iPSCs for Studying Infectious Diseases

The series Advances in Stem Cell Biology is a timely and expansive collection of comprehensive information and new discoveries in the field of stem cell biology. iPSCs for Studying Infectious Diseases, Volume 8 addresses how important induced pluripotent stems cells are and how can they can help treat certain infectious diseases. Somatic cells can be reprogrammed into induced pluripotent stem cells by the expression of specific transcription factors. These cells have been transforming biomedical research over the last 15 years. This volume will address the advances in research of how induced pluripotent stem cells are being used for treatment of different infectious diseases, such as corona virus, coxsackievirus, salmonella infection, influenza virus and much more. The volume is written for researchers and scientists in stem cell therapy, cell biology, regenerative medicine and organ transplantation; and is contributed by world-renowned authors in the field.

• Provides overview of the fast-moving field of stem cell biology and function, regenerative medicine, and therapeutics
• Covers infections by several pathogens, such as coronavirus, coxsackievirus, influenza virus, herpes simplex virus 1, T. gondii, T. cruzi, S. agalactiae, N. meningitidis, Salmonella, and more
• Is contributed by world-renowned experts in the field

• Language: English
• Publisher: Academic Press
• Release date: May 29, 2021
• ISBN: 9780128241899

Book preview

iPSCs for Studying Infectious Diseases, Volume 8

Editor

Alexander Birbrair

Table of Contents

Cover image

Title page

Advances in Stem Cell Biology

Copyright

Dedication

Contributors

About the editor

Preface

Chapter 1. The application of iPSCs to questions in virology: a historical perspective

A brief history of virology

Viruses as obligate parasites

The advent of cell biology

Stem cells, embryonic stem cells, and induced pluripotent stem cells

Current applications of iPSCs to virology

The family Caliciviridae

The family Coronaviridae

The family Flaviviridae

The family Hepadnaviridae

The family Hepeviridae

The family Herpesviridae

The family Orthomyxoviridae

The family Paramyxoviridae

The family Picornaviridae

The family Polyomaviridae

The family Retroviridae

The family Togaviridae

Future directions

Chapter 2. Transplantation of iPSC-derived neural progenitor cells promotes clinical recovery and repair in response to murine coronavirus-induced neurologic disease

Introduction

Conclusions

Chapter 3. iPSCs for modeling influenza infection

Introduction

IAV-induced cell death in iPSCs

Differentiation potentials of IAV-infected iPSCs

iPSC-derived tissues and organoids for modeling influenza infection

Concluding remarks

Chapter 4. Human induced pluripotent stem cells for modeling of herpes simplex virus 1 infections

Introduction

Concluding remarks

Future directions

Chapter 5. iPSCs for modeling coxsackievirus infection

Biology of coxsackieviruses

Coxsackievirus-associated disease in humans

Experimental models for coxsackievirus infection

iPSC modeling of coxsackievirus infection

Concluding remarks and future perspectives

Chapter 6. Pluripotent stem-cell-derived oligodendrocyte progenitors to model demyelination caused by Theiler’s murine encephalomyelitis virus and other viruses

Importance of myelin in the CNS

Virus-induced demyelination

Steps to myelination: OPC proliferation, migration, and maturation

Disruption of myelination by viruses

Induced pluripotent stem cells (iPSCs) as a model system to study demyelinating viruses

Conclusions and future perspectives

Chapter 7. iPSCs for modeling hepatotropic pathogen infections

The liver is a target organ for many pathogens

Hepatitis viruses

Plasmodium

Addressing open questions in hepatotropic infection research with HLCs

Systems integrating diverse hepatic cell types to improve liver pathogenesis studies

3D systems to study hepatotropic infections

Personalized modeling and treatment of hepatotropic infections

Limitations of iPSCs and future directions for the study of hepatotropic infections

