摘要
目的探讨低密度脂蛋白受体相关蛋白2(LRP2)在β淀粉样蛋白(Aβ)诱导的Alzheimer’s病(AD)细胞模型中的表达情况,及调控LRP2表达后对神经元存活影响及其对丝裂原活化蛋白激酶(MAPKs)信号通路的影响。方法采用20μmol Aβ1-42处理的神经母细胞瘤(SH-SY5Y)细胞作为AD细胞模型。采用Western blotting法检测LRP2蛋白表达情况。Scrambled siRNA和LRP2 siRNA(si LRP2)分别转染SH-SY5Y细胞48 h,随后予以Aβ1-42处理24 h,采用噻唑蓝比色法检测细胞活力,流式细胞仪检测细胞凋亡情况,Western blotting法检测ERK、p38及JNK的磷酸化水平。结果si LRP2+Aβ组细胞活力显著低于SC+Aβ组(P<0.05)。与处理前比较,Aβ1-42处理后LRP2蛋白表达水平显著降低(P<0.05)。与空白对照组比较,siRNA-1组、siRNA-2组LRP2 mRNA及蛋白表达水平均显著降低(P<0.05~0.01)。SC+Aβ组细胞凋亡率显著低于si LRP2+Aβ组(P<0.01)。SC+Aβ组及si LRP2+Aβ组(p-ERK1/2)/ERK与SC组及si LRP2组比较,差异无统计学意义(均P>0.05)。与SC+Aβ组相比,si LRP2+Aβ组(p-ERK1/2)/ERK亦无统计学差异(P>0.05)。SC+Aβ组及si LRP2+Aβ组p-p38/p38显著高于SC组及si LRP2组(均P<0.01)。与SC+Aβ组相比,si LRP2+Aβ组p-p38/p38无统计学差异(P>0.05)。SC+Aβ组及si LRP2+Aβ组p-JNK/JNK显著高于SC组及si LRP2组(均P<0.01)。与SC+Aβ组比较,si LRP2+Aβ组p-JNK/JNK显著增高(P<0.01)。未经JNK抑制剂处理的SC组、si LRP2组细胞的Cleaved Caspase-3相对表达与经JNK抑制剂处理的细胞差异无统计学意义(均P>0.05)。结论抑制LRP2后促进Aβ诱导的细胞凋亡,并且这种效应与ERK、p38磷酸化无明显相关,与JNK磷酸化相关,但JNK抑制剂不能逆转LRP2抑制导致的凋亡增加,提示JNK信号通路可能不直接发挥作用。
Objective To investigate the expression of low-density lipoprotein receptor-related protein 2(LRP2)inβ-amyloid protein(Aβ)induced Alzheimer’s disease(AD)cell model,and explore the effects to the survive of neurons and mitogen-activated protein kinase(MAPK)signaling pathway after regulating the expression of LRP2.Methods Aβ1-42-treated SH-SY5 Y cells were used as AD cell model,and the expression of LRP2 was detected by Western blotting.Scrambled siRNA and LRP2 siRNA(si LRP2)were used to transfect into SH-SY5 Y cells for 48 h,than treatment of Aβ1-42 for 24 h.The cell viability was detected by methyl thiazolyl tetrazolium,and apotosis was detected by flow cytometry.The phosphate expression of ERK,p38 and JNK were detected by Western blotting.Results The cell viability in si LRP2+Aβgroup was significant reduced than that in scramble(SC)+Aβgroup(P<0.05).Compared with that before Aβtreatment,the expression of LRP2 was significant reduced after Aβtreatment(P<0.05).Compared with those in blank control group,the expression of LRP2 mRNA and protein in siRNA-1 group and siRNA-2 group were significantly decreased(P<0.05-0.01).The apoptosis rate in SC+Aβgroup was significantly decreased than that in si LRP2+Aβgroup(P<0.01).The(p-ERK1/2)/ERK in SC+Aβgroup and si LRP2+Aβgroup were no statistical significance compared with those in SC group and si LRP2 group(all P>0.05).Compared with that in SC+Aβgroup,there was no statistical significance of(p-ERK1/2)/ERK in si LRP2+Aβgroup(P>0.05).p-p38/p38 in SC+Aβgroup and si LRP2+Aβgroup were significantly increased than that in SC group and si LRP2 group(all P<0.01).Compared with that in SC+Aβgroup,there was no statistical significance of p-p38/p38 in si LRP2+Aβgroup(P>0.05).p-JNK/JNK in SC+Aβgroup and si LRP2+Aβgroup were significantly increased than that in SC group and si LRP2 group(all P<0.01).Compared with that in SC+Aβgroup,p-JNK/JNK in si LRP2+Aβgroup was significantly increased(P<0.01).There was no statistical significance between the relative expression of
作者
王丽玲
李焰生
王鹏飞
WANG Li-ling;LI Yan-sheng;WANG Peng-fei(Department of Neurology,South Campus of Renji Hospital Affiliated to Shanghai Jiao Tong University School of Medicine,Shanghai 201112,China)
出处
《临床神经病学杂志》
CAS
2020年第1期40-46,共7页
Journal of Clinical Neurology
基金
仁济医院南院国自然培育基金(2017PYQA01)。
