United States
Environmental Protection
Agency
Health Effects Research
Laboratory
Research Triangle Park NC 27711
Research and Development
EPA/600/S1-85/004 Aug. 1985
SERA Project Summary
Respiratory Carcinogenicity of
Diesel Fuel Emissions
Alan M. Shefner, Bobby R. Collins, Arsen Fisks, Jean L Graf, and
Carol A. Thompson
A large-scale experiment utilizing
3000 hamsters was conducted to com-
pare the carcinogenicity of diesel ex-
haust particles (administered by 15
weekly intratracheal instillations) to that
of organic extracts of diesel particles,
coke oven emissions, roofing tar vola-
tiles and cigarette smoke condensate.
Appropriate solvent controls, untreated
controls and positive controls were
included in the design of the experiment.
The overall incidence of respiratory
tract tumors in any of the treatment
groups was not significantly higher than
in control hamsters. Treatment with
benzol a) pyrene resulted in significantly
higher tumor incidences, primarily
evidenced as polyps in the trachea,
lung, and larynx. Similarly, there were
no significant differences in the survival
rates of hamsters treated with test
materials from those of their respective
controls. Hamsters treated with test
materials generally showed significantly
lower mean body weights than control
animals. This effect began during the
15-week treatment period and frequent-
ly lasted into the post-treatment holding
period. Treated hamsters generally
showed a delay in time to reach maxi-
mum body weight when compared to
hamsters in control groups.
Treatment of hamsters with test
materials induced a variety of hyper-
plastic, proliferative and inflammatory
lesions of the respiratory tract. The
highest incidence rates and greatest
severity of the lesions were induced by
diesel exhaust particles and coke oven
emissions. Diesel exhaust extract and
benzo(a)pyrene were less reactive, and
cigarette smoke condensate and roofing
tar volatiles produced the lowest inci-
dence of respiratory tract lesions.
This Project Summary was developed
by EPA's Health Effects Research Labo-
ratory, Research Triangle Park, NC, to
announce key findings of the research
project that is fully documented in a
separate report of the same title (see
Project Report ordering information at
back).
Introduction and Summary
The purpose of this study was to assess
the carcinogenic potential of diesel engine
exhaust particles and organic solvent
extracts and to compare their carcinogen-
icity to that of other combustion products
of known epidemiologic significance. The
comparison materials included roofing
tar, coke oven emissions, and cigarette
smoke condensate. All materials and the
proper controls were given to hamsters
by intratracheal instiljation (Saffiotti tech-
nique) for 15 weeks. Animals were sacri-
ficed at 110 weeks of age (98-99 weeks
on study) and gross and histopathological
examinations were made.
In general, there were no significant
differences in the survival rates, organ
weights, and clinical pathology param-
eters related to the test compound or test
compound concentration within each
replicate experiment. However, the sur-
vival rates and presence of skin lesions
differed significantly between the two
replicate trials. These differences were
ascribed to the initial housing condition
used in the two replicate experiments. In
the first replicate the hamsters were
housed in groups of three during the first
five months of the experiment and indi-
vidually thereafter. A considerable degree
of fighting was seen among the hamsters
during the period of group housing,
resulting in skin lesions, severe secondary
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infections and death. During the second
replicate experiment, hamsters were
group housed for only one month before
the individual housing was established.
Hamsters treated with test materials
frequently showed significantly lower
mean body weights than their respective
solvent controls. This effect was usually
seen during the 15-week treatment period
and in many cases continued during the
post-treatment holding period. Inspection
of the computer-drawn body weight
curves shows that the body weight change
difference frequently was cuased by a
shifting (delay) in weight gain in test-
material treated hamsters rather than by
a reduction in the maximum body weight
achieved.
Treatment of hamsters for 15 weeks by
once weekly mtratracheal instillation of
the various test articles induced gross
and microscopic lesionsof the respiratory
tract at response rates that were related
to specific test substances.
Diesel emission particles administered
alone (DPOO) or in combination with
ferric oxide (DPFO) and coke oven emis-
sions (COFO) were the most reactive of
the substances tested and produced a
variety of pathologic lesions in the respir-
atory tract at incidence rates significantly
higher than their respective controls.
