Human eosinophils regulate human lung- and skin-derived fibroblast properties in vitro: a role for transforming growth factor beta (TGF-beta).

Eosinophils have been associated with fibrosis. To investigate their direct role in fibrosis, human peripheral blood eosinophil sonicate was added to human lung or dermal fibroblasts, and proliferation ([(3)H]thymidine) and collagen synthesis ([(3)H]proline) were evaluated. Proliferation was enhanced significantly in the monolayers in a dose-dependent manner. The activity of the eosinophil fibrogenic factor(s) remained unaltered when heated (56 degrees C, 30 min). Supernatants of cultured eosinophils (20 min or 18 hr) also enhanced lung fibroblast proliferation, indicating that the preformed mitogenic factor(s) can be released both promptly and with a long kinetic. Eosinophils significantly decreased collagen production in lung fibroblasts while increasing it in dermal fibroblasts. However, eosinophils containing matrix metalloproteinase 9 (zymography) in latent form and tissue inhibitors of metalloproteinases 1 and 2 (reverse zymography) did not influence either fibroblast matrix metalloproteinases or tissue inhibitors of metalloproteinases. Eosinophil sonicate added to skin and lung fibroblasts in tridimensional collagen lattices significantly enhanced lattice contraction. Transforming growth factor beta (TGF-beta) is a major fibrogenic cytokine produced by eosinophils. Therefore, to assess its role, eosinophil sonicate was preincubated with anti-TGF-beta neutralizing antibodies. This treatment partially inhibited proliferation of lung and collagen synthesis of dermal fibroblasts and suppressed the stimulation of lattice contraction, indicating the fibrogenic role of eosinophil-associated TGF-beta. In conclusion, we have shown that eosinophils act as direct modulatory cells in fibroblast proliferation, collagen synthesis, and lattice contraction, in part, through TGF-beta. These data corroborate the importance of eosinophils in skin and lung fibrosis.

Mast cells retain their responsiveness upon continuous and repetitive exposure to antigen.

To assess the effects of prolonged and repetitive exposure of antigen on mast cells (MC) we utilized our co-culture system in which rat peritoneal MC are kept viable and functionally active on 3T3 fibroblasts. MC were presensitized with IgE anti-dinitrophenyl-human serum albumin (anti-DNP-HSA), activated with the relevant antigen (DNP-HSA) (69.4% histamine release) and incubated for 7 days in medium alone or medium containing the antigen with or without Ca2+ ions. During the prolonged incubation MC released only small amounts of histamine (20-26 ng/plate/day) under all the different culture conditions. Moreover, these MC were not activated after each medium replacement with fresh antigen. When activated MC cultured in medium containing antigen and Ca2+ were presensitized prior to a second challenge performed on days 1, 2, 5 and 7, they released 35-45% histamine but were still partially desensitized compared to initially challenged MC. Immunofluorescence studies demonstrated that the MC unresponsiveness to a second challenge could be partially attributed to lack of membrane-associated IgE antibodies. Repetitive activation (four times) of MC at daily intervals did not abrogate their releasing capacity although the intracellular histamine content was diminished. When repetitive activation was performed 2 and 4 hr after the first challenge, a more intense MC refractoriness was observed (release of 17.3% and 21.1% histamine respectively versus 67.7%). It is suggested that MC activation occurs even after continuous exposure to antigen and that MC can respond to repetitive immunological stimulation.