To evaluate DAPI (4′,6-diamidino-2-phenylindole) as a nuclear tracer of stem cell migration and incorporation it was observed the pattern of retinal integration and differentiation of mesenchymal stem cells (MSCs) injected into the vitreous cavity of rat eyes with retinal injury. For this purpose adult rat retinas were submitted to laser damage followed by transplantation of DAPI-labeled BM-MSCs grafts and double-labeled DAPI and quantum dot-labeled BM-MSCs. To assess a possible DAPI diffusion as well as the integration and differentiation of DAPI-labeled BM-MSCs in laser-injured retina, host retinas were evaluated 8 weeks after injury/transplantation. It was demonstrated that, 8 weeks after the transplant, most of the retinal cells in all neural retinal presented nuclear DAPI labeling, specifically in the outer nuclear layer (ONL), inner nuclear layer (INL), and ganglion cell layer (GCL). Meanwhile, at this point, most of the double-labeled BM-MSCs (DAPI and quantum dot) remained in the vitreous cavity and no retinal cells presented the quantum dot marker. Based on these evidences we concluded that DAPI diffused to adjacent retinal cells while the nanocrystals remained labeling only the transplanted BM-MSCs. Therefore, DAPI is not a useful marker for stem cells in vivo tracing experiments because the DAPI released from dying cells in moment of the transplant are taken up by host cells in the tissue.
INTRODUCTIONuse in multicolor fluorescent techniques, mRNA in situ hybridization, and in vivo cell tracking experiments. Because stem cells can easily self-replicate and difTo label cells without affecting their morphology or function is a critical issue for investigating cell behavior, ferentiate into lineage-specific cells, such characteristics provide several and important challenges for cell labelgrowth, migration, and differentiation at a single-cell level. For all these purposes, researchers are always ing. To date, thymidine analogs, such as 5-bromo-2-deoxyuridine, iododeoxyuridine, and tritiated thymidine, searching for a stable, nontoxic labeling that also will not leak, or be transferred from cell to cell either in vivo, and DAPI, have been extensively used to label stem cells prior to transplantation. These dyes are used in orin tracing experiments, or in vitro.The blue fluorescent DAPI (4′,6-diamidino-2-phenylder to allow the fate of the labeled cells into damaged areas to be determined (5,21,26 nonmesenchymal derivatives such as neural cells, in a retinal lesion was induced by an Nd-YAG laser, using an average of 0.5 mJ energy. A Goldman three-mirror process called stem cell plasticity or transdifferentiation (19,22,32). Several reports have shown that bone marlens was used to produce approximately 15-20 YAG laser shots around the optic disc. The lesions were all row mesenchymal stem cells (BM-MSCs) differentiate into retinal neural cells in vivo and in vitro (11), and full-thickness retina and choroid disruptions, as observed by deep subretinal hemorrhage and sometimes when implanted at a site of inju...