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中华口腔医学研究杂志(电子版) ›› 2019, Vol. 13 ›› Issue (02) : 84 -89. doi: 10.3877/cma.j.issn.1674-1366.2019.02.004

所属专题： 文献；

基础研究

心理应激对大鼠髁突软骨中半胱氨酸天冬氨酸蛋白酶9表达的影响
董晶1, 宋文婷2, 郑纪伟1, 秦莹1, 孙晋虎1,†()   
  1. 1. 徐州医科大学口腔医学院　221004；徐州医科大学附属医院口腔科　221004
    2. 徐州医科大学口腔医学院　221004
  • 收稿日期:2018-08-31 出版日期:2019-04-01
  • 通信作者: 孙晋虎

Effect of psychological stress on the expression of caspase-9 in temporomandibular joint condyle cartilage in rats

Jing Dong1, Wenting Song2, Jiwei Zheng1, Ying Qin1, Jinhu Sun1,†()   

  1. 1. School of Stomatology, Xuzhou Medical University, Xuzhou 221004, China; Department of Stomatology, the Affiliated Hospital of Xuzhou Medical University, Xuzhou 221004, China
    2. School of Stomatology, Xuzhou Medical University, Xuzhou 221004, China
  • Received:2018-08-31 Published:2019-04-01
  • Corresponding author: Jinhu Sun
  • About author:
    Corresponding author: Sun Jinhu, Email:
  • Supported by:
    Key University Science Research Project of Jiangsu Province(12KJB320015); Xuzhou Science and Technology Project(KC14SH114, KC15SH015, KC16SL125)
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董晶, 宋文婷, 郑纪伟, 秦莹, 孙晋虎. 心理应激对大鼠髁突软骨中半胱氨酸天冬氨酸蛋白酶9表达的影响[J]. 中华口腔医学研究杂志(电子版), 2019, 13(02): 84-89.

Jing Dong, Wenting Song, Jiwei Zheng, Ying Qin, Jinhu Sun. Effect of psychological stress on the expression of caspase-9 in temporomandibular joint condyle cartilage in rats[J]. Chinese Journal of Stomatological Research(Electronic Edition), 2019, 13(02): 84-89.

目的

探讨心理应激对大鼠髁突软骨中半胱氨酸天冬氨酸蛋白酶9（caspase-9）表达的影响。

方法

将48只雄性SD大鼠用随机数表法随机分为6组：心理应激组（1、5和9周）及空白对照组（1、5和9周），每组各8只。应激组大鼠予以慢性不可预见性应激相应周数，应激完成后经旷场实验及糖水偏嗜度实验观察大鼠抑郁情况，数据通过重复测量的方差分析进行处理，随后处死相应时间点对照组及应激组大鼠，采用免疫组织化学染色及蛋白免疫印迹法了解髁突软骨细胞中caspase-9的表达情况，两对应时间点的数据通过两独立样本t检验进行分析。

结果

实验中，心理应激组大鼠应激1周后即已开始出现总移动距离减少（t = 8.676，P = 0.011）、中间格停留时间缩短（t = 5.091，P = 0.041）、糖水偏嗜度降低（t = 10.150，P<0.001）等抑郁样改变，且大鼠髁突表面不整、出现了细胞胶原纤维松解的病理表现；应激5周时大鼠髁突胶原破坏持续存在，应激9周时大鼠髁突内软骨细胞减少，出现了无细胞的空白区域；且各时间点应激组髁突中caspase-9的表达均高于相应时间点的空白对照组，差异具有统计学意义（P<0.001）。

结论

心理应激可导致大鼠出现活动减少、糖水偏嗜度降低等抑郁样改变，且大鼠髁突软骨中caspase-9的表达明显升高，这可能是应激导致颞下颌关节损伤的机制之一。

关键词: 应激，心理学, 颞下颌关节, 半胱氨酸天冬氨酸蛋白酶9, 免疫组织化学
Objective

To investigate the alteration of the expression of cysteine-dependent aspartate-specific proteases-9 (caspase-9) in the condylar cartilage of tempromadibualr joint (TMJ) of rats.

