The Role And Mechanism Of BDNF-ERK-CREB Signaling Pathway In PFOS Neurotoxicity

Objective: The aim is to investigate the neurotoxicity of PFOS and its mechanism, and to provide experimental evidence for further study on the neurotoxicity of PFOS.Methods: The experiment was divided into two groups, the control(0 ?M) and the PFOS groups(1, 10, 50, 100, 150, 200 ?M). The cell viability and time-efficiency of SH-SY5 Y were determined by CCK8, the morphology and rate of apoptosis were tested by AO-EB dual fluorescent labeling and Annexin-V FITC/PI double-staining, respectively. The effect of PFOS on oxidative stress damage in SH-SY5 Y cells was detected by UV spectrophotometry and fluorescence methods. QPCR method was used to detect the effects of PFOS on has-miR-16, has-miR-22, BDNF gene, ERK/1/2 gene, CREB gene, DNMT1 gene, DNMT3 a gene and DNMT3 b gene mRNA expression levels in SH-SY5 Y cells. Elisa was used to detect the expression level of BDNF protein on SH-SY5 Y cell supernatant. The protein expression levels such as TrkB, ERK1/2,pERK1/2, CREB, pCREB, DNMT1, DNMT3 a and DNMT3 b in SH-SY5 Y cell were detected by Western Blot method.Results: The outcomes of CCK8 test showed a drop of survival rate in a time-dose effective relationship. The apoptosis rate increased in correlating with the PFOS concentration for 48 hours after contamination indicated by both AO-EB dual fluorescent observation and Annexin-V FITC/PI dual-dyeing flow-cytometry. The results of UV and fluorescence spectrophotometry showed that PFOS induced SH-SY5 Y cell superoxidedismutase(SOD) and glutathione reductase(GSH) decreased, compared with the control, the highest group of PFOS decreased GSH from 8±0.12nmol/mg protein to 5.89±0.9 nmol/mg protein. While the total reactive oxygen species(ROS) and malondialdehyde(MDA) levels increased,compared with the control, the highest group of PFOS elevated MDA from 1.92±0.17 nmol/mg protein to 6.95±0.26 nmol/mg protein. QPCR and Western Blot results showed that PFOS disturbs BDNF-ERK-CREB signalling in association with increased microRNA-22 in SH-SY5 Y cells,compared with the control, the BDNF protein expression levels in culture supernant of 100?M group decreased from 60±1.2 ng/mL to 29±1.5ng/mL. Meanwhile, PFOS interfered the protein expression levels of DNMT1, DNMT3 a and DNMT3 b.Conclusions: 1. PFOS exposure decreased the cell viability of SH-SY5 Y cells, induced SH-SY5 Y cells apoptosis in a dose-dependent manner. 2. PFOS disturbed BDNF-ERK-CREB signaling pathway in association with increased microRNA-22 in SH-SY5 Y cells, and may be one of the mechanisms of the neurotoxicity of PFOS. 3. PFOS produced oxidative stress damage in SH-SY5 Y cells, and changed the expression levels of DNMT1, DNMT3 a and DNMT3 b, which may be the mechanisms of neurotoxicity induced by PFOS.