Inhaled particle retention in rats receiving low exposures of diesel exhaust.

To study the effects of a low concentration exposure on the retention and clearance of submicron particles from the lungs, we exposed male Fisher 344 rats to diesel exhaust diluted to 50 micrograms diesel exhaust particles (DP)/m3, 20 h/d, 7 d/wk for 52 wk. Lung burdens (amount of DP in lungs) and the alveolar macrophage burdens were measured up to 52 wk postexposure. By 1 yr postexposure at least 80% of the DP was eliminated from the lungs and similarly cleared from the lavaged pool of macrophages. The DP remaining in the lungs was observed in alveolar, parabronchial and paravascular maculae. In contrast to previous high concentration exposure studies, only trace amounts of particles were observed in the mediastinal lymph nodes. To study the concentration dependence of particle retention, rats were exposed to equivalent exposures of 18 d x mg DP/m3 delivered at 5700 micrograms/m3 for 3 d, 1600 micrograms/m3 for 12 d, 250 micrograms/m3 for 72 d, or 50 micrograms/m3 for 365 d. Higher lung and macrophage burdens were initially achieved with the brief, high concentration exposures. During the postexposure period, animals exposed to the higher concentrations cleared more of the lung burden. Exposure to lower concentrations resulted in higher long-term lung burdens. These results are consistent with a model of lung clearance in which the macrophage burden and the duration of exposure are both important to the formation of the maculae. In a brief high concentration exposure, the macrophage burden rises rapidly, but then declines rapidly. However, in longer low concentration exposures, the macrophage burden will not reach the same peak, but stays at intermediate levels during the exposure and stimulates a steady development of the lung maculae from particle-laden macrophages leaving the active pool of pulmonary phagocytes.

Retention and clearance of inhaled submicron carbon black particles.

Carbon black aerosols were used as a probe of the pulmonary retention and clearance of submicron particles. Male Fischer rats (COBS CD) were exposed for 20 h/d, 7 d/wk for 1, 3, or 6 wk to either 7 +/- 2 mg/m3 carbon black or filtered air. The submicron aerosol (mass median aerodynamic diameter, MMAD, 0.24 microns) was generated with a Wright dust feed-cyclone system. Lung and hilar lymph node particle burdens were determined immediately following the exposure and at preselected intervals up to 1 yr postexposure. After 1-, 3-, and 6-wk exposures, the lung burdens were 1.1 +/- 0.1, 3.5 +/- 0.2, and 5.9 +/- 0.1 mg, respectively. One year after a 1-, 3-, or 6-wk exposure, 8%, 46%, and 61% of the initial lung burden remained in the lungs. Initially, the hilar lymph nodes contained 0.2%, 0.9%, and 2.0% of the lung burdens in the 3 exposure groups, respectively. At 1 yr postexposure, particle translocation from the lungs led to a rise in lymph node burdens to 1%, 21%, and 27% of the initial lung burden. The retention of carbon black in both the lungs and lymph nodes combined was 9%, 67%, and 89% for the 1-, 3-, and 6-wk exposed animals. Lung clearance was modeled as a compartmental system consisting of four lung compartments and a regional lymph node compartment. The results from the model are similar for carbon black and diesel engine exhaust particles. However, the compartmental kinetics of carbon black differed in two ways: the deposition efficiency in the alveolar region was lower than that for diesel exhaust particles, and there was earlier transport of particles to the regional lymph nodes. These results showed that when lung burdens reached 0.8 mg, lung clearance was decreased by 50% and lymphatic transport of insoluble particles was increased.

Pulmonary phospholipidosis in rats respiring air containing diesel particulates.

Rats chronically exposed to diesel particulates (dp) or given intratracheally a single dose of dp show increased levels of phospholipids in the lungs and in pulmonary lavage fluid. Pulmonary phospholipidosis is accompanied by increased lecithin levels and by increased palmitate content in lecithin of both lungs and pulmonary lavage fluid. A de novo increase of pulmonary and hepatic phospholipid (PL) formation was detected 5 days after rats were treated with dp. We hypothesize that a dp-stressed lung releases a pulmonary lipogenic factor (PLF), which stimulates hepatic lipogenesis. This was further tested by an in vitro study in which primary cultures of free hepatocytes were incubated with [2-14C]acetate and various molecular weight fractions of a pulmonary homogenate from rats. The results from these studies indicated that in rat lung homogenates a PLF exists of greater than 100,000 Da molecular mass. The results also indicate that respired air containing a dp concentration of greater than 750 micrograms dp/m3 of air would result in a mild phospholipidosis in the lung, whereas a dp dose in respired air of 250 micrograms dp/m3 of air for 2 years did not alter pulmonary PL content in rats.

