Industrial Health 1997, 35, 301-324

REVIEW ARTICLE

Assessment of the Health Effects of Trichloroethylene

Takashi KANEKO, Pei-Yu WANG and Akio SATO*

Department of Environmental Health, Medical University of Yamanashi, Tamaho, Yamanashi 409-38, Japan

Received March 22, 1997 and accepted May 7, 1997

Abstract: The epidemiological studies performed thus far have presented only limited evidence for
the carcinogenicity of trichloroethylene (TRI) to humans. However, these studies had drawbacks
such as insufficient size of cohort, short observation period, and inadequate TRI exposure assessment;
therefore, no concrete conclusion has been reached concerning TRI carcinogenicity to humans.
Despite the limited epidemiological evidence as to the carcinogenicity of TRI, the International Agency
for Research on Cancer (IARC) has changed the carcinogenicity classification of TRI from Group 3
(not classifiable as to carcinogenicity to humans) to Group 2A (probably carcinogenic to humans).
In regard to the new classification by the IARC, the committee for occupational exposure limits of
the Japan Society for Occupational Health has made a proposal that it is too early to classify the
carcinogenicity of TRI as Group 2A and that it is proper to promote exposure control with the
carcinogenicity being classified as 2B for the moment. There are species differences in TRI
carcinogenicity, particularly between rats and mice. Although experimental studies have found no
evidence that TRI induces liver cancer in rats, there is ample evidence that TRI promotes the
development of liver cancer in mice, particularly in B6C3F~ mice. The carcinogenicity of TRI in this
strain of mice may be based on an epigenetic mechanism rather on a genotoxic mechanism and the
liver cancer may be induced only after TRI has been inhaled for a long period of time at concentrations
high enough to cause cytotoxicity. Conversely, with no reports showing TRI-induced renal tumors
in mice, the possibility has been suggested that this chemical induces such tumors in male rats. The
species differences are mainly accounted for by differences in the metabolism of TRI between rats
and mice. From a general survey of the literature, it can be concluded that TRI itself is not mutagenic.
However, the conjugation of TRI with glutathione (GSH), a minor pathway of TRI metabolism,
results in mutagenic metabolites in the kidney of rats. The acute toxicity of TRI is neurotoxicity
based on its anesthetic action. An exposure to extremely high levels of TRI may cause the liver and
kidney disorders. Repeated exposures to high levels of TRI may result in neuro-, hepato-, and/or
nephrotoxicity. The main symptoms appearing after chronic exposure at low levels are neurological
changes represented by subjective symptoms relating to central and autonomic nervous systems, or
by a lowered conduction velocity of the nerves or a prolonged latency of the nerve responses. For
the present, it is reasonable to use the neurological findings for establishing the reference values of
TRI for both work and general environments. A value of 25 ppm (135 mg/m3) is proposed as a
reference value for work environments, and 25-50 ppb (135-270 pug/m3)for the general environment
(111,000 of the value for work environment).

Key words: Trichloroethylene, Carcinogenicity, Mutagenicity, Risk assessment

*To whom correspondence should be addressed .

