Should intake of carbon monoxide be used as a guide to intake of other smoke constituents?

The relation between blood carboxyhaemoglobin (COHb) and plasma nicotine concentrations was studied in a group of 12 smokers smoking cigarettes of three levels of standard delivery. While the intake of carbon monoxide from a single cigarette was unrelated to the intake of nicotine, presmoking "trough" concentrations of the two substances (reflecting longer-term exposure) were highly correlated. Various other measures of nicotine exposure were at best only moderately correlated with blood nicotine concentrations. Thus trough COHb concentrations might be used to provide a reliable indication of the exposure to nicotine of individual smokers smoking the same type of cigarette, and of the relative exposure to nicotine of populations smoking cigarettes of different standard deliveries.     

Assessment of the Chemical Quality of Recreational Bathing Water in Argentina by Health Risk Analysis

Del Azul creek (Argentina) is a natural water body used for recreational bathing in which heavy metals and pesticides have been detected. The aim of the study is to estimate the probabilistic non-cancer and cancer risks by recreational bathing, applying U.S. Environmental Protection Agency models for aggregated (exposure through accidental oral intake of water and dermal contact) and cumulative risks (combined exposure to groups of substances) for bathers of 5, 10, 15, and 20 years old. Bathing in Del Azul creek does not generate risks at the concentrations and the exposure scenarios considered. Cypermethrin and arsenic and heptachlor were the riskiest non-cancer and cancer substances, respectively. Our study highlights the importance of considering both routes of exposure because of the great significance of the dermal route and because of the variability of population characteristics, as it has been stated in other studies. These considerations are highly significant for Argentina, where the quality of recreational water for other than microbiological causes is frequently evaluated based only on the harmful oral intake and applied to a hypothetical individual representative of the population.     

Critical evaluation and statistical validation of a hydroponic culture system for Arabidopsis thaliana.

Arabidopsis thaliana is one of the most widely used model organisms in plant sciences. Because of the increasing knowledge in the understanding of its molecular pathways, a reproducible and stable growth set-up for obtaining uniform plants becomes more important. In order to be able to easily harvest and study both roots and shoots, and to allow simple exposure to water-soluble toxic substances, a hydroponic system is the desired cultivation method for controlled plant growth. Based on earlier developed hydroponic cultivation protocols, a hydroponic set-up was optimized and statistically validated using linear mixed-effects models. In order to determine important components that influence the level of variability in a hydroponic set-up, stress-related indicators were examined at the biochemical as well as at the molecular level. It is highly recommended that statistical as well as biological assumptions are carried out before post-analyses are performed. Therefore, we suggest a model where factors that influence variability such as the usage of different pots and harvesting on different times are taken into account in the analyses. Furthermore, in contrast to what has been reported in earlier studies, our findings indicate that continuous aeration of the hydroponic solution is highly important.     

Muscle blood flow during isometric activity and its relation to muscle fatigue

The effect of isometric exercise on blood flow, blood pressure, intramuscular pressure as well as lactate and potassium efflux from exercising muscle was examined. The contractions performed were continuous or intermittent (5 s on, 5 s off) and varied between 5% and 50% maximal voluntary contraction (MVC). A knee-extensor and a hand-grip protocol were used. Evidence is presented that blood flow through the muscle is sufficient during low-level sustained contractions (less than 10% MVC). Despite this muscle fatigue occurs during prolonged contractions. One mechanism for this fatigue may be the disturbance of the potassium homeostasis. Such changes may also play a role in the development of fatigue during intermittent isometric contractions and even more so in the recovery from such exercise. In addition the role of impaired transport of substances within the muscle, due to long-lasting daily oedema formation, is discussed in relation to fatigue in highly repetitive, monotonous jobs.     

Teratogenic effects of environmental chemicals.

Despite the widespread distribution of a great many chemical substances in the environment, very few have been implicated in human teratogenicity, and most of these are drugs used at relatively high biological effect levels. Although several other types of environmental chemicals such as pesticides, solvents, and metals can be shown under laboratory conditions to have some teratogenic potential, there is little evidence that these, at present ambient concentrations and conditions of exposure, represent significant hazards to human intrauterine development. An exception to the latter generalization is methylmercury which, because of peculiarities in distribution, can reach high concentration in the human diet.     

