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OCR for page 145
Mode] Systems for the Evaluation of Toxic
Agents Affecting Aging or Age-Related
Diseases
.
The development of mode! systems in the field of aging is at
about the same stage today as the development of mode! systems to
detect potentially mutagenic environmental chemicals was in 1970.
Screening tests had shown that potentially mutagenic chemicals
belonged to a wide variety of chemical classes, for example, foods,
drugs, cosmetics, agricultural chemicals, and household chern~cals.
In the first phase of the development of short-term screen-
ing tests, appropriate mode! systems were used to obtain infor-
mation on the various mechanisms that produce genetic lesions
and to identify the types of genotoxic agents that would produce
gene mutations, chromosomal aberrations, and aneuploidy. Those
models were almost exclusively in vitro assays that used organisms
ranging from bacteria to mammalian cells in culture.
The second phase was the identification of mammalian models
that could be used to evaluate chemicals that had positive results
in in vitro tests to determine the effects of these chemicals on
somatic celb and germ cells. The later tests were selected to
provide a comprehensive screen in the mammal that could generate
a data base suitable for predicting human response qualitatively
and quantitatively.
The development of short-term tests for mutagenicity provides
145
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146
AGING IN TODAY'S ENVIRONMENT
a useful example for the development of appropriate model shy
terns to identify environmental chemicals that affect aging. The
procedure in using mode! systems to evaluate the effects of envi-
ronmental chemicals on aging is first, to identify short-term tests
that can be used to screen environmental chern~cals for their po-
tential to affect aging processes or age-associated diseases; second,
to screen chemicals; and third, to test those chemicals that had
positive results in short-term tests in mammalian model systems
to develop a data base for predicting human responses.
CONSIDERATIONS IN CHOOSING AND DESIGNING
MODEL SYSTEMS
The general lack of information about the fundamental nature
of aging processes poses problems for risk assessment of potential
environmental agents that promote aging. In the absence of reli-
able information, a prudent course might be to use multiple model
systems. The considerations in choosing model systems for as
sessing relationships between environmental agents and aging or
age-related diseases include:
Length of life or of assay.
. Previous use of the model in the study of aging and toxi-
cology.
. Knowledge of and ability to control for adventitious m
croorganisms.
netic analysis.
.
Knowledge of genetic characteristics and suitability for ge-
Capacity to maintain defined environmental conditions.
Knowledge of pathologic changes associated with aging.
Ready availability of the cells or organisms.
Economic feasibility.
Widespread use in other biologic disciplines.
Relevance to aspects of human aging.
Life span is by far the most widely used marker in assessing
the effectiveness of experimental interventions into the aging pros
cesses. If the end point is longevity, life-table data are required,
including life expectancy at birth and life span of the animal model
to be used. For many strains of mice and rats and for many inver-
tebrate species, sound life-table data are available. Species with
short life spans are distinctly advantageous for such uses. A short
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MODEL SYSTEMS FOR EVALUATING TOXIC AGENTS
147
life span enables the investigator to study the animals longitudi-
nally throughout their lives under well-defined conditions and to
analyze the data with the kiln of undertaking further studies within
the time frame of the investigator's scientific "life span. It must
be emphasized, however, that age-associated changes might occur
in a long-lived species like humans that do not occur in short-lived
species. Obviously, when such changes are to be studied in an
animal model, the time required for the changes to develop must
be considered in relation to the an~mal's life span.
Many invertebrates have life spans of only a few days. Such
organisms might be very useful in the early stages of a comprehen-
sive toxicity-testing regunen. If the assay involves measures other
than life span, it should also be of short duration. Fast assays cost
less than slow ones, yield results rapidly, and make it possible to
assay many compounds in a small space and a short tone.
The importance of keeping a mode} free of infectious diseases
is well illustrated by the reports of Paget and Lemon (1965), who
compared the longevity characteristics of conventionally main-
ta~ned Wistar rats with those of Wistar rats kept specific-pathogen-
free in a barrier facility. Both the median length of life and the life
span of the specific-pathogen-free rats were markedly greater. The
data suggest that infectious disease might distort the assessment
of the aging processes and thus compromise the value of the model.
