瘦素受体敲除引起大鼠脑组织小胶质细胞活化

Leptin receptor knockout induced microglial cell activation in rats

目的本文利用瘦素受体(LEPR)敲除大鼠,分析瘦素受体基因敲除后大鼠脑中小胶质细胞形态与功能的改变,探究瘦素受体在小胶质细胞中的功能作用。方法采用RT-PCR,蛋白印迹法,免疫组化法和免疫荧光法,观察大鼠瘦素受体敲除后体内外小胶质细胞活化状态。结果瘦素受体在小胶质细胞表达,基因敲除可以完全剔除小胶质细胞中LEPR蛋白;LEPR敲除增强大鼠LPS对刺激的炎症反应,存活率降低了75%;LEPR敲除大鼠脑中的活化的小胶质细胞比例明显增加;LEPR敲除的原代小胶质细胞不仅分泌更多炎性因子也增强了吞噬能力;Western blot发现PI3K/AKT在瘦素受体敲除大鼠脑组织蛋白中磷酸化明显增强。结论瘦素受体敲除后,大鼠小胶质细胞向促炎促吞噬的方向发展,揭示了LEPR/Leptin可能通过小胶质细胞调节神经炎症。

Objective Neuroinflammation is a phenotype of leptin receptor(LEPR)mutated mice(db/db)and raises the question of whether LEPR is involved in microglial activation.To examine the function of LEPR on microglia cells,microglial activation was comparatively analyzed in LEPR+/+and LEPR-/-rats.Methods RT-PCR,Western blot,immunohistochemistry,and immunofluorescence were used to examine microglia morphology,inflammatory factor secretion,and sensitivity to lipopolysaccharide(LPS)treatment in vitro and in vivo.Results First,LEPR was expressed in microglia from LEPR+/+rats,while LEPR protein was completely deleted in microglia from LEPR-/-rats.LEPR deletion enhanced sensitivity to LPS treatment and reduced survival rate of LEPR-/-rats by 75%.Activated microglia were significantly increased in brain tissue of LEPR-/-rats compared with LEPR+/+rats.LEPR deletion also increased the expression of inflammatory cytokines including interleukin(IL)-6,inducible nitric oxide synthase,and Interleu bin-1β(IL-1β),and enhanced phagocytotic ability.Phosphatidylinositol-3-kinase(PI3K)/AKT phosphorylation was significantly increased in microglia from LEPR-/-rats compared with LEPR+/+rats,suggesting that activation of the PI3K/AKT signal pathway could partly be the mechanism of microglia activation.Conclusions Deletion of LEPR in microglia induced its activation and suggests the involvement of neuroinflammation in rats.Our result also suggest that the LEPR/leptin axis plays important roles in neuroinflammation through microglia.