302
Introduction
Trichloroethylene (TRI) is an industrial solvent used
mainly for degreasing of metal parts. Its use in dry cleaning
is now uncommon. TRI may be found in printing inks,
varnishes, adhesives, paints, and lacquers. TRI has attracted
great attention among the general public because of its
leakage from industrial settings into the general environment,
particularly into underground water. The greatest concern
raised by both occupational and environmental health
professionals has been whether or not TRI is a human
carcinogen, i.e., whether long-term exposure to low doses
of TRI causes cancer in humans. In the present paper, the
health effects of TRI are discussed with a special emphasis
on its carcinogenicity.
Carcinogenicity of TRI
(1) Epidemiological studies
The results of the epidemiological studies of TRI
carcinogenicity are summarized in Table 1'~. The first cohort
study on the carcinogenicity of TRI was conducted in 1978
by Axelson et a1.2~in Sweden. This study was carried out
with a cohort of 518 persons exposed to TRI; no excess
death from cancer was demonstrated. A cohort study
conducted in Finland thereafter with 2,084 workers
occupationally exposed to TRI also found no evidence of
TRI carcinogenicity3~. Excess death from cancer was not
found in a cohort consisting of 2,646 employees of a
manufacturing plant using TRI4~. In this study, the cohort
had an observed cancer mortality of 21 as opposed to an
expected mortality of 36.7, with a standardized mortality
ratio (SMR) of 0.57.
Garabrant et al.5~conducted a retrospective cohort mortality
study with 14,067 workers of both sexes employed for four
or more years between 1958 and 1982 at an aircraft
manufacturing company in San Diego County, California.
The follow-up of the cohort members for a mean duration
of 15.8 years from the date of first employment resulted in
the successful tracing of 95% of the cohort and found 1,804
deaths through 1982. The mortality rates thus obtained were
compared with the U.S. national data and San Diego County
data in terms of race (white and nonwhite), sex, age, calendar
year, and cause of death. Mortality due to all cancers was
low (SMR 0.84). When the data were analyzed according
to the site of lesion, SMR with a value more than 1.0 was
calculated for the esophagus (1.14), pancreas (1.19), urinary
tract (1.10, malignant tumors in the urinary tract other than
T KANEKO et al.
the kidney [1.26]), and benign brain tumors or brain tumors
not evaluated for the degree of malignancy (1.10, brain tumor
not evaluated for the degree of malignancy [1.21]). The
lower limit of the 95% confidence interval (95% CI) of SMR
did not exceed 1.0 for any of these lesion sites. Garabrant
et al. concluded that no significant excesses in cause-specific
mortality was found. However, they also stated that because
of the study design, excess risks could not be detected for
those diseases that had latency periods in excess of 20 to 30
years. In this study, 37% of the subjects were exposed
exclusively to TRI, but workers exposed to other various
compounds were also included in the cohort.
Spirtas et al.6~carried out a retrospective cohort study at
an aircraft maintenance facility of an air force base in Utah,
the U.S., that employed 14,457 workers. Of these workers,
6,929 were exposed to TRI in the 1950s to 1960s. The
observed deaths among white workers were compared with
the expected number of deaths, based on the Utah white
population, and adjusted for age, sex, and calendar period.
Significant decreases were found in mortality from all causes
(SMR 0.92; 95% CI 0.90-0.95), all malignant neoplasms
(0.90; 0.83-0.97), ischemic heart disease (0.93; 0.88-0.98),
non-malignant respiratory disease (0.87; 0.76-0.98), and
accidents (0.61; 0.52-0.70). Mortality was increased for
multiple myeloma (MM) among the women (2.36; 0.87-
5.14), non-Hodgkin's lymphoma (NHL) among the women
(2.12; 1.02-3.90), and cancer of the biliary passages and
liver among the men who died after 1980 (3.58;1.16-8.36).
The detailed analysis of the 6,929 employees occupationally
exposed to TRI, the most widely used solvent at the base
during the 1950s and 1960s, did not show any significant
or persuasive association between several measures of
exposure to TRI and any excess of cancer. Among the women
employed in the departments in which fabric cleaning and
parachute repair operations were performed, more deaths
than expected occurred from MM and NHL. The inconsistent
mortality patterns by sex, multiple and overlapping
exposures, and small numbers made it difficult to ascribe
these excesses to any particular substance.
Axelson et al.'s expanded their initial epidemiological
study2>.The cohort of the updated study was 1,727 workers
who had used TRI and had undergone a urinalysis for
trichloroacetic acid (TCA) at least once during the period
1955 through 1975. Of these workers, 57 were excluded
from the data analysis because of failure in the follow-up
(e.g., incomplete documents); the remaining 1,670 workers
(1,421 males and 249 females) were enrolled in the study.
The mean TCA value was used as the level of exposure to
Industrial Health 1997, 35, 301-324
HEALTH EFFECTS OF TRICHLOROETHYLENE
Table 1. Epidemiological studies on the carcinogenicity
TRI when more than one TCA values were available. The
interval from the date when the first urinalysis for TCA was
made until retirement or 1979 was used as the duration of
exposure. The cohorts were monitored for deaths until 1986
(or 1987 for cancer death or cancer incidence). Risk was
expressed in terms of SMR or standardized incidence ratio
(SIR) by comparing the cancer mortality or incidence in
the cohorts with those in the nationwide population. Among
the men (n=1,421), the SMR for all mortalities was 0.97
(95% CI 0.86-1.10) and the SMR for mortalities from cancer
was 0.65 (0.47-1.10). The SMR for mortalities from
circulatory disorders (cardiovascular and cerebrovascular)
303
of trichloroethylene
was 1.17 (1.00-1.37), showing that the mortality rate for
circulatorydiseases was marginally higher in persons exposed
to TRI. However, no significant relationship between the
level of exposure, as determined from urinary TCA levels,
and the mortality from circulatory disorders was observed.
When the duration of exposure was divided into less than 2
years and not less than 2 years, the subjects exposed to TRI
for less than 2 years showed a higher SMR value for all
mortalities (1.36; 1.241.72) as well as for the mortality
from circulatory disorders (1.68; 1.242.27). In addition,
in the low exposure level group (TCA<49 mg/1), the SMR
was significantly higher both for all mortalities (1.49;1.13-
304
1.97) and for mortalities from circulatory disorders (1.79;
1.23-2.57). The incidence of all cancers in the exposed
cohort was similar to that in the general population (SIR
0.96; 0.80-1.16). When the incidence of all cancers was
analyzed according to the site of the lesion, the incidence
of nonmelanocytic skin cancer was higher in the exposed
cohort than in the nationwide population (SIR 2.36; 1.02-
4.65) with the higher incidence being particularly noted in
the low-exposure-level group (TCA<49 mg/l). The
incidences of liver cancer (SIR 1.14), larynx cancer (1.39),
prostate cancer (1.39), testis tumor (1.25), and non-Hodgkin
lymphoma (1.56) were higher than the expected values, but
no significant increase was found in the exposed cohort.
The number of subjects in the female subcohort was small
(249) and the information available was limited. The total
mortality rate in the exposed female cohort was higher than
that in the nationwide population (SMR 1.55; 1.02-2.31).
The observed number of mortalities was larger for malignant
tumors (SMR 1.53) and circulatory disorders (2.02) than
the expected number of mortalities, with no significant
increase in the exposed cohort. In all, for both genders, the
excess risks were largely confined to groups of workers with
lower exposure levels or short duration of exposure or both.
Axelsonet al. concludedthat their study provided no evidence
that TRI is a human carcinogen.
Anttila et al.8~compared workers occupationally exposed
to TRI, tetrachloroethylene (TETRA), and 1,1,1-
trichloroethane (1,1,1-TRI) whose exposure was identified
by analyzing biologicalsamples (TRI, urinary TCA; TETRA,
blood TETRA; 1,1,1-TRI, blood 1,1,1-TRI) with the
population in Finland with respect to the incidence of cancers
(SIR). Only the epidemiological study of TRI will be
described here. The cohort exposed to TRI was composed
of 1,698 males (31,552 person-years) and 1,391 females
(28,353 person-years). Each worker in the TRI cohort
underwent urinalysis for TCA 2.5 times on the average. The
median urinary TCA level was 48 mmolll for males and 60
mmolll for females (males<females) during the period 1965
through 1982, whereas the corresponding values were 80-
90 mmolll and 60-80 mmolll, respectively, before 1970
(males>females). The incidence of cancers (total, liver,
biliary tract, gallbladder, pancreas, kidney, bladder,
lymphohematopoietic tissue, brain and nervous system,
genital organs, and tissues likely to be exposed to TRI [oral
cavity,pharynx, larynx, nasal cavity, trachea, and lung]) was
investigated by using the cancer registry in Finland (1967-
1992). Tumors developed in 208 workers in the TRI cohort.
For workers exposed to TRI for 20 years or longer, the SIR
T KANEKO et al.
was significantly higher than 1.0 in the following: total
cancers (number of patients observed 60, SIR 1.97, 95% CI
1.20-2.20), stomach cancer (7, 2.98,1.20-2.20), liver cancer
(males: 3, 6.07,1.25-17.7; females: 3, 13.0, 2.68-37.9),
prostatic cancer (8, 3.57, 1.54-7.02), and
lymphohematopoietic tissue tumors (7, 2.98,1.20-6.14).
When the data were analyzed by taking into account the
level of TRI exposure, an increase in the incidence was noted
for rectal cancer (9, 2.34,1.07-4.44) in the group having
urinary TCA<100 mgll and for cancer of the cervix uteri
(5, 4.35, 1.41-10.1) in the group with urinary TCA> 100
mgll. However, when all the members of the exposed cohort
were used for comparison, an increased incidence was found
only in the cancer of cervix uteri (8, 2.4,1.1-4.8). Since
the study of Anttila et al. was carried out in Finland, where
the cancer registry and the dynamic statistics of the population
are accurately conducted with individual identification codes,
the obtained results are most likely reliable. However, no
account has been given in the report of Anttila et al.
concerning the increased incidence of stomach or uterine
(cervix) cancer in the TRI-exposed cohort, which has not
been found in animal experiments or in other epidemiological
studies. Whether or not the members in the exposed cohort
were representative of the Finnish people concerning the
living environment is unknown because the details of the
cohort are not given in the report.
Henschler et al.9~conducted a retrospective cohort study
on the relationship between exposure to TRI and renal cell
cancer in a cardboard factory in Germany. The study group
consisted of 169 men who had been exposed to TRI for at
least 1 year between 1956 and 1975. The average observation
period was 34 years. The control group was composed of
190 workers who worked at the same factory in the same
period without being exposed to TRI. The SMR was
calculated compared with the local population. By the closing
day of the study (December 31,1992), 50 members of the
exposed cohort had died (SMR 0.79), 15 of the 50 from
malignant neoplasms (1.01). Of these 15 workers, two died
of kidney cancer (SMR 1.01). In the control cohort, 52
workers had died by the end of the study period (1.03). Of
these workers, 15 died of malignant neoplasms (1.16), but
none of them died of kidney cancer. Three workers of the
control cohort died of brain tumors (9.38; 95% confidence
interval 1.93-27.37). As for the incidence of cancer, the
diagnosis of kidney cancer was made in five workers of the
exposed cohort: four with renal cell cancers and one with a
urothelial cancer of the renal pelvis. The time interval
between the TRI exposure and diagnosis was 18 years or
Industrial Health 1997, 35, 301-324
HEALTH EFFECTS OF TRICHLOROETHYLENE
longer. None of the control cohort was diagnosed as having
kidney cancer. The SIR calculated with the data of the Danish
cancer registry was 7.97 (2.59-18.59). The SIR for kidney
cancer, as calculated with the data of cancer registry in the
former West Germany, was 9.66 (3.14-22.55). After the end
of the observation period, two additional kidney tumors (one
renal cell and one urothelial cancer) were diagnosed among
the members of the exposed cohort. Based on the results of
this study, Henschler et al. concluded that the incidence of
kidney cancer was significantly elevated among workers
exposed to TRI. Nevertheless, none of the epidemiological
studies concerning TRI-related cancer have reported
relationship between TRI and kidney cancer2-4, 6,x,10-12)
Henschler et al. have emphasized that their exposed cohort
was exposed to TRI at high levels for prolonged periods of
time and that their study differed from other studies in that
the incidence (SIR) was used for evaluation in place of the
mortality (SMR). However, no relationship between
exposure to TRI and kidney cancer was found in the studies
of Axelson et al.') or Anttila et al.g) who also used the SIR
as an evaluation parameter.
In summary, a negative view is predominant
epidemiologically with respect to the carcinogenicity of TRI.
However, no unanimous conclusion has been reached
concerning the carcinogenicity of TRI in humans because
epidemiological studies conducted to date have the following
drawbacks: the cohort sizes were small, the durations of
observation were short, and the assessments of the exposure
to TRI were insufficient.
In 1995, the International Agency for Research on Cancer
(IARC)13) changed the carcinogenicity classification of TRI
from Group 3 (not classifiable as to carcinogenicity to
humans) to Group 2A (probably carcinogenic to humans),
stating that there is limited evidence to support the
carcinogenicity of TRI to humans. The Committee for
Occupational Exposure Limits of the Japan Society for
Occupational Health (JSOH)14), commenting on the IARC's
carcinogenicity classification of TRI, issued its own proposal,
which is summarized as follows: "In the evaluation of IARC
concerning the carcinogenicity of TRI, the description given
in 198715)is that while evidence for the carcinogenicity to
humans is inadequate, there is limited evidence for the
carcinogenicity to animals. On the other hand, the description
given in 199513)is that while evidence for the carcinogenicity
to humans is limited, there is sufficient evidence for the
carcinogenicity to animals. TRI was classified as Group 3
and Group 2A on the basis of overall evaluation in 1987
and 1995, respectively. This indicates that the overall
305
evaluation is based mainly on the epidemiological findings.
In this connection, placing great stress on the cohort studies,
the IARC cited the reports of Garabrant et al.5), Spirtas et
a1.6),Axelson et al.'), and Anttila et al.8), and attached
particular importance to the reports of Spirtas et al., Axelson
et al., and Anttila et al. (when revising its classification of
TRI carcinogenicity,the IARC did not take into consideration
the study of Henschler et al.9) for the reason that the study
was initiated after the observation of a cluster of renal cancer
cases). The results of the four studies are summarized by
the IARC13)as shown in Table 2. The agency put particular
a stress on the finding that the SIR or SMR for hepatobiliary
cancer and non-Hodgkin lymphoma exceeded 1.0 in the
studies of Spirtas et al., Axelson et al., and Anttila et al.
However, the lower confidence of the 95% confidence
interval of the SIR or SMR for such tumors was below 1.0.
Anttila et al. affirmed the carcinogenicity of TRI, whereas
Spirtas et al. and Axelson et al., unlike the IARC, did not
reach any affirmative conclusion. The JSOH thus judged
that it is too early to classify the TRI carcinogenicity as
Group 2A and that it is appropriate for the moment to promote
exposure control under a classification as Group 2B."
(2) Animal experiments
The U.S. National Cancer Institute (NCI)16)first published
the results of a carcinogenicity experiment in 1976. It was
found that TRI caused hepatocellular carcinoma in B6C3F1
mice. However, since the TRI used in the experiment
contained the stabilizer epichlorohydrin, a known mutagen
and carcinogen, the findings of the experiment were open
to question. In this connection, the U.S. National Toxicology
Program (NTP)17) conducted new carcinogenicity tests of
TRI. TRI not containing stabilizers was dissolved in corn
oil and given orally to F344/N rats (50 of each sex) and
B6C3F1 mice (50 of each sex), 5 days/week for up to 103
weeks. Based on the results of a preliminary experiment,
the dosage was set at 0, 500 or 1,000 mg/kg for rats and 0 or
1,000 mg/kg for mice. A group of 50 male and 50 female
rats were used as untreated control rats.
The survival of the TRI-treated F344/N rats was
significantly reduced (p<0.005) in comparison with the
untreated controls. An increased incidence of renal
adenocarcinomas was seen in males treated at 1,000 mg/kg
dose (p<0.05). Two males that received the 500 mg/kg dose
developed renal adenomas. The incidence of tumors in
female rats was not increased at any site. The fact that the
survival rate of treated rats was low (with 20% of deaths
due to failure in forced administration) makes the
306 T KANEKO et al.

Table 2. Summary of data from four cohort studies of trichloroethylene (IARC13))

interpretation of the results obtained with rats difficult.
Histopathological examination revealed increased incidences
of hepatocellular carcinomas in the TRI-treated B6C3F, mice:
in the males, hepatocellular carcinomas were found in 8/48
controls and 31/50 treated animals (p<0.001), and in females,
the carcinomas occurred in 2/48 controls and 13/49 treated
animals (p<0.05). The increase of hepatocellular adenomas
in TRI-treated mice was also noted (7148 vs. 14150 in males,
p<0.05; 4/48 vs. 16/49 in females, p=0.001).
Fukuda et al.'8 exposed female ICR mice and female
Sprague-Dawley rats to air containing TRI at 0, 50,150, or
450 ppm 7 hr/day, 5 days/week for up to 104 weeks, and
performed necropsies at the 107th week. The mean number
of lung tumors per animal was 3--4 times higher in mice
exposed to TRI at the concentration of 150 or 450 ppm than
in control mice. The incidence of lung adenocarcinomas
was significantly higher in these exposed mice than in the
control mice (p<0.05);1/49 for the 0 ppm, 8/50 for the 150
ppm, and 7146 for the 450 ppm groups. The tumor incidence
was not increased in any organs or tissues of the TRI-exposed
rats. The TRI used by Fukuda et al. contained
epichlorohydrin as a stabilizer. However, its concentration
was low; 0.04 and 0.12 ppm at TRI concentrations of 150
and 450 ppm, respectively. Although the possibility of a
combined effect of epichlorohydrin with TRI cannot be ruled
out entirely, the lung adenocarcinomas in mice in this
experiment are likely to have been induced by TRI. This
report is the first that indicated a correlation between inhaled
TRI and lung cancer in animals.
The experimental findings described above can be
summarized as follows: they provide ample evidence to
indicate that TRI promotes the development of liver cancer
in mice (B6C3F1 mice in particular) but no evidence of this
compound causing the cancer in rats. There is no report
that TRI caused kidney tumors in mice, but a possibility
that this compound causes such tumors in male rats is
suggested. All of the TRI carcinogenicity tests in animals
revealed distinct species differences (between mice and rats).
However, the IARC13~has concluded that there is sufficient
evidence for the carcinogenicity of TRI to animals.

