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The Effects of in Vitro and Aerosol Exposures to Cadmium on Phagocytosis by Rat Pulmonary Macrophages. GREENSPAN, B. J., and MORROW, P. E (1984). Fundam. Appl. Toxicol. 4, 48–57. Aerosol exposures of rats were performed to assess the in vivo effects of cadmium on the phagocytosis of latex particles by pulmonary macrophages. An in vitro assay for particle uptake was devised which allowed quantification of phagocytosis by adhering and nonadhering macrophages. In vitro exposure to CdCl2 caused dose-dependent decreases in viability (trypan blue exclusion), percentage cells with particles, total number of particles phagocytized, and the ability of the macrophages to adhere to a siliconized glass surface. In vivo effects were studied following 30-min aerosol exposures to 1.5 mg/m3 Cd (MMAD = 0.35 μm, σg = 1.45) or 5.0 mg/m3 Cd (MMAD = 0.45 μm, σg = 1.60) as CdCl2. Phagocytic activity in the in vitro test system was increased immediately and at Day 1 in the low exposure group. However, following exposure to 5.0 mg/m3 Cd, phagocytic activity was depressed until 8 days postexposure. The results show that cadmium is capable of modifying the phagocytic ability of macrophages in vivo as well as in vitro. Determinations of the total number of particles phagocytized were found to be more sensitive than percentage cells phagocytizing in detecting the effects of cadmium exposure.

Toxaphene Inhibition of Calmodulin-Dependent Calcium ATPase Activity in Rat Brain Synaptosomes. PRASADA RAO, K. S., TROTTMAN, C. H., MORROW, W., AND DESAIAH, D. (1986) Fundam. Appl. Toxicol. 6, 648–653. Effect of toxaphene on Ca+2-ATPase activity in rat brain synaptosomes was studied in vitro and in vivo. Ca+2-ATPase in calmodulin-depleted synaptosomes was inhibited in vitro to a maximum of about 50% at 150 μM toxaphenc. Substrate activation kinetics of Ca+2-ATPase in synaptosomes revealed that toxaphene inhibited the enzyme activity noncompetetively by decreasing Vmax values, without affecting the enzyme-substrate affinity. Toxaphene inhibited the calmodulin activated Ca+2-ATPase activity in a concentration-dependent manner with an IC50 of 10 μM, a concentration at which no significant effect was observed on basal enzyme activity. Nuclear and P2 fraction (synaptosomes) calmodulin levels were reduced significantly in toxaphene-treated rats. The synaptosomal Ca+2-ATPase was also reduced to about 45% in toxaphene-treated rats and the activity was restored to normal levels by the exogenously added calmodulin. These results suggest that toxaphene may cause synaptic dysfunction by in terfering with calmodulin and its regulation of neuronal calcium.

The application of α-cyclodextrin (α-CD) as an alternative vehicle for water insoluble and volatile chemicals was investigated in toxicity studies of p-chloro-α,α,α-trifluorotoluene (CTFT). Groups of F344 rats and B6C3F1 mice of each sex were administered CTFT (97% pure) by gavage in either corn oil or α-CD aqueous formulations daily for 14 consecutive days. The dose levels used were 10 (mice only), 50, 400, and 1000 mg/kg for corn oil vehicle and 10, 50, and 400 mg/kg (maximum achievable dose at gavage volume of 5 ml/kg) for α-CD vehicle. With both vehicles CTFT and a2u-globulin were found to accumulate in the male rat kidney after 14 days of exposure and a dose-related toxic nephropathy was observed at dose of 50 mg/kg or higher. The hepatocellular hypertrophy and cytoplasmic vacuolation of the adrenal cortex which appeared in dosed male and female rats were also found to be independent of vehicle. Clinical pathology findings suggested a mild anemia and cholestasis in rats. With both vehicles no tissue bioaccumulation of CTFT was found in male or female mice. Vehicle-independent hepatocellular hypertrophy and cholestasis were also observed in mice at doses of 400 and 1000 mg/kg. In conclusion, the α-CD vehicle does not affect the toxic responses of CTFT in both sexes of both species. The results of the studies suggest that α-CD may be an appropriate alternative vehicle for toxicity studies.

