不同月龄BALB/c小鼠感染甲型H1N1型流感病毒的炎性调控差异研究

Regulation of Inflammatory Mediators in BALB/c Mice of Different Ages Infected with the Influenza A H1N1 Virus

为对比不同月龄BALB/c小鼠感染甲型H1N1型流感病毒后的淋巴细胞相对含量和炎症因子调控差异,提供流感类病毒性疾病防控的基础研究证据,本研究选用2月、8月龄雌性BALB/c小鼠,分别感染1×10^(5)TCID_(50)/0.1mL的甲型H1N1型流感病毒PR8毒株,每天记录体重、肛温变化,于感染第7d处死解剖取材,计算脏器指数,qPCR技术检测肺组织流感病毒M基因拷贝数,流式细胞术检测全血CD3^(+)、CD4^(+)、CD8^(+)淋巴细胞含量及肺组织中IL-6、IFN-γ、MCP-1等炎症因子的分泌水平。结果显示:(1)感染相同剂量病毒7d后,2月龄小鼠有1例样本死亡,生存质量下降较大,8月龄小鼠全部存活,体重较第0d相比仅下降12.35%,感染后2月龄组小鼠胸腺指数明显下降(P<0.01),8月龄组变化不明显。(2)2月龄组染毒小鼠肺组织M基因拷贝数显著低于8月龄染毒组(P<0.05)。(3)染毒后,2月龄组小鼠体内IFN-γ、IFN-β、IL-6等促炎性细胞因子含量增长显著(P<0.05),IL-6、MCP-1、IL-23含量显著高于8月龄组(P<0.05);2月龄染毒组小鼠杀伤性T淋巴细胞CD8+相对比例也明显高于8月龄染毒组(P<0.05)。可推测2月龄小鼠在感染病原体后产生了较强的免疫反应以清除病毒,但免疫系统的过度反应导致体内IL-6等炎症因子大量分泌,可能出现了细胞因子风暴现象致使机体损伤严重,使2月龄实验组小鼠较8月龄相比出现了更差的状态与较高的死亡率。本研究提示低年龄段人群感染相同剂量的流感病毒时较高年龄段人群可能会出现更严重的病理损伤。

In order to compare the pathologic changes,specific immune response and regulation of proinflammatory factors in different-aged BALB/c mice infected with the influenza A H1N1 virus,provide basic research evidence for the prevention and control of influenza-based diseases,this study chose two-month-old and8-month-old female BALB/c mice and infected them with the PR8 strain of the influenza A H1N1 virus(10^(5)TCID_(50)/0.1 mL).Changes in bodyweight and anal temperature were recorded each day.Samples were dissected and the organ index was calculated 7 days after infection.The viral load in lung tissue was detected by quantitative polymerase chain reaction.The contents of cluster of differentiation(CD)3^(+),CD4^(+),and CD8^(+) lymphocytes in whole blood and secretion of proinflammatory factors(e.g.,interleukin(IL)-6,interferon-γand monocyte chemoattractant protein(MCP)-1)in lung tissue were detected by flow cytometry.The results showed that after seven days identical-dosage-infection,one 2-month-old mouse died,and the quality of life of the other 2-month-old mice decreased greatly.All 8-month-old mice survived and their bodyweight decreased12.35%compared with that at day-0.The thymus index of 2-month-old mice decreased significantly after infection(P<0.01)while there was no significant change in the 8-month-old group.The viral load in the lung tissue of 8-month-old mice was significantly higher than that of 2-month-old mice(P<0.05).After exposure,the contents of interferon-γ,interferon-βand IL-6 in the 2-month-old group increased significantly,the contents of IL-6,MCP-1 and IL-23 in the 8-month-old group were significantly lower than those in the 2-month-old group,and the relative proportion of cytotoxic T lymphocyte CD8+in the 8-month-old group was significantly lower than that in the 2-month-old group.The results indicted that two-month-old mice produced a strong immune response to eliminate the virus after infection with pathogens.However,an overreaction of the immune system led to secretion of many proinflammatory factors(e.g.,IL-6),which could lead to serious injury caused by the“cytokine storm”and higher mortality in 2-month-old mice than that in 8-month-old mice.Our findings suggest that the younger age group infected with the same dose of an influenza virus may suffer more serious pathologic damage than the higher age group.