Cultured human fibroblasts synthesize and secrete thrombospondin and incorporate it into extracellular matrix.

Abstract

Thrombospondin, a major glycoprotein released from a granules of thrombin-stimulated platelets, is a disulfidebonded trimer of 160-kilodalton subunits. Cultured human foreskin and fetal lung fibroblasts secreted thrombospondin (determined by enzyme-linked immunosorbent assay) into the culture medium in a time-dependent manner (15.7 and 5.8 Thrombospondin is a major platelet a granule glycoprotein that is secreted and then partially bound to platelet membranes when human platelets aggregate in response to thrombin (1)(2)(3)(4)(5)(6)(7)(8). Thrombospondin is a 450-kilodalton (kDal) protein and is composed of three large disulfide-linked subunits (9, 10). During platelet aggregation, thrombin-stimulated platelets develop a membrane-bound lectin-like activity (11-13), which originates from a granules and appears to play a role in mediating platelet aggregation by binding to a specific receptor on other platelets (14,15 3.7 X 10'°Bq). The radioactive postculture medium was removed, centrifuged at 8,000 x g for 2 min to remove cells and debris, and frozen at -35°C. The radioactive cell layers were washed twice, removed with a rubber policeman, pelleted by centrifugation, dissolved by boiling for 5 min in 2% NaDodSO4 containing protease inhibitors as described (20), and frozen until analyzed.In experiments in which the accumulation of thrombospondin antigen was measured by enzyme-linked immunosorbent assay (ELISA), fibroblasts were cultured in minimal essential medium containing 20% rabbit serum in 2-cm2 wells of multiwell plates. When the cells were confluent, the cells were washed, and the medium was replaced with 1 ml of fresh minimal essential medium containing 20% rabbit serum. At various times after the medium change, the postculture medium was removed, centrifuged at 8,000 X g for 2 min, and frozen until assayed. 998The publication costs ofthis article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact.

Macrophage Recognition and Phagocytosis of Apoptotic Fibroblasts Is Critically Dependent on Fibroblast-Derived Thrombospondin 1 and CD36

“…9 Previous studies have also shown that fibroblasts release small amounts of TSP1. 22,23 We confirmed and extended these findings by demonstrating that TSP1 is released by fibroblasts cultured in serum-free media but release substantially more TSP1 after exposure to FasL. Using a blind-well chemotaxis assay we have shown TSP1 derived from apoptotic fibroblasts is a potent chemoattractant for macrophages.…”

Glucose and insulin modify thrombospondin 1 expression and secretion in primary adipocytes from diet-induced obese rats

“…It was initially isolated from platelets and megakaryocytes [28], but was later detected in several cell types such as macrophages and adipocytes [21,22,30,32]. Nowadays, it is known that TSP-1 is a multifunctional protein composed by multiple structural domains [5,10], and it has been especially related with several anti-angiogenic pathways [20,23,24].…”

Weakening of the repressive YY-1 site on the thrombospondin-1 promoter via c-Jun/YY-1 interaction

“…Previous reports have documented that TSP-1 is synthesized by numerous cells and is widely distributed throughout all tissues (Mosher et al, 1982;Jaffe et al, 1983;Wight et al, 1985;Varani et al, 1989). TSP-1 levels in body fluid are very low under normal conditions, but can be transiently elevated under pathological conditions or stimulation by regulatory agents (Lahav et al, 1993;Stern et al, 1996;Yesner et al, 1996).…”