-6), Tumor necrosis factor α (TNF-α), and Interleukin 1ß (IL-1ß) expression in renal tissues and blocked the phosphorylation of P65 (NF-κB subunit) and IκB. These results were confirmed in vitro. Overall, QDG reduced Ang Ⅱ-induced elevated blood pressure and renal injury by inhibiting oxidative stress and inflammation caused by NOX1 and NF-κB pathways. The results of this study provide an experimental basis for the clinical application of QDG, and to open up a new direction for the clinical treatment of hypertension.

The protective effect of harpagoside on angiotensin II (Ang II)-induced blood-brain barrier leakage in vitro.

Hypertension and its associated dysfunction of the blood-brain barrier (BBB) contribute to cerebral small vessel disease (cSVD). Angiotensin II (Ang II), a vasoactive peptide of the renin-angiotensin system (RAS), is not only a pivotal molecular signal in hypertension but also causes BBB leakage, cSVD, and cognitive impair. Harpagoside, the major bioactive constituent of Scrophulariae Radix, has been commonly used for the treatment of multiple diseases including hypertension in China. The effect of harpagoside on Ang II-induced BBB damage is unclear. We employed an immortalized endothelial cell line (bEnd.3) to mimic a BBB monolayer model in vitro and investigated the effect of harpagoside on BBB and found that harpagoside alleviated Ang II-induced BBB destruction, inhibited Ang II-associated cytotoxicity in a concentration-dependent manner and attenuated Ang II-induced reactive oxygen species (ROS) impair by downregulation of Nox2, Nox4, and COX-2. Harpagoside prevented Ang II-induced apoptosis via keeping Bax/Bcl-2 balance, decreasing cytochrome c release, and inactivation of caspase-8, caspase-9, and caspase-3 (the mitochondria-dependent and death receptor-mediated apoptosis pathways). Moreover, harpagoside can alleviate Ang II-induced BBB damage through upregulation of tight junction proteins and decrease of caveolae-mediated endocytosis. Thus, harpagoside might be a potential drug to treat Ang II-induced cSVD.

Quyu Shengxin capsule (QSC) inhibits Ang-II-induced abnormal proliferation of VSMCs by down-regulating TGF-ß, VEGF, mTOR and JAK-STAT pathways.

ETHNOPHARMACOLOGICAL RELEVANCE: Quyu Shengxin capsule (QSC) is an herbal compound commonly used to treat blood stasis syndrome in China, and blood stasis syndrome is considered to be the root of cardiovascular diseases (CVD) in traditional Chinese medicine. However, the potential molecular mechanism of QSC is still unknown. AIM OF STUDY: To study the therapeutic effect of QSC on the abnormal proliferation of VSMCs induced by Ang-II, and to explore its possible mechanism of action. MATERIALS AND METHODS: Qualitative analysis and quality control of QSC through UPLC-MS/MS and UPLC. The rat thoracic aorta vascular smooth muscle cells (VSMCs) were cultured in vitro, and then stimulated with Angiotensin Ⅱ (Ang-II) (10-7 mol/L) for 24 h to establish a cardiovascular cell model. The cells were then treated with different concentrations of QSC drug-containing serum or normal goat serum. MTT assay was used to detect the viability of VSMCs and abnormal cell proliferation. In order to analyze the possible signal transduction pathways, the content of various factors in the supernatant of VSMCs was screened and determined by means of the Luminex liquid suspension chip detection platform, and the phosphoprotein profile in VSMCs was screened by Phospho Explorer antibody array. RESULTS: Compared with the model group, serum cell viability and inflammatory factor levels with QSC were significantly decreased (P < 0.001). In addition, the expression levels of TGF-ß, VEGF, mTOR and JAK-STAT in the QSC-containing serum treatment group were significantly lower than those in the model group. QSC may regulate the pathological process of CVD by reducing the levels of inflammatory mediators and cytokines, and protecting VSMCs from the abnormal proliferation induced by Ang-II. CONCLUSION: QSC inhibits Ang-II-induced abnormal proliferation of VSMCs, which is related to the down-regulation of TGF-ß, VEGF, mTOR and JAK-STAT pathways.

