Intrapleural instillation of tetracycline hydrochloride (TCN) is an effective means of achieving pleural fibrosis. However, its mechanism of action remains unknown. To evaluate the hypothesis that TCN stimulates pleural mesothelial cells to release growth-factor-like activity for fibroblasts we performed the following experiments. Rat visceral pleural mesothelial cells were incubated with TCN at doses ranging from 0.01 μg/ml to 100 mg/ml. The conditioned media (CM) were collected after incubation for 2 to 48 h. CM caused fibroblasts to increase incorporation of thymidine when compared with CM that was unexposed to TCN (p < 0.05). This growth-factor- like activity continued to be produced by mesothelial cells for 48 h after removal of TCN from the medium. There was a dose-response relationship since increasing doses of TCN to as much as 1 mg/ml caused increasing production of growth-factor-like activity without mesothelial cell injury as measured by trypan blue exclusion. The growth factor activity was a competence-type activity. It coeluted with human PDGF at a molecular weight of 31,000. It was heat-stable (100° C for 10 min) and sensitive to trypsin and papain but not to heat-inactivated trypsin. Addition of cycloheximide or actinomycin D inhibited its production. TCN did not have any direct effect on fibroblasts. Bleomycin CM did not contain growth-factor-like activity for fibroblasts. These data demonstrate that TCN stimulates mesothelial cells to release a growth-factor-like activity for fibroblasts. This phenomenon may play an important role in TCN-induced pleural fibrosis.