Outlook

Chapter 8. Use of human induced pluripotent stem cells (hiPSC)-derived neuronal models to study the neuropathogenesis of the protozoan parasite, Toxoplasma gondii

Introduction

Overview of principles and methods for generation of neurons from hiPSCs

T. gondii: biology of chronic infection in the brain

Future trends and direction: use of hiPSC-derived 2D and 3D models to model human parasitic infections

Chapter 9. Induced pluripotent stem cells for modeling Chagas disease

Cardiomyopathy

Chagas disease

Chagas disease pathogenesis

Clinical Chagas disease

Immune response in Chagas disease

Role of therapy in Chagas disease and relation to immune response

New approaches to therapy

Models to study Chagas disease

iPSC for modeling Chagas disease

Chapter 10. Induced pluripotent stem-cell derived brain-like endothelial cells to study host–pathogen interactions with the bacterial pathogens Streptococcus agalactiae and Neisseria meningitidis

Introduction

Bacterial meningitis

Brain endothelial cell models and infection

Current iPSC models

Group B Streptococcus

Bacterial interaction with iPSC-BECs

Current iPSC-BEC models and future outlook

iPSC based models and infections

Chapter 11. Human induced pluripotent stem cells for modeling of Salmonella infection

Introduction

iPSCs: bridging the gap between human and animal research

Establishing iPSC-derived cellular systems as a model for Salmonella infection

Differentiation of iPSCs to other Salmonella infection-relevant cell types

Using iPSCs to investigate the role of host genotype on Salmonella response phenotype

Using iPSCs for modeling the molecular consequences of human genetic variants

Future trends and directions

Conclusions

Index

Advances in Stem Cell Biology

Series Editor

Alexander Birbrair

Copyright

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Notices

Knowledge and best practice in this field are constantly changing. As new research and experience broaden our understanding, changes in research methods, professional practices, or medical treatment may become necessary.

Practitioners and researchers must always rely on their own experience and knowledge in evaluating and using any information, methods, compounds, or experiments described herein. In using such information or methods they should be mindful of their own safety and the safety of others, including parties for whom they have a professional responsibility.

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Dedication

This book is dedicated to my mother, Marina Sobolevsky, of blessed memory, who passed away during the creation of this volume. Professor of Mathematics at the State University of Ceará (UECE), she was loved by her colleagues and students, whom she inspired by her unique manner of teaching. All success in my career and personal life I owe to her.

My father Lev Birbrair and my beloved mom Marina Sobolevsky of blessed memory (July 28, 1959–June 3, 2020)

Contributors

Serkan Belkaya,     Department of Molecular Biology and Genetics, Faculty of Science, Bilkent University, Çankaya, Ankara, Turkey

David C. Bloom,     Department of Molecular Genetics & Microbiology, University of Florida College of Medicine, Gainesville, FL, United States

Guglielmo Bove,     Schaller Research Group, Center of Infectious Diseases, Department of Virology, Heidelberg University Hospital, Heidelberg, Germany

Adriana Bozzi

Instituto René Rachou, FIOCRUZ, Belo Horizonte, Brazil

Departamento de Ciências Biológicas, Universidade Estadual de Santa Cruz, UESC, Ilhéus, Brazil

Kevin M. Coombs

University of Manitoba, Department of Medical Microbiology and Infectious Diseases, Winnipeg, MB, Canada

Manitoba Centre for Proteomics & Systems Biology, Winnipeg, MB, Canada

Children’s Hospital Research Institute of Manitoba, Winnipeg, MB, Canada

Viet Loan Dao Thi,     Schaller Research Group, Center of Infectious Diseases, Department of Virology, Heidelberg University Hospital, Heidelberg, Germany

Matthew J. Demers,     Department of Psychiatry, Western Psychiatric Institute and Clinic, University of Pittsburgh School of Medicine, Pittsburgh, PA, United States

Leonardo D’Aiuto,     Department of Psychiatry, Western Psychiatric Institute and Clinic, University of Pittsburgh School of Medicine, Pittsburgh, PA, United States

