Stem Cells and Extracellular Matrices

Transcription

1 Colloquium series on stem Cell Biology Series Editor: Wenbin Deng Stem Cells and Extracellular Matrices Lakshmi Kiran Chelluri life sciences Morgan & Claypool life SCIEnCES

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3 Stem Cells and Extracellular Matrices

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5 Colloquium Series on Stem Cell Biology Editors Wenbin Deng, Ph.D., Department of Cell Biology and Human Anatomy, Institute for Pediatric Regenerative Medicine, School of Medicine, University of California, Davis This series is interested in covering the fundamental mechanisms of stem cell pluripotency and differentiation, and the strategies of translating fundamental developmental insights into discovery of new therapies. The emphasis is on the roles and potential advantages of stem cells in developing, sustaining and restoring tissue after injury or disease. Some of the topics included will be the signaling mechanisms of development and disease; the fundamentals of stem cell growth and differentiation; the utilities of adult (somatic) stem cells, induced pluripotent stem (ips) cells and human embryonic stem (ES) cells for disease modeling and drug discovery; and the prospects for applying the unique aspects of stem cells for regenerative medicine. We hope this series will provide the most accessible and current discussions of the key points and concepts in the field, and that students and researchers all over the world will find these in-depth reviews to be useful. Published titles (for future titles please see the website,

6 Copyright 2012 by Morgan & Claypool Life Sciences All rights reserved. No part of this publication may be reproduced, stored in a retrieval system, or transmitted in any form or by any means electronic, mechanical, photocopy, recording, or any other except for brief quotations in printed reviews, without the prior permission of the publisher. Stem Cells and Extracellular Matrices Lakshmi Kiran Chelluri ISBN: paperback ISBN: ebook DOI: /C00053ED1V01Y201204SCB001 A Publication in the Colloquium Series on Stem Cell Biology Lecture #1 Series Editor: Wenbin Deng, University of California Series ISSN Pending

7 Stem Cells and Extracellular Matrices Lakshmi Kiran Chelluri Department of Transplant Biology, Immunology & Stem Cell Lab Global Hospitals, Lakdi-ka-Pool, Hyderabad (A.P) India COLLOQUIUM SERIES ON STEM CELL BIOLOGY #1

8 vi Abstract Stem cells have great potential in regenerative medicine and tissue injury. Regulation of stem cell homeostasis in a 3D microenvironment is controlled by the niche components that influence stem cell fate, regulation, and function. It is therefore necessary to understand the mechanisms of cell cell interaction, molecular cross talk between stem cells and their extracellular matrix (ECM) environment. The adhesion molecules play a pivotal role in establishing the cell cell contact and subsequent integration with the ECM. This understanding is the basis for establishing design criteria for biomimetic. The integrated approach by biologists, material science engineers, biomedical engineers, and clinicians is the key in the development of tissue engineered constructs for effective translation to clinics. Keywords stem cells, extracellular matrices, tissue microenvironment, cell-based therapy, bio-mimetic, tissue engineering, biopolymers, cell cell interaction, integrins, tissue constructs, stem cell fate

9 vii Contents Abbreviations... ix 1. Introduction to Stem Cell Biology and Niche Components Basic Stem Cell Biology Types of Stem Cells Stem Cells and Differentiation The Living Environment of Stem Cells Extracellular Matrix (ECM) Soluble Factors Cell Cell Interaction Extracellular Matrix Proteins Forces ECM Structure and Organization Constituents of ECM Collagen A Structural Protein of ECM Elastin Fibers Proteoglycans and GAGs in the ECM Hyaluronan Adhesive Glycoproteins Basement Membrane Extract and Specialized Proteins Silk Design Strategies for Cell Matrix Engineering Biomechanical Attributes of an ECM Matrix Reactions and Interactions Matrix Designing for 3 Dimensional Stem Cell Cultures ECM Control, Regulation on Stem Cell Fate and Function Stem Cell Fate Regulators ECM as a Regulator of Stem Cell Fate... 25