Diesel extract (DEFO) and benzo(a)-
pyrene (BPFO) induced less respiratory
tract pathology than did diesel particles or
coke oven emissions but did produce
significant pathologic changes. Treatment
with BPFO produced granulomas and
mineralization in thoracic and mandibular
lymph nodes to a significant degree, an
effect not seen with other test materials.
Cigarette smoke condensate (CSFO)
and roofing tar volatiles (RTFO) induced
minimal changes above those produced
by the solvent-ferric oxide control treat-
ments. The incidence of respiratory tract
lesions induced by these two substances
was significantly lower than seen in
hamsters treated with the other test
materials.
A Chi Square statistic was used to
compare paired samples of pathologic
effects in order to address particular
questions of interest. Three such analyses
are shown in Table 1 and indicate that:
1. Diesel particles alone at a dose
level of 5 mg per week induce
significantly more pathology than
does a 5 mg per week treatment
consisting of 2.5 mg of diesel
particles admixed with 2.5 mg of
ferric oxide. That is, it is the specific
properties of diesel particles rather
than the total particle load that is
significant.
2. A consistent dose response rela-
tionship is seen when the effects
produced by 5.0 mg per week
treatments with diesel particles are
compared to those induced by 2.5
mg per week exposures.
3. The addition of an equal quantity of
ferric oxide to the 2.5 mg dose of
diesel particles generally increased
the response rates to a modest
degree but statistically significant
changes were only seen for one
class of lesions. That is, in the case
of diesel particles admixed with
ferric oxide the induced pathology
can be attributed to the diesel
particles rather than to an experi-
mentally produced dose particle.
Conclusions
The results of these studies indicate
that diesel particles with or without ferric
oxide and diesel extract with ferric oxide
were not carcinogenic for hamsters when
given intratracheally at 5.0, 2.5 and 1.25
mg doses once a week for 15 weeks.
Similarly, coke oven extract, cigarette
smoke condensate or roofing tar extracts
in mixture with ferric oxide were not
carcinogenic at 5.0, 2.5 and 1.25 mg
doses given over the same time period.
Benzo(a)pyrene plus ferric oxide caused
numerous polyps in the trachea and a few
carcinomas in the lung and trachea,
especially in the second replicate experi-
ment.
The presence of diesel particles in the
lung, pleuro, and trachea did causea high
incidence of lesions in these organs. The
lesions were generally mild to moderate
in severity and were dose related. The
lesions included inflammatory reactions,
granuloma formation, fibrosis, adenom-
atous and papillary hyperplasia, squa-
mous metaplasia, and particle accumula-
tion in the lung. Particle accumulation
and inflammatory reaction were seen in
the trachea, larynx and pleura in addition
to fibrosis and proliferation in the pleura.
The presence of these lesions in the lung,
pleura, and trachea did not appear to be
life threatening over the time course of
this experiment since there were no
significant differences between the sur-
vival rates of the hamsters given the test
articles and their respective controls.
Recommendations
Concern over a postulated increase in
the proportion and numbers of light-duty
diesel engine powered vehicles in the
total automotive fleet has provided the
impetus to studies of potential health
effects of an increase in emissions of
carbonaceous particulate matter which
might result. The U.S. Environmental
Protection Agency has responded by
sponsoring a number of studies to assess
such impacts of which this program
carried out at IIT Research Institute is
one.
The design of this current study incor-
porated not only the evaluation of diesel
emission particles as a potential respira-
tory carcinogen but also the ability to
Table 1.
Lesions
Comparison of Pathological Effects of Diesel Particle (DPOO) and Diesel Particle Plus
Ferric Oxide (DPFO) Test Materials
Treatment
Treatment
DPOO - 5 mg DPFO - 2.5 mgc DPOO - 2.5 mg DPFO - 2.5 mg
Pres ABS Pres ABS Pres ABS Pres ABS
Lungs
Inflam. Reaction
Granuloma
Fibrosis
Hyperplasia. Adem.
