Multimodal imaging of a humanized orthotopic model of hepatocellular carcinoma in immunodeficient mice

Abstract

The development of multimodal strategies for the treatment of hepatocellular carcinoma requires tractable animal models allowing for advanced in vivo imaging. Here, we characterize an orthotopic hepatocellular carcinoma model based on the injection of luciferase-expressing human hepatoma Huh-7 (Huh-7-Luc) cells in immunodeficient mice. Luciferase allows for an easy repeated monitoring of tumor growth by in vivo bioluminescence. The intrahepatic injection was more efficient than intrasplenic or intraportal injection in terms of survival, rate of orthotopic engraftment, and easiness. A positive correlation between luciferase activity and tumor size, evaluated by Magnetic Resonance Imaging, allowed to define the endpoint value for animal experimentation with this model. Response to standard of care, sorafenib or doxorubicin, were similar to those previously reported in the literature, with however a strong toxicity of doxorubicin. Tumor vascularization was visible by histology seven days after Huh-7-Luc transplantation and robustly developed at day 14 and day 21. The model was used to explore different imaging modalities, including microtomography, probe-based confocal laser endomicroscopy, full-field optical coherence tomography, and ultrasound imaging. Tumor engraftment was similar after echo-guided intrahepatic injection as after laparotomy. Collectively, this orthotopic hepatocellular carcinoma model enables the in vivo evaluation of chemotherapeutic and surgical approaches using multimodal imaging.

Non-invasive quantitative imaging of hepatocellular carcinoma growth in mice by micro-CT using liver-targeted iodinated nano-emulsions

“…Considering now the whole group of mice followed (Fig. 6 ), the growth capacity of HCC is logically subject-dependent, probably because, as we recently demonstrated by histology, some cells die during the first few days after injection and only part of the injected cells survive and engraft from D7, leading to the formation of the tumor 32 . However, the methodology undertaken allows emphasizing a growth factor of λ = 0.22 d −1 common to all mice, and that the difference between mice came from the moment when the tumor begins to growth.…”

“…Objectives of the current study are evaluating this innovative CT contrast agent for non-invasive HCC diagnosis in mice. Based on preclinical imaging, we will explore the feasibility in the detection and follow-up of HCC tumor implanted into the liver, comparing to the other modalities reported in literature for MRI 23 – 25 , 29 , and BLI 30 – 32 . Comparison between the different modalities is performed through the resolution of the nodule detection and the precision over a population studied, taken as absolute result.…”

Monitoring of Tumor Growth and Vascularization with Repetitive Ultrasonography in the Chicken Chorioallantoic-Membrane-Assay.

“…In contrast to radiological imaging like CT, ultrasound represents a cost-e cient alternative without the effects of ionizing radiation on either the egg or the CAM. Whilst MRI has no radiation, the increased cost factor as well as impaired imaging resolution due to movement of the embryo are obvious limitations 47 or bioluminescence detection (54). In contrast to the CAM-assay, in which neither the CAM nor the embryo are nociceptive, any evaluation of the tumor size in the mouse model results in stress or discomfort through repetitive handling, movement restriction or sedation.…”

Ultrasound‐Guided Approaches to Improve Orthotopic Mouse Xenograft Models for Hepatocellular Carcinoma

“…Compared to the surgical method (i.e., laparotomy implantation), this new method offers a better way to follow the 3R criteria (replace, refine, reduce) by improving the rate of success and reducing mortality. The tumor engraftment rate is 95% with image‐guided injection versus 86% with intrahepatic injection in the left lobe after laparotomy (Wu et al., ). A lower mortality rate is also obtained (0.01% versus 5.2% for echo‐guided injection and laparotomy, respectively).…”

“…Huh-7-Luc cells were generated in Thomas Baumert's laboratory as previously described (Wu et al, 2016). Briefly, HuH-7 cells (Japanese Collection of Research Bioresources Cell Bank, Osaka, Japan) were transduced with a retrovirus produced from a luciferaseencoding pCLNCX vector and cloned by limited dilution.…”