Abstract
Thrombospondin-1 (TSP-1), a multifunctional protein that is able to function as a negative regulator of solid tumor progression and angiogenesis, is normally present at a very low level but rapidly elevated in pathological tissues. To understand the cellular regulation of TSP-1 expression, the mode of it's expression in Hep3B, SK-HEP-1, and porcine aortic endothelial (PAE) cells was examined in the presence of all-trans retinoic acid (ATRA), interleukin-6 (IL-6), interferon-gamma (IFN-gamma), or phorbol 12-myristate 13-acetate (PMA). ATRA or IL-6 induced a dose-dependent increase of TSP-1 protein and mRNA levels in PAE cells, while they negatively regulated TSP-1 expression in the Hep3B and SK-HEP-1 cells. In contrast, PMA showed just the opposite effects on the TSP-1 expression in the same cells. IFN-gamma had little effect on TSP-1 level in Hep3B and PAE cells. The TSP-1 expression in SK-HEP-1 cells by these agents showed a close resemblance to that of liver cells rather than that of the endothelial cell line. Possible TSP-1 promoter-mediated responses by ATRA, IL-6, IFN-gamma, or PMA in Hep3B and PAE cells examined with luciferase activity of TSP-LUC reporter plasmid showed that levels of TSP-1 promoter activity were lower than that of the expressed TSP-1 protein and mRNA levels. Transfection of c-Jun and/or RARalpha expression vectors into Hep3B and PAE cells resulted in the enhanced TSP-1 promoter activity as well as the increments of of its protein and mRNA level. These results suggest that regulatory agents-induced TSP-1 expression may be attributed to mRNA stability and/or translational activation in concert with transcriptional activation and TSP-1 expression may be independently controlled via each signal pathway stimulated by PMA or ATRA.
Highlights
Thrombospondin-1 (TSP-1) is a homotrimeric glycoprotein synthesized and incorporated into extracellular matrix by numerous cells
The results demonstrated that TSP1 production in Hep3B, porcine aortic endothelial (PAE), and SK-HEP-1 cells is regulated in a cell-type specific manner by all-trans retinoic acid (ATRA), IL-6, IFN-γ, and phorbol 12-myristate 13-acetate (PMA)
To investigate the regulation of TSP-1 expression, Hep3B, SK-HEP-1, and PAE cells that express a sufficient level of TSP-1 protein were used and the culture media of each cells treated with regulatory agents; PMA, ATRA, IL-6, or IFN-γ were analyzed for TSP-1 by western blot analysis
Summary
Thrombospondin-1 (TSP-1) is a homotrimeric glycoprotein synthesized and incorporated into extracellular matrix by numerous cells. TSP-1 is a multi-function protein known to regulate cell growth, motility, and apoptosis under physiological and pathophysiological states including wound healing, angiogenesis, and neoplasia (Lawler, 1986; Bornstein, 1992; Lahav, 1993; Roberts, 1996; Chen et al, 2000). Overexpression of TSP-1 suppresses tumor growth, but normal level is not sufficient to block tumor growth (Bluel et al, 1999; Streit et al, 1999). Transfection of TSP-1 cDNA into a human breast carcinoma cell reduces primary tumor growth, metastatic potential, and angiogenesis (Weinstat-Saslow et al, 1994). TSP-1 level is inversely decreased to the increase of the proangiogenic factors, vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF-2) in human papilloma virus-16 positive cells (Bequet-Romero and Lopez-Ocejo, 2000). The regulation of TSP-1 synthesis is of critical importance for modulating various biological processes and for new approaches in clinical therapy
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