Gruber Helen E, Hoelscher Gretchen L, Bethea Synthia, Hanley Edward N
From the Department of Orthopaedic Surgery, Carolinas HealthCare System, Charlotte, NC.
Spine (Phila Pa 1976). 2015 Jun 15;40(12):876-82. doi: 10.1097/BRS.0000000000000936.
A study using cultured human annulus cells and human annular tissue.
To further explore and define mitochondrial mechanisms related to disc cell apoptosis in vitro and in vivo.
Mitochondrial-dependent intrinsic signaling pathways are a well-recognized component of apoptosis (programmed cell death). Disc cell apoptosis is important because it is a major mechanism by which cell numbers decrease during disc degeneration. Our objective was to further explore and define mitochondrial mechanisms related to disc cell apoptosis.
High-content screening techniques were used to study nuclear morphology and mitochondrial membrane potentials in cultured annulus cells. Gene expression in annulus tissue was studied with microarray analysis.
Cultured cells showed significantly increased nuclear size (an indicator of apoptosis) with increasing Thompson grade (P < 0.00001 by analysis of variance). A significant negative correlation for mitochondrial potential (which results from the difference in electrical potential generated by the electrochemical gradient across the inner membrane of the mitochondrion) versus Thompson grade was identified in cultured human annulus cells in control conditions (r = 0.356, P < 0.0001). When exposed to the K ionophore valinomycin at sublethal levels to induce apoptosis, a significant reduction in mitochondrial potential was identified versus nontreated cells. Gene expression patterns in more degenerated Thompson grade III, IV, and V discs versus healthier grade I and II discs showed significant upregulation of a number of genes with well-recognized apoptosis roles in mitochondrial potential decline (ITM2B, beta-2-microglobulin, and cathepsin B, DAP, GAS1, and PDCD5) and TNF-α associations (cathepsin B, RAC1, and PPT1).
Data presented here show the in vivo expression of apoptosis-related genes associated with the loss of mitochondrial membrane integrity and decreased mitochondrial membrane potential with increasing Thompson scores. These data, which mimic our novel, direct cell-based in vitro findings, stress the importance of mitochondrial changes related to apoptosis and TNF-α during human disc degeneration.
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一项使用培养的人椎间盘纤维环细胞和人纤维环组织的研究。
进一步探索和确定体内外与椎间盘细胞凋亡相关的线粒体机制。
线粒体依赖性内在信号通路是细胞凋亡（程序性细胞死亡）中一个公认的组成部分。椎间盘细胞凋亡很重要，因为它是椎间盘退变过程中细胞数量减少的主要机制。我们的目的是进一步探索和确定与椎间盘细胞凋亡相关的线粒体机制。
采用高内涵筛选技术研究培养的纤维环细胞的核形态和线粒体膜电位。用微阵列分析研究纤维环组织中的基因表达。
随着汤普森分级增加，培养的细胞显示核大小显著增加（细胞凋亡的一个指标）（方差分析，P < 0.00001）。在对照条件下培养的人纤维环细胞中，线粒体电位（由线粒体内膜跨膜电化学梯度产生的电位差）与汤普森分级呈显著负相关（r = 0.356，P < 0.0001）。当用亚致死水平的钾离子载体缬氨霉素处理以诱导细胞凋亡时，与未处理细胞相比，线粒体电位显著降低。与较健康的I级和II级椎间盘相比，退变程度更高的汤普森III级、IV级和V级椎间盘中的基因表达模式显示，许多在线粒体电位下降中具有公认凋亡作用的基因（ITM2B、β-2-微球蛋白、组织蛋白酶B、DAP、GAS1和PDCD5）以及与肿瘤坏死因子-α相关的基因（组织蛋白酶B、RAC1和PPT1）显著上调。
此处呈现的数据表明随着汤普森评分增加，与线粒体膜完整性丧失和线粒体膜电位降低相关的凋亡相关基因在体内表达。这些数据与我们基于细胞的新型直接体外研究结果相似，强调了线粒体变化在人类椎间盘退变过程中与凋亡和肿瘤坏死因子-α相关的重要性。
无。
