Idetification of an Endothelial cell growth inhibitory factor produced by oral cancer cell line cultured in protein-tree medium.

In order to study the effects of squamous cell carcinoma (SCC) on endothelial cell function, supernatants of HSC-3PF, a human SCC line able to growth without serum and any proteins, was examined the effect on cultured bovine artery endothelial cells (BAEC). It inhibited angiognesis by the BAEC in vitro three dimensional culture. The factor in the supernatant of HSC-3PF caused the inhibition of BAEC growth, named endothelial cell inhibitory factor (ECGIF), was partially purified by means of Mono-Q-anion exchange and gel filtrated column chromatography.<BR>ECGIF inhibited the proliferation of BAEC in a dose-dependent manner, and the inhibitory activity was reversible. The inhibitory effect was specific to endothelial cells, but not to other types of cells, such as smooth muscle cells, fibroblasts. ECGIF also affected cell morphology of BAEC, that is, it induced cell contraction. For further purification, ECGIF was applied on Superose 12 HR chromatography. The inhibitory activity was found in the flow through fraction. SDS-PAGE analysis of ECGIF showed that there detected 2 bands at 90-kDa and 70-kDa. To identify ECGIF with TGF-β₁ or TNF-α, we examined the colony formation of NRK-49F in soft agar in the absence of EGF and the growth of L-M cells. ECGIF stimulated the colony formation of NRK-49 F, but it did not affect L-M cell growth. Moreover the antibodies of TGF-β₁ or TNF-α failed to abolish the inhibition of BAEC growth by ECGIF. These results indicate that HSC-3PF produced ECGIF is distinguished from TGF-β and TNF-α, and regulate endothelial cell growth and morphology.