碱性成纤维细胞生长因子缓释微球对许旺细胞在小肠粘膜下层增殖活性的影响

Study on the Activities of SC Proliferation with Basic Fibroblast Growth Factor Controlled Release Microspheres Combined with Schwann Cells and Small Intestinal Submucosa

目的：观察碱性成纤维细胞生长因子（bFGF）缓释微球对许旺细胞（SC）在小肠粘膜下层（SIS）上增殖影响。方法运用双酶两步消化法对 SC 进行体外分离、培养、纯化,将 bFGF-PLGA 缓释微球（bFGF-PLGA-MS）与 SC 及 SIS进行体外复合培养,并以游离 bFGF 组及单纯培养液组进行对比,观察 bFGF-PLGA 缓释微球对 SC 在 SIS 上增殖影响。结果运用双酶两步消化的体外原代培养方法获取的许旺细胞数量较多,细胞纯度可达90%以上。细胞的体外增殖活性良好,细胞增殖周期约为6～7d,培养后2～7d 为对数生长期,第7d 后进入生长平台期； bFGF 缓释微球能持续地促进许旺细胞在 SIS 上的增殖；细胞增殖周期缩短,细胞能较稳定地保持着良好的活性；提高了细胞增殖指数,并使 SIS 上的 SC持续保持较高的增殖活性；细胞数量与 bFGF 含量的相关系数为0.988（P =0.02）,表明两者间有明显正相关关系。结论bFGF-PLGA 微球的药物缓释促进了SC 在SIS 上持续而稳定的增殖,为以SC 复合SIS 构建人工神经提供了有利条件。

Objective To observe the effects of basic fibroblast growth factor（ bFGF） micropheres on adhesion and proliferation and migration of schwann cells（SC） on scaffold of small intestinal submucosa（SIS）, and the activities of SC proliferation was observed. Methods Schwann cells was isolated ,cultured and purified in vitro using two-step enzymatic digestion method, To explore the effects of bFGF- PLGA- MS on adhesion and proliferation and migration of schwann cell（SC） on the scaffolds of SIS in vitro, and the free bFGF group and simple medium group was used as control. Results A larger number of active Schwann cells with the cell purity above 90% were obtained using two-step enzyme digestion method a of primary cells culture in vitro, the good activity of cells in vitro were confirmed, the cell cycle is about 6 ～ 7 days and the logarithmic growth phase is 2 ～ 7 days after culutre, The bFGF-MS was observed continuously promoting the adhesion and proliferation and migration of Schwann cells on SIS, the cell cycle of SC be shortened by bFGF-MS with a more stable and good activity of cells, according to flow cytometry, the bFGF microspheres increased cell proliferation index, and continuously maintained a higher proliferative activity of SC on SIS, it showed a clear positive correlation between the content of bFGF released from microspheres and cells counts, with the correlation coefficient of 0. 988 （P = 0. 02）. Conclusion The drug delivery of bFGF-PLGA-MS promotes Schwann cells sustained and stable adhesion and proliferation on the scaffold of small intestinal submucosa, matched with the drug release rate. Schwann cells are provided to be constructed artificial nerve combined with SIS.