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nadph对帕金森病中小胶质细胞吞噬α-突触核蛋白寡聚体的影响及其机制研究
首发时间：2024-06-21
摘要：目的：探究nadph对小胶质细胞吞噬α-突触核蛋白预制原纤维（α-synuclein preformed fibrils，pff）的作用及其分子机制。方法：（1）采用荧光微球（fluorescent microspheres）吞噬实验衡量小胶质细胞吞噬能力；通过免疫荧光分析不同浓度lps以及不同浓度nadph作用后吞噬荧光微球的小胶质细胞数量。（2）利用大肠杆菌体外表达、纯化α-synuclein单体，将单体经恒温振荡制备成α-syn pff。（3）通过免疫荧光和免疫印迹检测nadph处理后原代小胶质细胞和bv2细胞吞噬pff以及小胶质细胞对pff的降解能力。（4）采用免疫印迹和钙离子荧光探针分别检测nadph和pff作用后p2x7r蛋白表达以及激活情况；通过免疫印迹检测抑制剂或激动剂抑制或激动小胶质细胞p2x7r后小胶质细胞吞噬pff的作用。结果：（1）nadph能够逆转高浓度lps诱导的小胶质细胞吞噬能力降低，引起小胶质细胞对荧光微球的吞噬能力升高，并且呈浓度依赖性。（2）nadph促进小胶质细胞对pff的吞噬能力，并且小胶质细胞能够对pff进行有效降解。（3）nadph并不是通过影响p2x7r蛋白表达引起吞噬能力的改变；而是通过抑制p2x7r，促进小胶质细胞对pff的吞噬。结论：nadph能够缓解高浓度lps诱导的小胶质细胞吞噬能力降低。nadph通过抑制p2x7进而促进小胶质细胞对pff的吞噬能力。
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effects of nadph on microglial phagocytosis of α-synuclein oligers in parkinson\'s disease
abstract：objective:this study aimed to explore the effect of nadph on microglia phagocytosis of α-syn preformed fibrils (pff) and its molecular mechanism.methods:(1) use fluorescent microspheres phagocytosis experiment to measure the phagocytosis of microglia. the number of microglia phagocytosis fluorescent microspheres with different concentrations of lps and nadph was analyzed by immunofluorescence. (2)use the e.coli to produce the α-synuclein monomer and the monomer was purified. α-syn pff was prepared by constant temperature oscillation, (3) use the immunofluorescence and western blot to detect the phagocytosis of pff by primary microglia and bv2 cells after nadph treatment and the degradation ability of pff by microglia. (4) western blotting and ca2 fluorescence probes were used to detect the expression and activation of p2x7r after nadph and pff treatment.; used western blot to detect the phagocytosis of pff by microglia after inhibition or stimulation of p2x7r by inhibitors or agonists. results:(1) nadph can reverse the decrease in phagocytic ability of microglia induced by high concentration of lps, in a concentration-dependent manner. (2) nadph promoted the phagocytosis ability of microglia to pff and microglia could degrade pff effectively. (3) nadph and pff treatments did not affect the expression of p2x7r protein in microglia, and nadph and lps did not affect the expression of p2x7r protein in microglia, but nadphpromoted the phagocytosis of pff in microglia by inhibit the p2x7r.
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nadph对帕金森病中小胶质细胞吞噬α-突触核蛋白寡聚体的影响及其机制研究
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