Abstract:

Gingival mesenchymal stem cells（GMSCs）cell clusters and functional heterogeneity were analyzed through single cell RNA sequencing technology. Human GMSCs were isolated and cultured in vitro, GMSCs were sequenced by 10× scRNA-Seq platform. The cells were grouped according to the gene expression differences, and analyze the expression of surface markers and cell cycle differences in each cluster. Differences in gene expression and gene expression profile of osteogenic, lipogenic and chondrogenic differentiation were compared，and the interactions between different clusters of cells were also analyzed. Human GMSCs were successfully extracted and grew in a vortex like manner. The GMSCs were composed of 9 cell clusters. ENG, NT5E, THY1, ITGB1 and CD44 were highly expressed, and CD34, CD14, PECAM1 and PTPRC were negatively expressed. The proportion of G2/M phase of GMSCs was 41.59%. The G2/M phase ratio of cell cluster 6 was the lowest（16.44%）, while the cell cluster 2 was the highest with 99.08%. The expressions of adipogenic, chondrogenic and osteoblastic differentiation related genes were not identical in the 9 cell clusters. Nine clusters had different gene expression profiles, and 2/4 cell clusters showed high similarity, with relatively high expressions of TOP2A, UBE2C, KPNA2, CENPF genes. DKK1, KRT7 and ID1 genes were highly expressed in the cell cluster 5. MALAT1, FN1 and NEAT1 genes were highly expressed in the cell cluster 9. Moreover, GO analysis showed that each cell cluster enriched different pathways. The communication between cell cluster 9 and other cell clusters were the most active. The single-cell level study suggests that there are different functional clusters of GMSCs, and the proliferation capacity and genomic expression profile of each cluster were heterogeneous.

Key words: gingival mesenchymal stem cells, single cell RNA sequencing, cell clusters, functional heterogeneity