Qiliqiangxin alleviates Ang II-induced CMECs apoptosis by downregulating autophagy via the ErbB2-AKT-FoxO3a axis.

Our previous work revealed the protective effect of Qiliqiangxin (QLQX) on cardiac microvascular endothelial cells (CMECs), but the underlying mechanisms remain unclear. We aimed to investigate whether QLQX exerts its protective effect against high-concentration angiotensin II (Ang II)-induced CMEC apoptosis through the autophagy machinery. CMECs were cultured in high-concentration Ang II (1 µM) medium in the presence or absence of QLQX for 48 h. We found that QLQX obviously inhibited Ang II-triggered autophagosome synthesis and apoptosis in cultured CMECs. QLQX-mediated protection against Ang II-induced CMEC apoptosis was reversed by the autophagy activator rapamycin. Specifically, deletion of ATG7 in cultured CMECs indicated a detrimental role of autophagy in Ang II-induced CMEC apoptosis. QLQX reversed Ang II-mediated ErbB2 phosphorylation impairment. Furthermore, inhibition of ErbB2 phosphorylation with lapatinib in CMECs revealed that QLQX-induced downregulation of Ang II-activated autophagy and apoptosis was ErbB2 phosphorylation-dependent via the AKT-FoxO3a axis. Activation of ErbB2 phosphorylation by Neuregulin-1ß achieved a similar CMEC-protective effect as QLQX in high-concentration Ang II medium, and this effect was also abolished by autophagy activation. These results show that the CMEC-protective effect of QLQX under high-concentration Ang II conditions could be partly attributable to QLQX-mediated ErbB2 phosphorylation-dependent downregulation of autophagy via the AKT-FoxO3a axis.

Melatonin alleviates angiotensin-II-induced cardiac hypertrophy via activating MICU1 pathway.

Mitochondrial calcium uptake 1 (MICU1) is a pivotal molecule in maintaining mitochondrial homeostasis under stress conditions. However, it is unclear whether MICU1 attenuates mitochondrial stress in angiotensin II (Ang-II)-induced cardiac hypertrophy or if it has a role in the function of melatonin. Here, small-interfering RNAs against MICU1 or adenovirus-based plasmids encoding MICU1 were delivered into left ventricles of mice or incubated with neonatal murine ventricular myocytes (NMVMs) for 48 h. MICU1 expression was depressed in hypertrophic myocardia and MICU1 knockdown aggravated Ang-II-induced cardiac hypertrophy in vivo and in vitro. In contrast, MICU1 upregulation decreased cardiomyocyte susceptibility to hypertrophic stress. Ang-II administration, particularly in NMVMs with MICU1 knockdown, led to significantly increased reactive oxygen species (ROS) overload, altered mitochondrial morphology, and suppressed mitochondrial function, all of which were reversed by MICU1 supplementation. Moreover, peroxisome proliferator-activated receptor gamma coactivator 1-α (PGC-1α)/MICU1 expression in hypertrophic myocardia increased with melatonin. Melatonin ameliorated excessive ROS generation, promoted mitochondrial function, and attenuated cardiac hypertrophy in control but not MICU1 knockdown NMVMs or mice. Collectively, our results demonstrate that MICU1 attenuates Ang-II-induced cardiac hypertrophy by inhibiting mitochondria-derived oxidative stress. MICU1 activation may be the mechanism underlying melatonin-induced protection against myocardial hypertrophy.

Functional interactions between complex I and complex II with nNOS in regulating cardiac mitochondrial activity in sham and hypertensive rat hearts.

