平面和三维培养对兔髓核细胞增殖和基质合成影响的实验研究-东南大学学报医学版

平面和三维培养对兔髓核细胞增殖和基质合成影响的实验研究

单位：东南大学附属中大医院,骨科,江苏,南京,210009

分类号：R-33, Q813.1, R681.55

出版年·卷·期（页码）：2008·27·第三期（178-183）

摘要：

目的:探讨兔髓核细胞平面和三维培养对其增殖及基质合成的影响,为椎间盘组织工程寻找一种新的种子细胞获取方法.方法:体外培养兔髓核细胞,分为平面培养组和PLGA支架三维培养组两组,利用倒置显微镜、扫描电镜进行髓核细胞形态学观察,MTT法测定各组髓核细胞增殖情况,HE染色、免疫组化分析髓核细胞Ⅱ型胶原的表达,阿利新兰染色法测定培养液中髓核细胞合成的糖胺聚糖(GAG)含量.结果:经过两周的培养,两组髓核细胞均能够增殖;与平面培养组相比,三维培养组细胞增殖率、Ⅱ型胶原表达量、GAG合成量均增高(均P<0.01).结论:髓核细胞能够与PLGA支架很好地黏附并增殖;PLGA支架三维培养较平面培养能促进髓核细胞增殖,提高Ⅱ型胶原和GAG的合成量,可作为椎间盘组织工程种子细胞获取的一种新方法.

Objective To study the proliferation and matrix synthesis of the nucleus pulposus(NP) cells in monolayer vs 3D culture system and thus try to find a new way to supply seed cells for intervertebral disc tissue engineering.Methods Rabbit NP cells grouped into 2 groups were cultured in vitro for two weeks.Group 1,monolayer culture;group 2,three dimensional culture in PLGA scaffold.Morphological changes were observed under inverted microscope and scanning electron microscope.The proliferation of the NP cells in the each group was tested by MTT.The expression of type Ⅱ collagen in each group was analyzed by HE-staining and immunohistochemistry.The glycosaminoglycan(GAG) content synthesized by the NP cells in the culture medium were measured by histochemistry of alcian blue.Results Two weeks after wards,the proliferation of the NP cells in both monolayer and 3D culture system was observed.Compared with the monolayer culture group,the proliferation of the NP cells,the expression of type Ⅱ collagen and the synthesis of GAG increased significantly(all P<0.01) in the 3D culture group.Conclusion Rabbit NP cells can adhere to PLGA and proliferate.3D culture in PLGA scaffold can promote the proliferation of the NP cells,and increase the synthesis of type Ⅱ collagen and GAG.This method may become a new one to supply seed cells for intervertebral disc tissue engineering.

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