Monique Verstegen - Academia.edu (original) (raw)

Papers by Monique Verstegen

Bioengineering, Mar 7, 2022

This article is an open access article distributed under the terms and conditions of the Creative... more This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY

Best Practice & Research in Clinical Gastroenterology, Apr 1, 2017

Due to the complex function and structure of the liver, resourceful solutions for treating end-st... more Due to the complex function and structure of the liver, resourceful solutions for treating end-stage liver disease are required. Currently, liver transplantation is the only durable therapeutic option. However, due to a worldwide donor shortage, researchers have been looking in other fields for alternative sources of transplantable liver tissue. Recent advances in our understanding of liver physiology, stem cell and matrix biology have accelerated tissue engineering research. Most notable is the discovery of a culture system to grow liver-like organoids from human hepatic stem cells. The extensive expansion capacity of these stem cells has contributed greatly to the availability of hepatocyte-like cells for tissue engineering. In addition, new techniques are explored to obtain biological liver scaffolds from full size donor organs. This review summarizes these state-of-art techniques which may lay the groundwork towards recreating transplantable tissue from autologous or allogenic stem cells in the coming decade.

Liver Transplantation, Jun 20, 2019

Cell death is a natural process for the turnover of aged cells, but it can also arise as a result... more Cell death is a natural process for the turnover of aged cells, but it can also arise as a result of pathological conditions. Cell death is recognized as a key feature in both acute and chronic hepatobiliary diseases caused by drug, alcohol, and fat uptake; viral infection; or after surgical intervention. In the case of chronic disease, cell death can lead to (chronic) secondary inflammation, cirrhosis, and the progression to liver cancer. In liver transplantation, graft preservation and ischemia/reperfusion injury (IRI) are associated with acute cell death. In both cases, so-called programmed cell death modalities are involved. Several distinct types of programmed cell death have been described of which apoptosis and necroptosis are the most well known. Parenchymal liver cells, including hepatocytes and cholangiocytes, are susceptible to both apoptosis and necroptosis, which are triggered by distinct signal transduction pathways. Apoptosis is dependent on a proteolytic cascade of caspase enzymes, whereas necroptosis induction is caspase-independent. Moreover, different from the "silent" apoptotic cell death, necroptosis can cause a secondary inflammatory Accepted Article This article is protected by copyright. All rights reserved. cascade, so-called necroinflammation, triggered by the release of various damage-associated molecular patterns (DAMPs). These DAMPs activate the innate immune system, leading to both local and systemic inflammatory responses, which can even cause remote organ failure. Therapeutic targeting of necroptosis by pharmacological inhibitors, such as necrostatin-1 (Nec-1), shows variable effects in different disease models. In this review, we will discuss the mechanisms of necroptosis, and we will focus on liver transplantation and liver diseases, such as acute liver failure, fatty liver diseases, cholestatic liver diseases, chronic viral hepatitis, and primary liver cancer. Furthermore, we will review the clinical relevance of necroptotic cell death and therapeutic potential by targeting cell death in liver diseases. Cell death is a fundamental process that is essential in embryonic and (neo)natal development and homeostasis in all organs, including the liver. Cell death is a means of removing aged and

STAR protocols, Sep 1, 2023

Journal of Inherited Metabolic Disease, Nov 3, 2021

Take-down policy If you believe that this document breaches copyright please contact us providing... more Take-down policy If you believe that this document breaches copyright please contact us providing details, and we will remove access to the work immediately and investigate your claim.