Changes in hypothalamic noradrenaline and 5-hydroxytryptamine levels during the development of warm- and cold-adaptation in the rat

Hypothalamic 5-hydroxytryptamine (5-HT) and noradrenaline (NA) as well as plasma corticosterone levels were studied in male rats after 1, 2, 4 and 6 weeks of exposure to 4--7 or 30--31 degrees C. An increase of the NA concentration and a decrease of the 5-HT level was observed after the first week in both cold and warm environment together with an increase of plasma corticosterone levels in both groups. NA, 5-HT and plasma corticosterone levels returned to normal in cold-exposed animals by the 6th week whereas in warm-acclimated rats NA and corticosterone levels regained their initial values and 5-HT concentrations remained low. Changes by the end of the first week of exposure may result from the thermal stress. The low 5-HT levels of warm-adapted animals at the end of the 6th week were probably secondary to the process of adaptation.     

Intermittent High Glucose Stimulate MCP-l,IL-18, and PAI-1, but Inhibit Adiponectin Expression and Secretion in Adipocytes Dependent of ROS

Elevated circulating concentrations of interleukin-18 (IL-18), monocyte chemoattractant protein-1 (MCP-1), and plasminogen activator inhibitor-1 (PAI-1) and decrease of adiponectin are associated with obesity-related diseases. The mechanism that mediates the aberrant production of the adipokines remains poorly understood. The aim of this study was to investigate the effect of intermittent high glucose on the expression of IL-18, MCP-1, and PAI-1 and adiponectin in 3T3-L1 adipocytes. 3T3-L1 adipocytes were incubated for 24 h in media containing different glucose concentrations: 5 mmol/l, 20 mmol/l and a daily alternating 5 or 20 mmol/l glucose, with or without the addition of 1.0 mmol/l N-acetylcysteine (NAC). The expression and secretion of IL-18, MCP-1, PAI-1, and adiponectin were determined by real-time RT-PCR and ELISA, respectively. The production of reactive oxygen species (ROS) and 8-hydroxydeoxyguanosine (8-OHdG) were measured. Stable high glucose significantly increased expression and secretion of IL-18, MCP-1, and PAI-1, and reduced adiponectin expression and secretion compared to normal glucose conditions. These effects were significantly greater under intermittent high glucose conditions compared to stable high glucose. The level of ROS and 8-OHdG were significantly elevated under both intermittent and stable high glucose conditions, the effect being greater under intermittent high glucose. The intermittent glucose was more effective in triggering the generation of ROS than stable high glucose. The adding of the NAC, a specific pharmacological inhibitor of ROS, normalized the expression of these adipokines and the levels of ROS and 8-OHdG under both stable and intermittent glucose conditions. Intermittent high glucose induces a greater aberrant production of key adipokines than stable high glucose, and this effect seems to be related to over-production of ROS.     

Glycophorin A mutant frequency in radiation workers at the nuclear power plants and a hospital

We studied to assess the validity of the glycophorin A (GPA) mutant assay as a biological marker of the cumulative effects of chronic low doses of ionizing radiation. In 144 nuclear power plants workers and 32 hospital workers, information on confounding factors, such as age and cigarette smoking, was obtained through a self-administered questionnaire. The information on physical exposure doses was obtained from the registries for radiation exposure monitoring and control at each facility. The range of cumulative exposure doses were 0-12.02cGy. GPA mutant assay was performed by the BR6 method with modification using a FACScan flow cytometer. Potential confounders, such as, age and cigarette smoking habits showed increasing trends with GPA variants, but were not of statistical significance. The hospital workers showed higher frequency of the GPA NO variant than nuclear power plant workers. Significant dose-response relationships were found between cumulative exposure to radiation and variants levels by simple and multiple linear regression models. The slope of regression equation of the dose-response of nuclear power plants workers was much smaller than that of hospital workers. These findings suggest that there may be dose-rate effects. In a population exposed to chronic low-dose radiation, the GPA assay shows potential to be used as an effective biologic marker for assessing the cumulative exposure dose although it could not be able to see a dose relation below 10cGy of cumulative exposure dose.     