Response of pulmonary cellular defenses to the inhalation of high concentrations of diesel exhaust.

Rats were exposed to three concentrations of diluted diesel exhaust for 6 mo and 1 yr. Bronchopulmonary lavage was used to obtain the pulmonary phagocytes from the animals in order to study the response of the phagocytic defenses to the inhaled particulate. The cell numbers and volumes were measured using an electronic particle counter. The cell counts of alveolar macrophages (AM) were proportional to the concentration of diesel exhaust particulate (DP) in the chronic exposures. AM increased in the lungs in response to the rate of DP mass entering the lungs, rather than to the total DP burden in the lung. The geometric mean volumes of AM from the exposed and control animals were approximately 1100 micron3 at both 6 and 12 mo of exposure, although exposed cell-volume distributions skewed towards larger sizes. The AM volume distributions extended to 2000 micron3 in both control and 250 micrograms DP/m3 exposed animals and up to 5000 micron3 in cells from animals exposed to 750 and 1500 micrograms DP/m3. The volume distributions were found to be reproducible in equivalent control and exposed cell populations. Polymorphonuclear leukocytes were present in the lavaged cell populations from the animals exposed to 750 and 1500 micrograms/m3. In addition, at 1 yr of exposure, lymphocytes were also lavaged from animals exposed to 750 and 1500 micrograms DP/m3. Protein, beta-glucuronidase activity, and acid phosphatase activity were measured in the lavaged cells, and were elevated in the cells from animals exposed to 750 and 1500 micrograms/m3. The buoyant density of diesel-laden AM was found to be greater than that of control AM, and overlapped with the buoyant density of the polymorphonuclear leukocytes.

A spectrophotometric method for the quantitation of diesel exhaust particles in guinea pig lung.

Particulate material in diesel engine exhaust deposits in the lungs of exposed animals. A technique for measuring the amount of soot in the lungs would be useful for determining the rates of the deposition and clearance of the submicron particles. The paper describes the use of light extinction for quantitating diesel particles in aqueous suspension. Particles collected by electrostatic precipitation and finely suspended in 0.01 N NaOH by sonication strongly absorb visible light. The extinction of light at 750 nm is proportional to the mass concentration of particles, with an extinction coefficient of 28 +/- 1 cm2 mg-1. Lungs from guinea pigs exposed to dilute diesel exhaust were dried and digested in potassium hydroxide and ethanol. The insoluble particles were centrifuged and resuspended in water by sonication. The optical density of the suspension was compared to that of suspensions made from lungs of animals not exposed to diesel exhaust, with or without known amounts of particulate added at the beginning of digestion. A concentration-dependent increase in the total amount of particles per lung was found for guinea pigs exposed to 0, 269, 813 and 1530 micrograms m-3 diesel particles for 6 months.

Enoyl coenzyme A reduction by bovine mammary fatty acid synthetase. Specificity and other characteristics.

The NADPH-dependent enoyl coenzyme A reductase activity of bovine mammary fatty acid synthetase has been characterized with regard to substrate specificity and the product formed. A relatively high specificity for an unsubstituted, four-carbon, 2,3-enoyl chain in trans configuration is obtained. Reduction of trans-crotonyl-CoA results in butyrate, 50% of which is coenzyme A-bound. The reaction is subject to product inhibition, specifically by butyryl-CoA and NADP. Free coenzyme A, on the other hand, is an activator. The pH profile, susceptibility to inhibition by -SH reagents, the results of the relative activities obtained with substrate analogues and homologues, and the ready use of crotonyl-CoA as a primer in fatty acid synthesis are consistent with a mechanism in which the crotonyl group is transferred to an -SH group, is reduced, and then is either transferred back to CoA or hydrolyzed.