3D Liver Model Research Articles

112-LB: In Vitro DILI Risk Assessment of IDG16177, a Potent and Selective GPR40 Agonist, for the Treatment of Type 2 Diabetes

G protein-coupled receptor 40 (GPR40) , also known as free fatty acid receptor 1 (FFA1/FFAR1) , is expressed primarily on pancreatic β-cells and induces insulin secretion only at high glucose levels, making it a promising target for type 2 diabetes mellitus (T2DM) . Fasiglipam (TAK-875) , a partial agonist of GPR40, was discontinued in phase 3 due to hepatotoxicity despite its potent hypoglycemic effect. Previous studies of hepatic transporter inhibition, mitochondrial function, covalent protein binding and toxicity to human hepatocytes confirmed the lower drug-induced liver injury (DILI) potential of IDG16177 compared to fasiglifam, but bile acid (BA) analysis and transcription factor profiling were further performed. In a 3D human liver model, we indirectly evaluated the accumulation of BA in the liver by measuring the secretion of glycolic acid (GCA) , which is a major component of human BAs and has been identified as a significant increase in patients with DILI. Fasiglifam induced a significant accumulation of GCA at 4 μM lower than the human Cmax of 10 μM, whereas IDG16177 showed no significant accumulation at 1 μM higher than the predicted human Cmax of 0.3 μM. IDG16177 did not have a significant effect in quantitative evaluation of the activity of various transcription factors, whereas treatment with 10 uM fasiglifam for 24 hours significantly increased the activities of peroxisome proliferator activating receptor (PPAR) , activator protein 1 (AP-1) and nuclear respiratory factor 2 (NRF2) , which were highly correlated with liver disease pathogenesis/progression. Interestingly, treatment with 10 μM IDG16177 for 48 h increased the activity of FXR, a master regulator of BA metabolism that inhibits BA absorption system and BA synthesis and promotes BA enterohepatic circulation, by 1.58-fold. These results show that IDG16177 has the advantage of having a low DILI potential compared to fasiglifam in addition to its potent efficacy for the treatment of type 2 diabetes. Disclosure J. Yoon: None. H. Song: None. K. An: None. C. Hong: None. H. Kwak: None. H. Song: None.

Modeling of interstitial microwave hyperthermia for hepatic tumors using floating sleeve antenna.

Microwave hyperthermia is a treatment modality that uses microwaves to destroy cancer cells by increasing their temperature to 41-45°C. This study aims to design, model, and simulate a microwave sleeve antenna for hepatic (liver) hyperthermia. A floating sleeve antenna with 0.5 w input power was designed to resonate at 2.45GHz. The antenna was tested in six different 3D liver models. The models were varied from a very simple model without a tumor and blood vessels to complex models that contain realistic tumors and blood vessels. To test the capability of the proposed antenna for heating the interstitial tumors, the size, shape, and location of the tumor were changed. The specific absorption rate (SAR) and temperature were calculated for each model. The tumors' temperature was elevated between 43 and 45°C, while the temperature of the surrounding tissues was below 41°C. The Specific Absorption Rate (SAR) was between 29 and 30W/kg in the tumors and below 24W/Kg in the surrounding tissues. The return loss of the antenna varied from - 45 to - 25dB for the six models. The antenna could heat hepatic tumors with different sizes and locations. The heating process was performed in a short time by using a very low input power compared to all previous studies.

Registration Accuracy Using the Main Portal-vein Branch as Vascular Landmark Acquired by Intraoperative Ultrasound for 3D-Navigated Liver Resection

