ToxSci Advance Access originally published online on September 4, 2006
Toxicological Sciences 2006 94(2):439; doi:10.1093/toxsci/kfl099
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TO THE EDITOR
Department of Environmental Medicine and Department of Pediatrics/Neonatology, University of Rochester, 575 Elmwood Avenue, Medical Center Box 850, Rochester, New York 14642
E-mail: gunter_oberdorster{at}urmc.rochester.edu
Received July 31, 2006; accepted August 2, 2006
Phalen et al. (2006)
have opened an important discussion on a key question of in vitro dosing of cells with poorly soluble particles, focusing on tracheobronchial epithelial cells. Indeed, this paper is very timely, given the increasing number of publications using exorbitantly high in vitro doses of particles and extrapolating from those results potential adverse effects in exposed humans. In almost all cases, there is no, or only minimal, discussion about the all-important issue of the relevancy of applied doses to the real-world conditions. Therefore, it is commendable that Phalen et al. address this issue in a balanced way, pointing out that local deposited doses on the airways—under certain assumed exposure scenarios—can be several thousand times greater than average surface area doses. At the same time, they discuss the limitations and simplifications of their calculations and caution that their assumptions may not always be realistic, and they also emphasize the very small area of highly exposed cells in vivo. This latter point is very important, and it would have been useful to emphasize that out of 1,000,000 cells on a 12-mm culture dish, only less than 1% of the cells should receive the much greater than average dose by alternative method C in Figure 1 of Phalen et al. (assuming 25–30 hot spots; incidentally, the focal high dose area of 200 cells should be 0.01 mm2, not 1 mm2 as stated in the paper).
While the overall message of the paper is very informative, the recommendation to use a simple equation to calculate/predict surface deposition doses for aerosol particles to be applied to in vitro studies is only partly helpful and, in fact, can be very misleading without an additional cautionary note. Among the parameters in this predictive equation, such as surface area and deposition efficiency for specific regions of interest in the respiratory tract, is also the duration of exposure. While the difficulty of applying these calculations to type I cells of the alveolar region is pointed out (the equation should also not be used for alveolar macrophage and type II cell cultures), it is surprising that exposure duration is listed without any further comment. In vivo particle delivery by inhalation is gradual, over hours (Phalen et al. considered a 2-h exposure), days, weeks, etc, as opposed to seconds or less for in vitro dosing, and this raises the issue of "dose rate." This point is not addressed at all by the authors, which should have been an integral part in this discussion, especially since exposure duration is included in the predictive dose calculation. While a number of studies have established comparability between instillation and inhalation with regard to effects, in vitro studies are seldom of sufficient duration to account for the effect of dose rate and possible clearance that are present in vivo. The danger is that future in vitro studies might be designed by applying very long exposure durations in the simple formula and thereby justifying unrealistic high in vitro doses. With the possible exception of air-liquid interphase in vitro exposures by aerosol or continuous perfusion systems, a "bolus" effect will always be present in vitro. This effect will be amplified by extremely high doses, likely resulting in responses whose underlying mechanism may be different from that operating in vivo with gradually increasing deposited doses.
REFERENCES
Phalen RF, Oldham MJ, Nel AE. (2006) Tracheobronchial particle dose considerations for in vitro toxicology studies. Toxicol. Sci. 92:1126–132.
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