Prenatal exposure to airborne polychlorinated biphenyl congeners and male reproductive health.

STUDY QUESTION: Is fetal exposure to lower-chlorinated polychlorinated biphenyls (LC-PCBs) in indoor air of private homes built with PCB-containing materials associated with semen characteristics and testicular volume in adult men? SUMMARY ANSWER: We observed only marginal and inconsistent associations between maternal exposure to PCBs in indoor air and semen quality, testicular size and reproductive hormones in the adult offspring. WHAT IS KNOWN ALREADY: Recent studies have shown LC-PCBs to exhibit endocrine-disrupting properties and increase the risk of cryptorchidism. Although exposure to LC-PCBs in indoor air is relatively common, the long-term impact of prenatal exposure on male reproductive health has not yet been investigated. STUDY DESIGN, SIZE, DURATION: In this cohort study, participants were men (18+ years) whose mothers carried them while living in one of two residential areas where indoor air had been contaminated by LC-PCB evaporating from building materials in subsets of the apartments. Men were considered prenatally exposed if their mother had lived in a PCB-contaminated apartment and unexposed if their mother had lived in an uncontaminated apartment for a minimum of 1 year during the 3.6 years before conception or during the first trimester. Mothers of prenatally unexposed men could not have lived in a contaminated apartment at any point. Recruitment lasted from 2017 to 2019. In total, 73 exposed and 111 unexposed men gave a blood and semen sample. PARTICIPANTS/MATERIALS, SETTING, METHODS: Percentage differences in semen volume, sperm concentration, total sperm count, morphologically normal spermatozoa, progressively motile spermatozoa and DNA fragmentation index (DFI) between prenatally exposed and unexposed men were estimated using negative binomial regression. Associations with total and calculated free testosterone (CFT), LH and FSH were modeled using the linear regression. Odds of small testicular volume was estimated with logistic regression. MAIN RESULTS AND THE ROLE OF CHANCE: Overall, the results of this study were conflicting. No differences in semen volume, sperm concentration, testosterone and CFT were observed between the groups, but there were slight indications of lower total sperm count, increased FSH and risk of small testicles, alongside lower sperm DFI and a higher proportion of normal spermatozoa in men exposed to LCB-PCBs from indoor air during fetal life. There is no apparent biologically plausible explanation for the apparently improved measures of DNA fragmentation and morphology, and these findings may have occurred purely by chance. LIMITATIONS, REASONS FOR CAUTION: Owing to the indirect measure of exposure, lack of adjustment for paternal factors, the potential for self-selection due to known exposure status and fertility issues, inability to take time spent away from the residence, limited statistical power and lack of comparable literature, independent replication of the study in larger cohorts is warranted. WIDER IMPLICATIONS OF THE FINDINGS: While our findings may appear reassuring for the large number of people residing and/or working in buildings with indoor air contaminated with LC-PCBs, further efforts to understand the full range of health consequences of fetal LC-PCB exposure are needed. STUDY FUNDING/COMPETING INTEREST(S): The study was supported by the Independent Research Fund Denmark (ref no. 6110-00085B), Bispebjerg Hospital, Landsbyggefonden, Realdania (ref. no. PRJ-2017-00176), Grundejernes Investeringsfond (ref. no. 18-58) and Helsefonden (ref. no. 16-B-01-22 and 21-B-0412). K.S.H. was supported by FFIKA, Focused Research Effort on Chemicals in the Working Environment, from the Danish Government. The authors declare that they have no financial, personal or professional competing interests. TRIAL REGISTRATION NUMBER: Not applicable.

Risk assessment of consumer spray products using in vitro lung surfactant function inhibition, exposure modelling and chemical analysis.

Consumer spray products release aerosols that can potentially be inhaled and reach the deep parts of the lungs. A thin layer of liquid, containing a mixture of proteins and lipids known as lung surfactant, coats the alveoli. Inhibition of lung surfactant function can lead to acute loss of lung function. We focused on two groups of spray products; 8 cleaning and 13 impregnation products, and in the context of risk assessment, used an in vitro method for assessing inhibition of lung surfactant function. Original spray-cans were used to generate aerosols to measure aerodynamic particle size distribution. We recreated a real-life exposure scenario to estimate the alveolar deposited dose. Most impregnation products inhibited lung surfactant function at the lowest aerosolization rate, whereas only two cleaning products inhibited function at the highest rates. We used inhibitory dose and estimated alveolar deposition to calculate the margin of safety (MoS). The MoS for the inhibitory products was ≤1 for the impregnation products, while much larger for the cleaning products (>880). This risk assessment focused on the risk of lung surfactant function disruption and provides knowledge on an endpoint of lung toxicity that is not investigated by the currently available OECD test guidelines.

