Liu Quan, Yu Hong-Meng, Dai Chun-Fu, Li Wen, Zhu Ya-Ying, Gu Yu-Rong, Li Hua-Wei
Department of Otolaryngology, Central Laboratory, Eye Ear Nose and Throat Hospital, Fudan University, Shanghai 200031, China.
Sheng Li Xue Bao. 2010 Apr 25;62(2):115-21.
The objective of this study is to explore whether olfactory ensheathing cells (OECs) can promote the survival of newborn rat spiral ganglion cells (SGCs) and the underlying possible mechanisms. Co-culture of OECs from adult rats with SGCs from newborn rat cochlea was established and single culture of SGCs acted as control. OECs were obtained and purified based on their special rate of attachment which was different from the other harvested cell types during culture. OECs and SGCs were immunocytochemically characterized and confirmed by expression of low-affinity nerve growth factor receptor p75 or positive label of neuron-specific betaIII-tubulin. To investigate the mechanisms of the role of OECs in survival of SGCs, brain derived neurotrophic factor (BDNF) and anti-BDNF antibody (IgY) were added into the media of the co-cultures respectively, and the surviving SGCs were examined after treatment. Single layer of OECs (92% pure) was seen seven days after plating. Surviving SGCs, which extended their primary neurites, were found on the surface of the layer in the co-cultures. When OECs and SGCs were co-cultured, the number of surviving SGCs was significantly greater than that in the single culture (P<0.01). Nine days after culture, there was even no change in the number of surviving SGCs in the co-culture while the number reduced to almost zero in the single culture. In comparison with co-culture without treatment, addition of BDNF (500 pg/mL) into the media had no obvious promoting effect on the survival of SGCs. The number of surviving SGCs reduced significantly when anti-BDNF antibody was applied into the media of co-cultures (P<0.01). These results suggest that OECs can promote the survival of SGCs when they are co-cultured in vitro. BDNF released from OECs, as one of the survival factors, plays an important role in the survival of SGCs.
本研究的目的是探讨嗅鞘细胞（OECs）是否能促进新生大鼠螺旋神经节细胞（SGCs）的存活以及潜在的可能机制。将成年大鼠的OECs与新生大鼠耳蜗的SGCs进行共培养，并将SGCs的单培养作为对照。基于OECs在培养过程中与其他收获细胞类型不同的特殊贴壁率来获取和纯化OECs。通过低亲和力神经生长因子受体p75的表达或神经元特异性βIII-微管蛋白的阳性标记对OECs和SGCs进行免疫细胞化学表征和确认。为了研究OECs在SGCs存活中作用的机制，分别向共培养物的培养基中加入脑源性神经营养因子（BDNF）和抗BDNF抗体（IgY），并在处理后检查存活的SGCs。接种七天后可见单层的OECs（纯度为92%）。在共培养物中，在该层表面发现了伸出初级神经突的存活SGCs。当OECs和SGCs共培养时，存活的SGCs数量显著多于单培养中的数量（P<0.01）。培养九天后，共培养中存活的SGCs数量甚至没有变化，而单培养中的数量减少到几乎为零。与未处理的共培养相比，向培养基中添加BDNF（500 pg/mL）对SGCs的存活没有明显的促进作用。当将抗BDNF抗体应用于共培养物的培养基中时，存活的SGCs数量显著减少（P<0.01）。这些结果表明，OECs在体外共培养时能促进SGCs的存活。从OECs释放的BDNF作为存活因子之一，在SGCs的存活中起重要作用。