10 viii Stem Cells and Extracellular Matrices 3.2 Stem Cell Topography in Regulation of the ECM Signal Transduction Mechanisms in Stem Cell Fate Control of ECM in Stem Cell-based Tissue Engineering ECM and Stem Cell Cultures Embryonic Stem Cell Cultures in Natural ECM Embryonic Stem Cell Cultures in Synthetic ECM Hematopoietic Stem Cells in Natural and Synthetic ECM Undifferentiated and Differentiated MSCs in ECM MSCs to Osteogenic Differentiation in ECM Chondrogenic Differentiation of MSCs in ECM Neural Stem Cells in ECM Endothelial Progenitor Cells in ECM Adipose-Tissue-Derived Stem Cells in ECM Challenges of Stem Cells in ECM Conclusion Acknowledgments Bibliography Author Biography Titles of Related Interest... 58

11 ix Abbreviations BMP Bone Morphogenic Protein BSMC Bladder Smooth Muscle Cells EB Embryoid Bodies ECM Extracellular Matrix EGF Epidermal Growth Factor ESC Embryonic Stem Cells Gj Gap Junction hesc human Embryonic Stem Cell MEF Mouse Embryo Fibroblast mes mouse Embryonic Stem Cell MSC Mesenchymal Stem Cells NGF Nerve Growth Factor PCL Poly Caprolactone PEG Poly Ethylene Glycol PLA Polylactic Acid PLGA poly (lactic-co-glycolic acid) PMMA Polymethyl Methacrylate RGD Arg Gly Asp Shh Sonic hedgehog TGF-β Transforming Growth Factor-β TIP Tension Induced Proteins

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13 1 c h a p t e r 1 Introduction to Stem Cell Biology and Niche Components 1.1 BASIC STEM CELL BIOLOGY Stem cells are defined based on their inherent properties of both self-renewal and differentiation (Weissman, 2000). They reside in a distinct environment necessary for their maintenance and survival. Such a milieu is defined as the microenvironment or the stem cell niche. Post tissue injuries, stem cells that are intrinsic to the tissue, replace necrotic cells as a first line of defense. When the pool of endogenous stem cells is exhausted, exogenous circulating stem cells are signaled to replenish the pool and participate in tissue repair. Circulating stem cells which serve as essential internal repair system undergoes countless division to replenish worn out or damaged cells in the body (Li et al., 2005a). The signals that emanate from these primitive cells are dependent on the native tissue and type to express the boundaries of their niche in order to rescue and repair (Figure 1). FIGURE 1: The stem cell niche is controlled by various factors such as mitotic division rate, type of division such as symmetric vs. asymmetric and the orientation of the axes of mitosis. The effector mechanism is influenced by the extrinsic and intrinsic factors. These factors have impact on the maintenance of the contact with the extracellular matrix via integrins.

14 2 Stem Cells and Extracellular Matrices Types of Stem Cells Cells that are primitive in origin have been shown to possess a developmental potential to differentiate into cells representing all three embryonic germ layers viz., endoderm, mesoderm, and ectoderm. These primitive cells are found in almost all the tissues of the body. They are broadly termed as adult stem cells, embryonic stem cells, and induced pluripotent stem cells (ipscs). They can be further classified to totipotent, pluripotent, and multipotent cells. Totipotent cells have the capability of differentiating to all types of cells. Pluripotent cells can differentiate to all three germ layers. Multipotential cells can differentiate into many cell types such as skeletal, muscle, cardiac, and liver cells. All blood cells are classified as hematopoietic cells. Mesenchymal progenitor cells are quadripotential cells which have the capability of becoming a cartilage, fat, bone forming, and stromal cell types. It was surmised that stem cell system of brain is incapable of regeneration and the existing neurons are terminally differentiated cells. With technology advancing at a rapid stride, it was observed that some regions of adult brain do exhibit neurogenesis. This observation is based on the identification of progenitor cells responsible for both embryonic and postnatal neural development (Alvarez-Buylla and Lois, 1995; Gage et al., 1996; Weiss et al., 1996). The existence of stem cells in olfactory epithelium, gonads, gut has demonstrated mosaic in vivo-lineage leading to presumption that their properties could be harnessed and utilized in degenerating, ageing tissues (Figure 2). FIGURE 2: Stem cells are of three types, i.e., totipotent, pluripotent, multipotent. They have the propensity to form three germ layers of ectoderm, mesoderm, and endoderm. The progenitor cells of mesoderm can form cell types of ectoderm and endoderm.

