【作者】 傅文青； 陈哲； 刘楠； 张国兴；

【机构】 苏州大学医学部医学心理研究所；

【摘要】 目的:1、建立慢性温和非预期性应激(chronic unpredictable mild stress,CUMS)模型,观察慢性温和非预期性应激对C3H/He J鼠皮肤组织的影响。2、研究小鼠皮肤组织中免疫相关细胞因子TNF-α、IL-1β、IL6和细胞凋亡相关蛋白Bax、Bcl-2在心理应激过程中的变化。3、探讨心理应激诱导皮肤组织免疫紊乱和细胞凋亡的机制,为治疗斑秃疾病提供新的思路和实验依据。方法:1、采用C3H/He J雌鼠建立CUMS模型,与对照组(Control组)比较,观察心理应激组小鼠体重和行为学变化以及心理应激对小鼠皮肤组织的影响。2、在上述实验基础上加入腹腔注射抗氧化剂组,把所有实验小鼠随机分成成Control组、CUMS组和CUMS+抗氧化剂组(Tempol组)三组。在施加21天心理应激后,对三组小鼠进行体重测量和行为学评估,检测小鼠血浆皮质酮水平,观察小鼠皮肤组织学切片,用免疫组织化学染色的方法观察皮肤组织中免疫相关细胞因子TNF-α、IL-1β、IL6的表达并用Elisa试剂盒进行定量检测,再用Western Blot方法检测皮肤组织中凋亡相关蛋白Bax和Bcl-2的表达。结果:1、小鼠体重变化:与Control组相比,CUMS组和Tempol组的小鼠体重显著降低(P<0.01);Tempol组的小鼠体重相对于CUMS组也有明显降低(P<0.05)。2、行为学评估:在3分钟旷场实验中,CUMS组与Tempol组小鼠的总运动距离和修饰行为次数明显少于Control组小鼠(P<0.01),静止时间明显多于Control组小鼠(P<0.01)。3、血浆皮质酮:CUMS组与Tempol组小鼠的血浆皮质酮含量显著高于Control组小鼠(P<0.01)。4、CUMS组小鼠皮肤组织中出现了大量炎性细胞聚集现象,Tempol组和Control组小鼠只有少量聚集或者没有出现。5、免疫相关细胞因子:CUMS组小鼠皮肤组织中免疫相关细胞因子TNF-α、IL1β、IL6的含量显著高于Control组小鼠(P<0.05)与Tempol组小鼠(P<0.05)。6、细胞凋亡相关蛋白:CUMS组小鼠皮肤组织中促凋亡蛋白Bax的表达量显著高于Control组小鼠(P<0.05)与Tempol组小鼠(P<0.05),三组小鼠皮肤组织中抑凋亡蛋白Bcl-2的表达量无显著差异。结论:1、慢性温和非预期性应激模型是研究疾病发生机制中心理因素作用的最适宜模型。2、慢性温和非预期性应激诱导了C3H/He J鼠皮肤组织的免疫紊乱,造成TNF-α、IL-1β、IL6等免疫细胞因子的水平升高。3、慢性温和非预期性应激促进了C3H/He J鼠皮肤组织细胞的凋亡。4、TNF-α等免疫细胞因子与促细胞凋亡蛋白Bax表达具有正相关性。5、抗氧化剂Tempol可以逆转慢性温和非预期性应激对小鼠皮肤组织造成的影响。更多还原

【Abstract】 Objective:1. Establish chronic unpredictable mild stress(CUMS) model to observe the influence of mental stress on the skin tissue of mice. 2. Investigate the changes of immune related cytokines of TNF- α, IL-1 β, IL6 and apoptosis related proteins Bax, Bcl-2 in the skin tissue of mice by chronic mild unpredictable stress. 3. Investigate the mechanism of mental stress induced skin immunologic derangement and apoptosis, to provide new ideas and experimental evidence for the treatment of alopecia areata. Methods:1. Establish CUMS model with C3H/HeJ female mice, compared with the control group and then, observe the changes of weight, behavior and skin tissue of mice. 2. Added one group of intraperitoneal injection antioxidant,all animals were assigned to the following three groups randomly: Control group, CUMS group and CUMS+ antioxidant group(Tempol group). After 21 days’ mental stress, observe the behavior and weight change, detect the levels of plasma corticosterone in mice, observe the skin tissue morphology, use immunohistochemical method to observe the expressions of the immune related cytokines TNF-α, IL-1β and IL6 in skin tissue and quantitative determination by Elisa Kit, observe the expressions of apoptosis related protein Bax and Bcl-2 in skin tissue with Western Blot method. Results: 1. The weight change of mice: Compared with the Control group, the weight of mice in CUMS group and Tempol group reduced significantly(P<0.01); compared with the CUMS group the weight of mice in Tempol group also reduced significantly(P<0.05). 2. Behavioral assessment: Compared with Control group, the moving distances and the numbers of grooming behavior in 3min of Open-field test in CUMS group and Tempol group were significantly decreased(P<0.01), resting time increased significantly(P<0.01). 3. Plasma corticosterone: Compared with Control group, the plasma corticosterone levels of CUMS group and Tempol group were significantly increased(P<0.01). 4. A lot of inflammatory cells aggregated in the skin tissue of CUMS group mice, just little in the skin tissue of Control group and Tempol group. 5. Immune related cytokines: Compared with Control group and Tempol group, the TNF- α, IL-1β and IL6 in skin tissue of CUMS group were increased significantly(P<0.05). 6. Apoptosis related proteins: Compared with Control group and Tempol group, the expression of Bax which is a protein that promoted apoptosis in skin tissue of CUMS group were increased significantly(P<0.05), but the expression of Bcl-2 which is a protein that inhibited apoptosis in skin tissue was not much difference in the three groups of mice. Conclusion: 1. Chronic mild unpredictable stress model is an ideal model to research the role of mental factors in the pathogenesis of the disease. 2. Chronic mild unpredictable stress induced the immunologic derangement in the skin tissue of C3H/HeJ mice, it made the levels of TNF- α, IL-1β and IL6 in skin tissue increased. 3. Chronic mild unpredictable stress promoted the apoptosis in skin tissue. 4. The expression of TNF- α and apoptotic protein Bax has positive correlation. 5. The antioxidant Tempol could reverse the effect of chronic mild unpredictable stress caused to the skin tissue of mice.更多还原