Activation of heme oxygenase-1 by Ginkgo biloba extract differentially modulates endothelial and smooth muscle-like progenitor cells for vascular repair.

Vascular progenitors such as endothelial progenitor cells (EPCs) and smooth muscle-like progenitor cells (SMPCs) may play different roles in vascular repair. Ginkgo biloba extract (GBE) is an exogenous activator of heme oxygenase (HO)-1, which has been suggested to improve vascular repair; however, the detailed mechanisms have yet to be elucidated. This study aimed to investigate whether GBE can modulate different vascular progenitor cells by activating HO-1 for vascular repair. A bone marrow transplantation mouse model was used to evaluate the in vivo effects of GBE treatment on wire-injury induced neointimal hyperplasia, which is representative of impaired vascular repair. On day 14 of GBE treatment, the mice were subjected to wire injury of the femoral artery to identify vascular reendothelialization. Compared to the mice without treatment, neointimal hyperplasia was reduced in the mice that received GBE treatment for 28 days in a dose-dependent manner. Furthermore, GBE treatment increased bone marrow-derived EPCs, accelerated endothelial recovery, and reduced the number of SMPCs attached to vascular injury sites. The effects of GBE treatment on neointimal hyperplasia could be abolished by co-treatment with zinc protoporphyrin IX, an HO-1 inhibitor, suggesting the in vivo role of HO-1. In this in vitro study, treatment with GBE activated human early and late EPCs and suppressed SMPC migration. These effects were abolished by HO-1 siRNA and an HO-1 inhibitor. Furthermore, GBE induced the expression of HO-1 by activating PI3K/Akt/eNOS signaling in human late EPCs and via p38 pathways in SMPCs, suggesting that GBE can induce HO-1 in vitro through different molecular mechanisms in different vascular progenitor cells. Accordingly, GBE could activate early and late EPCs, suppress the migration of SMPCs, and improve in vivo vascular repair after mechanical injury by activating HO-1, suggesting the potential role of pharmacological HO-1 activators, such as GBE, for vascular protection in atherosclerotic diseases.

Increased risk of ventricular tachycardia and cardiovascular death in patients with myocarditis during the long-term follow-up: A national representative cohort from the National Health Insurance Research Database.

The incidence of acute myocarditis complicated with ventricular tachycardia (VT) is unknown. This study aimed to investigate the association between myocarditis and the incidence of VT and mortality. We also aimed to determine the independent predictors that increased the VT risk in those patients. From 2000 to 2004, 13,250 patients with a history of myocarditis were identified from the Taiwan National Health Insurance Research Database. The same number of individuals without heart disease with a matched sex and underlying diseases were selected as the control group. The long-term risks of life-threatening ventricular arrhythmias and mortality in patients with a history of myocarditis were investigated by an adjusted Cox proportional hazards regression. After a mean follow-up of 10.4 ±â€Š2.94 years (interquartile range: 12, 10.19-12), the myocarditis patients showed a higher incidence of new onset VT events compared with healthy controls (5.4% [519 per 100,000 person-year] in the myocarditis group vs, 0.47% [43 per 100,000 person-year] in the healthy controls; adjusted hazard ratio [HR]: 16.1, 95% confidence interval [CI]: 12.4-20.9; P < .001). A higher incidence of cardiovascular death was noted in the myocarditis group than healthy controls (6.52% vs 3.18%; HR: 2.42, 95% CI: 2.14-2.73; P < .001) after adjusting for the multivariate confounders including sex, age, underlying comorbidities, and medications. The results of this study suggested that there was higher incidence of life-threatening VT and mortality during the very long-term follow-up in patients with a history of myocarditis. Future work should focus on an in-depth risk stratification of VT in myocarditis patients.

Effects of pitavastatin versus atorvastatin on the peripheral endothelial progenitor cells and vascular endothelial growth factor in high-risk patients: a pilot prospective, double-blind, randomized study.

