Study on human adipose mesenchymal stem cells differentiating into retinal pigment epithelial-like cells and its in vivo application

Abstract

Background Stem cell transplantation represents a promising treatment option for patients suffering from degenerative disorders. Accumulating evidences indicate that mesenchymal stem cells (MSCs) are able to differentiate into retinal pigment epithelial (RPE) -like cells. However, MSCs are difficult to obtain. Human adipose mesenchymal stem cells (ADSCs) are proved to have similar properties to MSCs, but relevant study is less. Objective This study was to assess the feasibility of human ADSCs differentiating into RPE-like cells and the safety of its application in vivo. Methods The third generation of human ADSCs were incubated into 6-well plate, and 100 ng/ml epithelial growth factor, 50 μmol/L taurine and 5×10-7 mol/L retinoic acid were added into the medium 12 hours after cultured to induce the cells, and conventional cultured cells were used as the control group. Induced cells were traced with PKH26, and Pan-cytoke ratin (Pan-CK) monoclonal antibody was used to identify the cells under the fluorescence microscope. Induced RPE-like cell suspension of 1 μl was intravetreally injected in the right eyes of 6 BALB/c mice, and equal volume of PBS was used in the same way in another 6 mice. The animals were sacrificed 1 month after injection, and the retinal morphology was examined by histopathology under the optical microscope. The ultrastructure of retinal ganglion cells (RGCs) was examined by the transmission electron microscope. The use and care of the animals complied with Regulations for the Administration of Affair Concerning Experimental Animals by State Science and Technology Commission. Results Cultured human ADSCs grew well with the slender polygone shape. Cell membranes showed the red fluorescence for PKH26 after induced. In addition, Pan-CK was expressed in the cell membranes with the red fluorescence in the induced cells, but the response was absent in the control cells. One month after intravitreal injection, induced cells located on the retinal surface, and the retinal morphology was clear under the optical microscope. No abnormality in RGCs was seen under the transmission electron microscope. Conclusions Human ADSCs can differentiate into RPE-like cells after induction. PKH26 can mark induced cells well. There is no adverse effect of induced cells on retina after intravitreal injection in a short-term duration in mice. Key words: Cell differentiation; Adipose-derived mesenchymal stem cells; Pigment epithelium of eye/cytology; Stem cells transplantation; Retina/pathology; Ultrastructure; Mice, BALB/c