Journal of translational autoimmunity, 2020

This is a PDF file of an article that has undergone enhancements after acceptance, such as the ad... more This is a PDF file of an article that has undergone enhancements after acceptance, such as the addition of a cover page and metadata, and formatting for readability, but it is not yet the definitive version of record. This version will undergo additional copyediting, typesetting and review before it is published in its final form, but we are providing this version to give early visibility of the article. Please note that, during the production process, errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

MBio, Aug 25, 2020

Biliary atresia (BA) is a neonatal liver disease characterized by progressive fibroinflammatory o... more Biliary atresia (BA) is a neonatal liver disease characterized by progressive fibroinflammatory obliteration of both intrahepatic and extrahepatic bile ducts. The etiologies of BA remain largely unknown, but rotavirus infection has been implicated at least for a subset of patients, and this causal relation has been well demonstrated in mouse models. In this study, we aim to further consolidate this evidence in human biliary organoids. We obtained seven batches of human biliary organoids cultured from fetal liver, adult liver, and bile duct tissues. We found that these organoids are highly susceptible and support the full life cycle of rotavirus infection in three-dimensional culture. The robust infection triggers active virus-host interactions, including interferon-based host defense mechanisms and injury responses. We have observed direct cytopathogenesis in organoids upon rotavirus infection, which may partially recapitulate the development of BA. Importantly, we have demonstrated the efficacy of mycophenolic acid and interferon alpha but not ribavirin in inhibiting rotavirus in biliary organoids. Furthermore, neutralizing antibody targeting rotavirus VP7 protein effectively inhibits infection in organoids. Thus, we have substantiated the causal evidence of rotavirus inducing BA in humans and provided potential strategies to combat the disease. IMPORTANCE There is substantial evidence indicating the possible involvement of rotavirus in biliary atresia (BA) development, at least in a subset of patients, but concrete proof remains lacking. In a mouse model, it has been well demonstrated that rotavirus can infect the biliary epithelium to cause biliary inflammation and obstruction, representing the pathogenesis of BA in humans. By using recently developed organoids technology, we now have demonstrated that human biliary organoids are susceptible to rotavirus infection, and this provokes active virus-host interactions and causes severe cytopathogenesis. Thus, our model recapitulates some essential aspects of BA development. Furthermore, we have demonstrated that antiviral drugs and neutralizing antibodies are capable of counteracting the infection and BA-like morphological changes, suggesting their potential for mitigating BA in patients. KEYWORDS biliary atresia, rotavirus infection, human organoids B iliary atresia (BA) is characterized by progressive fibroinflammatory obliteration of the bile ducts, resulting in chronic cholestasis and biliary cirrhosis. It is one of the leading causes for liver transplantation in infants (1, 2). Exposure to rotavirus in mice has demonstrated the infection in biliary epithelium, resulting in BA-like biliary inflammation and obstruction (3). Nevertheless, whether rotavirus is a causal agent for BA in patients remains controversial, also because of a paucity of preclinical models. Organoid technology provides an excellent way forward here.

Stem Cells and Development, Jun 15, 2015

After organ transplantation, recipient T cells contribute to graft rejection. Mesenchymal stromal... more After organ transplantation, recipient T cells contribute to graft rejection. Mesenchymal stromal cells from the bone marrow (BM-MSCs) are known to suppress allogeneic T-cell responses, suggesting a possible clinical application of MSCs in organ transplantation. Human liver grafts harbor resident populations of MSCs (L-MSCs). We aimed to determine the immunosuppressive effects of these graft-derived MSCs on allogeneic T-cell responses and to compare these with the effects of BM-MSCs. BM-MSCs were harvested from aspirates and L-MSCs from liver graft perfusates. We cultured them for 21 days and compared their suppressive effects with the effects of BM-MSCs on allogeneic T-cell responses. Proliferation, cytotoxic degranulation, and interferon-gamma production of alloreactive T cells were more potently suppressed by L-MSCs than BM-MSCs. Suppression was mediated by both cell-cell contact and secreted factors. In addition, L-MSCs showed ex vivo a higher expression of PD-L1 than BM-MSCs, which was associated with inhibition of T-cell proliferation and cytotoxic degranulation in vitro. Blocking PD-L1 partly abrogated the inhibition of cytotoxic degranulation by L-MSCs. In addition, blocking indoleamine 2,3-dioxygenase partly abrogated the inhibitive effects of L-MSCs, but not BM-MSCs, on T-cell proliferation. In conclusion, liver graft-derived MSC suppression of allogeneic T-cell responses is stronger than BM-MSCs, which may be related to in situ priming and mobilization from the graft. These graft-derived MSCs may therefore be relevant in transplantation by promoting allohyporesponsiveness.

