凋亡相关蛋白Bcl-2、Bax在消炎痛所致胃黏膜细胞凋亡中的作用

目的研究在体情况下消炎痛（Indomethacin,IND）所致胃黏膜细胞凋亡过程中Bcl-2、Bax蛋白表达的变化,以探讨其黏膜损伤机制。方法 SD大鼠胃内灌注不同剂量IND,灌胃后3h处死,采用TUNEL标记技术检测黏膜细胞凋亡;应用免疫组化方法检测Bcl-2、Bax蛋白表达的变化。结果①TUNEL标记显示对照组大鼠胃黏膜仅见少量凋亡细胞,IND使凋亡细胞数明显增加,计算机图像分析显示单位面积内阳性细胞平均像素点30mg/kg组为对照组的6.3倍（P〈0.01）,60～120mg/kg组分别为对照组的8.0、12.6和17.1倍,明显高于对照组（P〈0.01）;②免疫组化染色显示对照组大鼠Bcl-2在胃腺体部呈中等至强阳性表达,平均灰度值为113.8±9.6;而Bax蛋白仅在胃腺体及基底部呈弱阳性表达,平均灰度值为128.5±6.1。30mg/kgIND使Bcl-2表达明显减弱（P〈0.05vs对照组）,灰度值为124.8±6.3,60～90mg/kg组呈中等至弱阳性表达,灰度值分别为153.1±10.1、174.1±8.6,120mg/kg组仅见散在弱阳性细胞分布于胃腺体部,灰度值为222.3±14.6,表达均明显低于对照组（P〈0.01）,表达水平变化同细胞凋亡显著负相关（r=-0.9771,P〈0.01）;Bax表达在30～90mg/kg组呈中等阳性,明显高于对照组（P〈0.05）,灰度值分别为107.4±4.2、90.1±5.6、72.8±6.3,120mg/kg组在胃腺体及基底部见大量强阳性染色细胞分布,灰度值为69.8±6.2,表达明显高于对照组（P〈0.01）,其变化与细胞凋亡明显正相关（r=0.9725,P〈0.01）。结论 IND使凋亡抑制蛋白Bcl-2表达减弱和促凋亡蛋白Bax表达增强,从而降低了Bcl-2/Bax,导致胃黏膜细胞凋亡。

Role of apoptosis-associated protein Bcl-2 and Bax in indomethacin-induced gastric mucosal cell apoptosis

Objective To determine the changing expressions of Bcl-2 and Bax protein during indomethacin(IND)-induced gastric mucosal cell apoptosis in vivo.Methods Healthy male Sprague-Dawley rats were treated intragastrically with four different doses of IND.The rats were killed 3 hours after IND administration and the TUNEL technique was applied to detect mucosal cell apoptosis.The changes of Bcl-2 and Bax protein expression were monitored immunohistochemically.Results TUNEL assay revealed only a few apoptotic cells in rats in control group.Oral administration of IND resulted in the appearance of massive TUNEL positive cells.Computer-aided image analysis showed the mean pixels in 30-120 mg/kg groups were 6.3-,8.0-,12.6-and 17.1-folds that in control group(P < 0.01 vs control).In rats of control group,immunohistochemistry showed that Bcl-2 protein was moderate to strong positive in the middle portion of gastric glands.The mean gray level was 113.8 ± 9.6.In contrast,Bax protein was detected only as a weak signal in the basal and body regions of gastric glands,and the mean gray level was 128.5 ± 6.1.IND administration caused a significant decrease in Bcl-2 expression in a dose-dependent manner.The mean gray level in 30mg/kg group was 124.8 ± 6.3(P < 0.05 vs control).The positive signals in 60-90 mg/kg groups were moderate to weak,being significantly lower than that in control group(153.1 ± 10.1,174.1 ± 8.6,P < 0.01).In 120 mg/kg group,only a few scattered positive cells were seen in the body region of gastric glands(222.3 ± 14.6,P < 0.01 vs control).The change of Bcl-2 expression was significantly negative-related with mucosal cell apoptosis(r =-0.9771,P < 0.01).The expression of Bax protein was significantly enhanced by IND treatment dose-dependently.In 30-90 mg/kg groups,the positive signals were weak to moderate(107.4 ± 4.2,90.1 ± 5.6,72.8 ± 6.3,P < 0.05 vs control).In rats treated with IND at the dose of 120 mg/kg,massive strong-positive cells were found in the basal and body regions of gastric glands(69.8 ± 6.2,P < 0.01 vs control).There was a significant positive relationship between the expression of Bax protein and cell apoptosis(r = 0.9725,P < 0.01).Conclusion IND administration weakened the expression of anti-apoptotic protein Bcl-2 and enhanced the expression of pro-apoptotic protein Bax,resulting in the decrease in Bcl-2/Bax ratio and mucosal cell apoptosis.