Growth kinetics and transplantation of human retinal progenitor cells.
We studied the growth kinetics of human retinal progenitor cells (hRPCs) isolated from donor tissue of different gestational ages (G.A.), determined whether hRPCs can be differentiated into mature photoreceptors and assessed their ability to integrate with degenerating host retina upon transplantation. Eyes (12-18 weeks G.A.) were obtained with IRB approval and retinas were enzymatically dissociated. Cells were expanded in vitro, counted at isolation and at each passage, and characterized using immunocytochemistry and PCR. GFP positive hRPCs were co-cultured with retinal explants from rd1 and rhodopsin -/- mice, or transplanted into B6 mice with retinal photocoagulation and rhodopsin -/- mice. Eyes were harvested for histological evaluation following transplantation. Our results show that hRPCs from 16 to 18 weeks G.A. had the longest survival in vitro and yielded the maximum number of cells, proliferating over at least 6 passages. These cells expressed the retinal stem cell markers nestin, Ki-67, PAX6 and Lhx2, and stained positively for photoreceptor markers upon differentiation with serum. Some of the GFP positive cells used for transplantation studies showed evidence of migration into the degenerative host retina and expressed rhodopsin. In conclusion, we have determined the growth kinetics of hRPCs and have shown that cells from donor tissue of 16-18 weeks G.A. exhibit the best proliferative dynamics under the specified conditions, and that hRPCs can also be differentiated along the photoreceptor lineage. Further, we have also demonstrated that following transplantation, some of these cells integrate within the host retina and differentiate to express rhodopsin, thereby supporting the potential utility of hRPC transplantation in the setting of retinal degenerative disorders.
Aftab
Unber
U
Schepens Eye Research Institute, Harvard Medical School, 20 Staniford Street, Boston, MA 02114, USA. unber.aftab@schepens.harvard.edu
Jiang
Caihui
C
Tucker
Budd
B
Kim
Ji Yeon
JY
Klassen
Henry
H
Miljan
Erik
E
Sinden
John
J
Young
Michael
M
D000818:Animals; D002454:Cell Differentiation; D049109:Cell Proliferation; D002470:Cell Survival; D002478:Cells, Cultured; D053686:Fetal Stem Cells / Q000166:cytology* / Q000378:metabolism / Q000637:transplantation; D016332:Fetal Tissue Transplantation / Q000379:methods*; D005865:Gestational Age; D006801:Humans; D051379:Mice; D018345:Mice, Knockout; D020419:Photoreceptor Cells, Vertebrate / Q000166:cytology / Q000378:metabolism; D016133:Polymerase Chain Reaction / Q000379:methods; D012160:Retina / Q000166:cytology / Q000196:embryology* / Q000378:metabolism / Q000637:transplantation; D012243:Rhodopsin / Q000378:metabolism; D033581:Stem Cell Transplantation / Q000379:methods*
D012243:Rhodopsin