Effect of ten-eleven translocation protein on the expression of amyloid precursor protein and β-site APP cleaving enzyme-1 in SH-SY5Y and IMR-32 cells in folic acid deficiency and the underlying mechanisms / 中华老年医学杂志

ABSTRACT

Objective To investigate the effect of ten-eleven translocation protein on the proliferation of human neuroblastoma cell lines SH-SY5Y and IMR 32 and the expression of amyloid precursor protein,PS1,and β site APP cleaving enzyme 1 in the absence of folic acid and possible mechanisms involved.Methods SH-SY5Y and IMR-32 cells were cultured in vitro and divided into the folic acid deficiency group (0 mg/L),the low folic acid group (1mg/L),and the normal control group (4mg/L).The MTT method was used to observe cell proliferation,and RT-PCR was adopted to detect the mRNA expression of APP,PS1,BACE1,DNMTs and TETs in the cells in real-time.Besides,we generated a stable low-level TET1 expression cell line,and compared the expression with that in a negative control group.Furthermore,the expression of fluorescent protein was observed by fluorescence inverted microscope,cell proliferation was measured by the MTT assay,and mRNA levels of TET1,APP,PS1,and BACE1 were detected by RT-PCR.Results (1) In the folic acid deficiency group and the low folic acid group,cell proliferation of SH-SY5Y after 120 h and of IMR-32 cell after 144 h significantly decreased (P＜0.001).The mRNA levels of APP,PS1,BACE1,DNMT1,DNMT3a,DNMT3b,TET1,TET2,and TET3 in SH-SY5Y cells increased (F=80.315,35.386,101.979,786.407,80.331,131.545,28.000,9.165,and 102.167,all P＜0.05);the mRNA levels of APP,PS1,BACE1,DNMT1,DNMT3b,TET1,TET2,and TET3 in IMR-32 cells also rose (F=12.283,93.669,40.815,157.234,24.835,147.594,54.794,and 73.068,all P＜0.05).(2) Generation of a stable low-level TET1 expression cell lineThe mRNA level of TET1 in the low expression group (SH-SY5Y-shTET1) was 0.25± 0.02,which was significantly lower than that in the negative control group (1.00±0.09) (P=0.007);the mRNA level of TET1 in the low expression group (IMR-32-shTET1) was 0.28 ±0.07,significantly lower than that in the negative control group (1.00±0.01) (P=0.003).(3)The proliferative ability of the low expression groups (SH-SY5Y-shTET1 and IMR-32-shTET1) was significantly higher than that in the negative control group (P＜0.01).The mRNA levels of APP and BACE1 decreased (P＜0.01 or P＜0.05)Conclusion In the human neuroblastoma cell lines SH-SY5Y and IMR-32,folic acid deficiency up-regulates the expression of TETs,increases the expression of APP and BACE1 in the cells by TET protein demethylation,and inhibits cell growth.