RESUMEN
The heated cigarette (HC) generates mainstream smoke by vaporizing the components of the tobacco rod using a carbon heat source at the cigarette tip. Mainstream smoke of HC contains markedly less chemical constituents compared to combusted cigarettes. Mainstream smoke from HC was generated under Health Canada Intense regimen and its biological effects were compared to those of Reference (3R4F) cigarettes, using nose-only 5-week and 13-week inhalation studies. In the 13-week study, SD rats were necropsied following exposure to mainstream smoke from each cigarette at 200, 600 or 1000 µg wet total particulate matter/L for 1 h/day, 7 days/week or following a 13-week recovery period. Histopathological changes in the respiratory tract were significantly lesser in HC groups; e.g. respiratory epithelial hyperplasia in the nasal cavity and accumulation of pigmented macrophages in alveoli. After a 13-week recovery, the lesions were completely or partially regressed, except for accumulation of pigmented macrophages in alveoli, in both HC and 3R4F groups. In the 5-week study, SD rats were necropsied following exposure to mainstream smoke of either cigarette at 600 or 1000 µg/L for 1 h, two times/day (with 30 min interval), 7 days/week or following a 4-week recovery period. Bronchoalveolar lavage fluid (BALF) analysis of neutrophil percentages and enzyme levels like γ-GT, ALP and LDH indicated that pulmonary inflammation was significantly less in HC groups compared to 3R4F groups. In conclusion, HC demonstrated significantly lower biological effects compared to 3R4F, based on the BALF parameters and histopathology.
Asunto(s)
Sistema Respiratorio/efectos de los fármacos , Productos de Tabaco , Contaminación por Humo de Tabaco/efectos adversos , Animales , Peso Corporal/efectos de los fármacos , Líquido del Lavado Bronquioalveolar/citología , Carboxihemoglobina/análisis , Recuento de Células , Femenino , Calor , Riñón/efectos de los fármacos , Riñón/patología , Hígado/efectos de los fármacos , Hígado/patología , Masculino , Miocardio/patología , Tamaño de los Órganos/efectos de los fármacos , Ratas Sprague-Dawley , Sistema Respiratorio/patología , Sistema Respiratorio/fisiopatología , Humo/efectos adversos , Humo/análisis , Nicotiana , Pruebas de Toxicidad SubcrónicaRESUMEN
Female C57BL/6 mice were exposed to mainstream cigarette smoke at 600 µg WTPM/L, 4 h/day and 5 days/week for up to 52 weeks. At 26, 52 and 65 weeks (52 weeks of exposure plus 13 weeks of no exposure), lungs were assessed for inflammation, function, histopathology and morphometry. Structural changes were observed and accompanied by altered lung function at 26 and 52 weeks (e.g. increase of static compliance and hysteresis, and decrease of elastance). Lung morphometry quantified significant increase in airspace enlargement at 52 weeks. Chronic smoke exposure induced inflammation in respiratory organs, e.g. mixed inflammatory cell infiltrates, perivascular lymphocyte infiltrates and pigmented alveolar macrophages in the lungs. Minimal or mild alveolar emphysema was diagnosed in 70% by 26 weeks or 80% by 52 weeks. After 13 weeks of recovery, most biochemical, histopathological and morphometrical alterations were restored, while emphysema was observed to persist at 18% incidence by 65 weeks. In conclusion, the employed exposure conditions induced emphysematous changes in the lungs, accompanied by altered lung function and morphological/histopathological changes. Following the 13 weeks of no exposure, morphological changes persisted, although some functional/biochemical alterations regressed.
Asunto(s)
Contaminantes Atmosféricos/toxicidad , Enfisema/inducido químicamente , Pulmón/efectos de los fármacos , Contaminación por Humo de Tabaco/efectos adversos , Glándulas Suprarrenales/efectos de los fármacos , Glándulas Suprarrenales/patología , Animales , Líquido del Lavado Bronquioalveolar/química , Líquido del Lavado Bronquioalveolar/citología , Carboxihemoglobina/análisis , Recuento de Células , Diferenciación Celular , Cotinina/sangre , Citocinas/metabolismo , Enfisema/patología , Enfisema/fisiopatología , Femenino , L-Lactato Deshidrogenasa/metabolismo , Pulmón/patología , Pulmón/fisiopatología , Ratones Endogámicos C57BL , Nicotina/sangre , Tamaño de los Órganos/efectos de los fármacosRESUMEN
A variety of exposure regimens of cigarette smoke have been used in animal models of lung diseases. In this study, we compared biological responses of smoke exposure in rats, using different smoke concentrations (wet total particulate matter [WTPM]), daily exposure durations, and total days of exposure. As a range-finding acute study, we first compared pulmonary responses between SD and F344 strains after a single nose-only exposure to mainstream cigarette smoke or LPS. Secondly, F344 rats were exposed to cigarette smoke for 2 or 13 weeks under the comparable daily exposure dose (WTPM concentration x daily exposure duration; according to Haber's rule) but at a different WTPM concentration or daily exposure duration. Blood carboxylhemoglobin was increased linearly to the WTPM concentration, while urinary nicotine plus cotinine value was higher for the longer daily exposure than the corresponding shorter exposure groups. Gamma glutamyl transferase activity in bronchoalveolar lavage fluid (BALF) was increased dose dependently after 2 and 13 weeks of cigarette smoke exposure, while the neutrophil content in BALF was not increased notably. Smoke-exposed groups showed reduced body weight gain and increased relative lung and heart weights. While BALF parameters and the relative lung weights suggest pulmonary responses, histopathological examination showed epithelial lesions mainly in the upper respiratory organs (nose and larynx). Collectively, the results indicate that, under the employed study design, the equivalent daily exposure dose (exposure concentration x duration) induces equivalent pulmonary responses in rats.