Industrial Health 1997, 35, 301-324

HEALTH EFFECTS OF TRICHLOROETHYLENE
Fig. Metabolic pathways of TRI (adapted from the IARC13~ with modifications)
(3) Mechanism of species differences in TRIcarcinogenicity
with special reference to its metabolism
A major portion of TRI absorbed into living bodies is
metabolized by cytochrome P450 systems (probably P450
2E119))to chloral hydrate (CH), which is then oxidized to
trichloroaceticacid (TCA) or reduced to trichloroethanol(TCE)
(Fig.). TCA and TCE are excreted in the urine either directly
(TCA) or after conjugationreaction(TCE). An epoxide (1,2,2-
trichlorooxyrane) was once postulated as an intermediate
during the oxidation of TRI to CH; however, later studies
on the biotransformation of TRI and other chlorinatedolefines
suggest a stepwise oxidation of TRI to CH, in which the
epoxide is not an obligatory intermediate2o,21>
Reduced glutathione (GSH), which is present in high
concentrations in most living cells and participates in a variety
of vital cellular reactions, protects cells from potentiallytoxic
electrophiles formed via the metabolism of xenobiotics. Such
reactions involving GSH have long been associated with
the process of detoxification. However, in recent years,
evidence has accumulated indicating that GSH conjugation
plays an important role in the formation of toxic metabolites
from a variety of chemicals includingTRI22>. A minor portion
of absorbed TRI is thought to be converted, via conjugation
307
with GSH, into either cytotoxic, genotoxic, or mutagenic
metabolites23~.According to Dekant24~,TRI is metabolized
to S-(1,2-dichlorovinyl) glutathione in the liver by GSH S-
transferase present in the microsomal or soluble fraction.
The conjugate is converted to S-(1,2-dichlorovinyl)-L-
cysteine, a cysteine S-conjugate, by the removal of glutamic
acid and glycine. This conjugate is transported to the kidney
and activated by f3-lyase to produce highly electrophilic
chlorothioketenef which can acylate the macromolecules,
producing nephrotoxicity (Fig.). Since mercapturic acids
N-acetyl-S-(1,2-dichlorovinyl)-L-cysteine and N-acetyl-S-
(2,2-dichlorovinyl)-L-cysteine have been identified in the
urine of workers exposed to TRI25~, humans may have the
capacity to form GSH conjugates.
It is clear that TRI orally administered at a high dose
induces hepatocellular carcinomas in mice. However, the
carcinogenicity of TRI in rats has not been demonstrated in
any carcinogenicity tests. Many attempts have been made
to elucidate the underlying mechanism of the species
differences in TRI carcinogenicity, focusing mainly on the
differences in TRI metabolism between mice and rats.
Stott et a1.26>
exposed mice (B6C3F1) and rats (Osborne-
Mendel) to TRI by inhalation for 6 hr at a concentration of
308
10 or 600 ppm. Although there was no species difference
between the mice and rats at the exposure concentration of
10 ppm, at the exposure level of 600 ppm, the amount of
TRI metabolized was 262% larger in the mice than in the
rats with the binding of TRI to macromolecules in the liver
being 322% greater in the mice than in the rats. In the same
report, the oral administration of TRI caused hepatocellular
toxicity in repeated dosing trials in mice. The hepatic effects
observed in mice treated with a maximum tolerated dose of
2,400 mg/kg/day of TRI for three days were primarily a
centrilobular hepatocellular swelling with focal
hepatocellular necrosis. These effects led to an enhanced
regenerative process as indicated by the increased hepatic
DNA synthesis activity (220% of control) and the incidence
of mitotic cells. In contrast to the mouse data, rats appeared
to be less sensitive to a maximum tolerated dose level of
TRI (1,100 mg/kg/day), displaying enhanced hepatic DNA
synthesis levels (175% of control) with no histopathology.
From these results, Stott et al. concluded that an epigenetic
mechanism of tumor formation is operative in B6C3F1 mice,
implying that a tumorigenic response to TRI exposure in
this species would only be evident upon chronic
administration of high, cytotoxic dose levels of TRI.
The genetic mechanism of carcinogenesis is characterized
by a direct interaction (alkylation or intercalation) of a
chemical with DNA, while epigenetic mechanisms of
carcinogenesis do not involve a direct interaction of a
chemical with DNA. An epigenetic mechanism that
characterizes the tumorigenic action of TRI in B6C3F1 mice
may be that of recurrent cytotoxicity. This mechanism of
tumorigenicity involves the enhancement of the normal
cellular mutation rate during regenerative activity (increased
DNA synthesis) in response to a cytotoxic challenge by TRI.
Thus, a tumorigenic response of animals to TRI is observed
only after prolonged exposure to toxic dose levels. Another
possible epigenetic mechanism is that TRI causes a
hypertrophic response in the liver and only enhances
preexisting oncogenic changes present in B6C3F1 mice.
Prout et a1.27~
gave [14C]TRI orally to rats (Osborne-Mendel
and Alderley Park Wistar) and mice (B6C3F1 and Swiss-
Webster) at doses of 10-2,000 mg/kg and observed the
toxicokinetics of TRI and its metabolites. The metabolism
of TRI in the mice was linear over the range of doses used,
whereas in the rats it became constant independent of dose
at 1,000 mg/kg and above (metabolism saturation). TRI given
at a dose of 10 mg/kg was almost completely metabolized
in both rats and mice, 60% of the dose being excreted in
urine with only 1 to 4% being eliminated unchanged in
T KANEKO et al.
expired air in the first 24 hr. At 2,000 mg/kg, however,
78% of the dose was eliminated unchanged in the rats, but
only 14% in the mice. The area under the concentration-
time curve (AUC) of TRI and its metabolites in the blood
calculated at a dose of 1,000 mg/kg was greater in mice
than in rats: the ratio of AUC in the mice relative to that in
the rats was 7.9 for TRI, 3.9 for trichloroethanol (TCE),
and 6.3 for trichloroacetic acid (TCA). This finding indicates
that the mice were exposed to significantly higher
concentrations of TRI metabolites than were the rats at high
dosages. In particular, the blood TCA levels in mice remained
high for over 30 hr. A high blood concentration of TCA is
known to induce hepatic peroxisome proliferation in mice
but is insufficient to induce this response in rats28~. The
results of this study suggest a link between TCA-induced
proliferation of peroxisomes and liver cancer28~,which forms
a basis of the species difference in response to TCA.
According to the study conducted by Elcombe et al.28>,
TRI administered to B6C3F1 and Alderley Park mice by
gavage for 10 consecutive days at doses of 500 to 1,500
mg/kg increased both the liver weight (175% of control)
and the synthesis of DNA (500% of control) in the liver. In
contrast, similar treatment of Osborne-Mendel rats and
Alderley Park rats resulted in a smaller increase in the liver
weight (130% of control) with no effect of TRI on the DNA
synthesis in either strain of rats. The increased DNA synthesis
in mice was probably not due to regenerative hyperplasia,
since no signs of necrosis were seen in these animals. While
the increased liver weight in rats appeared to be due to liver
cell enlargement (hypertrophy), both hypertrophy and
hyperplasia (cell proliferation) were operative in the increase
in mice; an increased number of mitotic cells (10 times the
mitotic index of control) was observed in parallel with the
increase in DNA synthesis in the TRI-treated mice. An
important observation was that TRI induced the peroxisomal
enzyme activities, catalase and cyanide-insensitive palmitoyl-
CoA oxidation (147 and 786% of control, respectively), in
mice but not in rats. Furthermore, increases in peroxisome
volume density (up to 1,110% of control) were observed in
the mice that received TRI. These observations suggest that
the species difference in the hepatocarcinogenicity of TRI
is due to a species difference in peroxisome proliferation
and cell proliferation, the peroxisome proliferation leading
to increased reactive oxygen species and DNA damage, and
the cell proliferation then acting to promote this lesion.
Nelson and Bu1129~evaluated the ability of TRI and its
metabolites to induce single-strand breaks in the hepatic DNA
of male B6C3F1 mice and Sprague-Dawley rats in vivo using
Industrial Health 1997, 35, 301-324
HEALTH EFFECTS OF TRICHLOROETHYLENE 309