Methods

The 48 male SD rats were randomly divided into 6 groups with random table method: blank control (CON) group (1, 5, and 9 weeks) and psychological stress (PS) group (1, 5, and 9 weeks) , the rats were treated by chronic unpredicted mild stress (CUMS) except those in the control group. After corresponding weeks of CUMS treatment, the open filed test and sucrose preference were performed to assess the depression state, the data were analyzed with repeated measures analysis of variance, then the rats were killed and the immunohistochemical staining as well as western blot were applied to detect the expression of caspase-9 in condylar. The data between CON group and PS group of the same time were processed by two independent sample t test.

Results

Compared with CON group, PS group had showed less distance movement (t= 8.676, P = 0.011) and less time spent in center (t = 5.091, P = 0.041) in the open field test and short of sucrose preference (t = 10.150, P<0.001) starting from the first week. Besides, edema and releases of collagen fiber in the condylar surface were observed in PS 1-week group. In PS 5-week group, the results also shown collagen fiber loosen, fracture of cracks in the condyle. In PS 9-week group, the cells in condylar decreased significantly, appearing the "blank" area without cells. The variation of expression on Caspase-9 in PS group, which were significantly higher than that in CON group at the same time (P<0.001) .

Conclusions

The CUMS can lead to SD rats changing behaviors and taking less surcose, and high expression of caspase-9 were shown in condylar cartilage of the stressed rats, which may be one of the mechanisams of TMJ damage induced by CUMS.

Key words: Stress, psychological, Temporomandibular joint, Caspase 9, Immunohistochemistry
图1 空白组及应激组不同时间点的大鼠旷场运动路径图
表1 空白组及应激组相应周数的大鼠旷场实验结果（±s，n = 8）
组别 移动总距离（cm） 中间格停留时间（s）
1周 5周 9周 1周 5周 9周
CON组 1 096.1 ± 551.1 1 138.5 ± 265.0 1 166.5 ± 496.8 3.9 ± 3.3 3.6 ± 2.2 3.4 ± 2.5
PS组 413.5 ± 354.8 492.7 ± 246.1 361.1 ± 278.7 1.1 ± 1.2 1.0 ± 1.5 1.1 ± 1.4
t值 8.676 25.457 16.146 5.091 8.580 5.115
P值 0.011 <0.001 <0.001 0.041 0.011 0.040
表2 空白组及应激组相应周数大鼠糖水偏嗜度的变化情况（%，±s，n = 8）
组别 1周 5周 9周
CON组 68.6 ± 3.5 68.4 ± 2.9 67.7 ± 2.8
PS组 55.0 ± 2.5 54.0 ± 2.8 50.4 ± 3.6
t值 9.116 10.150 10.568
P值 <0.001 <0.001 <0.001
图2 空白组及应激组相应周数的大鼠髁突免疫组织化学染色图像　黑色箭头为胶原纤维松解留下的裂隙，白色箭头为髁突细胞减少后留下的空白区域
图3 空白组及应激组相应周数的大鼠髁突中半胱氨酸天冬氨酸蛋白酶9（caspase-9）的表达情况　A：大鼠髁突中caspase-9的免疫组织化学染色情况，箭头所示为棕色或棕黄色的caspase-9阳性细胞；B：大鼠髁突中caspase-9阳性细胞百分率情况（aP<0.001）
图4 空白组及应激组相应周数的大鼠髁突中半胱氨酸天冬氨酸蛋白酶9（caspase-9）的表达情况　A：大鼠髁突中caspase-9免疫印迹表达，1、3、5：CON组1、5和9周；2、4、6：PS组1、5和9周；B：大鼠髁突中caspase-9相对蛋白量表达情况，aP<0.01
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