Jessica L. Forbester

Division of Infection and Immunity/Systems Immunity University Research Institute, Cardiff University, Cardiff, United Kingdom

MRC Weatherall Institute of Molecular Medicine, University of Oxford, John Radcliffe Hospital, Oxford, United Kingdom

Eric C. Freundt,     Department of Biology, The University of Tampa, Tampa, FL, United States

Sandra K. Halonen,     Department of Microbiology and Immunology, Montana State University, Bozeman, MT, United States

Brandon J. Kim,     University of Alabama, Department of Biological Sciences, Tuscaloosa, AL, United States

Paul R. Kinchington

Department of Ophthalmology, University of Pittsburgh School of Medicine, Pittsburgh, PA, United States

Department of Molecular Microbiology and Genetics, University of Pittsburgh, Pittsburgh, PA, United States

Thomas E. Lane,     Department of Neurobiology & Behavior, University of California, Irvine, Irvine, CA, United States

Jeanne F. Loring,     Department of Molecular Medicine, The Scripps Research Institute, La Jolla, CA, United States

Laura L. McIntyre,     Department of Molecular Biology & Biochemistry, University of California, Irvine, Irvine, CA, United States

James McNulty,     Department of Chemistry and Chemical Biology, McMaster University, Hamilton, ON, Canada

Ann-Kathrin Mehnert,     Schaller Research Group, Center of Infectious Diseases, Department of Virology, Heidelberg University Hospital, Heidelberg, Germany

Vishwajit L. Nimgaonkar,     Department of Psychiatry, Western Psychiatric Institute and Clinic, University of Pittsburgh School of Medicine, Pittsburgh, PA, United States

Warren C. Plaisted,     Genomics Institute of the Novartis Research Foundation, San Diego, CA, United States

Pavan Rajanahalli,     Department of Biology, The University of Tampa, Tampa, FL, United States

Duncan R. Smith,     Institute of Molecular Biosciences, Mahidol University, Salaya, Nakhon Pathom, Thailand

David A. Stevens

California Institute for Medical Research, San Jose, CA, United States

Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, CA, United States

Craig M. Walsh,     Department of Molecular Biology & Biochemistry, University of California, Irvine, Irvine, CA, United States

Maribeth A. Wesesky,     Department of Psychiatry, Western Psychiatric Institute and Clinic, University of Pittsburgh School of Medicine, Pittsburgh, PA, United States

Ali Zahedi-Amiri

University of Manitoba, Department of Medical Microbiology and Infectious Diseases, Winnipeg, MB, Canada

Manitoba Centre for Proteomics & Systems Biology, Winnipeg, MB, Canada

Wenxiao Zheng

Department of Psychiatry, Western Psychiatric Institute and Clinic, University of Pittsburgh School of Medicine, Pittsburgh, PA, United States

Department of Psychiatry, The Second Xiangya Hospital, Xiangya School of Medicine, Central South University, Changsha, China

About the editor

Alexander Birbrair

Dr. Alexander Birbrair received his bachelor's biomedical degree from Santa Cruz State University in Brazil. He completed his PhD in Neuroscience, in the field of stem cell biology, at the Wake Forest School of Medicine under the mentorship of Osvaldo Delbono. Then, he joined as a postdoc in stem cell biology at Paul Frenette's laboratory at Albert Einstein School of Medicine in New York. In 2016, he was appointed faculty at Federal University of Minas Gerais in Brazil, where he started his own lab. His laboratory is interested in understanding how the cellular components of different tissues function and control disease progression. His group explores the roles of specific cell populations in the tissue microenvironment by using state-of-the-art techniques. His research is funded by the Serrapilheira Institute, CNPq, CAPES, and FAPEMIG. In 2018, Alexander was elected affiliate member of the Brazilian Academy of Sciences (ABC), and in 2019, he was elected member of the Global Young Academy (GYA). He is the Founding Editor and Editor-in-Chief of Current Tissue Microenvironment Reports and Associate Editor of Molecular Biotechnology. Alexander also serves in the editorial board of several other international journals: Stem Cell Reviews and Reports, Stem Cell Research, Stem Cells and Development, and Histology and Histopathology.