The genetic characteristics of the animal mode] should be
known and the genotype should be stable to ensure reproducibility
and to ease interpretation in aging studies. An example of the
problems that can be encountered otherwise is illustrated by the
use of the popular ~outbred" strains of rats (e.g., the Sprague-
Dawley and Wistar strains) for aging research. These strains are
maintained by randomly (or not so randomly) mating members
of the stock; a procedure likely to result in each supplier having
a stock with genetic characteristics different from those of other
suppliers. The genetic heterogeneity and the investigator's lack of
awareness of its existence might result In erroneous interpretations,
for instance, if data are obtained on young rats from one supplier
and old rats from a different supplier.
Many invertebrate species have been used extensively in so-
phisticated genetic analyses involving both classical and molecu-
la~ genetics. For example, the metazoan organism with the best
understood genetics is Drosophila melanogaster. The extensive
differences in life expectancy and life span reported in different
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148
AGING IN TODAY'S ~VIRO~NT
stuclies on D. melanogaster relight wise In part from genetic drift
during laboratory propagation. Genetic drift can be avoided by
frequent referral to a common source or by maintenance of stocks
in a nongrowing state. The availability of a common reference
strain that can survive indefinitely while frozen, which eliminates
genetic drift, and its sophisticated genetics make Caenorha6ditis
elegans a system of choice also.
Riley (1981b) demonstrated that it ~ important to define and
control all environmental conditions. He found that over 65%0
of conventionally housed female C3H/HeJ mice had mammary
tumors at the age of 400 days, whereas fewer than logo of the mice
protected from the noise, odors, and other stressors common to
conventional anneal facilities had these tumors at the same age.
Riley related that finding to the difference in plasma corticosterone
concentrations, which were 15~500 ng/m! in the conventionally
housed mice and ~35 ng/m] in the mice housed in the protected
environment.
A major advantage of invertebrate organisms is their ability to
grow under completely defined environmental conditions. More-
over, the fungi and C. elegans can even be grown in chemically
defined media, albeit with some alteration in maximal growth ki-
netics and other life-history traits. This capability can simplify
nutritional studies on aging.
The use of cell, tissue, and organoid cultures as mode} systems
for the investigation of the aging effects of chemical, physical, and
nutritional agents on somatic cells has a number of attractive
features. Tissue-culture studies can simplify interpretation of the
results of in viva studies by avoiding such variables as the effects
of the neural, endocrine, and ~rnmune systems and the nutritional,
microbial, and pathologic status of the host animal.
Knowledge of the pathologic changes that occur with age in an
animal model is essential for both the design and the interpretation
of aging studies. Published data on age-related pathology are
available for some, but not all, animal models. In addition, the
pathologic causes of senescent death in most invertebrates have
not been well explored.
The immediate availability of a source of aged animals, main-
tained in carefully specified environments, would simplify many
studies. The lack of availability of aged animals of short-lived
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MODEL SYSTEMS FOR EVALUATING TOXIC AGENTS
149
strains does not present a serious problem, because in many re-
gards it is preferable for the investigator to be in control of the
animal's environment over its whole life.
Such considerations are usually of relatively minor importance
in invertebrates. Many species are readily available from a variety
of sources, including stock centers and individual laboratories (see
Appendix A). Many of these organisms can be shipped by regular
mail without special precautions, and several species, including
fungi and C. elegans, can be storer} indefinitely in the laboratory
without passage.
An advantage of the invertebrates is their relatively low cost.
For example, a complete life table can be constructed from a
population of 100 nematodes for a cost of about $50, including
overhead, technician time, and data analysis. A comparable study
using mice, however, costs at least $10,000, and about twice that
for using rats. Such cost considerations are a major force in the
development of biologic markers of aging other than life span.
All strains and species mentioned here are widely used in other
biologic disciplines. In particular, Neurospora crassa, C. elegans,
and D. metanogaster are among the most widely studied metazoan
organisms. The use of widely studied organisms is an important
consideration that was covered in detail in a previous report of the
National Research Council (1985~.
For a toxicity-test system to be truly valuable, it must closely
mimic the toxic response typical of an aging human population
or must represent a relevant end point of human aging or an
age-related disease. At our current poor level of unclerstanding
of aging, it is impossible to make completely informed decisions
about the validity of mode} systems. Many fundamental physio-
logic and molecular processes in humans are also present in many
invertebrates ~d other vertebrates, and there is no a priori reason
to expect aging processes to be different. Each model system must
be approached on its own merits, and validity assessment must be
based on background information on the disease state, the mode!
system, and the particular assay under analysis.
