The Role of Adult Bone Marrow–Derived Progenitor Cells in Experimental CNV Induced in TNFR1b–/–Knock–Out Mice

Abstract

Purpose: : The formation of CNV is closely related to both angiogenesis and vasculogenesis. An inflammatory component is also involved in the pathogenesis of ARMD. We combined the transplantation of whole bone marrow cells and laser–induced CNV in TNFR1b –/– mice to examine the development of CNV.

Methods: : Lethally irradiated adult TNFR1b –/– as well as C57BL/6 mice were used as recipients for bone marrow transplantation. Whole bone marrow mononuclear cells were obtained from gfp⁺ transgenic mice and injected into the tail vein of recipient mice. As a control, whole bone marrow obtained from C57BL/6 and TNFR1b –/– mice was transplanted to lethally irradiated C57BL/6 and TNFR1b –/– mice, respectively. CNV was induced by 4 separate laser burns of each eye. Two weeks later, mice were perfused transcardially with rhodamine–conjugated concanavalin–A (ConA). CNV was examined by dual–fluorescence scanning confocal microscopy of flatmounts. The area of CNV (µm²), as well as colocalization of gfp⁺ cells to the vasculature were determined.

Results: : One month after transplantation, FACS analysis showed 70% gfp⁺ circulating cells in recipient mice. Gfp⁺–cells were localized within and around the edge of CNV and within the optic nerve head. Some gfp⁺–cells with branching processes (F4/80–positive) were observed to infiltrate the overlying neurosensory retina. The area of neovascularization was significantly reduced in TNFR1b –/– in comparison to C57BL/6 recipients (36.3x10³+15.4 vs. 95x10³+26.6 µm²; p=0.013). Gfp⁺ fluorescence areas within the CNV were significantly reduced in TNFR1b –/– in comparison to C57BL/6 recipients (p=0.049). Many gfp⁺ cells in both groups were integrated into the neovascular tissue (co–localization of gfp⁺ and rhodamin–ConA fluorescence). However, no significant difference in co–localization areas (merged green and red images) were found between the both groups.

Conclusions: : Laser injury induced bone marrow–derived cell recruitment into the CNV lesions in both TNFR1b and C57BL/6 mice.CNV formation was reduced in TNFR1b –/– in comparison to C57BL/6 recipients. It remains to be determined how transplantation of TNFR1b –/– donor bone marrow may affect CNV–size after laser injury; and whether decreased CNV formation in TNFR1b recipients is due to decreased involvement of transplanted cells in the formation of new vessels or to other mechanisms associated with the absence of TNFR1b.

Keywords: age-related macular degeneration • choroid: neovascularization • transplantation