Preface

This book's initial title was iPSCs: Recent Advances. Nevertheless, because of the ongoing strong interest in this theme, we were capable of collecting more chapters than would fit in one single volume, covering induced pluripotent stem cells (iPSCs) biology from different perspectives. Therefore, the book was subdivided into several volumes.

This volume iPSCs for Studying Infectious Diseases offers contributions by known scientists and clinicians in the multidisciplinary areas of biological and medical research. The chapters bring up-to-date comprehensive overviews of current advances in the field. This book describes the use of iPSCs to model several infectious diseases in vitro, enabling us to study the cellular and molecular mechanisms involved in different infectious pathologies. Further insights into these mechanisms will have important implications for our understanding of infectious disease appearance, development, and progression. The authors focus on the modern state-of-the-art methodologies and the leading-edge concepts in the field of stem cell biology. In recent years, remarkable progress has been made in the obtention of iPSCs and their differentiation into several cell types, tissues, and organs using state-of-the-art techniques. These advantages facilitated identification of key targets and definition of the molecular basis of several disorders. Thus, the present book is an attempt to describe the most recent developments in the area of iPSCs biology, which is one of the rising hot topics in the field of molecular and cellular biology today. Here, we present a selected collection of detailed chapters on what we know so far about the use of iPSCs for modeling multiple infectious diseases. Eleven chapters written by experts in the field summarize the present knowledge about iPSCs for studying infectious diseases.

Duncan R. Smith from Mahidol University gives a historical perspective on the application of iPSCs in virology. Thomas E. Lane and colleagues from University of California discuss the use of iPSCs in coronavirus-induced neurologic disease. Ali Zahedi-Amiri and Kevin M. Coombs from University of Manitoba describe iPSCs for modeling influenza infection. Leonardo D'Aiuto and colleagues from University of Pittsburgh compile our understanding of iPSCs for modeling of herpes simplex virus 1 infections. Serkan Belkaya from Bilkent University updates us with what we know about iPSCs for modeling coxsackievirus infection. Eric C. Freundt and Pavan Rajanahalli from The University of Tampa summarize current knowledge on iPSCs to model Theiler's murine encephalomyelitis virus infection. Viet Loan Dao Thi and colleagues from Heidelberg University address the importance of iPSCs for modeling of hepatotropic pathogen infection. Sandra K. Halonen from Montana State University talks about the use of human iPSCs to study the neuropathogenesis of Toxoplasma gondii. Adriana Bozzi and David A Stevens from Stanford University focus on iPSCs for modeling Chagas disease. Brandon J Kim from the University of Alabama presents the use of iPSCs to study host–pathogen interactions with Streptococcus agalactiae and Neisseria meningitidis. Finally, Jessica L Forbester from the University of Oxford updates us on the use of iPSCs for modeling of Salmonella infection.

It is hoped that the articles published in this book will become a source of reference and inspiration for future research ideas. I would like to express my deep gratitude to my wife Veranika Ushakova, and Ms. Billie Jean Fernandez and Ms. Elisabeth Brown from Elsevier, who helped at every step of the execution of this project.