Exposure reconstruction for reducing uncertainty in risk assessment: example using MTBE biomarkers and a simple pharmacokinetic model

Adverse health risks from environmental agents are generally related to average (long-term) exposures. Because a given individual's contact with a pollutant is highly variable and dependent on activity patterns, local sources and exposure pathways, simple ‘snapshot’ measurements of surrounding environmental media may not accurately assign the exposure level. Furthermore, susceptibility to adverse effects from contaminants is considered highly variable in the population so that even similar environmental exposure levels may result in differential health outcomes in different individuals. The use of biomarker measurements coupled to knowledge of rates of uptake, metabolism and elimination has been suggested as a remedy for reducing this type of uncertainty. To demonstrate the utility of such an approach, we invoke results from a series of controlled human exposure tests and classical first-order rate kinetic calculations to estimate how well spot measurements of methyl tertiary butyl ether and the primary metabolite, tertiary butyl alcohol, can be expected to predict different hypothetical scenarios of previous exposures. We found that blood and breath biomarker measurements give similar results and that the biological damping effect of the metabolite production gives more stable estimates of previous exposure. We also explore the value of a potential urinary biomarker, 2-hydroxyisobutyrate suggested in the literature. We find that individual biomarker measurements are a valuable tool in reconstruction of previous exposures and that a simple pharmacokinetic model can identify the time frames over which an exogenous chemical and the related chemical biomarker are useful. These techniques could be applied to broader ranges of environmental contaminants to assess cumulative exposure risks if ADME (Absorption, Distribution, Metabolization and Excretion) is understood and systemic biomarkers can be measured.     

Limited use of Centritech Lab II Centrifuge in perfusion culture of rCHO cells for the production of recombinant antibody

Perfusion cultures of recombinant Chinese hamster ovary cells, producing recombinant antibody against the S surface antigen of Hepatitis B virus, were carried out in continuous and intermittent mode using a Centritech Lab II Centrifuge. In the continuous perfusion process, despite the absence of shear stress from the pump head, long-term operation was not possible because of continuously repeated exposure to oxygen limitation and low temperature, as well as shear stress from centrifugal force. In the intermittent perfusion processes, the frequency of cell-passage through the centrifuge was substantially reduced, compared with the continuous perfusion mode; however, the degree of reduction could not guarantee stable long-term operation. Although various operating parameters were applied in the intermittent perfusion cultures, high cell densities could not be maintained stably. In a single bioreactor culture system, a specific cell that is returned from the centrifuge to the bioreactor could be transferred from the bioreactor to the centrifuge again in the next cycle. These repetitive damages, caused by shear stress from the pump head and centrifugal force, as well as exposure to suboptimal conditions such as oxygen limitation and low temperature below 37 degrees C, were more serious at higher perfusion rates. Subsequently, damaged cells and dead cells were continuously accumulated in the bioreactor. Culture temperature shift from 37 to 33 degrees C increased antibody concentrations but showed inhibitory effects on cell growth. The negative effects of lowering culture temperature on cell growth overwhelmed the positive effects on antibody production. To protect cells from shear stress, Pluronic F-68 was 2-fold concentrated in the culture medium; nevertheless, a significantly higher concentration of Pluronic F-68 (2 g/L) may have inhibitory effects on cell growth.     

Dynamic resetting of the human circadian pacemaker by intermittent bright light

In humans, experimental studies of circadian resetting typically have been limited to lengthy episodes of exposure to continuous bright light. To evaluate the time course of the human endogenous circadian pacemaker's resetting response to brief episodes of intermittent bright light, we studied 16 subjects assigned to one of two intermittent lighting conditions in which the subjects were presented with intermittent episodes of bright-light exposure at 25- or 90-min intervals. The effective duration of bright-light exposure was 31% or 63% compared with a continuous 5-h bright-light stimulus. Exposure to intermittent bright light elicited almost as great a resetting response compared with 5 h of continuous bright light. We conclude that exposure to intermittent bright light produces robust phase shifts of the endogenous circadian pacemaker. Furthermore, these results demonstrate that humans, like other species, exhibit an enhanced sensitivity to the initial minutes of bright-light exposure.     

