Aβ_(42)蛋白与胶质细胞吞噬相关受体相互作用的研究

Research on the interaction of Aβ_(42) and phagocyticrelated receptors in microglia

目的探讨寡聚形态Aβ_(42)蛋白与胶质细胞吞噬相关受体SRB、CD36的体外结合情况,以及上述受体封闭状态下或内吞抑制剂存在条件下该细胞吞入Aβ_(42)数量的变化规律。查明富寡聚物Aβ_(42)(o Aβ_(42))环境对小胶质细胞的吞噬相关受体SRB、CD36、TNF-α、TNF-R基因表达量的影响。方法用酶联免疫法测定o Aβ_(42)与胶质细胞SRB、CD36受体的结合力。用SRB、CD36受体的功能性抗体,受体内吞(巨吞饮)抑制剂作用细胞,检测BV2细胞对o Aβ_(42)吞噬能力的变化,反转录法测定o Aβ_(42)影响下的BV2小胶质细胞吞噬相关基因表达量变化。结果 o Aβ_(42)与胶质细胞清道夫家族SRB、CD36受体在体外实验中特异性结合,SRB受体的阻断使小胶质细胞内吞o Aβ_(42)的能力显著下降,CD36受体阻断剂对细胞内吞量无显著影响,受体内吞(巨吞饮)抑制剂增加细胞对o Aβ_(42)摄取的量。o Aβ_(42)对人脑小胶质细胞吞噬相关受体的表达量有显著影响。结论 o Aβ_(42)与胶质细胞吞噬相关受体的相互作用可进一步影响细胞对病理蛋白的吞入和清除过程。该结论可为从分子水平调节胶质细胞与Aβ_(42)相互作用的受体激活剂,基因封闭剂等AD免疫疗法提供参考依据。

Objective To investigate the binding affinity between oligomer Aβ_（42）（o Aβ_（42）） and phagocytic-related receptors（SRB, CD36） in glia cells, assure the internalization pattern of Aβ_（42） in phagocytic receptors（SRB, CD36）-blocked and endocytosis-blocked microglia, and clarify the expression of phagocytosis-related receptor（SRB, CD36, TNF-α, TNF-R） after o Aβ_（42） incubation in microglia.Methods ELISA assay was used to detect the binding affinity between o Aβ_（42） and glia SRB/CD36 receptor in vitro. Receptors neutralization and endocytosis/macropinocytosis blockade was used to detect receptor blocking effect on o Aβ_（42） internalization process. Reverse transcription was used to assay the fold change of phagocytosis-related receptor expression in o Aβ_（42）-treated microglia. Results SRB and CD36 receptor showed high affinity with o Aβ_（42） in glia cells. Blockade of SRB receptor, but not CD36 receptor can decrease internalization of o Aβ_（42）. o Aβ_（42） incubation displayed remarkable effect on expression of phagocytosis-related receptors in BV2 cells. Conclusions Interaction between o Aβ_（42） and phagocytosis-related receptors in glia cells could influence the internalization and clearance of pathological protein in disease process. These findings provide reference to AD immunotherapy of receptor activator or gene blocker at molecular level.