Diabetes mellitus comprises a heterogeneous group of metabolic disorders with underlying hyperglycemia and secondary cardiovascular complications. Growing evidence suggests that vascular dysfunction is among the most important causes of diabetic cardiovascular disease. Therefore, we determined whether streptozotocin (STZ)-induced diabetes in mice affects blood pressure and cerebral arterial responsiveness to angiotensin (Ang) II and acetylcholine (ACh), which are important modulators of cerebrovascular autoregulation. Diabetes was induced using a single intraperitoneal injection of STZ (50 mg/kg). Blood pressure was measured in conscious mice using the indirect tail-cuff method. Functional studies of the isolated arteries' response to vasoactive substances were performed using a micro-organ-bath system at 60 days after STZ injection. Systolic, diastolic, and mean blood pressures significantly increased at days 45 and 60 in the STZ-induced diabetic mice. In the isolated basilar arteries, ACh-induced relaxation, which is dependent on nitric oxide (NO) production from endothelial cells, decreased. In contrast, Ang II-induced contraction, mediated via rho-kinase activation in the smooth muscle, increased in the diabetic mice. There was significantly greater relaxation in the precontracted isolated basilar arteries of diabetic mice that had been treated with Y27632, a rho-kinase inhibitor, than in the control mice arteries. Pretreatment with Nω-nitro-L-arginine (L-NAME), an NO synthase inhibitor, significantly enhanced Ang II-induced contraction and Y27632-induced relaxation in the control basilar arteries but not in the STZ-induced diabetic mice arteries. These results suggest that decreased NO bioavailability and enhanced rho-kinase activity in basilar arteries contribute to altered reactivity to ACh and Ang II, respectively, in STZ-induced diabetic mice.

Mercury (Hg) and cadmium (Cd) are the major toxic heavy metals and are known to induce neurotoxicity. Although many studies have shown that several heavy metals have neurotoxic effects, the cellular and molecular mechanisms thereof are still not clear. Oxidative stress is reported to be a common and important mechanism in cytotoxicity induced by heavy metals. However, the assays for identifying toxic mechanisms were not performed under the same experimental conditions, making it difficult to compare toxic properties of the heavy metals. In this study, we investigated the mechanisms underlying neurotoxicity induced by heavy metals and H 2 O 2 , focusing on cell death, cell proliferation, and oxidative stress under the same experimental condition. Our results showed that MeHg caused lactate dehydrogenase (LDH) release, caspase activation and cell-cycle alteration, and ROS generation in accordance with decreased cell viability. HgCl 2 caused LDH release and cell-cycle alteration, but not caspase activation. CdCl 2 had a remarkable effect on the cell cycle profiles without induction of LDH release, caspase activation, or ROS generation. Pretreatment with N-acetyl- l -cysteine (NAC) prevented the decrease in cell viability induced by MeHg and HgCl 2 , but not CdCl 2 . Our results demonstrate a clear difference in neurotoxic mechanisms induced by MeHg, HgCl 2 , CdCl 2 or H 2 O 2 in SH-SY5Y cells. Elucidating the characteristics and mechanisms of each heavy metal under the same experimental conditions will be helpful to understand the effect of heavy metals on health and to develop a more effective therapy for heavy metal poisoning.

