Expression of IL-2 receptor β and γ chains by human gingival fibroblasts and up-regulation of adhesion to neutrophils in response to IL-2

Abstract

To investigate the role of human gingival fibroblasts (HGF), the major constituents of gingival tissue in periodontal inflammatory disease, the expression of interleukin-2 receptor (IL-2R) alpha, beta, and gamma chains was examined. Immunohistochemistry showed a pronounced accumulation of CD8(+) T cells in the inflamed lamina propria of gingival tissue from patients with adult periodontitis. HGF express IL-2Rbeta and IL-2Rgamma at mRNA and protein levels, but the expression of IL-2Ralpha could not be detected, as assessed by reverse transcriptase-polymerase chain reaction and flow cytometry. IL-2Rbeta, and -gamma expressed on HGF were functionally active, as addition of neutralizing anti-IL-2Rbeta and -gamma antibodies caused inhibition of the IL-2-induced production of monocyte chemoattractant protein-1 (MCP-1), and addition of IL-2 induced phosphorylation of Janus tyrosine kinase 3, which is critical in signaling through IL-2Rgamma in HGF. The IL-2-induced MCP-1 production was significantly inhibited by pretreatment with neutralizing antibody to IL-15. Addition of IL-2 also induced a marked up-regulation of the expression of intercellular adhesion molecule-1 (ICAM-1) on the surface of HGF, which in turn, significantly augmented the adhesion of human neutrophils, which were inhibited by an anti-ICAM-1 antibody. These results suggest that HGF express functional IL-2Rbetagamma, respond to IL-2 from infiltrated T cells, and actively participate in the inflammatory process in the periodontal region and that IL-15 produced by HGF sustains IL-2-mediated signaling in HGF.

Endogenous IL-15 Sustains Recruitment of IL-2Rβ and Common γ and IL-2-Mediated Chemokine Production in Normal and Inflamed Human Gingival Fibroblasts

“…IFN-␥ treatment (1000 IU/ml for 3 days) induced the expression of IL-15R␣ mRNA, but had no effect on the expression of IL-2R␣ mRNA and surface expression of IL-15 (Ref. 16 and data not shown). In the present study, HGF were used as confluent monolayers at subculture levels 3-10, and inflamed HGF at longer subculture levels up to 15 did not alter the phenotype of IL-2R and IL-15R or the level of surface IL-15 expression (data not shown), indicating that inflammatory conditions prevalent in the tissue environment are not involved in the expressions of IL-2R␣ and IL-15R␣.…”

“…Constitutive activation of member(s) of the IFN regulatory factor family may occur in inflamed HGF. Periodontal disease is caused by infection with anaerobic Gramnegative organisms such as Porphyromonas gingivalis, and T and B cells as well as neutrophils are found in the dense inflammatory infiltrate in this disease (16,17). In addition, Th1-or Th2-type T cells and their associated cytokines contribute to the onset and healing of this disease (27).…”

“…Cells were lysed with a lysing buffer for 30 min at 4°C with mixing as described (16). After centrifugation, the supernatants were precleared by incubation with protein G-agarose (Santa Cruz Biotechnology) for 1 h at 4°C.…”

“…PBMCs were used as positive controls. PCR for IL-2R␣, IL-2R␤, ␥ c , and human GAPDH was performed as described (16). The primers used for IL-15R␣, IL-15, and 21 aa-IL-15 were as follows: IL-15R␣, forward 5Ј-GCCAGCGCCACCCTCCACAGTAA-3Ј and reverse 5Ј-GCCAGCGGGGGAGTTTGCCTGAC-3Ј (29); IL-15, forward 5Ј-GGCTTTGAGTAATGAGAATTTCGA-3Ј and reverse 5Ј-ATCAGTT GCAATCAAGAAGTGTTG-3Ј (17); and 21 aa-IL-15, forward 5Ј-GCCT TCATGGTATTGGGAAC-3Ј and reverse 5Ј-GAATCAATTGCAATCA AGAAGTG-3Ј (20).…”

“…It has been shown that human monocytes also bear functional IL-2R␤ and ␥ c (11), human fibroblasts from adult bone marrow, embryonic skin, and lung express IL-2R␣ and IL-2R␤ (12)(13)(14), and fibroblast-like synoviocytes express functional IL-2R␤ and ␥ c (15). In addition, we have recently shown that normal HGF express functional IL-2R␤ and ␥ c and that IL-2 stimulation induced production of MCP-1 and expression of ICAM-1 by HGF, resulting in the augmentation of ICAM-1-mediated neutrophil adhesion, and that the IL-2-induced MCP-1 production was significantly inhibited by pretreatment with anti-IL-15 neutralizing mAb (16). These observations suggest that IL-2 has a wide variety of biological activities in cells other than the lymphoid cell population, and that IL-15 expressed by HGF sustains IL-2-mediated signaling in HGF.…”