an alkaline unwinding assay. Doses of TM of 22-30 mmol/ suggests that a metabolic pathway from TRI to DCA may
kg were required to produce strand breaks in DNA in rats, exist only in mice. The blood DCA concentrations observed
whereas a dose of 11.4 mmol/kg was sufficient to increase in the mice given a carcinogenic dose of TRI (15 mmol/kg)
the rate of alkaline unwinding in mice. The TRI metabolites were of the same magnitude as those observed with
TCA, dichloroacetic acid (DCA) and chloral hydrate (CH) carcinogenic doses of DCA. In contrast, when TCA was
induced strand breaks in hepatic DNA in a dose-dependent given to animals, DCA was detected in the blood of both
manner in both species. The slopes of the dose-response species and the blood DCA level was higher in the rats than
curves and the order of potency of these metabolites differed in the mice. Based on these findings, Larson and Bull
significantly between rats and mice, suggesting that different suggested that it is highly likely that both TCA and DCA
mechanisms of single-strandbreak inductionmay be involved play a role in the induction of hepatic tumors in B6C3F1
in the two species. In addition, the dose of metabolites mice by TRI.
required to induce strand breaks was lower in mice than in The hepatocarcinogenicity of TRI is likely due to an
rats. In particular, a 100-fold difference was noted between epigenetic mechanism (promoter) rather than a genotoxic
the two species in the minimum dose of TCA required to mechanism (initiator). Liver cancer develops after TM has
induce the breaks. This finding coincides with the report that been given over extended periods of time at doses high
TCA is liable to cause peroxisomal proliferation in mice28'30) enough to produce cytotoxicity. However, the view that
The study of Nelson and Bull is another explanation for the the cancer is caused by a genetic mechanism cannot be ruled
different sensitivityof mice and rats to the hepatocarcinogemc out entirely since the binding of TRI to DNA was
effects of TRI. demonstrated in vitro in the presence of a metabolism-
Intercellular gap junctions are comprised of proteinaceous activating system21~,and since an increased induction of
plasma membrane channels that link the interiors of adjacent micronuclei36~ and DNA single-strandbreaks29> were observed
cells and permit cells to directly exchange small (<1,000 in vivo (mice and rats).
Daltons) molecules and ions. One cellular effect exhibited Why does the site of tumor development by TRI depend
by a number of nongenotoxic carcinogens and tumor on the route of administration? The oral administration of
promoters is the inhibition of gap-junction mediated TRI caused liver cancer in mice17~, whereas the main tumors
intercellular communication31~.Klauning et al.32~assessed caused in mice exposed to TRI by inhalation were pulmonary
the effects of TRI and its metabolites (TCA, TCE, and CH) tumors18~.The carcinogenicity of TRI is mediated not by
on gap-junction mediated intercellular communication in TRI itself but by its metabolites, DCA or TCA. The activity
cultured B6C3F1mouse and F344 rat hepatocytes. TRI and of cytochrome P450 enzymes that catalyze the metabolism
TCA inhibited intercellular communication in the mouse is greatest in the liver, with only weak activity in the lung
hepatocytes but not in the rat hepatocytes. TCE and CH and kidney. After oral administration, all of the amount of
had no effect on hepatocyte intercellular communication in TRI absorbed passes through the liver where TRI is processed
either rat or mouse cells. The inhibitory action of TCA was (first-pass metabolism), and the liver cells come into contact
strongerthan that of TRI. In addition, TRI appeared to require with active metabolites at high concentrations. In contrast,
cytochrome P450 metabolism by the mouse hepatocytes to TRI exposed to animals by inhalation is first metabolized
exhibit its inhibitory effect on the communication. Thus, in the pulmonary tissues, and it may thus cause injuries in
the inhibitory action of TRI was likely to be produced by the tissues if sufficient amounts are metabolized.
the metabolite TCA. A difference in the metabolic capacity In summary, the most probable explanation for the reason
between mice and rats (TRI metabolism, mice>rat25~)may why TRI-induced hepatocellular carcinoma occurs in mice
also be involved in the difference in the inhibitory action but not in rats is that the activity of cytochrome P450 enzymes
between both species. The findings of Klauning et al. also in the liver of mice is higher than in rats. The occurrence of
provide an explanation for the species differences in liver cancer depends on the enzyme activity, especially when
susceptibility to TRI-induced liver carcinogenesis. TRI is given to animals at a high dosage. This is because
There are reports that DCA, a metabolite of TRI, shows the metabolism in the liver is rate-limited by the enzyme
carcinogenicity in B6C3F1 mice33'34).In experiments with activity (capacity-limited metabolism) at such high doses37~.
rats (Sprague-Dawley)and mice (B6C3Fr),Larson and Bull35> Another reason is that the B6C3F1 mice used in many
demonstrated that DCA was detected in the blood of mice carcinogenicity tests have spontaneously occurring liver
but not in rats following TRI administration, a finding which cancer with high frequency.
310 T KANEKO et al.

(4) Mutagenicity to an unusually high MN frequency in the controls. A

After examining short-term mutagenicity test data in the subsequent replication of the 1-day 5,000 ppm TCE exposure
published literature, Jackson et al.38~summarized that TRI with rats again showed a highly significant increase in MN
containing no mutagenic stabilizers (e.g., epichlorohydrin, frequencies compared to controls.
1,2-epoxybutane, etc.) is unlikely to induce gene or Konietzko et al.43~reported a pathological rate of
chromosomal mutations or aneuploidy. hypodiploid cells in the lymphocytes of male workers
Metabolites of TRI were demonstrated to bind to DNA exposed to a high level (206 ppm) of TRI. There was a
in in vitro systems21' 39,40)According to Miller and significant correlation between the rate of hypodiploid cells
Guengerich20' 21>,TRI oxide is probably not the TRI metabolite and TRI exposure concentration (r=0.46, p<0.05). The rate
responsible for the irreversible binding to DNA. Concerning was 10.9% for the exposed group (27 workers) and 6.5%
the binding in a living system, while one report suggested for the unexposed group (10 workers). In addition, the rate
slight binding to hepatic DNA41>,the others were unable to of chromosomal breaks in mitotic cells was 5 times higher
give direct evidence of the formation of TRI-DNA adducts in the exposed group than in the unexposed group. However,
in the liver39,40~. a major drawback of this report is that the effect of smoking
As was discussed earlier, Nelson and Bull29>demonstrated was not taken into consideration.
the ability of TRI to induce single-strand breaks in the hepatic Nagaya et al.44~compared the frequency of SCE in the
DNA of male B6C3F1 mice and Sprague-Dawley rats in vivo, peripheral lymphocytes between 22 workers occupationally
using an alkaline unwinding assay. The strand breaks were exposed to TRI (mean TCA concentration in urine: 183.6
observed at doses that produced no observable hepatotoxic mg/l) and 22 control workers matched by age, sex and
effects as measured by serum aspartate aminotransferase smoking history. Although the smokers had a high frequency
and alanine aminotransferase levels. However, since strand of SCE, no difference in the smoking-adjusted frequency
breaks are not always induced by direct interaction between was found between the two groups. Seiji et al.45~ examined
chemical compounds and DNA42~,the finding of Nelson and the frequency of SCE in peripheral lymphocytes from 38
Bull does not necessarily mean that TRI acts as an initiator workers exposed to TRI at 7 ppm and 19 workers exposed
of the carcinogenicity. to a mixture of TRI and tetrachloroethylene (TETRA) (8
Kligerman et a1.36~performed inhalation studies using male and 17 ppm, respectively) (TRI + TETRA). The results
C57BL/6 mice and CD rats to determine whether TRI can were compared with the findings in control subjects matched
induce cytogenetic damage in vivo. Animals were exposed by age, sex, smoking habits and place of residence. No
to target concentrations of 0, 5, 500, or 5,000 ppm TRI significant increase in SCE frequencies was observed in
(reagent grade) for 6 hr. Tissue samples were taken between association with the exposure to TRI or TRI + TETRA.
18 and 19 hr after exposure. Cultured peripheral blood Cysteine S-conjugates, TRI metabolites via the GSH
lymphocytes (PBL) in rats and splenocytes in mice were pathway of TRI metabolism, have long been known to bind
analyzed for the induction of sister chromatid exchanges to DNA, and the conjugate from TRI is mutagenic in the
(SCE), chromosomal aberrations, and micronuclei (MN) in Ames bacterial mutation assay47~.However, according to
cytochalasin B-blocked binucleated cells. Bone marrow Green et al., kidney f3-lyaseactivity is far lower in humans
polychromatic erythrocytes (PCE) were analyzed for MN. than in mice and rats. It is thus uncertain whether the GSH
The only positive response observed was for MN in rat bone pathway in the metabolism of TRI has any relevance to TRI
marrow PCE. TRI caused a significant increase in MN at carcinogenicity in humans.
all concentrations, inducing an approximate four-fold increase
over the control levels at 5,000 ppm. TRI was also cytotoxic Toxicity Other than Carcinogenicity
in rats, causing a significant concentration-related decrease
in the ratio of PCE/normochromatic erythrocytes. In follow- The acute toxicity of TRI originates from its skin- and
up studies, CD rats were exposed for 6 hr/day over 4 mucous membrane-irritating action and anesthetic action.
consecutive days to 0, 5, 50 or 500 ppm TRI. No significant As is clear from the fact that TRI was formerly used as an
concentration-related increases in cytogenetic damage were inhalational anesthetic, it is a strong central nervous system
observed. While the MN frequencies in the 4-day study depressant. As the exposure level is increased, cognitive
were comparable to those at the equivalent concentrations ability and associated movement are affected together with
in the 1-day study, they were not significantly elevated due eye irritation, sleepiness, headache, and malaise. When a

