Stem Cells

481 Pages·19.561 MB·English

by  C.S. Potten (Eds.)

Contributors
.E Arciniegas
CRC Department of Medical Oncology, Christie Hospital NHS Trust, Wilmslow Road,
Manchester M20 9BX, UK
P. W. Barlow
Department of Agricultural ,secneicS University of Bristol Institute of Arable Crops ,hcraeseR
Long Ashton hcraeseR Station, Bristol BS18 9AF, UK
R. Barraclough
recnaC and Polio Research Fund ,seirotarobaL Department of Biochemistry, University of
,loopreviL OP xoB 147, Liverpool 96L 3BX, UK
A-M. Buckle
aimeakueL hcraeseR Fund, Cellular Development Unit, Department of Biochemistry and Applied
Molecular Biology, UMIST, Sackville Street, Manchester M60 1QD, UK
P. Cameron-Curry
draozoH Hughes Medical Institute, 107 West 168th Street, lOth floor, New York, NY
,23001 USA
F. M. F. van Dissel-Emiliani
Departnlent of Functional Morphology, Veterinary School, University of Utrecht, Postbus
80.15 ,7 3508 TD Utrecht, ehT Netherlands
C. Dulac
Institut d'Embryologie Cellulaire et Moleculaire, UMRC9924, 49bis, Avenue de al Belle
Gabrielle, 94736 Nogent-sur-Marne Cedex, France
Present address: Harvard University, Cambridge, MA, USA
D. G. Fernig
recnaC dna Polio Research Fund ,seirotarobaL Department of Biochemistry, University of
,loopreviL OP xoB 147, Liverpool 96L 3BX, UK
D. Francis
loohcS of eruP and Applied Biology, University of Wales egelloC Cardiff, PO xoB 915, Cardiff
C IF 3TL, UK
.J W. Grisham
Department of Pathology, BC # 7525, University of North Carolina at Chapel Hill, 303
ttilluB-suahkz~irB Building, Chapel Hill, NC 27599-7525, USA
C. M. Heyworth
CRC Department of latnemirepxE Haematology, nosretaP Institute for Cancer ,hcraeseR Christie
Hospital NHS Trust, Wilmslow Road, Manchester M20 9BX, UK
G. .E Jones
ehT Randall Institute, King's egelloC London, 26-29 Drury Lane, London WC2B 5RL, UK
D. B. Kohn
Division of hcraeseR Immunology and Bone Marrow Transplantation, Children's Hospital soL
,selegnA Departments of scirtaideP and Microbiology, University of Southern California School
of Medicine, 4650 Sunset ,draveluoB soL Angeles, CA 90027, USA
viii Contributors
.J Kummermehr
Institute of Radiobiology, GSF- murtnezsgnuhcsroF ff~r Umwelt und Gesundheit, Institut ffir
,eigoloibnelhartS Ingolstiidter .rtsdnaL ,1 D-8042 Neuherberg, Germany
R. M. Lavker
Department of Dermatology, University of Pennsylvania School of Medicine, Philadelphia, PA,
USA
M. Loeffler
t~ftitsnI fur Medizinische Informatik, Statist& dnu ,eigolomedipE t~ffisrevinU ,gizpieL essartsgibeiL
2 ,7 04103 Leipzig, Germany
B. I. Lord
CRC Department of Experimental Haematology, nosretaP Institute for Cancer ,hcraeseR Christie
Hospital NHS Trust, Wilmslow Road, Manchester M20 9BX, UK
S. J. Miller
Department of Dermatology, Johns Hopkins Medical School, 60I North Carolina Street,
,eromitlaB MD 21287-0900, USA
.J A. Nolta
Division of hcraeseR Immunology and Bone Marrow Transplantation, Children's Hospital soL
Angeles, Departments of scirtaideP and Microbiology, University of Southern California loohcS
of Medicine, 4650 Sunset ,draveluoB soL Angeles, CA 90027, USA
C. S. Potten
CRC Department of lailehtipE Biology, nosretaP Institute for Cancer ,hcraeseR Christie Hospital
NHS Trust, Wilmslow Road, Manchester M20 9BX, UK
D. G. de Rooij
Department of Cell Biology, Medical School, University of Utrecht, Postbus 80.157, 3508 TD
,thcertU ehT Netherlands
P. S. Rudland
