Influence of angiogenesis inhibitors, endostatin and PF-4, on lymphangiogenesis.
This study was designed to screen potential safe and effective inhibitors of lymphangiogenesis. Lymphatic endothelial cells from pig thoracic duct were isolated and cultured. The control group, 3 endostatin, and 3 PF-4 experimental groups were tested for effects on proliferation and distance of migration of the cultured cells by two methods (method of the scraping line and MTT assay), and observations by light, confocal, and electron microscopy were also made. Total cells migrating past the scrape line for the endostatin control group was 28.6 +/- 1.2 (mean +/- standard error) and the 3 endostatin experimental groups (50 ng/ml, 100 ng/ml, and 150 ng/ml), respectively, were 17.5 +/- 0.6, 10.5 +/- 0.5, and 4.8 +/- 0.3 (all p < 0.05 cf control). Migration distance for the endostatin control group was 381.7 +/- 9.67 microm, and the migration distance of the 3 endostatin experimental groups, respectively, were 252.9 +/- 5.58, 164.5 +/- 7.09, and 91.2 +/- 7.98 microm (all p < 0.05). Cell migration number for the PF-4 control group was 28.3 +/- 1.0 compared with doses of 40 ng/ml, 80 ng/ml, or 120 ng/ml of PF-4, respectively, were 13.6 +/- 0.7, 9.5 +/- 0.6, and 4.6 +/- 0.4 (all p < 0.05 cf control). Migration distance of cells in PF-4 control group was 419.9 +/- 5.87 microm, and the 3 PF-4 experimental groups, respectively, were 199.2 +/- 8.16, 152.5 +/- 7.28, and 104.2 +/- 6.70 microm (all p < 0.05 cf control). The MTT assay confirmed that as the concentrations of endostatin and PF-4 were increased, the inhibitory effect was increased. We conclude that endostatin and PF-4 are able to inhibit the migration and proliferation of lymphatic endothelial cells, and these effects are dose-dependent.