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HEALTH EFFECTS OF TRICHLOROETHYLENE
human is exposed to an extremely high level of TRI, its
anesthetic action may result in death. However, when a
person is acclimated to an odor specific to TRI, such
symptoms appear only rarely during exposure at a
concentration of 50 ppm or lower.
Regarding the health effects of chlorinated hydrocarbons,
hepatotoxicity has been a matter of great concern. However,
TRI is weakly hepatotoxic relative to such hepatotoxicants
as carbon tetrachloride and chloroform49~Nakajima et al.5o~
exposed rats to TRI by inhalation for 2 hr at a concentration
of 500-8,000 ppm and found that the alanine
aminotransferase activity in the plasma, an indicator of liver
function disorder, was not significantly increased until the
concentration was raised to 4,000 ppm.
The health effects of long-term exposure to TRI are chiefly
manifestedas neuropathy. Most of the individuals chronically
exposed to a certain level of TRI present subjective symptoms
related to the nervous system such as headache, sleepiness,
dizziness, malaise, nervousness, finger tremor, nausea, and
anorexia, for example. These symptoms appear in workers
exposed to TRI at 50 ppm or higher for extended periods of
timed-53~,
(1) Neurotoxicity
Rasmussen et al.54~examined the clinical neurological
manifestations after long-term exposure to degreasing
solvents, mainly TRI. Seventy metal degreasers exposed
to TRI at a relatively constant level (mean exposure
concentration in terms of urinary TCA level: recent years,
7.7 mg/l; 1950s-1970s, 40-60 mgll) were divided into three
groups according to the length of working years, i.e., low-
exposure group (mean 0.5 year), mid-exposure group (2.1
years) and high-exposure group (11.0 years). The most
marked finding was a highly significant dose-response
relation between solvent exposure and motor
dyscoordination, a finding that was retained after a
multivariate control of relevant confounders. Vibration
thresholds increased by solvent exposure at a bivariate level,
but the multivariate analysis showed that age explained most
of the increase. No significant trigeminal nerve dysfunction
was found.
Ruijten et a1.55~
performed neurological tests in 31 printing
workers exposed to TRI (mean age, 44 years; mean duration
of exposure, 16 years; cumulative exposure level, 704 ± 583
ppm-year) and 28 controls (mean age, 45 years). A slight
reduction (-1.1 mis) in the sural nerve conduction velocity
and a prolongation (0.4 ms) of the sural nerve refractory
period (mean of the controls 1.95 ms) were found. The
311
latency of the masseter reflex (mean 10.4 ms) had increased
(0.4 ms). With respect to the blink reflex, no prolongation
was found. No impairment was found in the functions of
motor and autonomic nerves. Based on these results, Ruijten
et al. concluded that long-term exposure to TRI at 35 ppm
may slightly affect the trigeminal and sural nerves. This
study also suggests that the refractory period is a sensitive
indicator of preclinical toxic neuropathies.
Feldman et al.56>
evaluated the use of the electrophysiologic
blink reflex as an indicator of neurotoxic effects of TCE in
occupationally exposed workers. The blink reflex was tested
in individual cases with documented histories of exposure
to TRI (n=18). When compared with the nonexposed
laboratory control values (n=30), the subjects with a
significant history of TRI exposure demonstrated the most
prolonged latencies (greater than or equal to 3 SD above
the nonexposed group mean) in the R 1 component of the
blink reflex measurement. Considering a latency value above
the mean ± 3 SD of the control group to be positive, the
blink reflex latency test allowed to detect the effect of TRI
on the trigeminal nerve with 50% sensitivity and 90%
specificity. The results of this study indicate that the
electrophysiological blink reflex has application in assessing
TRI exposure and in documenting the neurotoxic effects of
its exposure on trigeminal nerve functions in humans.
Liu et al.53~
examined 103 workers exposed to TRI (mostly
at less than 50 ppm) and 111 non-exposed control workers
for subjective symptoms, hematology, serum biochemistry,
and glucose, protein and occult blood in the urine. The
clinico-laboratory test results in the TRI-exposed group were
essentially normal, and no clinically significant effects of
TRI exposure were found in the blood or liver functions
among the exposed workers as compared with the controls.
The prevalence of the subjective symptoms was, however,
significantly higher in the exposed group than in the controls,
and a dose-response relationship was established in some
selected symptoms such as nausea, heavy feeling in the head,
forgetfulness, tremor in extremities, cramps in extremities
and dry mouth. These findings warrant further attention to
the effects of TRI, especially on the central and autonomic
nervous systems at concentrations lower than 50 ppm.
Taken together, the findings described above indicate that
the chronic effects of TRI exposure at low concentrations
present as neurological changes. In addition to neurological
tests such as the measurement of nerve conduction velocity
and its refractory period, subjective symptoms are useful
for detecting such neurological effects.
312
(2) Hepatotoxicity
TRI-induced liver injuries are discussed in the above
section, "Mechanism of species differences in TRI
carcinogenicity with special reference to its metabolism."
Hence, only the hepatic effect of occupational exposure to
TRI will be discussed here.
Takamatsu divided degreasing work with TRI into three
categories: high-exposure (150-250 ppm), mid-exposure
(50-100 ppm), and low-exposure (<50 ppm)50).TRI-related
liver function disorders were found in the workers engaged
in the high- and mid-exposure works, but not in the workers
engaged in the low-exposure work.
Liu et al.53~examined 101 workers exposed to TRI (<50
ppm) and 111 non-exposed control workers for serum
biochemistry, i.e., the activities of alanine aminotransferase
(ALT), aspartate aminotransferase (AST), y-
glutamyltranspeptidase(y GTP), alkaline phosphatase (ALP),
lactate dehydrogenase (LDH) and leucine aminopeptidase
(LAP) in the serum, the thymol turbidity test (TTT), the
zinc turbidity test (ZTT), and the total bilirubin content in
the serum. Significant differences (in either the mean of
values or the rate of abnormal/total cases) were observed
only in the activities of ALT,LAP and LDH. The deviations
from the normal ranges of liver function indicators were all
slight and of subclinical levels, except for two cases in which
both ALT and AST were elevated to a degree of clinical
significance (ALT 115; AST 75 Karmen units) in a 25-year
old nondrinking exposed man, and to subclinical levels (ALT
24; AST 32 Karmen units) in a nondrinking exposed man at
the age of 48 years. Since all the values in the other liver
function tests were essentially within the corresponding
normal ranges and there was no significant difference in
either the mean or the prevalence of abnormalvalues between
the exposed and control groups, the authors concluded that
the slight deviation from normal ranges were not associated
with TRI exposure.
Rasmussen et al.57~examined the dose-response
relationships between long-term exposure to TRI and a
battery of liver function tests among 99 metal degreasers.
The study group was divided into four groups with nearly
equal numbers of workers accordingto a cumulative exposure
index calculated using the number of hours of degreasing
work, multiplied by the number of years of exposure,
multiplied by 45 working weeks per year, i.e., group I
(reference group) less than one year, group II 1-2.8 years,
group III 2.9-6.7 years, and group IV 6.8-35.6 years. The
mean urinary TCA level in the highest exposure group was
7.7 mg/1with a maximum TCA level of 26 mg/l. The activity
T KANEKO et al.
of y glutamyltransferase in the serum was found to be
elevated concomitantly with increasing solvent exposure.
The significance of this relationship, however, was not able
to withstand a multiple regression analysis, with age and
alcohol abuse as confounding variables. The study indicates
no significant association between occupational TRI exposure
at levels lower than the current permissible exposure level
and the results of tests screening for early liver dysfunction.
To examine the early hepatic effects of chronic exposure
to low-level TRI, Nagaya et al.58>determined the total
cholesterol (T C) and high-density-lipoprotein cholesterol
(HDL-C) contents and the activities of AST, ALT and y-
GTP in the sera of 148 workers (a cross-sectional study)
and of 13 workers (a 2-year follow-up study) occupationally
exposed to TRI. In the cross-sectional study, three exposure
groups were defined by urinary total trichloro-compounds
(TTC) levels; a low-exposure group (TTC<10 mg/g creatinine,
n=49), a moderate-exposure group (10<-TTC<100, n=56),
and a high-exposure group (TTC>100, n=43). With
increasing exposure levels, the T-C and HDL-C levels were
slightly but not significantly (p=0.143 and 0.080 by ANOVA,
respectively) increased. The exposure, however, had no effect
on the activities of the three serum enzymes. In the follow-
up study, the fluctuations in TTC were well reflected in
subclinical changes in HDL-C, AST, and y-GTP, but not in
T -C or ALT. Based on these results, the authors suggested
that exposure to low-level TRI influences hepatic functions,
affecting cholesterol metabolism rather than causing hepatic
cell damage, and that these influences are subclinical and
reversible.
(3) Nephrotoxicity
In the TRI carcinogenicity test conducted by the U.S. NTP
with B6C3F1 mice and F344/N rats17~,renal adenocarcinoma
developed solely in male rats. In the study of Maltoni et
al.S9~,TRI was administered by inhalation to rats (Sprague-
Dawley) and mice (Swiss and B6C3F1) 7 hr/day, 5 days/
week, for 8 or 104 weeks at various concentrations. The
animals were kept under observation until spontaneous death.
The most relevant finding was the dose-related increased
incidence of Leydig cell tumors in male rats, and the onset
of few renal adenocarcinomas at the highest dose (600 ppm),
always in male rats. The renal adenocarcinomas were
preceded by and associated with a characteristic lesion of
the kidney: tubular cell karyomegaly (megalonucleocytosis).
Protein (hyaline) droplet nephropathy has been linked to
the a2-microglobulin specifically occurring in male rats60~.
Goldsworthy et al.61~ attempted to determine whether
Industrial Health 1997, 35, 301-324
HEALTH EFFECTS OF TRICHLOROETHYLENE
chlorinated hydrocarbon-induced protein droplet
accumulation was related to the male rat specific protein,
a2-microglobulin. Male and female F-344 rats were gavaged
for 10 days with TRI (1,000 mg/kg), tetrachloroethylene
(TETRA, 1,000 mg/kg) or pentachloroethane (PENTA, 150
mg/kg). Protein droplet accumulation in the cytoplasm of
the P2 segment of the proximal tubule was evident in the
male rats administered TETRA and PENTA, whereas the
droplet accumulation in the male rats treated with TRI did
not differ from that in the control rats. Immunohistochemical
staining for a2-microglobulin revealed a marked correlation
between the presence of a2-microglobulin and the protein
droplets. In addition, cell replication rates increased
specifically in the histologically damaged P2 segments of
the male rats treated with TETRA or PENTA,a fmding which
suggests that the increase in cell replication may be linked
to the tubular damage caused by male rat-specific a2-
microglobulin. Since compensatorycell divisionis postulated
to affect all of the stages of the carcinogenic process, the
increased incidence of renal tumors in male rats after TETRA
or PENTA treatment may be related to the protein droplet
nephropathy specifically occurring in male rats. The results
of the study of Goldsworthy et al. suggest that the
mechanisms involved in TCE-induced renal carcinogenicity,
if any, may differ from TETRA- and PENTA-induced renal
carcinogenicity.
Case reports on nephropathy induced by TRI have been
published sporadically. For example, David et al.62~ reported
a 34-year-old patient with acute renal failure that developed
after the completion of an 8-hr computer ribbon-cleaning
job using TRI. On admission (3 weeks after the exposure),
the patient presented a remarkable foot edema, proteinuria,