Manganese chloride (50–800 μg) was injected unilaterally into the right nostril of rats and its accumulation in the central nervous system (CNS) was monitored. Brain manganese levels were elevated in a dose-dependent, time-dependent, and tissue-dependent manner. Elevated levels of manganese were detected in the right olfactory bulb and olfactory tubercle within 12 hr after instillation and remained elevated for at least 3 days. As little as 100 μg of manganese chloride was sufficient to increase brain manganese levels. No changes were detected on the left side of the brain. The manganese content of the striatum, the target site for manganese neurotoxicity, was unchanged following acute administration, but was elevated when two injections were made 1 week apart. These results suggest that air-borne manganese can be retrogradely transported along olfactory neurons to the CNS and can reach deeper brain structures under appropriate exposure conditions.

The state of the art for modeling the retention of inhaled insoluble particles deposited in the alveolar region of the lung is briefly reviewed, and a new compartmental model of long-term retention is proposed. Wherever possible, this new model favors the replacement of simple first-order kinetics of particle transport processes in the lung by quantified mechanisms derived from or suggested by experimental data of published studies in lung physiology and histopathology. In particular, all macrophage-mediated transport processes, including classical alveolar clearance onto the mucociliary escalator, are modeled as dependent on actual macrophage mobility and are assumed to be influenced by the finite macro-phage life time. The mobility is predicted to decrease with increasing particle burden of the macrophage, and there is a limit to the macrophage capacity for accumulating burdens of insoluble particles by phagocytosis. Furthermore, at high particle burdens, macrophages will be pro-gressivity sequestered by irreversible aggregation and immobilization. Using published data on Fischer 344 rats for a quantitative demonstration of the patterns of the new model under chronic exposures, a basic set of model parameters predicts that, at moderate particle deposition rates, retention is limiting itself by establishing a steady state, and the alveolar burden is almost completely eliminated during the postexposure period. However, at high particle deposition rates, the alveolar particle burden increases continuously during the exposure period, and only a small fraction of the deposit is subject to clearance after termination of exposure. In qualitative terms, these are typical features of the “overload” effect which has been observed in a number of recent chronic aerosol inhalation exposure studies with animals.

Comparative Toxicity of Arsine Gas in B6C3F1 Mice, Fischer 344 Rats, and Syrian Golden Hamsters: System Organ Studies and Comparison of Clinical Indices of Exposure. BLAIR, P. C, THOMPSON, M. B., MORRISSEY, R. E., MOORMAN, M. P., SLOANE, R. A., AND FOWLER, B. a. (1990). Fundam. Appl. Toxicol. 14, 776–787. In order to examine possible species differences in response to arsine exposure, multiple inhalation studies consisting of acute (1-day), subacute (14- and 28-day), and subchronic (90-day) exposures to this agent were conducted using three different species of rodents. Evaluations of hematopoietic organs and alterations in the heme biosynthetic pathway were the focus of these studies. Species used were B6C3F1 mice (exposed 1, 14, or 90 days), Fischer 344 rats (exposed 14, 28, or 90 days), and Syrian Golden hamsters (exposed 28 days). All arsine exposures were at concentrations of 0.5, 2.5, or 5.0 ppm except for 90-day studies, in which concentrations were lowered to 0.025, 0.5, or 2.5 ppm. No changes in body weight gain were observed in either sex of mice or hamsters. The only decrease in body weight gain occurred in male rats exposed to 5.0 ppm arsine for 28 days. Significant exposure-related increases in relative spleen weights occurred in both sexes of mice and rats in the 0.5 (except 14-day female rats), 2.5, and 5.0 ppm exposure groups from all studies and in hamsters in the 2.5 and 5.0 ppm exposure groups. Generally, increases in relative liver weight occurred in fewer exposure groups and were of a lesser magnitude than increases in spleen weight. Other parameters affected included decreased packed cell volumes (mice, rats, and hamsters), hematol-ogy profiles (rats), and an increase in 6-aminolevulinic acid dehydratase activity in all species. Arsenic content was measured in livers of rats after 90 days of exposure. Concentrations increased in relation to atmospheric concentrations of arsine. Histopathological changes included increased hemosiderosis and extramedullary hematopoiesis in spleen and intracanalicular bile stasis (mice only) in liver. Additionally, bone marrow hyperplasia was observed in rats. Effects on other organs were not observed, suggesting that the hematopoietic system is the primary target for arsine. In conclusion, we have determined that the effects of arsine exposure upon mice, rats, and hamsters are similar. Most importantly, even though no effects on the hematopoietic system were observed following a single exposure to 0.5 ppm arsine which is 10 times the Threshold Limit Value (TLV) set by the American Conference of Governmental Industrial Hygienists, repeated exposure to 0.025 ppm (one-half the TLV) caused a significant anemia in rats.