Alexander Birbrair

Editor

Chapter 1: The application of iPSCs to questions in virology

a historical perspective

Duncan R. Smith     Institute of Molecular Biosciences, Mahidol University, Salaya, Nakhon Pathom, Thailand

Abstract

Viruses are obligate parasites in that they can only replicate within a living host cell. Thus the science of virology is largely dependent upon the requirement to be able to grow and propagate such host cells. While it is relatively simple to be able to grow and maintain suitable host cells for viruses that infect prokaryotic cells, the situation is more complicated when eukaryotic host cells are required for viral propagation. Studies on eukaryotic viruses are thus often a compromise between the ease of propagation of the host cell and the fidelity of the propagated cells to the bona fide host cell. Until recently the choice was largely between primary cells (high fidelity, low ease of propagation) or immortalized and transformed cells (low fidelity, high ease of propagation). More recently, the discovery of induced pluripotent stem cells (iPSCs), which have high fidelity and relatively high ease of propagation, has introduced a third option. This chapter will present the historical context of the application of iPSCs to questions in virology and describe how these cells are currently being used.

Keywords

Caliciviridae; Cell culture; Coronaviridae; Flaviviridae; Hepadnaviridae; Hepeviridae; Herpesviridae; Induced pluripotent stem cells; Orthomyxoviridae; Paramyxoviridae; Picornaviridae; Polyomaviridae; Retroviridae; Togaviridae; Virology

A brief history of virology

Viruses as obligate parasites

The advent of cell biology

Stem cells, embryonic stem cells, and induced pluripotent stem cells

Current applications of iPSCs to virology

The family Caliciviridae

The family Coronaviridae

The family Flaviviridae

The family Hepadnaviridae

The family Hepeviridae

The family Herpesviridae

The family Orthomyxoviridae

The family Paramyxoviridae

The family Picornaviridae

The family Polyomaviridae

The family Retroviridae

The family Togaviridae

Future directions

Acknowledgment

References

A brief history of virology

The field of microbiology has existed for nearly 350 years and is considered to have formally started in 1676 when the Dutch scientist Antionie van Leeuwenhoek first observed microbial life using handmade microscopes. The field of virology as a distinct subfield of microbiology has had a much shorter history. The roots of virology lie in the work of Dimitri Ivanovsky (1846–1920), who, in 1892, demonstrated the presence of a causal agent of tobacco mosaic disease that was smaller than any previously described infectious particle. From this point on, viruses were largely defined as an infectious agent that would pass through a filter that retained bacteria and required living cells rather than culture medium for propagation. The physical nature of the infectious agent remained largely unknown until Kausche, Pfankuck, and Ruska observed discrete particles of tobacco mosaic virus using an electron microscope in 1939 (Kausche et al., 1939). Even in the absence of an understanding of the nature of viruses, a number of viruses had been identified as disease agents before 1931 including foot and mouth disease virus by Leoffler and Frosch in 1898, yellow fever virus by Walter Reed in 1900, and rabies virus by Remlinger and colleagues in 1903. Even more strikingly, vaccines had been developed for a number of diseases that we now know are viral in origin including the use of cowpox virus for vaccination against smallpox by Jenner in 1796 and a vaccination against rabies developed by Pasteur in 1885 (see Fig. 1.1).

Viruses as obligate parasites

One of the defining moments in virology was when Milton Rivers proposed that viruses are obligate parasites (Rivers, 1927). Although initially controversial, the proposal accounted for the fact that successful virus amplification had previously only been achieved in embryonated eggs or laboratory animals. Maitland and Maitland demonstrated propagation of vaccinia virus in minced chicken kidney in a mixture of chicken serum and Tyrode’s solution (Maitland and Maitland, 1928), although they believed that this did not constitute a cell culture system. However, Rivers, Haagen, and Muckenfuss showed the requirement for live cells using a similar system (Rivers et al., 1929). Li and Rivers subsequently established that the virus could grow in minced chicken embryo tissue in Tyrode’s solution (a chemically defined medium), removing the need for a plasma component (Li and Rivers, 1930).

Figure 1.1 A brief history of virology.The figure shows some of the key points on the path to defining virology as a distinct area of study.

