苦瓜蛋白对柯萨奇B3病毒性心肌炎小鼠半胱天冬酶3活性及凋亡的抑制作用

Inhibitory Effects of Momordicin on the Activity of Caspase 3 and Apoptosis of Myocardium in BALB/C Mice with Coxsackievirus B3 Myocarditis

为研究苦瓜蛋白对凋亡的关键酶半胱天冬酶 3及凋亡的调节作用 ,用已经建立起来的方法从苦瓜果肉中提取苦瓜蛋白。将BALB/C小鼠分为 4组 :病毒对照组、正常对照组、药物对照组和药物治疗组 ,分别在第 0天、第 3天、第 7天和第 14天各处死 5只小鼠 ,第 2 1天全部处死动物 ,心肌组织半胱天冬酶 3活性测定按照CAL BIOCHEM公司的试剂说明书进行 ,并稍加改正 ,凋亡鉴定按照Oncogene公司的TdT DNA裂解片断末端原位标记试剂说明书进行。结果发现 ,①病毒对照组第 7天开始出现半胱天冬酶 3活性 (0 .6 3± 0 .2 1pmol/min ,n =5 ) ,第 14天的活性 (10 .9± 1.5pmol/min ,n =5 )显著高于第 7天 ,第 2 1天的活性 (12 .6± 1.3pmol/min ,n =5 )又高于第 14天。②药物治疗组 ,只有一个第 2 1天的标本有半胱天冬酶 3活性 (0 .4 1pmol/min) ,其它两组均未检测到此酶的活性。③DNA裂解片断末端原位标记法发现 ,病毒对照组在第 7天心肌中有少数细胞凋亡 ,第 14天、第 2 1天凋亡细胞明显增多 ,在非病变区域可发现单个凋亡的心肌细胞。治疗组未发现凋亡细胞 ,其它两组也未发现凋亡细胞。结果提示 ,CVB3病毒性心肌炎中发现有明显的凋亡现象 ,凋亡和半胱天冬酶 3发现于第 7天 ,且随着时间增加而逐渐加重(增强 ) ;苦瓜蛋白可显?

Aim To measurement the inhibitory effect of Momordicin on the activity of caspase 3 and apoptosis of myocardium in BALB/C mice with coxsackievirus B3 myocarditis. Method Momordicin was purificated from Momordica charantia L by our previous reported methods. Four group animals were used in this experiment, such as disease group (DG), normal group (NG), Momordicin group (MG), and Momordicin therapy group (MTG). Five animals were killed at the 3rd,the 7th,and the 14th days in each group, and all were killed at the 21st day. According to the method of Calbiochem Company, the activity of caspase 3 from the heart of BALB/C mice was measured, and apoptosis was measured by terminal transferase mediated DNA nick end labeling assay (TUNEL, the method provided by Oncogene Company). Results ①The activity of caspase 3 were measurable at the 7th day in the DG (0.63±0.21 pmol/min, n=5). The activity of caspase 3 at the 14th day (10.9±1.5 pmol/min, n=5) and the 21st day (12.6±1.3 pmol/min, n=5) are higer than the 7th day, and the activity of the 21st was higher than 14th day (P<0.05). ②Only a animal has the activity of caspase 3 at the 21st day (0.41 pmol/min) in the HTG, no caspase 3 activity were detected in the animals of MG and NG. ③Only a few apoptotic cells were found at the 7th day in the DG, and more apoptotic cells were found among cardiac muscles at 14th and 21th days. Single apoptotic cardiomyocytes were also found at the outside of the pathological change areas, and no apoptotic cells were found in the animals of NG, MG, and MTG. Conclusions Distinct apoptosis were found in CVB3 viral myocarditis; the activitys of caspase 3 and apoptosis were found at the 7th day, and higher after the 7th day, single apoptotic cardiomyocytes were also found. The high dose Momordicin has distinct inhibitory effect on the activity of caspase 3 and apoptosis.