Upregulation of intercellular adhesion molecule 1 and proinflammatory cytokines by the major surface proteins of Treponema maltophilum and Treponema lecithinolyticum, the phylogenetic group IV oral spirochetes associated with periodontitis and endodontic infections.
Treponema maltophilum and Treponema lecithinolyticum belong to the group IV oral spirochetes and are associated with endodontic infections, as well as periodontitis. Recently, the genes encoding the major surface proteins (Msps) of these bacteria (MspA and MspTL, respectively) were cloned and sequenced. The amino acid sequences of these proteins showed significant similarity. In this study we analyzed the functional role of these homologous proteins in human monocytic THP-1 cells and primary cultured periodontal ligament (PDL) cells using recombinant proteins. The complete genes encoding MspA and MspTL without the signal sequence were cloned into Escherichia coli by using the expression vector pQE-30. Fusion proteins tagged with N-terminal hexahistidine (recombinant MspA [rMspA] and rMspTL) were obtained, and any possible contamination of the recombinant proteins with E. coli endotoxin was removed by using polymyxin B-agarose. Flow cytometry showed that rMspA and rMspTL upregulated the expression of intercellular adhesion molecule 1 (ICAM-1) in both THP-1 and PDL cells. Expression of proinflammatory cytokines, such as interleukin-6 (IL-6) and IL-8, was also induced significantly in both cell types by the Msps, as determined by reverse transcription-PCR and an enzyme-linked immunosorbent assay, whereas IL-1beta synthesis could be detected only in the THP-1 cells. The upregulation of ICAM-1, IL-6, and IL-8 was completely inhibited by pretreating the cells with an NF-kappaB activation inhibitor, l-1-tosylamido-2-phenylethyl chloromethyl ketone. This suggests involvement of NF-kappaB activation. The increased ICAM-1 and IL-8 expression in the THP-1 cells obtained with rMsps was not inhibited in the presence of the IL-1 receptor antagonist (IL-1ra), a natural inhibitor of IL-1. Our results show that the Msps of the group IV oral spirochetes may play an important role in amplifying the local immune response by continuous inflammatory cell recruitment and retention at an infection site by stimulation of expression of ICAM-1 and proinflammatory cytokines.
Lee
Sung-Hoon
SH
Department of Oromaxillofacial Infection and Immunity, College of Dentistry, Seoul National University, 28 Yongon-Dong, Chongno-Gu, Seoul 110-749, Republic of Korea.
Kim
Kack-Kyun
KK
Choi
Bong-Kyu
BK
D002460:Cell Line; D003001:Cloning, Molecular; D016207:Cytokines / Q000096:biosynthesis* / Q000235:genetics; D004708:Endodontics; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000096:biosynthesis* / Q000235:genetics; D053590:Interleukin 1 Receptor Antagonist Protein; D008565:Membrane Proteins / Q000235:genetics / Q000502:physiology*; D016328:NF-kappa B / Q000502:physiology; D010518:Periodontitis / Q000382:microbiology*; D010802:Phylogeny; D012333:RNA, Messenger / Q000032:analysis; D012795:Sialoglycoproteins / Q000494:pharmacology; D014208:Treponema / Q000145:classification / Q000276:immunology*; D015854:Up-Regulation
D016207:Cytokines; C507763:IL1RN protein, human; D053590:Interleukin 1 Receptor Antagonist Protein; D008565:Membrane Proteins; D016328:NF-kappa B; D012333:RNA, Messenger; D012795:Sialoglycoproteins; D018799:Intercellular Adhesion Molecule-1