Influence of different blood serum concentration and growth time on olfactory ensheathing cells from the olfactory mucosa of adult rats

doi:10.3969/j.issn.2095-4344.2014.07.016

Abstract

Abstract:

BACKGROUND: Preliminary experiments have reported the influence of serum and nerve growth factor on olfactory ensheathing cells proliferation in vitro, but there are less studies concerning choice of serum concentration and growth time for in vitro culture of olfactory ensheathing cells.
OBJECTIVE: To find out the influence of different blood serum concentration and growth time on olfactory ensheathing cells from the olfactory mucosa of adult rats based on the growth curve of olfactory ensheathing cells.
METHODS: The olfactory ensheathing cells from the olfactory mucosa of adult rats were separated, culture and identified in vitro. Sulforhodamine B and microplate reader were employed to measure absorbance values and plot growth curve of olfactory ensheathing cells.
RESULTS AND CONCLUSION: When cultured for the same time in blood serum of different concentrations, absorbance values, especially in the groups 10%, 20%, 30%, 40%, tended to increase with time except the 0% group. When cultured in the same serum for different time, absorbance values increased within the first 9 days, then promoted rapidly in the groups 10%, 20%, 30%, 40% at 13 days, entered the plateau phase at 19 days, and decreased at 23 days; meanwhile, in the other groups (50%, 60%, 70%, 80%, 90%) the absorbance values peaked at the 13th day and then decreased gradually. These findings indicate that different serum concentrations and different growth time in vitro affect cell growth and survival of olfactory ensheathing cells significantly, which should be fully considered when cells are cultured in an in vitro condition.

中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

全文链接：

Key words: olfactory mucosa, cells, microglia, growth charts, actihaemyl, nerve growth factor

CLC Number:

R318

Zhang Zhi-qiang, Li Yi-nan, Zhou Ying, Yang Jian-dong, Feng Xin-min. Influence of different blood serum concentration and growth time on olfactory ensheathing cells from the olfactory mucosa of adult rats[J]. Chinese Journal of Tissue Engineering Research, 2014, 18(7): 1076-1081.

Figures/Tables 1

2.1 供体动物的术后存活情况动物经手术取出部分嗅黏膜后，一两天内呼吸频率加快，鼻腔内有少量血性分泌物，活动及饮食正常。手术切口于1周内愈合，术后2只SD大鼠均能长期存活。 2.2 嗅鞘细胞的形态学观察及免疫荧光染色培养的嗅黏膜细胞由不同形态的细胞所组成。动态观察发现这些细胞刚贴壁时为纺锤形，从胞体向外周伸出许多丝状伪足，相邻细胞的丝状伪足相互吸引。经三四次传代培养后细胞大多为长梭形，突起细长，数量多，并成为优势细胞，见图1。经嗅鞘细胞特异性免疫细胞化学和免疫荧光染色证实，大部分双极或梭形细胞呈GFAP和NGFPRp75免疫反应阳性为嗅鞘细胞(图1A-C)。 2.3 磺基罗丹明B法测定细胞生长曲线[10] 血清浓度对细胞生长影响的实验结果：血清浓度0%组的A值明显低于其他浓度组，血清浓度10%-90%组细胞均可以正常生长。随着血清浓度的增加，特别是血清浓度在10%-40%，对细胞生长起到明显的促进作用(相对于0%组，P=0.000)。当血清浓度继续增大，达到50%-90%时，对细胞生长仍有一定的促进作用(相对于0%组，P < 0.05)，只是对细胞生长促进的作用较血清浓度在10%到40%组明显变弱(P=0.000，图1D)。生长时间对细胞生长影响的实验结果：随着细胞生长时间的延长，除0%组A值只呈微弱的降低趋势外，其余浓度组A值呈现增加状态。各浓度组A值在5天开始增加。生长时间在5-9 d时，A值增加趋势较弱，第9天之后，增加趋势又有所增强。血清浓度10%-40%组，在第13天，细胞生长迅速增快，第19天左右进入一个平台期，在第23天，A值有减弱的趋势。而血清浓度50%-90%组，在第13天，细胞生长似乎达到了峰值，其后随着时间的延长，A值逐渐变小(图1E)。"

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