Stem Cells – From Hype to Real Hope

Khawaja Husnain Haider, Salim Aziz, MD

• Publisher: De Gruyter
• Year: 2018
• ISBN:
  9783110586916
• Edition: 1
• Topic: Medicine
• Subtopic: Pharmacology

This book is a compilation of the bench experience of leading experts from various research labs involved in the cutting edge area of research. The authors describe the use of stem cells both as part of the combinatorial therapeutic intervention approach and as tools (disease model) during drug development, highlighting the shift from a conventional symptomatic treatment strategy to addressing the root cause of the disease process.

The book is a continuum of the previously published book entitled "Stem Cells: from Drug to Drug Discovery" which was published in 2017.

Valeria A. Tsvelaya, Anna Gam, Jenna Aziz and Igor R. Efimov

1 Induced pluripotent stem-cell-derived cardiomyocytes (iPSC-CMs): novel diagnostic platform

Abstract: There is growing concern regarding the clinical relevance of therapy testing in immortalized cell lines and animal models. The advent of induced pluripotent stem cells allows conduction of early diagnostics and potential therapy testing in cell derived from a specific patient retaining his/her genetic background. This is especially important in studies of various heritable cardiomyopathies, which are difficult to recapitulate in animal models. A number of methods have been developed building upon the pioneering work of Yamanaka laboratory, aiming at reprogramming various somatic cells to cardiomyocytes and creating two- and three-dimensional models for investigation of cardiac physiology in a particular genetic background. Conventional models, such as short-lived primary cell preparations, monolayer cell cultures, coronary-perfused hearts and preparations, and, more recently, heart slices have unique advantages and limitations, especially in studies of long-term chronic conditions. The emerging induced pluripotent stem-cell-derived cardiomyocyte (iPSC-CM) platform holds significant potential to overcome these limitations while allowing patient-specific diagnostics and therapy development. This chapter compares conventional models with iPSC-CM, reviews cardiogenesis as the basis for reprogramming protocol development, focuses on the iPSC-CM disease modeling platforms for drug testing and patient specific theranostics, and concludes with clinical studies on transplantation of in vitro derived CMs.

Key words: Cardiomyocytes, Cardiogenesis, Differentiation, Drug discovery, Engineered, Heart, iPSCs, Model

1.1 Rationale for the emerging need for iPSC-CMs

Cardiomyocytes (CMs) are the major cell type that determines cardiac function. CMs have low proliferation and turnover rates, which make them hard to generate, grow to maturity, and study in vitro. In addition to those constraints, there is limited availability of both healthy and diseased human CMs, which makes it hard to construct in vitro models of the diseases, for drug testing, and design methods for in vivo applications [1, 2]. Other models that can be derived from human donor hearts, such as wedge preparations and cardiac slices, have limited availability and low throughput and often lack the desired genetic background. Animal models have significant species-dependent differences; for example, mouse heart rate is 10 times faster, which results in 5–10 times shorter QT-interval compared with human hearts. Murine CMs express species-specific ion channels and structural proteins. Additionally, increased heart rate in mice correlates with decreased contraction force, which is opposite to humans [1, 3]. Similar variability across various species renders animal models not entirely suitable for drug testing since the results do not always correlate with the drug response in humans. Additionally, animal cells and tissues cannot be safely used for implantation into humans due to immunogenic response. These constraints stimulated the research in human stem cells, which in turn led to the development of induced pluripotent stem cells (iPSCs). Pluripotent stem cells (PSCs) can be either embryonic stem cells (ESCs) or iPSCs. The first human iPSC-CMs (hiPSC-CMs) were reported in 2009 by Zhang et al. [4, 5]. Here, mainly iPSCs are discussed since the goal of this chapter is to describe the-state-of-art research with a focus on personalized medicine. Table 1.1.1 summarizes the current models used to study cardiac functions and highlights the advantages and limitations of each model.

Tab. 1.1.1: Comparison of models to study human cardiac function.

Model	Advantages	Limitations
Single cell	– Study of single ion channels	– No cell-cell interactions – Low viability
Cell monolayer (cell line)	– Cell-cell interactions – High viability – High reproducibility	– No 3D interactions – Change of phenotype over time
Primary cells	– Person specific/precision medicine	– Low viability – Limited availability
iPSC monolayer (can be incorporated in EHT, CMT, and biowires)	– Relatively easy to obtain – Potentially large quantities – Patient specific – Retain genetic background – Enables studying of heritable diseases – Enables early diagnostics	– Challenging to purify – Difficult to classify – Hard to obtain 100% of desired phenotype – Full maturity has not been reported
Slices	– 3D morphology – Accurate model for acute studies – Genetic modification (e.g., siRNA transfection)	– Limited availability – Cannot perform chronic studies
Wedges	– 3D physiology studies	– Low throughput – Limited availability

CMT= cardiac microtissue; iPSC= induced pluripotent stem cell; EHT= engineered heart tissue; 3D= three dimensional.

1.2 Cardiogenesis and iPSC differentiation

In order to differentiate stem cells into CMs, it is necessary to know how this process naturally occurs in vivo. To this day, the biggest challenge of the field is the inability to obtain mature CMs of a single phenotype. Therefore, here, the different stages of cardiogenesis are reviewed in parallel with what has been recapitulated in vitro to highlight the missing steps that could improve current reprogramming practices and address the aforementioned challenges.

The heart consists of several cell types, including CMs, fibroblasts, neurons, and vascular cells. CMs separate further into four lineages: atrial, ventricular, Purkinje, and nodal cells, each having unique electrophysiology, mechanics, and structure. Since during iPSC differentiation, it is unclear which CMs lineage is obtained, it is necessary to tune the differentiation protocol for a specific CMs lineage that recapitulates its embryonic developmental process.

Developmentally, cells destined for cardiogenesis arrive from a mesodermal germ layer, which are modified by transcriptional signaling molecules such as wingless/integrated (Wnt), fibroblast growth factor (FGF), transforming growth factor beta (TGFβ), bone morphogenic protein 4 (BMP4), activin, and nodal [6]. The gradients of these molecules either promote or inhibit different pathways based on spatiotemporal and functional cues. Wnt/β-catenin signaling contributes to the formation of cardiac progenitor cells, and its inhibition by adjacent cells through frizzled-related protein 2 (FRZP2) and Dickkopf 1 (DKK1) allows for further differentiation and specification into progenitor cells [6–9]. The heart is formed by contribution from two types of progenitor cells: first heart field (FHF) and second heart field (SHF). The cardiac field is specified by downstream transcription events that lead to the expression of cardiac-specific transcription factors [6, 9]. The FHF forms the primitive heart tube, which later transforms to the left ventricle (LV) and parts of both atria. The SHF forms the right ventricle (RV) and outflow tract and contributes to both atria as well [6, 10]. FG...

Table of contents

• Cover
• Title Page
• Copyright
• Dedication
• Preface
• Contents
• Contributing authors
• List of abbreviations
• 1 Induced pluripotent stem-cell-derived cardiomyocytes (iPSC-CMs): novel diagnostic platform
• 2 Preclinical large-animal models of cardiovascular regeneration
• 3 Cell-free therapy with stem cell secretions: protection, repair, and regeneration of the injured myocardium
• 4 Myoblasts provide safe and effective treatments for hereditary muscular dystrophies, cardiomyopathies, type 2 diabetes, solid tumors, and aging
• 5 Stem cells in ophthalmology
• 6 Stem cells for ocular therapies
• 7 Cell therapy for liver regeneration
• 8 Patient-specific induced pluripotent stem cells for cardiac disease modeling
• 9 Role of stem cells on evolution: a hypothesis
• Index