EXAMPLES OF MODEL SYSTEMS
~ Vitro Modem
The ability to maintain and grow cells and tissues outside the
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AGING IN TODAY'S ENVIRONMENT
body has progressed to the point where the effects of environmen-
tal agents on living cells can be studied in vitro. For the purposes
of screening environmental agents, the use of such assays can result
in substantial economnes and efficiencies particularly important
when alternative studies require aged animate of several long-lived
species or large numbers of dos~response and drug-interaction ex-
periments. Stock ceils and, in some cases, tissue explants can be
cryobiologically stored in large amounts. This permits repeated
assays with comparable materials and the sharing of common
stock materials by numerous laboratories. Moreover, such stocks
can be used to investigate cell-cell interactions, such as metabolic
cooperation and metabolic transformation. Fmally, tissue-culture
approaches can substantially reduce the numbers of animals re-
quired for experimentation. Such methods cannot, however, be
expected to elirn~nate the need for experimentation with intact
animals.
There are three general categories of methods: organoid cul-
tures, tissue explants, and cell culture. Organoid cultures involve
the short-term maintenance of viable intact segments of tissue,
for example, the full thickness of a segment of aorta. ~ tissue
explants, the early migration and proliferation of epithelioid and
fibroblastoid cell types can be observed. Cell cultures are of three
general types:
~ Primary clones and cultures, that is, proliferating colonies
and mass cultures of cells taken directly from the animal, usually
after enzymatic dispersion of biopsied tissue.
. Established, serially passaged cultures with relatively re-
producible cycles of growth in early phases, but with limited
replicative life span, and with a genetic makeup reflecting that
of the donor animal.
"Transformers cell cultures with indefinite replicative pm
tential and generally with altered genetic makeup.
.
Each of the materials described could prove useful for stud-
ies of the effects of environmental agents on aging. One general
approach would be to explore the toxic effect of an agent as a
function of donor age so as to detect unusual susceptibilities of the
cells and tissue of aged subjects. Another general approach would
be to use the tissue-culture methods after in viva treatments. If
a set of behaviors or phenotypes were observed with tissue from
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MODEL SYSTEMS FOR EVALUATING TOXIC AGENTS
151
young, treated subjects that proved to be comparable to that oW
served with untreated tissue from old anunals, an effect of the in
viva treatments on aging processes could be inferred.
An entirely different experimental paradigm could be based
on the hypothesis that established cultures with finite replicative
life span recapitulate the natural history of comparable cell types
in viva in aging animals the well-known sin vitro mode! of cellu-
lar agings first developed by Hayflick and Moorhead (1961~. The
appropriateness of such models for the study of aging is contro-
versial, but considerable evidence supports the proposition that
t.h~ z`~.~.~nll;~tion Of growth oh~erved in vitro corresponds to events
_ O
· · .
OccurRlilg In VlVO.
Of special interest would be the evaluation of agents that
exhibit unusual toxicity in putative stem cells or that accelerate
the terminal differentiation of such stem ceils, because an excessive
depletion of stem cells might seriously compromise the regenerative
potential of tissues in aging subjects. In such studies, it wall be
important to investigate a variety of cell types. Most research
in this field has concentrated on the in vitro aging of cultures
of fibroblastoid cells established from the fetal lung or from the
dermis of individual subjects of various ages. The precise cell types
of origin of such cultures are not clear. Thus, it will be difficult to
compare age-related changes in viva with those observed in vitro.
Considerable progress has been made in the culture of other
cell types, including epidermal keratinocytes, epidermal melano-
cytes, lenticular epitheliai cells, renal epithelial cells, chondrocytes,
arterial and venous endothelial cells, vascular smooth muscle cells,
skeletal muscle satellite cells and myoblasts, erythroid stem cells,
myeloid stem cells, T arid B lymphocytes, glial cells, and retinal
epithelial cells. Much of this progress has been associated with the
improved characterizations of optimal growth media, especially of
subsets of growth factors. In some instances, growth in chern~cally
defined or nearly defined media has been achieved.
