Objective To investigate the effect of the inhibition of 15-lipoxygenase (15-LOX)/15-hydroxyeicosatetraenoic acid (15-HETE) on the expression of Kv channels in the cerebral vasoconstriction induced by hypoxia. Methods Smooth muscle cells originated from the cerebral artery of Wistar rats were separated and cultured by enzyme digestion method. The cells were then assigned randomly into four groups: group A:control group, carotid artery smooth muscle cells (CASMCs) were cultured under normoxic condition for 48h; group B:hypoxia group, CASMCs were cultured under hypoxic condition for 48h; group C:15-LOX gene overexpression under normoxic condition group, 15-LOX gene of CASMC was overexpressed and then the CASMCs were cultured under normoxic condition for 48h; and group D:15-LOX gene knockout under hypoxic condition group, 15-LOX gene of CASMC was knocked out and then the CASMCs were cultured under hypoxic condition for 48 h. The production of 15-HETE in each group was detected by using enzyme-linked immunosorbent assay (ELISA), the expression of Kv2.1 channel messenger ribonucleic acid (mRNA) and protein were detected by using reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis respectively. Results After interference of 15-LOX gene on the CASMC, the production of 15-HETE and the expression of Kv2.1 were signiifcantly different from the normal cells group. The production of 15-HETE was higher and the expression of Kv2.1 channel mRNA and protein were lower in both hypoxia group and 15-LOX gene overexpression under normoxic condition group than control group; the production of 15-HETE was lower and the expression of Kv2.1 channel mRNA and protein were higher in 15-LOX gene knockout under hypoxic condition group than hypoxia group. Conclusion The expression of 15-LOX increased in hypoxia, which increased the production of 15-HETE, and then leading to cerebral vasoconstriction via enhancing Kv2.1 channel inhibition. So the inhibition of 15-LOX/15-HETE on the Kv channels expression plays an important role that leads to cerebral vasoconstriction by hypoxia.%目的探讨缺氧诱导的脑动脉收缩中，脑动脉平滑肌细胞上的15-脂加氧酶（15-lipoxygenase，15-LOX）/15-羟二十碳四烯酸（15-hydroxyeicosatetraenoic acid，15-HETE）对Kv通道的抑制作用。 方法健康Wistar大鼠，通过酶消化法分离培养脑动脉平滑肌细胞（carotid artery smooth muscle cel， CASMC），分为4组：A组为正常细胞常氧组（对照组）：将CASMC在常氧环境下常规培养48 h；B组为正常细胞缺氧组：在缺氧箱内培养48 h；C组为15-LOX基因过表达细胞常氧组：对细胞上的15-LOX进行基因过表达后放入常氧环境下培养48 h；D组为15-LOX基因敲除细胞缺氧组：对细胞上的15-LOX进行基因敲除后放入缺氧箱内培养48h。酶联免疫吸附测定（enzyme-linked immunosorbent assay，ELISA）法测定各组CASMC的15-HETE生成量；反转录聚合酶链反应（reverse transcription-polymerase chain reaction，RT-PCR）及Western-blot测定各组CASMC的Kv2.1信使RNA（messenger ribonucleic acid，mRNA）及蛋白质表达情况。 结果对细胞上的15-LOX基因进行干扰后，其15-HETE生成量及Kv2.1表达情况异于正常细胞组。与正常细胞常氧组比较，正常细胞缺氧组及15-LOX基因过表达细胞常氧组15-HETE生成量均增加， Kv2.1 mRNA及蛋白质表达均下调（P＜0.05）；与正常细胞缺氧组比较，15-LOX基因敲除细胞缺氧组15-HETE生成量减少，Kv2.1 mRNA及蛋白质表达均上调（P＜0.05）。 结论在缺氧诱导的脑动脉收缩中，脑动脉平滑肌细胞上的15-LOX表达增加，15-HETE生成量增加，对Kv通道的抑制作用增强，因此缺氧可能是通过15-LOX/15-HETE对Kv通道的抑制引起脑动脉收缩的。
展开▼
