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UBC Theses and Dissertations

Effect of kindlin-1 deficiency on inflammatory cytokine expression in gingival keratinocytes in the presence of oral biofilm Kehler, Angela


Kindler syndrome (KS) is an autosomal recessive skin disorder of unknown etiology resulting in congenital skin blisters, photosensitivity, generalized progressive poikiloderma, and mucosal alterations. Early onset and severe periodontal disease has been noted as a common clinical finding. Kindlin-1 is an intracellular focal adhesion protein that functions in cellular adhesion in the epidermis and mucosal tissues through interaction and activation of integrins. Kindlin-1 deficiency, as occurs in KS, results in impaired adhesion of the junctional epithelium to the basement membrane and may contribute to severe periodontal disease. We hypothesize that kindlin-1 deficiency may affect other aspects of keratinocyte behavior, including expression of cytokines and mediators involved in inflammation and repair. Expression of kindlin-1 was down-regulated in human gingival keratinocytes (HGK) and confirmed via Western blot analysis and RT-PCR. HGKs were exposed to varying concentrations of a native and heat-inactivated oral biofilm extract as well as a 3-week live oral biofilm. mRNA was isolated from control and kindlin-1 deficient HGKs and RT-PCR was used to assess relative gene expression of the following gene products: β6 integrin, fibronectin ED-A and ED-B, IL-1α, IL-1β, IL-6, IL-8, TNFα, TGFβ-1, TGFβ-3, MMP-1, MMP-2, MMP-9, and tenascin C. Kindlin-1 deficiency in HGKs was found to significantly up-regulate the expression of genes encoding for β6 integrin, IL-1α, and IL-1β. β6 integrin expression was increased in K1-deficient cells in the absence of biofilm treatment and when exposed to native biofilm extract. The pro-inflammatory cytokines IL-1α and IL-1β were significantly up-regulated in K1-deficient cells as compared to control cells in keratinocytes exposed to native oral biofilm extract, and in control cells exposed to heat- inactivated biofilm. ii MMP-2, MMP-9, IL-8, tenascin C, ED-B fibronectin, and TGFβ-3 were differentially regulated by the presence of biofilm extract or live biofilm in the gingival keratinocytes. TGFβ-3 and ED- B fibronectin expression were down-regulated in cell exposed to biofilm, while IL-1α, IL-1β, MMP-2, MMP-9, and tenascin C were up-regulated. Our findings suggest that the severe and early-onset periodontal disease found in patients with KS may be a result of enhanced expression of certain pro-inflammatory cytokines.

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