| ObjectiveOur study included comparative evaluations of MMP-9 levels in GCF among smoking and non-smoking patients with chronic periodontitis and healthy,to understand the effect of smoking on MMP-9 in gingival crevicular fluid in subjects with chronic periodontitis,to eventually offer new possibilities for the smoking prevention. This study was also aimed to investigate the influence of nicotine on the secretion of MMPs by HGFs and to evaluated the influence of nicotine of different concentration on the growth and proliferation of HGFs.So that we can understand the role of nicotine on MMPs secreted by gingival fibroblast in the process of peridontitis with the hope that it can provide new view smoking is a true risk factor for periodontitis.Methods1 To investigate the change of levels of MMP-9 in gingival crevicular fluid(GCF)among smoking and non-smoking patients with chronic periodontitis and healthy group. GCF samples were collected using strip filter paper.The levels of MMP-9 were estimated by ELISA.2 (1)Specimens of healthy human gingival tissue were obtained from clinical patients under orthodontic treatment,and primary culture of HGFs was performed using the tissue culture method and the improved enzyme digestion tissue culture. (2)The fourth-generation of HGF were inoculated into a 96-Pore culture plate and then challenged by nicotine at 10μg/ml(A),50μg/ml(B),100μg/ml(C),250μg/ml(D), 500μg/ml(E),1000μg/ml(F),2000μg/ml(G),We examed the proliferation activity of HGFs to observe the influence of nicotine on the growth and the proliferation of HGF by MTT assay. (3)The fourth-generation of HGFs were inoculated into a 24-pore culture plate with cover glass and then challenged by 100μg/ml nicotine,then the cover glass with HGFs were collected at different times(0,4,8,24h)and after 24h,different concentrations and detected the expression of MMP-9 by immunocytochemical staining,so that we can investigate the influence of nicotine on the secretion of MMPs by HGFs. Results1 In subjects with chronic periodontitis, the level of MMP-9 was higher in smoking group than non-smoking group(P<0.05). And the levels of MMP-9 were higher in subjects with chronic periodontitis than in healthy subjects (P<0.05).2 The result of MTT observing the influence of nicotine on the growth and proliferation of HGF is listed as follows:At the first day,compared with the negative control group,the relative inhibition rate of A, B, C, D,E, F, G is4.36%,6.03%,8.86%,11.51%,16.33%,21.67%,25.26%respectively.At the second day,the relative inhibition rate is 9.45%,14.66%,18.11%,22.76%,25.65%,34.12%, 39.52%respectively.At the third day,the relative inhibition rate is 11.97%,15.83%,20.31%,25.12%,28.89%,44.58%,50.66%respectively.3 Immunocytochemical staining detecting the expressionof MMP-9 by HGF which challenged by nicotine revealed as follows:MMP-9 antibody mainly exprssed in cytoplasma of HGF, the expression of MMP-9 of Oh,4h,8h,24 is 0.0481±0.0093, 0.0774±0.009,0.0980±0.0092,0.1296±0.0225, (P<0.01) respectively, and after 24h,0μg/ml,100μg/ml 1000μg/ml is 0.0570±0.0096,0.1296±0.0225,0.2161±0.0301 (P<0.01),expression trends increased gradually.Conclusion:1 Smoking has effects on MMP-9 in gingival crevicular fluid in subjects with chronic periodontitis.2 Nicotine can inhibited their proliferation,and over time and concentration to extend, the inhibition rate is getting higher and higher.3 Human gingival fibroblasts in normal circumstances can express a small amount of MMP-9,under the stimulation of nicotine,expression of MMP-9 volume increased. |
PDF Full Text Request