Introduction: The accuracy of image registration for 3D-navigated liver resection is crucial. Registration with intraoperative ultrasound (IOUS) using the right or left main portal-vein branch (PV) as landmark has the potential to optimize the accuracy and reduce errors compared to traditional liver surface landmark acquisition. Aim of this study was to analyze the IOUS registration accuracy regarding the distance to the portal vein branches and compare the use of a commercial vs. an semi-automatic in-house 3d-liver model.MethodsImage analysis of the intraoperative use of the IOUS registration process in patients who underwent 3D-navigated combined liver resection/thermal ablation. Results: 108 measurements were performed. Accuracy defined by the mean registration error was 0.75cm±0.48cm, using the left portal vein branch it was 0.65cm± 0.40cm, the right branch 0.95cm±0.54cm(p=0.001). Correlation between distance to the acquired portal vein branch landmarks and the resulting registration error was low (r2=0.36) showing a high accuracy in a distance up to 6cm to the PV. Registration error comparing the commercial 3D-liver model vs. the semi-automatic in-house 3D-liver model was not different (0.91±0.51 vs 0.99±0.58,p=0.64) Conclusion: IOUS registration algorithm shows a high accuracy in a distance up to 6cm to the region of interest in the left liver lobe using in-house 3D-image reconstruction. Accuracy in the right liver lobe was lower.

3D Patient Specific Models Enhance Surgeon’s Understanding of Preoperative Anatomy in Liver Tumors

Introduction: Liver surgery may be complex due to anatomical variants and anatomical disturbances by tumors. Preoperative planning is guided by cross-sectional imaging and the limitations of 2D imaging. Herein, we aimed to determine the impact of using 3D models on surgeons’ understanding of preoperative anatomy and surgical outcomes. Methods: Prospective single center study. Patients undergoing liver resection were included and classified based on diagnosis and by Lee et al complexity score. Preoperative cross-sectional imaging was used for the creation of 3D digital and printed models for each patient. A 5-point Likert scale survey was administered to surgeons to assess understanding of the patient anatomy. The survey was answered at two time points: before and after 3D liver model completion. Operative variables were recorded. Results: 30 patients were included, 13 (43.3%) were female and median age was 51 (IQR 32-65) years. 10 patients were living donors and 12 were diagnosed with primary liver cancer and 8 were diagnosed with metastatic disease. The most common planned procedures were right hepatectomy (44%) and right trisectionectomy (27%). Surgeons “understood” patients’ liver anatomy on 9, 3, and 5 cases before 3D education and increased to “completely understood” in 6 (p=0.7), 5 (p=0.2), and 9 (p=0.02) cases after 3D education in donor, metastatic disease, and primary liver cancer groups, respectively (Figure 1). Liver surgery complexity score, estimated blood loss, and length of stay were similar between groups. Conclusion: Patient specific 3D anatomic models improved clinical planning for the surgeon particularly in those with primary liver disease.

Efficiency of 3D imaging in children with abdominal echinococcosis

BACKGROUND: Laparoscopy in the treatment of abdominal echinococcosis is accompanied by complications. Therefore, studies on optimizing surgical approaches that reduce intra- and postoperative complications in liver echinococcosis are extremely relevant.
 AIM: This study aimed to assess the possibility of using three-dimensional (3D) technologies in children with abdominal echinococcosis to determine whether it can increase the efficiency of laparoscopic treatment and reduce complications.
 MATERIALS AND METHODS: A prospective analysis was conducted from 2013 to 2019 among 43 children with isolated liver echinococcosis who underwent multiport laparoscopic echinococcectomy. In the preoperative period, 25 patients from the main group used a complex of modern 3D technology: creating a 3D reconstruction of a liver with a parasitic cyst and then printing a 3D model of an organ with vessels and bile ducts.
 RESULTS: The use of virtual computer visualization with the 3D reconstruction of the parasitic cyst and adjacent blood vessels with bile ducts made it possible to produce a 3D liver model. This approach provided the possibility of personalized laparoscopic access and precision in performing surgeries. Postoperatively, residual cavity (n = 1, 4.0%) was observed in the main group and biliary fistula and residual cavity (n = 2, 11.1%) in the comparison group
 CONCLUSIONS: Thus, the use of 3D technologies in children with abdominal echinococcosis can increase the efficiency of laparoscopic treatment and reduce the number of early and late complications.

3D printing of liver models to assist difficult surgical operations

In the effort of a surgeon to overcome in the best possible way a difficult surgical event such as the removal of tumors from liver, it is required the use of all possible imaging-diagnostic tools to familiarize the doctor with the current patient’s situation. Even though tomography images are implying the 3D geometry of the human body they are not providing in a manageable way specific 3D model structures of separate organs. The present work concerns the use of 3D modeling techniques to produce 3D physical model of both liver and its internal geometry (vessels, tumor) using a 3D printer.

