The effects of cyclosporin on the collagenolytic activity of gingival fibroblasts.

BACKGROUND: The immunosuppressive agent cyclosporin is associated with a number of major side-effects including the development of gingival overgrowth. Although the pathogenesis of cyclosporin-induced gingival overgrowth remains unclear, it has been suggested that the finely regulated balance between extracellular matrix synthesis and degradation may be disturbed, resulting in an accumulation of excess connective tissue components within the gingival tissue. The aim of this study was to investigate the effect of cyclosporin on matrix metalloproteinases (MMP)-1 and tissue inhibitors of MMP (TIMP)-1 expression at the mRNA, protein, and enzyme activity levels. METHODS: Gingival fibroblasts were grown to confluence and then cultured in serum-free medium supplemented with cyclosporin over the concentration range of 0 to 2000 ng/ml. MMP-1 and TIMP-1 mRNA levels in cultures were determined by reverse transcription polymerase chain reaction (RT-PCR), protein levels in whole conditioned medium were assessed by enzyme-linked immunosorbent assay (ELISA), and collagenolytic activity determined using a 3H-acetylated type I collagen degradation assay. Tissue mRNA levels in normal and overgrown gingiva were also determined by RT-PCR. RESULTS: Results indicated that cyclosporin inhibited MMP-1 expression at both the mRNA and protein level in a dose- and time-dependent fashion. The effects on TIMP-1 expression were less clear, cyclosporin inhibiting mRNA expression, but having no effect on TIMP-1 protein levels at any concentration studied. Addition of the drug resulted in reduced levels of collagenolytic activity in the culture medium. MMP-1 mRNA expression was significantly reduced in overgrown compared to normal tissue. CONCLUSIONS: These results add support to the hypothesis that the accumulation of collagen seen in gingival overgrowth can be explained by a cyclosporin-induced inhibition of collagenolytic activity within the gingival tissues.

The role of soluble interleukin (IL)-6 receptor in mediating the effects of IL-6 on matrix metalloproteinase-1 and tissue inhibitor of metalloproteinase-1 expression by gingival fibroblasts.

BACKGROUND: Interleukin-6 (IL-6) is a multifunctional cytokine thought to play a role in the tissue destruction that characterizes periodontal disease. IL-6 exerts its cellular effects through a cell-surface receptor which also exists in a soluble form (sIL-6r). This study investigated the effects of IL-6 on matrix metalloproteinase (MMP)-1 activity in gingival fibroblast cultures, specifically determining the role of the sIL-6r in mediating these actions. METHODS: Fibroblasts were grown to confluence, washed in Hank's balanced saline solution (HBSS), and then cultured for 72 hours in serum-free medium supplemented with 0.2% bovine serum albumin, 1 microgram/ml Escherichia coli LPS and containing various combinations of IL-6 and its soluble receptor over the concentration range 0 to 1,000 ng/ml. MMP-1 and tissue inhibitor of MMP (TIMP)-1 protein levels in the conditioned medium were assessed by enzyme-linked immunosorbent assay (ELISA) and collagenolytic activity determined using a 3H-acetylated type I collagen degradation assay. RESULTS: Results indicated that the addition of IL-6 alone to cultures, over the concentration range 0 to 1,000 ng/ml, had no significant effect on MMP-1 protein expression. However, addition of IL-6 in combination with its soluble receptor resulted in a statistically significant, dose-dependent upregulation in MMP-1 expression. The IL-6/sIL-6r combination also induced a significant increase in collagenolytic activity in cultures. IL-6 and sIL-6r, either alone or in combination, had no marked effect on TIMP expression or cell growth. CONCLUSIONS: These data strongly suggest that future clinical studies investigating the role of IL-6 in periodontal disease must also determine the levels of sIL-6r within the periodontal tissues.