Methylmercury (MeHg) is a persistent environmental contaminant that has been reported worldwide. MeHg exposure has been reported to lead to increased risk of cardiovascular diseases; however, the mechanisms underlying the toxic effects of MeHg on the cardiovascular system have not been well elucidated. We have previously reported that mice exposed to MeHg had increased blood pressure along with impaired endothelium-dependent vasodilation. In this study, we investigated the toxic effects of MeHg on a human endothelial cell line, EA.hy926. In addition, we have tried to elucidate the role of myristoylated alanine-rich C kinase substrate (MARCKS) in the MeHg toxicity mechanism in EA.hy926 cells. Cells exposed to MeHg (0.1–10 µM) for 24 hr showed decreased cell viability in a dose-dependent manner. Treatment with submaximal concentrations of MeHg decreased cell migration in the wound healing assay, tube formation on Matrigel and spontaneous nitric oxide (NO) production of EA.hy926 cells. MeHg exposure also elicited a decrease in MARCKS expression and an increase in MARCKS phosphorylation. MARCKS knockdown or MARCKS overexpression in EA.hy926 cells altered not only cell functions, such as migration, tube formation and NO production, but also MeHg-induced decrease in cell viability and NO production. These results suggest the broad role played by MARCKS in endothelial cell functions and the involvement of MARCKS in MeHg-induced toxicity.

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Methylmercury (MeHg) is an environmental pollutant that shows severe toxicity to humans and animals. However, the molecular mechanisms mediating MeHg toxicity are not completely understood. We have previously reported that the MARCKS protein is involved in the MeHg toxicity to SH-SY5Y neuroblastoma and EA.hy926 vascular endothelial cell lines. In addition, calpain, a Ca2+-dependent protease, is suggested to be associated with the MeHg toxicity. Because MARCKS is known as a substrate of calpain, we studied the relation between calpain activation and cleavage of MARCKS and its role in MeHg toxicity. In SH-SY5Y cells, MeHg decreased cell viability along with increased calcium mobilization, calpain activation and a decrease in MARCKS amounts. However, pretreatment with calpain inhibitors attenuated the decrease in cell viability and MARCKS amount induced only by 1 µM but not by 3 µM MeHg. In cells with a MARCKS knockdown, calpain inhibitors failed to attenuate the decrease in cell viability caused by MeHg. In EA.hy926 cells, although MeHg caused calcium mobilization and a decrease in MARCKS levels, calpain activation was not observed. These results indicate that the participation of calpain in the regulation of MARCKS amounts is dependent on the cell type and concentration of MeHg. In SH-SY5Y cells, calpain-mediated proteolysis of MARCKS is involved in cytotoxicity induced by a low concentration of MeHg.

The aim of this study was to evaluate, for the first time, the antagonistic effects of Gingko biloba leaf (GB) and Sophora japonica L. flower bud (SJ) extracts on cerebral vasoconstriction in response to KCl, extracellular Ca[Formula: see text], histamine, 5-hydroxytryptamine (5-HT), 9,11-dideoxy-9[Formula: see text],11[Formula: see text]-methanoepoxy prostaglandin (PG) F[Formula: see text](U46619) and bradykinin (BK), in order to explain their traditional application for diseases associated with cerebral vasospasm. Isolated porcine basilar arteries (PBA) and endothelial cells from them were used as the study materials. Neither SJ nor GB had any effect on the contractions induced by KCl and extracellular Ca[Formula: see text]. SJ significantly inhibited the contraction induced by histamine, 5-HT, U46619 and BK, whereas GB inhibited histamine-induced contraction, but had no effects on the contractions induced by 5-HT, U46619 and BK. In the presence of diphenhydramine (a H receptor antagonist), ketanserin (a 5-HT receptor antagonist) and ONO-3708 (a thromboxane (TX) A/PG receptor antagonist), the inhibitory effects of these extracts on the contractions induced by histamine, 5-HT and U46619 were abolished. SJ significantly inhibited the contractions induced by BK and PGF[Formula: see text], but in the presence of ONO-3708 (10[Formula: see text] M) had no effect on them. BK enhanced the production of PGF[Formula: see text] from cultured PBA endothelium cells, and SJ significantly attenuated this enhancement. These results suggest that SJ and GB have a H-antagonistic effect, and that SJ also attenuates cerebral vasoconstriction mediated via 5-HT and TXA/PG receptors. These findings appear to explain why SJ has been used traditionally as a therapeutic medication for cerebral vasospasm after cerebral hemorrhage.