The advent of cell biology

The use of minced animal tissues in defined media dominated much of virology in the 1940s and 1950. Importantly, Enders, Weller, and Robbins showed that poliovirus could be grown in cultured cells that were not nerve cells (Weller et al., 1949), and this was instrumental in developing the first polio vaccines, with the original injectable Salk inactivated vaccine (Salk et al., 1955) and the oral live attenuated Sabin vaccine (Sabin et al., 1960) to protect against poliomyelitis being produced in minced rhesus macaque monkey kidney cells. However, as eloquently stated by Tom Curtis, "By 1960, scientists and vaccine manufacturers knew that monkey kidneys were sewers of simian viruses (Curtis, 2004). In particular, it is estimated that millions around the world were exposed to polio vaccines contaminated with Simian virus 40 (SV40). Questions over the safety of polio vaccines led to a shift of production to African green money kidneys cells and finally to a vaccine produced in the well-characterized Vero cell line (Montagnon, 1989). The polio vaccine story highlighted the problem of using primary cells—the possible presence of endogenous viruses. A second major drawback of using primary cells is their relatively limited useful life span. Primary cells are not able to replicate indefinitely, and after a period in culture, the cells become senescent and eventually die and thus must be continually replaced with newly sourced tissues. The concept of a defined life span for cells was first promoted by Leonard Hayflick based on his work with normal human diploid cells. Hayflick proposed that normal somatic cells had an inherent replication capacity of 40   +   10 cells divisions, after which the cells become senescent and die (Hayflick, 1965). This intrinsic replication capacity is now termed the Hayflick limit," and in 2009, Blackburn, Greider, and Szostak shared the Nobel Prize in Physiology or Medicine for their work on telomeres and telomerase, an enzyme linked with the biological counting mechanism of cellular replication (Varela and Blasco, 2010).

The Hayflick limit was proposed to explain the behavior of normal diploid cells, as there were already cell lines that did not conform to this limit. The first bona fide cell line capable of continuous culture was the mouse strain L, generated by W.R. Earle in 1940 from mouse subcutaneous areolar and adipose tissue (Earle et al., 1943). A clone from this strain (L929) generated in 1948 from the 95th subculture was subsequently the first cloned cell line developed (Sanford et al., 1948). In the following years, a number of immortalized or transformed cell lines capable of continuous growth were produced, including HeLa (Scherer et al., 1953), CHO (Tjio and Puck, 1958), MDCK (the isolation of this line was not published by Madin and Darby, but it was subsequently used (Green, 1962) and characterized (Gaush et al., 1966) by others), and WI-38 (Hayflick, 1965), the last of which was developed by Hayflick himself.

Currently there are a large number of cell lines capable of continuous growth. A main central repository for cell lines, the American Type Culture collection (ATCC), maintains over 4000 cell lines. These cells are easy to propagate and expand and have thus driven virus research for the last 60 or more years. Cell lines are either immortalized or immortalized and transformed. Immortalized cells generally have achieved stable telomeres through the expression of telomerase activity (Bodnar et al., 1998), while transformed cells additionally have undergone neoplastic transformation. In this regard, as these cells have acquired properties not normally possessed by the corresponding primary cell, immortalized and transformed cells cannot be considered as normal cells. In particular, transformed cells often express proteins not normally found in the original cell type and conversely can fail to express proteins that are normally expressed (Pan et al., 2009).

The ability of a virus to productively infect a particular cell depends upon the susceptibility of the cell, as well as the permissiveness of the cell. Susceptibility indicates that a particular virus can enter into a cell, while permissiveness indicates that viral replication, packaging, and cellular egress can occur. In this regard, the deranged protein expression found in immortalized and transformed cells can lead to the derivation of susceptible and permissive cell lines from tissues that are not normally target tissues of infection. Conversely, cell lines derived from a known viral target tissue might be refractory to infection. Much of virology is therefore dependent upon less than satisfactory model systems in which virus/cell line pairings are based on utility, rather than being a reflection of true tropism. That said, it should be noted that a similar criticism applies to studies on human pathogenic viruses conducted in animals, in which the pathology may only poorly reflect the pathogenesis seen in humans (Ruiz et al., 2017).

