乙酸体外损伤大鼠结肠模型的建立及褪黑素的影响

Establishment of rat colon injury model with acetic acid in vitro and the protective effect of melatonin

目的建立一种简单的乙酸体外损伤大鼠结肠的实验模型并观察褪黑素的作用。方法制备乙酸损伤大鼠结肠体外模型,观察0、0.1%、0.2%、0.4%浓度的乙酸作用5、10、15、20、25、30 m in的结肠粘膜损伤情况,并检测培养液中反映粘膜损伤程度的乳酸脱氢酶(LDH)、丙二醛(MDA)、粘液含量及结肠粘膜AB染色情况。并以0.4%乙酸作用30m in为实验模型,观察具有抗结肠炎作用的褪黑素(终浓度为0.1、1.0、10 mmol.L-1)对模型中上述实验指标的影响。结果随着乙酸浓度增加和作用时间延长,培养液LDH含量逐渐增高,大鼠结肠粘膜损伤与乙酸存在明显的量效和时效关系,同时可见结肠粘膜水肿、肠壁变薄和点片状上皮脱失,培养液MDA含量增加和粘液含量减少,AB染色显示粘膜上皮细胞内粘液蛋白无明显减少。预先给予不同浓度褪黑素可减少LDH释放、增加粘液和降低MDA含量。结论该模型中大鼠结肠粘膜损伤与乙酸存在量效和时效关系,乙酸通过氧化损伤细胞膜并削弱粘膜粘液屏障。褪黑素通过抗氧化作用增强粘液屏障功能,减轻乙酸的直接损伤。

Aim To build a simple experiment model of rat colon injury induced by acetic acid in vitro and to observe the effects of melatonin on this model. Methods On the basis of the establishment of rat colon injury model with acetic acid in vitro, we observed the colon mucosal damage caused by different concentrations of acetic acid （0,0. 1% ,0. 2% and 0. 4% ） for different time（ 5,10, 15,20,25 and 30 min ） , determined the content of lactic dehydrogenase （ LDH ） , malondiadehyde （ MDA ） and mucus in medium, and examined the histological changes of colon mucosa by Alacian Blue method. On the basis of the establishment of this model, the experiment was designed into normal group,model group（acetic acid of 0. 4% , time for 30 min） and melatonin treatment group（ the final concentration O. 1,1.0, 10 mmol · L^- 1 ）, and the indicators described above were detected to investigate the protective effects of melatonin. Results The LDH content of medium elevated gradually and the colon mucosal epithelial cells were injured by acetic acid in dose-and time-dependent manner, the mucosal edema, colon-wall depth and epithelium lose were observed at the same time, the MDA content of medium enhanced and mucus reduced correspondingly, but no significant change of the mucin expression in mucosal epithelial cells were observed. Pretreatment with melatonin reduced the release of LDH and MDA while promoted the secretion of mucus. Conclusion Colon mucosa of rats was injured by acetic acid in dose-and time-dependent manner in vitro. Acetic acid can impair the mucosal-mucus barrier by oxidating injured cell memberane. Melatonin can strengthen the barrier function of colon mucosa by its anti-oxidant action, attenuate the direct damage on colon mucosa of acetic acid.