Cancer and Polio Research Fund ,seirotarobaL Department of Biochemistry, University of
,loopreviL PO xoB 147, Liverpool L69 3BX, UK
A. M. Schor
Department of Dental Surgery and ,ygolotnodoireP ehT Dental School, Park ,ecalP University
of Dundee, Dundee DD1 4HR, UK
S. L. Schor
Department of Dental Surgery and ,ygolotnodoireP ehT Dental School, Park ,ecalP University
of Dundee, Dundee DD1 4HR, UK
.J A. Smith
Cancer and Polio Research Fund ,seirotarobaL Department of Biochemistry, University of
,loopreviL PO xoB 147, Liverpool L69 3 BX, UK
T-T. Sun
lailehtipE Biology Unit, Ronald .O Perelman Department of Dermatology and Department of
,ygolocamrahP Kaplan Comprehensive Cancer Center, New York University Medical ,loohcS
New York, NY, USA
N. G. Testa
CRC Department of latnemirepxE Haematology, nosretaP Institute for Cancer ,hcraeseR Christie
Hospital NHS Trust, Wilmslow Road, Manchester M20 9BX, UK
S. S. Thorgeirsson
Laboratory of Experimental ,sisenegonicraC National Cancer Institute/National Institutes of
Health, Building ,73 Room 3C ,82 Bethesda, MD 20892-0001, USA
Contributors ix
K-R. Trott
tnemtrapeD of Radiation ,ygoloiB St s'wemolohtraB Hospital Medical ,egelloC esuohretrahC
,erauqS nodnoL ECIM 6BQ, UK
D. .J Watt
tnemtrapeD of Anatomy, Charing ssorC dna Westminster Medical ,loohcS Fulham ecalaP ,daoR
nodnoL W6 8RF, UK
A. D. Whetton
aimeakueL hcraeseR ,dnuF ralulleC tnempoleveD Unit, Department of yrtsimehcoiB dna Applied
Molecular ,ygoloiB UMIST, Sackville ,teertS Manchester M60 1QD, UK
N. A. Wright
tnemtrapeD of ,ygolohtapotsiH Royal etaudargtsoP Medical School Hammersmith Hospital
uD Cane Road, London W12 ONN, UK
Preface
In I983 I edited a volume, published by Churchill Livingstone, consisting of ten chapters
on stem cells in a variety of invertebrate and vertebrate systems, entitled Stem Cells-
their identification and characterisation. The book was successful and quickly went out of
print. I decided in 1993 that perhaps it was time to produce a new, updated book on
the subject perhaps covering fields that were not dealt with in I983. There have been
a number of significant developments since that time. Little was known then, for example,
about the complex interacting network of cytokines and growth factors that regulate
the proliferation and differentiation of stem cells. Much of the complexity of these signals
si now well understood in systems such as the mammalian bone marrow. Furthermore
the use of a variety of newly developed molecular biological techniques and probes has
made possible dramatic advances in our ability to study the lineage development
that si derived as a consequence of divisions of the stem cells. Also in the last few years
there have been significant increases of interest in identifying, characterizing and isolating
stem cell populations from a variety of tissues. Major granting bodies have targeted
stem cells and stem cell concepts amongst the topics identified as high priority for
funding. A major impetus here has been the desire to identify these all-important cells
ni tissues so that strategies for targeting these permanent lineage ancestor cells for gene
therapy may be developed. Also there has been an increased interest, arising from the
fact that many of the regulatory growth factors and cytokines have been identified, in
manipulating the tissue stem cells in various clinical situations including, for example,
the normal tissue stem cells during cancer therapy.
