Engraftment of adult neural progenitor cells transplanted to rat retina injured by transient ischemia

PURPOSE. To optimize delivery parameters for achieving engraftment, migration, and differentiation of adult neural progenitor cells transplanted to the retinas of rats after transient retinal ischemia. METHODS. Retinal ischemia was induced by transiently raising the intraocular pressure. Some animals then received transplantation of green fluorescent protein (GFP)-expressing cells derived from the adult rat hippocampus and were allowed to recover for 6 hours to 9 weeks. Retinal cryosections were prepared for TUNEL analysis to determine the time course of ischemia-induced cell death, and some sections were prepared for immunohistochemistry for retinal neuronal antigens. RESULTS. TUNEL analysis revealed that ischemia-induced cell death peaked at 24 hours. By 96 hours, the inner nuclear (INL) and ganglion cell (GCL) layers were largely obliterated in the central retina, sparing peripheral regions. By 2 weeks after transplantation, numerous GFP-expressing cells had engrafted into the host retina, migrated to the inner retina, and extended processes. At 4 weeks, many GFP-labeled cells were present throughout the INL and displayed horizontal-, bipolar-, and amacrine cell-like morphologies. GFP-expressing cells were also present in the GCL with fibers extending into the nerve fiber layer. At 5 weeks, many GFP-expressing cells were present at the optic nerve head, and some GFP-labeled fibers were present in the optic nerve, occasionally passing through the full extent of the lamina cribrosa. Only rarely were GFPexpressing cells found that coexpressed retinal phenotypic markers at any time point examined. CONCLUSIONS. Adult hippocampus-derived neural progenitor cells transplanted to the subretinal space readily engraft into a host retina that has undergone ischemic injury. Many cells migrate to specific retinal cellular layers and undergo limited morphologic differentiation reminiscent of retinal neurons, including extension of processes into the optic nerve. Concurrent control studies demonstrate that optimal engraftment is achieved by subretinal delivery within a specific temporal window. These results imply that certain inductive cues may be regulated after injury, and they demonstrate the potential for adult neural progenitor cell transplantation for the treatment of retinal neurodegenerative diseases. (Invest Ophthalmol Vis Sci. 2003;44:3194 -3201) DOI:10.1167/iovs.02-0875 N eurodegenerative diseases of the inner retina are major causes of blindness worldwide. These include the glaucomas, which primarily affect retinal ganglion cells, and ischemic retinopathies (including diabetic and hypertensive retinopathies) that affect other populations of inner retinal neurons as well. 1,2 Current therapies for inner retinal degenerations are largely retardant and are not sufficient to restore visual function after severe impairment of inner retinal circuitry. Numerous neuroprotective strategies have been used in an effort to prolong the survival of inner retinal neurons damaged in rodent models of inner retinal degeneration. 3-13 However, many neurodegenerative diseases of the inner retina, such as primary open-angle glaucoma (POAG) are quite protracted, extending over many months to years. Experimental models for these diseases have, by necessity, compressed this time course dramatically. The mechanisms of ganglion cell death (and potential neuroprotection) have been examined extensively in models of optic nerve transection or crush 14 -17 and in models using either acute ischemia followed by reperfusion 18 -20 or chronic elevation of intraocular pressure (IOP). 12,13 These experimental models also often result in degeneration of cells in the inner nuclear layer (INL). However, preservation of these cells and inner retinal circuitry is still problematic. Recent advances in stem cell biology have invigorated the potential for achieving partial restoration of visual function after inner retinal neurodegeneration by augmenting the remaining inner retinal circuitry. It has been demonstrated that neural progenitor cells can be isolated from near the ventricular wall of the adult brain, as well as from the adult hippocampus

Bone Marrow Transplantation Transfers Age-Related Susceptibility to Neovascular Remodeling in Murine Laser- Induced Choroidal Neovascularization