Nitric oxide (NO) affects mitochondrial activity through its interactions with complexes. Here, we investigated regulations of complex I (C-I) and complex II (C-II) by neuronal NO synthase (nNOS) in the presence of fatty acid supplementation and the impact on left ventricular (LV) mitochondrial activity from sham and angiotensin II (Ang-II)-induced hypertensive (HTN) rats. Our results showed that nNOS protein was expressed in sham and HTN LV mitochondrial enriched fraction. In sham, oxygen consumption rate (OCR) and intracellular ATP were increased by palmitic acid (PA) or palmitoyl-carnitine (PC). nNOS inhibitor, S-methyl-l-thiocitrulline (SMTC), did not affect OCR or cellular ATP increment by PA or PC. However, SMTC increased OCR with PA + malonate (a C-II inhibitor), but not with PA + rotenone (a C-I inhibitor), indicating that nNOS attenuates C-I with fatty acid supplementation. Indeed, SMTC increased C-I activity but not that of C-II. Conversely, nNOS-derived NO was increased by rotenone + PA in LV myocytes. In HTN, PC increased the activity of C-I but reduced that of C-II, consequently OCR was reduced. SMTC increased both C-I and C-II activities with PC, resulted in OCR enhancement in the mitochondria. Notably, SMTC increased OCR only with rotenone, suggesting that nNOS modulates C-II-mediated OCR in HTN. nNOS-derived NO was partially reduced by malonate + PA. Taken together, nNOS attenuates C-I-mediated mitochondrial OCR in the presence of fatty acid in sham and C-I modulates nNOS activity. In HTN, nNOS attenuates C-I and C-II activities whereas interactions between nNOS and C-II maintain mitochondrial activity.

Cyclodextrin Prevents Abdominal Aortic Aneurysm via Activation of Vascular Smooth Muscle Cell Transcription Factor EB.

BACKGROUND: Abdominal aortic aneurysm (AAA) is a severe aortic disease with a high mortality rate in the event of rupture. Pharmacological therapy is needed to inhibit AAA expansion and prevent aneurysm rupture. Transcription factor EB (TFEB), a master regulator of autophagy and lysosome biogenesis, is critical to maintain cell homeostasis. In this study, we aim to investigate the role of vascular smooth muscle cell (VSMC) TFEB in the development of AAA and establish TFEB as a novel target to treat AAA. METHODS: The expression of TFEB was measured in human and mouse aortic aneurysm samples. We used loss/gain-of-function approaches to understand the role of TFEB in VSMC survival and explored the underlying mechanisms through transcriptome and functional studies. Using VSMC-selective Tfeb knockout mice and different mouse AAA models, we determined the role of VSMC TFEB and a TFEB activator in AAA in vivo. RESULTS: We found that TFEB is downregulated in both human and mouse aortic aneurysm lesions. TFEB potently inhibits apoptosis in VSMCs, and transcriptome analysis revealed that TFEB regulates apoptotic signaling pathways, especially apoptosis inhibitor B-cell lymphoma 2. B-cell lymphoma 2 is significantly upregulated by TFEB and is required for TFEB to inhibit VSMC apoptosis. We consistently observed that TFEB deficiency increases VSMC apoptosis and promotes AAA formation in different mouse AAA models. Furthermore, we demonstrated that 2-hydroxypropyl-ß-cyclodextrin, a clinical agent used to enhance the solubility of drugs, activates TFEB and inhibits AAA formation and progression in mice. Last, we found that 2-hydroxypropyl-ß-cyclodextrin inhibits AAA in a VSMC TFEB-dependent manner in mouse models. CONCLUSIONS: Our study demonstrated that TFEB protects against VSMC apoptosis and AAA. TFEB activation by 2-hydroxypropyl-ß-cyclodextrin may be a promising therapeutic strategy for the prevention and treatment of AAA.

α-Linolenic acid but not linolenic acid protects against hypertension: critical role of SIRT3 and autophagic flux.