In Vitro Toxicity Screening of an Inclusive Panel of Engineered Nanomaterials using an Advanced 3D Liver Model.

In Vitro Toxicity Screening of an Inclusive Panel of Engineered Nanomaterials using an Advanced 3D Liver Model.

Predicting drug-induced liver injury in vitro by combining human 3D liver models and organ-on-chip technology

Predicting drug-induced liver injury in vitro by combining human 3D liver models and organ-on-chip technology

CBCT-based navigation system for open liver surgery: Accurate guidance toward mobile and deformable targets with a semi-rigid organ approximation and electromagnetic tracking of the liver.

PurposeThe surgical navigation system that provides guidance throughout the surgery can facilitate safer and more radical liver resections, but such a system should also be able to handle organ motion. This work investigates the accuracy of intraoperative surgical guidance during open liver resection, with a semi‐rigid organ approximation and electromagnetic tracking of the target area.MethodsThe suggested navigation technique incorporates a preoperative 3D liver model based on diagnostic 4D MRI scan, intraoperative contrast‐enhanced CBCT imaging and electromagnetic (EM) tracking of the liver surface, as well as surgical instruments, by means of six degrees‐of‐freedom micro‐EM sensors.ResultsThe system was evaluated during surgeries with 35 patients and resulted in an accurate and intuitive real‐time visualization of liver anatomy and tumor's location, confirmed by intraoperative checks on visible anatomical landmarks. Based on accuracy measurements verified by intraoperative CBCT, the system’s average accuracy was 4.0 ± 3.0 mm, while the total surgical delay due to navigation stayed below 20 min.ConclusionsThe electromagnetic navigation system for open liver surgery developed in this work allows for accurate localization of liver lesions and critical anatomical structures surrounding the resection area, even when the liver was manipulated. However, further clinically integrating the method requires shortening the guidance‐related surgical delay, which can be achieved by shifting to faster intraoperative imaging like ultrasound. Our approach is adaptable to navigation on other mobile and deformable organs, and therefore may benefit various clinical applications.

Using virtual 3D-models in surgical planning: workflow of an immersive virtual reality application in liver surgery

PurposeThree-dimensional (3D) surgical planning is widely accepted in liver surgery. Currently, the 3D reconstructions are usually presented as 3D PDF data on regular monitors. 3D-printed liver models are sometimes used for education and planning.MethodsWe developed an immersive virtual reality (VR) application that enables the presentation of preoperative 3D models. The 3D reconstructions are exported as STL files and easily imported into the application, which creates the virtual model automatically. The presentation is possible in “OpenVR”-ready VR headsets. To interact with the 3D liver model, VR controllers are used. Scaling is possible, as well as changing the opacity from invisible over transparent to fully opaque. In addition, the surgeon can draw potential resection lines on the surface of the liver. All these functions can be used in a single or multi-user mode.ResultsFive highly experienced HPB surgeons of our department evaluated the VR application after using it for the very first time and considered it helpful according to the “System Usability Scale” (SUS) with a score of 76.6%. Especially with the subitem “necessary learning effort,” it was shown that the application is easy to use.ConclusionWe introduce an immersive, interactive presentation of medical volume data for preoperative 3D liver surgery planning. The application is easy to use and may have advantages over 3D PDF and 3D print in preoperative liver surgery planning. Prospective trials are needed to evaluate the optimal presentation mode of 3D liver models.

A Critical Perspective on 3D Liver Models for Drug Metabolism and Toxicology Studies.

