Neuroprotective effects of bone marrow stromal cells in cocultures of fetal dopaminergic neuronal cultures

Abstract

Objective: The primary objective was to compare the neuroprotection conferred by diffusible factors to those mediated by direct cell-cell contacts in cocultures of bone marrow stromal cells (BMSC) and dopaminergic (DA) neurons treated with methyl-phenyl-pyridinium (MPP+). Methods: Fetal midbrain (MB) cell cultures of DA neurons were cocultured directly with green fluorescent protein (GFP+)-expressing BMSC or in bilayer cultures separated by a 0.4-µm pore semipermeable membrane. Endpoints of toxicity included survival of tyrosine hydroxylase (TH+) immunoreactive cells, lengths of the TH+ neurites, and [3H]-DA uptake. Neurotrophic factors released into media were measured. A number of cells coexpressing GFP+ and neuronal markers were counted to assess possible fusion with or transdifferentiation into DA neurons. Results: DA neurons in cocultures (mixed and bilayer) sustained significantly less damage compared to the effects of MPP+ on MB monolayer cultures. Neurite length and [3H]-DA uptake were significantly greater in bilayer cultures than in monolayer MB cell cultures. Mixed cocultures (with MB and GFP+ BMSC in direct contact) were also protected against MPP+. Levels of glial cell-derived neurotrophic factor were found to be elevated in the bilayer culture media. There were no TH+ cells that coexpressed GFP, but 7-10 percent of neurons coexpressed GFP and neuron-specific nuclear antigen suggesting possible fusion of GFP+ BMSC with non-TH+ neurons. Conclusion: Neuroprotection was observed to an equal extent in both mixed cultures and bilayer cultures in which MB and BMSC cultures were separated by a membrane. Therefore, diffusible factors elaborated by BMSC are sufficient for mitigating neurotoxicity to DA neurons. Keywords: Mesenchymal stem cells, Methyl-phenyl-pyridinium, Growth factors, Cell fusion DOI:10.5055/jndr.2013.0010