PURPOSE: To investigate and compare the effects of two different protocols on inducing bone marrow stromal cells (BMSCs) to differentiate into retinal neurons. METHODS: BMSCs from adult rat femurs were cultured and passaged 3 times. The cells were then induced by either the basic fibroblast growth factor (bFGF) protocol or by co-culturing with neonatal rat retina. Morphological changes were monitored and immunocytochemistry was used to detect the expression of neuronal and retinal neuron-specific markers. RESULTS: The bFGF protocol induced a maximum of 74% of the BMSCs to assume a neuronal-like morphology, but this was also associated with significant cell detachment during the 7-day culture period. By comparison, with the co-culture method only a maximum of 10% of cells became neuronal-like, but without marked cell detachment. The cells expressed microtubule-associated protein-2 (MAP-2, a neuronal marker) and Thy1.1 (a retinal ganglion cell marker) at all time points under both protocols. The percentage of MAP-2- and Thy1.1-positive cells was much higher following bFGF induction compared to co-culture induction. Following bFGF induction a small number of opsin-positive cells (a photoreceptor marker) were observed only on day 1. These cells were first observed at day 5 with co-culturing. The expression of protein kinase Calpha (a bipolar cell marker) and glial fibrillary acidic protein was not detected after either protocol. CONCLUSION: BMSCs can express retinal neuron-specific phenotypic markers in response to bFGF induction or retinal co-cultures in vitro. When a short induction time is used the bFGF induction is superior, albeit with certain limitations, to retinal co-cultures for inducing BMSCs to express retinal neuron-specific markers. Copyright 2009 S. Karger AG, Basel.
PURPOSE: To investigate and compare the effects of two different protocols on inducing bone marrow stromal cells (BMSCs) to differentiate into retinal neurons. METHODS: BMSCs from adult rat femurs were cultured and passaged 3 times. The cells were then induced by either the basic fibroblast growth factor (bFGF) protocol or by co-culturing with neonatal rat retina. Morphological changes were monitored and immunocytochemistry was used to detect the expression of neuronal and retinal neuron-specific markers. RESULTS: The bFGF protocol induced a maximum of 74% of the BMSCs to assume a neuronal-like morphology, but this was also associated with significant cell detachment during the 7-day culture period. By comparison, with the co-culture method only a maximum of 10% of cells became neuronal-like, but without marked cell detachment. The cells expressed microtubule-associated protein-2 (MAP-2, a neuronal marker) and Thy1.1 (a retinal ganglion cell marker) at all time points under both protocols. The percentage of MAP-2- and Thy1.1-positive cells was much higher following bFGF induction compared to co-culture induction. Following bFGF induction a small number of opsin-positive cells (a photoreceptor marker) were observed only on day 1. These cells were first observed at day 5 with co-culturing. The expression of protein kinase Calpha (a bipolar cell marker) and glial fibrillary acidic protein was not detected after either protocol. CONCLUSION: BMSCs can express retinal neuron-specific phenotypic markers in response to bFGF induction or retinal co-cultures in vitro. When a short induction time is used the bFGF induction is superior, albeit with certain limitations, to retinal co-cultures for inducing BMSCs to express retinal neuron-specific markers. Copyright 2009 S. Karger AG, Basel.