Micronucleus induction in V79 cells after direct exposure to whole cigarette smoke

Abstract

Previous investigations on the effects of cigarette smoke on cultured cells have used mainly smoke condensate dissolved in culture medium. A system has been designed which allows direct exposure of cells to fresh cigarette smoke, without an intervening layer of growth medium between the cells and the smoke. Preliminary results have been obtained which demonstrate the viability of the system. V79 cells were cultured on porous membranes (Transwell; Costar). During smoke exposure only the lower surface of each Transwell is supplied with culture medium from the bottom of the culture chambers. In this way the cells had direct contact with the atmosphere at the upper surface and could be exposed directly to the test compound. The constructed exposure system consists of a smoke generator and an exposure unit containing six Transwells, the latter contained in an incubator. Cigarette smoke was generated using a standard 2 s, 35 ml puff once per min. The puff is diluted with conditioned air from the incubator and injected into the exposure unit. Following exposure of the cells to air only for 3 h there was no effect upon V79 cell viability. However, after exposure to smoke containing between 88 and 224 mg/m3 particulate matter, an inhibition of cell proliferation and induction of micronuclei was measured. When a Cambridge filter pad was placed between the cigarette and the cell exposure system to remove particulate matter cell proliferation was also reduced and an increased frequency of micronuclei above the control value was measured.

In-vitro cell exposure studies for the assessment of nanoparticle toxicity in the lung—A dialog between aerosol science and biology

“…A variety of such ALI cell exposure systems have been described in the literature Mohr, 1999, 2000;Aufderheide, 2005;Bé ruBé et al, 2009;Fukano et al, 2004;Massey et al, 1998: Bitterle et al, 2006Mülhopt et al, 2008;Rothen-Rutishauser et al, 2009aLenz et al, 2009). While most of them rely on diffusion and/or gravitational settling as deposition mechanisms, some exposure systems have been introduced utilizing electrostatic deposition for bipolarly or unipolarly charged particles (Savi et al, 2008;Stevens et al, 2008;de Bruijne et al, 2009).…”

“…The CULTEX s glass modules have been used in a wide variety of studies on the biological effects of complex mixtures like diesel exhaust, cigarette smoke, volatile compounds and particles after direct exposure of cells cultivated at the airliquid interface (ALI) (Aufderheide et al, , 2003aAufderheide, 2005Aufderheide, , 2008Aufderheide and Mohr, 2004;Diabaté et al, 2008;Fukano et al, 2004;Massey et al, 1998;Pariselli et al, 2006Pariselli et al, , 2009aPariselli et al, , 2009bSeagrave et al, 2005Seagrave et al, , 2007Wolz et al, 2002). A very important feature in ALI exposure systems is the supply of the cell cultures with nutrients.…”

Characterisation of an aerosol exposure system to evaluate the genotoxicity of whole mainstream cigarette smoke using the in vitro γH2AX assay by high content screening

“…The selection of the RM20S® 8-syringe smoking machine as the WMCS exposure system was based on previous in vitro studies [5,24] and thorough evaluations of precision, accuracy, repeatability and reproducibility [25,26]. The smoking exposure system is schematically represented in Figure 1.…”

Cigarette Mainstream Smoke: The Evolution of Methods and Devices for Generation, Exposure and Collection

“…What would one see if the evolution of methods and devices for the generation and collection of, and exposure to, cigarette mainstream smoke was attentively followed, critically appraised and put into proper perspective? For instance: 1843 First meaningful experimental studies on tobacco "combustion" (1, 2); correct proportional elemental composition of nicotine established 1908 The first cigarette mainstream smoke inhalation study with rabbits (3) 1936 Bradford's "robot for mass smoking" using standard parameters, namely a 35-mL puff in 2 sec every minute (4) 1939 The first mouse skin painting study with genuine cigarette smoke condensate (5) 1959 The Cambridge glass fiber filter suggested for the quantitative collection of mainstream smoke particulate matter (6) 1959 The preparative 600-port "mammoth" machine for smoking up to 20,000 cigarettes per day (7) 1967 The linear 20-port syringe smoking machine (8) adopted by the U.S. tobacco industry as single design for routine analytical studies 1967 The commencement of formal cigarette testing by the U.S. Federal Trade Commission (FTC) using the "Cambridge Filter Method" (9) 1977 The first edition of ISO 3308, defining smoking parameters and specifying requirements for a routine analytical smoking machine (10) 1991 A CORESTA collaborative study (11) completed with harmonized smoking machines complying with ISO 3308:1991 (12) 1998 The first direct exposure of a mammalian lung cell line to cigarette mainstream smoke at the air-liquid interface (13). This list, however, is just a glimpse into the multifarious developments of the devices and methods under review; and there is more to it.…”

“…On the technical side, it was noted that a considerable portion of particulate matter was obviously lost by deposition, especially in the tubing of the smoke distribution lines; the overall loss was estimated to be as high as 36%. A study similar to the one just discussed (13) for "an exposure device", which obviously followed the principles underlying the exposure chamber described by MASSEY et al (13) though with a quite differently designed top portion featuring a horizontal discshaped metal plate for homogeneous smoke dispersion and three holes in the base for housing inserts. This chamber (Figure 49) -called "BAT exposure chamber" in several publications -was used in a number of BAT research projects (472)(473)(474)(475)(476)(477), discussed below, in combination with the Borgwaldt smoking machine RM20S.…”