A biomimetic liver model recapitulating bio-physical properties and tumour stroma interactions in hepatocellular carcinoma

Abstract

23 24 SUMMARY: 25 A protocol for a novel 3D biomimetic HCC model with accompanying fibrotic stromal 26 compartment and vasculature, to study endocrine and paracrine signaling in liver cancer. The 27 model uses physiological relevant hydrogels in ratios mimicking the bio-physical properties of the 28 stromal extracellular matrix, which is an active mediator of cellular interactions, tumor growth 29 and metastasis. 30 31 ABSTRACT: 32 Hepatocellular carcinoma (HCC) is a primary liver tumor developing in the wake of chronic liver 33 disease. Chronic liver disease and inflammation leads to a fibrotic environment actively 34 supporting and driving hepatocarcinogenesis. Insight into hepatocarcinogenesis in terms of the 35 interplay between the tumor stroma micro-environment and tumor cells is thus of considerable 36 importance. Three-dimensional (3D) cell culture models are proposed as the missing link 37 between current in vitro 2D cell culture models and in vivo animal models. Our aim was to design 38 a novel 3D biomimetic HCC model with accompanying fibrotic stromal compartment and 39 vasculature. Physiologically relevant hydrogels such as collagen and fibrinogen were 40 incorporated to mimicking the bio-physical properties of the tumor ECM. In our model LX2 and 41HepG2 cells embedded in a hydrogel matrix were seeded onto the inverted insert membrane of 42 a Transwell™ system. HUVEC cells were then seeded onto the opposite side of the membrane. 43 Three formulations consisting of ECM-hydrogels embedded with cells were prepared and the bio-44 physical properties determined by rheology. Cell viability was determined by the AlamarBlue® 45 assay over 21-days. The effect of the chemotherapeutic drug doxorubicin was evaluated in both 46 a 2D co-culture and our 3D model for a period of 72h. We show that this model is viable for 25-47 129 premalignant and tumor microenvironment by incorporating some of the key players in the 130 development of HCC. These include endothelial cells, hepatic stellate cells and malignant 131 hepatocytes, grown in a microenvironment composed of physiologically relevant hydrogels. With : Collagen gels 4 mg/mL containing 1.0 x 10 5 cells/mL seeded onto inserts in varying 442 volumes. (A) Insert on the left 100 µL, middle 150 µL and right 200 µL, directly after seeding. 443 Bubbles in the gels are still present. (B) Insert on the left 200 µL, middle 150 µL and right 100 µL, 444 after 60 min crosslinking. All gels regardless of volume still appear flat. 445 446 Combination collagen and Fibrinogen 447Based on the results from the collagen and fibrinogen concentration ranges the effect of 448 combining the collagen and fibrinogen was evaluated. Three inserts were setup with the 449 following, 4 mg/mL collagen, 20 mg/mL fibrinogen and a 1:1 ration of collagen (4 mg/mL) and 450 fibrinogen (20 mg/mL), see figure 6. Directly after seeding the hydrogels, we observed that the 451 insert with collagen alone still had a flat appearance combined with the occurrence of bubbles 452 within the gel. The inserts ...

3D Bioprinted cancer models: Revolutionizing personalized cancer therapy

“…This model showed chemotherapeutic drug resistance commonly seen in HCC patients. This 3D tumor construct could provide a reliable new platform to investigate multifocal HCC that contribute to early stages of cancer metastasis [214] . Also, a 3D printing model of intrahepatic vessel was developed with the application in a navigation surgery of hepatocellular carcinoma and provided an early proof-of-concept for further development of vessel like structures in bioprinted liver cancer constructs [215] .…”

Responsive Hydrogels Based on Triggered Click Reactions for Liver Cancer

“…[122,123] These polymer nanocomposites can be further modified into stimulus-response systems based on mechanisms that trigger the release of exogenous stimuli to achieve effective biological distribution and controlled release of drugs at specific sites. [124][125][126][127][128][129] Drug delivery systems based on nanocomposite hydrogels can be triggered by varying internal or external stimuli to achieve controlled release of drugs, as shown in Figure 5. More precisely, we summarize some internal and external stimuli that hydrogels can apply to transfer the stage of degradation or conformational, which can be utilized to achieve nanodrug delivery for cancer research.…”