大麻素受体2抑制对口腔黏膜天疱疮棘层松解影响的研究
Cannabinoid receptor 2 inhibition on acantholysis in oral mucosal pemphigus
作者：刘慧娟, 宋鹏, 侯亚丽, 霍晓, 秘利进, 刘春艳
Author：Liu Huijuan, Song Peng, Hou Yali, Huo Xiao, Mi Lijin, Liu Chunyan
收稿日期：2025-04-24 年卷（期）页码：2025,43(6):829-829-836
期刊名称：华西口腔医学杂志
Journal Name：West China Journal of Stomatology
关键字：大麻素受体,自身免疫性疾病,天疱疮,桥粒黏蛋白,HaCaT细胞,
Key words：cannabinoid receptor,autoimmune diseases,pemphigus,desmoglein,HaCaT cell,
基金项目：河北省医学科学研究课题(20241093)
中文摘要
目的 利用大麻素受体2（CB2）抑制剂和天疱疮血清共培养HaCaT细胞，探讨CB2抑制对HaCaT细胞桥粒黏蛋白3（DSG3）的影响。 方法 采用免疫组织化学染色观察天疱疮患者和正常受试者黏膜中CB受体表达，酶联免疫吸附实验（ELISA）检测天疱疮患者和正常受试者血清中CB2含量，对天疱疮患者血清CB2浓度与DSG表达进行相关性分析。CCK-8检测不同浓度AM630对HaCaT细胞的抑制作用，计算AM630对HaCaT细胞的半数抑制浓度（IC50）并作为抑制剂实验浓度。将正常受试者、天疱疮患者血清分别与完全培养液1∶1比例共培养HaCaT细胞作为阴性对照组和天疱疮组，完全培养液培养的HaCaT细胞作为对照组，天疱疮组培养液稀释AM630至IC50浓度后培养的HaCaT细胞作为AM630组。实时荧光定量聚合酶链反应（PCR）、蛋白印迹（Western blot）检测4组CB2、DSG3、β-catenin基因和蛋白的表达。细胞解离实验观察AM630对HaCaT细胞黏附功能的影响。 结果 免疫组织化学染色显示天疱疮黏膜与正常黏膜组织中CB2表达差异有统计学意义（P<0.000 1），CB1表达差异无统计学意义。ELISA检测表明，正常受试者与天疱疮患者血清CB2的表达差异有统计学意义（P<0.001）。天疱疮患者血清CB2表达与DSG3表达呈正相关（r=0.831，P=0.003）。CCK-8检测表明，AM630对HaCaT细胞的半数抑制浓度IC50为0.55 μmol/L。实时荧光定量PCR、Western blot结果显示，天疱疮组CB2、DSG3表达升高，β-catenin表达降低，与AM630组差异有统计学意义（P<0.05）。 结论 口腔黏膜天疱疮中CB2高表达，AM630抑制天疱疮引起的CB2、DSG3高表达及β-catenin低表达，可为天疱疮治疗提供新的治疗靶点和思路。
英文摘要
ObjectiveThe aim of this study is to determine the effect of cannabinoid receptor (CB) 2 inhibitor on desmoglein 3 (DSG3) expression in HaCaT cells co-cultured with pemphigus serum.MethodsImmunohistochemical staining was used to compare CB expression in pemphigus patients and normal individuals. Enzyme-linked immunosorbent assay (ELISA) was employed to quantify the concentration of CB2 in the serum of pemphigus patients and normal individuals. A correlation analysis was performed to examine the relationship between the serum CB2 and DSG of pemphigus patients. The CCK-8 assay was used to evaluate the inhibitory effect of AM630 on HaCaT cells, and the half-maximal inhibitory concentration (IC50) value was utilized to determine the experimental concentration. Serum from normal individuals (negative control group) and pemphigus patients (pemphigus group) was co-cultured with HaCaT cells at a 1∶1 ratio. HaCaT cells cultured in complete medium were used as the control group. HaCaT cells in the pemphigus group treated with AM630 were employed as the AM630 group. Real-time polymerase chain reaction (PCR) and Western blot were conducted to assess the expression levels of CB2, DSG3, and β-catenin. Cell dissociation experiments were conducted to evaluate the effect of AM630 on the adhesion of HaCaT cells.ResultsImmunohistochemistry revealed significant differences in CB2 expression between pemphigus and normal mucosa (P<0 .000 1), but no difference was found in cb1 expression. elisa analysis revealed a statistically significant difference in the expression levels of cb2 in the serum between normal individuals and pemphigus patients (P<0 .001). the expression of cb2 in the serum of pemphigus patients exhibited a significant positive correlation with that of dsg3 (r=0.831,P=0.003). The CCK-8 assay indicated that the IC50of AM630 on HaCaT cells was 0.55 μmol/L. Real-time PCR and Western blot showed that the expression levels of CB2 and DSG3 increased in the pemphigus group, while the expression level of β-catenin decreased compared with that in the AM630 groups (P<0 .05).ConclusionCB2 is highly expressed in oral mucosal pemphigus. AM630 inhibits overexpression of CB2 and DSG3 and underexpression of β-catenin levels, which can provide new therapeutic targets for pemphigus.
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