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Jose-Carlos Pastor, Amar K Singh, Ivan Fernandez-Bueno, David Rodriguez-Crespo, Maite Garcia-Gutierrez, Manuel Gayoso, Girish K Srivastava; Human adipose tissue derived mesenchymal stem cells (hAD-MSCs) together with different combinations of factors delay the degeneration of human neuroretina in indirect co-culture system. Invest. Ophthalmol. Vis. Sci. 2014;55(13):5748.
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To analyze the neuroprotective effect of hAD-MSCs together with factors [vasoactive intestinal peptide (VIP), nicotinamide (NIC) and all-trans-retinoic acid (ATRA)] (patent pending) on in vitro degeneration of human neuroretinal (hNR) explants.
Study was approved by ethical committee of the University of Valladolid (Spain). hAD-MSCs were isolated from donated human lipo-aspirates and maintained in culture and characterized as previously described by us (Singh et al. Histol Histopathol. 2013). hNR explants (6x8mm) were prepared from donated human post mortem eyes (Fernandez-Bueno et al. Exp Eye Res. 2012). Retinal pigment epithelial cell condition medium (RCM) was collected by centrifuging the supernatant of ARPE19 cell culture at 48 hours. hAD-MSCs were seeded (30,000 cells/cm2) on the bottom of Transwell culture plates for 24 hrs to grow. Then, hNR explants were placed with photoreceptors side downwards on the porous membrane of trans-insert and co-cultured in Dulbecco's Modified Eagle Medium: Nutrient Mixture F-12 medium with 50% RCM for 6 days in 7 different combinations of factors with two controls as follows: RCM only (control-1), RCM+MSCs (control-2), control-2 +VIP (5μM), control-2 +NIC (10mM), control-2 +ATRA (5μM), control-2 +VIP+NIC, control-2 +VIP+ATRA, control-2 +NIC+ATRA and control-2 +VIP+NIC+ATRA. Epoxy resin-embedded semi-thin sections of hNR explants from each condition were used to evaluate the effects of different combinations.
hNR explants of control-1 were badly affected and disorganized (Fig.1). General structure of hNR explants of control-2 was significantly better that that of the control-1.The general structure of hNR explants cultivated with the combinations VIP+NIC and VIP+NIC+ATRA in the presence of RCM was significantly better preserved (Fig.2).
hAD-MSCs plus combination of factors (VIP+NIC or VIP+NIC+ATRA) preserve the neuroretinal degeneration under in vitro conditions and might delay the degeneration of neuroretina in vivo. This study could further be extended to in vivo animal models of retinal degeneration before going ahead to any clinical trials in retinal degenerative diseases.
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