title: Establishment and characterization of a novel primary hepatocellular carcinoma cell line with metastatic ability in vivo.
abstract: BACKGROUND
  Hepatocellular carcinoma (HCC) is a highly aggressive and heterogeneous disease. HCC cell lines established from different patients would be useful in elucidating the molecular pathogenesis. However, success of HCC primary culture establishment remains at low rate. We aim to establish and characterize HCC primary culture and the derived cell line.
  
  
  METHODS
  Fresh tumor tissues were collected from 30 HCC patients. Culture conditions were optimized for the attachment and growth of the isolated hepatocytes. Granulin-epithelin precursor (GEP), a growth factor reported to associate with cancer stem cell properties, was examined by flow cytometry to elucidate its role on primary culture establishment. The primary cell line was characterized in detail.
  
  
  RESULTS
  Cells isolated from 16 out of 30 HCC cases (53%) had viability more than 70% and were subject to subsequent in vitro culture. 7 out of 16 cases (44%) could give rise to cells that were able to attach and grow in culture. GEP expression levels significantly correlated with the viability of isolated hepatocytes and success rate of subsequent primary culture establishment. Cells from HCC patient 21 grew and expanded rapidly in vitro and was selected to be further characterized. The line, designated HCC21, derived from a Hong Kong Chinese female patient with HCC at Stage II. The cells exhibited typical epithelial morphology and expressed albumin, AFP and HBV antigens. The cell line was authenticated by short tandem repeat analysis. Comparative genome hybridization analysis revealed chromosomal loss at 1p35-p36, 1q44, 2q11.2-q24.3, 2q37, 4q12-q13.3, 4q21.21-q35.2, 8p12-p23, 15q11.2-q14, 15q24-q26, 16p12.1-p13.3, 16q, 17p, 22q and gain at 1q21-q43 in both HCC21 cells and the original clinical tumor specimen. Sequence analysis revealed p53 gene mutation. Subcutaneous injection of HCC21 cells into immunodeficient mice showed that the cells were able to form tumors at the primary injection sites and metastatic tumors in the peritoneal cavity.
  
  
  CONCLUSIONS
  The newly established cell line could serve as useful in vitro and in vivo models for studying primary HCC that possess metastasis ability.
authors.affiliation: Department of Surgery, The University of Hong Kong, Hong Kong, China ; Center for Cancer Research, The University of Hong Kong, Hong Kong, China.
authors.forename: Phyllis Fung-Yi
authors.initials: PF
authors.lastname: Cheung
authors.affiliation: Department of Surgery, The University of Hong Kong, Hong Kong, China ; Center for Cancer Research, The University of Hong Kong, Hong Kong, China.
authors.forename: Chi Wai
authors.initials: CW
authors.lastname: Yip
authors.affiliation: Department of Surgery, The University of Hong Kong, Hong Kong, China.
authors.forename: Linda Wing-Chi
authors.initials: LW
authors.lastname: Ng
authors.affiliation: Department of Anatomical and Cellular Pathology, The Chinese University of Hong Kong, Hong Kong, China.
authors.forename: Kwok Wai
authors.initials: KW
authors.lastname: Lo
authors.affiliation: Department of Anatomical and Cellular Pathology, The Chinese University of Hong Kong, Hong Kong, China.
authors.forename: Nathalie
authors.initials: N
authors.lastname: Wong
authors.affiliation: Department of Obstetrics and Gynaecology, The Chinese University of Hong Kong, Hong Kong, China.
authors.forename: Kwong Wai
authors.initials: KW
authors.lastname: Choy
authors.affiliation: Department of Anatomical and Cellular Pathology, The Chinese University of Hong Kong, Hong Kong, China.
authors.forename: Chit
authors.initials: C
authors.lastname: Chow
authors.affiliation: Department of Pathology, Tuen Mun Hospital, Hong Kong, China.
authors.forename: Kui Fat
authors.initials: KF
authors.lastname: Chan
authors.affiliation: Department of Surgery, The University of Hong Kong, Hong Kong, China.
authors.forename: Tan To
authors.initials: TT
authors.lastname: Cheung
authors.affiliation: Department of Surgery, The University of Hong Kong, Hong Kong, China ; Center for Cancer Research, The University of Hong Kong, Hong Kong, China ; State Key Laboratory for Liver Research, The University of Hong Kong, Hong Kong, China.
authors.forename: Ronnie Tung-Ping
authors.initials: RT
authors.lastname: Poon
authors.affiliation: Department of Surgery, The University of Hong Kong, Hong Kong, China ; Center for Cancer Research, The University of Hong Kong, Hong Kong, China ; State Key Laboratory for Liver Research, The University of Hong Kong, Hong Kong, China.
authors.forename: Sheung Tat
authors.initials: ST
authors.lastname: Fan
authors.affiliation: Department of Surgery, The University of Hong Kong, Hong Kong, China ; Center for Cancer Research, The University of Hong Kong, Hong Kong, China ; State Key Laboratory for Liver Research, The University of Hong Kong, Hong Kong, China.
authors.forename: Siu Tim
authors.initials: ST
authors.lastname: Cheung
keywords: Cell line establishment; Granulin-epithelin precursor; Hepatocellular carcinoma