Impaired function of epithelial plakophilin-2 is associated with periodontal disease.
Impaired function of epithelial plakophilin-2 is associated with periodontal disease.
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DOI：
10.1111/jre.12918
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发表时间：
2021-12
影响因子：
3.5
通讯作者：
Zhang S
中科院分区：
文献类型：
--
作者：
Yu N;Zhang J;Phillips ST;Offenbacher S;Zhang S
Plakophilin-2 （PKP2）是一种细胞内桥粒锚定蛋白，在一项全基因组关联研究中，PKP2的遗传变异与牙龈卟啉单胞菌(P.gingivalis)-显性牙周生态失调有关。在本研究中，我们比较了牙周炎牙龈活检患者和无牙周炎患者的离体PKP2表达，并评估了PKP2在牙龈上皮屏障功能中的体外作用以及牙龈假单胞菌调节PKP2表达的机制。利用逆转录定量实时PCR （RT-qPCR）技术，我们检测了11名牙周病健康患者、10名实验性牙周炎患者和10名慢性牙周炎患者的牙龈活检组织中PKP2 mRNA的表达水平。免疫组织化学检测牙龈组织PKP2蛋白的表达。然后，我们用包括牙龈假单胞菌、直弯曲杆菌和各种toll样受体激动剂在内的细菌攻击原代牙龈上皮细胞。Western blot和免疫荧光染色检测蛋白表达。阻断蛋白酶途径的抑制剂被用于检测牙龈假单胞菌介导的PKP2蛋白降解。我们还使用慢病毒短发夹RNA （shRNA）敲除内源性上皮细胞PKP2，并评估细胞增殖、扩散和屏障功能。牙周炎患者的PKP2 mRNA含量比健康对照组低约2倍（p < 0.05）。PKP2蛋白主要在牙龈上皮中表达。在原代牙龈上皮细胞中，牙龈假单胞菌侵染后PKP2 mRNA水平升高，蛋白表达降低，提示牙龈假单胞菌具有蛋白降解机制。半胱氨酸蛋白酶抑制剂极大地减弱了牙龈卟啉菌介导的PKP2蛋白降解。缺乏PKP2的上皮细胞表现出细胞增殖和扩散受到抑制，不能形成单层。最后，牙龈假单胞菌损害了牙龈上皮屏障功能。PKP2似乎对维持牙龈上皮屏障功能至关重要，并且容易被牙龈假单胞菌产生的半胱氨酸蛋白酶降解。我们的研究结果已经确定了牙龈假单胞菌通过促进PKP2降解来损害上皮屏障功能的机制。
Plakophilin-2 (PKP2) is an intracellular desmosomal anchoring protein that has been implicated in a genome-wide association study, in which genetic variants of PKP2 are associated with Porphyromonas gingivalis (P.gingivalis)-dominant periodontal dysbiosis. In this study, we compared the ex vivo PKP2 expression in periodontitis gingival biopsies to periodontitis-free subjects and assessed the in vitro role of PKP2 in gingival epithelial barrier function and the mechanism by which P.gingivalis modulates PKP2 expression. Using reverse transcription quantitative real-time PCR (RT-qPCR), we determined PKP2 mRNA expression levels in gingival biopsies collected from 11 periodontally healthy, 10 experimental gingivitis, and 10 chronic periodontitis subjects. PKP2 protein expression in gingival biopsies was detected by immunohistochemistry. We then challenged primary gingival epithelial cells with bacteria including P.gingivalis, Campylobacter rectus, and various Toll-like receptor agonists. Western blot and immunofluorescence staining were used to detect protein expression. Inhibitors blocking proteases pathways were tested for P.gingivalis-mediated PKP2 protein degradations. We also knocked down endogenous epithelial PKP2 using lentiviral short-hairpin RNA (shRNA) and evaluated cell proliferation, spreading, and barrier function. Periodontitis gingival biopsies had approximately twofold less PKP2 mRNA than did healthy controls (p < .05). PKP2 protein was predominantly expressed in gingival epithelium. In primary gingival epithelial cells, P.gingivalis challenge increased PKP2 mRNA levels, while protein expression decreased, which suggests that P.gingivalis has a protein degradation mechanism. Cysteine proteases inhibitors greatly attenuated P.gingivalis-m ediated PKP2 protein degradation. Epithelial cells with deficient PKP2 exhibited inhibited cell proliferation and spreading and failed to form monolayers. Finally, P.gingivalis impaired gingival epithelial barrier function. PKP2 appears to be critical in maintaining gingival epithelial barrier function and is susceptible to degradation by cysteine proteases produced by P.gingivalis. Our findings have identified a mechanism by which P.gingivalis impairs epithelial barrier function by promoting PKP2 degradation.
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影响因子：
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