Cancer Research, Apr 4, 2023

Hepatocellular carcinoma (HCC) is the most common type of liver cancer. Its incidence is increasi... more Hepatocellular carcinoma (HCC) is the most common type of liver cancer. Its incidence is increasing, and is closely related to advanced liver disease. Interactions in the HCC microenvironment between tumor cells and the associated stroma actively regulate tumor initiation, progression, metastasis, and therapy response. In the present study, we used the OrganoPlate graft to establish a co-culture system consisting of dissociated HCC tumors (HCC 1-8) and cell lines, HCC derived fibroblasts and vasculature. Cultures were prepared and validated by assessing their response to Sorafenib and Lenvatinib (72 hours). Cultures had their viability (alamar blue assay), and chemokine/cytokine levels in the supernatant (Luminex) determined. In addition, the organization of the vasculature in the tumor compartment was studied through immunostainings, confocal imaging, and subsequent morphological analyses. HCC models were characterized by a range of specific markers, tumor (albumin), endothelial (CD31 and VE-Cadherin) and stromal (aSMA) cells. CD31 immunostained cultures were imaged, and morphology changes quantified. Sorafenib and Lenvatinib induced changes in the tumor vasculature area and organization. Hereby, we present vascularized patient-derived HCC models that include relevant cellular players of the HCC microenvironment. These co-cultures are highly suitable for studying specific cell types as well as patient-specific responses. We envision that this patient derived model will evolve to become a platform for understanding the interplay between angiogenesis, stroma and immune infiltrate in HCC. Citation Format: Orsola Mocellin, Abbie Robinson, Aleksandra Olczyk, Stephane Treillard, Thomas Olivier, Chee Ng, Jeroen Heijmans, Désiréé Goubert, Arthur Stok, Gilles van Tienderen, Monique Verstegen, Sebastian Trietsch, Henriëtte Lanz, Paul Vulto, Jos Joore, Karla S. Queiroz. Sorafenib and Lenvatinib induce vascular responses in patient derived HCC on-Chip models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 5876.

bioRxiv (Cold Spring Harbor Laboratory), Jun 10, 2020

Journal of Hepatology, Jul 1, 2022

Cancers, Nov 1, 2019

Primary liver cancer, consisting predominantly of hepatocellular carcinoma (HCC) and cholangiocar... more Primary liver cancer, consisting predominantly of hepatocellular carcinoma (HCC) and cholangiocarcinoma (CCA), remains one of the most lethal malignancies worldwide. This high malignancy is related to the complex and dynamic interactions between tumour cells, stromal cells and the extracellular environment. Novel in vitro models that can recapitulate the tumour are essential in increasing our understanding of liver cancer. Herein, primary liver cancer-derived organoids have opened up new avenues due to their patient-specificity, self-organizing ability and potential recapitulation of many of the tumour properties. Organoids are solely of epithelial origin, but incorporation into co-culture models can enable the investigation of the cellular component of the tumour microenvironment. However, the extracellular component also plays a vital role in cancer progression and representation is lacking within current in vitro models. In this review, organoid technology is discussed in the context of liver cancer models through comparisons to other cell culture systems. In addition, the role of the tumour extracellular environment in primary liver cancer will be highlighted with an emphasis on its importance in in vitro modelling. Converging novel organoid-based models with models incorporating the native tumour microenvironment could lead to experimental models that can better recapitulate liver tumours in vivo.

