Identification of the Differentially Expressed microRNAs Involved in Cleft Palate Induced by Retinoic Acid (RA) in Mouse Model

Abstract

Identifying the diff erentially expressed miRNAs in cleft palate, in order to study the molecular mechanism in the development and progress of cleft palate. C57BL/6J mice were used to establish the RA induced cleft palate mouse model. Nine pairs of tissues were obtained on embryonic day 15.5 (E15.5). Total RNA was extracted and miRNAs microarray chip was used to screen the miRNAs. Real-time quantitative PCR (RT-qPCR) was used to verify the miRNAs microarray chip results. Cleft palate related genes targeted by the miRNAs were predicted by TargetScan and miRTarBase, and functional annotation clustering of Gene Ontology (GO) term and KEGG signaling pathways in DAVID were used to analysis these target genes. 1265 miRNAs were identifi ed in cleft palate, and among them 31 were diff erentially expressed (p < 0.01, fold change > 1.5), including 17 up-regulated and 14 down-regulated miRNAs in cleft palate. 7 up-regulated miRNAs (mmu-miR181a-5p, mmu-miR-410-3p, mmu-miR-3960, mmu-miR-1224-5p, mmu-miR-3970, mmu-let-7e-5p and mmu-miR-1907) and 3 down-regulated miRNAs (mmu-miR-140-3p, mmu-miR-351-5p and mmu-miR-503-5p) were validated by RT-qPCR, and mmumiR-181a-5p and mmu-miR-410-3p were in concordance with those of miRNAs microarray chip detection. 484 target genes were predicted and proven by TargetScan and miRTarBase. GO term showed that RA-induced cleft palate was associated with enrichments in miRNAs involved in embryo development, osteoblast diff erentiation and so on. KEGG signaling pathways analysis indicated that the diff erentially expressed miRNAs were involved in MAPK, TGFβ and WNT signaling pathways. 10 diff erentially expressed miRNAs in cleft palate have been identifi ed. These miRNAs and their target genes may become new therapeutic targets for cleft palate.