FIGURE 6.

Decorin induces the anti-angiogenic factor thrombospondin-1 both in cell cultures and in tumor xenografts. A and B, media conditioned by HeLa exposed to either vehicle (control) or decorin (500 nm) for 24 h were evaluated for the presence of secreted thrombospondin-1 and perlecan (as control). Notice that the levels of thrombospondin-1 are significantly elevated by exogenous decorin (***, p < 0.001, n = 4). The values were obtained by utilizing dot blots with serial dilutions of conditioned media (400–12.5 μl) and anti-thrombospondin-1 or anti-perlecan antibodies and reported as the relative signal intensities of either secreted thrombospondin-1 or perlecan, respectively. C, Western blot of cellular thrombospondin-1 from HeLa cells treated with or without decorin as indicated. D, tumor volume of HeLa xenografts at day 23. Mice bearing HeLa xenografts were treated with intraperitoneal injections of decorin (5 mg/kg) every other day over a period of 23 days (***, p < 0.001, n = 6 per group). E, induction of thrombospondin-1 in the tumor xenografts by systemic delivery of decorin. The images show representative frozen sections of HeLa xenografts at day 23 post-decorin treatment or control immunostained for thrombospondin-1. All micrographs were imaged using the same exposure and gain. The right panels are three-dimensional surface plots of the corresponding images to the left, which were generated with ImageJ software as described before (47). Bar = ∼10 μm. F and G, qPCR analysis of mRNA extracted from HeLa xenografts at day 23 post-decorin or control treatment. Data are from three independent trials run in quadruplicate and normalized to the endogenous housekeeping gene ACTB. (***, p < 0.001).