Thrombin is a multifunctional serine protease that has a crucial role in blood coagulation. It is also a potent mesenchymal cell mitogen and chemoattractant and might therefore have an important role in the recruitment and local proliferation of mesenchymal cells at sites of tissue injury. We hypothesized that thrombin might also affect the deposition of connective tissue proteins at these sites by directly stimulating fibroblast procollagen production. To address this hypothesis, the effect of thrombin on procollagen production and gene expression by human foetal lung fibroblasts was assessed over 48 h. Thrombin stimulated procollagen production at concentrations of 1 nM and above, with maximal increases of between 60% and 117% at 10 nM thrombin. These effects of thrombin were, at least in part, due to increased steady-state levels of α 1 (I) procollagen mRNA. They could furthermore be reproduced with thrombin receptor-activating peptides for the protease-activated receptor 1 (PAR-1) and were completely abolished when thrombin was rendered proteolytically inactive with the specific inhibitors d -Phe-Pro-ArgCH 2 Cl and hirudin, indicating that thrombin is mediating these effects via the proteolytic activation of PAR-1. These results suggest that thrombin might influence the deposition of connective tissue proteins during normal wound healing and the development of tissue fibrosis by stimulating fibroblast procollagen production.

Transforming growth factor (TGF) β 2 gene expression was examined in murine, rat and human lung by in situ hybridization with riboprobes. Hybridization signal was observed in a variety of cells with the sense probe, and Northern-blot analysis with this probe demonstrated the presence of a novel 3.5 kb transcript. This first report suggesting the existance of a natural TGFβ 2 antisense transcript raises the possibility that such a transcript may play a role in regulating TGFβ 2 production.

The transforming growth factor-β (TGFβ) family of mediators consists of five closely related isoforms, of which three are present in mammals. TGFβ 1 has been shown to exert a biphasic effect on the proliferation of several cell types, including fibroblasts, with stimulation at low concentrations and inhibition at higher concentrations. The stimulatory effects are well characterized, but the mechanisms by which TGFβ 1 inhibits cell proliferation are incompletely understood. In the present study we have compared the effects of all three mammalian TGFβ isoforms on human lung fibroblast proliferation, and have elucidated the role of the TGFβ-induced synthesis of prostaglandin E 2 (PGE 2 ) in mediating their actions. All three isoforms stimulated fibroblast proliferation with maximal effects at 5 pg/ml (0.2 pM) and an order of potency of TGFβ 3 > TGFβ 2 > TGFβ 1 . At higher concentrations, proliferation declined, and at 40 pg/ml and above all isoforms inhibited fibroblast proliferation. Again TGFβ 3 was the most potent, but there were no significant differences between the inhibitory effects of TGFβ 1 and TGFβ 2 . Addition of indomethacin, an inhibitor of PGE 2 synthesis, did not alter the proliferative activity of any of the TGFβ isoforms, but completely overcame their inhibitory effects, restoring the stimulatory actions observed at lower TGFβ concentrations. All TGFβ isoforms stimulated PGE 2 synthesis; TGFβ 3 was approximately twice as potent as TGFβ 1 and TGFβ 2 , each of which had similar effects. These data suggest that the inhibition of fibroblast proliferation at higher concentrations of TGFβ isoforms may be mediated by autocrine stimulation of PGE 2 synthesis.