When I decided to put together this book I contacted lla the original contributors to ask
whether they felt there were developments that warranted a new review in this 1996
book. Most responded, either indicating that they thought there were new topics to
review or suggesting alternative authors and topics, some did not - which I interpreted
to indicate a lack of new developments in that particular field. I have also endeavoured
to cover some topics in this new book not covered in the previous one, examples of
which are obvious from the contents page. I was particularly interested in trying to
solicit a chapter describing what si known about precursor cells or stem cells in insect
systems. The extensive work on the genetics and morphogenic signals determining tissue
development of Drosophila suggested that the concept of stem cells in this system would
be interesting. Somewhat surprisingly to me, those working in the field tend not
to think of these developmental processes in terms of cell lineages and lineage ancestor
or stem cells but rather in terms of morphogens and topography (cell tissue interfaces
or boundaries) determining development and cell fate. Unfortunately, in spite of
contacting successively about ten authors in the field, I could not persuade anyone to
xii Preface
put together a chapter. I am not sure how one interprets this reticence. There were a
few other fields where I encountered similar difficulties.
I have managed to recruit altogether 51 chapters for this new book. Inevitably, it has
taken a considerable time to achieve this and to acquire the promised chapters from the
various authors. This was exacerbated by the consecutive chain of prospective authors
that I sometimes had to contact as indicated above. To those authors who did respond
quickly and were diligent in submitting their chapters by the initial deadlines, I offer my
sincere apologies for the length of time that it has taken to collect the remaining chapters
and produce the book. I think the only mitigating factor si that inevitably the book is
more valuable and comprehensive as a consequence of waiting for important contribu-
tions. Most of the initial contributors have had the opportunity to update their chapters
in the intervening period.
It si still not possible to identify stem cells in most tissues by either their morphological
characteristics or by the use of a specific marker. Numerous attempts have been made
to find stem cell specific markers, but the problem here may be that what characterizes
these cells si more likely to be the absence of specific features than the presence
of something that can be identified by a marker or probe. In the absence of such abilities,
finding these elusive cells si somewhat analogous to the needle in the haystack problem.
However, there are approaches that can be adopted to enrich these cells. In several
systems one of the major problems in studying stem cells si the fact that the techniques
available involve perturbation of the system, which inevitably causes changes in the
behaviour and characteristics (and possibly the number) of the stem cells which are to
be studied, resulting in a situation analogous to the Heisenberg uncertainty principle in
the field of subatomic particle and quantum physics.
There si still considerable variability or context dependence in the definition of stem
cells and the operation of the definition to identify and study stem cells amongst different
investigators. Although stem cells inherently and by definition are proliferative cells, this
si a weak and ineffective criterion to use on its own. The ability to divide a large number
of times and maintain a tissue throughout the life of an animal si a stronger criterion
for defining these cells but difficult to study directly and as a consequence usually
inferred indirectly. There are two further criteria which are virtually inherent in the point
just raised. Firstly, that if a stem cell si to divide a large number of times and maintain
a tissue, it must consequently also maintain its own numbers. This property of
self-maintenance si again difficult to study experimentally, usually requiring serial
transplantation or subcloning procedures. It si a cardinal property of stem cells that they
maintain their numbers and that the self-maintenance probability si subject to control
and hence variation. The second feature arising from the ability of stem cells to maintain
the cellularity of the tissue, results from the fact that tissues commonly contain a variety
of cell types, which implies that the stem cells are capable of generating cells that
differentiate down a variety of lineages. This si clearly demonstrable in some of the
more extensively studied tissues, such as bone marrow and the gastrointestinal tract. It
si a topic that si further discussed in several chapters dealing with other tissues in this
book.