15 Introduction to Stem Cell Biology and Niche Components Stem Cells and Differentiation Unspecialized stem cells giving rise to specialized cells undergo the process called differentiation. Differentiation pathway of the cell goes through several stages to become more specialized at each step. This differentiation process is controlled by both intrinsic and extrinsic factors. The intrinsic signals come from within the cell s own genes and extrinsic signals are provided by the other cells, and their physical contact with neighboring cells and certain molecules which constitutes the stem cell niche (Knoblich, 2008). These complex interactions of signals during differentiation process cause the cells to acquire innate changes in their DNA make-up, by switching the responses of the newly acquired phenotype. Such a mechanism is termed as epigenetic changing mechanisms. These restrict DNA expression in the cell that which can be passed on through cell division to the progeny. The structural proteins of the niche including the components of ECM cytoskeleton help in unequal partitioning of cell fate determinants (Broadus and Doe, 1997). A number of transcriptional factors which make up the stem cell niche control the fate of the daughter cells produced from the stem cell division. During differentiation, the proliferative potential of stem cell is lost, as specialized differentiated features are acquired. Two major models have been proposed to explain the differentiation potential of stem cells; the hierarchical model (Koeffler et al., 1981) and the continuum model (Chomarat et al., 2003). According to the hierarchical model, stem cells generate differentiated cells through a unidirectional series of cell fate transitions progressing from the stem cell through progenitor cells and finally to precursor cells. The most primitive stem cell has high proliferative potential but reduced capacity to differentiate, whereas precursor cells have little or no proliferation potential but differentiate readily. With every pace in the transition of stem cell to precursor cell, proliferative potential diminishes while the capacity to differentiate augments. In this model stem cells are quiescent and remain in the G0 phase of the cell cycle until they enter the G1 phase either arbitrarily or under environmental influences such as growth factors, cytokines, adhesion molecules or cell cell contact provided by stem cell niche. Once stimulated to proliferate, the stem cells can either progress through the progenitor and precursor stages or revert to quiescent state until a new stimulus arises. However, progenitor and precursors cells once formed cannot regress to stem cell state (Koeffler et al., 1981) (Figure 3). In continuum model, stem cells are continuously and reversibly altering their state and phenotype of stem cells and can shift from one state to another. They can revert back too, to the original phenotype which is termed as de-differentiation. This change in state is supposed to be caused by chromatin remodeling associated with cell cycle changes, which consecutively alters the surface phenotype of stem cells reversibly, thus determining the way the stem cells respond to environmental stimuli. Quiescent stem cells may be stimulated to enter the G1 phase and thereafter either proceed through the pathway of differentiation or revert to quiescent state (Chomarat et al., 2003).

16 4 Stem Cells and Extracellular Matrices FIGURE 3: Hierarchical model of stem cell division. Stem cell(s) can divide asymmetrically to maintain their number while giving rise to transit amplifying cells (TA). They reduce over a period of time. These TA cells terminally differentiate (TD) and are programmed to die in a tissue-specific time span. FIGURE 4: Stem cells are continuously and reversibly altering their state and phenotype in an inverse correlation between stem cells and progenitor cells. During engraftment, the stem cells decrease and the progenitors increase.