Although dietary α-linolenic acid (ALA) or linolenic acid (LA) intake was reported to be epidemiologically associated with a lower prevalence of hypertension, recent clinical trials have yielded conflicting results. Comparable experimental evidence for the roles of these two different fatty acids is still lacking and the underlying mechanisms need to be further elucidated. Our data showed that ALA but not LA supplementation alleviated systolic blood pressure elevation and improved ACh-induced, endothelium-dependent vasodilation in both spontaneously hypertensive rats (SHRs) and AngII-induced hypertensive mice. In addition, SHRs displayed reduced vascular Sirtuin 3 (SIRT3) expression, subsequent superoxide dismutase 2 (SOD2) hyperacetylation and mitochondrial ROS overproduction, all of which were ameliorated by ALA but not LA supplementation. In primary cultured endothelial cells, ALA treatment directly inhibited SIRT3 reduction, SOD2 hyperacetylation, mitochondrial ROS overproduction and alleviated autophagic flux impairment induced by AngII plus TNFα treatment. However, these beneficial effects of ALA were completely blocked by silencing SIRT3. Restoration of autophagic flux by rapamycin also inhibited mitochondrial ROS overproduction in endothelial cells exposed to AngII plus TNFα. More interestingly, SIRT3 KO mice developed severe hypertension in response to a low dose of AngII infusion, while ALA supplementation lost its anti-hypertensive and endothelium-protective effects on these mice. Our findings suggest that ALA but not LA supplementation improves endothelial dysfunction and diminishes experimental hypertension by rescuing SIRT3 impairment to restore autophagic flux and mitochondrial redox balance in endothelial cells.

COA-Cl prevented TGF-ß1-induced CTGF expression by Akt dephosphorylation in normal human dermal fibroblasts, and it attenuated skin fibrosis in mice models of systemic sclerosis.

BACKGROUND: Systemic sclerosis (SSc) is characterized by fibrosis of the skin and internal organs. Although transforming growth factor (TGF)-ß1-induced connective tissue growth factor (CTGF/CCN2) expression has been presented in SSc fibrosis, the therapeutic potential of targeting CTGF in SSc has not been fully explored. COA-Cl is a novel nucleic acid analog, which is reported to have pleiotropic beneficial biologic effects. OBJECTIVE: We examine the effects of COA-Cl on TGF-ß1-induced CTGF expression in normal human dermal fibroblast (NHDF). We also examined the effects of COA-Cl on CTGF expression in a mouse SSc model of angiotensin II (Ang II)-induced skin fibrosis. METHODS: NHDF was cultured for in vitro experiments. For in vivo experiments, C57BL/6J mice were treated with Ang II for 14 days by subcutaneous osmotic pump. Quantitative real-time polymerase chain reaction, western blot analysis, immunohistochemical staining and immunofluorescence staining were performed to examine the expression levels of CTGF and phosphorylation levels of Smad2/3, ERK1/2 and Akt. RESULTS: COA-Cl attenuated the TGF-ß1-induced expression of both CTGF mRNA and protein in NHDF. Although COA-Cl did not alter the TGF-ß1-induced phosphorylation of Smad2/3 or ERK1/2, it reduced the TGF-ß1-induced phosphorylation levels of Akt in NHDF. Notably, COA-Cl dephosphorylated the Akt of lysates of TGF-ß1-treated NHDF. COA-Cl reduced the levels of CTGF mRNA, CTGF protein, dermal thickness, collagen content and Akt phosphorylation in the skin of mice SSc model. CONCLUSION: These results imply that the inhibition of TGF-ß1-induced CTGF expression by COA-Cl may be a therapeutic approach for SSc.

Dietary Nitrate Reduces Blood Pressure in Rats With Angiotensin II-Induced Hypertension via Mechanisms That Involve Reduction of Sympathetic Hyperactivity.