Stem cells, embryonic stem cells, and induced pluripotent stem cells

A stem cell has the capacity to self-renew and to give rise to all of the differentiated cell types of the organism. This concept is almost as old as the field of virology. In his book Anthropogenie, published in 1874, Ernst Haeckel (1834–1919) proposed that a fertilized egg be called a "stammzelle" (or stem cell) (Haeckel, 1874). Around the same time, the field of hematopoiesis (the generation of the cells of the blood) was revolutionized after Paul Erlich (1845–1915) developed the methods to specifically stain different blood cell types (for a review of Erlich’s contributions to histochemistry, see (Buchwalow et al., 2015)). In particular, this work triggered a debate as to whether red and white blood cells had a common precursor. On the side of those who believed in a single precursor, Pappenheim (Pappenheim, 1896) used the term stem cell to describe the postulated precursor. In the following years, a number of studies pointed toward the existence of a blood stem cell. For example, Florence Sabin working with irradiated animals provided strong evidence of blood stem cells, but did not identify the cells specifically (Sabin et al., 1932). In 1963, Till and McCulloch published a study (Becker et al., 1963) that showed that one type of cell in the blood was capable of differentiating into three distinct lineages (erythrocytic, granulocytic, and megakaryocytic). While not directly using the term stem cell, the first identification of stem cells is commonly credited to them. However, hematopoietic stem cells are not totipotent (capable of differentiating into all cell types including extraembryonic tissues) or pluripotent (capable of differentiating into cells of the three germ layers), but they are multipotent (capable of differentiating into a number of related cell types). The first pluripotent stem cells were isolated and cultured by Evans and Kaufmann from mouse blastocysts (Evans and Kaufman, 1981), and the first human pluripotent stem cells were produced from human blastocysts in 1998 by James Thompson (Thomson et al., 1998). Human embryonic stem cells are produced from potentially viable human embryos, and as such their production and use remain controversial (Lo and Parham, 2009).

In 2006, Takahashi and Yamanaka provided a solution to the problems associated with the use of embryonic stem cells. Working with 24 genes identified as being important to embryonic cell function, they showed that the presence of four of these genes was sufficient to reprogram a mouse somatic cell to an embryonic stem cell-like phenotype (Takahashi and Yamanaka, 2006). These four factors, called the Yamanaka factors, consisted of Oct3/4, Sox2, Klf4, and c-Myc. These first-generation cells, however, were not fully pluripotent in that they could neither produce functional chimeras nor contribute to the germ line (Takahashi and Yamanaka, 2006). Improved methodologies published the following year by three groups (Maherali et al., 2007; Okita et al., 2007; Wernig et al., 2007) were able to generate fully pluripotent cells, termed induced pluripotent stem cells (iPSC), which were functionally identical to embryonic stem cells. In the same year, human iPSCs generated from somatic cells (fibroblasts) were reported from Yamanaka’s group using the same factors (Takahashi et al., 2007), as well as by the group of James Thompson using Oct4, Sox2, Nanog, and Lin28 (Yu et al., 2007). There is considerable research ongoing in developing iPSCs using different factors, cell types, and protocols, as well as the development of protocols to differentiate iPSCs into different cell types (Liu et al., 2020). However, crucially, the development and widespread use of iPSCs put these cells into the hands of virologists who, for the first time, were able to look at the cellular events ongoing during virus infection in a cell line that could be differentiated into a bona fide cell type (See Fig. 1.2).

Current applications of iPSCs to virology

A search of the relevant literature undertaken in late March 2020 identified more than 100 studies that used iPSCs to address questions in virology (Table 1.1). Not included in the analysis were studies that used viruses such as a lentivirus (Takenaka et al., 2010) or Sendai virus (Simara et al., 2014) to generate iPSCs, or those that use a virus as a tool to investigate non-infection-related questions (Naaman et al., 2018).