A final experimental paradigm would be to use, in culture,
postreplicative terminally differentiated celb to investigate agents
for their potential to accelerate age-related alterations observed in
viva. Such cell types might be derived through in vitro terminal
differentiation, for example, or normal or transformed embryonic
neuroblasts or myoblasts. A major concern, however, would be
the extent to which the experimental milieu reflected in viva con-
ditions.
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152
AGING IN TODAY'S ENVIRONMENT
Nonmam~an Animal Modem
Numerous invertebrate species fulfill ~nany of the criteria out-
lined earlier and have been widely used as modem for the study of
aging. Some have also been used In studies of genotoxicity. More
important, several have already been used to examine toxic effects
on aging or age-related processes.
The most widely used and perhaps the only commonly ac-
cepted biomarker of aging is life span (Comfort, 19793. Life span
has also been the end point most often assayed in invertebrates,
although other end points, such as accumulation of lipofusc~n and
the end of reproductive ability, have also been used. Because of
their short life spans, relative ease of use, and relatively low cost,
invertebrate organisms will likely be important ~ the initial phases
of a test system to detect environmental toxins that might affect
aging or age-related diseases.
Life-table data are a sine qua non of aging research and are
therefore essential in a test system to assess toxic effects on aging
or age-relatec! diseases. Because invertebrates are often relatively
short-lived, it is easy to collect reliable data on the invertebrate
of choice. Because such data might not be representative of other
genotypes or environments, however, it is often best to choose
an organism on which adequate life-table data have already been
collected by many investigators over an extended period and under
a variety of experimental conditions.
Invertebrates for which adequate life tables are available in-
clude fungi, especially N. crassa and Podospora anserine (Esser
and Bockelman, 1985; Munkres, 1985~; protozoa, such as Parame-
cium telraurelia and Tetrabymena pyriformis (Sm~th-Sonneborn,
1985a,b,c); rotifers (Barrows and Kokkonen, 1985~; the nematodes
C. elegans, Turbatrix aceti, and C. briggsce (Johnson, 1984; John-
son and Simpson, 1985; Russell and Jacobson, 1985~; and insects
(tints, 1985a), especially D. melanogaster (Baker et al., 1985;
Lints, 1985b), but also Musca domestics (Cheeky, 1985; Sohal and
McArthur, 1985), Ha6ro6racon juglandis, and Tribolium confusum
(Soliman, 1985~. Several other invertebrate animal species, less
widely studied (tints, 1985c; Mitchell and Johnson, 1984), and
plants (Nooden and Thompson, 1985), whose mode of senescent
action has been widely explicated, have also been used as models
for the study of aging.
Of the organisms that have been widely used in aging research,
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MODEL SYSTEMS FOR EKALUAT~G TOXIC AGENTS
153
four are of particular interest because they fulfill many of the
criteria outlined earlier: P. tetraureiia, which has a replicative
life span of about 200 cell divisions, encompassing about 40 days
(Smith-Sonneborn, 1985b,c); C. elegans, which has a mean life
span of 1~30 days, depending on temperature and food (Johnson
and Simpson, 1985~; D. melanogaster, which survives an average
of about 40 days (Baker et al., 1985~; and M. domestica, which has
a life span of only about 2 weeks (Cheeky, 1985~. Only two (the
nematode C. elegans and the fruit fly D. metanogaster) are widely
used in other kinds of biologic research and are amenable to the
wide range of molecular and genetic analyses currently available
in the biotechnolog~c armory.
Some protozoa have little ability to continue mitotic replica-
tion in the absence of mating and so have been used to study
the phenomenon of finite proliferative life span, usually termed
clonal aging (Sm~th-Sonneborn, 1984~. Protozoa display a spec-
trum of finite replicative divisions among different species, ranging
from about 40 divisions to apparent clonal immortality. Probably
the best-studied species in aging research is Paramecium aure-
lia, whose age-related morphologic changes have been described
during clonal aging, including rn~cronuclear, macronuclear, and cy-
toplasmic changes. Functional changes in the rate of macromolec-
ular syntheses have been reported. Paramecium has been used in
assessing the effects of environmental insults, particularly radia-
tion, on clonal life span and replication rate (Sm~th-Sonneborn,
1985c). Classical genetics is available for many protozoa, includ-
ing Paramecium. Molecular genetic analysis is well developed in
some species, but is almost completely lacking in Paramecium,
and studies are limited to a few laboratories.
