Mechanism of c-Jun-targeted regulation of miR-182-5p transcription in upper limb functional rehabilitation of hemiplegic cerebral palsy
WANG You
XU Yun-xian
WU Qi-hong
CAI Kai-yin
WU Lin-lin
LIU Li-ru
XU Kai-shou
TANG Hong-mei
Abstract：Objective To investigate whether the transcription factor c-Jun can directly target miR-182-5p,identify the specific binding sites,and explore whether constraint-induced movement therapy(CIMT)can improve upper limb motor function in hemiplegic cerebral palsy(HCP)mice by promoting c-Jun/miR-182-5p transcription.Methods Lentivirus-mediated c-Jun knockdown and overexpression Neuro-2a cell lines were constructed to estab-lish stable cell models.Western blot was used to verify c-Jun protein expression,while RT-qPCR was employed to measure miR-182-5p expression.Chromatin immunoprecipitation(ChIP)was performed to identify the binding sites of c-Jun on the miR-182-5p promoter region.Subsequently,an HCP mouse model was established and subjected to CIMT intervention for 2 weeks.Behavioral tests were conducted to evaluate the effect of CIMT,and RT-qPCR was used to assess c-Jun and miR-182-5p expression in the left motor cortex(M1)of mice.Results Lentiviral transfection of c-Jun knockdown siRNA(si3,viral titer:5E+8 TU/mL)significantly reduced c-Jun protein expression in Neuro-2a cells(P<0.01),whereas c-Jun overexpression(OE,viral titer:5E+8 TU/mL)markedly increased c-Jun expres-sion(P<0.001).miR-182-5p transcription decreased significantly with reduced c-Jun expression(P<0.01)and increased significantly with elevated c-Jun expression(P≤0.001),demonstrating a synergistic relationship.Further studies revealed that c-Jun binds to multiple sites in the-1,000 to-1 bp upstream region of the miR-182-5p pro-moter,with the most prominent binding at Site 2(-71 to-65 bp)(P<0.001).Behavioral tests showed that,com-pared to the HCP group,the HCP+CIMT group exhibited significantly prolonged retention times in the fatigue rotarod test(P<0.01)and the upper limb suspension test(P<0.01).Additionally,c-Jun and miR-182-5p transcription levels in the M1 region of the brain were significantly elevated in the HCP+CIMT group(P<0.001).Conclusion The transcrip-tion factor c-Jun enhances miR-182-5p transcription by binding to its upstream promoter region(-71 to-65 bp).At the transcriptional level,the c-Jun/miR-182-5p regulatory axis provides a molecular basis for CIMT to improve upper limb function in HCP.
Keywords：c-JunmiR-182-5ptranscriptional regulationhemiplegic cerebral palsyconstraint-induced movement therapy
Publication Date：2025-03-15
Online Publishing Date：2025-08-15(First online date of this platform, not the publication date of the document)
Pages：8( 399-406 )