RESUMEN
Cigarette smoke exposures in mice have been conducted under various exposure conditions using different strains as animal models of smoke-related diseases. We exposed cigarette smoke to two strains of mice [C57BL/6J (C57) and AKR/J (AKR)] under two different exposure regimens (1 h or 4 h/day) at equivalent daily exposure amount (concentration × time). After 2 weeks exposure, mice were evaluated using exposure markers and biological responses. Smoke exposure suppressed respiratory parameters dependent on exposure concentration. The 1-h regimen groups generally showed a greater degree of respiratory suppression and relatively lower exposure markers of urinary nicotine metabolites than the corresponding 4-h regimen groups. Tidal volume was more suppressed in AKR compared to C57, while respiratory rate was more suppressed in C57. Plasma exposure markers and respiratory parameters suggested that C57 inhaled more volume of smoke than AKR. Changes in bronchoalveolar lavage fluid (BALF) cytology and enzyme parameters were most noticeable in the 1 h AKR groups. In BALF cytokine concentration, TARC concentration in C57 was higher than AKR, while KC and MCP-1 in AKR were higher than C57. Relative lung/body weight ratio in smoke-exposed C57 was generally higher, as well as the incidence and severity of lesions in respiratory organs compared to AKR. In summary, C57 appeared to inhale relatively more smoke and displayed greater inflammatory changes in respiratory tract than AKR. Comparison of exposure regimens suggests that a longer exposure duration at lower WTPM concentration might deliver a larger dose of smoke than a shorter exposure duration at higher WTPM concentration.
Asunto(s)
Exposición por Inhalación/efectos adversos , Sistema Respiratorio/efectos de los fármacos , Contaminación por Humo de Tabaco/efectos adversos , Animales , Biomarcadores/sangre , Biomarcadores/orina , Líquido del Lavado Bronquioalveolar/citología , Líquido del Lavado Bronquioalveolar/inmunología , Citocinas/análisis , Femenino , Exposición por Inhalación/análisis , Ratones , Ratones Endogámicos AKR , Ratones Endogámicos C57BL , Nicotina/metabolismo , Nicotina/orina , Neumonía/etiología , Neumonía/inmunología , Neumonía/patología , Frecuencia Respiratoria/efectos de los fármacos , Sistema Respiratorio/inmunología , Sistema Respiratorio/patología , Especificidad de la Especie , Volumen de Ventilación Pulmonar/efectos de los fármacos , Factores de Tiempo , Contaminación por Humo de Tabaco/análisisRESUMEN
In vitro testing can facilitate the rapid assessment of next generation nicotine delivery products (NGPs) with comparisons to combustible tobacco products. In vitro assays for cytotoxicity and oxidative stress were employed at BAT (UK) and JT (Japan) to test total particulate matter (TPM) of a scientific reference cigarette and aerosol collected mass (ACM) of a commercially available E-cigarette and two tobacco heating products (THP). 3R4F TPMs were generated using the Health Canada intense (HCI) regimen, a modified regime (mHCI) for the THP ACMs and the CORESTA recommended method no. 81 for the E-cigarette ACM. Human lung cells were exposed to the test product TPM/ACMs at concentrations between 0-200 µg/ml followed by the employment of commercially available assays for endpoint analysis that included reactive oxygen species (ROS) generation, the glutathione ratio (GSH:GSSG), activation of the antioxidant response elements (ARE) and cellular viability. TPM/ACM nicotine concentrations were quantified using a UPLC-PDA technique. At both laboratories the 3R4F TPM induced significant and dose-dependent responses in all in vitro assays, whereas no significant responses could be measured for the NGP ACMs. In conclusion, both laboratories obtained comparable results across all endpoints therefore demonstrating the utility of the in vitro techniques combined with standardised test products to support the assessment of NGPs.