Several experimental and clinical studies have shown that dietary nitrate supplementation can increase nitric oxide bioavailability. In the oral cavity, commensal bacteria reduce nitrate to nitrite, which is subsequently absorbed into the circulation where reduction to nitric oxide by enzymatic systems occur. Although it is well-known that boosting the nitrate-nitrite-nitric oxide pathway can improve cardiovascular, renal, and metabolic functions and that sympathoexcitation contributes to the development of the same disorders, the potential effects of dietary nitrate on sympathetic activity remain to be elucidated. In this study, we hypothesized that treatment with inorganic nitrate could prevent the increase in sympathetic nerve activity in an experimental model of Ang II (angiotensin II)-induced hypertension. Multiple in vivo approaches were combined, that is, Wistar rats orally treated with the nitric oxide synthase inhibitor L-NAME (N(G)-nitro-L-arginine methyl ester, 0.5 g/L) and implanted with subcutaneous osmotic minipump for continuous delivery of Ang II (120 ng/kg per minute; 14 days). Simultaneously, rats were supplemented with sodium nitrate (10 mmol/L) or placebo (sodium chloride; 10 mmol/L) in the drinking water. Blood pressure, heart rate, and renal sympathetic nerve activity were recorded. In placebo-treated rats, Ang II+L-NAME treatment-induced arterial hypertension, which was linked with reduced spontaneous baroreflex sensitivity and increased renal sympathetic nerve activity, as well as upregulation of AT1Rs (Ang II type-1 receptors) in the rostral ventrolateral medulla. Supplementation with nitrate normalized the expression of AT1Rs in rostral ventrolateral medulla and reduced sympathetic nerve activity, which was associated with attenuated development of hypertension. In conclusion, chronic dietary nitrate supplementation blunted the development of hypertension via mechanisms that involve reduction of sympathetic outflow.

Acute and Chronic Antihypertensive Effect of Fractions, Tiliroside and Scopoletin from Malva parviflora.

Hypertension is a disease of high prevalence and morbidity where vascular inflammation and associated oxidative stress (endothelial dysfunction) is the underlying cause of this pathology. We are reporting the antihypertensive activity of extracts and fractions of Malva parviflora in mice with chronic and acute hypertension. Also, the treatments of this plant were able to counteract the kidney inflammation and associated oxidative stress. The chronic hypertension model consisted of administration of angiotensin II (AGII) during 12 weeks, causing a sustained increase in systolic (SBP) or diastolic (DBP) pressure, with values of pharmacological constants of: ED50 = 0.038 mg/kg y Emax = 135 mmHg for SBP and ED50 = 0.046 mg/kg y Emax = 98 mmHg for DBP. The chronic hypertension caused the inflammation and lipid peroxidation in kidneys, measured by of tissue level of cytokines such as interleukin-1ß (IL-1ß), IL-6, Tumor Necrosis Factor-α (TNF-α), IL-10 and malondialdehyde, and treatments for M. parviflora were able to modulate these parameters. The chemical fractionation allowed to identify three compounds: oleanolic acid, tiliroside and scopoletin, which were tested in a model of acute hypertension. The pharmacodynamic parameters for SBP were ED50 = 0.01 and 0.12 mg/kg while Emax = 33.22 and 37.74 mmHg for scopoletin and tiliroside, respectively; whereas that for DBP data were ED50 = 0.01 and 0.02 mg/kg; with an Emax = 7.00 and 6.24 mmHg, in the same order. M. parviflora, is able to counteract the effect of chronic and acute administration of AGII, on hypertension, but also the inflammatory and oxidative damage in the kidney. The oleanolic acid, scopoletin and tiliroside are the compounds responsible for such activities.

The standardized extract of Nigella sativa and its major ingredient, thymoquinone, ameliorates angiotensin II-induced hypertension in rats.

Background This study investigated the effect of hydroalcoholic extract of Nigella sativa (N. sativa) and its active component, thymoquinone (TQ) on hypertension induced by angiotensin II (AngII), the main product of renin-angiotensin system (RAS). Methods Seven animal groups (n=7 for each group) were used as follows: (1) control, (2) AngII (300 ng/kg), (3) AngII+losartan (Los; 10 mg/kg), (4) TQ (40 mg/kg)+AngII, and (5-7) three doses of N. sativa (200, 400, and 600 mg/kg)+AngII. Los and AngII were injected intravenously; TQ and extracts were injected intraperitoneally. In TQ and N. sativa-treated groups, 30 min after injection of the extract and TQ, AngII was injected. Cardiovascular parameters were recorded by power lab system after cannulation of femoral artery. The maximum changes (∆) of systolic blood pressure (SBP), mean arterial pressure (MAP), and heart rate (HR) were calculated and used for statistical analysis. Results AngII significantly increased maximal ∆SBP, ∆MAP, and ∆HR compared with the control (p<0.001), and these effects significantly were blunted by Los. TQ and two higher doses (400 and 600 mg/kg) of N. sativa significantly could antagonize effect of AngII on ∆SBP, ∆MAP (p<0.05 to p<0.001). AngII-induced changes of HR are also significantly decreased by TQ and dose 600 mg/kg of extract (p<0.01 and p<0.05, respectively). Conclusions The N. sativa and its component TQ have the beneficial effect on hypertension probably due to attenuation cardiovascular effects of AngII.