and a conspicuous degree of hematuria and pyuria. Renal
tubular necrosis secondary to allergic interstitial nephritis
was histologically established by biopsy. The solvent used
for the work was TRI of 99.5% purity, but the details of
TRI exposure were unknown (David et al. estimated the
exposure concentration to be 166-3,700 ppm).
A comparison was made concerning the urine levels of
total protein and f3 -microglobulinbetween 104 male workers
occupationally exposed to TRI (15 ppm) and 102 male
nonexposed controls63~.The urinary total protein levels of
the exposed group, rather than the urinary fl-microglobulin
level, was slightly higher than those of the controls. However,
the difference in total protein level between the two groups
was not significant. The mean total protein level for the
35- to 44-year-old workers (62.5 ± 21.3 mg/g creatinine;
n=23) was significantly higher than that for corresponding
313
controls (50.6 ± 17.0 mglg creatinine; n=20). These results
suggest that the adverse effects of occupational exposure to
TRI on the kidney, if any, are mild and glomerular rather
than tubular.
(4) Pulmonary toxicity (including association with lung
cancer)
Forkert and Birch64>examined the time course of TRI-
induced pulmonary injuries, focusing on the morphological
changes and the covalent binding of [14C]TRIsoon after
administration of a single dose of TRI (2,000 mg/kg) to CD-
1 male mice. At one hour after treatment, Clara cells of the
bronchiolar epithelium exhibited necrotic changes involving
the mitochondria and endoplasmic reticulum. Dilatation of
the endoplasmic reticulum became more severe at two hours
after TRI administration, and by 4 hr, distended cisternae
coalesced to form small vacuoles within the cytoplasmic
matrix of the Clara cells. The severity of cellular damage
increased between 8 and 12 hr, and by 24 hr, the majority of
Clara cells within the airway were severely vacuolated. The
covalent binding of [14C]TRIto lung macromolecules was
evident at one hour and peaked at four hours. The peak
binding (142.6 ± 31.8 nmollg of wet weight) represented
approximately 20% of [14C]TRI distributed to the lung.
Although the levels of binding in the liver were at all times
greaterthan those in the lung, the liver injuries were relatively
insignificant. The results demonstrated a positive correlation
between the onset of Clara cell injury and the formation of
reactive metabolites, as assessed by the covalent binding of
[14C]TRI. The selective damage to Clara cells by TRI
suggests that TRI undergoes metabolic activation by
cytochrome P450 existing in the cells.
When animals are exposed to TRI by inhalation, Clara cells
are directly in contact with TRI in the inhaled air together
withTRI carriedthere by the bloodstream. It is unclearwhether
pulmonarytumors, believedto be caused by TRI'8'59>, originate
from Clara cells. One reason why lung tumors are frequently
caused in mice may lie in the high frequency of spontaneous
adenomatosis in some strains of mice. It must be borne in
mind that ICR mice used by Fukuda et al.18> and Swiss mice
used by Maltoni et al.59~ have lung adenomas with relatively
high frequency. However,no data are availablefor comparing
TRI metabolism in the lung between rats and mice.
Nichols et al.6S~examined the susceptibility of pulmonary
cells and the mechanisms of cytotoxicity of TRI using
enriched subpopulations of isolated rabbit (New Zealand
white) lung cells incubated with TRI. The ability of TRI to
induce selective pneumotoxicity was evaluated by its
314
bioactivation in three cell types, i.e., Clara cells, alveolar
type II cells,and alveolar macrophages. TRI at concentrations
of 5 and 10 mM was nonselectively and dose-dependently
cytotoxic to Clara cells, type II cells and alveolar
macrophages. Since the cytotoxicity of TRI was not
ameliorated by 1-aminobenzotriazole, an inhibitor of
cytochrome P450, Nichols et al. ruled out the bioactivation
of TRI by P450. However, such a conclusion may be
premature because the concentration employed in this study
was too high (5 or 10 mM) to make a conclusion about the
metabolism inhibition.
In the study of Odum et a1.66),female CD-1 mice exposed
to TRI (6 hr/day) at concentrations of 20-2,000 ppm
developed a highly specific lung lesion after a single
exposure, characterized by vacuolation of the Clara cells,
the number of cells affected increasing with increasing dose
level. At the highest dose levels, pyknosis of the Clara cells
was apparent. After 5 days of repeated exposure, the lesion
had resolved but the exposure of mice following a 2-day
break resulted in recurrence of the lesion. The changes in
mouse lung Clara cells were accompanied by a marked loss
of cytochrome P450 activities. In contrast, no morphological
changes were seen in the lungs of rats (female A1pk:APfSD)
exposed to 500 or 1,000 ppm TRI. Isolated mouse lung
Clara cells metabolized TRI to CH, TCE and TCA. CH
was the major metabolite. TCE glucuronidewas not detected.
The activity of UDP-glucuronosyltransferase was compared
in mouse lung Clara cells and hepatocytes using two phenolic
substrates and TCE. Hepatocytes readily formed
glucuronides from all three substrates, whereas Clara cells
were only active with the two phenolic substrates. When
the three major metabolites of TRI, CH (inhalation), TCE
(inhalation) and TCA (intraperitoneal administration) were
each administered to mice, only CH had an effect on mouse
lung, causing a lesion (Clara cell) identical to that seen with
TRI. The lesions produced by CH at a concentration of 100
ppm were equivalent to the changes caused by TRI at 1,000
ppm. Based on these results, Odum et al. proposed that the
failure of Clara cells to conjugate TCE leads to an
accumulation of CH, resulting in cytotoxicity. The known
genotoxicity of CH suggests that this lesion may be related
to the development of lung tumors in mice exposed to TRI
by inhalation.
Since TRI entering the body by inhalation comes into
directcontact with the bronchioli where Clara cells containing
cytochrome P450 exist most abundantly in the pulmonary
tissues, cytotoxicity is likely to develop at the same sites.
According to Smith et al.67),the number of Clara cells is in
T KANEKO et al.
the order of mice>rats>humans, and human Clara cells lack
smooth endoplasmic reticulum, where P450 is localized.
Thus, the response of humans to TRI is likely to differ from
that of mice.
(5) Fetal toxicity (including teratogenicity)
In 1981, the groundwater for a small area in the Tucson
Valley, Arizona, U.S. was found to be contaminated with
TRI. Goldberg et al.68)compared the prevalence of congenital
heart disease in children between parents who were exposed
to TRI through TRI-contaminated wells and parents who
were never exposed to the contaminated water. The
proportion of infants with congenital heart disease as
compared with the live births was significantly higher for
mothers in the contaminated water area than for mothers
with no exposure. The odds ratio for parents of case children
was three times that for parents of control children (p<0.005).
Goldberg et al. concluded that their data suggest a significant
association but not a cause and effect relation between
congenital heart disease and parental exposure to water
contaminated with TRI.
To experimentally verify the epidemiologic phenomenon
suggesting a greater than expected number of pediatric
patients with congenital heart disease in areas where the
drinking water was contaminated by TRI, Dawson et al.69)
infused TRI as a saline solution (15 or 1,500 mg/l) through
a catheter into the gravid uterus by an intraperitoneal osmotic
pump to the developing rat fetus in utero during the period
of organ differentiation and development (7th gestation day).
Although only a very small number of congenital heart
anomalies (3%) were found in the control group (saline
solution only), the incidence of anomalies was found to be
9% and 14% in the lower and the higher dose groups,
respectively (p<0.05). While a variety of cardiac defects
appeared, only a single noncardiac anomaly was found. From
these findings, Dawson et al. concluded that TRI appeared
to be a specific cardiac teratogen. From the experiment
simultaneously conducted, Dawson et al. reported that 1,1-
dichloroethylene was a much stronger heart teratogen: the
chemical caused congenital heart anomalies almost to the
same extent as TRI at 1/10 the dose of TRI. In this
experiment, however, neither the TRI- nor the 1,1-
dichloroethylene-treated group differed from the control
group in the number of resorbed, dead or live fetuses. It
may be true that a high dose of TRI causes congenital heart
anomalies in rats. However, the animal experiment of
Dawson et al. does not provide sufficient evidence to establish
a relationship between congenital heart disease and
Industrial Health 1997, 35, 301-324
HEALTH EFFECTS OF TRICHLOROETHYLENE
groundwater contaminated with TRI in humans. TRI was
injected directly into the gravid uterus at a high dose level
in this experiment. In humans; the concentration of TRI in
drinking water is about 300 ppb at the highest, and the
compound absorbed by, pregnant mothers reaches the fetuses
after passing through the =mothers'liver where the first-pass
metabolism, :prevails.
Another epidemiologic study was conducted by Deane
et al.70~
to investigate a suspected cluster of adverse outcomes
of pregnancies conceived in 1980-1981 among women who
resided in a census tract in Santa Clara County, California
that was thought to be exposed to drinking water from a
well contaminated by TRI. A comparison census tract that
received water from a different source was selected on the
basis of demographic comparability. The cluster was
confirmed; the odds ratio for spontaneous abortion was 2.3
(95% CI 1.3-4.2) after adjustment by multiple logistic
regression for maternal risk factors, including maternal age,
alcohol consumption, smoking, and prior fetal loss. The
relative risk for congenital malformations was 3.1(1.1-10.4).
Because of the lack of precise information on the timing
and extent of contamination, the pattern of spontaneous
abortion rates throughout the study period cannot be used
to support or refute a causal inference. Subsequently, Deane
et al.71 investigated the relation between spontaneous
abortions and consumption of tapwater in the same cohort
as used in the previous study70~.The results suggested an
association between spontaneous abortions and reported cold
tapwater consumption. The observed effect was not due to
maternal risk factors, nor was it°a function of consumption
of bottled water. After controlling for bottled water, the
odds ratio for consumption of tap water was 3.4(0.6-19.4).
Windham et al.72~conducted a large case-control study
(n=1,926) in Santa Clara, California concerning the
relationship between spontaneous abortion (SAB) and
exposure to organic solvents. Exposure to solvents was
ascertained by a telephone interview that asked about the
occupational use of 18 specific solvents or products, as well
as an open-ended `other' solvent category. The adjusted
odds ratio for the use of any solvent was 1.1(95% CI 0.8-
1.5). Solvents for which at least a doubled crude risk of
SAB was found included tetrachloroethylene(4.7;1.1-21.1),
TRI (3.1; 0.9-10.4), and paint thinners (2.3;1.0-5.1). The
odds ratio for TRI was 7.7 (1.3-47.4) when low levels of
exposure (<0.5 hr/week) were excluded. Comparing
exposure greater than 10 hr per week versus less did not
show consistent dose-response effects. The household use
of solvent-containing products was generally not strongly
315
associated with SAB, nor did it appear to confound the
association seen with occupational use. Windham et al.
concluded from this study that occupational exposure to at
least some solvents appeared to be associated with SAB.
In the experimental study of Cosby et al.73~,female B6D2F1
mice were gavaged from days 1 to 5, 6 to 10, or 11 to 15
(day 1=vaginal plug) with TRI in corn oil at 0, 1/100, and
1110 of the oral LD50 (2,402 mg/kg). Litters were counted,
sexed, weighed, and measured for crown-rump length until
weaning on day 21, and some offspring were allowed to
develop to 6 weeks of age. At this time, a minimum of two
litters from each dose group were killed and the gonads were
removed, weighed, and preserved in Bouin's fixative. Litters
