Isolation And Characterization Of EpCAM~+and CD90~+Cells In Human Hepatocellular Carcinoma

Objective The expression of CD90+/EpCAM+in hepatocellular carcinoma(HCC) cell lines was examined by fluorescence activated cell sorter (FACS). Toisolate CD90+/EpCAM+cells from human hepatocellular carcinoma (HCC) celllines and study its stem cell property.Methods Five kinds of HCC cell lines (HepG2, SMMC-7721, Bel-7404,Bel-7403and SK-Hep-1) were collected and cultured in DMEM medium.FACS was used to detect the expression of CD90and EpCAM in five celllines.CD90+/EpCAM+cells were sorted from HCC cell lines. CCK-8assay wasused to observe the proliferation capacity of CD90/EpCAM positive cell andthe inhibition rate of cisplatin on CD90/EpCAM positive cell. The colony-formation ability of sorted cells were observed in terms of tablet clone formationtest.Results66.02%of hepatoma cell line Sk-Hep-1showed CD90+expression while0.15%of hepatoma cell line HepG2showed CD90+expression, and54.94%of hepatoma cell line Bel-7404showed EpCAM+expression.Culture in vitro showed that after4days the proliferation capacity of CD90+/EpCAM+cells was significantly higher than that of CD90-/EpCAM-andunsorted cells. And the inhibition rates of CD90+/EpCAM+cells was lower thanthose of CD90-/EpCAM-and unsorted cells at the same concentration ofcisplatin. The inhibition rates of CD90+, CD90-and unsorted cells were (36.88±1.44)%,（64.67±1.72）%and（50.71±1.65）%, CD90+cells showedlower inhibition than CD90-cells and unsorted cells(P＜0.05). The inhibitionrates of EpCAM+cells, EpCAM-cells and unsorted cells were （21.73±2.81）%,（49.32±6.23）%and（39.51±9.74）%, EpCAM+cells showed lowerinhibition than EpCAM-cells and unsorted cells (P＜0.05). The pelletizing rateof CD90+cells was （18.63±2.41）%in the table colony formation assay(P＜0.05), and it was significantly higher than that of CD90-cells and unsorted cells.The pelletizing rate of EpCAM+cells was （21.53±3.27）%in the tablecolony formation assay(P＜0.05), and it was significantly higher than that ofEpCAM-cells and unsorted cells.Conclusion In this study, the results show that CD90and EpCAM areindependently expressed in HCC cell lines. CD90+and EpCAM+subpopulationsorted from HCC cell lines has properties of liver cancer stem cell, whichpossessed the ability of self-renew, proliferation and drug resistence,furthermore, they had stronger ability of forming clonies.