In vitro prediction of clinical signs of respiratory toxicity in rats following inhalation exposure.

To date there are no OECD validated alternative approaches to study toxicity following inhalation exposure to airborne chemicals. The available OECD test guidelines for acute inhalation toxicity aim to estimate a value of the lethal air concentration of the test chemical leading to the death of 50% of the exposed animals (LC50), to satisfy hazard classification and labelling requirements. This paper explores the view that alternative approaches must compare to outcomes of existing guideline methods to become accepted and implemented in a regulatory context. This case study describes the initiatives taken to validate the lung surfactant bioassay, an in vitro cell-free method, and discusses the challenges faced. While the lung surfactant bioassay could not predict the GHS classification for acute inhalation toxicity of 26 chemicals, the assay successfully predicted the clinical signs of respiratory toxicity observed during or shortly after exposure in vivo as reported in registration dossiers. The lung surfactant bioassay is a promising alternative approach to assess the potential of chemicals to cause changes to respiration remaining after exposure (indicating decreased lung function), and can be combined with other test methods in an integrated approach to testing and assessment of inhaled substances.

Chronic maternal inflammation or high-fat-feeding programs offspring obesity in a sex-dependent manner.

BACKGROUND/OBJECTIVES: The current world-wide obesity epidemic partially results from a vicious circle whereby maternal obesity during pregnancy predisposes the offspring for accelerated weight gain and development of metabolic syndrome. Here we investigate whether low-grade inflammation, characteristic of the obese state, provides a causal role for this disastrous fetal programming in mice. METHODS: We exposed pregnant and lactating C57BL/6JBom female mice to either high-fat diet (HFD), or continuous infusion of lipopolysaccharide (LPS), a potent trigger of innate immunity, and studied offspring phenotypes. RESULTS: Both maternal LPS or HFD treatments rendered the offspring hyperphagic and inept of coping with a HFD challenge during adulthood, increasing their adiposity and weight gain. The metabolic effects were more pronounced in female offspring, while exposed male offspring mounted a larger inflammatory response to HFD at adulthood. CONCLUSIONS: This supports our hypothesis and highlights the programming potential of inflammation in obese pregnancies.

Airway exposure to multi-walled carbon nanotubes disrupts the female reproductive cycle without affecting pregnancy outcomes in mice.

BACKGROUND: The use of multiwalled carbon nanotubes (MWCNT) is increasing due to a growing use in a variety of products across several industries. Thus, occupational exposure is also of increasing concern, particularly since airway exposure to MWCNTs can induce sustained pulmonary acute phase response and inflammation in experimental animals, which may affect female reproduction. This proof-of-principle study therefore aimed to investigate if lung exposure by intratracheal instillation of the MWCNT NM-400 would affect the estrous cycle and reproductive function in female mice. RESULTS: Estrous cycle regularity was investigated by comparing vaginal smears before and after exposure to 67 µg of NM-400, whereas reproductive function was analyzed by measuring time to delivery of litters after instillation of 2, 18 or 67 µg of NM-400. Compared to normal estrous cycling determined prior to exposure, exposure to MWCNT significantly prolonged the estrous cycle during which exposure took place, but significantly shortened the estrous cycle immediately after the exposed cycle. No consistent effects were seen on time to delivery of litter or other gestational or litter parameters, such as litter size, sex ratio, implantations and implantation loss. CONCLUSION: Lung exposure to MWCNT interfered with estrous cycling. Effects caused by MWCNTs depended on the time of exposure: the estrous stage was particularly sensitive to exposure, as animals exposed during this stage showed a higher incidence of irregular cycling after exposure. Our data indicates that MWCNT exposure may interfere with events leading to ovulation.

