Abstract
The bone marrow (BM) is home to different stem/progenitor populations, including tissue-committed stem cells. In this context, we have cocultured BM-derived stem cells (BMSC) in order to investigate their differentiation capacity towards the retinal pigment epithelial (RPE) lineage in vitro . Furthermore, pre-differentiated BMSC were transplanted into the pharmacologically damaged subretinal space to determine their rescue ability in vivo . BM was harvested from the tibias and femurs of adult GFP + C57BL/6 mice. Differentiated hematopoietic cells were removed by lineage depletion, and CD45 - BMSCs were separated by magnetic activated cell sorting (MACS). To induce differentiation, the cells were then cocultured with murine RPE for 10 days, and retinal markers were assessed using immunohistochemistry (IHC). To induce retinal degeneration, mice were treated with sodium iodate (NaIO 3 ). Seven days later, approx. 60,000 pre-differentiated GFP + BMSC, sorted by FACS, were transplanted subretinally. Optical coherence tomography (OCT) was used to follow the transplants and to quantify the retinal thickness over time. Visual acuity was measured concurrently using the optokinetic reflex (OKR). Finally, IHC was performed to investigate the expression of retina-specific markers in the transplants. CD45 - BMSC adopted an RPE-like elongated morphology and showed expression of the RPE markers RPE65 and bestrophin after coculture. After transplantation of CD45 - BMSC, visual acuity increased in individual animals compared to the contralateral control eye, but did not reach baseline levels. Additionally, no significant increase in retinal thickness in the transplanted eye was found. However, the cells were detectable in the subretinal space for up to 28 days and expressed the RPE markers RPE65 and bestrophin. In summary, the BMSC differentiated into RPE-like cells but were not able to restore visual function or rescue retinal morphology after subretinal transplantation.
Highlights
Replacement of the degenerated retinal pigment epithelium (RPE) before irreversible degeneration of the foveal photoreceptors occurs was considered as a potential therapy for age-related macular degeneration (AMD) nearly 20 years ago [1]
The CD45- bone marrow-derived stem cells (BMSCs) showed no expression of the glia marker GFAP, the early neuronal marker βIII- tubulin or the late neuronal marker MAP-2 after coculture with RPE
To avoid contamination by feeder cells that had taken up GFP, the sorting parameters were selected with appropriate controls
Summary
Replacement of the degenerated retinal pigment epithelium (RPE) before irreversible degeneration of the foveal photoreceptors occurs was considered as a potential therapy for age-related macular degeneration (AMD) nearly 20 years ago [1]. New findings have recently contradicted the central dogmas of commitment of adult SC, including bone marrow-derived stem cells (BMSCs), by showing their plasticity to differentiate across tissue-lineage boundaries, irrespective of classical germ layer designations [6]. In this context transdifferentiation into non-hematopoietic, including retinal, cell types was documented [7,8,9]. BMSCs home to the subretinal space if systemically transferred or endogenously mobilized where they express RPE lineage markers [13,14,15]
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