BACKGROUND: Circulating endothelial progenitor cells (EPCs) reflect endothelial repair capacity and may be a significant marker for the clinical outcomes of cardiovascular disease. While some high-dose statin treatments may improve endothelial function, it is not known whether different statins may have similar effects on EPCs.This study aimed to investigate the potential class effects of different statin treatment including pitavastatin and atorvastatin on circulating EPCs in clinical setting. METHODS: A pilot prospective, double-blind, randomized study was conducted to evaluate the ordinary dose of pitavastatin (2 mg daily) or atorvastatin (10 mg daily) treatment for 12 weeks on circulating EPCs in patients with cardiovascular risk such as hypercholesterolemia and type 2 diabetes mellitus (T2DM). Additional in vitro study was conducted to clarify the direct effects of both statins on EPCs from the patients. RESULTS: A total of 26 patients (19 with T2DM) completed the study. While the lipid-lowering effects were similar in both treatments, the counts of circulating CD34+KDR+EPCs were significantly increased (from 0.021 ± 0.015 to 0.054 ± 0.044% of gated mononuclear cells, P < 0.05) only by pitavastatin treatment. Besides, plasma asymmetric dimethylarginine level was reduced (from 0.68 ± 0.10 to 0.53 ± 0.12 µmol/L, P < 0.05) by atorvastatin, and plasma vascular endothelial growth factor (VEGF) level was increased (from 74.33 ± 32.26 to 98.65 ± 46.64 pg/mL, P < 0.05) by pitavastatin. In the in vitro study, while both statins increased endothelial nitric oxide synthase (eNOS) expression, only pitavastatin increased the phosphorylation of eNOS in EPCs. Pitavastatin but not atorvastatin ameliorated the adhesion ability of early EPCs and the migration and tube formation capacities of late EPCs. CONCLUSIONS: While both statins similarly reduced plasma lipids, only pitavastatin increased plasma VEGF level and circulating EPCs in high-risk patients, which is probably related to the differential pleiotropic effects of different statins. TRIAL REGISTRATION: This trial is registered at ClinicalTrials.gov, NCT01386853.

Low-dose dextromethorphan, a NADPH oxidase inhibitor, reduces blood pressure and enhances vascular protection in experimental hypertension.

BACKGROUND: Vascular oxidative stress may be increased with age and aggravate endothelial dysfunction and vascular injury in hypertension. This study aimed to investigate the effects of dextromethorphan (DM), a NADPH oxidase inhibitor, either alone or in combination treatment, on blood pressure (BP) and vascular protection in aged spontaneous hypertensive rats (SHRs). METHODOLOGY/PRINCIPAL FINDINGS: Eighteen-week-old WKY rats and SHRs were housed for 2 weeks. SHRs were randomly assigned to one of the 12 groups: untreated; DM monotherapy with 1, 5 or 25 mg/kg/day; amlodipine (AM, a calcium channel blocker) monotherapy with 1 or 5 mg/kg/day; and combination therapy of DM 1, 5 or 25 mg/kg/day with AM 1 or 5 mg/kg/day individually for 4 weeks. The in vitro effects of DM were also examined. In SHRs, AM monotherapy dose-dependently reduced arterial systolic BP. DM in various doses significantly and similarly reduced arterial systolic BP. Combination of DM with AM gave additive effects on BP reduction. DM, either alone or in combination with AM, improved aortic endothelial function indicated by ex vivo acetylcholine-induced relaxation. The combination of low-dose DM with AM gave most significant inhibition on aortic wall thickness in SHRs. Plasma total antioxidant status was significantly increased by all the therapies except for the combination of high-dose DM with high-dose AM. Serum nitrite and nitrate level was significantly reduced by AM but not by DM or the combination of DM with AM. Furthermore, in vitro treatment with DM reduced angiotensin II-induced reactive oxygen species and NADPH oxidase activation in human aortic endothelial cells. CONCLUSIONS/SIGNIFICANCE: Treatment of DM reduced BP and enhanced vascular protection probably by inhibiting vascular NADPH oxidase in aged hypertensive animals with or without AM treatment. It provides the potential rationale to a novel combination treatment with low-dose DM and AM in clinical hypertension.

Monascus purpureus-fermented rice inhibits tumor necrosis factor-α-induced upregulation of matrix metalloproteinase 2 and 9 in human aortic smooth muscle cells.

