W.H. Steinhagen

Effect of single or repeated formaldehyde exposure on minute volume of B6C3F1 mice and F-344 rats☆

Abstract To characterize species differences in sensory irritation to formaldehyde (HCHO) and to determine if tidal volume ( V T ) compensation parallels the respiratory rate (ƒ) depression, ƒ, V T , and minute (V E ) of B6C3F1 mice and F-344 rats were measured during 10-min, head-only HCHO exposures. Comparisons also were made between naive (no previous HCHO exposure) and pretreated animals (HCHO, 2, 6, or 15 ppm, 6 hr/day for 4 days). HCHO exposure elicited immediate and simultaneous decreases in ƒ and V E in a concentration-dependent manner. No significant compensation in V T was observed in either species except the 15-ppm-pretreated rats. While ƒ and V E remained maximally depressed in mice for the remainder of the exposure period, recovery occurred in rats. In addition to tolerance development, rats were also less sensitive to HCHO. The RD50 (concentration at which ƒ decreased by 50%) of naive rats was 31.7 ppm vs 4.9 ppm for mice. Similar RD50 values and time-response curves for V E and ƒ were observed among naive and pretreated groups, although the baseline ƒ of pretreated animals was significantly depressed as compared to naive animals. In summary, HCHO exposure depressed V E in both mice and rats. But, because of greater sensitivity and lack of tolerance, mice were able to minimize inhalation of HCHO more than rats. This species difference may contribute to differences in respiratory tract toxicity from inhaled HCHO.

Sensory irritation structure-activity study of inhaled aldehydes in b6c3f1 and swiss-webster mice.

The sensory irritation potential of a series of saturated and unsaturated aliphatic and cyclic aldehydes was investigated in B6C3F1 and Swiss-Webster mice. With the reflex decrease in respiratory rate as the endpoint response, alpha, beta-unsaturated aliphatic aldehydes yielded RD50 values (concentration which elicits a 50% decrease in respiratory rate) ranging from 1 to 5 ppm while saturated aliphatic aldehydes with two or more carbons produced RD50 values from 750 to 4200 ppm. Cyclic aldehydes produced intermediate RD50 values which ranged from 60 to 400 ppm. No statistically significant differences were found between concentration-response curves of B6C3F1 and Swiss-Webster mice. Saturated aliphatic aldehydes with two or more carbons were nearly 1000 times less potent than formaldehyde. Although the mechanisms responsible for stimulation of trigeminal nerve endings by airborne chemicals are poorly understood, several hypotheses may help to explain the differences seen in this study. For example, the sensory irritation potency of the saturated aliphatic aldehydes diminished with their reported dehydration constants which may determine the degree to which these aldehydes crosslink with receptor proteins. The sensory irritation potency of acrolein and crotonaldehyde was probably due to 1,2 or 1,4 addition reactions. Additionally, molecular conformation and a recently published physical mechanism may contribute to sensory irritation responses, particularly for the less reactive aldehydes. Tentative threshold limit values (TLVs), based upon prevention of sensory irritation, were extrapolated from the RD50 values of Swiss-Webster mice. With the exception of crotonaldehyde, good agreement was found with currently published TLVs.

Sensory irritation response to inhaled aldehydes after formaldehyde pretreatment

Pretreatment of Fischer-344 (F-344) rats with formaldehyde (HCHO) induces significant cross tolerance to the sensory irritation properties of Cl2. The purpose of this study was to determine if HCHO pretreatment would cause sensory irritation cross tolerance to other inhaled aldehydes. Male F-344 rats, weighing 190 to 210 g, were pretreated with 15 ppm HCHO, 6 hr/day for 9 days, and challenged on the 10th day with a saturated (acetaldehyde, propionaldehyde, and butyraldehyde), unsaturated (acrolein and crotonaldehyde), or cyclic (cyclohexanecarboxaldehyde, 3-cyclohexene-1-carboxaldehyde, and benzaldehyde) aldehyde. The sensory irritation response in these animals was quantified by measuring respiratory rate depression in a head-only inhalation chamber using plethysmographic techniques. Control animals were challenged identically without prior pretreatment. In naive (nonpretreated) animals, the concentration eliciting a 50% decrease in respiratory rate (RD50) was 23 ppm or less for unsaturated aliphatic aldehydes. For cyclic and saturated aliphatic aldehydes, the RD50 ranged from 600 to 1000 ppm and 3000 to 6800 ppm, respectively. Formaldehyde pretreatment resulted in cross tolerance only with acetaldehyde (RD50 increased 3.5-fold) and acrolein (RD50 increased 5-fold). These results indicate that the development of cross tolerance following HCHO pretreatment is not a general phenomenon. Prediction of acceptable concentrations of occupational exposure for the prevention of sensory irritation in humans has been based primarily on RD50 data for mice. Comparison of the RD50 values obtained for rats in this investigation with previously published results for mice varied by over one-half an order of magnitude, thereby disputing the usefulness of data from F-344 rats in setting threshold limit values for the prevention of sensory irritation.