Prenatal maternal stress and atopic diseases in the child: a systematic review of observational human studies.

BACKGROUND: A growing number of studies suggest that maternal stress during pregnancy promotes atopic disorders in the offspring. This is the first systematic review to address prenatal maternal stress (PNMS) and the subsequent risk of atopy-related outcomes in the child. METHODS: The review was performed in accordance to the PRISMA criteria. We searched and selected studies in PubMed, Scopus, Embase and PsychINFO until November 2014. RESULTS: Sixteen (with 25 analyses) of 426 identified articles met the review criteria. Five main PNMS exposures (negative life events, anxiety/depression, bereavement, distress and job strain) and five main atopic outcomes (asthma, wheeze, atopic dermatitis, allergic rhinitis and IgE) were assessed across the studies. Overall, 21 of the 25 analyses suggested a positive association between PNMS and atopic outcomes. Of the 11 exposure-response analyses reported, six found statistically significant trends. CONCLUSION: This systematic review suggests a relationship between maternal stress during pregnancy and atopic disorders in the child. However, the existing studies are of diverse quality. The wide definitions of often self-reported stress exposures imply a substantial risk for information bias and false-positive results. Research comparing objective and subjective measures of PNMS exposure as well as objective measures for atopic outcome is needed.

Psychosocial job strain and risk of congenital malformations in offspring--a Danish National cohort study.

OBJECTIVE: To investigate if maternal exposure to psychosocial job strain at work (high demands and low control) measured by questionnaire early in pregnancy (median week 15) is associated with malformations in the offspring. DESIGN: Population-based cohort study. SETTING: The Danish National Birth Cohort. POPULATION: A cohort of 60,386 singleton children with full information on mother's occupational status, exposure to psychosocial job strain and all covariates during pregnancy. METHODS: Logistic regression analysis was used to calculate the odds of congenital malformations as a function of job strain with adjustment for maternal age, body mass index, parity, smoking, alcohol use, manual versus nonmanual work, maternal serious disease and gestational age at interview. MAIN OUTCOME MEASURES: Circulatory malformation, musculoskeletal malformation or any malformation. RESULTS: Logistic regression analyses, both crude and adjusted, indicated no associations between working under high strain and giving birth to a child with circulatory malformation (adjusted odds ratio [OR] 1.04, 95% confidence interval [95% CI] 0.75-1.44), musculoskeletal malformation (aOR 0.88, 95% CI 0.71-1.10) or any malformation (aOR 0.99, 95% CI 0.85-1.15). Supplementary analyses including restriction to first-borns and a stratified analysis with respect to manual and nonmanual work did not change the results. CONCLUSIONS: Association between exposure to high job strain during pregnancy and elevated risk of circulatory, muscle and any malformations is not supported by this study.

Male-mediated infertility in sons of building painters and gardeners: a nationwide register-based follow-up study.

AIM: To investigate whether sons of gardeners and building painters have increased risk of infertility in comparison with sons of bricklayers, carpenters and electricians. METHODS: Participants were men born 1965-1984 in Denmark whose fathers the year before birth had worked as gardeners, painters, bricklayers, carpenters or electricians (N=22,978). Cases of infertility were identified by Danish registers, and participants were followed-up for up to 24 years after their 20th birthday. RESULTS: Sons of gardeners did not have increased risk of infertility. Hazard ratios for sons of painters fluctuated around the null in main analyses but were 1.6 (98% CI: 1.0-2.5) and 1.7 (95% CI: 0.9-3.2) in the subset of participants with smallest risk of paternal exposure misclassification. CONCLUSIONS: Working as gardener or building painter was not related to increased risk of infertility among the next generation of males in main analyses. However, inherent limitations in data may have attenuated true associations.

Gestational chronic mild stress: effects on acoustic startle in male offspring of rats.

