VDAC2相关的线粒体凋亡通路在严重烧伤大鼠心肌损伤中的作用及其机制

Voltage dependent anion channel 2 involved mitochondrial apoptosis and its possible regulatory signal pathway in hearts of rats with severe scalds

目的探讨电压依赖负离子通道2（VDAC2）相关的线粒体凋亡途径在严重烧伤大鼠心肌损伤的作用及其调控机制。方法清洁级Wistar大鼠60只（体重198～219g），按随机数字表法分为假伤组、烫伤组各30只。伤后第1、7、14天收集大鼠血清及心肌组织。酶联免疫吸附（ELISA）法检测血清内心肌钙蛋白I（cTnI）水平，Western印迹法检测总蛋白内VDAC2、Bel-2、Bax、细胞色素C、磷脂酰肌醇3激酶（P13K）、磷酸化糖原合成酶激酶3β（β-GSK3p）、己糖激酶2（HK2）的表达水平，检测胞质蛋白内细胞色素c的表达水平。结果烫伤组与假伤组第1、7、14天cTnI浓度分别为（1．41±0．25）、（1．93±0．53）、（1．62±0．34）μg／L与（0．53±0．23）、（0．43±0．23）、（0．41±0．22）μg/L，差异均有统计学意义（均P〈0．05）；Bax／Bel-2比值分别为（3．360±0．173）、（2．736±0．341）、（1．290±0．234）与（0．623±0．044）、（0．698±0．064）、（0．718±0．063），差异均有统计学意义（均P〈0．05）。烫伤组与假伤组第1、7、14天VDAC2表达水平分别为0．070±0．009、0．007±0．002、0．009±0．004和0．328±0．020、0．291±0．025、0．302±0．037，胞质内细胞色素C表达分别为0．418±0．030、1．685±0．169、0．300±0．037和0．022±0．007、0．030±0．011、0．098±0．014，差异均有统计学意义（均P〈0．05）。烫伤组与假伤组第14天P13K表达水平为0．083±0．015与0．328±0．011，差异有统计学意义（P〈0．05）。各时相点烫伤组p-GSrd13表达水平分别为0．098±0．014、0．064±0．002、0．074±0．010，均低于假伤组的0．446±0．031、0．476±0．054、0．442±0．041，差异均有统计学意义（均P〈0．05）。烫伤组第7、14天HK2表达水平分别为0．390±0．027、0．267±0．018，均低于假伤组的0．611±0．070、0．490±0．042，差异均有统计学意义（均P〈0．05）。结论VDAC2相关线粒体凋亡途径加重严重烫伤后的心肌损伤，该过程可能受P13K—GSK3β-HK2信号通路调控。

Objective To explore the role of vohage dependent anion channel 2 （VDAC2） involved mitochondrial apoptosis in heart injury of rats with severe scald injury and elucidate its possible regulatory signal pathway. Methods A total of 60 Wistar rats were divided into sham scald group （ n = 30） and scald group （ n = 30） according to a random digital table. Blood and heart tissue samples were harvested at Day 1, 7, 14 post scalding. Myocardial injury was assessed with cardiac troponin Ⅰ （cTnI） by enzyme-linked immunosorbent assay （ELISA）. Mitochondrial apoptosis activation was evaluated by the expressions of Bax/ Bcl-2 ratio, cytoplasmic cytochrome C and VDAC2. And the levels of phosphatidylinositol 3-kinase, p-Glycogen Synthase Kinase-3β and hexokinase 2 protein were determined by Western blot. Results The serum levels of cTnI were significantly higher in scald group than those in sham scald group at Day 1,7, 14 （（1.41 ±0.25） vs （0.53 ±0.23） μg/L, （1.93 ±0.53） vs （0.43±0.23）μg/L, （1.62 ±0.34） vs （0. 41 ± 0. 22） μg/L respectively, all P 〈 0.05 ）. Compared with sham scald group, Bax/Bcl-2 ratio increased significantly in scald group at Day 1,7 day post-scalding （ 3. 360 ± 0. 173 vs 0. 623 ± 0. 044,2. 736 ± 0. 341 vs 0. 698 ± 0. 064, 1. 290 ± 0. 234 vs 0. 718 ± 0. 063 respectively, all P 〈 0. 05 ）, VDAC2 protein level in scald group decreased significantly at Day 1, 7, 14 （0. 070 ± 0. 009 vs 0. 328 ± 0. 026, 0. 007 ±0. 002 vs 0. 291 ± 0. 025, 0. 009 ± 0. 004 vs 0. 302 ± 0. 037 respectively, all P 〈 0. 05 ）, the cytoplasmic levels of eytoehrome increased significantly in scald group at Day 1, 7, 14 （0. 418 ± 0. 030 vs 0. 022 ±0. 007, 1. 685 ±0. 169 vs 0. 030 ±0. 011, 0. 300 ±0. 037 vs 0. 098 ±0. 014 respectively, all P 〈 0. 05）, the expression of PI3K was significantly lower in scald group at Day 14 post-scalding （0. 083 ± 0. 015 vs 0. 328 ±0. 011, P 〈0.05） , the expressions of p-GSK3β all reduced significantly at Day 1,7, 14 （0. 098 ±0. 014 vs 0. 446 ± 0. 031, 0. 064 ± 0. 002 vs 0. 476 ± 0. 054,0. 074 ± 0. 010 vs 0. 442 ± 0. 041, respectively, all P 〈 0. 05 ） and the expressions of HK2 were lower at Day 7, 14 post-scalding （0. 390 ± 0. 027 vs 0. 611 ± 0. 070, 0. 267 ± 0. 018 vs 0. 490 ± 0. 042, respectively, all P 〈 0. 05 ）. Conclusions VDAC2 involved mitoehondrial apoptosis is activated in myoeardium after severe scalds. And it may be regulated by the pathway of PI3K-GSK-HK2.