The poor predictability of human liver toxicity is still causing high attrition rates of drug candidates in the pharmaceutical industry at the non-clinical, clinical, and post-marketing authorization stages. This is in part caused by animal models that fail to predict various human adverse drug reactions (ADRs), resulting in undetected hepatotoxicity at the non-clinical phase of drug development. In an effort to increase the prediction of human hepatotoxicity, different approaches to enhance the physiological relevance of hepatic in vitro systems are being pursued. Three-dimensional (3D) or microfluidic technologies allow to better recapitulate hepatocyte organization and cell-matrix contacts, to include additional cell types, to incorporate fluid flow and to create gradients of oxygen and nutrients, which have led to improved differentiated cell phenotype and functionality. This comprehensive review addresses the drug-induced hepatotoxicity mechanisms and the currently available 3D liver in vitro models, their characteristics, as well as their advantages and limitations for human hepatotoxicity assessment. In addition, since toxic responses are greatly dependent on the culture model, a comparative analysis of the toxicity studies performed using two-dimensional (2D) and 3D in vitro strategies with recognized hepatotoxic compounds, such as paracetamol, diclofenac, and troglitazone is performed, further highlighting the need for harmonization of the respective characterization methods. Finally, taking a step forward, we propose a roadmap for the assessment of drugs hepatotoxicity based on fully characterized fit-for-purpose in vitro models, taking advantage of the best of each model, which will ultimately contribute to more informed decision-making in the drug development and risk assessment fields.

In Vitro Three-Dimensional Liver Models for Nanomaterial DNA Damage Assessment.

Whilst the liver possesses the ability to repair and restore sections of damaged tissue following acute injury, prolonged exposure to engineered nanomaterials (ENM) may induce repetitive injury leading to chronic liver disease. Screening ENM cytotoxicity using 3D liver models has recently been performed, but a significant challenge has been the application of such in vitro models for evaluating ENM associated genotoxicity; a vital component of regulatory human health risk assessment. This review considers the benefits, limitations, and adaptations of specific in vitro approaches to assess DNA damage in the liver, whilst identifying critical advancements required to support a multitude of biochemical endpoints, focusing on nano(geno)toxicology (e.g., secondary genotoxicity, DNA damage, and repair following prolonged or repeated exposures).

A 3D microfluidic liver model for high throughput compound toxicity screening in the OrganoPlate®

We report the development, automation and validation of a 3D, microfluidic liver-on-a-chip for high throughput hepatotoxicity screening, the OrganoPlate LiverTox™. The model is comprised of aggregates of induced pluripotent stem cell (iPSC)-derived hepatocytes (iHep) seeded in an extracellular matrix in the organ channel and co-cultured with endothelial cells and THP-1 monoblasts differentiated to macrophages seeded in the vascular channel of the 96 well Mimetas OrganoPlate 2-lane. A key component of high throughput screening is automation and we report a protocol to seed, dose, collect and replenish media and add assay reagents in the OrganoPlate 2-lane using a standard laboratory liquid handling robot. A combination of secretome measurements and image-based analysis was used to demonstrate stable 15 day cell viability, albumin and urea secretion. Over the same time-period, CYP3A4 activity increased and alpha-fetoprotein secretion decreased suggesting further maturation of the iHeps. Troglitazone, a clinical hepatotoxin, was chosen as a control compound for validation studies. Albumin, urea, hepatocyte nuclear size and viability staining provided Robust Z’factors > 0.2 in plates treated 72 h with 180 μM troglitazone compared with a vehicle control. The viability assay provided the most robust statistic for a Robust Z’ factor = 0.6. A small library of 159 compounds with known liver effects was added to the OrganoPlate LiverTox model for 72 h at 50 μM and the Toxicological Prioritization scores were calculated. A follow up dose-response evaluation of select hits revealed the albumin assay to be the most sensitive in calculating TC50 values. This platform provides a robust, novel model which can be used for high throughput hepatotoxicity screening.

Validation of a coupled electromagnetic and thermal model for estimating temperatures during magnetic nanoparticle hyperthermia

