Impaired function of epithelial plakophilin-2 is associated with periodontal disease

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.

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.

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-mediated 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. Conclusions: 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.