Principles of Tissue Engineering

Robert Lanza,Robert Langer,Joseph P. Vacanti

• Publisher: Academic Press
• Year: 2011
• ISBN:
  9780080548845
• Edition: 3
• Topic: Biological Sciences
• Subtopic: Biotechnology
• Index: Biological Sciences

First published in 1997, Principles of Tissue Engineering is the widely recognized definitive resource in the field. The third edition provides a much needed update of the rapid progress that has been achieved in the field, combining the prerequisites for a general understanding of tissue growth and development, the tools and theoretical information needed to design tissues and organs, as well as a presentation by the world's experts of what is currently known about each specific organ system. This edition includes greatly expanded focus on stem cells, including adult and embryonic stem cells and progenitor populations that may soon lead to new tissue engineering therapies for heart disease, diabetes, and a wide variety of other diseases that afflict humanity. This up-to-date coverage of stem cell biology and other emerging technologies is complemented by a series of new chapters on recent clinical experience in applying tissue engineering. The result is a comprehensive textbook that we believe will be useful to students and experts alike.New to this edition: *Includes new chapters on biomaterial-protein interactions, nanocomposite and three-dimensional scaffolds, skin substitutes, spinal cord, vision enhancement, and heart valves*Expanded coverage of adult and embryonic stem cells of the cardiovascular, hematopoietic, musculoskeletal, nervous, and other organ systems

Part I

The Basis of Growth and Differentiation

Chapter Five Molecular Biology of the Cell

Jonathan Slack

I. Introduction

II. The Cell Nucleus

III. The Cytoplasm

IV. Growth and Death

V. Cytoskeleton

VI. Cell Adhesion Molecules

VII. Extracellular Matrix

VIII. Culture Media

IX. Cells in Tissues

X. Further Reading

I INTRODUCTION

This chapter is a general introduction to the properties of animal or human cells. It deals with gene expression, metabolism, protein synthesis and secretion, membrane properties, response to extracellular factors, cell division, properties of the cytoskeleton, cell adhesion, and the extracellular matrix. It shows how these cellular properties underlie the specific conditions required for successful tissue culture. In particular cells require effective access to nutrients, removal of waste products, and their growth and behavior are controlled by a variety of extracellular hormones and growth factors present in the medium. The properties of individual cells are also the basis for understanding how cells can become organized into tissues, which are normally composed of more than one cell type and have a specific microarchitecture appropriate to their function.

To a naive observer the term tissue engineering might seem a contradiction. The word engineering conjures up a vision of making objects from hard components, such as metals, plastics, concrete, and silicon, that are mechanically robust and will withstand a range of environmental conditions. The components themselves are often relatively simple, and the complexity of a system emerges from the number and connectivity of the parts. By contrast, the cells of living organisms are themselves highly delicate and highly complex. Despite our knowledge of a vast amount of molecular biological detail concerning cell structure and function, their properties are still understood only in qualitative terms, and so any application using cells involves a lot of craft skill as well as rational design. What follows is a very brief account of cell properties intended for newcomers to tissue engineering who have an engineering or physical science background. It is intended to alert readers to some of the issues involved in working with cells and to pave the way for understanding how cells form tissues and organs, topics dealt with in more detail in the later chapters. Because it comprises very general material, it is not specifically referenced, although some further reading is provided at the end.

Cells are the basic building blocks of living organisms, in the sense that they can survive in isolation. Some organisms, such as bacteria, protozoa, and many algae, actually consist of single free-living cells. But most cells are constituents of multicellular organisms, which, though they can survive in isolation, need very carefully controlled conditions to do so. A typical animal cell suspended in liquid will be a sphere of the order of about 20 microns in diameter (Fig. 5.1).

FIG. 5.1 Structure of a generalized animal cell.

(From paternityexperts.com website.)

Most cells will not grow well in suspension, and so they are usually grown attached to a substrate, where they flatten and may be quite large in horizontal dimensions but only a few microns in vertical dimension. All eukaryotic cells contain a nucleus, in which is located the genetic material that ultimately controls everything the cell is composed of and all the activities it carries out. This is surrounded by cytoplasm, which has a very complex structure and contain substructures called organelles that are devoted to specific biochemical functions. The outer surface of the cell is the plasma membrane, which is of crucial importance as the frontier across which all materials must pass on their way in or out. The complexity of a single cell is awesome, since it will contain thousands of different types of protein molecules, arranged in many very complex, multimolecular aggregates comprising both hydrophobic and aqueous phases, and also many thousands of low-molecular-weight metabolites, including sugars, amino acids, nucleotides, fatty acids, and phospholipids, among many others. Although some individual steps of metabolism may be near to thermodynamic equilibrium, the cell as a whole is very far from equilibrium and is maintained in this condition by a continuous interchange of substances with the environment. Nutrients are chemically transformed, with release of energy that is used to maintain the structure of the cell and to synthesize the tens of thousands of different macromolecules on which its continued existence depends. Maintaining cells in a healthy state means to provide them continously with all the substances they need, in the right overall environment of substrate, temperature, and osmolarity, and also continuously to remove all potentially toxic waste products.

