Bertil Pettersson

Effects of tobacco smoke compounds on the noradrenaline induced oxidative metabolism in isolated brown fat cells

The effect on cell metabolism of 320 individual smoke components have been investigated by measuring their inhibition of noradrenaline induced respiration in isolated hamster brown fat cells. The compounds are representative of the gaseous and semivolatile phases of tobacco smoke. The strongest inhibitors were found within the groups of aliphatic alcohols, aldehydes and acids, of alkylated phenols and indoles and of alpha,beta-unsaturated aliphatic aldehydes and ketones. Some of the aliphatic aldehydes and acids significantly increased the basal respiration of the cells, probably by acting as substrates and/or uncoupling of mitochondrial respiratory control.

Evaluation of the biological activity of cigarette‐smoke condensate fractions using six in vitro short‐term tests

The biological activity of the volatile part of the particulate phase of cigarette-smoke condensate, the semivolatile fraction, has been examined, since the constituents of this material are accessible to selective filtration. Such a process offers a possibility to reduce the biological activity of total cigarette smoke without appreciably affecting the taste. Cigarette-smoke condensate, obtained from domestic American blend type cigarettes, was therefore separated into a nonvolatile and a semivolatile fraction, and the latter was fractionated by liquid-liquid extractions into four subfractions; acids, phenols, bases, and neutrals. The biological activity of these fractions was investigated using six in vitro short-term tests, of which two, the Ames test and the induction of sister chromatid exchanges, provided information on their genotoxicity, and the other four provided information on their cytotoxicity by measuring inhibition of cell growth, inhibition of oxidative metabolism, membrane damage, and ciliotoxicity. Sister chromatid exchanges were found to be induced by the total condensate, the nonvolatile and the semivolatile fractions, and the subfractions derived from the semivolatile fraction, except the bases. The Ames test showed the total condensate and the nonvolatile fraction to contain direct-acting base-pair mutagens as well as indirect-acting frameshift mutagens. While the semivolatile fraction was found nonmutagenic, two of its subfractions, acids and phenols, were shown to contain base-pair mutagens, which did not require metabolic activation. The total condensate and the nonvolatile and semivolatile fractions showed similar activity in the four cytotoxicity tests. Of the semivolatile subfractions, the acids and the phenols exhibited the highest activity and the bases the lowest; the toxicity observed for the neutrals varied with the test system used.

In vitro studies of biological effects of cigarette smoke condensate: I. Genotoxic and cytotoxic effects of neutral, semivolatile constituents

Much of the biological activity of cigarette smoke resides in the neutral fraction of the particulate phase. Since the volatile constituents of this material, the semivolatiles, are accessible to selective filtration, some of the biological activity of cigarette smoke might be reduced. In view of this, the genotoxic and cytotoxic effects and the chemical composition of the semivolatile neutral material of a cigarette smoke condensate was investigated. Cigarette smoke condensate obtained from domestic American blend type cigarettes, was separated into a volatile, a nonvolatile and a semivolatile fraction. The semivolatile constituents were fractionated by liquid-liquid extraction into 4 subfractions: acids, phenols, bases and neutrals. The neutral material was separated further by silica gel chromatography into 7 subfractions of varying polarity. The major components of these were identified by gas chromatography-mass spectrometry. These fractions were studied using 4 in vitro short-term tests, of which 2, the Ames test and induction of sister-chromatid exchanges, provided information on their genotoxicity and the other 2 provided information on their cytotoxicity by measuring inhibition of cell growth and inhibition of oxidative metabolism. Sister-chromatid exchanges were induced by the neutral fraction and the 7 subfractions, the activities of which increased with increasing polarity. Neither the total neutral material, nor the subfractions showed any mutagenic activity in the Ames test. The cytotoxic effect of the fractions of medium polarity, was greater than that of the total neutral material, while the most and the least polar fractions were less toxic.