Cucumis sativus Aqueous Fraction Inhibits Angiotensin II-Induced Inflammation and Oxidative Stress In Vitro.

Inflammation and oxidative stress play major roles in endothelial dysfunction, and are key factors in the progression of cardiovascular diseases. The aim of this study was to evaluate in vitro the effect of three subfractions (SFs) from the Cucumis sativus aqueous fraction to reduce inflammatory factors and oxidative stress induced by angiotensin II (Ang II) in human microvascular endothelial cells-1 (HMEC-1) cells. The cells were cultured with different concentrations of Ang II and 0.08 or 10 µg/mL of SF1, SF2, or SF3, or 10 µmol of losartan as a control. IL-6 (Interleukin 6) concentration was quantified. To identify the most effective SF combinations, HMEC-1 cells were cultured as described above in the presence of four combinations of SF1 and SF3. Then, the effects of the most effective combination on the expression of adhesion molecules, the production of reactive oxygen species (ROS), and the bioavailability of nitric oxide (NO) were evaluated. Finally, a mass spectrometry analysis was performed. Both SF1 and SF3 subfractions decreased the induction of IL-6 by Ang II, and C4 (SF1 and SF3, 10 µg/mL each) was the most effective combination to inhibit the production of IL-6. Additionally, C4 prevented the expression of adhesion molecules, reduced the production of ROS, and increased the bioavailability of NO. Glycine, arginine, asparagine, lysine, and aspartic acid were the main components of both subfractions. These results demonstrate that C4 has anti-inflammatory and antioxidant effects.

Treatment with salvianolic acid B restores endothelial function in angiotensin II-induced hypertensive mice.

Salvianolic acid B (Sal B) is one of the most abundant phenolic acids derived from the root of Danshen with potent anti-oxidative properties. The present study examined the vasoprotective effect of Sal B in hypertensive mice induced by angiotensin II (Ang II). Sal B (25mg/kg/day) was administered via oral gavage for 11days to Ang II (1.2mg/kg/day)-infused C57BL/6J mice (8-10weeks old). The vascular reactivity (both endothelium-dependent relaxations and contractions) in mouse arteries was examined by wire myography. The production of reactive oxygen species (ROS), protein level and localization of angiotensin AT1 receptors and the proteins involved in ROS formation were evaluated using dihydroethidium (DHE) fluorescence, lucigenin-enhanced chemiluminescence, immunohistochemistry and Western blotting, respectively. The changes of ROS generating proteins were also assessed in vitro in human umbilical vein endothelial cells (HUVECs) exposed to Ang II with and without co-treatment with Sal B (0.1-10nM). Oral administration of Sal B reversed the Ang II-induced elevation of arterial systolic blood pressure in mice, augmented the impaired endothelium-dependent relaxations and attenuated the exaggerated endothelium-dependent contractions in both aortas and renal arteries of Ang II-infused mice. In addition, Sal B treatment normalized the elevated levels of AT1 receptors, NADPH oxidase subunits (NOx-2 and NOx-4) and nitrotyrosine in arteries of Ang II-infused mice or in Ang II-treated HUVECs. In summary, the present study provided additional evidence demonstrating that Sal B treatment for 11days reverses the impaired endothelial function and with a marked inhibition of AT1 receptor-dependent vascular oxidative stress. This vasoprotective and anti-oxidative action of Sal B most likely contributes to the anti-hypertensive action of the plant-derived compound.

Sinomenine Attenuates Angiotensin II-Induced Autophagy via Inhibition of P47-Phox Translocation to the Membrane and Influences Reactive Oxygen Species Generation in Podocytes.