II THE CELL NUCLEUS

The nucleus contains the genes that control the life of the cell. A gene is a sequence of DNA that codes for a protein, or for a nontranslated RNA, and it is usually considered also to include the associated regulatory sequences as well as the coding region itself. The vast majority of eukaryotic genes are located in the nuclear chromosomes, although a few genes are also carried in the DNA of mitochondria and chloroplasts. The genes encoding nontranslated RNAs include those for ribosomal (transfer) RNAs and also a large number of microRNAs that are probably involved in controlling expression of protein-coding genes. The total number of protein-coding genes in vertebrate animals is about 30,000, and every nucleus contains all the genes, irreversible DNA modifications being confined to cells of the immune system in respect of the genes encoding antibodies and T-cell receptors.

The DNA is complexed into a higher-order structure called chromatin by the binding of basic proteins called histones. Protein-coding genes are transcribed into messenger RNA (mRNA) by the enzyme RNA polymerase II. Transcription commences at a transcription start sequence and finishes at a transcription termination sequence. Genes are usually divided into several exons, each of which codes for a part of the mature mRNA. The primary RNA transcript is extensively processed before it moves from the nucleus to the cytoplasm. It acquires a “cap” of methyl guanosine at the 5′ end and a polyA tail at the 3′ end both of which stabilize the message by protecting it from attack by exonucleases. The DNA sequences in between the exons are called introns, and the portions of the initial transcript complementary to the introns are removed by splicing reactions catalyzed by snRNPs (small nuclear ribonucleoprotein particles). It is possible for the same gene to produce several different mRNAs as a result of alternative splicing, whereby different combinations of exons are spliced together from the primary transcript. In the cytoplasm the mature mRNA is translated into a polypeptide by the ribosomes. The mRNA still contains a 5′ leader sequence and a 3′ untranslated sequence flanking the protein-coding region, and these untranslated regions may contain specific sequences responsible for translational control or intracellular localization.

Control of Gene Expression

There are many genes whose products are required in all tissues at all times, for example, those concerned with basic cell structure, protein synthesis, or metabolism. These are referred to as housekeeping genes. But there are many others whose products are specific to particular cell types, and indeed the various cell types differ from each other because they contain different repertoires of proteins. This means that the control of gene expression is central to tissue engineering. Control may be exerted at several points. Most common is control of transcription, and we often speak of genes being “on” or “off” in particular situations, meaning that they are or are not being transcribed. There are also many examples of translational regulation, where the mRNA exists in the cytoplasm but is not translated into protein until some condition is satisfied. Control may also be exerted at the stage of nuclear RNA processing or indirectly via the stability of individual mRNAs or proteins.

Control of transcription depends on regulatory sequences within the DNA and on proteins called transcription factors that interact with these sequences. The promoter region of a gene is the region just upstream from the transcription start site to which the RNA polymerase binds. The RNA polymerase is accompanied by a set of general transcription factors, which together make up a transcri...

Table of contents

• Cover image
• Title page
• Table of Contents
• Copyright
• CONTRIBUTORS
• FOREWORD
• PREFACE
• PREFACE TO THE SECOND EDITION
• PREFACE TO THE FIRST EDITION
• Introduction to Tissue Engineering
• Part I: The Basis of Growth and Differentiation
• Part II: In Vitro Control of Tissue Development
• Part III: In Vivo Synthesis of Tissues and Organs
• Part IV: Models for Tissue Engineering
• Part V: Biomaterials in Tissue Engineering
• Part VI: Transplantation of Engineered Cells and Tissues
• Part VII: Stem Cells
• Part VIII: Gene Therapy
• Part IX: Breast
• Part X: Cardiovascular System
• Part XI: Endocrinology and Metabolism
• Part XII: Gastrointestinal System
• Part XIII: Hematopoietic System
• Part XIV: Kidney and Genitourinary System
• Part XV: Musculoskeletal System
• Part XVI: Nervous System
• Part XVII: Ophthalmic Applications
• Part XVIII: Oral/Dental Applications
• Part XIX: Respiratory System
• Part XX: Skin
• Part XXI: Clinical Experience
• Part XXII: Regulation and Ethics
• Epilogue
• INDEX