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P.A. Stephano, Y. Guo, P. Saloupis, S. Shaw, M. Mahoney, D. Rickman; Neural Progenitor Cell Transplantation in the Ischemic Retina With Sustained BDNF Delivery . Invest. Ophthalmol. Vis. Sci. 2003;44(13):1018.
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Purpose: To determine the extent of engraftment and differentiation of adult neural progenitor cells (NPCs) transplanted to the ischemic retina under conditions of minimal injury and in response to sustained release of BDNF. Methods: We used female Fischer 344 rats and well-characterized NPCs derived from the adult rat hippocampus and expressing GFP (a gift of Dr. F. Gage). Animals were subjected to visually-confirmed ischemia, induced by inserting a needle attached to a saline drip into the anterior chamber and increasing the intraocular pressure to 110 mm Hg for up to 60 min. Dissociated NPCs of varying amount (3 µl of 5 x 103 – 5 x 104 cells) were transplanted into the subretinal space immediately following ischemia. Some rats also received subretinal injection of biocompatible, degradable microspheres containing either BDNF (100ng/µl) or BSA. Animals were allowed to recover for 2-11 wk, then perfused with 4% PFA. Eyes were removed and retinas were cryosectioned for analysis. Sections were stained for retinal cell type-specific markers including rhodopsin, recoverin, PKC, calbindin, calretinin, parvalbumin, or Thy1.1, using the appropriate Cy3-conjugated secondary antibody. Results: Successful transient ischemia and reperfusion was achieved, as confirmed by examination of the retinal arteries. After 2 wk, engraftment of transplanted GFP-expressing cells was observed following an injury as brief as 60 minutes and injection of as few as 5 x 103 cells. By four weeks, cells had migrated to the inner retina and extended processes. At six weeks, GFP-expressing cells showed limited differentiation with morphological features of horizontal, bipolar, and amacrine cells. In animals co-transplanted with BDNF-releasing microspheres, we observed transplanted cells with an enhanced degree of morphological differentiation, relative to animals receiving BSA-containing microspheres. Expression of retinal cell type-specific antigens, however, was only modestly enhanced. Conclusions: Cell replacement is a novel but, as yet, unrealized therapeutic strategy for neurodegenerative disease. Here, we have shown successful engraftment and limited differentiation of transplanted cells under conditions of minimal ischemic injury and number of transplanted cells. Additionally, co-transplantation of sustained release BDNF microspheres appears to modestly enhance the differentiation of transplanted cells. These results suggest that a combination of therapeutic approaches may prove beneficial in the design of cell transplantation strategies.
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