An evaluation of the utility of four in vitro short term tests for predicting the cytotoxicity of individual compounds derived from tobacco smoke

An evaluation of results obtained earlier and now complemented to provide information on the activity of 305 compounds in four in vitro tests has been undertaken. Biological data based on the effects of these compounds on cell multiplication, oxidative metabolism, ciliary activity and membrane permeability are compared with a view to clarifying intersystem similarities and differences and, on the basis of a mean activity parameter, evaluating structure-activity and functionality-activity relationships.High mean activity is observed for 59 compounds, of which 18 are phenols, 14 aldehydes, 8 N-heterocyclics, 7 alcohols and 5 hydrocarbons. The medium and low mean activity groups comprise 105 and 131 members, respectively, and both include representatives of all functionalities examined. Delineation of the 305 compounds using 45 descriptors, and computer-assisted matching of these and any combination of them against the mean activity showed the most toxic single descriptor group to be terpenoids followed by indoles and naphthalenes, and the most toxic two-descriptor group to be α, β-unsaturated carbonyls followed by n-alkyl alcohols, aldehydes and acids.Examination of intersystem similarities and differences, using a high-medium-low scale, shows that all four systems give the same result for 35 percent of the compounds, three systems for 41 to 48 percent of the compounds, and two systems for 53 to 64 percent of the compounds. Of all compounds, 16 percent exhibit a high activity in one system and a low activity in the other three, or vice versa. Nearly, half od these discrepancies are caused by the membrane permeability system showing diverging results, while none of them are related to the brown fat cell system. The last test system is found to best represent the mean activity obtained from the four systems and the one of choice if reducing the number of tests from four to one.

Isolated hamster brown fat cells: a bioassay for toxicity tests at the cellular level. Effect of cigarette smoke condensates from high and low tar yield cigarettes.

A test system for the detection of substances with cell-irritating or cell-damaging effects using isolated brown fat cells is presented and the effects of cigarette smoke condensates (CSC) from high and low tar yield cigarettes on this system examined. The condensates inhibited noradrenaline stimulated respiration in a dose-dependent manner. Ethanol solutions of the condensates had a stronger inhibitory effect than DMSO solutions. The CSC strongly inhibited the mitochondrial function but other cellular functions were also impaired.

The evaluation of air sample extracts on the phagocytosis of alveolar macrophages and studies of macrophage-mediated mutagenesis in co-cultivated V79 Chinese hamster cells

Abstract The purpose of utilizing rabbit pulmonary alveolar macrophages (PAMs) in screening air sample extracts for cytotoxicity, was to evaluate particulate air pollutants from a biological point of view, using a relevant cell type. Phagocytic capacity of PAMs was used as the test parameter. The cytotoxicity of extracts of particulate air pollutants (φ 50 ) was determined and expressed as μg particles per mL medium. The mean values of samples representing five tested categories of environmental conditions was evaluated: a rural area, IC = 471 μ g/mL; a city street, IC 50 = 270 μ g/mL; a roof of a five-storied city building, IC 50 = 124 μ g/mL; a tunnel for automotive traffic, IC 50 = 143 μ g/mL; gasoline exhaust particles, IC 50 = 5 μ g/mL. Since air sample extracts have been shown to be mutagenic, the influence of particle ingestion (phagocytosis) on macrophage-mediated mutagenesis of polycyclic aromatic hydrocarbons in V79 Chinese hamster cells was studied. The results suggest that the mutagenicity of 7,8-dihydroxy-7,8-dihydro-benzo(α)pyrene is enhanced tenfold when PAMs are stimulated with particles. The mechanism by which the phagocytic uptake of particles by the PAMs enhanced the mutagenicity of the PAH can so far only be a matter for speculation.

SUCCINATE OXIDATION IN HAMSTER BROWN ADIPOCYTES.

In a suspension of isolated hamster brown adipocytes succinate can be oxidized at a high rate. This oxidation capacity is found to be mainly extracellular and can be totally blocked by the SH-reagent DTNB. In the intact cell succinate is found to be oxidized very slow probably as a function of a low transport rate over the membrane(s). Succinate potentiates, however, NE-induced respiration by supplying a condensing partner for entry into the citric acid cycle, thereby promoting a higher rate and extent of acetyl CoA oxidation.