摘要
背景:生长因子参与调节嗅鞘细胞增殖、分化及代谢功能,因此有望通过应用其合理的序贯刺激形式,达到细胞数量地大量扩增。目的:观察牛垂体提取物、碱性成纤维细胞生长因子及Forskolin对嗅鞘细胞增殖的影响及剂量-效应关系。方法:采用原代培养技术,从新生大鼠的嗅球分离培养嗅鞘细胞,并用差速贴壁+化学药物+胰酶快速消化法纯化培养嗅鞘细胞,添加牛垂体提取物、碱性成纤维细胞生长因子及Forskolin 3种促增殖因子及其组合。MTT法检测各组嗅鞘细胞生长的情况。根据NGFRp75抗体的免疫细胞化学染色统计嗅鞘细胞的纯度和计算嗅鞘细胞的增殖率。结果与结论:在嗅鞘细胞原代培养和纯化后,添加牛垂体提取物具有促进嗅鞘细胞增殖的效应,但未见典型的剂量-效应关系;添加碱性成纤维细胞生长因子具有促进嗅鞘细胞增殖的效应,且呈现典型的剂量-效应关系;添加Forskolin并不具有促进嗅鞘细胞增殖的效应。说明合理应用不同质量浓度的碱性成纤维细胞生长因子与Forskolin进行联合刺激,能有效促进细胞的增殖。结果表明序贯的使用合适浓度的生长因子组合,将是有效促进嗅鞘细胞增殖的合理策略,有助于解决脊髓损伤治疗中种子细胞来源不足这一问题。
BACKGROUND：Growth factors participate in regulating proliferation,differentiation and metabolism of olfactory ensheathing cells（OECs）.Therefore,reasonable sequential stimulation form can be used to reach multiple amplification of cell number.OBJECTIVE：To investigate the effect of basic fibroblast growth factor（bFGF）,bovine pituitary extract and Forskolin on OECs proliferation as well as dose-effect relationship.METHODS：The OECs were isolated from the neonatal rat olfactory bulb using primary culture technique.The OECs were purified using differential velocity adherent ＋ chemicals ＋ trypsin digestion method.The bFGF,bovine pituitary extract and Forskolin alone and their combination were added.MTT assay was utilized to detect OEC growth in each group.The purity of the OECs was evaluated and growth rate of OECs was calculated using NGFR p75 immunocytochemistry.RESULTS AND CONCLUSION：Following primary culture and purification of OECs,bovine pituitary extract could facilitate OECs growth,but typical dose-effect relationship was not found.bFGF could facilitate OECs growth,which presented typical dose-effect relationship.An additional Forskolin could not promote OEC proliferation.These indicated that combined use of bFGF and Forskolin in a proper concentration significantly enhanced cell proliferation.Results suggest that sequent use of combined growth factors in a proper concentration would be a reasonable strategy to effectively enhance cell proliferation and this might be an ideal way in dealing with seed cell scarcity for treatment with spinal cord injury.
出处
《中国组织工程研究与临床康复》
CAS
CSCD
北大核心
2010年第40期7447-7452,共6页
Journal of Clinical Rehabilitative Tissue Engineering Research