Stem Cells and Development, Sep 15, 2017

Liver transplantation is the only effective treatment for end-stage liver disease, but absolute d... more Liver transplantation is the only effective treatment for end-stage liver disease, but absolute donor shortage remains a limiting factor. Recent advances in tissue engineering focus on generation of native extracellular matrix (ECM) by decellularized complete livers in animal models. Although proof of concept has been reported for human livers, this study aims to perform whole liver decellularization in a clinically relevant series using controlled machine perfusion. In this study, we describe a mild nondestructive decellularization protocol, effective in 11 discarded human whole liver grafts to generate constructs that reliably maintain hepatic architecture and ECM components using machine perfusion, while completely removing cellular DNA and RNA. The decellularization process preserved the ultrastructural ECM components confirmed by histology, electron microscopy, and proteomic analysis. Anatomical characteristics of the native microvascular network and biliary drainage of the liver were confirmed by contrast computed tomography scanning. Decellularized vascular matrix remained suitable for normal suturing and no major histocompatibility complex molecules were detected, suggesting absence of allo-reactivity when used for transplantation. After extensive washing, decellularized scaffolds were nontoxic for cells after reseeding human mesenchymal stromal or umbilical vein endothelial endothelium cells. Indeed, evidence of effective recellularization of the vascular lining was obtained. In conclusion, we established an effective method to generate clinically applicable liver scaffolds from human discarded whole liver grafts and show proof of concept that reseeding of normal human cells in the scaffold is feasible. This supports new opportunities for bioengineering of transplantable grafts in the future.

Excessive alcohol consumption increases the risk of developing liver cancer, but the mechanism th... more Excessive alcohol consumption increases the risk of developing liver cancer, but the mechanism through which alcohol drives carcinogenesis is as yet unknown. Here, we determined the mutational consequences of chronic alcohol use on the genome of human liver stem cells prior to cancer development. No change in base substitution rate or spectrum could be detected. Analysis of the trunk mutations in an alcohol-related liver tumor by multi-site whole-genome sequencing confirms the absence of specific alcohol-induced mutational signatures driving the development of liver cancer. However, we did identify an enrichment of nonsynonymous base substitutions in cancer genes in stem cells of the cirrhotic livers, such as recurrent nonsense mutations in PTPRK that disturb Epidermal Growth Factor (EGF)signaling. Our results thus suggest that chronic alcohol use does not contribute to carcinogenesis through altered mutagenicity, but instead induces microenvironment changes which provide a 'fertile ground' for selection of cells with oncogenic mutations. INTRODUCTION Alcohol consumption is an important risk factor for the development of various cancer types, including hepatocellular carcinoma (HCC), and causes an estimated 400,000 cancer-related deaths worldwide each year 1-3. In spite of the clear link between alcohol intake and tumorigenesis, the underlying mechanism remains debated and mainly revolves around two hypotheses. The first hypothesis suggests that alcohol consumption may contribute to the development of cancer through an increased mutation accumulation in the genome 4. Consistently, the first metabolite of ethanol, acetaldehyde, is highly carcinogenic 5-7 and can also contribute to the formation of mutagenic reactive oxygen species (ROS) 8-11. Analysis of a large number of tumor exomes and genomes showed that alcohol intake is associated with an increased mutation load and different mutational characteristics 12-15. The second hypothesis .

Transplantation, Jul 1, 2014

(dsDNA-EC), both with Lyovec as a transfection reagent. Optimal concentration and incubation time... more (dsDNA-EC), both with Lyovec as a transfection reagent. Optimal concentration and incubation time were established. mRNA was isolated and via PCR, RT-qPCR and gel electrophoreses expression was determined. In resting primary tubular epithelial cells, expression of DAI and AIM2 could not be detected while these were clearly up-regulated upon stimulation with both dsDNA-EC and poly(I:C).[fi gure 1] DDX41, MRE11, LLRFIP1, and Ku70 were present but no altered regulation took place after stimulation. cGAS was only up-regulated upon poly(I:C) simulation. After stimulation with dsDNA-EC or poly(I:C), up-regulation of IFI16, ABCF1, and STING took place. We conclude that primary tubular epithelial cells do express several cytosolic dsDNA receptors, which can be triggered by synthetic PRR ligands. How BK virus can evade these functioning receptors remains to be elucidated.

