DIFFERENTIAL PROLIFERATION OF RAT LUNG FIBROBLASTS INDUCED BY THE PLATELET-DERIVED GROWTH FACTOR-AA, FACTOR-AB, AND FACTOR-BB ISOFORMS SECRETED BY RAT ALVEOLAR MACROPHAGES

Platelet-derived growth factor (PDGF)-like molecules secreted by alveolar macrophages have been postulated to be mediators of lung fibrogenesis since these cytokines stimulate the proliferation and chemotaxis of lung fibroblasts. We are studying the biology and biochemistry of rat macrophage-derived PDGF and have identified for the first time the specific isoforms of PDGF (-AA, -AB, and -BB) that these macrophages secreted in vitro following activation with either chrysotile asbestos or carbonyl iron spheres. Subsequently, the proliferative response of rat lung fibroblasts (RLF) to the different PDGF isoforms was established. Using several antibodies raised against the distinct isoforms, we established that two different PDGF-like factors with molecular masses of 30 to 34 kD and 16 to 18 kD were contained in alveolar macrophage-conditioned medium. Within each of these molecular mass regions was a mixture of all three PDGF isoforms. We estimated that the 30- to 34-kD PDGF was mainly PDGF-BB (approximately 50%), while the remaining consisted of PDGF-AA (approximately 13%) and PDGF-AB (approximately 37%). Purified recombinant PDGF isoforms were tested for their ability to stimulate the growth of early-passage RLF and Swiss 3T3 cells in a 3-day cell proliferation assay. PDGF-BB and PDGF-AB were the most potent inducers of RLF proliferation and stimulated growth half-maximally at approximately 1 ng/ml and approximately 7 ng/ml, respectively. While these two B-chain-containing dimers stimulated lung fibroblast growth by as much as 150% above control medium, the PDGF-AA homodimer stimulated lung fibroblast proliferation less than 25% above control medium at the highest concentrations tested (20 ng/ml). In contrast, Swiss 3T3 cells proliferated in the presence of all PDGF isoforms, with half-maximal stimulation values between 1 and 3 ng/ml PDGF. Alveolar macrophage-conditioned medium fractionated by gel filtration chromatography in 1 M acetic acid contained two peaks of growth-stimulatory activity for RLF with apparent molecular masses of 30 to 34 kD and 16 to 18 kD. Both of these PDGF-like factors competed for specific [I-125]PDGF-BB binding to RLF. The growth-promoting activity of these two PDGF-like factors was inhibited 60 to 80% by anti-PDGF-BB/AB. These data demonstrate that macrophages produce the three PDGF isoforms and underscore the need to identify the specific isoforms of PDGF in order to understand the biology of a paracrine link that may be operative between the macrophage as an effector cell and the fibroblast target.