Effects of hGAG on aggressiveness of B16F10 tumor cells.

<p>(A) Chemical structure of hGAG. (B) Representative metastatic nodules on lung tissues. B16F10 tumor cells were treated with hGAG at the indicated concentration for 24 hours/37°C and injected into C57BL/6J mice through the tail vein. After 23 days, mice were sacrificed and metastatic nodules on lung surface were photographed. Metastatic nodules were counted under a dissecting microscope. Values are expressed as the mean ± SD. hGAG treatment reduces <i>in vivo</i> metastatic capacity of B16F10 tumor cells in mice. (C) Representative HE staining of lung tissue sections. Paraffin-embedded formalin-fixed lung tissues from each group were prepared and the sections were H&E stained. Arrows indicated tumor cells on each section. (D) Wound healing. B16F10 monolayer cells at 90–95% confluence were serum starved for 24 h and then carefully wounded using sterilized pipette tips (t = 0 h). After removing detached cells, cells were incubated with medium, tumor necrosis factor (TNFα, final conc. 50 nM), or TNFα in combination with hGAG at the indicated concentration for 24 h at 37°C and photographed immediately (t = 24 h). (E) TF levels in the plasma from mice assessed by ELISA. hGAG-treated B16F10 tumor cells injected into mice produced reduced level of TF. Data was expressed as mean ± S.E. (3–5 independent experiments). *<i>p</i><0.05, **<i>p</i><0.01.