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目的开展肺炎链球菌自溶酶(LytA)生物安全性的动物实验研究,为其申报生物制品鉴定提供实验数据。方法制备纯化的重组表达LytA蛋白,以6.5μg、13.0μg和19.5μg 3种剂量黏膜免疫BALB/c小鼠,根据《中华人民共和国国家标准食品安全性毒理学评价程序》(GB15193.1-1994)进行LytA的90 d喂养试验、传统致畸试验及遗传毒性试验检测,以分析其生物安全性。结果 90 d喂养试验结果显示各实验组小鼠与对照组小鼠的体重增长趋势一致,且各时间点检测的小鼠体重均无显著性差异(P>0.05)。传统致畸试验检测发现各实验组小鼠与对照组小鼠心脏、肺、肝脏、肾、脾、胃、子宫重量、红细胞和白细胞数目、谷丙转氨酶、尿素氮、总胆固醇、甘油三脂、血糖、总蛋白、白蛋白及球蛋白等指标均无明显差异(P>0.05)。小鼠骨髓嗜多染红细胞微核试验结果显示LytA经鼻腔黏膜免疫小鼠无遗传毒性。结论利用90 d喂养试验、传统致畸试验及遗传毒性试验证明LytA鼻腔黏膜免疫BALB/c小鼠安全、毒副作用不明显,有良好的生物安全性,值得进一步研究。
Objective To carry out animal experiments on the biosafety of Streptococcus pneumoniae lytA and provide experimental data for the identification of biological products. Methods Purified recombinant LytA protein was prepared and BALB / c mice were immunized with mucosal mucosa at doses of 6.5μg, 13.0μg and 19.5μg respectively. According to the “National Standard Food Safety Toxicology Evaluation Program of the People’s Republic of China” (GB15193.1-1994 ) For 90 days LytA feeding test, traditional teratogenicity test and genotoxicity test to analyze its biosafety. Results The results of 90-day feeding showed that the trend of body weight gain was consistent among mice in the experimental group and control group, and there was no significant difference in the body weight of the mice tested at each time point (P> 0.05). The results of traditional teratogenicity test showed that the weight of the heart, lung, liver, kidney, spleen, stomach and uterus, the numbers of red blood cells and white blood cells, alanine aminotransferase, urea nitrogen, total cholesterol, Blood glucose, total protein, albumin and globulin and other indicators were not significantly different (P> 0.05). Mouse bone marrow polychromatic erythrocyte micronucleus test results showed that LytA nasal mucosal immunization mice without genotoxicity. Conclusion 90 days feeding test, traditional teratogenicity test and genotoxicity test showed that BALB / c mice immunized with LytA nasal mucosa are safe and have no obvious toxic and side effects and good biosecurity, which deserves further study.