Open Access

ARTICLE

Behavior of mesenchymal stem cells stained with 4’, 6-diamidino-2-phenylindole dihydrochloride (DAPI) in osteogenic and non osteogenic cultures

by N.M. OCARINO*, A. BOZZI**, R.D.O. PEREIRA*, N.M. BREYNER**, V.L. SILVA*, P. CASTANHEIRA**, A.M. GOES**, R

* Departamento de Clínica e Cirurgia Veterinárias, Escola de Veterinária da Universidade Federal de Minas Gerais, Brazil.
** Departamento de Bioquímica e Imunologia do Instituto de Ciências Biológicas da Universidade Federal de Minas Gerais, Brazil.
Address correspondence to: Dr. R. Serakides. Departamento de Clínica e Cirugia Veterinárias, Escola de Veterinária da Universidade Federal de Minas Gerais, Avenida Presidente Antonio Carlos, 6627, CEP 30.161-970, Belo Horizonte, Minas Gerais, BRASIL. E-mail: serakide@dedalus.lcc.ufmg.br

BIOCELL 2008, 32(2), 175-183. https://doi.org/10.32604/biocell.2008.32.175

Abstract

4’, 6-diamidino-2-phenylindole dihydrochloride (DAPI) is a DNA dye widely used to mark and trace stem cells in therapy. We here studied the effect of DAPI staining on the behavior of mesenchymal stem cells cultured in either a control, non-osteogenic medium or in an osteogenic differentiation medium. In the control medium, the number of stem cells/field, as well as the number of fluorescent cells/field increased up to the sixth day in both control and DAPI-treated cultures. Afterwards, both the number of fluorescent cells and their fluorescence intensity decreased. Control cells were fusiform and with some long extensions that apparently linked them to neighboring cells, while DAPI-treated cells were mostly round cells with fine and short extensions. The trypan-blue exclusion method showed 99% cell viability in both groups, however, both alkaline phosphatase activity and the thiazolyl blue formazan assay (indicative of mitochondrial metabolism) gave significantly lower values in DAPI-marked cells. The mitochondrial mass, as indicated by specific staining and flow cytometry, showed no differences between groups. Mesenchymal stem cells gave origin to mineralized nodules in the osteogenic differentiation medium and there were not DAPI-marked cells on the ninth day of culture. Alkaline phosphatase activity, viability assay and number of cells/field and of mineralized nodules/field were similar in both groups. So, DAPI treatment did not change cell viability and proliferation during osteogenic differentiation of mesenchymal stem cells. However, since these cells loose DAPI marking after 9 days in osteogenic cultures suggests that DAPI may not be an effective marker for mesenchymal stem cells implanted in bone tissue for long periods.

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