a Leptin regulation of inflammatory responses in human gingival fibroblasts

Phd ThesisObesity and type 2 diabetes mellitus (T2DM) are positively associated with the destructive, chronic inflammatory disease periodontitis. The adipokine leptin is elevated in obesity and T2DM, and promotes inflammatory responses. Gingival fibroblasts are implicated in the pathogenesis of periodontitis because inflammatory stimuli can drive these cells towards destructive, inflammatory responses. The aim of this study was to identify whether leptin promotes inflammatory responses in gingival fibroblasts, focussing on the extracellular matrix-degrading matrix metalloproteinases (MMPs), and their inhibitors (TIMPs). Primary human gingival fibroblasts (hGFs) were isolated from gingival tissue and cultured in vitro. RT-PCR, ELISA and flow cytometry were used to assess MMP, TIMP and TLR expression; hGF signalling (±chemical pathway inhibitors) was assessed by Western blotting. hGFs constitutively expressed numerous MMPs and TIMPs. Leptin increased the expression of MMP-1, MMP-3, MMP-8 and MMP-14, but not TIMPs, in hGFs. Leptin and interleukin-1 or the TLR2 agonist pam2CSK4 synergistically increased MMP-1 and MMP-3 production by hGFs; in contrast, leptin and Escherichia coli LPS regulated MMP production in a donor-dependent manner. TLR4 was detected on the surface of hGFs from all donors tested, suggesting that differential responses to E. coli LPS were not due to absent cell surface TLR4. Overall, these results suggest that leptin promotes an ECM-degrading hGF response, which is further enhanced under inflammatory conditions. Leptin activated multiple intracellular signalling pathways, but the MAPK pathway and ERK in particular, regulated leptin-stimulated MMP-1 expression in hGFs. Genome-wide expression profiling revealed that leptin enhanced the expression of genes in hGFs whose products function in inflammation; similarly, leptin enhanced the inflammatory gene expression profile induced by IL-1 in hGFs. Leptin+interleukin-1 did not promote collagen degradation in human gingival connective tissue explants. In conclusion, leptin enhances wide-ranging inflammatory responses in hGFs in a context-dependent manner. This response could be a mechanistic link between obesity, T2DM and periodontitis.Oral and Dental Research Trus