体外培养人退变髓核细胞的生物学性状

背景：椎间盘退行性变后很难自行修复，研究退变髓核细胞的生物学特性可为研究椎间盘退变机制、组织工程椎间盘构建、基因治疗等提供理论基础。 目的：观察体外培养的人退变髓核细胞的生物学特性。 方法：分离培养人退变椎间盘髓核细胞，采用光镜、电镜观察细胞的形态和超微结构，荧光定量PCR技术检测髓核细胞Ⅱ型胶原和糖胺多糖mRNA的表达。ELISA法检测细胞培养上清中人Ⅱ型胶原水平，DMMB比色法检测细胞培养上清中糖胺多糖水平。 结果与结论：体外培养的传2代内的退变椎间盘髓核细胞结构与原代细胞相似，传3代后的细胞出现退变及凋亡改变。对体外培养第1代的退变髓核细胞和正常髓核细胞的Ⅱ型胶原和糖胺多糖进行检测发现，退变髓核细胞Ⅱ型胶原、糖胺多糖mRNA水平及细胞外基质中Ⅱ型胶原和糖胺多糖的表达均明显低于正常髓核细胞，呈现去分化趋势。

Biological properties of human degenerated nucleus pulposus cells cultured in vitro

BACKGROUND: Degeneration of intervertebral disc hardly self-repairs. Studying the biological properties of the degenerated nucleus pulposus cells can provide theoretical basis for studying the mechanism underlying intervertebral disc degeneration, construction of intervertebral disc by tissue engineering, and gene therapy. OBJECTIVE:To observe the biological properties of human degenerated nucleus pulposus cells cultured in vitro. METHODS: Human degenerated nucleus pulposus cells were isolated and cultured. Cell morphology and ultrastructure were observed by microscopy and electron microscopy. TypeⅡcollagen and glycosaminoglycan mRNA expression in the nucleus pulposus cells was detected by fluorescence quantitative polymerase chain reaction technique. TypeⅡcollagen content in the supernatant was detected by ELISA method. Glycosaminoglycan content in the supernatant was determined by DMMB method. RESULTS AND CONCLUSION:The structure of degenerated nucleus pulposus cells of passages 1, 2 cultured in vitro was similar to that of primary cells. Cells of passage 3 or high exhibited degenerative and apoptotic changes. Type Ⅱcollagen and glycosaminoglycan mRNA expression in the nucleus pulposus cells and typeⅡcollagen and glycosaminoglycan content in the supernatant, exhibiting a tendency of dedifferentiation, were obviously lower in the degenerated nucleus pulposus cells than in the normal nucleus pulposus cells.