15-HETE影响大鼠脑动脉平滑肌细胞内钙离子浓度的机制

Effect of 15-hydroxyeicosatetraenoic acid on calcium ion concentration of cerebral arterial smooth muscle cells in rats

目的 观察15-羟化二十烷四烯酸（15-HETE）对脑动脉平滑肌细胞内钙离子浓度（[Ca^2＋]）的影响，进一步探讨15-HETE引起[Ca^2＋]，变化的钙来源，从而明确15-HETE引起脑动脉平滑肌收缩的机制。方法酶法分离大鼠脑动脉平滑肌细胞，分为15-HETE组与对照组，15-HETE组添加15-HETE处理；对照组正常培养，不做其他处理。激光共聚焦技术测定15-HETE对[Ca^2＋]i的影响：进一步通过阻断外钙内流和耗竭内钙，探明15-HETE引起钙动员的来源；应用血管环技术从功能上判定细胞外钙对15-HETE引起的颈内动脉环收缩有无影响。结果15-HETE组与对照组相比，[Ca^2＋]，明显增加，差异有统计学意SL（P〈0．05）；预先加入硝苯地平、镧离子及改用无钙液阻断外钙内流后，15-HETE组[Ca^2＋]，仍明显高于对照组，差异有统计学意义（P＜0．05）；而预先加入咖啡因耗竭细胞内钙后，15-HETE组[Ca2＋]，较对照组差异无统计学意义（P＞0．05）；采用无钙液去除细胞外钙后，15-HETE引起的血管环张力增加与有钙液中比较差异无统计学意义（P〉0．05）。结论15-HETE可通过促使内钙释放而使[Ca^2＋]，增加，进而引起大鼠脑动脉平滑肌收缩。

Objective To observe the effect of 15-hydroxyeicosatetraenoic acid （15-HETE） on the intracellular calcium ion （[Ca2＋]i） concentration of cerebral arterial smooth muscle and discuss the source of （[Ca2＋]i） mobilization induced by 15-HETE to reveal the mechanism underlying the vasoconstriction aroused by 15-HETE. Methods First, papain and collagenase were employed to isolate the cerebral artery smooth muscle cells （SMCs） from the rats. The cells were separated into experimental group （treated with 15-HETE） and control group. Confocal laser scanning microscope was used to investigate the （[Ca2＋]i） signaling in cultured SMCs. Then, Ca2＋ channel blockers and Ca2＋ store depletion agent were added to identify the source of Ca2＋ transients in SMCs of the 2 groups. Finally, internal carotid artery （ICA） rings were used to observe the effect of non-Ca2＋ solution on 15-HETE-induced ICA vasoconstriction on both groups. Results Compared with the control group, 15-HETE treatment group enjoyed a significantly increased level of （[Ca2＋]i） in cultured SMCs （P〈0.05）. After the addition of Ca2＋ channel blockers （nifedipine, lanthanum ion） and calcium-free solution, 15-HETE treatment group still enjoyed a significantly increased level of （[Ca2＋]i） in cultured SMCs as compared with the control group （P〈0.05）, while after the addition of Ca2＋ store depletion agent （caffeine）, no obvious difference was noted between the 2 groups （P〉0.05）. Non-Ca2＋ solution had no effect on 15-HETE induced vasoconstriction. Conclusion 15-HETE may activate Ca2＋ releasing from intracellular stores to rise. （[Ca2＋]i） level in SMCs, which subsequently induce the cons.Wiction of cerebral arterial smooth muscle.