MiR-144-3p Regulates TH Expression And Neural Cell Differentiation By Targeting FOXO1 3'UTR

Purpose 1.To investigate the role of serum differentially expressed microRNAs on the regulation of FOXO1 expression in children with ADHD.2.ADHD animal model overexpressing and interfering with miR-144-3P in the prefrontal cortex of SHR rats,observe changes in SHR hyperkinetic symptoms and expression levels of FOXO1 and TH.3.Analyze the expression of FOXO1 in cells with different degrees of differentiation,and analyze the level of cell differentiation after overexpression and interference with miR-144-3P Methods 1.Collect serum sample from 40 untreated children with ADHD and 120 healthy children with the same conditions to form a 1 to 3 matching pair.Microarrays were used to screen out differentially expressed microRNAs.Serum samples were recollected and RT-qPCR verified for differentially expressed microRNAs.2.MicroRNAs targeted for regulation of FOXO1 were analyzed and screened from differential microRNAs through the bioinformatics website.Luciferas activity experiments verified whether microRNAs targeted to FOXO1 3?UTR.3.Compare the expression levels of miR-144-3P and FOXO1 in the prefrontal cortex of WKY and SHR rats.SHR rat brain prefrontal cortex was injected with miR-144-3p-virus,ADHD-like behaviors of SHR rats were observed through open field test and lat maze test.PFC brain tissues were taken and the expression levels of FOXO1 and TH were detected at the mRNA and protein levels,respectively.4.Differential expression of FOXO1 mRNA in PC12 H and PC12 L cells was detected by QPCR.PC12 L cells were infected with miR-144-3p-virus,and the effect on cell differentiation was observed.Results 1.In the serum of ADHD children,25 microRNAs with different expressions were screened.We further screened four microRNAs with significantly different expressions,which were miR-20a-5p,miR-20b-5p,miR-106b-5p,and miR-144-3p.The qPCR verification results showed that compared with the control group,the expressions of the four miRNAs were all down-regulated,and the differences were statistically significant.2.Based on the mirSVR score and PhastCons score,we further screened the microRNAs that may have a greater effect on FOXO1 3?UTR,namely miR-144-3p.Luciferas activity experiments showed that miR-144-3p directly targets FOXO1 3'UTR.3.Compared with WKY rats,miR-144-3p was significantly lower expressed in SHR PFC and FOXO1 was significantly higher expressed.In the open field test,the number of trellises crossed and times of standing with hind limbs of miR-144-3p-virus group were significantly lower than control group;In the lat maze,the number of miR-144-3p-virus group crossing corners and standing with hind legs was significantly lower than control group.PFC was used to detect the expression of miR-144-3p and FOXO1,and the results showed that compared with the control group,miR-144-3p was significantly increased and FOXO1 was significantly decreased,and the differences were statistically significant.4.The expression of FOXO1 mRNA in PC12 L cells was significantly higher than that in PC12 H cells.The synaptic length of PC12 L cells in miR-144-3p-virus group was significantly higher than that in control group,and the difference was statistically significant.Conclusion 1.Compared with healthy children,miR-20a-5p,miR-20b-5p,miR-106b-5p,and miR-144-3p are significantly lower expressed in the serum of children with ADHD,which can provide a basis for clinical diagnosis of ADHD.2.miR-144-3p affects TH expression and neuronal differentiation through targeted regulation of FOXO1 3?UTR,which may play an important role in the development of ADHD.