Role of autophagy in Japanese encephalitis viral (JEV) infection is not well known. In the present study, we reported the role of autophagy flux in microglia activation, neurobehavioral function and neuronal death using a mouse model of JEV. Markers for autophagy (LC3-II/I, SQSTM1/P62, phos-Akt, phos-AMPK), and neuronal death (cleaved caspase 12, H2Ax, polyubiquitin) were investigated by western blot at 1, 3 and 7 days post inoculation. Cathepsin D was measured in cerebral cotex of JEV infected mice spectrophotometrically. Microglia activation and pro-inflammatory cytokines (IL1β, TNF-α, IFNγ, IL6) were measured by immunohistochemistry, western blot and qPCR analysis. In order to determine the neuroinflammatory changes and autophagy mediated neuronal cell death, BV2-microglia and N2a-neuronal cells were used. Autophagy activation marker LC3-II/I and its substrate SQSTM1/P62 were significantly increased while cathepsin D activity was decreased on day 7 post inoculation in cerebral cortex. Microglia in cortex were activated and showed higher expression of proinflammatory mRNA of IL1β, TNF-α, IFNγ and IL6, with increased DNA damage (H2AX) and neuronal cell death pathways in hippocampus and neurobehavioral dysfunction. Similar observations on JEV infection mediated autophagy flux inhibition and neuronal cell death was found in N2a neuronal cell. Collectively, our study provides evidence on the role of autophagy regulation, microglial activation and neurodegeneration following JEV infection.

自噬在日本脑炎病毒（JEV）感染中的作用尚不清楚。在本研究中，我们报道了使用JEV小鼠模型自噬通量在小胶质细胞激活，神经行为功能和神经元死亡中的作用。在接种后1、3和7天，通过蛋白质印迹研究了自噬的标志物（LC3-II / I，SQSTM1 / P62，phos-Akt，phos-AMPK）和神经元死亡（裂解的胱天蛋白酶12，H2Ax，多泛素）。用分光光度法在感染JEV的小鼠的大脑皮质中测量组织蛋白酶D。通过免疫组织化学，western印迹和qPCR分析测量小胶质细胞的活化和促炎细胞因子（IL1β，TNF-α，IFNγ，IL6）。为了确定神经炎性变化和自噬介导的神经元细胞死亡，使用了BV2-小胶质细胞和N2a-神经元细胞。自噬激活标记LC3-II / I及其底物SQSTM1 / P62显着增加，而脑皮层接种后第7天组织蛋白酶D活性降低。皮层中的小胶质细胞被激活，并显示出IL1β，TNF-α，IFNγ和IL6的促炎性mRNA的较高表达，并增加了海马的DNA损伤（H2AX）和神经元细胞死亡途径以及神经行为功能障碍。在N2a神经元细胞中发现了对JEV感染介导的自噬通量抑制和神经元细胞死亡的类似观察。总的来说，我们的研究为JEV感染后自噬调节，小胶质细胞活化和神经变性的作用提供了证据。皮层中的小胶质细胞被激活，并显示出IL1β，TNF-α，IFNγ和IL6的促炎性mRNA的较高表达，并增加了海马的DNA损伤（H2AX）和神经元细胞死亡途径以及神经行为功能障碍。在N2a神经元细胞中发现了对JEV感染介导的自噬通量抑制和神经元细胞死亡的类似观察。总的来说，我们的研究为JEV感染后自噬调节，小胶质细胞活化和神经变性的作用提供了证据。皮层中的小胶质细胞被激活，并显示出IL1β，TNF-α，IFNγ和IL6的促炎性mRNA的较高表达，并增加了海马的DNA损伤（H2AX）和神经元细胞死亡途径以及神经行为功能障碍。在N2a神经元细胞中发现了对JEV感染介导的自噬通量抑制和神经元细胞死亡的类似观察。总的来说，我们的研究为JEV感染后自噬调节，小胶质细胞活化和神经变性的作用提供了证据。