Tracing study of DAPI Labeled Mesenchymal Stem Cells in Vivo and in Vitro

Objective To explore labeling efficiency of DAPI labeled mesenchymal stem cells(MSCs) in vivo and in vitro,and provide technical foundation for the study of cell fate determination in the course of MSCs transplantation repairing ischemic diseases.Methods DAPI was added to the culture medium at a final concentration of 50 μg/ml for 30 minutes,when the cultured MSCs in passage 3 reached 50% confluency.For in vitro study,MSCs were collected during the initial period and after1,2,3 and 4 weeks.The Intensity and pattern of fluorescence,as well as the labeling efficiency,were evaluated just after labeling and again after 1,2,3 and 4 weeks.The scale was calculated based on the fraction of positive MSCs to the total MSCs attached to the bottom.For in vivo study,labeled MSCs were transplanted into ischemic hind limb in rats,and samples were collected 1,2,3 and 4 weeks after transplantation.These tissue sections were observed under the same microscope with the same filter,and the numbers of labeled MSCs were evaluated.Results DAPI achieved outstanding initial labeling efficiency for the cultured MSCs(all 100%).However,the labeling efficiency after 3,4 weeks decreased to 20% and 5% for DAPI in vitro,respectively.Intensive labeled MSCs in tissue sections were identified 1,2 weeks after MSCs transplantation,the numbers of labeled MSCs after 3 weeks decreased obviously.Conclusion An advantage of DAPI is the high labeling efficiency via simple procedure.The labeled MSCs were easily detected,and the initial labeling efficiency was 100%.However,disadvantage of DAPI is the loss of labeling intensity due to cell proliferation and division.DAPI may be used only for quantitative analysis for a short period.