四逆汤对大鼠肠缺血再灌注后肠粘膜细胞凋亡的影响及神经酰胺机制

目的　研究四逆汤(SND)对大鼠肠缺血再灌注 (II/R后肠粘膜细胞凋亡的影响,并从神经酰胺信使通路探讨其抗凋亡机制。方法　24只SD大鼠随机分为 3组(n=8):对照组(仅分离不阻断肠系膜上动脉)、模型组 (阻断肠系膜上动脉 1h后再灌注 3h)、SND组(每天 3g·kg-1 SND灌胃,连续 3天后手术)。取回肠末端组织行电镜检查;检测肠粘膜组织SOD活性、MDA及神经酰胺含量的变化,TUNEL法检测肠粘膜上皮细胞的凋亡指数,RT PCR法分析肠粘膜组织鞘磷脂酶(SMase)基因表达的变化。结果　模型组电镜下可发现许多典型凋亡的上皮细胞,凋亡指数达 30 82%6 34%,SND预处理后凋亡指数为 14 91%±5 40%,明显低于模型组(P<0 01);模型组SOD活性明显低于对照组 (<0 01),MDA与神经酰胺含量以及SMase的基因表达明显高于对照组(P<0 01),且神经酰胺含量与凋亡指数具有良好的正相关 (r=0 852,P<0 01),与SOD活性具有负相关(r= -0 775,P<0 01 )。SND预处理能明显增强SOD活性,降低肠粘膜组织MDA及神经酰胺含量,减少肠粘膜组织SMase的基因表达(P<0 01)。SND组凋亡指数与神经酰胺的含量亦呈显著的正相关 (r=0 832,P<0 01 )。结论　SND具有抗II/R后肠粘膜细胞凋亡的作用,其作用与它清楚氧自由基、抑制SMase的基因表达、减少神经酰胺的生成有关。

Effects of sini decoction on intestinal mucosa cell apoptosis and its ceramide mechanism after intestinal ischemia reperfusion in rats

Aim To investigate the effects of sini decoction(SND) on intestinal mucosa cell apoptosis and its mechanism after intestinal ischemia reperfusion in rats.Methods Twenty-four Sprague-Dawley rats of both sexes were randomly divided into 3 equal groups:①control group in which sham operation was performed;②model group in which intestinal I/R was produced by clamping super mesenteric artery(SMA) for 1 hour and declamping SMA for 3 hours;③SND group in which SND(3 g·kg -1)was given via stomach tube for 3 days before operation. A strip of small intestine was taken from distal end of ileum for electron microscopic examination; SOD activity, MDA and ceramide concentration of intestinal mucosa were detected;Apoptosis of intestinal mucosa cell was examined by TUNEL method and Sphingomyelinase (SMase) mRNA expression was analyzed by RT-PCR.Results In model group, many typical apoptotic epithelial cells were seen under electron microscopic, apoptosis index was higher than that in control group(P<0.01). SOD activity of model group was significantly lower than that of control group(P<0.01), but MDA and ceramide content as well as SMase mRNA expression were obviously higher than those in control group(P<0.01).In addition, In model group, ceramide content was positively correlated to apoptosis index(r=0.852,P<0.01),and negativel y to SOD activity (r=-0.775,P< 0.01).The pretreatment of SND change d the above abnormal phenomena significantly(P<0.01). Conclusion SND can reduce intestinal mucosa cell apoptosis by removing oxygen free r adical, downregulation of SMase mRNA expression and suppression of ceramide production.