17 Introduction to Stem Cell Biology and Niche Components 5 Interestingly, cells that have proceeded to progenitor stages can still revert to stem cell state, thus opening up the possibility for one stem cell to generate cells of different lineages depending on the microenvironmental stimuli (Figure 4). Stem cells can be induced to differentiate once removed from the mouse embryo fibroblast (MEF) feeders and introduced with bioactive signals. This is done either directly or through the FIGURE 5: Schematic diagram of the induced differentiation protocol used for myeloid cell generation from hescs and hipscs.

18 Stem Cells and Extracellular Matrices formation of embryoid bodies (EBs). EBs are small clumps of stem cell colonies grown in suspension which form three dimensional (3D) spheroid bodies representing a differentiation model with the widest spectrum of cell types that can be achieved in in vitro conditions. Differentiating cells within the EBs enjoy cell cell interaction and paracrine effects of the three embryonic germ layers, in developing 3D microenvironment that mimics to the temporal and spatial processes taking place in the developing embryo. Although differentiation is possible, most differentiation systems rely on EB formation. Once cells start to differentiate, a variety of bioactive manipulations can be applied to control and direct the differentiation process. In general, these include (Figure 5): Soluble signals, such as growth factors, hormones, and cell conditioned media; Genetic modifications, such as over-expression of transcription factors known to derive stem cells into the desired cell type; Direct or indirect co-culturing with other developing or mature somatic cell populations; Physical stimuli, such as mechanical forces, temperature, and oxygenation changes. Whether differentiation is allowed to occur spontaneously or in a directed manner, the resultant cell population in most known protocols is still heterogeneous. 1.2 THE LIVING ENVIRONMENT OF STEM CELLS EXTRACELLULAR MATRIX (ECM) Extracellular matrix is simply the living environment of the cells. It is a network of proteins and carbohydrates that binds cells together. It supports and surrounds cells regulating the cell activities and provides lattice structure/framework for cell movement. There are essentially five classes of macromolecules that makeup the stem cell framework, i.e., collagens, elastic fibers, proteoglycans, hyaluronan, and adhesive glycoproteins. These can be mixed up in different proportions for different functions. The ECM influences cell development, movement, and differentiation (Tanentzapf et al., 2007; Hayashi et al., 2007). It is a reservoir of extracellular signaling molecules and coordinates cellular functions through signaling with specific cellular adhesion receptors such as integrins. The cell adhesion receptors transmit mechanical stimuli from the extracellular matrix to the cytoskeleton. Abundant ECMs are present at the time when primitive cells start differentiating and migrating toward terminally differentiated cell, wherein the ECM proteins start declining with the progression of the differentiation process (Letourneau et al., 1994; Lathia et al., 2007) (Figure 6). The natural living environment of cells comprises of four niche components which dictates the ultimate fate and function of stem cells. They include (a) soluble factors, (b) direct cell cell contact, (c) extracellular matrices, and (d) forces. Extensive work is in progress to develop synthetic living microenvironment keeping the tissue architecture in mind and the regulatory constraints in using xeno-free reagent base.

19 Introduction to Stem Cell Biology and Niche Components 7 FIGURE 6: Schematic representation of the constituents of the ECM networking with the plasma membrane and the cell interaction mechanisms through the integrins Soluble Factors Soluble factors, such as epithelial growth factor (EGF), nerve growth factor (NGF), basic fibroblast growth factor (bfgf), bone morphogenic proteins (BMPs) and Wnts, aid in the immobilization of cells by direct anchoring onto the cellular membranes (Rider et al., 2006) and increasing the local protein concentration. This attribute has been conveniently employed in tissue engineering in the development of the stem cell niches. Immobilization of growth factors aids via the mechanism of hindering diffusion and receptor-mediated endocytosis. It is pertinent and critical to use these soluble factors for immobilization onto the matrix-binding domain in order to mediate and regulate the downstream signaling events more effectively and in a sustained fashion. This concept was directly applied by using EGF onto a biomaterial surface such as poly-methyl methacrylate (PMMA/ hydrogels) to achieve high protein concentration and sustained signaling in cultures of primary rat hepatocytes (Kuhl and Griffith-Cima, 1996). Further, there is an enhanced differentiation potential