Matrix regulation of skeletal cell apoptosis - Role of calcium and phosphate ions
Matrix regulation of skeletal cell apoptosis - Role of calcium and phosphate ions
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DOI：
10.1074/jbc.m006492200
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发表时间：
2001-06-08
影响因子：
4.8
通讯作者：
Shapiro, IM
中科院分区：
文献类型：
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作者：
Adams, CS;Mansfield, K;Shapiro, IM
先前，我们注意到无机磷酸盐（Pi），骨细胞外基质的主要组分，诱导成骨细胞凋亡（Meleti，Z.，夏皮罗岛M.，和亚当斯，C. S.（2000）Bolts（NY）27，359-366）。由于Ca ~（2+）沿着P-i在骨吸收过程中从骨中释放，我们提出了Ca ~（2+）调节P-i介导的成骨细胞凋亡的假设。为了验证这一假设，将成骨细胞与两种离子一起孵育，并测定细胞死亡。我们注意到，0.1-1 mM的培养基Ca 2+浓度（[Ca 2 +]（e））的适度增加引起培养成骨细胞的P-1依赖性死亡的显著和快速增强。单独升高[Ca 2 +]（e）对成骨细胞活力没有影响，而Ca 2+通道阻滞剂未能抑制离子对处理细胞的杀伤。这些结果表明，P-i介导的细胞死亡不依赖于胞质Ca 2+浓度的持续增加。末端dUTP缺口末端标记分析和测量的caspase-3活性的离子对处理的细胞表明，死亡是凋亡;凋亡证实使用caspase-3和核酸内切酶抑制剂。测定细胞线粒体膜电位和胞浆Ca ~（2+）浓度。在与[Ca 2 +]（e）和Pi孵育后，注意到线粒体荧光的减少，表明离子降低线粒体跨膜电位。继线粒体膜电位的下降，有一个短暂的升高，在胞浆Ca 2+浓度。研究结果表明，离子对在质膜水平上合谋诱导细胞内变化，导致线粒体功能丧失。随后的细胞内Ca 2+浓度的增加可能会引发下游事件，导致成骨细胞凋亡。
Previously, we noted that inorganic phosphate (P-i), a major component of bone extracellular matrix, induced osteoblast apoptosis (Meleti, Z., Shapiro, I. M., and Adams, C. S. (2000) Bolts (NY) 27, 359-366). Since Ca2+ along with P-i is released from bone during the resorption process, we advanced the hypothesis that Ca2+ modulates P-i-mediated osteoblast apoptosis. To test this hypothesis, osteoblasts were incubated with both ions, and cell death was determined. We noted that a modest increase in the medium Ca2+ concentrations ([Ca2+](e)) of 0.1-1 mM caused a profound and rapid enhancement in P-i-dependent death of cultured osteoblasts. An elevation in [Ca2+](e) alone had no effect on osteoblast viability, whereas Ca2+ channel blockers failed to inhibit killing of ion pair-treated cells. These results indicated that P-i-mediated cell death is not dependent on a sustained increase in the cytosolic Ca2+ concentration. Terminal dUTP nick-end labeling analysis and measurement of caspase-3 activity of the ion pair-treated cells suggested that death was apoptotic; Apoptosis was confirmed using caspase-3 and endonuclease inhibitors. The mitochondrial membrane potential and cytosolic Ca2+ status of the treated cells were evaluated. After incubation with [Ca2+](e) and P-i, a decrease in mitochondrial fluorescence was noted, suggesting that the ions decreased the mitochondrial transmembrane potential. Subsequent to the fall in mitochondrial membrane potential, there was a transient elevation in the cytosolic Ca2+ concentration. Results of the study suggest that the ion pair conspire at the level of the plasma membrane to induce intracellular changes that result in loss of mitochondrial function. The subsequent increase in the cytosolic Ca2+ concentration may trigger downstream events that transduce osteoblast apoptosis.