Effects of chronic restraint stress on the expression of N6-methyladenosine and related enzymes in the hippocampus of mice

doi:10.16098/j.issn.0529-1356.2023.02.003

Abstract

Abstract:

Objective To investigate the effect of chronic restraint stress on the expression of N6-methyladenosine (m6A）and related enzymes in the hippocampus of mice. Methods Twenty C57BL/6J male mice were randomly divided into control group and chronic restraint stress (CRS) group, the model group was given for 3 weeks chronic restraint stress to establish a mouse anxiety model. Open field test and elevated plus maze test were used to detect anxiety-like behavior; Immunohistochemistry and m6A RNA methylation assay were used to detect the expression changes of mouse hippocampal m6A; Western blotting and Real-time PCR were used to analyze hippocampal m6A related enzymes expression. Results 1.The behavioral results showed that, compared with the control group, the CRS group showed significantly reduced time spent in the center of the open field(P<0.01), the CRS group showed significantly reduced exploration time in the open arm of elevated plus maze (P<0.0001); 2. Immunohistochemical results showed that, compared with the control group, the hippocampal m6A content in the CRS group reduced significantly (P<0.001); The results of the m6A RNA methylation assay showed that, compared with the control group, the CRS group showed significantly reduced amount of hippocampal m6A(P<0.05); 3. Real-time PCR results showed that the expression of hippocampal demethylase anaplastic lymphoma kinase B(AlkB) homolog 5(ALKBH5) (P<0.001) and fat mass and obestity associated protein(FTO) (P<0.05) in the CRS group significantly up-regulated, the expression of methylase Wilms’tumour 1-associating protein(WTAP) (P<0.05) was significantly down-regulated compared with the control group; The expression of m6A methylation binding protein YTH domaincontaining family protein 3(YTHDF3) (P<0.05) and YTH domaincontaining protein 2(YTHDC2)(P<0.01) was significantly up-regulated. Western blotting result showed that, compared with the control group, the mouse hippocampal demethylase ALKBH5 (P<0.05) and FTO (P<0.05) expression in the CRS group significantly up-regulated, the expression of WTAP (P<0.01) was significantly down-regulated; m6A methylation binding protein YTHDF3 (P<0.01) and YTHDC2 (P<0.05) were significantly up-regulated. Conclusion In the anxiety model induced by chronic restraint stress, the expression of m6A in the hippocampus of mice is down-regulated. The mechanism may be related to the up-regulation of the m6A demethylase ALKBH5 and FTO or the down-regulation of the methylase WTAP.

Key words:

Chronic restraint stress, Anxiety, N6-methyladenosine, Anaplastic lymphoma kinase B homolog 5, Fat mass and obesity-associated protein, Wilms’ tumour 1-associating protein, Western blotting, Mouse

CLC Number:

R322.81

ZHANG Yuan SHI Meng-meng HE Jia-bei WU Qiong ZHONG Xiao-lin XIANG Yu-yan XU Yang HE Jie . Effects of chronic restraint stress on the expression of N6-methyladenosine and related enzymes in the hippocampus of mice[J]. Acta Anatomica Sinica, 2023, 54(2): 142-148.

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