Purchase this article with an account.
Jose-Carlos Pastor, Amar K. Singh, Girish K. Srivastava, David Rodríguez, Maite García-Gutierrez; Adipose derived Mesenchymal stem cells partially rescue the Mitomycin C treated ARPE19 from cell death in co-culture. Invest. Ophthalmol. Vis. Sci. 2012;53(14):5843.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
Some retinal degenerations are primarily caused by damage at the Retinal Pigment Epithelial (RPE) layer. Thus its replacement becomes a target for therapeutical approaches including in cell therapy. Stem cells are promising option. Adipose derived mesenchymal stem cells (AD-MSCs) secret growth factors which play important role in the maintenance of neuroretina and RPE and their possible use for maintaining the viability of RPE cells has been suggested. The aim of this study was to investigate the possibility of rescuing mitomycin C (MMC) induced cell death in ARPE19 cells by AD-MSCs in indirect co-culture condition.
ARPE19 cells and AD-MSCs were cultured under standard condition (5% CO2, humid air, 37º C in complete DMEM/F12 and complete low glucose DMEM with glutamine medium). ARPE19 cells were treated with MMC in different concentrations (50µg/ml, 100µg/ml and 200µg/ml) for 2 hours to induce cell death. Cell death and viability assay were done by cell Viability/cytotoxicity Assay Kit for Animal Live & Dead Cells (Biotium Inc., USA). ARPE19 cells were seeded in 24 well plates and incubated for 24 hours. Then ARPE19 cells were incubated with MMC, then washed with PBS to remove any residual MMC. AD-MSCs were seeded on the transwells. After 3 days of co-culture, to evaluate ARPE19 cells’ growth and proliferation, 10% Alamar Blue (AB) ( AbD Serotec, Oxford, UK) was added to the growth medium and incubated for 5 hours. AB fluorescence intensity was measured with excitation wavelength at 560 nm and emission at 590 nm.
MMC (50µg/ml) treated ARPE19 cells start to die after 2 weeks and almost dies completely after 3 weeks. Cell toxicity increases significantly (p <0,05) along with increase of MMC concentration. When MMC treated ARPE19 cells grown with AD-MSCs grow better with respect to MMC treated ARPE19 cells alone. Mean fluorescence intensities significant increase from 12,720.9 (SD:2,884.6) to 17,106.1 (SD:1,803.0), 10,088.5 (SD:2615.6) to 14,838.5 (SD:2557.4) and 5,238.2 (SD:647.9) to 8,295.7 (SD:780.9) in 50, 100, and 200µg/ml MMC treated groups.
Results show that AD-MSCs increase the cell survival of MMC treated ARPE19 cells probably by secreting growth factors in the medium which is shared by both types of cells. This study would help in the study of effect of direct subretinal transplantation of AD-MSCs over the RPE degeneration process in AMD patients
This PDF is available to Subscribers Only