Nematodes, particularly C. elegans and T. aceti, have been
widely studied as models of metazoan aging (Johnson, 1984; John-
son and Simpson, 1985; Russell and Jacobson, 1985~. The somatic
cells of adult nematodes are all postreplicative; mean life spans
are a few weeks; and kinetics of death display the Gompertzian
increase in mortality seen in higher metazoans. A variety of mor-
phologic, behavioral, physiologic, and molecular changes occur
over the life span of the nematode (Johnson and Simpson, 1985~;
some uremic changes observed in the mammalian aging processes,
such as loss of general motor ability (Bolanowski et al., 1981;
Johnson, 1987) and lipofuscin accumulation (Clokey and JacoW
son, 1986~. The animals can be grown on a simple bacterial diet
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AGING IN TODAY'S ENVIRONMENT
or in a completely defined medium (Johnson, 1984; Russell and
Jacobson, 1985~.
C. elegans is the object of study in about 50 laboratories
throughout the world. Sophisticated classical and molecular ge-
netic analyses on this organism are available (Russell and JacoW
son, 1985), including genetic transformation (Fire, 1986~. The
entire cell lineage, from one-cell stage to adult, has been described
(SuIston et al., 1983~. Mutagenesis by transposable elements and
DNA transformation are available. A range of genetic variants
with lengthened life span are also available (Friedman and John-
son, in press; Johnson, 1987~.
Nematodes have been widely used as models in genetic tox-
icology, as well as in the ascertainment of the effects of drug
treatments on life span (Johnson, 1984; Johnson and Foltz, 1987~.
It would be inappropriate to expect to mode! all aging processes
of humans in any invertebrate. For example, all somatic cells in
C. elegans adults are postreplicative and therefore do not mimic
replicating mammalian cells.
D. melanogaster, the most widely studied invertebrate species,
has been the most widely used in aging research (Baker et al.,
1985~. Drosophila has life spans of a few months and can be main-
ta~ned in the laboratory conveniently under well-defined growth
conditions. A wide variety of morphologic, behavioral, physiologic,
and molecular changes with age have been described (Baker et al.,
1985), some of which parallel changes observed in mammals.
Most important, the fruit fly has been intensively studied
for over 70 years as a genetic mode} and for over 60 years as a
general genetic mode! of metazoan aging (tints, 1978~. Studies
have been completed with a wide variety of dietary supplements,
environmental insults, and potential toxins to assess the effect
of these substances on life span (Baker et al., 1985~. Sophisti-
cated molecular genetic techniques are available, including almost
routine transposable-element mutagenesis and molecular cloning
(Spradling and Rubin, 1982~. More is known about the genome
of this organism than about that of any other higher metazoan,
including the human. Moreover, Tong-lived strains of Drosophila
have been derived by selective breeding (I,uckinbill et al., 1984;
Rose, 19843.
Except for Drosophila, the house fly Musca fdomestica is the
most widely used insect mode} for aging research (tints, 1984~.
Musca is easy to maintain, and mean life spans are around 20
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MODEL SYSTEMS FOR EVALUATING TOXIC AGENTS
155
days. Molecular assays can be performed on single flies. A variety
of morphologic, behavioral, molecular, and physiologic charac-
teristics display changes over the life span (Cheeky, 1985~. The
free-radical theory of aging has been most directly examined in
Musca with detailed physiologic and molecular assays (Cheeky,
1985; Sohal, 1981~. Drugs have been used to modify the life span
and physiology in an effort to test the free-radical mode] further.
Unfortunately, almost no classical or molecular genetic analysis of
the house fly is available (Cheeky, 1985~.
The fungi N. crassa and P. anserine have been widely ex-
ploited both in aging research and in classical and molecular
genetic analyses. The haploid nature of the fungi makes them
especially amenable to the identification of some types of muta-
tions, and both of these fungi have been used to characterize the
only senescence phenomenon understood at the molecular level
(Esser and Bockelman, 19853. Most molecular genetic techniques
are also ava~iable for use with these fungi.
