Zhang Lei, Tan Xuefeng, Dong Chuanming, Zou Linqing, Zhao Heyan, Zhang Xinhua, Tian Meiling, Jin Guohua
Department of Anatomy and Neurobiology, The Jiangsu Key Laboratory of Neuroregeneration, Nantong University, Nantong 226001, People's Republic of China.
Int J Dev Neurosci. 2012 Oct;30(6):471-7. doi: 10.1016/j.ijdevneu.2012.05.006. Epub 2012 Jun 5.
We isolated and expanded fibroblast-like cells from the Wharton's jelly of human umbilical cord successfully. Immunocytochemistry showed that they were positive for several markers of mesenchymal stem cells (CD73, CD90, and CD105) and integrin markers (CD29 and CD44), but negative for a hematopoietic cell maker (CD45) and an endothelial cell marker (CD31). Their differentiation into osteocytes and adipocytes under specific conditions indicated that they had multi-lineage differentiation potential. Therefore these results proved that the cells we obtained from Wharton's jelly were human umbilical cord mensenchymal stem cells (hUCMSCs). Using immunocytochemistry and Western blotting analysis, we found that after treatment with neuronal induction medium [NIM; consisting of brain-derived neurotrophic factor (BDNF) and low-serum media] for 14 days, hUCMSCs expressed a neuronal specific marker, microtubule associated protein 2 (MAP2), and extended neurite-like processes. After treatment with NIM, supplemented with hippocampal cholinergic neurostimulating peptide (HCNP) or rat denervated hippocampal extract [rDHE; derived from rat fimbria fornix (FF) transected hippocampus], hUCMSCs expressed choline acetytransferase (ChAT) and this action could be enhanced when cells were cultured with NIM, supplemented with HCNP and rDHE in combination. ELISA showed that these ChAT-positive cells could secrete acetylcholine (ACh). These findings indicate that hUCMSCs possess the potential of differentiation into functional ChAT-positive cells in vitro and provide a new candidate of cells for the cell transplantation to treat Alzheimer's disease (AD).
我们成功地从人脐带华通氏胶中分离并扩增出成纤维细胞样细胞。免疫细胞化学显示，它们对间充质干细胞的几种标志物（CD73、CD90和CD105）以及整合素标志物（CD29和CD44）呈阳性，但对造血细胞标志物（CD45）和内皮细胞标志物（CD31）呈阴性。它们在特定条件下分化为骨细胞和脂肪细胞，表明它们具有多向分化潜能。因此，这些结果证明我们从华通氏胶中获得的细胞是人脐带间充质干细胞（hUCMSCs）。通过免疫细胞化学和蛋白质印迹分析，我们发现用神经元诱导培养基[NIM；由脑源性神经营养因子（BDNF）和低血清培养基组成]处理14天后，hUCMSCs表达神经元特异性标志物微管相关蛋白2（MAP2），并伸出神经突样突起。用补充了海马胆碱能神经刺激肽（HCNP）或大鼠去神经海马提取物[rDHE；源自切断海马穹窿海马伞（FF）的大鼠海马]的NIM处理后，hUCMSCs表达胆碱乙酰转移酶（ChAT），当细胞与补充了HCNP和rDHE组合的NIM一起培养时，这种作用会增强。酶联免疫吸附测定（ELISA）表明，这些ChAT阳性细胞可以分泌乙酰胆碱（ACh）。这些发现表明，hUCMSCs在体外具有分化为功能性ChAT阳性细胞的潜力，并为细胞移植治疗阿尔茨海默病（AD）提供了一种新的细胞候选物。
