Isolation and Characterization of Mesenchymal Stem Cells from Human Gingiva

Abstract

A new nonimmortalized cell line (MSC-GING) was isolated from the gingiva of a 35-year-old healthy donor and characterized. The analysis of different properties was carried out at the 6th, 7th, 13th, 18th, 20th, and 23rd passages. During long-term cultivation, the proportion of aging cells according to the activity of β-galactosidase increased. The cloning efficiency of MSC-GING cells was significantly reduced, and the number of aging β-galactosidase-positive cells was gradually increased during cell prolonged cultivation. Growth curves showed active cell proliferation at the sixth passage. It was significantly declined by the 18th and 20th passages. Karyotypic analysis at the 7th and 18th passages showed the presence of a diploid number of chromosomes (46) in metaphase plates. At passage 7, however, the proportion of cells with a normal diploid karyotype 46, XX, was 50 ± 5%. Other cells had a clonal rearrangement inv (10) (q21.1q25.1), an inversion of the long arm of chromosome 10. Their number significantly decreased during cultivation up to passage 18, while the proportion of cells with a normal diploid karyotype increased. The cell number with surface antigens common for human mesenchymal stem cells (CD44, CD73, CD90, CD105, HLA-ABC) and vimentin was high at passages 6 and 20, while CD34, CD45, and HLA-DR positive cells were very rare. It is noteworthy that no cells carrying markers of undifferentiated embryonic stem cells (Oct-4, SSEA-4, and SOX2) are observed at passages 6 and 20. It was shown that MSC-GING cells were able to differentiate in osteogenic and chondrogenic directions. The ability to differentiate in the adipogenic direction was manifested only at the level of glut4 gene expression. Induction of neuronal differentiation increased the expression the nse gene (neurospecific elonase). In general, the results confirmed the MSC status for the obtained line, but revealed significant karyotypic instability in the early passages, which decreased during the replicative aging.