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[ADIPOSE-DERIVED STEM CELLS DIFFERENTIATION INTO NEURON-LIKE CELLS INDUCED BY CO-CULTURE WITH SCHWANN CELLS].

OBJECTIVE
To investigate the differentiation of rat adipose-derived stem cells (ADSCs) into neuron- like cells by indirect co-culture with Schwann cells (SCs) in vitro so as to look for the ideal seed cells for tissue engineering.


METHODS
SCs were isolated from sciatic nerves of 1-2 days old Sprague-Dawley rats with enzymatic digestion method. Immunofluorescence staining was used to identify SCs with the marker S-100. ADSCs were isolated from the epididymal fat pads of adult male Sprague-Dawley rats by means of differential attachment. And the cell phenotypes (CD29, CD34, CD45, CD73, CD90, and CD105) of ADSCs at passage 3 were determined by flow cytometry analysis. Primary SCs and ADSCs at passage 3 were co-cultured at a ratio of 2:1 in Transwell culture dishes (experimental group), and ADSCs cultured alone served as control group. Immunofluorescence and flow cytometry were adopted to investigate the neural differentiation of ADSCs at 14 days. The expression differences for neuron-specific enolase (NSE), microtubule-associated protein 2 (MAP2), neuronal nuclei protein (NeuN), and glial fibrillary acidic protein (GFAP) were detected, and the percentage of positive cells was calculated.


RESULTS
ADSCs were successfully extracted and can passage in a considerable large amount. Flow cytometry analysis showed that ADSCs at passage 3 were positive for CD29, CD90, CD73, and CD 105 expression, but negative for CD34 and CD45 expression. The ADSCs of the experimental group showed contraction of nucleus, increasing of soma refraction , and several long and thick protrusions of cell body. The cell shape had no obvious change in the control group. Both immunofluorescence and flow cytometry analysis results showed the expressions of MAP2, NSE, NeuN, and GFAP at 14 days after co-cultured with SCs, and the positive cell ratios were significantly higher than those in the control group (P < 0.01).


CONCLUSION
Co-culture with SCs not only can promote the survival regeneration of ADSCs, but also can induce the differentiation of ADSCs into neuron-like cells.