BACKGROUND/AIMS: Sinomenine, a pure alkaloid extracted from the Chinese medicinal plant Sinomenium acutum, and sinomenine hydrochloride (SN) has been successfully used for the therapy of rheumatoid arthritis (RA) and kidney diseases. Autophagy is a cytoprotective mechanism used by podocytes and other cells to alleviate the effects of oxidative stress, and angiotensin II (Ang II) significantly promotes podocyte autophagy. However, excessive autophagy may lead to cell death and podocyte depletion. The present study evaluated the effect of SN in podocytes induced by Ang II. METHODS: Podocytes were pretreated with graded concentrations (10(-8) M ∼ 10(-4) M) of SN and then stimulated with Ang II. The LC3B protein and the p47-phox membrane fraction were measured by Western blot. Autolysosomes were assessed by transmission electron microscopy. FACS was used to quantify the ROS produced by podocytes. The translocation of p47-phox to the membrane was investigated by immunofluorescence. RESULTS: The 10(-8) M ∼ 10(-4) M of SN alone did not effect ROS generation or podocyte autophagy. The 10(-8) M and 10(-6) M SN attenuated Ang II-induced autophagy in podocytes. Furthermore, SN decreased the level of ROS generation in Ang II-induced podocytes via inhibition of NOX subunit p47-phox translocation to the membrane. CONCLUSION: The appropriate concentration of SN attenuated Ang II-induced podocyte autophagy through ROS generation, at least in part, by regulating NOX subunit p47-phox translocation to the membrane.

Dipeptidyl peptidase-4 inhibitor decreases abdominal aortic aneurysm formation through GLP-1-dependent monocytic activity in mice.

Abdominal aortic aneurysm (AAA) is a life-threatening situation affecting almost 10% of elders. There has been no effective medication for AAA other than surgical intervention. Dipeptidyl peptidase-4 (DPP-4) inhibitors have been shown to have a protective effect on cardiovascular disease. Whether DPP-4 inhibitors may be beneficial in the treatment of AAA is unclear. We investigated the effects of DPP-4 inhibitor sitagliptin on the angiotensin II (Ang II)-infused AAA formation in apoE-deficient (apoE-/-) mice. Mice with induced AAA were treated with placebo or 2.5, 5 or 10 mg/kg/day sitagliptin. Ang II-infused apoE-/- mice exhibited a 55.6% incidence of AAA formation, but treatment with sitagliptin decreased AAA formation. Specifically, administered sitagliptin in Ang II-infused mice exhibited decreased expansion of the suprarenal aorta, reduced elastin lamina degradation of the aorta, and diminished vascular inflammation by macrophage infiltration. Treatment with sitagliptin decreased gelatinolytic activity and apoptotic cells in aorta tissues. Sitaglipitn, additionally, was associated with increased levels of plasma active glucagon-like peptide-1 (GLP-1). In vitro studies, GLP-1 decreased reactive oxygen species (ROS) production, cell migration, and MMP-2 as well as MMP-9 activity in Ang II-stimulated monocytic cells. The results conclude that oral administration of sitagliptin can prevent abdominal aortic aneurysm formation in Ang II-infused apoE-/-mice, at least in part, by increasing of GLP-1 activity, decreasing MMP-2 and MMP-9 production from macrophage infiltration. The results indicate that sitagliptin may have therapeutic potential in preventing the development of AAA.

Amelioration of Atherosclerosis by the New Medicinal Mushroom Grifola gargal Singer.

The beneficial effects of edible mushrooms for improving chronic intractable diseases have been documented. However, the antiatherogenic activity of the new medicinal mushroom Grifola gargal is unknown. Therefore, we evaluated whether Grifola gargal can prevent or delay the progression of atherosclerosis. Atherosclerosis was induced in ApoE lipoprotein-deficient mice by subcutaneous infusion of angiotensin II. Grifola gargal extract (GGE) was prepared and intraperitoneally injected. The weight of heart and vessels, dilatation/atheroma formation of thoracic and abdominal aorta, the percentage of peripheral granulocytes, and the blood concentration of MCP-1/CCL2 were significantly reduced in mice treated with GGE compared to untreated mice. By contrast, the percentage of regulatory T cells and the plasma concentration of SDF-1/CXCL12 were significantly increased in mice treated with the mushroom extract compared to untreated mice. In vitro, GGE significantly increased the secretion of SDF-1/CXCL12, VEGF, and TGF-ß1 from fibroblasts compared to control. This study demonstrated for the first time that Grifola gargal therapy can enhance regulatory T cells and ameliorate atherosclerosis in mice.

