ADSCs-CM对MPP+诱导的SH-SY5Y细胞神经保护作用及其机制

NEUROPROTECTIVE EFFECT OF ADSCs-CM ON MPP+-INDUCED SH-SY5Y CELLS AND ITS MECHANISM OF ACTION

目的 探讨人脂肪间充质干细胞条件培养基(ADSCs-CM)对1-甲基-4-苯基吡啶(MPP+)诱导的SH-SY5Y细胞神经保护作用及其机制。方法 将SH-SY5Y细胞分为Ctrl组、PD组、CM组,Ctrl组置于基础培养基中培养,PD组于MPP+(1.0 mmol/L)+基础培养基中培养,CM组于MPP+(1.0 mmol/L)+ADSCs-CM培养,均培养24 h。分别使用活性氧(ROS)试剂盒、三磷酸腺苷(ATP)试剂盒、线粒体复合物Ⅰ(CⅠ)活性试剂盒检测各组SH-SY5Y细胞中ROS含量、ATP总量和线粒体CⅠ活性;使用单丹磺酰尸胺(MDC)荧光法观察各组SH-SY5Y细胞自噬率;采用Western blot方法检测各组SH-SY5Y细胞线粒体自噬相关蛋白P62、LC3的表达水平。结果 PD组与Ctrl组、CM组比较,SH-SY5Y细胞的ROS含量、ATP总量、线粒体CⅠ的活性、细胞自噬率、P62、LC3Ⅱ/Ⅰ蛋白相对表达量比较差异均有显著性(tLSD=3.23~12.61,P<0.05)。结论 ADSCs-CM可改善MPP+诱导的SH-SY5Y细胞中线粒体的功能障碍,其作用机制可能是通过影响自噬相关蛋白P62、LC3的表达实现的。

Objective To investigate the neuroprotective effect of human adipose-derived mesenchymal stem cells-conditioned medium(ADSCs-CM)on 1-methyl-4-phenylpyridine(MPP+)-induced SH-SY5Y cells and the underlying mechanism.Methods SH-SY5Y cells were divided into three groups for 24 h culture:control group(basic culture medium),PD group(1.0 mmol/L MPP++basic culture medium),and CM group(1.0 mmol/L MPP++ADSCs-CM).The content of reactive oxygen species(ROS)in SH-SY5Y cells was determined using a ROS assay kit.The total amount of adenosine triphosphate(ATP)in SH-SY5Y cells was measured using an ATP assay kit.The activity of mitochondrial complexⅠ(CⅠ)in SH-SY5Y cells was determined using a mitochondrial CⅠactivity assay kit.The autophagy rate of SH-SY5Y cells was observed by the monodansylcadave-rine fluorescence method.Western blot was used to measure the expression of mitochondrial autophagy-related proteins(P62 and LC3)in SH-SY5Y cells.Results Compared with the control and CM groups,the PD group showed significant differences in ROS content,total ATP amount,mitochondrial CⅠactivity,autophagy rate,and the relative expression levels of P62 and LC3Ⅱ/Ⅰproteins in SH-SY5Y cells(t LSD=3.23-12.61,P<0.05).Conclusion ADSCs-CM can improve MPP+-induced mitochond-rial dysfunction in SH-SY5Y cells,possibly through influencing the expression of autophagy-related proteins P62 and LC3.