Yang Hongxu, Zhang Mian, Wang Xin, Zhang Hongyun, Zhang Jing, Jing Lei, Liao Lifan, Wang Meiqing
State Key Laboratory of Military Stomatology, Department of Oral Anatomy and Physiology and TMD, School of Stomatology, the Fourth Military Medical University, 145 Changle West Road, Xi'an, China.
PLoS One. 2015 Nov 3;10(11):e0141774. doi: 10.1371/journal.pone.0141774. eCollection 2015.
To determine if temporomandibular joint chondrocyte apoptosis is induced in rats with dental biomechanical stimulation and what a role TNF takes.
Thirty-two rats were divided into 4 groups (n = 8/group) and exposed to incisor mal-occlusion induced by unilateral anterior crossbite biomechanical stimulation. Two groups were sampled at 2 or 4 weeks. The other two groups were treated with local injections of a TNF inhibitor or PBS into the temporomandibular joints area at 2 weeks and then sampled at 4 weeks. Twenty-four rats either served as unilateral anterior crossbite mock operation controls (n = 8/group) with sampling at 2 or 4 weeks or received a local injection of the TNF inhibitor at 2 weeks with sampling at 4 weeks. Chondrocytes were isolated from the temporomandibular joints of 6 additional rats and treated with TNF in vitro. Joint samples were assessed using Hematoxylin&eosin, Safranin O, TUNEL and immunohistochemistry staining, real-time PCR, fluorogenic activity assays and Western blot analyses. The isolated chondrocytes were also analyzed by flow cytometry.
Unilateral anterior crossbite stimulation led to temporomandibular joint cartilage degradation, associated with an increase in TUNEL-positive chondrocytes number, caspase-9 expression levels, and the release of cytochrome c from mitochondria at 2 weeks without changes in TNF and caspase-8 levels until after 4 weeks. TNF stimulated apoptosis of the isolated chondrocytes and up-regulated caspase-8 expression, but did not change caspase-9 expression levels. Local injection of TNF inhibitor down-regulated caspase-8 expression and reduced TUNEL-positive cell number, but did not reverse cartilage thickness reduction, caspase-9 up-regulation or cytochrome c release.
Unilateral anterior crossbite stimulation induces mitochondrion-mediated apoptosis of articular chondrocytes. TNF accelerated the unilateral anterior crossbite induced chondrocytes apoptosis via death-receptor pathway. However, anti-TNF therapy does not prevent cartilage loss in this model of temporomandibular joint.
确定在接受牙齿生物力学刺激的大鼠中是否会诱导颞下颌关节软骨细胞凋亡，以及肿瘤坏死因子（TNF）发挥何种作用。
将32只大鼠分为4组（每组n = 8），通过单侧前牙反合生物力学刺激诱导切牙错合。两组在2周或4周时取样。另外两组在2周时在颞下颌关节区域局部注射TNF抑制剂或磷酸盐缓冲液（PBS），然后在4周时取样。24只大鼠要么作为单侧前牙反合假手术对照组（每组n = 8）在2周或4周时取样，要么在2周时接受TNF抑制剂局部注射并在4周时取样。从另外6只大鼠的颞下颌关节分离软骨细胞并在体外进行TNF处理。使用苏木精-伊红染色、番红O染色、TUNEL染色和免疫组织化学染色、实时聚合酶链反应（PCR）、荧光活性测定和蛋白质免疫印迹分析对关节样本进行评估。分离的软骨细胞也通过流式细胞术进行分析。
单侧前牙反合刺激导致颞下颌关节软骨降解，在2周时TUNEL阳性软骨细胞数量增加、半胱天冬酶-9表达水平升高以及细胞色素c从线粒体释放，而TNF和半胱天冬酶-8水平直到4周后才发生变化。TNF刺激分离的软骨细胞凋亡并上调半胱天冬酶-8表达，但未改变半胱天冬酶-9表达水平。局部注射TNF抑制剂下调半胱天冬酶-8表达并减少TUNEL阳性细胞数量，但未逆转软骨厚度减少、半胱天冬酶-9上调或细胞色素c释放。
单侧前牙反合刺激诱导关节软骨细胞发生线粒体介导的凋亡。TNF通过死亡受体途径加速单侧前牙反合诱导的软骨细胞凋亡。然而，在这种颞下颌关节模型中，抗TNF治疗并不能防止软骨丢失。
