Inhibition of tissue-reparing signaling by P.gingivalis infection

牙龈假单胞菌是一种牙周病的革兰氏阴性杆菌，可侵袭多种宿主细胞，如上皮细胞和成纤维细胞。感染牙周组织的慢性炎症会导致组织破坏。在这项研究中，我们研究了该病原菌的两个主要毒力因子：菌毛和一种名为牙龈痛的蛋白酶对感染宿主组织的破坏和抑制再生的作用。首先，我们构建了牙龈假单胞菌菌毛缺失和牙周疼痛缺失突变体。利用这两个突变体，我们发现：1)菌毛突变体和牙周疼痛缺失突变体均表现出对人牙周膜细胞(PDLS)侵袭能力的降低；2)野生型Pggvalis在无血清培养基中的感染导致PDLS从培养皿中脱落。3)感染野生型牙龈假单胞菌后，整合素相关信号分子如FAK和PXLIN的降解。4)通过划痕实验，我们发现牙龈假单胞菌的感染强烈地抑制了受损的牙周膜细胞单层的再生。细菌感染抑制了细胞的运动和生长。5)姜辣素缺失突变体对感染细胞单层的再生没有表现出抑制作用。另一方面，菌毛缺失突变体确实抑制了牙周组织的再生过程，这些结果有力地表明牙龈痛在抑制感染牙周组织的再生过程中起着重要作用。

P.gingivalis, a periodontopathic Gram-negative bacteria, is known to invade into a variety of host cells such as epithelial cells and fibroblasts. A chronic inflammation of infected periodontal tissue results in tissue destruction. In this study, we investigated the involvement of the two major virulent factors of this pathogen ; fimbriae and a protease called gingipain on the destruction and inhibition of regeneration of infected host tissues.At first, we constructed fimbriae-deficient and gingipain-deficient mutants of P.gingivalis. Using these two mutants, we found that :1)Both fimbriae- and gingipain-deficient mutants of P.gingivalis showed decreased invasion in human periodontal ligament cells (PDLs).2)Infection of wild type of P.gingivalis in serum-free medium resulted in a detachment of PDLs from the culture dishes. However, gingipain-deficient mutant did not induce such detachment of PDLs.3)Infection of wild type of P.gingivalis induced a degradation of integrin-associated signaling molecules such as FAK and paxillin. However, the degradation of FAK and paxillin was not observed during infection with gingipain-deficient mutant.4)By using a scratch assay, we found that the infection of P.gingivalis strongly inhibited the regeneration of damaged monolayers of PDLs. The bacterial infection inhibited the cellular movement as well as the cellular growth.5)Gingipain-deficient mutant did not show the inhibitory effect on the regeneration of the infected cellular monolayers. On the other hand, fimbriae-deficient mutant did inhibit the regeneration process.These results strongly suggested that gingipain of P.gingivalis plays an important role in the inhibition of regeneration of the infected periodontal tissues.