High-fat diet- and angiotensin II-induced aneurysm concurrently elicits splenic hypertrophy.

BACKGROUND: Angiotensin II (Ang II) and high-fat diet are implicated in causing pathological changes in the vascular endothelium, brain, kidney and liver. The association of aneurysm leading to histopathological changes in the splenic compartment remains elusive. Further, the salubrious credentials of antioxidants, especially α-tocopherol and ß-carotene in the resolution of splenic pathology have not been investigated. METHODS: Four-month-old Apoe(-/-) mice were used in the induction of aneurysm by infusing Ang II, and subsequently were orally administered with α-tocopherol and ß-carotene-enriched diet for 60 days. RESULTS: We observed splenomegaly in Ang II-infused aneurysm and high-fat diet-supplemented mice as compared to normal mice. These observations were further confirmed through histopathological investigations, demonstrating splenic follicular hypertrophy. We observed a remarkable decrease in the size of spleen in α-tocopherol and ß-carotene-treated Apoe(-/-) mice as compared with Ang II-treated animals. Furthermore, no marked changes in the histopathological splenic sections were seen in the ß-carotene-treated group. However, hyperplasia and proliferation of immature lymphocytes in the follicles were observed in the α-tocopherol-treated animals. We found that CD4+ T-cell levels were increased in the high-fat diet group relative to the control group and were decreased in the ß-carotene-treated animals. CONCLUSIONS: Our study provides evidence that Ang II infusion and high-fat supplementation induces abdominal aortic aneurysm that has pathological implications to the spleen. The use of ß-carotene but not α-tocopherol as an antioxidant markedly ameliorates the pathological changes in spleen.

Regulating effect of tea polyphenols on endothelin, intracellular calcium concentration, and mitochondrial membrane potential in vascular endothelial cells injured by Angiotensin II.

BACKGROUND: Polyphenols are the main active component of tea and are considered an antiatherosclerosis agent, protecting vascular endothelial cells (VECs) from injury and preventing cardiovascular diseases. Endothelin level, intracellular calcium concentration, and mitochondrial membrane potential in VECs directly reflect the function and injury status of cells. The objective of this study was to study the regulating effects of tea polyphenols on these factors in VECs injured by angiotensin II (Ang-II) and explore the protective effect of tea polyphenols on human VECs. METHODS: In this study, human aortic vascular endothelial cells were divided into 4 groups: (1) a control group; (2) Ang-II group: Ang-II at a concentration of 10(-7) mol/L was added to the cells; (3) low-concentration tea polyphenols + Ang-II group: tea polyphenols at a concentration of 5 mg/L was added in addition to Ang-II; (4) high-concentration tea polyphenols + Ang-II group: tea polyphenols at a concentration of 25 mg/L was added in addition to Ang-II. One hundred microliters of supernatant was extracted before treatment and at 0.5, 6, and 24 hours after treatment in each group to establish the content of endothelin. RESULTS: The results showed the following: (1) tea polyphenols decreased the expression of endothelin-1 messenger RNA, which was increased by Ang-II (P < 0.01). (2) Tea polyphenols inhibited endothelin secretion induced by Ang-II (P < 0.01), and the inhibition of low-concentration tea polyphenols was superior to that of high concentration tea polyphenols (P < 0.01). (3) Tea polyphenols ameliorated the changes in intracellular calcium concentration and mitochondrial membrane potential induced by Ang-II (P < 0.01). CONCLUSIONS: This study suggests that tea polyphenols may effectively protect VECs against injury by regulating endothelin, intracellular calcium concentration, and mitochondrial membrane potential in VECs injured by Ang-II. Additionally, lower dose would be used clinically rather than the higher dose for obtaining better results.