OBJECTIVES: Inflammation is associated with atherosclerosis. Cholestin (Monascus purpureus-fermented rice) contains a naturally occurring statin, which has lipid-modulating, anti-inflammatory and antioxidative effects. This study aimed to investigate the effects of Cholestin extract on the expression of matrix metalloproteinase (MMP)-2 and MMP-9 by tumor necrosis factor (TNF)-α-treated human aortic smooth muscle cells (HASMCs). METHODS: Zymography, reverse transcription polymerase chain reaction and immunoblot analyses were used for analysis of MMP expression of TNF-α-stimulated HASMCs. Gel shift assay was used for analysis of transcription factor nuclear factor-κB (NF-κB) activation. Intracellular reactive oxygen species (ROS) generation was also analysed. KEY FINDINGS: The supplement of HASMCs with Cholestin extract significantly suppresses enzymatic activities of MMP-2 and MMP-9 in TNF-α-stimulated HASMCs. RT-PCR and immunoblot analyses show that Cholestin extract significantly attenuates TNF-α-induced mRNA and protein expressions of MMP-2 and MMP-9. Gel shift assays show that Cholestin treatment reduces TNF-α-activated NF-κB. Furthermore, Cholestin also attenuates intracellular ROS generation in TNF-α-treated HASMCs. The supplement with an ROS scavenger N-acetyl-cysteine (glutathione precursor) gives similar results to Cholestin. CONCLUSIONS: Cholestin reduces TNF-α-stimulated MMP-2 and MMP-9 expression as well as downregulating NF-κB activation and intracellular ROS formation in HASMCs, supporting the notion that the natural compound Cholestin may have potential application in clinical atherosclerosis disease.

Nrf-2 mediated heme oxygenase-1 expression, an antioxidant-independent mechanism, contributes to anti-atherogenesis and vascular protective effects of Ginkgo biloba extract.

AIMS: Vascular protective effects of Ginkgo biloba extract (GBE) may involve both antioxidant-related and anti-inflammatory mechanisms. GBE was recently suggested as a heme oxygenase (HO)-1 inducer. The role of HO-1 in anti-atherogenesis and related vascular protective effects of GBE awaited further clarification. METHODS AND RESULTS: Tumor necrosis factor (TNF)-α was used to stimulate adhesiveness of human aortic endothelial cells (HAECs) to monocytes, an in vitro sign simulating atherogenesis. Pretreatment with GBE reduced TNF-α-stimulated endothelial adhesiveness, which could be attenuated by HO-1 inhibitors ZnPP IX or SnPP IX. GBE increased HO-1 expression and enzyme activity in HAECs. Pretreatment with MAP kinase inhibitor SB203580 significantly reduced GBE-induced HO-1 expression. Furthermore, GBE activated the translocation of the transcription factor nuclear factor-erythroid 2-related factor 2 (Nrf2), and increased its binding to the antioxidant response element (ARE) of the HO-1 gene. Pretreatment with PEG-SOD or other antioxidant reagents did not alter GBE-induced endothelial HO-1 expression. In vivo study also showed that GBE treatment could reduce leukocyte adherence to injury arteries, and enhance HO-1 expression in circulating monocytes and in arteries after wire injury, suggesting the in vivo induction of HO-1 by GBE. CONCLUSION: GBE could inhibit cytokine-induced endothelial adhesiveness by inducing HO-1 expression via the activation of p38 and Nrf-2 pathways, a mechanism in which oxidative stress is not directly involved. GBE might exert its anti-atherogenesis and vascular protective effects by inducing vascular HO-1 expression.

Ginkgo biloba extract inhibits endotoxin-induced human aortic smooth muscle cell proliferation via suppression of toll-like receptor 4 expression and NADPH oxidase activation.

Toll-like receptor 4 (TLR4) initiates the inflammatory response in blood vessels in reaction to immune stimuli such as lipopolysaccharide (LPS) produced by gram-negative bacteria. LPS-induced proliferation and functional perturbation in vascular smooth muscle cells play important roles during atherogenesis. Ginkgo biloba extract is an antiatherothrombotic Chinese herbal medicine with anti-inflammatory properties. The effects of G. biloba extract on LPS-induced proliferation and TLR4 expression and the underlying mechanisms for these actions, in human aortic smooth muscle cells (HASMCs), were examined in vitro. LPS-induced proliferation was mediated by the expression of TLR4 in HASMCs. LPS increased the expression of TLR4 in HASMCs, and this effect was mediated by the activation of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, phosphorylation of intracellular mitogen-activated protein kinases (MAPKs), and increases in the cytoplasmic level of HuR and TLR4 mRNA stability. G. biloba extract inhibited LPS-induced HASMC proliferation and decreased the expression of TLR4 by inhibiting LPS-induced NADPH oxidase activation, mRNA stabilization, and MAPK signaling pathways. These results suggest that LPS-induced TLR4 expression contributes to HASMC proliferation and that G. biloba inhibits LPS-stimulated proliferation of HASMCs by decreasing TLR4 expression.