The role of lipopolysaccharide in regulating mitochondrial fusion of SH-SY5Y cells in Alzheimer's disease
Objective To investigate the role of lipopolysaccharide(LPS)in regulating mitochondrial fusion of Human neuroblastoma(SH-SY5Y)cells in Alzheimer's disease(AD).Methods SH-SY5Y cell line was cultured routinely.During the logarithmic phase,SH-SY5Y cells were treated with 25,50 and 100 μg/ml LPS for 48 h,and were set as 25 μg/ml group,50 μg/ml group and 75 μg/ml group,and a blank control group was set.The cell survival rate was detected by cell counting kit-8(CCK-8)method,the mitochondrial damage was detected by light microscope,the changes of superoxide dismutase(SOD),malondialdehyde(MDA),nitric oxide(NO),and tumor necrosis factor-α(TNF-α)were detected by enzyme linked immunosorbent assay(ELISA),and the expression and protein expression of dynamin-related protein 1(Drp1),mitofusin 1(Mfn1),mitofusin 2(Mfn2)and optic atrophy 1(Opa1)micro ribonucleic acid(mRNA)were detected by real-time PCR method.The effects of LPS on survival and β-amyloid protein(Aβ)content of SH-SY5Y cells,mitochondrial morphology of SH-SY5Y cells,intracellular SOD,MDA,NO and TNF-α in SH-SY5Y cells,and intracellular mRNA expression in SH-SY5Y cells were analyzed.Results The results of CCK-8 assay showed that compared with the blank control group,the survival rate of SH-SY5Y cells in 25,50 and 100 μg/ml groups decreased significantly and the Aβ content increased significantly at 48 h of administration.With the increase of LPS dose,the survival rate of SH-SY5Y cells decreased in a dose-dependent manner,and the Aβ content increased in a dose-dependent manner,and the differences were statistically significant(P<0.05).The light microscope results showed that compared with the blank control group,mitochondria in SH-SY5Y cells in the 25,50 and 100 μg/ml groups showed significant enlargement and roundness,increased volume,swelling and vacuolization,accompanied by the occurrence of mitochondrial autophagy at 48 h of administration.ELISA results showed that compared with the blank control group,SOD expression in SH-SY5Y cells in 25,50 and 100 μg/ml groups significantly decreased by 24.16%,69.08%and 123.78%,MDA expression significantly increased by 37.65%,52.13%and 101.85%,NO expression significantly increased by 54.34%,119.08%and 167.78%,and TNF-α expression significantly increased by 54.34%,119.08%and 167.78%at 48 h of administration.With the increase of LPS dose,the changes of SOD contents were all dose-dependently decreased,and the changes of NO,MDA and TNF-α contents were all dose-dependently increased.The differences were statistically significant(P<0.05).Real-Time PCR results showed that,compared with the blank control group,the intracellular Drp1,Mnf1 and Mnf2 mRNA contents in SH-SY5Y cells in the 25,50 and 100 μg/ml groups increased significantly with the increase of the LPS dose at 48 h of administration,and the intracellular Opa1 mRNA content in SH-SY5Y cells significantly decreased with the increase of the LPS dose.The difference was statistically significant(P<0.05).Conclusion LPS can significantly accelerate the mitochondrial fusion and division of SH-SY5Y cells and reduce the survival rate of cells,which plays an important role in the pathogenesis of Alzheimer's disease.
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