SCI (spinal cord injury) is a complex and serious neurological disease with no efficient treatment. NSC (neural stem cells) have the potential for self-renewal, proliferation and differentiation into all types of nerve cells. The aim of our study is to evaluate the effect of SCE (spinal cord extracts) from injured spinal cord on the differentiation of rat embryonic NSC and to clarify its potential mechanism. Here, NSC were isolated and cultured with SCE. The experiments were divided into four groups, including NSC + sham, NSC + SCE, NSC + SCE + DMSO (dimethyl sulfoxide), NSC + SCE + DAPT (N-[N-(3,5-difluorophenacetyl)-1-alanyl]-S-Phenyl-glycinet-butylester). The Notch1 (notch receptor 1) and Hes1 (hes family bHLH transcription factor 1) mRNA expression was analyzed by qPCR (quantitative real-time PCR) analysis. The protein expression levels of GFAP (glial fibrillary acidic protein) and NSE (nestin) were evaluated by immunofluorescence staining. Cell differentiation of NSC was induced by using neurobasal medium. The results showed that the NSC were successfully identified, and could proliferate to form spherical aggregates and was passaged continuously and steadily in vitro. The NSC at fifth generation were positively stained with NSE, and was capable of differentiating into NSE-positive cells and GFAP-positive cells. SCE treatment could upregulate the mRNA expression levels of Notch1 and Hes1, but inhibited the differentiation of NSC into neurons. DAPT could down-regulate the mRNA expression of Notch1 and Hes1 in NSC. Mechanically, DAPT targeting Notch signal pathway could facilitate NSC differentiation into neurons. Together, our data highlighted that SCE suppresses the differentiation of rat embryonic NSC by regulating the Notch signaling pathway, and DAPT treatment can reverse the effect of SCE related differentiation.
Keywords: DAPT; Neural stem cells; Notch; differentiation; spinal cord extracts.
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