From Cell Biology to Tissue Engineering


Development, differentiation, and vascular components of subcutaneous and intrahepatic Hepa129 tumors in a mouse model of hepatocellular carcinoma

Richard T. Robertson1,3, Paula M. Gutierrez1, Janie L. Baratta1, Kristoffer Thordarson1, Joshua Braslow1, Sherry M. Haynes2 and Kenneth J. Longmuir2,3

1Department of Anatomy and Neurobiology, 2Department of Physiology and Biophysics and 3The Chao Family Cancer Center, School of Medicine, University of California, Irvine, CA, USA

Offprint requests to: Richard T. Robertson, Department of Anatomy and Neurobiology, School of Medicine, University of California, Irvine, CA 92697-1280, USA. e-mail: rtrobert@uci.edu

Summary. Tumor models in mice offer opportunities for understanding tumor formation and development of therapeutic treatments for hepatocellular carcinoma. In this study, subcutaneous or intra-hepatic Hepa129 tumors were established in C3H mice. Tumor growth was determined by daily measurements of subcutaneous tumors and post-mortem studies of subcutaneous and intrahepatic tumors. Administration of Edu was used to determine cell generation dates of tumor cells. Immunohistochemistry with antibodies directed at CD31 or CD34, and intravenous injection of labeled tomato lectin revealed tumor vasculature. Tissue sections also were processed for immunohistochemistry using a panel of antibodies to proteoglycans. Comparison of Edu labeled cells with immunoreactivity allowed determination of development and differentiation of tumor cells after cell generation. Subcutaneous and intrahepatic tumors displayed similar growth over 3 weeks. Immunohistochemistry showed strong labeling for glypican-3, 9BA12, and chondroitin sulfate of tumors in both loci, while normal liver was negative. Tumor regions containing Edu labeled cells did not show significant immuno-histochemical labeling for the tumor markers until 2-3 days after Edu treatment; overlap of Edu labeled cells and immunohistochemically labeled tumor regions appeared to reach a maximum at 5 days after Edu treatment. Ectopic subcutaneous tumors displayed vascular ingrowth as the tumor cells expressed immunocytochemical markers; subcutaneous tumors displayed significantly more vascular elements than did intrahepatic tumors. Histol Histopathol 31, 403-413 (2016)

Key words: Chondroitin sulfate, Edu labeling, Glypican-3, Proteoglycan, Vascular labeling

DOI: 10.14670/HH-11-684