The MRNA M6A Reader YTHDF2 Suppresses The Proliferation And Migration Of Pterygium Fibroblasts

Purpose: To study the expression difference of m6 A and m6 A methylation-related enzymes in pterygium and conjunctival.Investigate the role of m6 A methylation in the occurrence and development of pterygium.To preliminarily explore the effect of YTHDF2 on the proliferation and migration of pterygium fibroblasts and provide a new theoretical basis for the study of the pathogenesis of pterygium.Methods:1.42 pairs of pterygium and conjunctival tissues were collected.Using q RT-PCR,assessed the expression level of m6 A,m6A demethylase(FTO and ALKBH5),methyltransferase(METTL3,METTL14 and RBM15),and methylated reading protein(YTHDF1,YTHDF2,YTHDF3,YTHDC1,YTHDC2 and ELF3)in pterygium and conjunctival tissue.2.Human primary pterygium fibroblasts(HPFs)and Human conjunctival fibroblasts(HCon Fs)were cultured and identified by immunofluorescence technique.The expressions of YTHDF2,ELF3,ALKBH5,YTHDC1 and YTHDC2 in pterygium fibroblasts and conjunctival fibroblasts were detected by q RT-PCR.According to the results,using the immunohistochemical techniques detected the expression and distribution of YTHDF2 in pterygium and conjunctival tissues.3.The YTHDF2 overexpressed recombinant plasmid was constructed and transfected into pterygium fibroblasts.q RT-PCR and Western blotting detected RNA and protein expression levels after YTHDF2 transfection.4.CCK8 and scratch tests were used to detect the effect of YTHDF2 overexpression on HPFs proliferation and migration.Results:1.The expression of m6 A in pterygium tissue was significantly higher than that in conjunctival tissue(P < 0.01),methylated reading protein(YTHDF2,YTHDC1,YTHDC2 and ELF3)and demethylated enzyme ALKBH5 were decreased in pterygium(P < 0.05).There was no difference in the expression of FTO,METTL3,METTL14,RBM15,YTHDF1 and YTHDF3 in pterygium and conjunctival tissues(P >0.05).2.The expression of the methylated reading protein(YTHDF2,ELF3)and demethylase ALKBH5 decreased in HPFs(P < 0.05),while there was no difference in the expression of the methylated reading protein(YTHDC1,YTHDC2)between HPFs and HCon Fs(P > 0.05).3.After the recombinant plasmid overexpressing YTHDF2 was constructed and successfully transfected into HPFs,the m RNA and protein levels of YTHDF2 in HPFs were detected to be higher than the control group(P<0.05).4.CCK8 and Scratch test results showed that the proliferation and migration of the HPFs-YTHDF2 cell line decreased significantly compared with the HPFs-Vector cell line(P <0.001).Conclusions: This study suggests that m6 A methylation is involved in the occurrence and development of pterygium,and the elevated overall expression of m6 A may be related to the decrease of YTHDF2 expression in pterygium.YTHDF2 may act as an inhibitor of HPFs proliferation and migration.This finding provides new insights into understanding the role of m6 A modification in pterygium.