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30 mg/kg组为对照组的6.3倍(P＜0.01),60～120 mg/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±6.1.30 mg/kg IND使Bcl-2表达明显减弱(P＜0.05 vs对照组),灰度值为124.8±6.3,60-90 mg/kg组呈中等至弱阳性表达,灰度值分别为153.1±10.1、174.1±8.6,120 mg/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30～90 mg/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±6.3,120 mg/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.%Objective To determine the changing expressions of Bel-2 and Bax protein during in-dotnethacin (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 unmunohistochemically. 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, immuno-histochemistry 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 concrol 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 vscontrol). 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-depen-dently. 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.
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