摘要
目的:探讨异氟烷对心肌细胞血管内皮生长因子(vascular endothelial growth factor,VEGF)直接调节作用的细胞内信号转导途径,进一步揭示异氟烷心肌保护的分子机制。方法:分离培养原代SD大鼠心肌细胞,随机分为4组。对照组(CON组),细胞不经任何处理;异氟烷组(ISO组),细胞经1.4%的异氟烷处理6 h;蛋白激酶C(PKC)抑制剂组(CAL组),细胞经calphostin C 50 nmol.L-1孵育6 h;PKC抑制剂+异氟烷组(IC组),细胞经calphostin C及异氟烷共同作用6 h。对心肌细胞处理结束后采用Elisa法测定细胞培养液VEGF浓度,采用Western blotting法测定心肌细胞PKC亚型的表达。结果:与CON组比较,ISO组VEGF值显著升高(P<0.01);CAL组经PKC抑制剂calphostin C处理后,VEGF含量显著降低,明显低于ISO组(P<0.01);CAL组和CON组比较差异无统计学意义。异氟烷显著诱导PKC-epsilon(PKCε)激活转位,与CON组比较,1.4%的异氟烷组PKCα、PKCδ和PKCζ亚型在细胞质和细胞膜的表达均无统计学差异,但细胞膜PKCε亚型浓度显著升高,而细胞质的浓度显著降低(P<0.01)。结论:异氟烷通过蛋白激酶PKC-epsilon途径诱导心肌细胞分泌VEGF,有可能是异氟烷心肌保护机制之一。
Objective： To investigate the effect of isoflurane on VEGF expression and associated signaling pathway in cultured rat cardiac myocytes.Methods： Primary cultures of rat cardiomyocytes were randomized into the following groups.The control group cells were incubated without any treatment.The isoflurane group cells were treated with isoflurane at 1.4% for 6 hours.The PKC inhibitor group cells were treated with PKC inhibitor calphostin C at a final concentration of 50 nmol·L-1.The PKC inhibitor ＋ isoflurane group cells were treated with 50 nmol·L-1 calphostin C and 1.4% isoflurane for 6 hours.VEGF expressions were identified by enzyme-linked immunosorbent assay and PKC isoforms expressions were determined by Western blotting.Results： The effect of isoflurane on VEGF upregulation was blocked by PKC inhibitor calphostin C,however,calphostin did not alter VEGF expression（P0.05）.Isoflurane induced PKCε activation and translocation.Immunoblot analysis revealed that the immunoreactivity for PKCε but not for PKCα,PKCδ and PKCζ increased significantly in the membrane fractions and deceased significantly in the cytosol fractions from cells treated with the 1.4% isoflurane,when compared with non-treatment cells（P0.01）.Conclusion： Isoflurane induces VEGF secretion in cardiac myocytes through translocation of PKCε from the cytosol to the cell membrane,suggesting a possible novel mechanism of isoflurane induced cardiac protection.
出处
《东南大学学报（医学版）》
CAS
2011年第5期699-702,共4页
Journal of Southeast University(Medical Science Edition)
基金
湖北省科技攻关计划(2009)(302131741)
