目的 评价复方谷氨酰胺肠溶胶囊（CGEC）对Beagle犬化疗性肠炎模型的影响。方法 Beagle犬随机分为对照组、模型组、美沙拉嗪肠溶片（MECT，阳性药，每只500 mg）组和CGEC低、中、高剂量(48、96、192 mg·kg^-1)组，各组Beagle犬ig给予相应剂量药物，对照组和模型组分别给予与高剂量组同等剂量的肠溶明胶空心胶囊，每天1次，连续7 d。除对照组外，其余各组Beagle犬均于给药第3天开始连续5 d ip 8 mgk·g^-1 5-氟尿嘧啶（5-FU）诱导肠炎模型，对照组给予等体积的0.9%氯化钠注射液。每天观察各组Beagle犬的一般状况表现；于给药后第7天称质量，观察动物摄食量；苏木精-伊红（HE）染色观察回肠组织结构的病理变化，采用BX43+MD50数码医学图像分析系统测定回肠绒毛高度及隐窝深度；试剂盒法检测回肠组织白细胞介素（IL）-1β、肿瘤坏死因子（TNF）-α的水平；免疫组化检测回肠组织中增殖细胞核抗原（PCNA）和G蛋白偶联受体5（Lgr5）表达；体外培养人骨髓间充质干细胞（HMSC）至第3代，诱导分化为肠道干细胞，分为对照组、模型组、CGEC(10、20、40 mg·mL^-1)组、CGEC(20 mg·mL^-1)+FH535[Wnt/β-连环蛋白（β-catenin）抑制剂，50μmol·L^-1]组，除对照组外，其余各组均在给药同时添加3μg·mL^-1 5-FU造模；48 h后通过Western blotting实验检测Lgr5、β-catenin、Wnt1、糖原合成酶激酶-3β（GSK3β）的蛋白表达。结果 模型组动物造模第1～3天呕吐大量未消化的食糜或黄色黏稠液体，摄食量减少，自主活动减少，粪便呈水样便，造模第4天粪便呈脓血水样便；CGEC中、高剂量组动物自主活动减少，摄食量减少，呕吐少量未消化的食糜。模型组回肠黏膜上皮变性、坏死，黏膜层充血，肠腺上皮细胞变性、坏死，CGEC各组可明显缓解上述症状。与模型组比较，CGEC各剂量组动物体质量显著增加，摄食量显著增加（P<0.01），结肠组织IL-1β、TNF-α含量均显著降低（P<0.01）；中、高剂量组隐窝深度与绒毛高度的比值显著增加（P<0.01）；免疫组化结果显示，CGEC低、中、高剂量能够显著增加回肠组织中PCNA的表达（P<0.01），中、高剂量能够显著增加回肠组织中Lrg5的表达（P<0.05、0.01）；细胞实验结果显示，与模型组比较，CGEC 20、40 mg·mL^-1均能够增加Lgr5、β-catenin、Wnt1和GSK3β的蛋白表达（P<0.05、0.01）；当联用Wnt/β-catenin抑制剂时，能够显著削弱CGEC(20 mg·mL^-1)对Lgr5、β-catenin、Wnt1和GSK3β的促进作用（P<0.05、0.01）。结论 CGEC对化疗性肠炎具有显著治疗作用，且Wnt/β-catenin信号通路介导了其对肠黏膜的保护作用。

Objective To evaluate the effect of Compound Glutamine Enteric-coated Capsules（CGEC） on the chemotherapy-induced enteritis model in Beagle dogs. Methods Beagle dogs were randomly divided into the control group, the model group, the mesalazine enteric-coated tablets（MECT, positive drug, 500 mg per dog） group, and the low-, medium-, and high-dose CGEC(48, 96, and 192 mg·kg^-1) groups. Each group of Beagle dogs was ig administered the corresponding dose of the drug, and the control group and the model group were respectively given the same dose of enteric-coated gelatin capsules as the high-dose group, once a day for 7 consecutive days. Except for the control group, the other groups of Beagle dogs were intraperitoneally（ip） administered 8 mg·kg^-1 5-FU to induce the enteritis model from the third day of administration for 5 consecutive days, while the control group was given the same volume of 0.9% sodium chloride injection. The general condition of each group of Beagle dogs was observed daily; on the 7 th day after administration, the body weight was measured, and the food intake was observed. The pathological changes of the ileal tissue structure were observed by hematoxylin-eosin（HE） staining, and the height of the ileal villi and the depth of the crypts were measured by BX43 + MD50 digital medical image analysis system. The levels of interleukin（IL）-1β and tumor necrosis factor（TNF）-α in the ileal tissue were detected by kit method. The expression of proliferating cell nuclear antigen（PCNA） and G protein-coupled receptor 5（Lgr5） in the ileal tissue was detected by immunohistochemistry. Human bone marrow mesenchymal stem cells（HMSC） were cultured in vitro to the third generation and induced to differentiate into intestinal stem cells, and were divided into the control group, the model group, the CGEC(10, 20, 40 mg·mL^-1) groups, and the CGEC(20 mg·mL^-1) + FH535 [Wnt/β-catenin inhibitor, 50 μmol·L^-1] group. Except for the control group, the other groups were simultaneously administered 3 μg·mL^-1 5-FU to establish the model during administration. After 48 h, the protein expression of Lgr5, β-catenin, Wnt1, and glycogen synthase kinase-3β（GSK3β） was detected by Western blotting. Results On the first to third days of modeling, the model group of animals vomited a large amount of undigested chyme or yellow viscous liquid, had reduced food intake, decreased spontaneous activity, and watery stools. On the fourth day of modeling, the stools were purulent bloody water. In the medium-and high-dose CGEC groups, the animals had decreased spontaneous activity, reduced food intake, and vomited a small amount of undigested chyme. The ileal mucosal epithelium of the model group was degenerated and necrotic, the mucosal layer was congested, and the intestinal gland epithelial cells were degenerated and necrotic. CGEC groups could significantly alleviate the above symptoms. Compared with the model group, the body weight of the animals in each dose group of CGEC significantly increased, and the food intake significantly increased（P < 0.01）. The content of IL-1β and TNF-α in the colonic tissue was significantly decreased（P < 0.01）. The ratio of crypt depth to villus height in the medium-and highdose groups was significantly increased（P < 0.01）. Immunohistochemical results showed that low-, medium-, and high-dose CGEC could significantly increase the expression of PCNA in the ileal tissue（P < 0.01）, and medium-and high-dose CGEC could significantly increase the expression of Lgr5 in the ileal tissue（P < 0.05, 0.01）. Cell experiments showed that compared with the model group, CGEC 20 and 40 mg·mL^-1 could increase the protein expression of Lgr5, β-catenin, Wnt1, and GSK3β（P < 0.05, 0.01）. When combined with the Wnt/β-catenin inhibitor, the promoting effect of CGEC(20 mg·mL^-1) on Lgr5, β-catenin, Wnt1, and GSK3β was significantly weakened（P < 0.05, 0.01）. Conclusion CGEC has a significant therapeutic effect on chemotherapy-induced enteritis, and the Wnt/β-catenin signaling mediates its protective effect on intestinal mucosa.

R965

湖南省自然科学基金资助项目(2024JJ8140)