Citation: Espinosa-Heidmann DG, Malek G, Mettu PS, et al. Bone marrow transplantation transfers age-related susceptibility to neovascular remodeling in murine laser-induced choroidal neovascularization. Invest Ophthalmol Vis Sci. 2013;54:7439-7449. DOI:10.1167/iovs.13-12546 PURPOSE. Neovascular remodeling (NVR), the progression of small capillaries into large-caliber arterioles with perivascular fibrosis, represents a major therapeutic challenge in neovascular age-related macular degeneration (AMD). Neovascular remodeling occurs after laser-induced choroidal neovascularization (CNV) in aged but not young mice. Additionally, bone marrowderived cells, including macrophages, endothelial precursor cells, and mesenchymal precursor cells, contribute to CNV severity. In this study, we investigated the impact of aged bone marrow transplantation (BMT) on the degree of fibrosis, size, and vascular morphology of CNV lesions in a mouse model of laser-induced CNV. METHODS. Young (2 months) and old (16 months) mice were transplanted with green fluorescent protein (GFP)-labeled bone marrow isolated from either young or old donors. Laser CNV was induced 1 month following transplant, and eyes were analyzed via choroidal flat mounts and immunohistochemistry 1 month postlaser. The identity of cells infiltrating CNV lesions was determined using specific markers for the labeled transplanted cells (GFPþ), macrophages (F4/80þ), perivascular mesenchymal-derived cells (smooth muscle actin, SMAþ), and endothelial cells (CD31þ). RESULTS. Bone marrow transplantation from aged mice transferred susceptibility to NVR into young recipients. Inversely, transplantation of young marrow into old mice prevented NVR, preserving small size and minimal fibrosis. Mice with NVR demonstrated a greater relative contribution of marrow-derived SMAþ perivascular mesenchymal cells as compared to other cells. CONCLUSIONS. Our findings indicate that the status of bone marrow is an important determining factor of neovascular severity. Furthermore, we find that perivascular mesenchymal cells, rather than endothelial cells, derived from aged bone marrow may contribute to increased CNV severity in this murine model of experimental neovascularization

Estrogen receptor β protects against in vivo injury in RPE cells

Epidemiological data suggest that estrogen deficiency in postmenopausal women may contribute to the severity of AMD. We discovered that 17β-estradiol (E 2) was a crucial regulator of the severity of extracellular matrix turnover (ECM) dysregulation both in vivo and in vitro. We also found in vitro that the presence of estrogen receptor (ER)β regulates MMP-2 activity. Therefore in an attempt to delineate the role of the ER subtypes, female estrogen receptor knockout (ERKO) mice were fed a high-fat diet, and the eyes were exposed to seven 5-second doses of nonphototoxic levels of blue-green light over 2 weeks. Three months after cessation of blue light treatment, transmission electron microscopy was performed to assess severity of deposits, Bruchs membrane changes, and choriocapillaris endothelial morphology. We found that changes in the trimolecular complex of pro-MMP-2, MMP-14 and TIMP-2 correlated with increased Bruch's membrane thickening or sub-retinal deposit formation (basal laminar deposits) in ERKOβ mice. In addition RPE isolated from ERKOβ mice had an increase in expression of total collagen and a decrease in MMP-2 activity. Finally we found that ERK an intermediate signaling molecule in the MMP pathway was activated in RPE isolated from ERKOβ mice. These data suggest that mice which lack ERβ are more susceptible to in vivo injury associated with environmental light and high fat diet

Bone Marrow Transplantation Transfers Age-Related Susceptibility to Neovascular Remodeling in Murine Laser-Induced Choroidal Neovascularization

PURPOSE. Neovascular remodeling (NVR), the progression of small capillaries into large-caliber arterioles with perivascular fibrosis, represents a major therapeutic challenge in neovascular age-related macular degeneration (AMD). Neovascular remodeling occurs after laser-induced choroidal neovascularization (CNV) in aged but not young mice. Additionally, bone marrow–derived cells, including macrophages, endothelial precursor cells, and mesenchymal precursor cells, contribute to CNV severity. In this study, we investigated the impact of aged bone marrow transplantation (BMT) on the degree of fibrosis, size, and vascular morphology of CNV lesions in a mouse model of laser-induced CNV. METHODS. Young (2 months) and old (16 months) mice were transplanted with green fluorescent protein (GFP)-labeled bone marrow isolated from either young or old donors. Laser CNV was induced 1 month following transplant, and eyes were analyzed via choroidal flat mounts and immunohistochemistry 1 month postlaser. The identity of cells infiltrating CNV lesions was determined using specific markers for the labeled transplanted cells (GFP+), macrophages (F4/80+), perivascular mesenchymal-derived cells (smooth muscle actin, SMA+), and endothelial cells (CD31+). RESULTS. Bone marrow transplantation from aged mice transferred susceptibility to NVR into young recipients. Inversely, transplantation of young marrow into old mice prevented NVR, preserving small size and minimal fibrosis. Mice with NVR demonstrated a greater relative contribution of marrow-derived SMA+ perivascular mesenchymal cells as compared to other cells. CONCLUSIONS. Our findings indicate that the status of bone marrow is an important determining factor of neovascular severity. Furthermore, we find that perivascular mesenchymal cells, rather than endothelial cells, derived from aged bone marrow may contribute to increased CNV severity in this murine model of experimental neovascularization