体外培养人正常椎间盘髓核细胞与退变髓核细胞的生物学性状比较：可以为干预退变髓核细胞提供最佳时机吗？

Biological comparison between in vitro cultured degenerative and normal nucleus pulposus cells: When is the best time to intervene degenerative nucleus pulposus cells?

背景:人体椎间盘是一个承受载荷却又缺乏血管的结构,因而容易发生退行性变,但椎间盘退变的机制尚不明确。目的:通过体外培养人正常椎间盘髓核细胞与退变髓核细胞生物学性状比较,认识退变髓核细胞的细胞学退变时机。方法:分离、培养人正常及退变椎间盘髓核细胞,对两种细胞采用光镜、电镜等形态学方法进行大体形态和超微结构观察,采用生长曲线和XTT实验研究两种细胞的生长动力学差异,测定髓核细胞的活力和细胞Ⅱ型胶原及糖胺多糖的mRNA表达。结果与结论:退变椎间盘细胞至少要比正常椎间盘细胞提前2代出现形态学老化表现。退变髓核细胞表现为G1期阻滞,使细胞不能进入S期,细胞有丝分裂受到抑制。退变髓核细胞总体来说,生长要比正常髓核细胞快,但老化也较快。退变髓核细胞自第1代开始,Ⅱ型胶原和糖胺多糖的mRNA表达比同期正常髓核细胞低得多。说明在体外培养条件下,退变髓核细胞持续增殖能力低,更容易衰老、凋亡。提示退变髓核细胞体外培养衰老较快,进行干预试验逆转椎间盘退变的最佳时机为传2代之前的细胞。

BACKGROUND： Human intervertebral disc can bear load but lack vessels. The mechanism of intervertebral disc degeneration remains unclear. OBJECTIVE： To explore the mechanism of disc degeneration through comparison of different biological properties between normal and degenerative nucleus pulposus cells of intervertebral disc. METHODS： Human normal and degenerative intervertebral disc nucleus pulposus cells were isolated and cultured. Light microscopy, electron microscopy and other morphological methods were used to observe the general morphology and ultrastructure of the cells; growth curve and XTT experiment were used to compare the differences in growth kinetics; the vitality and cell-type Ⅱ collagen and glycosaminoglycan （GAG） mRNA expression was determined. RESULTS AND CONCLUSION： The degenerative nucleus cells displayed aging appearance at least 2 generations earlier than the normal nucleus cells. Degenerative nucleus pulposus cells showed G1 phase arrest, and the cell could not enter S phase; cell mitosis was inhibited. The growth of degenerative nucleus pulposus cells was faster than normal cells but also aged rapidly. Since the first generation, the degenerative nucleus pulposus cells expressed lower Ⅱ collagen and GAG mRNA than the normal nucleus pulposus cells at the same period. Results show that under in vitro culture conditions, the degenerative nucleus pulposus cells had low proliferative capacity and appeared aging and apoptosis. The best time to reverse the disc degeneration for the intervention trial is before the two-generation.