Several questions concerning stem cells remain unresolved, for example to what extent
their attributes are inherently determined and to what extent they are governed by the
environment in which the cell finds itself. It is quite clear that environmental (niche)
determinants play an important role in the functional capabilities of stem cells. However,
Preface xiii
it equally seems likely that there are intrinsic differences between stem cells and cells
later in the proliferative lineage; thus it si probable that both intrinsic and extrinsic factors
are involved. In spite of many years of discussion and investigation it si still not clear how
differentiation of stem cells si achieved and what controls this process, i.e. symmetric
versus asymmetric divisions, stochastic versus deterministic processes, environmental
versus inherent signals. Another question that si commonly raised si to what extent the
differentiation and proliferative capacities (life span) of a stem cell are limited. Bone
marrow stem cells can clearly be demonstrated to have a division potential far in excess
of that required for day-to-day replacement of haematopoietic cells. Intestinal stem cells
seem likely to have a division potential of about 1000 in the mouse- a large number
by any criteria. It si surprising that with their large numbers, extensive division potential
and short cell cycle, stem cells in the small intestine rarely develop tumours. This must
indicate efficient damage detection and protection mechanisms. Bone marrow stem cells
seem to have an extraordinarily broad repertoire of differentiation. Whether further
unknown elements of their repertoire could be unmasked provided the correct signals
were given remains unanswered, but si perhaps unlikely bearing in mind the efforts that
have gone into studying this question. This topic also has important implications in
terms of understanding the cellular elements involved in ageing. The role that
stem cells play in the development of cancers si also an important area, subject to much
debate and comment. In tissues such as the gastrointestinal tract with its highly mobile
and dynamic cell lineages with their short life expectancy in the tissue, the stem cells
would seem to be the only candidates present in the tissue long enough to play a role
at least in the initiation stage in the development of cancer. Once the stem cell si initiated
it would produce a lineage of similarly altered cells, and it si conceivable that subsequent
stages in carcinogenic transformation may involve cells other than stem cells. However,
even here in the gastrointestinal tract, the probability would be, based on the rapid
turnover and the time it would take for a transformed cell to grow into a micro-turnout
that turnouts are unlikely to arise in any other cells than the stem cells or very early
lineage cells. These and many other topics are raised and discussed in the various
chapters to be found in this volume.
I should like to thank lla the contributors for their chapters and for the patience shown
by those who submitted their chapters some considerable time ago. It would not be
possible to put together such a book without dedicated skilled secretarial help, and for
that I am very grateful. I should like to thank the publishers for help in producing this
volume and finally I should like to thank my wife and family for their support and
understanding over the years.
1 Stem cells and cellular pedigrees-a conceptual
introduction
Markus Loeffler and Christopher S. Potten*
Institute of Medical Informatics, Statistics and Epidemiology, Leipzig,
Germany: *CRC Department of Epithelial Cell Biology, Paterson Institute
for Cancer ,hcraeseR Manchester, UK
SUMMARY
In this chapter, we consider some of the problems involved in current discussions on
stem cells in adult mammalian tissues. The present concepts involve a number of pitfalls,
logical, semantic and classification problems. This indicates the necessity for new and
well defined concepts that are amenable to experimental analysis.
One of the major difficulties in considering stem cells si that they are defined in terms
of their functional capabilities which can only be assessed by testing the abilities of the
cells, which itself may alter their characteristics during the assay procedure; a situation
similar to the uncertainty principle in physics. Hence, a proper description requires the
measurement i.e. manipulation process itself to be taken into account.
If such context-dependent interactions exist between the manipulation and measure-
ment process and the challenged stem cells, the question of the number of stem cells in
a tissue has to be posed in a new way. Rather than obtaining a single number, one might
end up with different numbers under different circumstances, all being complementary.
This might suggest that sternness si not a property but a spectrum of capabilities from
which to choose. This concept might facilitate a reconciliation between the different and
sometimes opposing experimental results. Given certain experimental evidence, we have
attempted to provide a novel concept to describe structured cell populations in tissues
involving stem cells, transit cells and mature cells. It si based on the primary
assumption that the proliferation and differentiation/maturation processes are in principle
independent in the sense that each may proceed without necessarily affecting the other.
