Objective To investigate the effect of transcription factor specific protein5(Sp5)silencing on Wnt signaling pathway correlated factors and cell proliferation ability in mouse embryo palatal mesenchymal cells(mEPMCs).Methods mEPMCs of 14.5 d pregnant C57BL/6J mice were isolated and cultured in vitro.Cell source was identified by immunofluorescence staining.Lentivirus transfection technique was used to silence the expression of Sp5 gene in mEPMCs,and the transfection efficiency was verified by Western blot assay.Follow-up experiments were set up with the blank control group,the no-load virus group and the slience group(the Sp5-shRNA group).The protein and mRNA expression levels of β-catenin,GSK-3β,Wnt3a and CyclinD1 were detected by Western blot assay and RT-qPCR after transfection for 72 h in each group.Cell proliferation capacity was detected by CCK-8.The proliferation rate of 5-Ethynyl-2'-deoxyuridine(EdU)positive cells was detected by immunofluorescence assay.Cell cycle was detected by flow cytometry.Results mEPMCs were successfully isolated,and Sp5 expression was silenced.Western blot and RT-qPCR results showed that the protein and mRNA expressions of β-catenin,GSK-3β,Wnt3a and CyclinD1 were significantly higher in the Sp5-shRNA group than those in the blank control group and the no-loaded virus group(P<0.05).The proliferative ability and the proliferative rate of EdU positive cells were higher in the Sp5-shRNA group than those in the blank control group and the no-loaded virus group(P<0.05).The proportion of mEPMCs in S phase was higher in the Sp5-shRNA group than that in the blank control group and the no-loaded virus group(P<0.05).Conclusion Sp5 in silenced mEPMCs can participate in palate development and promote the proliferation of mEPMCs by regulating Wnt signaling pathway.