Cell density regulates thrombospondin-1 production in malignant glioma cells

Thrombospondin-1 (TSP-1) is a multifunctional matrix protein implicated in cancer cell adhesion, migration, and invasion, inhibition of angiogenesis, and activation of latent transforming growth factor-beta (TGF-beta). The effect of cell density was investigated on the production of TSP-1, basic fibroblast growth factor (bFGF), and vascular endothelial growth factor (VEGF) by two glioblastoma cell lines. The effect of TGF-beta was also examined. The amount of intracellular TSP-1 protein decreased significantly as the cell density increased in cultures of both T98G and A172 cells. The amount of intracellular TSP-1 was highest in sparse tumor cell cultures and lowest in densely confluent tumor cell cultures. The maximum reduction of TSP-1 protein production was 56.8% and 44.6% in T98G and A172 cells, respectively. The cell density did not affect the production of bFGF or VEGF. TGF-beta2 treatment did not affect the production of TSP-1, bFGF, or VEGF proteins. Treatment with excess TGF-beta2 resulted in a slight but significant decrease (22%; P < 0.02) of TGF-beta2 production by A172 cells, but not by T98G cells. The present results indicate that the production of TSP-1 protein is regulated by cell density of glioblastoma cells, while that of angiogenic factors is not affected by tumor cell density. This suggests that high tumor cell density may tilt the angiogenic balance in favor of angiogenesis.