Many invertebrate species can be grown under rigorously de-
fined culture conditions. C. etegans con be cultured on chemically
defined media and cells of D. me~anogaster can be grown in cul-
ture. Although many invertebrates have been examined in aging
research, D. meianogaster and C. elegant are widely used not only
in aging but also in other fields, and thus should be emphasized.
The lack of adequate pathology is a major limitation in the
use of many invertebrates. For example, only sparse pathologic
descriptions at death are available for Drosophila at the elec-
tron microscopic level (Baker et al., 1985) and for M. domestics
(Cheeky, 1985; Sohal and McArthur, 1985~. Only a minimal de-
scription, at the electron microscopic level, of senescent changes
in C. elegans has been made (Johnson, 1984~.
Invertebrates are phylogenetically far removed from mammals
and might therefore slider from them in fundamental ways. Al-
though many studies of basic biochemical and physiologic events
have shown that these processes are conserved over large phylo-
genetic distances, it is clear that not all physiologic processes are
conserved. The lack of information on the nature of basic aging
processes in any metazoan makes it impossible to determine, a
priori, the relevance of any organismic mode! to human aging.
Nevertheless, in some important applications, such as assessment
of toxic effects on life span and modification of life span by toxic
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AGING IN TODAY'S ENVIRONMENT
agents, invertebrates are useful modem and are cost-effective al-
ternatives to mammalian modem.
Mammalian Modem
Most mammals do not fully meet the criteria of a short life
span, relative ease of maintenance in a defined environment, wide
use In biologic research, and suitability for a wide variety of molec-
ular and genetic analyses. Mice and rats best fulfill these and other
criteria for aging research (National Research Council, 1981b), and
indeed they have been and are widely used In aging research.
Many vertebrate species, such as rabbits (National Research
Council, 1981b), lack good lif~table data, although they are often
used as models for atherogenesm research. It is difficult to obtain
a rabbit more than 5 years old, not because of spontaneous death,
but because breeders usually kill the animate by that age. Two
studies have addressed the longevity characteristics of rabbits; one
indicated a life span of 8 years (Weisbroth, 1974) and the other of
13 years (Flower, 1931~. Before rabbits can be electively used in
aging research, further information concerning their longevity is
necessary.
Even when excellent life-table data are available, in,restig~
tors often fad! to make use of them ~ designing their aging stud-
ies. Prominent biochemists and metabolic physiologists cornrnonly
draw conclusions about age-related changes in a biochemical pros
cess on the basis of a study limited to 2-month-old and ~month-old
rats in a strain with a life span of 48 months. Although such stud-
ies are of value, a broader range of ages should be studied if the
influence of age on biochemical activity is to be aclequately defined.
The value of published pathologic data is iilllstrated by the
following example. The male Fischer 344 rat ~ a popular mode!
for aging studies. More than 50~o of these rats have testicular
interstitial cell tumors at the age of 18 months, and by the age of
24 months almost all have the tumors (Coleman et al., 1977; Yu
et al., 1982~. That information is critically important in designing
studies on the aging of the male reproductive system. It is also
important to carry out pathologic analyses in the same animals on
which physiologic or biochemical measurements are being made.
For instance, if the concentration of sequin parathyroid hormone
increases in some but not all old male Fischer 344 rats, it Is
important to know whether this increase could be secondary to a
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MODEL SYSTEMS FOR EVALUATING TOXIC AGENTS
157
coexisting disease process, for example, a severe grade of chronic
nephropathy.
Rats have the advantage of providing more biologic material
per animal than mice without markedly increasing the ma~nte-
nance cost or space requirement. A disadvantage is the small
number of inbred strains available (National Research Council,
1981b). The Fischer 344 strain is the major inbred rat strain
that has been used in aging research. It has several advantages
in addition to genetic homogeneity: it does not become obese
with advancing age (Bertrand et al., 1980a), extensive data on its
age-related disease processes have been published (Coleman et al.,
1977; Maeda et al., 1985), and there is a sizable body of life-table
data (Masoro, 1980; Yu et al., 1982, 1985~.
The major disadvantage is the occurrence of progressive
chronic nephropathy and renal failure in animals fed ad libitum
at a relatively early age (Maeda et al., 1985~. Restricting the
rat strain to 60% of the ad libitum food intake prevents the pros
gressive chronic nephropathy. The National Institute on Aging is
developing Brown-Norway, Fischer 344 Fat hybrids for the purpose
of partially circumventing chronic nephropathy. It is already clear
that a controlled and defined dietary progam is necessary in aging
studies.
