敌草快中毒性肠病动物模型的建立与鉴定

Establishment and identification of animal model of toxic intestinal disease by diquat

关键词: 敌草快(DQ)  灌胃  肠道损伤  动物模型

Keywords: diquat(DQ)  gavage  intestinal injury  animal model

摘 要: 目的 建立更为合适、操作简便且结果稳定的敌草快(DQ)中毒肠道损伤模型。方法 选择 8~10 周龄雄性 C57BL/6J 小鼠, 随机分为对照组、低剂量染毒组(7 日总剂量 100 mg/kg) 和高剂量染毒组(7 日总剂量 200 mg/kg), 每组 3 只小鼠,所有小鼠按 8 μl/g 给予一次性灌胃染毒,对照组给予氯化钠注射液,低、高剂量组 DQ 浓度分别为 12.5 mg/ml 和 25.0 mg/ml。 观察小鼠一般情况,分析小鼠结肠大体形态、体质量、肠道组织病理学、超微结构(电镜)及氧化应激指标(T-AOC、 MDA)变化。结果 (1)一般情况: 与 100 mg/kg DQ 组相比, 200 mg/kg DQ 组小鼠早期即出现精神萎靡、反应迟钝、攻击力下降等表现,小鼠 7 d 存活率更低。(2)结肠大体形态:对照组小鼠结肠表面光滑、半透明,含清晰粪团; 100 mg/kg DQ 组结肠呈淡黄色,透明度下降,伴少量稀便; 200 mg/kg DQ 组结肠长度显著缩短(P<0.001),表面出现散在出血点,管腔内充满稀便。(3)组织病理学:光镜下,染毒第 2 天 200 mg/kg DQ 组小鼠空肠即出现绒毛排列紊乱、肌层变薄及炎性细胞浸润,100 mg/kg DQ 组损伤出现较晚(第 4 天)。电镜显示, 200 mg/kg DQ 组小鼠结肠微绒毛出现脱落融合、细胞间隙增宽,肠道物理屏障严重受损。(4)血清氧化应激标志物: 200 mg/kg DQ 组血清总抗氧化能力(T-AOC)在染毒第 1、3、5 天显著升高(P<0.05),丙二醛(MDA)水平于第 3 天升高。结论 一次性灌胃染毒可以成功构建敌草快中毒性急性肠道损伤小鼠模型,其中 200 mg/kg DQ 组可诱导结肠缩短、微绒毛脱落、炎性细胞浸润及氧化应激激活等典型病理特征。

Abstract: Objective To establish a more suitable, easy to operate, and stable intestinal injury model caused by acute poisoning of diquat. Methods Male C57BL/6J mice aged 8—10 weeks were randomly divided into control group, low dose groups (with a total dose of 100mg/kg for 7 days), and high dose group (with a total dose of 200 mg/kg for 7 days), 3 mice in each group, the mice received a single gavage dose of 8 μl/g experimental solution, the control mice received saline only, while the mice of poisoned groups were received 12.5 mg/ml (low dose group)or 25.0 mg/ml (high dose group)DQ solution. Then, observe the abnormal changes of mice such as general conditions, body weight, and colon morphology including histopathological changes, ultrastructure (electron microscopy), and the changes in oxidative stress markers (T-AOC, MDA)as well. Results The results showed that(1)General condition: compared with the low dose DQ exposure group (100 mg/kg), mice of the high dose group (200 mg/kg)showed early manifestations such as mental fatigue, delayed response, and decreased aggression, and had a lower 7-day survival rate. (2)Gross morphology of colon: the colon surface of the control group mice was smooth, semi-translucent and contained clear fecal masses; in 100 mg/kg DQ group, the colon appeared pale yellow with decreased transparency, with small amount of loose stools; while in the mice of 200 mg/kg DQ group, the colon length was significantly shortened (P<0.001), with scattered bleeding points appearing on the surface and the lumen was filled with loose stools. (3) Histopathology: under light microscope, on the 2nd day of DQ exposure, the mice of 200 mg/kg DQ group showed disordered villus arrangement, thinning of muscular layer, and inflammatory cell infiltration in the jejunum, whereas the mice of 100 mg/kg group those changes mentioned above appeared later (on the 4th day). Electron microscope observation revealed that the colon microvilli in the mice of 200 mg/kg DQ group had shed and fused, widened intercellular spaces, and severely damaged intestinal physical barriers. (4)Serum oxidative stress markers: the serum total antioxidant capacity (T-AOC) in the mice of 200 mg/kg DQ group was significantly increased on the 1st, 3rd, and 5th days after DQ exposure (P<0.05), and malondialdehyde (MDA)level increased on the 3rd day as well. Conclusion The results suggested that a one-time gavage could successfully establish a DQ induced toxic acute intestinal injury mouse model, and the 200 mg/kg DQ group could induce typical pathological features such as colon shortening, microvillus shedding, inflammatory cell infiltration, and oxidative stress activation.