Effect of Growth Regulators and Herbicides on Photosynthetic Partial Reactions in Isolated Leaf Cells

The effect of indoleacetic acid, gibberellic acid, 2,4-dichloro-phenoxyacetic acid and amitrole at various concentration levels ranging from 0.05 to 1.0 mM on the photosynthetic evolution of O₂ and ¹⁴CO₂ fixation by isolated leaf cells was studied. The plant growth regulators enhanced O₂ evolution and ¹⁴CO₂ fixation at low concentrations and were inhibitory beyond a critical level. The amitrole had an inhibitory effect at all the concentration levels used. All the substances exhibited similar patterns of effect on the ferricyanide reduction by isolated chloroplasts and on the electron transport rates of sub-chloro-plast particles containing PS-I and PS-II independently, under non-phosphorylating conditions. As was seen from the response in all the three electron transport systems of the chloroplast studied, the electron transport chain connecting PS-II and PS-I could be considered as a possible site of action at least for the growth regulating substances as it is the only part that is common to all the three reactions. The phosphorylation associated with this part of the electron transport was “inhibited” by the substances even at the lowest concentration used. The stimulation of non-phosphorylating electron flow, with a simultaneous reduction in the rate of ATP synthesis, at low concentration levels indicated that these substances played a possible uncoupling role. The amitrole on the other hand appeared to have a generalized non-specific inhibitory action on all the partial reactions of photosynthesis.     

Paraquat and menadione exposure of rainbow trout (Oncorhynchus mykiss)--studies of effects on the pentose-phosphate shunt and thiamine levels in liver and kidney

Possible xenobiotic interactions with thiamine were studied in salmonid fish, by repeatedly injecting two model substances, paraquat and menadione, into juvenile rainbow trout (Oncorhynchus mykiss). These two substances were chosen because of their well-known ability to redox-cycle and cause depletion of NADPH in several biological systems. Depletion of NADPH increases metabolism through the pentose-phosphate shunt and may thereby increase the need for thiamine diphosphate by heightened transketolase activity. A special food was produced with lower thiamine content than commercial food, usually enriched with thiamine, which could mask an effect on the thiamine level. After 9 weeks of exposure, glucose-6-phosphate dehydrogenase, transketolase, glutathione reductase and ethoxyresorufin O-deethylase were analysed in liver and kidney cellular sub-fractions as well as analysis of total thiamine concentrations in liver, kidney and muscle. The results showed that paraquat caused a large increase in hepatic glutathione reductase activity and induced hepatic glucose-6-phosphate dehydrogenase activity, i.e., the rate-limiting enzyme in the oxidative part of the pentose-phosphate shunt. Despite this paraquat exposure did not affect transketolase activity and total thiamine concentration.     

Indices of oxidative stress. 2. Lipid peroxides

Two methods of the determination of lipid peroxidation products have been compared which are based on Fe(II) oxidation by them at acid pH values in the presence of xylenol orange which binds Fe(III) have been compared. The first method uses cumene hydropeoxide as an internal standard. In the second one, lipid peroxides are previously reduced by triphenylphosphine and these substances content is measured as a difference of the production of complexes with xylenol orange and iron ions in the control (with reduction) and experimental sample (without reduction). The optimization of measurement conditions is described. The levels of lipid peroxides in goldfish tissues assayed simultaneously by two methods were similar. The method with cumene hydroperoxide needs less amounts of biological material; moreover, there is no necessity in a calibration curve. Effects of hyperoxia on lipid peroxide levels in goldfish tissues were studied with the cumene method. Within the first hours of hyperoxia this index increased 13-times in the liver and 2-times in the brain and muscle. The further exposure rebounded this parameter to the initial level. Levels of lipid peroxides positively correlated with levels of end products of lipid peroxidation (thiobarbiturate acid reactive substances) in the goldfish tissues. The method of quantification of lipid peroxides with cumene is recommended for wide using in biological investigations.     

