Vamsi K. Gullapalli

The Transplantation of Human Fetal Neuroretinal Cells in Advanced Retinitis Pigmentosa Patients: Results of a Long-Term Safety Study ☆

The purpose of this study was to determine the long-term safety of transplanting human fetal neuroretinal cells (14 to 18 week gestational age) into a series of patients with advanced retinitis pigmentosa (RP). After obtaining informed consent, both hosts and mothers of donors were screened for transmissible diseases. Pre- and postoperative clinical exams, visual acuity, electroretinograms, and fluorescein angiograms were performed and visual field testing was attempted in each case. Surgically, an anterior approach through pars plana ciliaris was used. A retinotomy was performed in the paramacular area and a two-function cannula was introduced into the subretinal space to deliver a suspension of donor cells. The cell suspension carried approximately 4000 cells/microl; the volume injected did not exceed 150 microl. The patients were examined for periods ranging from 12 to 40 months posttransplantation. To date, no evidence of inflammation, infection, or overt rejection of the graft was noted in the host eye, neither was any change observed in the contralateral, unoperated eye. In conclusion, neuroretinal cells were injected into the subretinal space of 14 patients with advanced RP with no clinical appearance of detrimental effects at the time of surgery or up to 40 months postinjection except in 1 patient who developed retinal detachment. This sets the stage for a phase II clinical trial to determine the possible beneficial effects of this procedure in patients blinded by degenerative retinal disease.

Cell-Deposited Matrix Improves Retinal Pigment Epithelium Survival on Aged Submacular Human Bruch's Membrane

PURPOSE To determine whether resurfacing submacular human Bruchs membrane with a cell-deposited extracellular matrix (ECM) improves retinal pigment epithelial (RPE) survival. METHODS Bovine corneal endothelial (BCE) cells were seeded onto the inner collagenous layer of submacular Bruchs membrane explants of human donor eyes to allow ECM deposition. Control explants from fellow eyes were cultured in medium only. The deposited ECM was exposed by removing BCE. Fetal RPE cells were then cultured on these explants for 1, 14, or 21 days. The explants were analyzed quantitatively by light microscopy and scanning electron microscopy. Surviving RPE cells from explants cultured for 21 days were harvested to compare bestrophin and RPE65 mRNA expression. Mass spectroscopy was performed on BCE-ECM to examine the protein composition. RESULTS The BCE-treated explants showed significantly higher RPE nuclear density than did the control explants at all time points. RPE expressed more differentiated features on BCE-treated explants than on untreated explants, but expressed very little mRNA for bestrophin or RPE65. The untreated young (<50 years) and African American submacular Bruchs membrane explants supported significantly higher RPE nuclear densities (NDs) than did the Caucasian explants. These differences were reduced or nonexistent in the BCE-ECM-treated explants. Proteins identified in the BCE-ECM included ECM proteins, ECM-associated proteins, cell membrane proteins, and intracellular proteins. CONCLUSIONS Increased RPE survival can be achieved on aged submacular human Bruchs membrane by resurfacing the latter with a cell-deposited ECM. Caucasian eyes seem to benefit the most, as cell survival is the worst on submacular Bruchs membrane in these eyes.

Hematopoietically derived retinal perivascular microglia initiate uveoretinitis in experimental autoimmune uveitis.

Abstract Background: Hematopoietically derived cells in the retina were studied for the expression of molecules associated with antigen presentation. Methods: Bone marrow cells of (Lewis × Brown Norway) F1 rats (LBNF1) were transplanted to sublethally irradiated Brown Norway (BN) rats to construct chimeric rats (LBNF1→BN). Each of 21 established chimeras received an adoptive transfer of uveitogenic Lewis T lymphocytes. Three rats were killed on each of 7 consecutive days. The right eye of each rat was processed for flat-mount preparation of the retina; the left eye of each was frozen for cryostat sectioning. All tissues were then stained with one of the following antibodies: OX-3 (Lewis-specific MHC class II marker), anti-ICAM, anti-B7-1, anti-TNF-α or anti-IL-1β. Results: Initial clinical signs of EAU appeared first on day 4; by day 6, full-blown EAU was noted. The flat-mount preparations revealed the presence of OX-3+ cells in the retina, perivascularly exhibiting dendritic morphology on day 2. These cells were observed in the retinal nerve fiber layer (NFL). No B7-1+, ICAM-1+, TNF-α+ or IL-1β+ cells were detected. Cryostat sections revealed positive cell staining of perivascular microglia and astrocytes in the retinal NFL with anti-IL-1β and anti-TNF-α antibodies. Conclusions: Since only perivascular bone marrow-derived cells are seen to express MHC class II molecules prior to onset of EAU, and since these cells also generate the cytokines IL-1β and TNF-α, it appears that initial presentation of antigen in the retina could be by these cells.

Stem Cell Therapy for Retinal Disease Treatment: An Update

At this time, the main targets of stem cell therapy for retinal degenerative disease are age-related macular degeneration, Stargardt disease, and retinitis pigmentosa. The goal of stem cell therapy is to either to “rescue” the surviving retinal cells (by providing the necessary support or generating neurotrophic agents) and/or to “replace” the cells that have degenerated. Stem cells being used in ongoing early human trials to treat degenerative retinal disease include induced pluripotent stem cell (iPSC)-derived retinal pigment epithelium (RPE), embryonic stem cell-derived RPE, iPSC-neural progenitor cells, bone marrow-derived stem cells, and human central nervous system derived stem cells among others. It is too early to judge the outcome of these sources of tissue, but early results are positive. Continuing research in various aspects of transplantation- establishing cell lines without danger of tumor formation or immune rejection, refining surgical techniques and instruments, and identifying factors that promote cell survival, differentiation, and integration of the transplanted cells, should allow for rapid and continued progress in the field.