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Background A hallmark of atherosclerosis is progressive intimal thickening (namely neointimal hyperplasia), which leads to occlusive vascular diseases. Over-production of reactive oxygen species (ROS) and alteration of Ca 2 + signaling are among the key factors contributing to neointimal growth. In the present study, we investigated the role of TRPM2, a ROS-sensitive Ca 2 + entry channel, in neointimal hyperplasia. Methods and results Perivascular cuffs were used to induce neointimal hyperplasia in rat/mouse arteries. Immunostaining showed numerous TRPM2-positive smooth muscle cells in neointimal regions. ROS were over-produced and PCNA-positive proliferating cells were numerous in the neointimal regions. The neointimal hyperplasia was substantially reduced in Trpm2 knockout mice compared with wild-type mice. In the cultured rat/mouse aortic smooth muscle cells, H 2 O 2 treatment was found to stimulate cell proliferation and migration. The effect of H 2 O 2 was reduced by a TRPM2-specific blocking antibody TM2E3 or Trpm2 knockout. The signaling molecules downstream of TRPM2 were found to be Axl and Akt. Conclusions These data suggest a critical functional role of TRPM2 in the progression of neointimal hyperplasia. The study also highlights the possibility of targeting TRPM2 as a potential therapeutic option for the treatment of occlusive vascular diseases.

Luteolin is an antioxidative, antitumor and anti-inflammatory flavone. It has been shown to reduce endothelial dysfunction, but the mechanism is not clear. We set out to explore the effects of luteolin on apoptosis and its mechanism of action in endothelial cells. The effect of luteolin on pyrogallol-induced superoxide stress and the subsequent apoptosis was studied in the mouse heart capillary endothelial cell line H5V and human umbilical vein endothelial cells, by the use of flow cytometry, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide, Hoechst staining, and western blot. Pyrogallol (0–400 μm) dose-dependently induced reactive oxygen species production, cytotoxicity, an annexin V–fluorescein isothiocyanate increase, mitochondrial transmembrane depolarization and DNA condensation in both H5V and human umbilical vein endothelial cells; these actions were reversed by luteolin (0.78–50 μm) in a concentration-dependent manner. Luteolin suppressed the poly (ADP-ribose) polymerase activation, caspase-8 cleavage and p38 mitogen-activated protein kinase activation triggered by pyrogallol, and stimulated the extracellular signal-regulated kinase signaling pathway to counteract the pyrogallol-induced apoptotic signals. Luteolin is an effective agent for the protection of endothelial cells from superoxide stress-induced apoptosis via the extracellular signal-regulated kinase signaling pathway.

Ethnopharmacological relevance Total flavonoids of Flos Chrysanthemi (TFFC) are known to modulate vascular functions, but their effect on endothelial cells injured by oxidative stress is unknown. Our objective was to investigate the vasoprotective effect and mechanism of action of TFFC on rat mesenteric artery exposed to superoxide anions produced by pyrogallol. Materials and methods The vasoprotective effect and mechanism of action of TFFC on primary cultured rat mesenteric arterial endothelial cells and small mesenteric arteries was investigated using small-vessel myography, fluorescent Ca2+ measurement, fluorescent membrane potential measurement and oxidative fluorescent studies. Results Experiments using small-vessel myography of third-order rat mesenteric arterial rings showed that pretreatment with pyrogallol (10–1000 μM), an auto-oxidizing source of superoxide anions, dose-dependently decreased ACh-induced endothelium-dependent relaxation. TFFC (2.5–320 mg/L) evoked a concentration-dependent dilation (pD2: 29.6 ± 0.276 mg/L), which was weakened by ChTX plus apamin. TFFC markedly attenuated the inhibition of vasorelaxation induced by pyrogallol (Emax elevated from 50.4 ± 7.36% to 86.2 ± 3.61%, and pD2 increased from 6.74 ± 0.06 to 7.28 ± 0.12). Furthermore, in primary cultured endothelial cells, fluorescent Ca2+ measurement, fluorescent membrane potential measurement and oxidative fluorescent studies demonstrated that ACh-induced endothelial Ca2+ influx and hyperpolarization were significantly weakened by the increased basal superoxide level induced by pyrogallol. When the endothelial cells were concurrently exposed to TFFC, the impairment effect of oxidative stress on ACh-induced Ca2+ influx, hyperpolarization and vasorelaxation were attenuated due to its superoxide-lowering activity. Conclusion This study shows that oxidative stress has a pronounced deleterious effect on EDHF-mediated vasorelaxation to ACh in rat mesenteric artery. TFFC has vasodilating effect and protects EDHF-mediated vasodilator reactivity from oxidative stress. Thus, our experiments suggest that TFFC is potentially useful for the development of therapeutic treatments for cardiovascular diseases associated with oxidative stress.

