建立微小隐孢子虫感染小鼠模型方法的研究

Study on the optimal conditions to establish the mouse model infected with Cryptosporidium parvum oocysts

目的探索微小隐孢子虫传代和扩增模型小鼠的最佳性别、日龄、免疫抑制时间和免疫抑制剂地塞米松磷酸钠(DEXp)的最佳剂量。方法25、50、75日龄雌性和雄性BALB/c小鼠各3组,免疫抑制3 d后接种卵囊;各剂量组小鼠1 L饮水中含DEXp分别为1 mg、5 mg、10 mg、15 mg、20 mg、25 mg;各免疫抑制时间组小鼠分别在免疫抑制前6 d、3 d和免疫抑制后0 d、3 d、6 d、9 d感染卵囊。以上各组小鼠经口感染保存时间和方法相同的微小隐孢子虫卵囊的剂量均为2.0×105个,感染后逐日收集鼠粪,用饱和蔗糖漂浮法分离纯化卵囊。结果雌性小鼠排卵囊强度均较同日龄雄性小鼠大,而不同日龄小鼠中又以50日龄(26 g-28g)小鼠排卵囊量最多;各剂量组小鼠中,15 mg/L组排卵囊强度较其他各组大;免疫抑制时间组中,免疫抑制3 d感染组排卵囊强度最大。结论适合于卵囊扩增和传代的最佳鼠模型是50日龄(26-28g)雌性小鼠;DEXp的最佳剂量是15 mg/L;最佳免疫抑制时间是感染前3 d。

The aim of this study is to investigate various conditions that may influence the establishment of the mouse model infected with Cryptosporidium par＇oum oocytes, such as the optimal sex, age and body weight of mice and the time and dosage of application with immunosuppressive agent, such as dexamethasome phosphate （DEXp）. Three groups of male and female BALB/c mice of 25, 50 and 75 days of age were divided. The oocysts were inoculated to all groups of mice when they were under application of immunosuppressive agent for 3 days, and different concentrations of DEXp （ 1mg/L, 5 mg/L, 10 mg/L, 20 mg/L and 25 mg/L） were used to immunosuppress mice. On the 3 rd and 6th day before immunosuppression with DEXp as well as on the first, 3 rd, 6 th, 9 th day after immunosuppression, 6 groups of mice were inoculated orally with oocysts, and the dosage of inoculated oocysts in each group was 2. 0× 10^5 for each mouse, which were kept at the same time with the same method at 4℃ and in 2.5 % potassium dichromate. On the second day after infection, fecal materials of mice were collected, from which oocysts were isolated and purified with saturated sucrose flotation technique. It was found that the intensity of oocyst discharge from female mice was greater than those from male mice of the same age, and in the mice with different ages, the greatest amount of oocyst discharge occurred in mice of 50 days of age （body weight： 28-28 g）. Meanwhile, the greatest amount of oocyst discharges in group of mice with different dosages and different duration for immunosuppressor application occurred in a dose of 15 mg/L of DEXp and in mice under immunosuppression for 3 days respectively. It was demonstrated from the above observations that the optimal sex and age（weight） of mice to establish mouse model infected with C. parvum oocysts were female and 50 days old （body weight 26-28 g）, and the optimal dose of DEXp to be used and the duration of application with immunosuppressor are 15 mg/L of DEXp and the third day after application of immunosuppressor, respectively.