Alterations in large and small proteoglycans in bleomycin-induced pulmonary fibrosis in rats

In bleomycin (BM)-induced lung fibrosis, alterations have been shown in the expression and deposition of small proteoglycans (PGs). Less, however, is known about changes in large PGs. We investigated changes in large aggregat ing (versican [VS]), basement membrane (heparan sulfate proteoglycan [HSPG] ), and small (biglycan and fibromodulin) PGs during the development of BM-i nduced pulmonary fibrosis. BM (1.5 U) was instilled intratracheally into ma le Sprague-Dawley rats. Control rats received saline. At 7, 14, and 28 d af ter administration of BM, lungs were excised; one lung was fixed in formali n and 5-mu m sections were cut and stained with hematoxylin-eosin. The othe r lung was used for PG extraction. PGs were extracted with guanidine and we re separated through composite gel polyacrylamide gel electrophoresis (PACE ) (large PGs) and sodium dodecylsulfate-PAGE (small PGs). Gels were either stained or electrophoretically transferred and probed with antibodies to VS , HSPG, biglycan, and fibromodulin. Histologic samples showed prominent inf lammation, with abundant proteinaceous material, most evident in the sample s obtained at 7 and 14 d after administration of BM. By 28 d after BM, much of the inflammatory response had resolved, and heterogeneous distribution of fibrosis was observed. Immunoblots showed a relative abundance of VS at 7 and 14 d. Control lungs stained minimally for VS. Levels of HSPG, biglyca n, and fibromodulin were increased maximally at 14 d after administration o f BM. Immunocytochemistry showed intense immunostaining of biglycan and fib romodulin in the areas of injured lung tissue from rats 14 and 28 d after B M administration. Control lungs revealed minimal staining for small PGs. Ou r findings indicate that changes in all subclasses of PGs occur during the development of BM-induced pulmonary fibrosis.