were also assessed for developmental abnormalities. No
maternal or reproductive effect of TRI was seen at either
dose level. In a second series of studies, TRI and its
metabolites DCA, TCA and TCE were added to culture media
to assess the toxic effects on in vitro fertilization (IVF) in
mice. TRI at a concentration of 1,000 mg/l did not affect
the success rate of IVF, whereas the metabolites decreased
the rate in a concentration-dependent manner.
Hayasaka et al.74)gave oral TRI to S1c:ICR mice at a dose
of 2 mUkg once or 1.25 mUkg twice at a 4-hr interval on
gestation days 10-13, and observed its fetotoxicity. Fetal
malformations (mainly cleft palate) developed at both dose
levels. Moreover, the weights of fetuses from mice given
TRI twice at 1.25 mUkg were significantly lower than those
from control mice. However, the body weights of pregnant
mice were concomitantly decreased by the Till administration
at either dose. Accordingly, it can be said from this study
that such a high dose of TRI as to cause general toxicity in
dams (weight loss) can produce a teratogenic effect as well
as developmental toxicity.
In the study of Coberly et al.75~,pregnant Swiss ICR mice
were treated with TRI, orally (0.1 and 1.0 µg/kg and 48.3
and 483 mg/kg) or intraperitoneally (0.01, 0.02 and 10 µg/
kg), when embryos were traversing the pronuclear stages
of development. Pregnant animals treated with the vehicle
(water or olive oil) served as the control. No treatment-
related effect was seen on the total number of embryos
recovered from oviducts at the 2- to 4-cell stage of
development. The 4-cell embryos were tested for cellular
proliferative disadvantage by the embryo chimera assay in
which embryos from treated mothers (experimental embryos)
were paired with embryos from nontreated mothers (control
embryos) to form aggregation chimeras. TRI did not affect
the development of embryos regardless of the route of
administration across all the doses examined. Since the
316
embryo chimera assay is believed to be a sensitive technique
for detecting the reproductive toxicity of chemical
compounds, Coberly et al. concluded that the reproductive
toxicity of TRI is almost negligible.
Smith et al.76~evaluated the developmental effects of TCA,
a metabolite of TRI, in pregnant Long-Evans rats. Animals
were administered TCA by oral intubation on gestation days
6-15 (plug=0) at 0, 330, 800, or 1,800 mg/kg/day.
The control rats were treated with distilled water. Maternal
parameters included clinical signs, weight change, and gross
evaluation of organ weights and uterine contents at necropsy
(day 20). There were no maternal deaths associated with
toxicity prior to sacrifice. Weight gain during treatment
was reduced at 800 mg/kg or higher. Spleen and kidney
weights were increased in a dose-related manner. The mean
percent of resorbed implants per litter was 34, 62 and 90 at
800, and 1,800 mg/kg, respectively. Live fetuses
showed dose-dependent reductions in weight and length.
The mean frequency of soft tissue malformations in the live
fetuses ranged from 9% at 330 mg/kg to 97% at 1,800 mg/
kg. These were principally in the cardiovascular system
(interventricular septal defect and levocardia). Skeletal
malformations were found only at 1,200 and 1,800 mg/kg
and were mainly in the orbit. The results of this study indicate
that TCA is developmentally toxic in the pregnant rat at
doses of 330 mg/kg and above.
Taken altogether, the findings above suggest that whereas
the incidence of spontaneous abortion and the birth rate of
infants with congenital heart disease were reported to be
higher in areas where the drinking water was contaminated
with TRI than in uncontaminated areas, the fetal toxicity of
TM (teratogenicity or developmental toxicity) has not been
confirmed at a likely dose in human exposure.
(6) Immunotoxicity
The long-term oral administration of TRI at high doses
induced liver cancer in mice16'17). Such a large dose is likely
to diminish the function of the immune surveillance system,
which may play a role in the development of tumors. Wright
et al.77~
reported a decrease in the activities of natural killer
(NK), natural cytotoxic (NC) and natural P815 killer (NPK)
cells following a 3-day administration of TRI at a massive
dose (Sprague-Dawley rats, 5 mmol/kg/day; B6C3F1 mice,
10 mmol/kg/day). This high TRI dose decreased the
splenocyte count in the rats and the relative spleen weights
in the mice. The results of this study suggest that
compromised immune function may play a part in the
carcinogenic responses in experimental animals exposed to
T KANEKO et al.
a massive dose of TRI.
Anicteric hepatitis and uveitis are reported to have occurred
in a 48-year-old woman engaged in degreasing work with
TRI78).According to this report, the patient had been exposed
to TRI at concentrations of 40-800 ppm for 3 hr daily. She
showed a 4- to 5-fold increase in serum alkaline phosphatase
(ALP) activity and a 3-fold increase in serum transaminase
activity, with an elevation of body temperature to 38.5°C.
Serologic tests ruled out hepatitis A and B, and serologic
tests for EB virus and cytomegalovirus were also negative.
While the acute hepatitis slowly resolved about 6 weeks
later, pain in the eyes and impaired vision then appeared. A
diagnosis of bilateral anterior uveitis was made. These ocular
symptoms were improved with corticosteroid treatment.
Because all of the symptoms disappeared with the test values
being restored to normal 6 months after the onset of hepatitis,
the patient returned to work. Two weeks after resuming
work (TRI level 550 ppm), a malaise appeared again with a
transient increase in serum ALP activity. For this reason,
the worker was relocated to an office work section. She
was followed for more than one year thereafter, and no
abnormality was noted. According to the authors, no cause
other than an occupational exposure to TRI was found
concerning the development of hepatitis and uveitis. The
hepatic injury owing to high levels of TRI exposure is
acknowledged, but the relationship between the uveitis and
TRI exposure remains to be clarified. However, since not a
few cases of TRI-related scleroderma-like disorders have
been reported7~85~, it is of interest that a worker exposed to
TRI developed uveitis, a disorder regarded as one of the
autoimmune diseases.
The inhabitants of Tucson Valley,Arizona, U.S. had used,
as living water,ground water containingTRI at concentrations
of 5-500 ppb or higher, for 25 years. Kilburn and Warshaw86>
applied the criteria of systemic lupus erythematosus (SLE)
(arthritis, Raynaud's phenomenon, sore mouth-repeated
ulcers, anemia, malar rash, skin lesions, pleurisy signs, hair
loss, protein in urine, and seizure or convulsion) in 362
subjects who had used the contaminated water (exposed
group) and 158 inhabitants matched to the exposed group
members by sex, age and educational level, serving as the
control group. The frequencies of arthritis, Raynaud's
phenomenon, malar rash with sunlight, skin lesions, and
seizure or convulsion were significantlyhigher in the exposed
group than in the control group. The number of subjects
who complained of four or more symptoms in the exposed
group was also 2.3 times larger than that in the control group.
These symptoms predominated in females in the exposed
Industrial Health 1997, 35, 301-324
HEALTH EFFECTS OF TRICHLOROETHYLENE
group. Furthermore, the titers of antinuclear autoantibodies
measured by fluorescence (FANA) in serum greater than
1:80 were approximately 2.3 times higher in these females.
The acute rheumatic arthritis (ARA) score and FANA rank
were correlated with a coefficient of 0.1251, r2=0.0205
(p<0.036) in females. The authors concluded that long-term
low-dose exposure to TRI in contaminated well water
significantly increasedthe symptoms of lupus erythematosus
as perceived by the ARA score and the increased FANA
titers.
Reports suggesting the development of pneumatosis
cystoides intestinalis (PCI), a relatively uncommon disease
characterized by the multiple occurrence of gas-filled cysts
within the wall of the intestine, following occupational TRI
exposure have been made by Japanese investigators87,88>.
Yamaguchi et al.87~ carried out a case-control study to assess
the association between PCI and working conditions,
includingoccupationalexposure to organic solvents. Thirteen
patients with primary PCI were individually matched with
5 controls by sex, age, and admission year. It was found
that there was a close association between the development
of primary PCI and occupational exposure to TRI. While
only 5 of 65 controls (7.7%) were exposed to TRI, 12 of the
13 patients with PCI (92.3%) were found to have been
exposed occupationally to TRI, resulting in the odds ratio
of 144. Moreover, the healing and recurrence of PCI in
these patients substantially paralleled the profile of their
occupational exposure to TRI. Two pairs of patients with
PCI had been working in the same factory, where they had
degreased camera lenses with TRI. Since this study was
conducted in one area of Japan, it is premature to conclude
that a causal relationship between PCI and TRI has been
established on the basis of this study. However, the exposure
odds ratio of 144 is too high to consider the relationship to
be accidental. Sato et al.88)collected the cases of PCI on a
national basis during a 5-year-period from 1979-1983, and
found that TRI exposure-associated primary PCI cases were
distributed almost throughout Japan. The relationship
between PCI and occupation was studied with the 66 patients
thus collected. Informationconcerning their occupation was
obtained from 37 (56.1%) patients; 16 patients had secondary
PCI and 21 had primary PCI. The secondary PCI was
associated with collagen diseases such as progressive
systemic sclerosis in 6 patients (37.5%) who were all females
and were affected in the small intestine. No particular
characteristicswere noted in the occupationsof the secondary
PCI patients. Primary PCI, occurring more frequently in
females (15121), affected predominantly the large intestine
317
(20121),locally involving the sigmoid colon in the majority
(14) of the patients. Most of the patients with primary PCI
(16121) were factory workers, of whom 15 (71.4%) were
engaged in the degreasing of manufacturing products with
TRI. From the high percentage of TRI workers among the
patients with primary PCI, Sato et al. concluded that
occupational exposure to this agent constitutes an etiological
factor in the development of this disease. However, since
reports of TRI-associated PCI have so far been limited to
Japan, the evidence is still not enough to conclude that there
is a causal relationshipbetween primary PCI and occupational
TRI exposure.
Several studies have thus referred to TRI-related
autoimmune diseases (e.g., multiple sclerosis). However,
much more evidence is needed to establish a causal
relationshipbetween autoimmunediseases and TRI exposure.
Health Risk Assessment of TRI Toxicity
When, as in the case of TRI carcinogenicity, no
epidemiologicaldata are available, carcinogenicity to humans
is estimated by extrapolating animal data to humans. In
such a case, route-route and high dose-low dose differences
must be taken into account, in addition to the species
difference in toxicokinetics. The greatest barrier to animal-
human extrapolation underlies the difference in lifespan
between both species. Eight hours a day is not biologically
equivalent between rodents and humans. The same length
of time occupying the lifespan of each species (mice and
rats, 1,000 days; humans, 80 years) is a 29-fold difference.
TRI must undergo metabolic activation to exhibit
carcinogenicity. The bioactivation may be mediated by either
the cytochrome P450 pathway or GSH conjugation pathway.
However, it is not clear which pathway is detoxification or
activation. Moreover, the concentration of an ultimate
metabolite in the target tissue is not always proportional to
the amount of total metabolites. When the dose is increased,
the relative rate for each metabolic pathway is changed,
thereby changing the relationship between activation and
detoxification. At present, for convenience only, the amount
of a certain TRI metabolite is used as the effective dose
(EFD) for carcinogenicity by assuming that the EFD is
proportional to body surface area.
TRI induced hepatocellular carcinomas in B6C3F1
