Department of Plastic and Reconstructive Surgery, Shanghai General Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200080, P.R. China.
Department of Laboratory Animal Science, Shanghai Jiao Tong University School of Medicine, Shanghai 200025, P.R. China.
Mol Med Rep. 2017 Nov;16(5):6600-6607. doi: 10.3892/mmr.2017.7427. Epub 2017 Sep 5.
Schwann cells (SCs) are indispensable for cell therapy and tissue engineering of the peripheral nervous system. Easy access to activated, highly proliferative SCs is necessary for clinical applications. The present study developed a fast, efficient method for obtaining highly purified SCs from the peripheral nerve of a mature Rhesus monkey. The common peroneal nerves of 4‑year‑old Rhesus monkeys were harvested and subjected to in vitro pre‑degeneration in a modified SC culture medium (SCCM). The nerve pieces were subsequently treated enzymatically to dissociate the cells and then cultured for 2 days in SCCM. Cultured cells were treated with purification medium containing Ara‑C to assist in restricting the overgrowth of fibroblast‑like cells, for 24 h. After another 24‑h cultivation period, the cells were subsequently treated with a multiplex collagenase, which enabled SC detachment over fibroblast detachment, and thereby facilitated SC isolation. Finally, SCs were cultured in SCCM. The cell yield was determined by cell counting following enzyme digestion and SC purity was determined from the percentage of SCs with respect to the total number of cells. Following purification, 96.3±3.9% of cells were identified as SCs. In vitro pre‑degeneration in the presence of basic‑fibroblast growth factor, heregulin β1 and forskolin maximized the purity and yield of SCs that could be obtained from monkey peroneal nerves. The present study identified a novel technique that can efficiently isolate and purify SCs from mature monkey nerves based on in vitro pre‑degeneration.
许旺细胞（SCs）对于周围神经系统的细胞治疗和组织工程是不可或缺的。为了临床应用，需要容易获得激活的、高增殖性的SCs。本研究开发了一种从成熟恒河猴周围神经中获得高纯度SCs 的快速、有效的方法。从 4 岁恒河猴的腓总神经中获取外周神经，并在改良的SCs 培养基（SCCM）中进行体外预退变。随后，用酶处理神经段以解离细胞，然后在 SCCM 中培养 2 天。在含有 Ara-C 的纯化培养基中处理培养的细胞，以帮助限制成纤维细胞样细胞的过度生长，持续 24 小时。经过另一个 24 小时的培养期后，用多酶胶原酶处理细胞，使SCs 脱离而纤维母细胞不受影响，从而促进SCs 的分离。最后，SCs 在 SCCM 中培养。通过酶消化后细胞计数来确定细胞产量，并根据SCs 数量占总细胞数量的百分比确定SCs 的纯度。经过纯化后，96.3±3.9%的细胞被鉴定为SCs。在碱性成纤维细胞生长因子、heregulin β1 和 forskolin 的存在下进行体外预退变，可最大限度地提高从猴腓总神经中获得的SCs 的纯度和产量。本研究确定了一种新的技术，可以从成熟猴神经中有效分离和纯化SCs。