Collectively, the studies investigated 25 different viruses in 18 genera belonging to 12 virus families and utilized a number of different cell types (Fig. 1.3). More than half of the studies investigated only three viruses, namely Zika virus (ZIKV), Hepatitis C virus (HCV), and Hepatitis B virus (HBV). The family Flaviviridae accounted for over half of all studies, and ZIKV alone was the subject of a quarter of all studies.

Figure 1.2 The application of iPSCs to questions in virology.The figure shows the overall route through which iPSCs are reprogrammed from somatic cells and can be used in virology.

Table 1.1

The family Caliciviridae

The family Caliciviridae consists of 11 genera, Bavovirus, Lagovirus, Minovirus, Nacovirus, Nebovirus, Norovirus, Recovirus, Salovirus, Sapovirus, Valovirus, and Vesivirus (Vinje et al., 2019). The viruses in this family are nonenveloped with a single-stranded, positive sense RNA genome. In terms of human health, the genus Norovirus is the most important. This genus contains a single virus species, Norwalk virus, but noroviruses are genetically very diverse with multiple genogroups and genotypes (Atmar, 2010). Noroviruses are transmitted primarily by the fecal–oral route and can cause both endemic and epidemic gastroenteritis. Noroviruses have traditionally been very difficult to culture, and it was only recently that a methodology was established to culture noroviruses using stem-cell-derived epithelial cell cultures, with the stem cells being obtained from intestinal crypts from tissues obtained at biopsy or surgery (Ettayebi et al., 2016). To overcome the limitations of a culture system requiring adult stem cells, Sato and colleagues successfully derived intestinal epithelial cells from iPSCs (Sato et al., 2019). It is likely that iPSCs will result in rapid advances in our understanding of noroviruses given this significant advance.

Figure 1.3 The utilization of iPSCs.iPSCs and cells differentiated from them have been used in studies on a number of different viruses.

The family Coronaviridae

The family Coronaviridae has two subfamilies, the Letovirinae and the Orthocoronavirinae. The subfamily Orthocoronavirinae contains four genera, Alphacoronavirus, Betacoronavirus, Deltacoronavirus, and Gammacoronavirus (ICTV Master Species list 2018b.v2, available at talk.ictvonline.org/files/master-species-list/m/msl/8266). The viruses in this family of 39 species consist of enveloped viruses with a positive sense, single-stranded RNA genome. Members of genus Betacoronavirus include the Severe acute respiratory syndrome-related coronavirus, and both SARS and SARS-CoV2 belong to this species of virus (Coronaviridae Study Group, 2020). The genus Betacoronavirus also contains the species Murine coronavirus, to which mouse hepatitis virus, a common virus infecting laboratory mice, belongs. Mangale and colleagues undertook a comparative analysis of the susceptibility to mouse hepatitis virus of ex vivo derived neural precursor cells (NPC) and NPCs derived through differentiation of iPSCs (Mangale et al., 2017). The authors found that although the iPSC-NPCs were functionally equivalent, there was reduced susceptibility to the neurotropic mouse hepatitis virus. This potentially has implications in using NPCs to treat neurological disorders.

The family Flaviviridae

The family Flaviviridae consists of four genera, Flavivirus, Pestivirus, Hepacivirus, and Pegivirus (Simmonds et al., 2017), and collectively the family has more than 60 virus species assigned to it (Simmonds et al., 2017). The members of this family all have a single-stranded positive sense RNA as their genomic material, and the viruses are enveloped. The family includes a number of viral species that are significant human pathogens with broad distribution including yellow fever virus, dengue virus, West Nile virus, and Zika virus in the genus Flavivirus, and Hepatitis C virus in the genus Hepacivirus. Studies have been undertaken on five members of the genus Flavivirus, including