Aims: The aim of this study was to investigate the effect of alcohol on rat artery and its underlying mechanism. Methods: The tension of isolated Sprague-Dawley rat thoracic aortic rings and the pressure of rat mesenteric arterial beds perfused with different concentrations of alcohol (0.1–7.0‰) were measured. Results: At resting tensions, alcohol caused a concentration-dependent relaxation on endothelium-denuded aortic rings precontracted with KCl (6×10−2 mol/L) or phenylephrine (PE, 10−6 mol/L), and this effect was most evident on rings at a resting tension of 3 g. Alcohol induced much less vasodilation on endothelium-intact rings. Alcohol inhibited the CaCl2-induced contraction of endothelium-denuded aortic rings precontracted with KCl or PE. Incubation of rings with dantrolene (5×10−5 mol/L), a ryanodine receptor blocker, or 2-aminoethyl diphenylborinate (7.5×10−5 mol/L), an IP3 receptor blocker, attenuated the vasodilating effect of alcohol on rings precontracted with PE. Alcohol also concentration-dependently relaxed rat mesenteric arterial beds precontracted with KCl (6×10−2 mol/L) or PE (10−5 mol/L), which was more potent on endothelium-denuded than on endothelium-intact beds. Conclusions: Alcohol has a vasodilating effect on rat artery depending on the resting tension. Both extracellular and intracellular Ca2+ mobilization of vascular smooth muscle cells are involved in the vascular effect of alcohol.

Transient receptor potential (TRP) superfamily is a superfamily of cation channels that can be divided into seven subfamilies. TRPM2 is the second member of the TRPM subfamily, which includes eight members, namely TRPM1-8. TRPM2 is widely expressed in excitable and non-excitable cells, where it forms a Ca(2+)-permeable cation channel and performs diverse cellular functions. TRPM2 channels are activated by ADP-ribose (ADPR), Ca(2+), H2O2 and other reactive oxygen species (ROS). It is established that TRPM2 serves as a cellular sensor for oxidative stress, mediating oxidative stress-induced [Ca(2+)]i increase and contributing to pathological processes in many cell types. Accumulating evidence has indicated that TRPM2 is a potential therapeutic target for oxidative stress-related diseases. This review will highlight recent progress in this field.

Luteolin is an antioxidative, antitumor and anti-inflammatory flavone. It has been shown to reduce endothelial dysfunction, but the mechanism is not clear. We set out to explore the effects of luteolin on apoptosis and its mechanism of action in endothelial cells. The effect of luteolin on pyrogallol-induced superoxide stress and the subsequent apoptosis was studied in the mouse heart capillary endothelial cell line H5V and human umbilical vein endothelial cells, by the use of flow cytometry, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide, Hoechst staining, and western blot. Pyrogallol (0-400 μm) dose-dependently induced reactive oxygen species production, cytotoxicity, an annexin V-fluorescein isothiocyanate increase, mitochondrial transmembrane depolarization and DNA condensation in both H5V and human umbilical vein endothelial cells; these actions were reversed by luteolin (0.78-50 μm) in a concentration-dependent manner. Luteolin suppressed the poly (ADP-ribose) polymerase activation, caspase-8 cleavage and p38 mitogen-activated protein kinase activation triggered by pyrogallol, and stimulated the extracellular signal-regulated kinase signaling pathway to counteract the pyrogallol-induced apoptotic signals. Luteolin is an effective agent for the protection of endothelial cells from superoxide stress-induced apoptosis via the extracellular signal-regulated kinase signaling pathway.

Nonreceptor tyrosine kinases have an increasingly appreciated role in cardiac injury and protection. To investigate novel tasks for members of the Tec family of nonreceptor tyrosine kinases in cardiac phenotype, we examined the behavior of the Tec isoform in myocardial ischemic injury. Ischemia-reperfusion, but not cardiac protective agents, induced altered intracellular localization of Tec, highlighting distinct actions of this protein compared with other isoforms, such as Bmx, in the same model. Tec is abundantly expressed in cardiac myocytes and assumes a diffuse intracellular localization under basal conditions but is recruited to striated structures upon various stimuli, including ATP. To characterize Tec signaling targets in vivo, we performed an exhaustive proteomic analysis of Tec-binding partners. These experiments expand the role of the Tec family in the heart, identifying the Tec isoform as an ischemic injury-induced isoform, and map the subproteome of its interactors in isolated cells.

Vascular oxidative injury is involved in numerous cardiovascular diseases, particularly in hypertension and atherosclerosis. Statins -- a family of lipid lowering drugs -- appear to display non-lip...