mice'7.89). The U.S. Environmental Protection Agency
(EPA)90) extrapolated this animal data (gavage test) to humans
using a linearized multistage model in order to estimate the
human risk of TRI exposure. The same agency later estimated
318
the risk of TRI exposure by inhalation using the EFD
calculated by a compartment mode191~.
Computer-assisted physiologically-based toxicokinetic
(PBTK) models are now widely accepted as a useful tool
for extrapolating animal data to humans, using parameters
such as body weight, Vmax and Km. Cronin et a1.92>and
Fisher and Allen93> used a PBTK model coupled with a
linearized multistage model to derive the human TRI
carcinogenic risk extrapolation from animal data. This
technique is certainly useful if the differences between
animals (B6C3F1 mice) and humans are only quantitative.
However, if, as in the case of TRI carcinogenicity, a high concentrations as to cause cytotoxicity.
qualitative difference that cannot be overcome is present,
such a technique cannot be considered to significantly
improve the reliability of risk assessment. As in the risk
assessment by the U.S. EPA, the assessment performed by
the PBTK modeling does not have reliable implications in
that both risk assessments were based on the same
carcinogenicity test where B6C3F1 mice were used" s9),as
discussed below.
Abelson94~ has severely criticized the risk assessment
performed by the U.S. EPA9o,9" as follows: "In calculating
the risks of human cancer and establishing regulations, the
U.S. EPA makes a series of conservative assumptions that
have no sound scientific basis. Exaggerations of risks are
inherent in the procedures by which rodents are employed
as stand-ins for humans. Dependence on results of maximum
tolerated doses in the most sensitive strain or species of rodent
is questionable. Important examples involve B6C3F1 mice,
which in contrast to humans in developed countries, have a
high control incidence of liver cancer. Those mice often
respond with liver tumors when exposed to large doses of a
substance. Other rodents are less often affected. Questionable
assessments involving liver cancer in B6C3F1 mice include
the risks posed by TRI, TETRA, methylene chloride,
butadiene, and phenobarbital." Steinberg and DeSessol~ have
also questioned the EPA's risk assessment based on a linear
extrapolation from the effects of high doses in rodents to
risks for humans at low doses, stressing that, instead of a
straight-line extrapolation model, a threshold model may
be more appropriate in the case of risk assessment of TRI
carcinogenicity. In other words, since the carcinogenicity
of TRI is preceded by cytotoxicity (necrosis), there may
exist a threshold for the carcinogenicity.
T KANEKO et al.
Proposal for Air Quality Standard of TRI
(1) Occupational exposure limit for workers
Since TRI has not been confirmed to be a human
carcinogen, and since the carcinogenicity (liver cancer) of
TRI to B6C3F1 mice may be based on an epigenetic rather
than genotoxic mechanism, it is not appropriate to establish
the occupational exposure limit of TRI in occupational
settings presupposing that TRI is a carcinogen. TRI
carcinogenicity, if any, would be induced only after this
chemical has been inhaled for a long period of time at such
The health effects of long-term exposure to TRI are chiefly
manifested as neuropathy, and thus the occupational exposure
limit for TRI could be established on the basis of its
neurotoxicity. The current exposure limit proposed by the
American Conference Governmental Industrial Hygienists
(ACGIH) (threshold limit value, TLV) and the JSOH
(occupational exposure limit, OEL) is 50 ppm (270 mg/m3).
Evidence has accumulated that a long-term exposure to TRI
at 50 ppm will cause neurotoxic effects to industrial workers,
suggesting that the value should be lowered.
In this connection, the study of Ahlmark and Forssman9S~
is of utmost importance because they examined 122 workers
for the neurotoxic effects (subjective symptoms) of TRI in
relation to the urinary levels of TCA. They found that the
workers who excreted more than 30 mg of TCA per liter of
urine manifested certain symptoms to a greater extent than
others. The symptoms most commonly complained of were
abnormal fatigue and increasedneed of sleep. Diffuse gastric
symptoms such as tympanism (meteorism), a sensation of
dryness in the mouth, a feeling of sickness, and intolerance
to alcoholic beverages were also frequently complained of.
Ahlmark and Forssman summarized their own work as
follows: "One may say that a constant exposure to TRI which
gives rise to an excretion of TCA of less than 20 mg per
liter of urine does not entail, or entails only in exceptional
cases, any definite TRI effects, whereas at the values of over
20 mg per liter of urine one sees such effects in an increasing
percentage of cases directly proportional to the increasing
excretion." However, the readers should pay attention to
the finding that among 21 workers who excreted 11-20 mg/
1of TCA, 7 were judged to have a `possible effect' and 5 an
`actual effect.' This indicates that the TCA level of 20 mg/
1 corresponds to a LOAEL (least observed adverse effect
level) rather than a NOAEL (no-observed adverse effect
level). Ahlmark and Forssman used `morning urine' for the
urinalysis for TCA. It was not defined when the `morning
Industrial Health 1997, 35, 301-324
HEALTH EFFECTS OF TRICHLOROETHYLENE
urine' was collected, in the morning before the beginning
of work or during the work in the forenoon. Although the
exposure concentration of TRI corresponding to 20 mg/l of
TCA in `morningurine' cannot have been exactly determined,
it can well be said that the concentration may be higher than
10 ppm but lower than 30 ppm. Indeed, Ahlmark and
Friberg96~proposed 30 ppm as the maximum allowable
concentration of TRI on the basis of urinalysis for TCA.
According to Liu et al.53~,the prevalence of subjective
symptoms in workers exposed to TRI around 50 ppm was
significantly high in comparison to control workers. A dose-
response relationship could be established in some selected
symptoms such as nausea, a heavy-headed feeling,
forgetfulness, tremor in extremities, and dry mouth. These
findings suggest that the effects of TRI on the central and
autonomic nervous systems appear at concentrations lower
than 50 ppm.
According to Ruijten et al.55~,occupational exposure to
TRI (cumulativeexposure level, 704 ± 583 ppm-year) caused
a slight reduction (- I .1 m/s) in the sural nerve conduction
velocity and a prolongation (0.4 ms) of the sural nerve
refractory period (mean of the controls 1.95 ms). Ruijten
et al. concluded that long-term exposure to TRI at 35 ppm
may slightly affect the trigeminal and sural nerves.
From the findings described above, the present authors
consider that the reference value of TRI in the work
environment should be lowered to below 50 ppm, presumably
to 25 ppm (135 mg/m3). Incidentally, the U.S. National
Institute for Occupational Safety and Health (NIOSH)
recommends 25 ppm as the recommended exposure limit
(REL) for TRI.
(2) Environmental air quality standard for the general
population
The contamination of air or water with TRI in the general
environment is related to industrial activities, with the
compound thereby incurring health effects not only on
industrial workers but also on the general population.
According to Brown et al.97~,TRI exposure in the general
environment allows a great margin of safety concerning TRI-
induced hepatic injuries and subsequent liver cancer
development. Hence, it is reasonable to establish the
environmental air standard (EAS) for TRI from health effects
other than its carcinogenicity. Among such health effects
caused by long-term exposure to TRI at low doses, its
neurological effect may be the most appropriate one for
consideration. Since the reference value of TRI for industrial
workers is based on relatively sufficient data, an EAS for
319
the general, population can also be established on the basis
of the reference value employed in the occupational settings.
The ACGIH defined the TLUs as "airborne concentrations
under which it is believed that nearly all workers may be
repeatedly exposed day after day without adverse health
effects." Furthermore, the ACGIH states that "because of
wide variation in individual susceptibility, a small percentage
of workers may experience discomfort from some substances
at concentrations at or below the threshold limit; a smaller
percentage may be affected more seriously by aggravation
of a pre-existing condition or by development of an
occupational illness.... Individuals may also be
hypersusceptible or otherwise unusually more responsive
to some industrial chemicals because of genetic factors, age,
personal habits (smoking, alcohol, or other drugs),
medication, or previous exposures. Such workers may not
be adequately protected from certain chemicals at
concentrations at or below the threshold limits." This
indicates that the reference value in the industrial workplace
has been established on the condition that it carries some
risk a priori. The OELs of the JSOH are essentially the
same as the TLUs of the ACGIH.
To establish an EAS for the general population on the
basis of the reference value for healthy workers engaged in
work with a definite sense of duty, a wide margin of
uncertainty factor must be taken into consideration. With
these factors in mind, the EAS can be calculated as a guide
from the following equation:
EAS = (Reference value for workers)
x (8124) x (2401365) x (40180) x 10-2,
(1) (2) (3) (4)
where
(1) conversion from occupational exposure (8 hr) to
exposure of the general population (24 hr),
(2) conversion from the number of yearly workdays (240
days) to the number of days a year (365 days),
(3) conversion of occupational exposure (40 years) to
lifelong exposure of the general population (80 years),
and
(4) a total uncertainty factor for conversion from
occupational exposure to exposure of the general
population.
The total of (1) x (2) x (3) x (4) is approximately 10-3.
That is, 10-3 of the reference value for occupational TRI
exposure will be a guide for the EAS for the general
population. The present authors propose a value falling in
the range of 25-50 ppb (135-270 ,ug/m3) as an EAS of TRI.
320
Values within this range may be scientifically sound as the
EAS, since a wide range of safety (uncertainty factor=1,000)
is already taken into account. What value is adopted as the
EAS of TRI is a question of risk management.
The above equation simplyproposes a procedure for setting
up the EAS of TRI on the basis of the reference value for
occupational TRI exposure. The only reason for adopting
40 years as the duration of occupational exposure is that
the occupational lives of the Japanese are considered to be
at 20-60 years of age (a spread of 40 years). Although it is
true that very few workers are engaged in the same job for
40 years, a sufficient margin of safety will be ensured as
the result of the assumption that the duration of exposure is
40 years. There is no precise reason for the uncertainty
factor of 102. One `10' is an uncertainty factor in
consideration of the differences in susceptibility between
industrialworkers (healthy adults) and the general population
including infants, elderly persons and highly susceptible
persons. In addition, while a recovery period of 16 hr exists
in occupationally exposed workers, the exposure of the
general population lasts for 24 hr without intermission. This
is taken into consideration in this uncertainty factor. The
other ` 10' is included from the personal view of the present
authors that the reference value for occupational exposure
is near to the LOAEL rather than to the NOAEL. The
uncertainty factor converting from LOAEL to NOAEL is
normally considered to be `10.' Lastly, it should be borne
in mind that the above equation is applicable to only TRI
on the condition that the compound is not classified as a
human carcinogen. However, the IARC has changed the
carcinogenicity classification of TRI from Group 3 to Group
2A. A new uncertainty factor may be needed to be taken
into account in the future, if this compound is identified as
a human carcinogen based on a genotoxic mechanism.
Acknowledgment
This work was financially supported in part by the Japan
Environment Agency.
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