AS30D Model of Hepatocellular Carcinoma: Tumorigenicity and Preliminary Characterization by Imaging, Histopathology, and Immunohistochemistry

Abstract

Purpose This study was designed to determine the tumorigenicity of the AS30D HCC cell line following orthotopic injection into rat liver and preliminarily characterize the tumor model by both magnetic resonance imaging (MRI) and ultrasound (US) as well as histopathology and immunohistochemistry. Materials AS30D cell line in vitro proliferation was assessed by using MTT assay. Female rats (N = 5) underwent injection of the AS30D cell line into one site in the liver. Rats subsequently underwent MR imaging at days 7 and 14 to assess tumor establishment and volume. One rat underwent US of the liver at day 7. Rats were euthanized at day 7 or 14 and livers were subjected to gross, histopathologic (H&E), and immunohistochemical (CD31) analysis to assess for tumor growth and neovascularization. Results AS30D cell line demonstrated an in vitro doubling time of 33.2 ± 5.3 h. MR imaging demonstrated hyperintense T2-weighted and hypointense T1-weighted lesions with tumor induction in five of five and three of three sites at days 7 and 14, respectively. The mean (SD) tumor volume was 126.1 ± 36.2 mm3 at day 7 (N = 5). US of the liver demonstrated a well-circumscribed, hypoechoic mass and comparison of tumor dimensions agreed well with MRI. Analysis of H&E- and CD31-stained sections demonstrated moderate-high grade epithelial tumors with minimal tumor necrosis and evidence of diffuse intratumoral and peritumoral neovascularization by day 7. Conclusions AS30D HCC cell line is tumorigenic following orthotopic injection into rat liver and can be used to generate an early vascularizing, slower-growing rat HCC tumor model.

Role for Putative Hepatocellular Carcinoma Stem Cell Subpopulations in Biological Response to Incomplete Thermal Ablation: In Vitro and In Vivo Pilot Study

“…All liver/ tumor specimens were placed in 10 % neutral buffered formalin, embedded in paraffin, and sectioned with a microtome for histopathologic and immunohistochemical analysis. Paraffin-embedded sections were stained with antibodies against CD44 (1:250) or CD90 (1:100) using methods previously described [42]. All sections were reviewed by an experienced pathologist (>20 years) in a blinded and random fashion to assess tumor/liver immunostaining [42].…”

“…Paraffin-embedded sections were stained with antibodies against CD44 (1:250) or CD90 (1:100) using methods previously described [42]. All sections were reviewed by an experienced pathologist (>20 years) in a blinded and random fashion to assess tumor/liver immunostaining [42]. Digital images were captured with a Leica DMLB microscope (Leica Microsystems) equipped with a MicroPublisher 3.3 RTV camera (Q-Imaging, Surrey, BC) and the MetaVue Imaging System (v.6.3r2; Universal Imaging Corp., Downington, PA).…”

Molecular Bioluminescence Imaging as a Noninvasive Tool for Monitoring Tumor Growth and Therapeutic Response to MRI-Guided Laser Ablation in a Rat Model of Hepatocellular Carcinoma

“…Finally, the techniques outlined warrant expansion to other small animal models of HCC such as the Morris hepatoma or AS30D models. 41,42 …”

Establishment of Animal Models with Orthotopic Hepatocellular Carcinoma

“…Modeling and validation methods are very important in preclinical studies of HCC. Therefore, several molecular imaging methods, which include computed tomography (CT), ultrasound (US) imaging in radiology, positron emission tomography (PET) and single photon emission computed tomography (SPECT) in nuclear medicine, bioluminescence imaging (BLI) in biology, hematoxylin and eosin (H&E), and immunohistochemistry (IHC) in histology, are commonly employed in preclinical research and in designing clinical trials [15][16][17][18][19]. Moreover, in an effort to establish the orthotopic HCC animal models, methodological challenges involving the use of various cell lines, with or without premedication such as an immunosuppressant, and different selections of HCC cell lines have been introduced by several laboratories worldwide [20][21][22][23].…”