Organ-Specific Hematopoietic Changes Induced by a Recombinant Human Interferon-α in Mice. ROSENTHAL, G. J., STRANAHAN, R. P., ILL, THOMPSON, M., BLAIR, P., GERMOLEC, D. R., COMMENT, C. E., SCHWAB, K., AND LUSTER, M. I. (1990). Fundam. Appl. Toxicol. 14, 666–675. Interferon-a (IFN-α) is a naturally occurring cytokine that mediates numerous biological activities and has demonstrated therapeutic potential in a variety of malignancies. Encouraging activity against HIV-1 replication has also been observed with IFN-α in the treatment of AIDS, although hematotoxicity has been a frequently observed side effect. In addition, in vitro studies have suggested that IFN-α may function as a down-regulator of myelopoiesis. A recombinant hybrid of subtypes of human IFN-α, rHuIFN-αA/D, has antiviral activity in mu-rine cells in vitro and In vivo. This study examines the effect of acute and subchronic exposure to rHuIFN-αA/D on hemopoietic and immune parameters in C57B1/6 mice. IFN-a was administered ip at 0, 1000, 10,000, and 100,000 units/day for either 1 or 10 consecutive days. Many of the known effects of IFN-α in humans such as anemia, leukopenia, and thrombocytopenia were observed in mice following subchronic exposure, with the latter two effects also manifested following acute exposure. Further analysis showed that this leukopenia was not selective. Both splenic and bone marrow cells were examined following 10 days of dosing with the high dose of IFN-α. Lymphocytes were reduced in both compartments, while granulocytes were increased in both compartments. Bone marrow cells programmed to differentiate into granulocytes (CFU-G) were suppressed, while macrophage progenitors (CFU-M) were stimulated. Erythroid cells decreased in the marrow but increased in the spleen, suggesting that the microenvironmerit may play a significant role in the effect of IFN-α. The proliferative capacity of both B and T splenic lymphocytes was significantly suppressed in a dose-related fashion following multiple exposure to IFN-a. Clinically, IFN-a is most often given in multiple doses and the present data suggest that such a regimen is toxic to both erythroid and myeloid cells, as well as being immunotoxic to splenic B and T lymphocytes.