Studies of immune function and host resistance in B6C3F1 mice exposed to formaldehyde

A series of immune function and host resistance parameters were examined in female B6C3F1 mice following a 21-day (6 hr/day) inhalation exposure to 15 ppm of formaldehyde (HCHO). Immune parameters examined included delayed hypersensitivity to keyhole limpet hemocyanin, antibody plaque-forming cell response to sheep erythrocytes (T-lymphocyte-dependent antigen) and TNP-Ficoll (T-lymphocyte-independent antigen), lymphoid organ weights and histopathology, routine hematology, bone marrow cellularity and CFU progenitor cell enumeration, lymphocyte subpopulation quantitation by cell surface markers, mitogen-induced lymphocyte blastogenesis, macrophage function parameters, and host resistance to challenge with the bacterium Listeria monocytogenes and transplantable tumor cells. Lymphoid organ weight, bone marrow cellularity, and hematology parameters were unchanged in HCHO exposed mice. Similarly, the percentage of T and B lymphocytes and their proliferative responses to mitogens were not significantly altered. Antibody (IgM) plaque-forming cell response following antigen challenge was unchanged. Macrophage function was normal although some evidence of enhanced H2O2 production associated with elevated bactericidal activity was observed in resident macrophages. Resistance to challenge with the bacteria Listeria monocytogenes was significantly enhanced, while resistance to tumor challenge remained unchanged. No evidence of immunosuppression following short-term exposure to HCHO was observed.

Macrocytic-megaloblastic anemia in male B6C3F1 mice following chronic exposure to 1,3-butadiene

In the present study exposure to 1,3-butadiene (BD) resulted in a macrocytic-megaloblastic anemia in male B6C3F1 mice following chronic inhalation of 1250 ppm for 6 to 24 weeks. Treatment-related changes evident after 6 weeks of exposure included a decrease in circulating erythrocytes, total hemoglobin, and hematocrit and an increase in mean corpuscular volume. A leukopenia, due primarily to a decrease in segmented neutrophils, and a five- to sixfold increase in circulating micronuclei were observed after 6 and 24 weeks of exposure. These changes were not accompanied by a significant alteration in mean corpuscular hemoglobin concentration, an increase in circulating reticulocytes, or circulating nucleated erythrocytes. A consistent treatment-related alteration in bone marrow cellularity was not found. However, flow cytofluorometric analysis of bone marrow DNA cell cycle kinetics revealed a 44% increase in proliferative index relative to controls, due primarily to an increase in the proportion of cells in S phase. These findings are consistent with a treatment-related macrocytic-megaloblastic anemia and indicate the bone marrow to be an important target organ for BD toxicity.

Macrocytic-megaloblastic anemia in male NIH Swiss mice following repeated exposure to 1,3-butadiene

Thymic lymphoma/leukemia is the major cause of death in B6C3F1 mice chronically exposed to 1,3-butadiene (BD). Similar to radiation-induced murine thymic lymphoma, the bone marrow is also a major target organ. Because of the association of murine thymic lymphoma with endogenous type-C murine leukemia retroviruses (MuLV) present in the germ line of most strains of laboratory mice, including B6C3F1 and its parent strains, we examined the effects of BD exposure on NIH Swiss mice which do not possess intact endogenous ecotropic MuLV. Male NIH Swiss mice exhibited a macrocytic-megaloblastic anemia following inhalation of 1250 ppm BD for 6 weeks. Treatment-related changes included decreases in circulating erythrocytes, total hemoglobin, and hematocrit and an increase in mean corpuscular volume. An eightfold increase in circulating micronuclei was also observed. The anemia was not accompanied by a significant alteration in mean corpuscular hemoglobin concentration, an increase in circulating reticulocytes, or an increase in circulating nucleated erythrocytes. These findings are consistent with a treatment-related macrocytic-megaloblastic anemia and indicate that the bone marrow is an important target for BD toxicity in mice independent of MuLV background and expression.

Susceptibility to 1,3-butadiene-induced leukemogenesis correlates with endogenous ecotropic retroviral background in the mouse

Previous studies have revealed marked differences in the pattern of carcinogenesis between rats and mice exposed to 1,3-butadiene (BD) that do not appear to be readily explained on the basis of pharmacokinetics or metabolism. Chronic exposure of B6C3F1 mice to BD produces a high incidence of thymic lymphoma (TL) that is not observed in rats. The potential of the endogenous ecotropic retroviral background to influence susceptibility to BD leukemogenesis was examined by comparing the incidence of TL between B6C3F1 and NIH swiss mice. Proviral ecotropic sequences are truncated in the NIH Swiss mouse, and the virus is not expressed. Chronic exposure to BD (1250 ppm) for up to 1 year resulted in a fourfold difference in the incidence of TL between B6C3F1 (57%) and NIH Swiss (14%) mice. These results provide presumptive evidence for retrovirus involvement since NIH Swiss mice lack ecotropic viruses and appear to be relatively resistant to induction of lymphoma by BD.

