参附注射液对糖尿病大鼠心肌缺血再灌注时PTEN和PI3K表达的影响

Effects of Shenfu injectio on expression of PTEN and PDK during myocardial ischemia-reperfusion in diabetic rats

目的探讨参附注射液（SFI）对糖尿病大鼠心肌缺血再灌注时第10染色体同源丢失性磷酸酶．张力蛋白酶基因（PTEN）和磷脂酰肌醇-3激酶（PI3K）表达的影响。方法健康成年雄性SD大鼠，体重220～280g，单次腹腔注射1％链脲佐菌素-柠檬酸盐缓冲液60mg／kg制备糖尿病模型，取糖尿病模型制备成功的大鼠30只，随机分为3组（n=10）：假手术组（S组）、缺血再灌注组（IR组）和SFI组。IR组和SFI组采用结扎冠状动脉左前降支30min，再灌注120min的方法建立心肌缺血再灌注模型，S组只穿线不结扎；SFI组开胸前以10ml·kg^-1·h^-1的速率静脉输注SFI，开胸后以3ml·kg^-1·h^-1的速率静脉输注至术毕，S组和IR组均静脉输注等容量乳酸钠林格氏液和羟乙基淀粉。于再灌注120min时处死大鼠，取左心室心肌组织，计算心肌梗死面积。采用TUNEL法检测心肌细胞凋亡情况，计算细胞凋亡指数。采用免疫组织化学法测定心肌组织PTEN和PI3K的表达水平，并计算其比值（PTEN／PI3K）。观察心肌组织病理学结果。细胞凋亡指数与PTEN／PI3K行直线相关分析。结果与S组比较，IR组和SFI组心肌梗死面积增加，细胞凋亡指数升高，PTEN和PI3K的表达上凋（P〈0．05或0．01），SFI组PTEN／PI3K降低（P〈0．05）；与IR组比较，SFI组心肌梗死面积减小，细胞凋亡指数降低，PTEN表达下调，PI3K表达上调，VFEN／PI3K降低（P〈0．05）；SFI组心肌损伤程度比IR组减轻。细胞凋亡指数与PTEN／PI3K呈正相关（r=0．452，P〈0．05）。结论SFI减轻糖尿病大鼠心肌缺血再灌注损伤的机制与下调心肌组织PTEN表达，上调PI3K表达，从而激活PI3K／Akt信号通路有关。

Objective To investigate the effect of Shenfu injectio （SFI） on the expression of phosphatase and tensin homolog deleted on chromosome 10 （PTEN） and phosphatidylinositol 3-kinase （PI3K） during myocardial ischemia-reperfusion （IR） in diabetic rats. Methods Thirty adult male diabetic SD rats weighing 220- 280 g were used in this study. Diabetes mellitus was induced with intraperitoneal streptozotocin 60 mg/kg and confirmed by fasting blood glucose 〉 16.7 mmol/L. The animals were randomly divided into 3 groups （ n = 10 eacb）：group Ⅰ sham operation （group S）; group Ⅱ IR and group Ⅲ SFI. Myocardial IR was produced by occlusion of left anterior descending branch （LAD） of coronary artery for 30 min followed by 120 min reperfusion in group IR and SFI. LAD was exposed but not occluded in group S. SFI was infused iv at 10ml·kg^-1·h^-1 before opening the thoracic cavity and at 3ml·kg^-1·h^-1 after opening the thoracic cavity in group SFI until the end of operation. Equal volume of Lactated Ringer＇s solution and hydroxyethyl starch was infused instead of SFI in group S and IR. The rats were killed and hearts removed at 120 min of reperfusion for microscopic examination and determination of cardiomyocyte apoptosis （by TUNEL）and expression of FIEN and PI3K （by immunohistochemical method）. PTEN/PI3K ratio, myocardial infarct size of left ventricle and apoptosis index were calculated. Correlation between apoptosis index and PTEN/PI3K ratio was analyzed. Results Myocardial infarct size and apoptosis index were significantly increased, while expression of PTEN and PI3K was up-regulated in group IR and SFI as compared with group S （P 〈 0.05 or 0.01）. PTEN/PI3K ratio was significantly decreased in group SFI as compared with group S （ P 〈 0.05 ） . Myocardial infarct size and apoptosis index were significantly decreased, PTEN expression was down-regulated, PI3K expression was up-regulated and PTEN/PI3K ratio was significantly decreased in group SFI as compared with group IR（ P 〈 0.05）. Myocardial pathological damage was attenuated in group SFI as compared with group IR. Apoptosis index was positively correlated to PTEN/PI3K ratio （ r = 0.452, P 〈 0.05）. Conclusion SFI can attenuate myocardial IR injury via down-regulating the expression of PTEN, up- regulating the expression of PI3K and activating PI3K/Akt signal pathway in diabetic rats.