Other rats that have been extensively used for aging research
are the Sprague-Dawley, Waster, and Long-Evans (Masoro, 1980~.
They are not inbred strains and are not genetically homogeneous.
It is more appropriate to call them stocks than strains (National
Research Council, 1981b). Life-table data are available (Masoro,
1980~. A comprehensive report (Anver et al., 1982) on the age-
related disease processes of the male Sprague-Dawley rat is avail-
able. A disacivantage of the male Sprague-Dawley and Wistar rats,
compared with male Fischer 344 rats, is that they become obese
with advancing age (National Research Council, 1981b).
Because they are small, mice are relatively inexpensive to
maintain and clo not require a large space. Moreover, many inbred
strains are available, and some are even available as aged animals
(see Appendix A), which facilitates genetic exploration of aging
(National Research Council, 1981b). A disadvantage of mice is
that only a small amount of biologic material can be obtained
from a single animal. Good life-table data are available on sev-
eral strains (National Research Council, l9Blb): C57BI,/6N Nia
males, BAI`B/cN Nia males, CB6F males, C57BL/6] females,
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AGING IN TODAY'S ENVIRONA~3NT
DBA2/] females, B6D2F females, C57BL/63 males, DBA2/]
males, B6D2F~ males, C3Hf/B~ females, BALB/c An B3f fe-
males, C3CF females, C3Hf/B] mates, BALB/c An B6f males,
and C3CF~ males.
Inbred strains often suffer from a single major dmease process
(e.g., cancer of the liver or chronic nephropathy), and the presence
of this major disease process in most if not all the animals compli-
cates gerontologic interpretation. Fit hybrid strains offer a partial
solution to the problem and can be readily produced in large num-
bers. McClearn et al. (1970) have developed a more genetically
heterogeneous stock of mice that is systematically maintained.
Their procedure yields a genetically stable population without the
drawback of inbred strains and thus probably provides a mode!
for aging research in which genetic variation is readily accessible.
Recombinant inbred strains (Bailey, 1981) should also be more
widely used. Because of the genetic identity of each member of
a population, some individuals can be sacrificed while identical
siblings are maintained for survival and analyses on living animal
(Johnson, 1987~.
If old animals are available from suppliers, the investigator
must also make certain of the availability of accurate "formation
on the lifetime environment of the animals, including dietary his-
tory and data on the monitoring of infectious disease. Usually,
such data are not available on long-lived animal models of ad-
vanced age. Indeed, the lack of well-defined animals and the cost
of their purchase and maintenance have made it difficult to carry
out aging research on adequately characterized long-lived animal
models. Reasonable life-table data are available on the beagle and
the thoroughbred horse.
With regard to nonhuman primates, the two principal mod-
els that appear to be emerging are the pig-tailed macaque mon-
key (Macaca nemestrina) (Bowden, 1979) and the rhesus monkey
(`Macaca mulatta) (Davis and Leathers, 1985~. It might be de-
sirable to develop aging cohorts of the chimpanzee because, from
a biochemical and genetic point of view, chimpanzees are closely
related to man; but practical considerations limit the feasibility of
this model.
The availability of tissues and fluids from a selection of mam-
malian species of contrasting life spans, ranging from the short-
lived murine species to the long-lived primates, would permit a
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MODEL SYSTEMS FOR EVALUATING TOXIC AGENTS
159
systematic comparative approach to the study of aging. For ex-
ample, it would permit the differentiation of markers that appear
to be related primarily to chronologic rather than biologic age,
and thus provide a stronger rationale for the choice of assays for
the effects of agents that might alter aging processes.
Epidemiologic Models
The expression "experiments of nature" refers to contrasts in
the exposure of human populations to toxic agents that provide
an opportunity to assess the impact of toxic exposures on the risk
of disease and death in the human population. Many reports of
acute and chronic releases of toxic agents into the human environ-
ment gave rise to marked contrasts In toxic exposures and disease
outcomes. They include the mercury contamination of Minimata
Bay in Japan, the atomic blasts in Hiroshima and Nagasaki, and
smog episodes in London.