Purpose Alternating magnetic field (AMF) tissue interaction models are generally not validated. Our aim was to develop and validate a coupled electromagnetic and thermal model for estimating temperatures in large organs during magnetic nanoparticle hyperthermia (MNH). Materials and methods Coupled finite element electromagnetic and thermal model validation was performed by comparing the results to experimental data obtained from temperatures measured in homogeneous agar gel phantoms exposed to an AMF at fixed frequency (155 ± 10 kHz). The validated model was applied to a three-dimensional (3D) rabbit liver built from computed tomography (CT) images to investigate the contribution of nanoparticle heating and nonspecific eddy current heating as a function of AMF amplitude. Results Computed temperatures from the model were in excellent agreement with temperatures calculated using the analytical method (error < 1%) and temperatures measured in phantoms (maximum absolute error <2% at each probe location). The 3D rabbit liver model for a fixed concentration of 5 mg Fe/cm3 of tumor revealed a maximum temperature ∼44 °C in tumor and ∼40 °C in liver at AMF amplitude of ∼12 kA/m (peak). Conclusion A validated coupled electromagnetic and thermal model was developed to estimate temperatures due to eddy current heating in homogeneous tissue phantoms. The validated model was successfully used to analyze temperature distribution in complex rabbit liver tumor geometry during MNH. In future, model validation should be extended to heterogeneous tissue phantoms, and include heat sink effects from major blood vessels.

Human liver model systems in a dish.

The human adult liver has a multi-cellular structure consisting of large lobes subdivided into lobules containing portal triads and hepatic cords lined by specialized blood vessels. Vital hepatic functions include filtering blood, metabolizing drugs, and production of bile and blood plasma proteins like albumin, among many other functions, which are generally dependent on the location or zone in which the hepatocyte resides in the liver. Due to the liver's intricate structure, there are many challenges to design differentiation protocols to generate more mature functional hepatocytes from human stem cells and maintain the long-term viability and functionality of primary hepatocytes. To this end, recent advancements in three-dimensional (3D) stem cell culture have accelerated the generation of a human miniature liver system, also known as liver organoids, with polarized epithelial cells, supportive cell types and extra-cellular matrix deposition by translating knowledge gained in studies of animal organogenesis and regeneration. To facilitate the efforts to study human development and disease using in vitro hepatic models, a thorough understanding of state-of-art protocols and underlying rationales is essential. Here, we review rapidly evolving 3D liver models, mainly focusing on organoid models differentiated from human cells.

Evaluation of a novel navigation platform for laparoscopic liver surgery with organ deformation compensation using injected fiducials.

In laparoscopic liver resection, surgeons conventionally rely on anatomical landmarks detected through a laparoscope, preoperative volumetric images and laparoscopic ultrasound to compensate for the challenges of minimally invasive access. Image guidance using optical tracking and registration procedures is a promising tool, although often undermined by its inaccuracy. This study evaluates a novel surgical navigation solution that can compensate for liver deformations using an accurate and effective registration method. The proposed solution relies on a robotic C-arm to perform registration to preoperative CT/MRI image data and allows for intraoperative updates during resection using fluoroscopic images. Navigation is offered both as a 3D liver model with real-time instrument visualization, as well as an augmented reality overlay on the laparoscope camera view. Testing was conducted through a pre-clinical trial which included four porcine models. Accuracy of the navigation system was measured through two evaluation methods: liver surface fiducials reprojection and a comparison between planned and navigated resection margins. Target Registration Error with the fiducials evaluation shows that the accuracy in the vicinity of the lesion was 3.78±1.89mm. Resection margin evaluations resulted in an overall median accuracy of 4.44mm with a maximum error of 9.75mm over the four subjects. The presented solution is accurate enough to be potentially clinically beneficial for surgical guidance in laparoscopic liver surgery.

Feasibility of Augmented Reality–Guided Transjugular Intrahepatic Portosystemic Shunt

PurposeTo investigate an augmented reality (AR)–guided endovascular puncture to facilitate successful transjugular intrahepatic portosystemic shunt (TIPS). Materials and MethodsAn AR navigation system for TIPS was designed. Three-dimensional (3D) liver models including portal and hepatic vein anatomy were extracted from preoperative CT images. The 3D models, intraoperative subjects, and electromagnetic tracking information of the puncture needles were integrated through the system calibration. In the AR head-mounted display, the 3D models were overlaid on the subjects, which was a liver phantom in the first phase and live beagle dogs in the second phase. One life-size liver phantom and 9 beagle dogs were used in the experiments. Imaging after puncture was performed to validate whether the needle tip accessed the target hepatic vein successfully. ResultsEndovascular punctures of the portal vein of the liver phantom were repeated 30 times under the guidance of the AR system, and the puncture needle successfully accessed the target vein during each attempt. In the experiments of live canine subjects, the punctures were successful in 2 attempts in 7 beagle dogs and in 1 attempt in the remaining 2 dogs. The puncture time of needle from hepatic vein to portal vein was 5–10 s in the phantom experiments and 10–30 s in the canine experiments. ConclusionsThe feasibility of AR-based navigation facilitating accurate and successful portal vein access in preclinical models of TIPS was validated.

