LATENT TRANSFORMING GROWTH-FACTOR-BETA-1 AND ITS BINDING-PROTEIN ARE COMPONENTS OF EXTRACELLULAR-MATRIX MICROFIBRILS

We studied the localization of latent transforming growth factor-beta 1 (TGF-beta 1) and its binding protein (LTBP-1) in the extracellular m atrix of cultured human fibroblasts by immunofluorescence and immunoel ectron microscopy. Immunofluorescence of confluent fibroblast cultures indicated that LTBP-1 localizes to extracellular fibrillar structures resembling fibronectin-collagen matrix, Similar fibrillar structures were detected in cells stained with antibodies specific for TGF-beta 1 propeptide (beta 1-LAP). Both LTBP-1 and beta 1-LAP co-localized with fibronectin in double immunofluorescence analysis, These fibrillar st ructures mere resistant to extraction with sodium deoxycholate, which is further evidence that LTBP-1 and large latent TGF-beta 1 complexes are integral components of the extracellular matrix, SV-40-transformed human fibroblasts lacked extracellular LTBP-1 fibers, EM analysis rev ealed approximate to 10-nm-thick microfibrils that were labeled by ant i-LTBP at 90-140-nm intervals. In addition, LTBP-1 was found in struct ures that were heavily labeled for fibronectin. The accumulation of LT BP-1 in the fibronectin matrix could be reconstituted in vitro. When i solated matrix components were immobilized on nitrocellulose and incub ated with fibroblast conditioned medium, LTBP-1 from the medium associ ated with cellular fibronectin but not with heparan or chondroitin sul fate, vitronectin, tenascin, laminin, or collagen I or IV, The associa tion of LTBP-1 with cellular fibronectin was abolished by treatment of the medium with plasmin, which cleaves LTBP-1 and inhibits its assemb ly to matrix, The present results indicate that latent TGF-beta 1 comp lexes are components of the extracellular matrix and suggest that alte rations of the pericellular matrix could result in aberrant TGF-beta s ignaling.