Hyperplasia, Papil.
Metaplasia, Oil.
Metaplasia, Squa.
Proliferation, Fibro.
Pleura
Inflam. Reaction
Fibrosis
Proliferation
Mononucl. Cell Infiltrate
76
71
29
71
27
38
a
11
33
62
40
14
14
19
61
19
63
52
82
79
51
22
44
70
55
40
14
54
7
6
4
7
10
27
38
4
39"
54"
80"
40"
87"
88"
90
87
81"
64"
53
87"
47
23
8
42
4
8
1
3
7
21
28
5
43"
67"
82"
48"
86"
82"
89"
87"
78"
64"
57"
80"
55
40
14
54
7
6
4
7
10
27
38
4
39
54"
80
40
87
88
90
87
81
64
53
87
"Significantly different from DPOO 5 mg.
^Significantly different from DPOO 2.5 mg.
"DPFO - 2.5 mg = 2.5 mg diesel particles + 2.5 mg ferric oxide.
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compare any effects produced to those of
an organic solvent extract of the diesel
particles and to other combustion pro-
ducts of known epidemiologic significance.
These other materials included for com-
parability and ranking of relative health
risks were coke oven emissions, roofing
tar volatiles, and cigarette smoke con-
densate.
Under the conditions of this experiment
(once weekly intratracheal administration
of test materials for 15 weeks and
termination at two years) none of the test
materials were carcinogenic. The positive
control compound, benzo(a)pyrene, in-
duced tumors, primarily polyps, in a
significant proportion of treated hamsters
using the same treatment regimen.
However, the absence of carcinogenic
response to diesel particles and the other
test substances under the conditions of
this experiment and in the test species
utilized does not permit definite conclu-
sions about their lack of carcinogenicity
to human beings. The limitations on the
total dose that can be effectively admin-
istered, the limited life span of the
hamsters as test species, total number of
animals treated, and possible species
differences in sensitivity all mitigate
against definitive conclusions. The lack of
reported tumor induction in long-term
experiments in which test animals have
been exposed to diesel exhaust particles
by the inhalation route is in agreement
with the results of this study.
The instillation of diesel exhaust par-
ticles, with or without admixture of fe203
carrier particles, produced inflammatory
and proliferative responses in various
portions of the respiratory tract. These
responses were similar in degree to those
induced by exposure to coke oven emis-
sions and more severe than those induced
by diesel particle extracts, roofing tar
volatiles, and cigarette smoke conden-
sate.
While the induced respiratory tract
pathology did not affect survival rates in
otherwise unstressed hamsters, the
effects of diesel particle exposure on the
ability of animals to respond to other
stressors should be evaluated. Intratra-
cheal administration with diesel particles
provides a useful method for loading the
respiratory tract of test animals with
known and reproducible burdens. Meas-
ures of effect which could have signifi-
cance for public health include:
• Additive effects in individuals with
reduced respiratory function.
• Long-term consequences of the depo-
sition of diesel particles for which
longer lived test species would be
necessary.
• Cocarcinogenic properties of diesel
particles.
• Respiratory function under work or
exercise stress.
• Reduction in immunocompetence.
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Alan M. Shefner, Bobby R. Collins, Arsen Fisks, Jean L. Graf, and Carol A.
Thompson are with IIT Research Institute. Chicago, IL 60616).
Judith A. Graham is the EPA Project Officer (see below).
The complete report, entitled "Respiratory Carcinogenicity of Diesel Fuel
Emissions," (Order No. PB 85-228 120/AS; Cost: $25.00, subject to change)
will be available only from:
National Technical Information Service
5285 Port Royal Road
Springfield, VA 22161
Telephone: 703-487-4650
The EPA Project Officer can be contacted at:
Health Effects Research Laboratory
U.S. Environmental Protection Agency
Research Triangle Park, NC 27711
United States Center for Environmental Research
Environmental Protection Information
Agency Cincinnati OH 45268
Official Business
Penalty for Private Use $300
EPA/600/S1-85/004
0000329 PS
U S ENVIR PROTECTION AGENCY
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