Searching for vasculogenic mimicry-associated differentially expressed genes in human hepatocellular carcinoma cell line MHCC97 using functional grouping cDNA microarray

Abstract

Objective To observe vasculogenic mimicry formation difference in human hepatocellular carcinoma cell lines MHCC97-H/L with different metastatic potentials under three-dimensional culture condition,and to study extracellular matrix and adhesion molecules-related mechanism of vasculogenic mimicry.Methods Three-dimensional cell culture system of MHCC97-H/L was established to observe tubelike structures by inverted microscope.Human umbilical vein endothelial cell(HUVEC),human hepatocellular carcinoma cell line Hep3B with non-metastatic potentials and normal human hepatic cell line HL-7702 were taken as control.Gene expression profiles of MHCC97-H and MHCC97-L were detected by Oligo extracellular matrix and adhesion molecules microarray analysis.Two differentially expressed genes were verified with semi-quantitative reverse transcriptionpolymerase chain reaction(RT-PCR)and Western blot.All data were analyzed using two sample t test.Results After a 24-hour three-dimensional culture,the length of tubelike structures was(474 ± 16)mm/cm2 in MHCC97-H,which were significantly longer than(320 ±41)mm/cm2 in MHCC97-L(t =6.119,P ＜0.05).Hep3B and HL-7702 could not form tubelike structures.Compared with MHCC97-L,the expression of 7 genes were up-regulated and the expression of 3 genes were down-regulated among a total of 113 angiogenesis genes in MHCC97-H.Two of the genes with differential expressions including tenascin C and extracellular matrix protein 1 were further validated by RT-PCR and Western blot.Conclusion MHCC97-H has stronger ability to form vasculogenic mimicry than MHCC97-L,and this perhaps correlates with the differential expression of certain extracellular matrix and adhesion molecules-related genes in MHCC97-H. Key words: Liver neoplasms; Vasculogenic mimicry; Three-dimensional culture; Gene chip