Stem cells may divide without maturation, while cells approaching functional
competence may mature but do not divide. In contrast, transit cells divide and mature
showing intermediate properties between stem cells and mature functional cells. The
need to describe this transition process and the variable coupling between proliferation
and maturation leads us to formulate a screw model of cell and tissue organization.
This concept si illustrated for the intestinal epithelium. Reference si made also to other
tissues including the basal epidermal cell layer and the haematopoietic system.
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2 Markus Loeffler and Christopher .S Potten
INTRODUCTION
At present there si no experimental way to decide if a given cell in a functional mammalian
tissue is a stem cell or not. There are also no morphological criteria to identify such
cells. This si partly due to lack of appropriate experimental techniques, and partly due
to some conceptual problems. At the present stage it si helpful to discuss sternness as
a latent variable which cannot directly be observed and which can only be deduced
retrospectively on the basis of some indirect evidence based on measurable observable
parameters in specific experimental settings. If one accepts sternness as a hidden property,
it becomes obvious that one has to talk about stem cells and stem cell properties within
the framework of concepts and models. This clearly implies that there cannot be a canonic
unique definition but that a wide variety of models and concepts can be imagined, and
have in fact to be proposed to describe various features of stem cell systems.
Furthermore, it si evident that such models will differ in their attitude, their methodology
and the set of phenomena on which they focus. At present there si no generally accepted
standard stem cell model.
This introductory chapter is designed to serve as a framework for the stem cell models
discussed in subsequent chapters. Starting with some general definitions, we try to set
up criteria that should be fulfilled for stem cells. We point out the conceptual distinction
between stem cells and transit cells before we discuss the problem of obtaining
measurements on stem cells. We further discuss the basic elements of models used to
describe cellular hierarchies and stem cells. In undertaking this exercise we will highlight
our present reflections on this topic but also try to relate these to concepts suggested
by other authors. In this respect we will extend ideas discussed in a previous
paper (Potten and Loeffler, 1990).
DEFINITIONS
General definitions and concepts
In order to understand the full meaning and implications of the definition of stem cells, we
need to consider some subsidiary definitions. The most important ones are associated with
differentiation and maturation.
Differentiation
Differentiation can be defined as a qualitative change in the cellular phenotype that is the
consequence of the onset of synthesis of new gene products, i.e. the non-cyclic (new)
changes in gene expression that lead ultimately to functional competence (see Lajtha,
1979c). It may be recognized by a change in the morphology of the cell or by the
appearance of changes in enzyme activity or protein composition. Since it is a qualitative
change, a cell can be said to be differentiated only relative to another cell, and during
its life a cell may be capable of undergoing several differentiation events. Differentiation
si commonly identified by the detection of a novel protein. The ability to define a cell
Stem cells and cellular pedigrees- a conceptual introduction 3
as differentiated thus clearly depends on the sensitivity of the detection procedures. A
few molecules of a novel protein may be detectable, as may the changes in the
messenger RNA responsible for these molecules, but ultimately the differentiation event
involves a change in the repression/activation of the genome, i.e. in transcription, and
this may approximate to a quantal phenomenon. According to this definition cells
developing from a primitive stage to functional competence may undergo many, even
a series of, differentiation events each linked to a novel change in the gene activation
pattern. In many circumstances, it may be practically helpful to consider only some
primary key (marker) genes as relevant indicators of differentiation, particularly if
secondary genes are activated subsequently.
Maturation
Maturation in contrast can be regarded as a quantitative change in the cellular phenotype
or the cellular constituent proteins leading to functional competence (see Lajtha, 1979c).
