TOXICOLOGICAL HIGHLIGHT Site-Specific, Dose-Dependent Transitional Analysis of Toxicologic Mechanisms: The Interplay of Local Metabolic and Physicochemical Saturation

When animal exposures are used to assess human risk to inhaled chemicals, the animals generally receive higher dose rates over shorter periods of time. This difference in dose rate and duration of exposure, when applied to risk extrapolation, creates a key challenge: the challenge of accounting for the impact of dose and dose rate on possible transitions in mechanisms of toxicity (Slikker et al., 2004a,b). One such transition and the related microdosimetry are the subject of this issue’s highlighted article Increasing Exposure Levels Cause an Abrupt Change in the Absorption and Metabolism of Acutely Inhaled Benzo(a)pyrene in the Isolated, Ventilated, and Perfused Lung of the Rat by Ewing et al. (2006). This latest article is the culmination, for these investigators, of over a decade of mechanistic research investigating the inhalation of highly lipophilic carcinogens such as polycyclic aromatic hydrocarbons (PAHs) and tobacco-specific nitrosamines (Gerde and Scott, 2001; Gerde et al., 1991, 1993a,b,c, 1997, 1998a,b, 2001, 2004). When inhaled, these highly lipophilic compounds show carcinogenetic activity primarily at the cellular site of entry into the body. However, at cumulative doses even much higher than those required to induce lung tumors in humans, inhalation exposures in laboratory animals generally fail to induce respiratory tract tumors (Coggins, 2001). At the epithelial cell, the initial site of entry, the dose, and the dose rate of PAHs and nitrosamines appear to change the mechanism of toxicity. The epithelial cell may experience two types of saturation: metabolic saturation and physicochemical saturation (Gerde et al., 1991). The physicochemical saturation, in the thicker epithelia of the conducting airways, is proposed to be related to a slow passage of the highly lipophilic PAHs; the metabolic saturation is proposed to be specific to epithelial cell metabolism. In order to measure the interplay of metabolic saturation and physicochemical saturation, the authors use a three-part experimental approach: (1) nanoporous carrier particles are used to control, in peripheral bronchi and bronchioles, epithelial cell microdosimetry of tritium-labeled, benzo(a)pyrene (BaP); (2) the isolated perfused lung (IPL) of the rat is used to sample lung-related metabolites of BaP; and (3) for inhalation by the IPL, a dry powder generator is used to reproducibly generate a high-concentration bolus of airborne carrier particles. The carrier particles are nanoporous spheres of silica (pore diameter 1⁄4 80 nm, sphere diameter 1⁄4 3000 nm); the porosity is sufficient to allow a 10003 increase in mass loading of BaP (3.4–3500 fg per particle) without significant change in the lung deposition site (as defined by aerodynamic particle size). The isolated perfused rat lung was suspended in a humidified, artificial thoracic chamber, ventilated at 75 breaths/min (85 ml/min), and perfused at 19.8 ml/min with Krebs–Ringer bicarbonate buffer (Gerde et al., 2004). By exposing an IPL to an aerosol of carrier particles, generated with a DustGun aerosol generator (Gerde et al., 2004), reproducible mass loadings of BaP were achieved for groups of three lungs at low-, medium-, and high-dose levels: 2.2, 36, and 84,000 ng per lung. For each lung, the values measured were lung volume, carrier particle count per lung, and, in lung tissue and perfusate, the concentration of radioactive BaP and metabolites. The key contributions of these measurements are twofold: a demonstration of the application of microdosimetry and a demonstration that, at the epithelial cells closest to each carrier particle, not the metabolism but the absorption of BaP saturates at the medium-dose level. The carrier particle deposition allowed, in adjacent epithelial cells, realistic estimates of the concentration of BaP in and above the cell compared to the whole-body dose; for the medium dose, the microdose at the epithelial cell level was 2,300,000nM compared to the whole-body average dose of 0.44nM. For epithelial cells below, or near to, a low-dose carrier particle, the cells absorbed BaP at a rate proportional to the local concentration of BaP (a firstorder process). At the mediumor high-exposure levels, the cell or cells absorbed BaP at a constant rate, independent of the local concentration (a zero-order process). At high exposures, 1 To whom correspondence should be addressed. E-mail: moss@ciit.org.