British Journal of Haematology, Aug 21, 2003

A single dose (0AE3 lg) of recombinant human thrombopoietin (TPO) was injected into sublethal irr... more A single dose (0AE3 lg) of recombinant human thrombopoietin (TPO) was injected into sublethal irradiated non-obese diabetic/severe combined immunodeficient (NOD/SCID) mice immediately after transplantation of 1AE5 • 10 5 purified CD34 + umbilical cord blood (UCB) cells. Bone marrow (BM) was analysed for human cells by immunophenotyping and colony culture at d 35. TPO treatment produced a two-to sixfold increase in the frequency and number of human CD45 + cells. The lineage distributions among the human cells were similar irrespective of TPO treatment; however, a prominent increase was observed in CD71 + GpAcells, reflecting the proliferative stimulus provided by TPO. The frequency of immature CD34 + cells and human granulocyte-macrophage colony-forming units and erythroid burst-forming units in TPO-treated mice was similar to that of untreated mice, but their absolute numbers had increased proportionally to the increase in human cells. The results demonstrate that human TPO is a major limiting factor for multilineage outgrowth of human UCB cells in NOD/SCID mice and can be conveniently supplemented by single-dose treatment immediately after transplantation. TPO did not affect the survival of mice after transplantation and did not significantly increase the number of immature CD34 + CD38cells; secondary transplantation revealed that TPO administration also had no significant effect on longterm repopulation. The findings demonstrate that human TPO is required for proper outgrowth of human haematopoietic stem cells after transplantation. In addition, a single administration of TPO may improve the efficiency and reproducibility of the NOD/SCID mouse assay for human immature transplantable progenitor cells.

Springer eBooks, 2015

Hepatitis C virus (HCV) is a leading cause of chronic hepatitis and infects approximately three t... more Hepatitis C virus (HCV) is a leading cause of chronic hepatitis and infects approximately three to four million people per year, about 170 million infected people in total, making it one of the major global health problems. In a minority of cases HCV is cleared spontaneously, but in most of the infected individuals infection progresses to a chronic state associated with high risk to develop liver cirrhosis, hepatocellular cancer, or liver failure. The treatment of HCV infection has evolved over the years. Interferon (IFN)-α in combination with ribavirin has been used for decades as standard therapy. More recently, a new standard-of-care treatment has been approved based on a triple combination with either HCV protease inhibitor telaprevir or boceprevir. In addition, various options for all-oral, IFN-free regimens are currently being evaluated. Despite substantial improvement of sustained virological response rates, some intrinsic limitations of these new direct-acting antivirals, including serious side effects, the risk of resistance development and high cost, urge the development of alternative or additional therapeutic strategies. Gene therapy represents a feasible alternative treatment. Small RNA technology, including RNA interference (RNAi) techniques and antisense approaches, is one of the potentially promising ways to investigate viral and host cell factors that are involved in HCV infection and replication. With this, newly developed gene therapy regimens will be provided to treat HCV. In this chapter, a comprehensive overview guides you through the current developments and applications of RNAi and microRNA-based gene therapy strategies in HCV treatment.

Journal of Molecular Medicine, Jan 20, 2022

Fatty liver disease has grown into a major global health burden, attributed to multi-factors incl... more Fatty liver disease has grown into a major global health burden, attributed to multi-factors including sedentary lifestyle, obesogenic diet and prevalence of metabolic disorders. The lack of robust experimental models is hampering the research and therapeutic development for fatty liver disease. This study aims to develop an organoid-based 3D culture model to recapitulate key features of fatty liver disease focusing on intracellular lipid accumulation and metabolic dysregulation. We used human liver-derived intrahepatic cholangiocyte organoids and hepatocyte differentiated organoids. These organoids were exposed to lactate, pyruvate, and octanoic acid (LPO) for inducing lipid accumulation and mitochondrial impairment. Lipid accumulation resulted in alternations of gene transcription with major effects on metabolic pathways, including triglyceride and glucose level increase, which is consistent with metabolic changes in fatty liver disease patients. Interestingly, lipid accumulation affected mitochondria as shown by morphological transitions, alternations in expression of mitochondrial encoded genes, and reduction of ATP production. Meanwhile, we found treatment with obeticholic acid and metformin can alleviate fat accumulation in organoids. This study demonstrated that LPO exposure can induce lipid accumulation and associated metabolic dysregulation in human liver-derived organoids. This provides an innovative model for studying fatty liver disease and testing potential therapeutics.