高压氧对大鼠溃疡性结肠炎自噬及炎症反应的影响

Experimental research on the effects of hyperbaric oxygen on ulcerative colitis in rats

目的 探讨高压氧（hyprbaric oxygen,HBO）对大鼠溃疡性结肠炎（ulcerative colitis,UC）自噬及炎症反应的作用.方法 SPF级Wistar大鼠30只,按照数字表法随机分为3组：对照组、模型组和HBO组,每组10只.采用2,4,6-三硝基苯磺酸（three nitrobenzene sulfonic acid,TNBs）诱导实验性大鼠UC模型,HBO治疗14 d后,观察各组大鼠死亡情况,并解剖各组大鼠UC肠组织,计算其结肠指数及溃疡面积.应用HE染色观察各组大鼠结肠组织病理形态,Western blotting检测病变肠黏膜组织中自噬相关基因Beclin1及微管相关蛋白1轻链3Ⅱ（light chain 3 Ⅱ,LC3Ⅱ）表达水平,ELISA检测血清白细胞介素（interleukin,IL）-4、5、6、10及肿瘤坏死因子-α（tumor necrosis factor-α,TNF-α）活性的影响.结果 模型组黏膜表面起伏不平,可见大量炎性细胞浸润,腺体萎缩,可见溃疡面形成,黏膜下层大量炎性浸润;HBO组结肠黏膜仍可见炎性细胞浸润,但明显少于模型组. 与对照组比较,模型组大鼠结肠指数、溃疡面积均显著升高,体质量显著降低,差异有统计学意义（P ＜0.01）;与模型组比较,HBO组大鼠结肠指数和溃疡面积均显著降低,体质量显著升高,差异均有统计学意义（P＜0.05） 与对照组比较,模型组大鼠IL-4、IL-5、IL-6及IL-10水平显著降低,而TNF-α水平显著升高,差异均有统计学意义（P＜0.01）;与模型组比较,HBO组大鼠IL-4、IL-5、IL-6、IL-10表达水平显著升高,而TNF-α水平显著降低,差异均有统计学意义（P＜0.05）.与对照组和HBO组比较,模型组大鼠Beclin1及LC3Ⅱ表达水平明显降低,差异有统计学意义（P＜0.01） .结论 HBO可明显缓解大鼠UC结肠组织的损伤,其作用机制可能与炎症反应的抑制和自噬水平的上调相关.

Objective To explore the effects of hyperbaric oxygen （HBO） on autophagy and inflammation of ulcerative colitis （UC） rats.Methods SPF grade Wistar rats were randomly divided into 3 groups：the control group,the model group and the hyperbaric oxygen （HBO） group,each consisting of 10 animals.The ulcerative colitis （UC） rat model was established by using 2,4,6,trinitrobenzene sulfonic acid （TNBs）.Fourteen days after HBO therapy,mortality rates of each group were evaluated and then anatomy of UC colon tissue was performed in all the groups,and colon index and size of colitis were calculated.HE staining was used to observe the pathological morphology of colon tissue in various groups.Western blotting was performed to assess the expression levels of autophagy-related proteins in intestinal mucosa tissue,such as Beclin1 and microtubule-associated protein 1 light chain 3 Ⅱ （LC3Ⅱ）.ELISA was conducted to detect effects on serum IL-4,5,6 and 10,as well as tumor necrosis factor-α （TNF-α）.Results The mucosal surface of the model group was uneven,displaying infiltration by large quantities of inflammatory cells and gland atrophy.Ulcer surface was also seen,and there was infiltration by a large number of inflammatory cells below submucosa.The inflammatory cell infiltration in the colonic mucosa of the HBO group was significantly less than that of the model group.Compared with those of the model group,colon index and ulcer size of the HBO group significantly withered,and body weight significantly increased,and statistical significance could be noticed,when comparisons were made between them （P ＜ 0.05）.Compared with those of the control group,the levels of IL-4,IL-5,IL-6 and IL-10 in the model group were significantly decreased,while TNF levels were significantly increased,and statistical significance could also be seen,when comparisons were made between them （P ＜0.05）.As compared with those of the model group,the levels of IL-4,IL-5,IL-6,IL-10 in the HBO group increased considerably,while TNF-α expression level was significantly reduced,also with statistical significance （P ＜ 0.05）.Western blotting detection results showed that compared with those of the control group and the HBO group,the expression levels of Beclin1 and LC3Ⅱ were significantly lower than those of the model group,also with statistical significance （P＜0.01）.Conclusion HBO could obviously alleviate colon tissue damage of UC rats,and the possible mechanism of which might be related to the inhibition of inflammatory reaction and the improvement of autophagy.