Enhancing the integration and efficacy of retinal cell therapy
Fatemeh- Sadat Hosseini-Mazinani 1 , Mohammad Kazemi Ashtiani2 , Leila Satarian3 *, Hossein Baharvand4
- 1. Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
- 2- Department of Cell Engineering, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
- 1- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran
- 1- Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, Tehran, Iran. 3- Department of Developmental Biology, University of Science and Culture, Tehran, Iran
Abstract: Human pluripotent stem cell-derived retinal pigmented epithelium (hPSC-RPE) transplantation has become an attractive option for retinal degenerative diseases. However, numerous pathological changes in Bruch's membrane (BrM), make extracellular matrix (ECM) proteins out of the cell’s reach, resulting in poor integration of transplanted cells in diseased eyes. In vitro experiments have revealed that conditioned medium (CM) derived from endothelial cells (EC) significantly increased RPE survival on AMD BrM explants due to increase in ECM deposition. These findings lead us to hypothesize that whether hESC-RPE preconditioning with EC-CM and/or RPE-ECM, could increase ECM deposition and integration after transplantation in animal models.
Methods: To figure out the effects of EC and RPE secreted factors on hPSC-RPE proliferation and monolayer formation, 5×104 cells/cm2 were cultured in one of four conditions: (1) control; (2) 50% EC-CM; (3) 50% RPE-CM; and (4) 1 μg/ml RPE-ECM. On day 9, all the groups were collected and stained in primary antibodies against Ki-67 and ZO1. To evaluate the effect of preconditioning on RPE integration, after 21 days the GFP+ RPE cells were digested and directly injected into the subretinal space of C57Bl/6 mice model of retinal degeneration (n = 24). After 7 days, the eyes were examined for green fluorescent area (% GFA) related to transplanted cells. To assess spatial visual acuity after hPSC-RPE transplantation, 1×105 cells were injected into the subretinal space of RCS rats (n = 70) and the visual acuity was measured at 30 to 90 days post-transplantation using OptoDrum system.
Results: Our data demonstrated that preconditioning with EC-CM and RPE-ECM significantly increase the proliferation rate of hPSC-RPE compared to the control group. Furthermore, RPE cells in both groups showed significantly more engraftment than the control group in mice as evidenced by median GFA evaluation. Interestingly the increased engraftment translated into improved spatial-visual acuity (1.5 and 2.25 fold respectively) in RCS rats. Also, in vivo immunofluorescence assay indicated that preconditioned hPSC-RPE not only remained in a non-proliferative state but also supported cone photoreceptor survival.
Conclusion: These results suggested that preconditioning hPSC-RPE with EC-CM and/or RPE-ECM improves their integration after subretinal transplantation in the models.
