Neural differentiation of rat adipose tissue-derived stem cells in vitro

Abstract

Objective To probe into the potential of neural differentiation of rat adipose-derived stem cells (ADSCs) in vitroto provide foundation to restore erectile dysfunction (ED) caused by cavernous nerves (CNs) injury.Methods Adipose tissue from SD rat abdomen and caputepididymidis was digested with collagenase type Ⅰ,followed by filtering and centrifugation.The isolated adipose stromal cells were cultured in dishes.ADSCs markers were measured by flow cytometry.Cells at the third passage were used for in vitro differentiation. Adipogenic and neuronal differentiation was induced by incubation of ADSCs with different induction media.Oil red O staining was carried out to identify adipose cells,and immunofluorescence to detect protein of glial fibrillary acidic protein (GFAP) and beta tubulin Ⅲ.Results Flow cytometry demonstrated that the ADSCs expressed CD44,but CD45 and CD34 were weakly expressed,and their positive rate was 96.4％,1.7％,and 0.9％ respectively.Adipose cells as evidenced by the lipid droplet were dyed red by oil red O dyeing.Neuron-like cells were evidenced by neuronal morphology and the presence of neuronal markers including GFAP and β-tubulin Ⅲ.The induction rate of GFAP and β-tubulin Ⅲ by method 1 [epidermal growth factor (EGF),basic fibroblast growth factor (bFGF),brain-derived neurotrophic factor (BDNF) + indomethacin,insulin,and 3-isobutyl-1-methylxanthine (IBMX)]and method 2 ( EGF,bFGF + indomethacin,insulin,and IBMX) was (74.0 ± 3.3 ) ％ and (65.3 ±2.1 ) ％,and (51.0 ± 1.2) ％ and (41.0 ± 1.1 ) ％ respectively.The number of the positive cells exhibited significant difference after the induction ( P ＜ 0.01 ).Conclusion ADSCs can be easily obtained from a small amount fat tissue and developed in culture.By using the method 1,the induction rate is high and induction time is short in neural differentiation of ADSCs.ADSCs may be an alternative source of stem cell therapy for neuropathic ED. Key words: Adipose tissue-derived stem cells; Neural cells; Differentiation