We then examined levels of CD44 and Hedgehog signaling proteins (Ptch1, Shh and Gli1) in HCC cell lines grown in a monolayer or in a spheroid condition by Western blot analysis and found that levels of CD44 and Hedgehog signaling proteins were higher in the spheroid cells than in the monolayer cells (Fig. 7d). This evidence concerns the gene CD44 and hepatocellular carcinoma.