NH3 concentrations in the expired air of the rat: Importance to inhalation toxicology

Abstract Endogenously produced ammonia (NH 3 ) has been quantitated in the expired air of dogs and humans but data for rodents are not available. NH 3 in expired air may have importance in toxicology studies with inhaled gases, vapors, or aerosols because of its ability to form secondary reaction products. Depending upon the reaction product(s) formed, the toxicity of an airborne agent may be increased or decreased. The objective of this study was to quantitate NH 3 in the expired air of unanesthetized Fischer-344 rats, a species commonly used in inhalation toxicity studies. Whole body plethysmography and an indophenol, colorimetric assay were used to quantitate breath NH 3 . Concentrations of NH 3 ranged from 10 to 353 ppb ( x = 78 ppb ) in nose-breathing animals and 34 to 744 ppb ( x = 286 ppb ) in tracheal cannulated animals. These differences were attributed to the absorptive effect of the moist, upper respiratory tract in nose-breathers which was absent in animals with tracheal cannulas. These results compared favorably with published reports using larger species. The presence of NH 3 in the expired air of the rat suggests that reaction products may be formed with a variety of airborne chemicals. This may be particularly significant in low-level, chronic inhalation studies.

Sensory irritation tolerance development to chlorine in F-344 rats following repeated inhalation.

Abstract Upper respiratory tract irritation (sensory irritation) by airborne chemicals in animals causes a decrease in respiratory rate with no compensatory increase in tidal volume. This condition is an important respiratory tract defense mechanism. The purpose of this investigation was to determine whether sensory irritation tolerance would develop in F-344 rats following repeated Cl 2 inhalation. Male, F-344 rats were divided into four groups (40 to 50 animals/group) and exposed to 0, 1, 5, or 10 ppm Cl 2 , 6 hr/day, 5 days/week for 2 weeks. Sensory irritation was quantified 16 to 24 hr following the last day of Cl 2 pretreatment by measuring respiratory rate depression during exposure of groups of three to four animals for 10 min to various Cl 2 concentrations (1 to 760 ppm). Concentration-response curves were compared to the curve obtained from rats receiving no pretreatment (naive group). Tolerance was seen after all Cl 2 pretreatments and was greatest in the 10-ppm group. When compared to the non-pretreated group, all Cl 2 -pretreated rats had concentration-response curves with decreased slopes demonstrating increasing tolerance with increasing Cl 2 challenge concentration. The concentration of Cl 2 needed to elicit a reflex decrease in respiratory rate of 50% (RD 50 ) was 25 ppm in non-pretreated rats whereas Cl 2 pretreatments of 1, 5, or 10 ppm increased the RD 50 values to 90, 71, and 454 ppm, respectively. Tolerance development to sensory irritation is important because an organisms ability to perceive the presence or lung damaging concentrations of irritants may be compromised. Since the current threshold limit value for Cl 2 is 1 ppm, indications of tolerance development of rats exposed to Cl 2 at 1 ppm may be relevant to humans exposed repeatedly to this concentration.

Effect of short-term inhalation exposure to 1,3-butadiene on murine immune functions

Interest in 1,3-butadiene (BD) as a potential immunomodulator was prompted by reports of an increased incidence of neoplasia in humans exposed to BD during the manufacture of styrene-butadiene synthetic rubber, and by a recent study which demonstrated a high incidence of thymic lymphomas in B6C3F1 mice. B6C3F1 mice were exposed to 1250 ppm BD by inhalation 6 hr per day, 5 days per week, for 6 or 12 weeks. Immune function assays were selected to evaluate specific humoral and cell-mediated immunity and spontaneous cytotoxicity; lymphoid organ histopathology was also evaluated. A slight decrease in antibody plaque-forming cells (PFC) per spleen was observed in exposed mice, although PFC per 10(6) splenic lymphocytes was normal. Significant extramedullary hematopoiesis and erythroid hyperplasia was observed in spleens from exposed mice, and correlated with a twofold increase in thymidine incorporation in spontaneously proliferating splenocytes. No differences in proliferation to alloantigens were demonstrable between control and BD-exposed splenocytes. Mitogenesis by phytohemagglutinin, Concanavalin A, and lipo polysaccharide was suppressed in splenocytes from exposed mice, but may have been due to the cellular dilution effect of hematopoietic activity. Cytotoxic T-lymphocyte generation was suppressed after a 6-week exposure to BD, but was comparable to controls after 12 weeks of exposure. No differences in spontaneous cytotoxicity were observed between control and exposed mice. Overall, no persistent immunological defects were detectable after inhalation exposure to this tumorigenic agent.