The Disposition of Coal Dusts in the Lungs and Tracheobronchial Lymph Nodes of Dogs. Morrow, P.E. and Yuile, C.L. (1982). Fundam. Appl. Toxicol. 2:300–305. The pulmonary disposition, histopathology and lymphatic uptake of anthracite (Tamaqua) and bituminous (Lower Kittaning) coal dusts were measured as part of a pulmonary retention study which revealed a mean half-time of 1.92 years in dogs (Morrow et al. 1981). After brief (1-2.5 hr) exposures to either natural or neutron-activated coals having an average airborne mass concentration of {small tilde}90 mg m−3 and a 1.8 μm mass median aerodynamic diameter (σg 2.5), dogs (n=12) were serially sacrificed up to 52 weeks after exposure. Coal dusts were found only in the lungs and pulmonary lymph nodes. The coals were considered indistinguishable as to their pulmonary clearance and disposition and lymphatic uptake. All coals in the lung were associated mainly with the peribronchiolar and perivascular lymphatics or connective tissue spaces, and some were found in alveolar macrophages. The lymphatic uptake of coal dusts followed the powder function 0.55 t0.613 where t is in weeks and uptake is expressed as percent of the initial alveolar burden. In terms of pulmonary dust clearance, only 4 percent of the initial alveolar burden appeared to have been translocated to the tracheobronchial lymph nodes in the first 50 weeks, but this constituted {small tilde}14 percent of the total alveolar clearance. Histopathologically, one distinction was found: animals exposed to the highest level of neutron-activated anthracite showed patchy hyaline thickening of some small blood vessels and alveolar septa. The response was low grade, probably exposure-related, but otherwise unremarkable.

Alveolar Retention and Clearance of Insoluble Particles in Rats Simulated by a New Physiology-Oriented Compartmental Kinetics Model. STOBER, W., MORROW, P. E., AND MORAWIETZ, G. (1990). Fundam Appl. Toxtcol. 15,329–349. A physiology-oriented compartmental kinetics model of alveolar retention of inhaled insoluble paniculate matter in rat lungs was proposed in a recent paper, (W. Stober, P. E. Morrow, and M. D. Hoover, 1989, Fundam. Appl. Toxicol. 13, 823–843), and the retention patterns obtained with the model for a hypothetical set of input data appeared to simulate phenomena which were observed in inhalation studies with Fischer 344 rats. The present paper represents the results of applying the new model for simulations of the actual experimental retention data of five different inhalation studies with Fischer 344 rats exposed to three different materials. The experimental data showed that model adjustments had to be made in order to account for clearance effects that appeared to be influenced by the age of the animals. After these adjustments were made and an appropriate set of values for the model parameters describing the respective exposure conditions was used, the model was constrained to represent the empirical data of all of the studies by one unique set of parameter values. Changes in particular values of this set were considered to be acceptable only if they reflected changes of relevant properties of the inhaled paniculate matter. The final simulations did not completely comply with this self-imposed criterion. However, the degree of compliance and the simulation quality achieved with a minimum of parameter variations seem to be unprecedented in retention modeling. The results of the study encourage attempts for further refining the present model

Various chemicals and proteins of industrial importance are known to cause respiratory allergy, with occupational asthma being the most important manifestation of the disease. This paper describes clinical syndromes, mechanisms associated with occupational respiratory hypersensitivity, and methods available currently for the prospective identification of potential respiratory allergens. Certain classes of chemicals are commonly associated with occupational respiratory allergy. There is insufficient information, however, to predict respiratory sensitization potential from analysis of structure alone, although reactivity with proteins is likely to be relevant. As yet there exist no fully validated or widely applied predictive methods or internationally harmonized guidelines. The most promising predictive animal methods are the mouse IgE test and guinea pig models. Work in mice has focused upon events occurring during the induction phase of sensitization following primary encounter with the test chemical. In contrast, guinea pig models have been used primarily to identify respiratory allergens (chemicals or proteins) as a function of elicitation reactions induced in previously sensitized animals. Given the possible serious health manifestations of respiratory allergy, early identification of respiratory sensitizers is urgently required. The two methods should, as a priority, be developed further and the production of a detailed protocol for these methods be undertaken to facilitate further validation. Together, this information will allow for two types of risk assessment associated with respiratory allergy: the risk that exposure to a material will (1) induce sensitization in an individual and (2) elicit allergic reactions in a previously sensitized individual.