Thus the degree of maturation, in principle, could be measured on a quantitative scale,
e.g. of the amount of a specific protein per cell. A differentiated cell matures with the
passage of time to form a functionally competent cell for that particular tissue. Its passage
through time and space could in principle be mapped, as new differentiation events occur
changing the path of the cell. This relationship is illustrated in Figure .1
The terms differentiation and maturation are often used in a loose fashion, inter-
changeably, with a consequent potential for confusion. It is also common to see the term
terminal differentiation used without an adequate definition of its meaning, which is
presumably an implicit indication of either the activation of the last differentiation event
in the cell's life history (e.g. involucrin synthesis for epidermal keratinocytes) or more
likely an indication of terminal maturation, i.e. the accumulation of differentiated
Figure 1 The course of an individual cell can be described in a differentiation-maturation
diagram. Acquisition of a qualitatively new marker si defined as differentiation (A), while the
trajectory for a given marker (from 0 to A) or, for a set of markers (from A to n) si defined
as maturation. Different maturation/differentiation paths may lead to the same state (n).
4 Markus Loeffler dna Christopher .S Potten
product(s) consistent with the final functional role of the cell (e.g. terminal keratinization,
or cornification for epidermal keratinocytes).
Proliferation
Proliferation si a process involving a sequential pattern of (cyclic, repeating) changes in
gene expression leading ultimately to the physical division of the cells. This is in contrast
with cell growth, which involves an increase in cell size or mass. In order to identify a
proliferating cell these changes have to be detected, and sensitivity problems similar to
those associated with differentiation are encountered. The changes may be represented
by discrete step-wise changes in the cellular concentration ,fo or by sharp peak alterations
,ni proliferation gene products. Many of these changes can be, and indeed have been,
mapped on a time scale represented by the interval in time between two subsequent cell
divisions, i.e. mapped in relation to the cell cycle. A large number of the gene products
of these proliferation-associated genes (which include many cellular oncogenes S-phase
and mitotic enzymes, cyclins etc.) have been mapped as transition points in the cell cycle.
Traditionally the four major transition points, the onset and termination of DNA
synthesis and mitosis have been used to identify proliferative cells, but many other
transition points may be equally valid.
There are certain difficulties in distinguishing cells on the basis of our definitions of
differentiation and proliferation. The first thing to note about these two processes si that
they are not necessarily mutually exclusive. Indeed many cells in the adult body may
exhibit differentiation markers, and hence be differentiated relative to cells earlier in tissue
development, and yet they also proliferate. Certainly many cells in bone marrow exhibit
both properties. Haematopoietic stem cells in the bone marrow are differentiated relative
to embryonic stem cells. The stem cells in surface epithelia may be differentiated relative
to the bone marrow stem cells and vice versa. The characterizations of the state of
proliferation and differentiation are dependent upon the ability to identify changing
patterns in gene expression and gene products. If these changes are of a cyclical nature
they may be associated with proliferation. However, the cells under consideration may
divide only once or we may have no knowledge of their previous history, in which case
we are unable to tell if a particular gene product has been produced cyclically. Hence,
it si more useful to define proliferation on the basis of the appearance of gene
products associated with DNA replication, or the cell division process, which are in fact
produced in a cyclic fashion. This implies a knowledge of many or lla the metabolic
processes associated with, and leading to cell division.
The distinction between differentiation, maturation and proliferation appears important
as the development from stem cells to functionally competent cells can be viewed as a
transition from one extreme (prolif: yes; diff/mat: no) to the opposite extreme (prolif: no;
diff/mat: yes). The transition takes place through states of coexistence with some
flexibility to accelerate or slow down one or both processes. It si this flexibility that
permits cells to be stimulated to differentiate and stop proliferation and vice versa. Below
we will introduce the assumption that proliferation, differentiation and maturation are
not strictly coupled and in many circumstances should be considered independent of
each other.
A special consideration here is to what extent a differentiated cell can dedifferentiate,
whether this involves a switching off of the already activated differentiation genes (this