新生鼠坏死性小肠结肠炎模型建立方法的改进及评价

Improvement and Evaluation of Necrotizing Enterocolitis Models in Neonatal Rats

目的通过改进和比较国内、外常用的坏死性小肠结肠炎(Necrotizing Enterocolitis,NEC)动物模型建立方法,明确简便有效的NEC建模方法。方法采用出生2 h内新生SD大鼠,随机分为5组,对照组10只,实验组每组20只。A组与代母鼠同笼鼠乳喂养,且未行任何干预;B组人工喂养+缺氧冷刺激+(脂多糖)LPS灌胃(5 mg/kg体质量);C组人工喂养+缺氧冷刺激+LPS灌胃(10 mg/kg);D组人工喂养+缺氧冷刺激+LPS腹腔注射(2 mg/kg);E组人工喂养+缺氧冷刺激+LPS腹腔注射(5 mg/kg)。每日观察新生鼠的活动状况和体质量变化。实验结束后取小肠组织行苏木素-伊红染色、评价小肠病理损伤程度;检测小肠组织内肿瘤坏死因子α(TNF-α)水平。结果实验组新生鼠均出现不同程度的活动减少,腹胀腹泻,黑便,体质量减轻。病理评分显示实验组较对照组新生鼠病理损伤评分显著增高(P<0.05)。LPS腹腔注射组(D、E)较经口给药组(B、C)NEC发病率更高(P<0.05),但LPS大剂量(5 mg/kg)腹腔注射组较其他组,非NEC相关性致死率明显增高(P<0.05)。结论在人工喂养、缺氧冷刺激的基础上结合小剂量LPS(2 mg/kg)腹腔注射建立NEC的方法更具有可操作性、稳定性。

Objective The present study aimed to modify and evaluate necrotizing enterocolitis(NEC) models widely used in rats, and to determine the better method. Method Ninety rat pups were divided into five groups: Group A as control(n=10). Group B to E(n=20, respectively), rats were raised with simulated milk and subjected to hypoxia, cold stress, LPS with different ways and doses(Group B p.o. LPS 5 mg/kg;Group C p.o.LPS 10 mg/kg;Group D i.p. LPS 2.0 mg/kg;Group E i.p. LPS 5 mg/kg) per 12 hours for 72 hours. Followed by observing and recording body weight, survival time and sample collection, gross examination, HE staining, NEC injury scored, TNF-α levels in the intestine and kidneys were detected. Result We found that Group A rats were normal, but rats from groups B-E exhibited NEC clinical characteristics, but to varying degrees in different groups. The Group D(i.p.LPS 2.0 mg/kg) has comparatively lower mortality rate, no-NEC-associated death rate, higher morbidity rate and moderate TNF-α level in intestine and kidneys and NEC injury score when compared to other groups. Conclusion The modified method that through combination of formula feeding, hypoxia, cold stress, and low-dose LPS intraperitoneal injection is an optimal strategy of modeling rat pup NEC.