Establishment of a hepatocellular carcinoma cell line with unique metastatic characteristics through in vivo selection and screening for metastasis-related genes through cDNA microarray

Abstract

Purpose: To establish a hepatocellular carcinoma (HCC) cell line from lung metastatic lesions of human HCC in nude mice so as to provide a suitable model for the study of lung-metastasis-related molecular mechanisms. Methods: HCC clone cells MHCC97-H were inoculated into BALB/c nude mice, and the pulmonary metastatic lesions were harvested and re-implanted into nude mice for the second round of in vivo selection. The same procedure was repeated twice. A new cell line from the third round of lung metastases was established. Results: A human HCC cell line with unique metastatic characteristics was established by in vivo selection. This cell line, designated as HCCLM3, was polygonal epithelial cell with hypotriploid karyotype and population doubling time of 34.9 h. The cells were positive for alpha fetoprotein (AFP), albumin, cytokeratin 8 (CK8), and negative for hepatitis B surface antigen (HBsAg) by immunocytochemistry. Fluorescence polymerase chain reaction (PCR) showed HBV DNA integration in the cellular genome. When 5·10 6 cells were injected subcutaneously into nude mice, tumorigenicity was 100%, with a latency period of 11±1 days. Five weeks after s.c. injection, the pulmonary metastatic rate was 100%, the median number of lung metastases being 121 per mouse. After orthotopic implantation of tumor tissue into nude mouse liver for 35 days, widespread loco-regional and distant metastases occurred, with 100% abdominal wall metastases, 80% intra-abdominal cavity metastases, 100% intrahe-patic metastases, 70% diaphragm metastases, and 100% pulmonary metastases. The median number of lung metastatic lesions was 268 per mouse. Gene expression profile of HCCLM3 was compared by cDNA microarray with MHCC97-L, a clonal cell strain from the same parental cell line but with low metastatic potential; 25 differentially expressed genes were identified, 18 of which showed decreased expression and seven increased expression in HCCLM3, including the decreased expression of cell cycle control gene Rb2, mismatch repair gene hMSH2, and signal transduction gene protein kinase C b2, and increased expression of signal transduction gene MAP kinase, kinase 6. Conclusions: A new HCC cell line characterized by high pulmonary metastases via s.c. and orthotopic inoculation was established, which provides a new model for the study of liver cancer metastasis. Its gene expression profile could help in the understanding of the mechanism of metastasis and provide potential targets for anti-metastasis intervention.

EF24 Suppresses Invasion and Migration of Hepatocellular Carcinoma CellsIn Vitrovia Inhibiting the Phosphorylation of Src

“…The HCCLM-3 cell line with high metastatic potential, derived from MHCC97 parental cells [16], was provided by the Liver Cancer Institute, Shanghai Medical College of Fudan University (Shanghai, China). HCCLM-3 and HepG2 cells were cultured in Dulbecco's modified Eagle's medium (HyClone; Logan, UT, USA) supplemented with 10% fetal bovine serum (FBS; Gibco; Carlsbad, CA, USA) and 1% antibiotic (100 IU/mL penicillin and 100  μ g/mL streptomycin; Mediatech, Inc., Manassas, VA, USA).…”

Special AT-rich DNA-binding protein-1 expression is associated with liver cancer metastasis

“…SATB1 mRNA and protein expression levels were also analyzed in two different liver cancer cell lines, MHCC-97H (high metastatic potential) and HepG2 (low metastatic potential) (30,31). SATB1 mRNA and protein expression was higher in the MHCC-97H cell line than the HepG2 cell line (P=0.002 and P=0.001, respectively).…”

γ-tocotrienol inhibits angiogenesis-dependent growth of human hepatocellular carcinoma through abrogation of AKT/mTOR pathway in an orthotopic mouse model

“…HUVECs were cultured in Medium 200 supplemented with LSGS. HCCLM3 cell lines were a kind gift from Professor Zhao-You Tang at the Liver Cancer Institute (Zhongshan Hospital, Fudan University, Shanghai) and have been described previously[43]. HCCLM3 were cultured in high glucose DMEM containing 1× antibiotic-antimycotic solution with 10% FBS.…”