3D Liver Models: Biofabrication of Hepatic Constructs by 3D Bioprinting of a Cell‐Laden Thermogel: An Effective Tool to Assess Drug‐Induced Hepatotoxic Response (Adv. Healthcare Mater. 21/2020)

3D Liver Models: Biofabrication of Hepatic Constructs by 3D Bioprinting of a Cell‐Laden Thermogel: An Effective Tool to Assess Drug‐Induced Hepatotoxic Response (Adv. Healthcare Mater. 21/2020)

Hydrogel microfluidic-based liver-on-a-chip: Mimicking the mass transfer and structural features of liver.

Liver is fed by nutrition via diffusion across the vascular wall from blood flow. However, hepatocytes in liver models are directly exposed to the perfusion culture medium, where the shear stress reduces the cell viability and liver-specific functions. By mimicking the mass transfer and structural features of hepatic lobule, we designed a microfluidic liver-on-a-chip based on the di-acrylated pluronic F127 hydrogel. In the hydrogel chip, hepatocellular carcinoma HepG2 and human hepatic stellate cell LX-2 were statically cultured inside the microwells on the outer channel. These hepatic cells were fed by the diffused medium from the adjacent but separated inner channel with endothelial cell monolayers, which was perfused by the medium with physiologically relevant shear stress. As found, the hepatic cells in the liver-on-a-chip rapidly formed spheroids within 1-dayincubation and expressed about one to two-fold higher viability/liver-specific functions than the corresponding static culture for at least 8 days. Moreover, the presence of endothelial cells also contributed to the expression of liver-specific functions in the liver-on-a-chip. Therefore, the proposed liver-on-a-chip provides a new concept for construction of 3D liver models in vitro, and shows the potential value for a variety of applications including bio-artificial livers and drug toxicity screening.

Adaptation of the in vitro micronucleus assay for genotoxicity testing using 3D liver models supporting longer-term exposure durations.

Following advancements in the field of genotoxicology, it has become widely accepted that 3D models are not only more physiologically relevant but also have the capacity to elucidate more complex biological processes that standard 2D monocultures are unable to. Whilst 3D liver models have been developed to evaluate the short-term genotoxicity of chemicals, the aim of this study was to develop a 3D model that could be used with the regulatory accepted in vitro micronucleus (MN) following low-dose, longer-term (5 days) exposure to engineered nanomaterials (ENMs). A comparison study was carried out between advanced models generated from two commonly used liver cell lines, namely HepaRG and HepG2, in spheroid format. While both spheroid systems displayed good liver functionality and viability over 14 days, the HepaRG spheroids lacked the capacity to actively proliferate and, therefore, were considered unsuitable for use with the MN assay. This study further demonstrated the efficacy of the in vitro 3D HepG2 model to be used for short-term (24 h) exposures to genotoxic chemicals, aflatoxin B1 (AFB1) and methyl-methanesulfonate (MMS). The 3D HepG2 liver spheroids were shown to be more sensitive to DNA damage induced by AFB1 and MMS when compared to the HepG2 2D monoculture. This 3D model was further developed to allow for longer-term (5 day) ENM exposure. Four days after seeding, HepG2 spheroids were exposed to Zinc Oxide ENM (0–2 µg/ml) for 5 days and assessed using both the cytokinesis-block MN (CBMN) version of the MN assay and the mononuclear MN assay. Following a 5-day exposure, differences in MN frequency were observed between the CBMN and mononuclear MN assay, demonstrating that DNA damage induced within the first few cell cycles is distributed across the mononucleated cell population. Together, this study demonstrates the necessity to adapt the MN assay accordingly, to allow for the accurate assessment of genotoxicity following longer-term, low-dose ENM exposure.