Analysis of interleukin-activated human gingival fibroblasts: modulation of chemokine responses by female hormones.

BACKGROUND: The female sex hormones are known to affect the response of numerous tissues to an immune challenge. Because such hormones normally fluctuate during puberty, pregnancy, and the menstrual cycle, more information about the hormonal modulation of such responses in the oral cavity is needed. Gingival fibroblasts (GF), major components of the oral tissues, are potentially sources for inflammatory mediators. METHODS: Macroarrays specific for cytokines and related proteins were used to examine the regulation of gene expression in GF under serum-free, resting conditions, after immune challenge with interleukin-1beta (IL-1beta), and in the presence of IL-1beta plus a progestin, +/-17beta-estradiol. Additional studies used enzymelinked immunosorbent assays (ELISAs) to test for secreted chemokines after the same treatments. RESULTS: Of the 392 genes on the macroarray, 66 were up- or downregulated at least 2-fold relative to the unstimulated control in an average of six different sub-lines. Chemokines represented the largest group (18%) of these regulated genes. Numerous genes whose expression was upregulated by IL-1beta were modulated downward by IL-1beta plus progestin, +/-17beta-estradiol. Measurements of the secretion of IL-8, a CXC chemokine, and MCP-1, a CC chemokine, confirmed the inhibitory effect of a progestin on these genes. CONCLUSIONS: Gingival fibroblasts are active participants in the immune response in the oral cavity, and may potentially produce many chemokine signals after exposure to IL-1beta. GF can attract neutrophils, monocytes, eosinophils, and fibroblasts to the area of injury, and aid in the wound repair process. The concentration of female sex hormones, especially progestin, may significantly affect these signaling systems.

The effects of progesterone on matrix metalloproteinases in cultured human gingival fibroblasts.

BACKGROUND: Although pregnancy gingivitis is widely believed to result from elevated hormone concentrations, the mechanism(s) involved in the etiology of this condition remain unknown. Paradoxically, despite the apparent inflammation for a prolonged period, pregnancy gingivitis rarely progresses to periodontitis and usually resolves postpartum. We used several methods to test in vitro the hypothesis that the elevated progesterone levels of pregnancy might inhibit the production of some of the matrix metalloproteinases (MMPs) that are responsible for periodontal destruction. METHODS: Cultured human gingival fibroblasts (GF) were tested in phenol red-free, serum-free medium with or without the progestogen, medroxyprogesterone acetate (MPA; 10(-6) M), using interleukin-1beta (IL-1beta) to initiate immune responses and MMP production. These MMP responses were examined by macroarrays, reverse transcription-polymerase chain reaction (RT-PCR), zymograms, and enzyme-linked immunosorbent assay (ELISA). RESULTS: Array analysis showed that pretreatment of GF with MPA reduced mRNA induction for MMPs-1, -3, and -10 in response to 6 to 8 hours incubation with IL-1beta. RT-PCR confirmed, that after 24 hours with IL-1beta , GF pretreated with MPA had undetectable levels of mRNA for MMPs-1, -2, -3, -7, -10, and -13. Zymograms of culture media from this 24-hour period showed reduction in several proteolytic activities. Examination of such 24-hour media using ELISA for MMP-3 and pro-MMP-13 confirmed that secretion of these enzymes was upregulated by IL-1beta and modulated downward by pretreatment with MPA. CONCLUSIONS: Production by GF of numerous MMPs in response to IL-1beta was significantly reduced by progesterone. This steroidal modulation of proteolytic enzymes could help to explain why pregnancy gingivitis typically is not characterized by progression to periodontitis.