The first of the weli-publicized air-pollution incidents that re-
sulted in marked increases in illness and death, mostly among the
elderly, occurred in the Meuse River Valley in Belgium in 1930.
This heavily industrialized area was seriously affected when ac-
cumulating air contaminants, trapped by an inversion resulting
from a thick, cold fog, caused 60 deaths and illness in thousands of
residents. The incident at Donora, Pennsylvania, in 1948 resulted
from a similar inversion that covered a wide area of the northeast-
ern United States, including the industrial community of Donora.
Of the population of 14,000, 20 died (compared with the expected
2 deaths for the period) and 43% fell ill. Again, the elderly were
the most seriously affected. Similar episodes of smog-induced mor-
tality in London as early as 1873 and in New York City during the
1950s and 1960s have been described (Amour, 1986~.
In all those episodes, the most vulnerable people were the el-
derly and those with pre-existing disease of the cardiopulmonary
system. Organic mercury compounds, specifically methyl and
ethyl mercury salts, are of toxicologic importance because they
pass through the blood-brain barrier, accumulate in the brain,
and can produce irreversible damage to the central nervous sys-
tem. Chronic exposure to these compounds might occur in the
workplace or in the general environment.
Intense interest in the toxicity of methyl mercury developed in
the 1950s, when a neurologic illness, now called Minimata disease,
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160
AGING IN TODAY'S ENVIRONMENT
appeared In the families of fishermen living around Minimata Bay
in Japan. The bay water was contaminated by mercury waste
(inorganic and methyl mercury) from a local chemical plant. In-
vestigation determined that the illness resulted from the repeated
ingestion of large quantities of fish in which the mercury concen-
tration was thousands of tunes that in the water (Goyer, 1986~.
The cancer experience of the Japanese atom~c-bomb survivors
in Hiroshima and Nagasaki has been carefully assessed in rela-
tion to the estimates] whol~body doses received by individual
survivors. The Life Span Study sample of the Radiation Ef-
fects Research Foundation includes 82,000 atomic-bomb survivors
and 27,000 nonexposed comparison persons, among whom 19,606
deaths had occurred by 1974 (Beebe, 1979; Beebe et al., 1978~.
These studies have been extremely informative about radia-
tion-related solid tumors as well as leukemia. For example, they
documented the striking difference in the minimal latent periods,
with the minimal period for solid tumors being about 10 years,
compared with 2 years for leukern~a. Age at time of exposure
appears to be a strong determinant of leukemia risk; the greatest
absolute risk is experienced by those exposed at ages ~9 and 50
years and over.
Leukemia was the first cancer reported in excess among
atomic-bomb survivors, but in the most recent followup stud-
ies, the later-appear~ng solid tumors appear to be ~ important
as leukemia in terms of absolute risk. Most of the excem cancer
deaths from solid tumors among the atomic-bomb survivors have
occurred in those over 35 at the time of the blast. It appears,
therefore, that the radiation effect manifested as lung, breast, and
other solid tumors is observed only in people who have reached
the age range normally associated with the incidence of cancer.
LIFESPAN MODULATION BY DRUG T1lEATMI:NT
The effects of toxic agents on aging or aging processes have
not been widely studied (Schneider and Reed, 1985~. Some of the
best studies carried out so far have concentrated on the effects
of drugs that alter oxidative stress. Usually, mean life span and
often maximal life span are used as the end points in assessing
the effectiveness of experimental interventions in such studies,
although in a few cases lipofuscin accumulation and other end
points have been used (BaTin, 1982~.
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MODEL SYSTEMS FOR EVALUATING TOXIC AGENTS
161
Although several examples of extension of mean life span in
mice, in nematodes (Balin, 1982), in Drosophila (Baker et al.,
1985), and in house flies (Sohal and Allen, 1986) have been re-
ported, the studies have not been highly replicated and have some-
times been complicated by design flaws. For instance, treatment
often affects only mean and not maximal life span, or treatment
might reduce early life trauma and death, but have no effect on
aging itself. Studies might also be complicated by effects of the
drug treatment on food intake, in that food intake or body weight
is significantly decreased by the drug (BaTin, 1982~. No single drug
treatment or dietary additive has been reliably shown to extend
life span in any organism.
Representative terms from entire chapter:
aging research