Asunto(s)
Aerosoles/análisis , Supervivencia Celular/efectos de los fármacos , Células Epiteliales/efectos de los fármacos , Pulmón/efectos de los fármacos , Nicotina/análisis , Material Particulado/análisis , Productos de Tabaco/análisis , Sistemas Electrónicos de Liberación de Nicotina , Humanos , Japón , Reino UnidoRESUMEN
Test cigarette (prototype "heated" cigarette) was evaluated on its dermal tumor promotion activity in SENCAR mice relative to conventional 3R4F cigarette. Mainstream cigarette smoke was generated under the modified Health Canada Intensive Regimen, and smoke condensate (CSCs) were collected using cold traps and extracted with acetone. Female mice received a topical application of 7,12-dimehtylbenz(a)anthracene (DMBA) as the tumor initiator on the back skin during Week 1. Subsequently, CSC was repeatedly applied as the tumor promoter at 5 doses, up to 30 mg tar/application, three times per week for 30 weeks. Test groups showed a clearly longer latency at lower doses (⩽15 mg), but the difference was less clear at higher doses (⩾22.5 mg), while mortalities were not affected throughout the study. Test groups also had consistently lower incidence and multiplicity of neoplasms, as well as lower incidences of non-neoplastic changes (e.g., inflammations and squamous epithelial hyperplasia on the site of application). The group without DMBA initiation did not induce any neoplasm but the respective Reference group showed an increase in tumorigenicity. In conclusion, the study demonstrated significant reduction in dermal irritancy and tumorigenicity of Test CSC compared to Reference CSC.
Asunto(s)
Transformación Celular Neoplásica/efectos de los fármacos , Fumar/efectos adversos , Productos de Tabaco/efectos adversos , 9,10-Dimetil-1,2-benzantraceno/administración & dosificación , 9,10-Dimetil-1,2-benzantraceno/toxicidad , Administración Cutánea , Animales , Peso Corporal/efectos de los fármacos , Pruebas de Carcinogenicidad , Carcinógenos/toxicidad , Transformación Celular Neoplásica/patología , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Femenino , Ratones , Ratones Endogámicos SENCAR , Tamaño de los Órganos/efectos de los fármacos , Piel/efectos de los fármacos , Piel/patología , Neoplasias Cutáneas/inducido químicamente , Neoplasias Cutáneas/patología , Productos de Tabaco/análisisRESUMEN
For the biological evaluation of cigarette smoke in vitro, the particulate phase (PP) and the gas vapor phase (GVP) of mainstream smoke have usually been collected individually and exposed to biological material such as cultured cells. Using this traditional method, the GVP is collected by bubbling in an aqueous solution such as phosphate-buffered saline (PBS). In such a way the water-insoluble GVP fraction is excluded from the GVP, meaning that the toxic potential of the water-insoluble GVP fraction has hardly been investigated so far. In our experiments we used a direct exposure method to expose cells at the air-liquid interface (ALI) to the water-insoluble GVP fraction for demonstrating its toxicological/biological activity. In order to isolate the water-insoluble GVP fraction from mainstream smoke, the GVP was passed through 6 impingers connected in series with PBS. After direct exposure of Chinese hamster ovary cells (CHO-K1) with the water-insoluble GVP fraction in the CULTEX(®) system its cytotoxicity was assayed by using the neutral red uptake assay. The water-insoluble GVP fraction was proven to be less cytotoxic than the water-soluble GVP fraction, but showed a significant effect in a dose-dependent manner. The results of this study showed that the direct exposure of cultivated cells at the air-liquid interface offers the possibility to analyze the biological and toxicological activities of all fractions of cigarette smoke including the water-insoluble GVP fraction.
Asunto(s)
Contaminantes Atmosféricos/toxicidad , Material Particulado/toxicidad , Contaminación por Humo de Tabaco/efectos adversos , Pruebas de Toxicidad/métodos , Aire/análisis , Animales , Células CHO , Técnicas de Cultivo de Célula , Supervivencia Celular/efectos de los fármacos , Cricetulus , Diseño de Equipo , Solubilidad , Pruebas de Toxicidad/instrumentación , Volatilización , Agua/químicaRESUMEN
Previous studies on the biological assessment of cigarette smoke (CS) mainly focused on the total particulate matter (TPM) collected using a Cambridge filter or gas vapor phase (GVP) bubbled through phosphate-buffered saline (PBS). To study the effects of native CS in vitro, direct exposure methods have been developed. Meanwhile, in vitro micronucleus (MN) assays have been reported to evaluate the mutagenicity of CS. The objective of this research is to investigate the MN-inducing activity of whole smoke (WS) and GVP using a whole smoke exposure system, CULTEX((R)), which allows direct exposure of cultured cells to native CS at the air/liquid interface (ALI). CS was generated according to the International Organization for Standardization (ISO; 35ml, 2s, once per 60s) or the Health Canada Intensive (HCI; 55ml, 2s, once per 30s, with complete ventilation block) regimens and Chinese hamster lung (CHL/IU) cells were then exposed to this smoke. Dosages were adjusted according to the amount of smoke entering the actual exposure position. Under both smoking regimens, WS and GVP from 2R4F reference cigarettes induced MN responses. The concept of the dosage and similar dose-response relationships between theoretical and monitored dosage values under the two regimens enabled us to compare the MN-inducing activities of cigarettes in the direct exposure assay, even in the case of various experimental settings or different TPM amounts. MN-inducing activities of 2R4F under the ISO regimen seemed to be higher than those under HCI estimated by the TPM equivalent calculated values.