Effect of Angiostatin on the in vitro Melanoma Invasion

Abstract

Abstract: : Purpose: To characterize invasive activity in vitro of murine melanoma cell lines and the effect of angiostatin on invasiveness. Methods: Matrigel© basement membrane matrix invasion chambers were filled with 500µL bicarbonate-based culture media. The chambers were rehydrated for 2h at 37°C and the media was removed. B16LS9 and Queens melanoma cells were washed with PBS, trypsinized, centrifuged and transferred to DMEM in concentrations of 1X10₅ in 100µL. Each 100µL solution was added to the Matrigel© invasion chamber well and 450µL of DMEM was placed in the bottom of each chamber. Each melanoma cell line was added to a well alone, with 50nM glu-plasminogen, with 250nM angiostatin or with 50nM glu-plasminogen plus 250nM angiostatin. Each concentration was repeated in triplicate. The cells were incubated at 37°C for six hours and then non-invading cells were removed with a cotton swab. The invading cells were stained with Diff-Quick. Five high-power fields were counted using a micrometer. Results: The results showing number of invading cells are shown below. Results  

Conclusions: B16LS9 and Queens melanoma cells invade Matrigel© synthetic basement membranes. Plasminogen enhances invasion of B16LS9 cells, and angiostatin reduces invasion of B16LS9 and Queens cells.

Keywords: 464 melanoma • 610 tumors • 506 pathology: experimental