An experimental design for the controlled modulation of intracellular GSH levels in cultured hepatocytes

This work proposes a practical experimental approach that allows the rapid in situ generation of a wide range of intracellular GSH concentrations in the intact hepatocyte under highly reproducible conditions. The strategy involves the use of diethyl maleate, a thiol-reactive electrophile that causes rapid and extensive GSH depletion, as well as GSH monoethylester, a GSH analogue that is readily taken up by cells and deesterified intracellularly to render GSH. For both agents, we have analyzed (i) the minimal exposure time required to produce a maximal and dose-related effect on intracellular GSH without altering hepatocyte viability or subsequent survival in culture, and (ii) the relative stability of the GSH levels achieved after removal of both modulators from the culture medium. Results show that with the appropriate timing of exposure, hepatocytes from the same preparation can be quickly synchronized to provide an extreme intracellular GSH concentration gradient (from 0.1- to 4-fold the normal liver content), which simulates an in vitro GSH dose dependency curve and remains stable for at least the subsequent 4 h of culture. The experimental design, which can be easily adapted to different cell types, provides an improved testing protocol to evaluate under reliable conditions the dose-response relationships of the thiol-redox state in a variety of biological processes. It has been applied here to investigate the role of endogenous GSH content in the covalent binding of N-acetyl-p-benzoquinoneimine, the reactive species formed during CYP-dependent bioactivation of acetaminophen, to hepatic proteins.     

The distribution of (5E)-(10S)-10,19-dihydroercalciol and its metabolites in serum of rats

The distribution of radioactivity in serum of male rats (275 g) after oral administration of tritium labelled (5E)-(10S)-10,19-dihydroercalciol (dihydrotachysterol2) has been studied as a function of time and dose level. Nine distinct radioactive substances, chromatographing in the range between calciol and calcitriol, could be demonstrated. No attempt was made to establish chemical structures. After a single dose of 1.84 nmol, (5E)-(10S)-10,19-dihydroercalciol is very rapidly metabolized to more polar forms. The time course of appearance, ranging between 30 min and 1 day after administration, and the polarity of these substances indicated that they might be formed in sequence. The highest serum concentrations of the major substances occurred between 2 h and 10 h after administration, but compared with the dosage they were very low. In response to daily administration the concentrations of the major substances achieved steady-state levels within 1-4 days. The metabolism of (5E)-(10S)-10,19-dihydroercalciol was apparently not affected by its nutritional status at the dose level studied. After single administration of progressively increasing doses, ranging from 1.84 nmol to 1.84 mumol, the relative increments of the concentrations of the major substances rose in proportion to the relative increases of the dosage. The mechanisms responsible for the appearance of these substances in serum were found to be closely related. At dose levels up to 18.4 nmol feedback control was apparently absent.     

Relationships between cold hardiness and diapause, and between glycerol and free amino acid contents in overwintering larvae of the oriental corn borer, Ostrinia furnacalis

To elucidate the relationship between diapause and cold hardiness in the oriental corn borer, Ostrinia furnacalis, the levels of various substances, cold hardiness and respiration were measured in diapausing and post-diapausing overwintering larvae. Under field conditions, diapause terminated between November and January, although O(2) consumption, measured at 20 degrees C in the laboratory, remained at a high level from October to January. Glycerol content was low during October and November but greatly increased during December and January. Serine was the most abundant of the free amino acids, and its concentrations were especially high during October and November, while the concentration of alanine increased in December and January. Under laboratory conditions, glycerol levels were low in diapausing larvae, and in post-diapausing larvae that were acclimated at either high temperatures or under anaerobic conditions, while they were high in post-diapausing larvae kept under aerobic, low temperature conditions. The survival rate (cold hardiness) was strongly correlated with glycerol content but not with serine or alanine levels. These results suggest that O. furnacalis has a highly developed cold hardiness mechanism in which termination of diapause enables the larvae to increase glycerol levels when the temperature decreases.     

