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目的 研究穿孔素 (PFP)缺陷小鼠对流感病毒感染及其免疫功能的影响。方法 PFP缺陷鼠 (pko)和对照B6鼠鼻腔内接种流感病毒A/PR/ 8/ 34 (H1N1)鼠肺适应株 ,测定LD50 和致死率。建立流感病毒肺炎模型 ,感染后 3、5、7、10、14、2 1d采取血清、脾脏和 /或肺。应用MDCK单层细胞培养测定肺匀浆病毒空斑数 (PFU) ,LDH释放试验测定脾细胞NK和CTL活性 ,并比较脾细胞培养诱生IFN γ、TNF α水平和血清血凝抑制效价。结果 pko和B6鼠流感病毒LD50 分别为 10 3 .50 和 10 3 .3 4PFU ,10 4 PFU所致病死率及Kaplan Meier生存曲线无统计差异 ,但pko鼠肺内病毒清除延迟。B6鼠感染后 3d ,NK活性明显升高。pko鼠感染前后则均无NK活性 ,感染后 7dCTL活性为B6鼠的 6 3.3%。pko和B6鼠感染后 3、7d ,脾细胞培养诱生IFN γ、TNF α水平及感染后 7、14、2 1d ,血清血凝抑制效价差异无显著性。结论 PFP参与了流感病毒感染的免疫控制过程 ,但并非唯一的主要免疫因素。pko鼠NK细胞毒活性缺失、CTL活性降低 ,但对淋巴细胞IFN γ、TNF α产生及血凝抑制效价的影响较小
Objective To study the effect of perforin (PFP) -deficient mice on influenza virus infection and its immune function. Methods PFP-deficient mice (pko) and control B6 mice were inoculated intranasally with influenza virus A / PR / 8/34 (H1N1) murine lung-adapted strain to determine LD50 and lethality. The model of influenza virus pneumonia was established, serum, spleen and / or lung were taken at 3, 5, 7, 10, 14, 21 days after infection. The number of plaque virus (PFU) in lung homogenate was measured by monolayer culture of MDCK. The activities of NK and CTL in spleen cells were determined by LDH release assay. The IFNγ, TNFα levels induced by splenocytes culture and the inhibition of serum hemagglutination were compared. Results The LD50 of pKO and B6 murine influenza virus were 103.50 and 103. 34 PFU respectively. There was no statistical difference in mortality and Kaplan Meier survival curves between 10 4 PFU and 10 pfu, but the virus clearance in pko mice was delayed. Three days after B6 infection, NK activity was significantly increased. There was no NK activity in pko mice before and after infection, and CTL activity in 7 days after infection was 63.3% in B6 mice. At 3 and 7 days after infection with pko and B6 mice, the levels of IFN-γ and TNF-α induced by spleen cell culture and the titer of serum hemagglutination inhibition were not significantly different at 7, 14 and 21 days after infection. Conclusion PFP is involved in the immune control of influenza virus infection, but it is not the only major immunological factor. The NK cell cytotoxic activity of pko mice was absent and the activity of CTL was decreased, but the effects on IFNγ, TNFα production and hemagglutination inhibition titers of lymphocytes were less