The Effect Of MiR-30a-3p On The Proliferation And Migration Of Enteric Neural Crest Cells And Its Mechanism

As the "second brain" of mammals,the enteric nervous system plays a powerful role in regulating the function of the gastrointestinal tract.Intestinal neurons and glial cells are derived from the neural crest.Neural crest cells migrate to the foregut during the embryonic period,and then continue to proliferate,migrate,and differentiate along the intestinal wall,and finally form the enteric nervous system.The formation of the enteric nervous system includes a series of complex processes involving many signal pathways,and miRNAs are also involved as non-coding RNAs.miR-30a-3p is a member of the miR-30 a family.It has been found to be reduced in expression in colon and rectal cancer and other cancer tissues,and its main role is to inhibit cell proliferation and migration,and accelerate cell apoptosis.Studies have shown that MiR-30a-3p is abnormally expressed in Hirschsprung disease caused by the developmental disorder of neural crest cells,but its effect on the migration and proliferation of enteric neural crest cells is not yet known.Therefore,this study used SH-SY5 Y cells as a model of enteric neural crest cells to study the effects and mechanisms of miR-30a-3p on the proliferation,migration and apoptosis of enteric neural crest cells.First,CCK-8,Ed U,RTqPCR,Western Blot,Transwell technology and flow cytometry were used to detect the proliferation,migration,and cell cycle of enteric neural crest cells after transfection with miR-30a-3p mimics and inhibitors.The target gene of miR-30a-3p is predicted by Targetscan,MiRdb,MiRBase and other websites,and then the targeting relationship is detected by the luciferase reporter test,and the expression level of miR-30a-3p and PTEN are quantified by RT-qPCR.Finally,the target gene was determined.This study obtained the following results:1.After the transfection of miR-30a-3p mimic in SH-SY5 Y cells,overexpression of miR-30a-3p reduced the proliferation activity of SH-SY5 Y cells,the m RNA transcription and protein expression of proliferation-related genes were reduced,and cell migration was inhibited,but no significant effect on cell cycle and apoptosis.After transfection with miR-30a-3p inhibitor,inhibiting the expression of miR-30a-3p increased the proliferation activity of SH-SY5 Y cells,and cell migration was also promoted.2.The luciferase report test and RT-qPCR test proved that the 3’-UTR region of the PTEN gene has miR-30a-3p binding sites,and the expression of PTEN was regulated by miR-30a-3p.3.After transfection of siPTEN,inhibiting the expression of PTEN gene promoted the proliferation activity of SH-SY5 Y cells,increased the m RNA transcription and protein expression of proliferation-related genes,and promoted cell migration.4.Co-transfection of miR-30a-3p mimic and siPTEN will inhibit the promotion of cell proliferation caused by siPTEN to a certain extent,and will inhibit the promotion of cell migration caused by siPTEN.In summary,in enteric neural crest cells,miR-30a-3p inhibits cell proliferation and migration,and PTEN also inhibits cell proliferation and migration;miR-30a-3p can target to bind the 3’-UTR region of PTEN;Overexpression of miR-30a-3p up-regulates the expression of PTEN,thereby inhibiting the proliferation and migration of enteric neural crest cells.This research used miR-30a-3p as the target to explore its role in enteric neural crest cells,and explored its regulatory relationship with PTEN gene,These results will further provide the theoretical basis for clarifying the regulatory mechanism of miRNA on enteric neural crest cells.