| Research Abstract |
We have developed a novel method to label live neural stem cells with green fluorescent protein (GFP) using an enhancer element of the nestin gene. Similarly, we have constructed the TN-GFP reporter, in which GFP expression is induced in dopaminergic neurons under the control of the tyrosine hydroxylase (TN) gene promoter. Using these transgenes, we isolated neural stem cells and dopaminergic neurons and transplanted them into the striatum of a rat model of the Parkinson's disease. We found that these cells survived well and differentiated into mature dopaminergic neurons in the host striatum, resulting in a significant functional recovery. Moreover, we introduced the TN-GFP reporter into mouse ES cells in order to label and isolate dopaminergic neurons derived from ES cells. GFP-positive cells were transplanted into the striatum of a rat model of Parkinson's disease. Consequently, amphetamine-induced rotation behavior was significantly decreased, indicating that these GFP-based methods are useful to isolate dopaminergic neurons derived from ES cells. However, these methods need introduction of a transgene, which is hardly applicable to human cells before the transplantation into patients. We then performed a large-scale screening for monoclonal antibodies that recognize cell surface molecules expressed in neural stem cells or its progenies. We chose two clones based on their staining pattern on tissue sections and FACS analyses for further analyses. Using an expression-cloning method, we cloned cDNAs encoding the antigens reactive with these antibodies, and identified the EprinB1 and M6a genes. There is a possibility that these proteins might be involved in proliferation and/or differentiation of neural stem cells. It is also likely that these antibodies are useful for functional studies of these proteins and for FACS-isolation of neural stem cells and neurons.
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