EFFECTS OF TRANSFORMING GROWTH-FACTOR-BETA-1 AND PHORBOL ESTER ON PAI-1 AND PA GENES IN HUMAN LUNG-CELLS

Transforming growth factor-beta (TGF-beta) mediates the production of extracellular matrix proteins, proteases and protease inhibitors in ep ithelial cells. Both TGF-beta and phorbol-L2-myristate-13-acetate (PMA ) exert both positive and negative effects on mitogenesis in these as well as other cell types. Phorbol esters act through stimulation of pr otein kinase C (PKC) and are among the most potent tumor promoters kno wn. The present study was conducted to determine whether the effect of TGF-beta in human non-small cell lung cancer (NSCLC) and normal human bronchial epithelial (NHBE) cells parallels that of the phorbol ester s and whether this effect of TGF-beta involves PKC. TGF-beta 1 and PMA increased expression of TGF-beta 1 mRNA 24 hr after their addition to both NSCLC and NHBE cells. The effects of these agents on expression of the mRNAs for TGF-beta 2 and TGF-beta 3 were more complex; while TG F-beta 2 and TGF-beta 3 mRNAs increased transiently in response to TGF -beta 1 in NHBE cells and TGF-beta 3 mRNA increased transiently in som e NSCLC tells, expression of these mRNAs decreased in most of these ce lls in response to PMA with the exception of the carcinoid NCI-H727 wh ere TGF-beta 2 mRNA increased dramatically. TGF-beta 1 and PMA both ca used a persistent increase in expression of the mRNAs for both plasmin ogen activator inhibitor-1 (PAI-1) and plasminogen activator (PA) up t o 24 hr in most NSCLC cells, with the increase in PAI-1 mRNA beginning several hours before that of PA mRNA. In contrast, while TGF-beta 1 a lso increased expression of PAI-1 mRNA in NHBE cells, the expression o f PA mRNA decreased simultaneously. The effect of PMA on PAI-1 and PA mRNAs was opposite of TGF-beta 1 in these cells, with expression of PA I-1 mRNA decreasing and PA mRNA increasing after addition of PR IA. Th ese data show that there is parallel regulation of the genes for TGF-b eta 1, PAI-1 and PA by TGF-beta 1 and PMA in NSCLC, but differential r egulation of the genes for PAI-1 and PA by these agents in NHBE cells. The responses of the mRNAs and proteins of TGF-beta 1, PAI-1 and PA t o TGF-beta 1 and PMA were inhibited by the serine/threonine kinase inh ibitor H7 in NSCLC cells. Treatment of NSCLC cells with TGF-beta 1 and PMA resulted in a persistent increase in the expression of fibronecti n mRNA and protein. This response was blocked by the addition of H7. I nhibition of these effects by H7 in NSCLC cells suggests that H7 block s TGF-beta responses hy inhibiting a protein serine/threonine kinase(s ), Because the effects of TGF-beta and PMA on the different TGF-beta i soforms, PA, PAI and fibronectin in NHBE and NSCLC cells are complex, our data suggest that there are distinct mechanisms for controlling th e different TSF-beta isoforms, PA, PAI and extracellular matrix protei ns in normal lung and lung cancer cells.