Regenerative effect of adipose derived mesenchymal stem cell on ganglion cells in the hypoxic organotypic retina culture

Abstract

Purpose : To examine whether ischemic retinal ganglion cells (RGC) will be salvaged from apoptosis by co-culture with adipose-derived mesenchymal stem cells (ADSC) by direct contact or by paracrine activity.

Methods : Retinas of wild type C57Bl/6 mice were harvested. Deprived of arterial oxygen supply, retinas were cultured as ex vivo organotypic cultures on an insert membrane. First, number of surviving RGC was evaluated at different time points (T0, T48, T72 hours) by Brn3a staining and confocal microscopy. Then, the therapeutic potential of ADSC on ischemic retinas was evaluated either by co-culture of ADSC with organotypic retinas with direct contact of ADSC (ADSC seeded above the retina) or by a co culture of ADSC with organotypic retinas allowing a paracrine activity of ADSC (ADSC seeded on tissue culture plate below the retina) (n=7). Retinas were flat-mounted and the number of surviving RGC was assessed using Brn3a staining and confocal microscopy. We also examined the cytokine secretion profile of ADSC related to their paracrine effect on ischemic retina by analyzing the protein expression in ADSC-conditioned medium using protein array.

Results : We demonstrated a time-dependent decrease in number of viable RGC in non-treated ischemic retinas evident by Brn3a+ staining. When co cultured with ADSC, the number of surviving RGC was significantly higher in treatment groups compared to controls of retinas treated with standard medium (Brn3a+ cells treated with ADSCs seeded above the retina = 29±10.9, p<0.001, Brn3a+ cells treated with ADSCs seeded below the retina = 29±7.2, p=0.001, controls = 4.5±2.5 SD). Evaluating ADSCs cytokines secretion profile, we showed secretion of anti apoptotic and pro- proliferative cytokines (threshold >1.4).

Conclusions : Our results demonstrate that transplantation of ADSC in a co culture with organotypic ischemic retinas significantly improved recovery of the RGC. The effect was similar when ADSC were seeded above and below the retina suggesting the beneficial effect seen was related to paracrine activitiy of ADSC rather than to direct contact of ADSC. These data correlated with secretion profile of ADSCs anti-apoptotic and pro proliferative cytokines. In summary, ADSC could direct future development of autologous cell therapy modalities to repair ischemic retinal and/or optic nerve damage.

This is a 2021 ARVO Annual Meeting abstract.