Serum bile acids as a sensitive biological marker for evaluating hepatic effects of organic solvents

Serum bile acids (SBAs) are suggested as a potentially sensitive and specific indicator of liver function which, accordingly, could provide an early indication of hepatobiliary dysfunction. This offers advantages over more traditional parameters of liver integrity/function. Recent studies have shown that occupational exposure to low levels of halogenated aliphatic or non-halogenated aromatic solvents is associated with significant increases in SBA levels. As this has often been evident in the absence of any effect on conventional parameters of hepatobiliary integrity/function, elevated SBA levels may well be regarded as a sensitive biological marker of exposure/effect of these compounds. In addition, it may be considered that they provide an early indicator of solvent-induced changes in hepatobiliary function. Extensive studies with experimental animals have also provided supporting evidence for these observations in solvent-exposed individuals. Investigations of the mechanisms at cellular and subcellular levels by which these increases occur have suggested that these effects are likely to be the result of selective, dose-related and reversible inhibition of bile acid uptake at the sinusoidal domain of the hepatocyte plasma membrane. Increased concentrations of SBA under low levels of exposure to different solvents have been demonstrated to be a short-lived and reversible effect which is not accompanied by any other evidence of liver damage. Therefore, it could be assumed that it is unlikely that there would be pathological sequelae to these effects, although the longer term ramifications of such effects have not been thoroughly investigated. Nevertheless, the available evidence indicates that investigation of SBA in solvent-exposed workers could provide useful indications of exposure and effect.     

Correlation between bacterial haemoglobin gene (vgb) and aeration: their effect on the growth and alpha-amylase activity in transformed Enterobacter aerogenes

AIMS: To evaluate the effects of bacterial haemoglobin on bacterial growth and alpha-amylase formation under different aeration conditions. METHODS AND RESULTS: Enterobacter aerogenes was transformed with the gene encoding Vitreoscilla (bacterial) haemoglobin, vgb. The growth kinetics and ability to synthesize alpha-amylase enzyme were investigated in this transformed Enterobacter strain as well as in two other Enterobacter control strains that do not harbour the vgb gene. Such comparison was made under variable aeration conditions, using the agitation rate as a measure of aeration. The expression of bacterial haemoglobin-supported cell growth determined as O.D.600 and cell viability in addition to the alpha-amylase production. These positive effects of bacterial haemoglobin were observed under both low and high aerations, but at different extents. CONCLUSIONS: In addition to improving cell growth under low aeration, the bacterial haemoglobin is able to promote bacterial cell tolerance during exposure to high oxygen tension. SIGNIFICANCE AND IMPACT OF THE STUDY: The expression of bacterial haemoglobin is advantageous in reducing the burden of certain toxic conditions such as high oxygen levels. It may have the same impact on some environmental toxic substances. This, haemoglobin biotechnology can be extended to induce enzymes of pollutants degradation or production of some useful industrial substances.     

Chromosomal damage in human diploid fibroblasts by intermittent exposure to extremely low-frequency electromagnetic fields

Environmental exposure to extremely low-frequency electromagnetic fields (ELF-EMFs) has been implicated in the development of cancer in humans. An important basis for assessing a potential cancer risk due to ELF-EMF exposure is knowledge of biological effects on human cells at the chromosomal level. Therefore, we investigated in the present study the effect of intermittent ELF electromagnetic fields (50 Hz, sinusoidal, 5'field-on/10'field-off, 2-24 h, 1 mT) on the induction of micronuclei (MN) and chromosomal aberrations in cultured human fibroblasts. ELF-EMF radiation resulted in a time-dependent increase of micronuclei, which became significant after 10 h of intermittent exposure at a flux density of 1 mT. After approximately 15 h a constant level of micronuclei of about three times the basal level was reached. In addition, chromosomal aberrations were increased up to 10-fold above basal levels. Our data strongly indicate a clastogenic potential of intermittent low-frequency electromagnetic fields, which may lead to considerable chromosomal damage in dividing cells.