The Researches On Expression Of Nuclear Factor-κB Dependent Inflammatory Mediators In Oral Lichen Planus And Corresponding Signaling Pathways

BackgroundOral lichen planus(OLP) is a chronic inflammatory oral mucosal disease ofunknown etiology. The histology of OLP is characterized by a dense subepitheliallympho-histiocytic infiltrate, increasing numbers of intra-epithelial lymphocytes, anddegeneration of basal keratinocytes. Although there are no consistent serological changesassociated with OLP, it has been widely accepted that some immunological aberrancesinvolved in development. Many biochemical mediateors, including cytokines,growth factors, cell adhension molecules, matrix metalloproteinase, etc. play a keyrole in the pathogenesis of oral lichen planus. Among these mediators, cytokines, theproteins secreted by various cell types, have been paid high attention for the uniquerole in mediateing immune and inflammatory processes. Recent studies have shownan increased production of pro-inflammatory cytokines including TNF-α,IL-1,IL-2,IL-6 and so on in patients with OLP, suggesting a pathogenic role of theirdisorganization in this disease. However, exact mechanism of their regulation andrelease remains unclear up to now. A pivotal pathogenic role in the pathodenesis oflong-lasting inflammatory processes is played by the activation of nuclear factor-κB (NF-κB), a primary transcription factor which upon translocation to the nucleus,binds to promoter regions of different genes encoding immune andpro-inflammatory mediators. Consequently, the study of NF-κB and its regulatinginflammatory mediator gene and screening sensitive biomarkers would be valuablein uncovering the pathogenesis of OLP, and provide new strategies and medicationfor OLP therapies.Objectives1. To explore the expression of NF-κB-dependent inflammatory mediatorsincluding tuomor necrosis factor-α(TNF-α), interlukin-6 (IL-6), interlukin-8 (IL-8),matrix metalloproteinase-9 (MMP-9) in serum and saliva, in the pathogenesis anddevelopment of OLP, and to screen sensitive biomarkers for monitoring diseaseactivity and therapeutic responses in OLP.2. To detect the expression of NF-κB and MMP-9 in the lesion of OLP, and toreveal the involvement of NF-κB and MMP-9 in pathogenesis of OLP.3. To investigat NF-κB signaling pathways involved in MMP-9 induced by TNF-αin peripheral blood monouclear cells in patients with OLP.Material and Methods1. A total of 30 Chinese patients with OLP (14 reticular form, 16 erosive form),and 30 age-sex-matched controls were enrolled in this study. Serum and their pairingwhole unstimulated saliva (WUS) samples were collected by the standard technique.The level of TNF-α, IL-6, IL-8, and MMP-9 wasscreened by enzyme-linkedimmunosorbent assay (ELISA), respectively. The differences between groups andthe correlation of these parameters between serum with saliva were statisticallyanalyzed.2. Using immunohistochemistry, the present study explored the expression ofNF-κdp65 and MMP-9 in 22 cases (11 reticular form, 11 erosive form) of OLP and10 cases of healthy controls, and analysed the correlation between NF-κd3 p65 andMMP-9. 3. PBMC were isolated from blood speicimens from 15 patients with OLP and 15healthy controls,and cultured with or TNF-αin vitro. Detect the activation of NF-κBand the supematant level of MMP-9 in PBMC by immunohistochemistry and ELISAmethod. Examine the effects and mechanism of TNF-αinduced activation of NF-κBand expression of MMP-9 in PBMC. Pyrrolidine dithiocarbamate(PDTC) was usedas the intervention factor to observe blocking the activation of NF-κB andexpression of MMP-9 in PBMC.Results1. Detection of the serum and salivary levels of TNF-α,IL-6,IL-8,MMP-9 inOLP:①A significantly increased levels of these mediators, both in serum and saliva,was found in OLP group in comparison with those in healthy subjects. With regardsto the subtypes of this illness, salivary TNF-α, IL-6, IL-8, MMP-9 were elevated ineither reticular or erosive form of OLP than those in the normal control, while theserum concentrations of these cytokines were merely increased in erosive form ofOLP. Salivary IL-8 and MMP-9 level in erosive form was inclined than that in thereticular form roup, with no difference of salivary TNF-α, IL-6 between those twosubtypes.②A positive statistical correlations existed between these cytokines inserum with saliva, while the levels of these cytokines in saliva inclined much moreremarkably than in serum.2. The positive incidence of NF-κB p65 expressed in OLP was 77.3％,and theexpression in healthy controls was 10％. The positive incidence of NF-κB p65 inreticular form was 54.5％; in erosive form was l00％.The positive incidence ofNF-κB p65 were significantly higher in erosive form than that in reticular form (P=0.04＜0.05) .The positive incidence of MMP-9 expressed in OLP was 63.6％,and the expression in healthy controls was 10％. The positive incidence of MMP-9in reticular form group was 36.4％; in erosive form group was 90.9％. The positiveincidence of MMP-9 were significantly higher in erosive form than that in reticularform (P=0.03＜0.05) .In addition, there is a positive coincidence between NF-κB p65 and MMP-9 (x=0.680, P=0.00＜0.05).3. PDTC can inhibit TNF-αinduced the expression of MMP-9 in PBMC fromOLP patients. Treat PBMC with TNF-在(10ng/ml) for-24h, the positive incidence ofNF-rd3 p65 expressed and the superant levels of MMP-9 in OLP group wer elevatedsimultaneously.While pretreated PBMC with PDTC, the expression of NF-κB p65and MMP-9 were inhibited.Conclusion1. Saliva-based test may be a more cost-effective adjunctive tool in the diagnosisand follow-up of OLR2. Salivary IL-8 and MMP-9 levels can be a promising biomarker for themonitoring of the disease activity of OLP.3. NF-κB is a major and essential factor in regulating the high expression ofMMP-9, and play a critical role in the pathogenesis of OLR The inhibition of NF-κBactivation may be an new target for therapies of OLP.