Toxicological comparisons of three styles of a commercial U.S. cigarette (Marlboro with the 1R4F reference cigarette.

Toxicological comparisons were made of three commercial cigarettes, namely Marlboro full flavor, Marlboro Lights, and Marlboro Ultra Lights, with the 1R4F reference cigarette. The main comparison was a 90-d inhalation study with mainstream smoke at 150 mg total particulate matter per cubic meter, in Sprague-Dawley rats using 6 h/d and 7 d/w exposures. The principal endpoint was histopathology of the respiratory tract, along with examinations of free lung cell counts after broncho-alveolar lavage. Additional studies on mainstream smoke included Salmonella mutagenicity, cytotoxicity of particulate and gas/vapor phases, and analytical chemistry. The exposures produced effectively the same responses in each of the four groups, and the histopathology results in the commercial cigarette groups were also effectively the same. The 1R4F was also tested at 75 and 200 mg/m(3), and most of the histopathology results obtained here showed dose-response relationships. The free lung cell responses were similar in the 1R4F/commercial cigarette comparison, and there were dose-related changes in the 1R4F groups, most notably for neutrophils. Most of the changes produced in the 90-d of exposure were resolved in a 42-d post-inhalation period. Responses in the in vitro and analytical assays for the four cigarettes were in general similar, when data were expressed either per mg TPM or per mg tar yield. There were judged to be no toxicologically meaningful differences between the profiles evaluated at similar smoke concentrations for the three commercial cigarettes and for the 1R4F using these assays.

Evaluation of the micronucleus assay in bone marrow and peripheral blood of rats for the determination of cigarette mainstream-smoke activity.

The mammalian in vivo micronucleus assay is widely used as part of the genotoxicity testing battery required during the development of new drugs. As such, the in vivo micronucleus assay has been used in a battery of assays for the assessment of cigarette ingredients or design modifications to help ensure that there is no increase in risk or any new risk introduced by these additions or modifications. The present series of studies was conducted to optimize and evaluate this assay for the assessment of the effects of mainstream smoke on the micronucleus frequency in the bone marrow and peripheral blood of rats. In a first experiment, the optimal conditions for performing the micronucleus assay in these tissues were determined. This was done by use of two compounds known for their micronucleus-inducing activity, i.e., the clastogen cyclophosphamide and the aneugen colchicine. In a second experiment, the effects of tube restraint on untreated control rats were investigated. In a third experiment, the optimal conditions were used to assess the clastogenic/aneugenic activity of cigarette smoke in Sprague-Dawley rats. The rat micronucleus assay in both bone marrow and peripheral blood is able to detect clastogenic and aneugenic activity. The flow cytometric determination of micronucleated cells in rat blood is at least as sensitive as determinations in bone marrow. No statistically significant differences were observed in micronucleus frequencies between rats with and without the additional stress of tube restraint; however, the cautious approach would be to use a fresh-air-exposed group (with tube restraint) as the negative control in inhalation experiments. Using the conditions identified as optimal in the above-mentioned experiments, the micronucleus assay was not able to detect effects induced by smoke from conventional cigarettes. Nevertheless, the micronucleus assay will remain a valuable tool as part of a testing battery used to investigate possible adverse effects related to product modifications.

Toxicological comparisons of cigarettes containing different amounts of vanillin.

Vanillin is a flavoring agent used in cigarettes. Previous toxicological examinations of the effects on the addition of vanillin to tobacco used mixtures with several other flavoring agents. In the present work, toxicological comparisons were made of experimental cigarettes containing no added vanillin against otherwise similar cigarettes with three different amounts of vanillin added to the tobacco. The main toxicological comparison was a subchronic inhalation study with mainstream smoke in Sprague-Dawley rats (exposures of 150 mg/m3 of total particulate matter, 6 h exposure per day, for 90 consecutive days). Vanillin concentrations in the tobacco of the 4 cigarette types at the end of the study were 0, 67, 1233, and 3109 ppm. Additional studies with mainstream smoke were Salmonella mutagenicity (5 bacterial strains, both with and without metabolic activation, particulate phase only), cytotoxicity of both particulate and gas/vapor phases (using the neutral red uptake assay), and analytical chemistry (49 analytes, including 5 metals). Similar responses were seen across the four cigarette types, and the responses were similar to those previously described in the scientific literature. At the same smoke concentration, the inhalation exposures produced effectively the same responses, in each of the four groups. Most of the changes produced in the 90 days of exposure were resolved in a 42-day postinhalation period. The addition of vanillin to tobacco at inclusion rates up to 3109 ppm did not influence a broad range of toxicological endpoints.

High-resolution X-ray microtomography for the detection of lung tumors in living mice.

In the present study, the feasibility of applying high-resolution microtomography (micro-CT) for the detection of lung tumors was investigated in live mice at an early and more advanced stage of tumor development. The chest area of anesthesized mice was scanned by X-ray micro-CT. In mice with a minor and heavy tumor load, micro-CT proved to be a fast and noninvasive imaging device for the detection of lung tumors. After validation of the CT data by histologic sectioning, it was shown that the majority of tumors could be distinguished in the reconstructed virtual slices obtained by micro-CT. The data from micro-CT were also confirmed by visual inspection of the inflated and excised lungs postmortem. In vivo micro-CT opens broad perspectives for imaging tumor development and its progression in a noninvasive way. Micro-CT also allows for longitudinal evaluation of the treatment of lung cancer by drugs.