Mechanical stress-induced apoptosis of endplate chondrocytes in organ-cultured mouse intervertebral discs -: An ex vivo study
Mechanical stress-induced apoptosis of endplate chondrocytes in organ-cultured mouse intervertebral discs -: An ex vivo study
复制标题
DOI：
10.1097/00007632-200307150-00010
复制
发表时间：
2003-07-15
期刊：
影响因子：
3
通讯作者：
Yoshikawa, H
中科院分区：
文献类型：
--
作者：
Ariga, K;Yonenobu, K;Yoshikawa, H
研究设计.在器官培养物中对小鼠椎间盘施加不同量的静态机械负荷。缺口末端标记法检测细胞凋亡。两种丝裂原活化蛋白激酶（MAPK）抑制剂加入到培养基中。目的建立机械应力诱导软骨细胞凋亡的实验模型，探讨MAPK和p38在应力诱导终板软骨细胞凋亡中的作用。软骨终板（CEP）退行性变化的原因尚不清楚。作者先前使用小鼠模型的研究结果表明，软骨终板中的细胞凋亡可能在椎间盘退变中起作用，机械应力可能诱导细胞凋亡。如果终板软骨细胞的凋亡参与了椎间盘退变的级联反应，那么机械应力如何诱导细胞凋亡在预防椎间盘退变中应该是重要的。然而，机械应力诱导细胞凋亡的机制尚不清楚。收获小鼠尾椎盘并进行器官培养。将各种静态压缩载荷（0、0.2、0.4、0.8和1.0MPa）施加在置于培养瓶中的椎间盘上24小时。石蜡包埋切片收获的光盘进行染色，使用番红-O和缺口末端标记程序。在每个椎间盘的软骨终板和连接纤维环中计数凋亡细胞。另外。在培养液中加入MAPK抑制剂U 0126和p38抑制剂SB 202190，观察它们对机械负荷诱导终板软骨细胞凋亡的调节作用。组织学上，加载的椎间盘隆起，椎间隙变窄。细胞凋亡是不存在的光盘没有负载，但在加载光盘（负载重量，1.0 MPa）是特别明显的。凋亡细胞的数量增加取决于负载的重量。两种MAPK抑制剂均显着增加凋亡细胞数量。结论。在器官培养中，静态机械负荷可诱导软骨细胞凋亡，尤其是软骨终板。细胞凋亡发生类似于以前的研究结果，使用体内模型。因此，该培养系统反映了体内证实的细胞凋亡。由于生物活性试剂，如MAPK抑制剂，可以简单地添加到培养基中，该系统可能是一个有用的方法，用于检测因素，影响机械应力诱导的细胞凋亡。两种MAPK抑制剂均增加细胞凋亡的发生。这表明这两种MAPK可以对抗机械应力诱导的凋亡途径。
Study Design. Various amounts of static mechanical load were applied to mouse intervertebral discs in organ cultures. The apoptosis then was examined using nick end labeling. Two mitogen-activated protein kinase (MAPK) inhibitors were added to the medium.Objectives. To establish an experimental model for detecting factors regulating chondrocyte apoptosis induced by mechanical stress, and to determine the role of MAPK and p38 in the stress induced apoptotic pathway of endplate chondrocytes.Summary of Background Data. The cause of degenerative change in the cartilaginous endplate (CEP) remains unclear. The authors' previous findings using a mouse model suggested that apoptosis in the cartilaginous endplate may play a role in intervertebral disc degeneration, and the mechanical stress may induce apoptosis. If apoptosis of endplate chondrocytes is involved in the cascade of intervertebral disc degeneration, then how apoptosis is induced by mechanical stress should be important in preventing disc degeneration. However, the mechanism of apoptosis induced by mechanical stress remains unclear.Methods. Mouse coccygeal discs were harvested and organ cultured. Various static compression loads (0, 0.2, 0.4, 0.8, and 1.0 MPa) were applied on intervertebral discs placed in culture bottles were 24 hours. Paraffin-embedded sections of the harvested discs were stained using Safranin-O and the nick end labeling procedure. The apoptotic cells were counted in the cartilaginous endplate and junctional anulus fibrosus of each intervertebral disc. In addition. U0126 (MAPK inhibitor) and SB202190 (p38 inhibitor) were added to the culture medium to determine their regulatory roles in the apoptosis of endplate chondrocytes induced by mechanical load.Results. Histologically, loaded discs became bulged, and the disc space became narrow. Apoptosis was absent in discs without load, but was particularly noticeable in loaded discs (load weight, 1.0 MPa). The number of apoptotic cells increased depending on the weight of the load. The two MAPK inhibitors significantly increased the number of apoptotic cells.Conclusions. Chondrocyte apoptosis was induced using static mechanical load especially in the cartilaginous endplate in an organ culture. Apoptosis occurred similarly to previous findings using an in vivo model. This culture system thus reflected the apoptosis demonstrated in vivo. Because biologically active reagents such as MAPK inhibitors can be simply added to culture media, this system may be a useful method for detecting factors that influence apoptosis induced by mechanical stress. Both MAPK inhibitors increased the occurrence of apoptosis. This suggests that these two MAPKs can counteract the apoptotic pathway induced by mechanical stress.