家兔膀胱黏膜下注射卡介菌多糖核酸的免疫效应

Immune response after injecting polysaccharide nucleic acid of Bacillus Calmette-Guerin under mucous membrane of bladder in rabbits

目的:探讨家兔膀胱黏膜下注射卡介菌多糖核酸(polysaccharide nucleic acid fraction of Bacillus Calmette-Guerin,BCG-PSN)所致的免疫效应变化,并寻求最合适注射的药物剂量。方法:将实验家兔随机分成5组,分别为BCG-PSN大、中、小(0.35、0.175、0.0875 mg/ml)剂量膀胱黏膜下注射组,BCG(0.35 mg/ml)膀胱黏膜下注射对照组和BCG-PSN(0.35 mg/ml)膀胱灌注对照组。分别在用药前和用药后1周、2周、1和3个月应用流式细胞术和酶联免疫吸附测定法(ELISA)对家兔外周血中T淋巴细胞亚群(CD4^+、CD8^+)和细胞因子(IL-2、TNF-α、IFN-γ)进行测定;通过H-E、Masson染色对家兔膀胱壁组织进行病理检查。结果:(1)注射BCG-PSN各组CD4^+、CD8^+ T细胞的数量随用药剂量增加而增加,同时随时间延长逐步增加,用药后2周达到高峰,用药后3个月基本恢复至原水平;BCG注射对照组变化趋势与之相同,BCG-PSN灌注对照组的CD4^+ T细胞数量明显低于大剂量BCG-PSN注射组(P<0.05),各组的CD4^+/CD8^+变化趋势相似。(2)注射BCG-PSN各组的细胞因子(IL-2、TNF-α、IFN-γ)变化与CD4^+、CD8^+ T细胞的变化趋势相同,BCG-PSN灌注对照组的细胞因子数最明显低于大、中剂量BCG-PSN注射组(P<0.05)。(3)各用药组膀胱黏膜下均见大量淋巴细胞浸润,BCG注射对照组家兔膀胱壁组织出现纤维化改变。结论:膀胱黏膜下注射BCG-PSN所致的免疫效应与注射BCG相似,优于BCG-PSN膀胱灌注,且不引起膀胱壁纤维化;以0.35 mg/ml注射效果最明显。

Objective： To investigate the immune response after injecting polysaccharide nucleic acid of Bacillus Calmette-Guerin （BCG-PSN） under the mucous membrane of bladder in rabbits and to search for the most suitable dose of BCG-PSN. Methods： The rabbits were randomly divided into 5 groups： high dose （0.35 mg/ml） BCG-PSN group, middle dose （0. 175 mg/ml） BCG-PSN group, low dose （0. 0875 mg/ml） BCG-PSN group, BCG （0.35 mg/ml） control group and BCG-PSN （0.35 mg/ml） intra-bladder perfusion control group. T lymphocyte subsets （ CD4^＋ , CD8^＋ ） in the peripheral blood were determined by flow cytometry before and 1 week ,2 weeks, 1 month and 3 months after the treatment; the levels of IL-2, TNF-α, and IFN-γ of peripheral blood were determined by enzyme-linked immunosorbent assay ; H-E and Masson staining was used for the pathological examination of the bladder. Results： （1） BCG-PSN injection increased the number of CD4^＋ and CD8^＋ T lymphocytes in a time- and dose-dependent manner, with the peak numbers appeared 2 weeks after BCG-PSN injection ; the numbers restored to the normal levels 3 months after BCG-PSN injection. BCG control group had a similar changing pattern to the BCG-PSN injection group. The number of CD4^＋ T cells BCG-PSN perfusion group was significantly lower than that in the high-dose BCG-PSN injection group （ P 〈0.05 ）. The changing tendencies of CD4^＋/CD8^＋ were similar in both groups. （2） The changes of IL-2, TNF-α, and IFN-γ/ levels in BCG-PSN injection groups were similar to those of CD4^＋ and CD8^＋ T cells; and the levels in BCG-PSN perfusion group were significantly lower than those of the high- and median-dose of BCG-PSN injection groups （ P 〈 0. 05 ）. （3） T lymphocytes infiltration of mucous memberance uncer bladder was observed in all groups after the treatment. The BCG injection control group had fibrosis of the bladder wall. Conclusions： The immune response of BCG-PSN injection under the mucous membrane o bladder can cause similar immune response to that of BCG; the response is superior to that of BCG-PSN perfusion; and BCG-PSN causes no bladder wall fibrosis. The optimal dose of BCG-PSN is 0.35 mg/ml.