Studies Of LPS On Cytoxicity And Production Of Inflammatory Cytokines In Human PDLC

Periodontal disease is characterized by the loss of periodontal ligament and alveolar bone in periodontal tissue. Therefore, it is very important to study the role of the periodontal ligament in the pathogenesis of periodontal disease. The periodontal ligament consists mainly of periodontal ligament cell (PDLC) and collagen, and PDLC exhibit high ALPase activity. On the other hand, the gram-negative anaerobic bacteria Pg.. Pi and Fn have been believed to be important periodontopathic bacteria. Furthem~iore, many investigations have indicated that the LPS may lead to destruction of the periodontal tissue by a direct action or by inducing inflammatory reactions indirectly. Previously, it has been reported that Pg.. Pi.. En LPS stimulate the induction of IL-i a~ IL-i f3.. IL-6.. IL-8 and TNF- ci by human gingival fibroblasts. However, the significance of Pg. Pi.. Fn LPS to periodontal ligament cell remains unknown. In this study, the effects of Pg.. Pi.. Fn LPS on ALPase activity and production of inflammatory cytokines of IL-6 and TNF- a stimulated by Pg.. Pi.. Fn LPS in human PDLC were examined. 1. Effects of LPS on the proliferation of human PDLC To evaluate the effects of Pg.. Pi.. En LPS on the proliferation of PDLC, we used MTT colormetic method to determine the change of PDLC amount. The results showed that the growth inhibitory effects were found only when considerably higher concentrations of LPS were given. In contrast, treatment with low concentration of LPS exhibited the enchancement of proliferation. Therefore, the growth inhibitory effects of LPS are mainly relevent to the concentrations. 2. Effects of LPS on ALPase activity of human PDLC 4 To observe the effects of Pg Pi En LPS on ALPase activity of PDLC, we employed cell culture technique and enzyme kinetics methods. The study indicated that all LPS diminished the ALPase activity of PDLC at all concentrations examined and suppressed the ALPase activity of PDLC, significantly different from control. So LPS could induce the subtype change and inhibit the PDLC differentiation. 3. The production of LL-6 and TNF- a in PDLC culture stimulated with LPS To test the effects of Pg~. Pi~ En LPS on IL-6 and TNF- a production by PDLC , we applied cell culture technique and ELISA method. The results illustrated that secretion of IL.-6 and TNF- a into the culture medium was detected at 6h. The effects of LPS onIL-6 and TNF- a production were time- dependent manner in 12 or 24h. Therefor, PDLC could be revolved in the periodontitis development by induction of IL-6 and TNF- a? 4. Effects of Pg LPS on mCD14 expression on human PDLC in vitro To investgate the effects of Pg LPS on mCD14 expression on human PDLC , we used cell culture technique and the streptavidin-biotin-enzyme complex kit The results suggested positivity for mCD14 was found on healthy human PDLC and LPS could increased the expression of mCD 14. 5. Distribution of mCDL4 in inflamed gingiva and periodontium To study mCD14 expression in inflamed gingiva and periodontium, we perfoimed the immunohistochemistty staining of tissue with the streptavidin-biotin-enzyme technique. Irnmunohistochemistiy staining revealed no positivity for mCD14 from clinically healthy gingiva; while positivity for mCD14 was found on epithelial from inflamed gingiva and periodontium. It assumed that epithelial with positivity for mCD14 could play a role in transmit...