An increasing number of scientific studies indicate that maternal stress during pregnancy influences fetal development of the nervous system and thereby the behavioural phenotype. We have previously reported attenuated prepulse inhibition (PPI) of the startle reaction in adult female rats derived from dams exposed to chronic mild stress (CMS) during gestation. In humans, decreased PPI has been reported to be associated with anxiety. Because of its potential translational value across species, the modulation of startle reactivity may be a useful tool in examining altered emotional reactivity following prenatal insults. The present study aimed at investigating whether prenatally stressed male offspring would display altered startle phenotype. Stress was induced by maternal gestational exposure to alternating procedures, i.e. CMS. At the age of 3 months, half of the offspring were blood sampled under restraint. At the age of 6 months, i.e. three months later, all animals were tested in the acoustic startle and the light enhanced startle (LES) paradigm. Control and CMS male offspring showed similar basal startle and LES levels. Maternal gestational exposure to the relatively mild, variable paradigm of stressors affected the PPI response pattern in male rats. In prenatally manipulated males, the PPI response differed statistically significantly, depending on prior exposure to an episode of postnatal acute stress (blood sampling under restraint). In contrast, the PPI response in control males was unaffected by this postnatal experience. The present work supports the hypothesis that the maternal environment is a long-term determinant of phenotypic differences in sensitivity to stressors.

Influence of diurnal phase on startle response in adult rats exposed to dexamethasone in utero.

Depression and pathological anxiety disorders are among the most prevalent neurological diseases in the world and can be precipitated and exacerbated by stress. Prenatal stress alters both behavioral and endocrine responses to stressful stimuli in later life. We have previously observed increased basal acoustic startle response (ASR) in Wistar rats exposed to stress or dexamethasone (DEX) in utero when tested during the light phase of the circadian rhythm, and decreased prepulse inhibition (PPI) in similar animals tested during the dark phase of the cycle. We speculated that this observation of increased basal startle might be influenced by diurnal phase. In the present study, adult female Sprague Dawley rats, stressed prenatally with DEX (200 µg/kg, gestational days 14-21) and postnatally by blood sampling under restraint, were tested for the ASR during both circadian phases (light and dark). Basal startle was increased in animals tested both during the light and the dark phases of the cycle. We hereby replicated our earlier findings in a new strain and laboratory, thus strengthening the validity of our model regarding prenatal stress effects on ASR in female offspring. Our results indicate that observation of increased basal ASR is not solely dependent on diurnal phase. We found no difference in hippocampal glucocorticoid and mineral corticoid receptor expression between groups.

Reduced mobility but unaffected startle response in female rats exposed to prenatal dexamethasone: different sides to a phenotype.

An adverse fetal environment is strongly associated with behavioral and emotional development in later life, and environmental interactions with the genome are essential in the development of pathophysiology. This implicates that a genetic vulnerability or other predisposition may interact with the environment and stressful life events to trigger mental disease. The startle reflex is highly sensitive to fear and anxiety in humans and animals. Elevated startle magnitude has been proposed as a marker for neurodevelopmental disorders. We have recently established an animal model for possible development of anxiety, where female rats are exposed to two stressful life events, during prenatal life and as adolescents, respectively. A blood sampling procedure 3 months prior to startle testing has previously been found to increase basal startle, but only in prenatally stressed rats. As the experimental procedure of acoustic startle response (ASR) measurement resembles the aversive blood sampling procedure, this suggests that effects on ASR may be caused by aversive contextual similarities between blood sampling under restraint and the ASR test. In the present study, postnatal blood sampling was replaced by another dissimilar stressful event. Animals exposed to a high prenatal glucocorticoid level (i.e. 150 mug dexamethasone/kg) were statistically significantly more immobile in the forced swim test (FST) than animals exposed to a lower level of dexamethasone (50 mug/kg) and control animals. Exposure to a novel contextual stressor at 3 months of age (FST) was unassociated with changes in basal startle. These data suggest, since the high prenatal dexamethasone group showed increased immobility in the FST but coped equally well with controls in the ASR, that the outcome of environmental influences is determined by the individual circumstances as different situations require different coping abilities in the same individual.

Prenatal and adult stress interplay--behavioral implications.

The origin of adult behavior and the possible pathogenesis of psychiatric disorders remain elusive, but extensive research indicates that interaction of genes and environment play a crucial role for adult phenotype. Differences in susceptibility may arise by earlier experiences and genomic variables, either alone or in combination. The acoustic startle response (ASR) has been shown to be altered in patients with several psychiatric diseases, a change that could result from a persistent sensitization caused by chronic arousal secondary to a traumatic incident. The current work hypothesized that a single aversive procedure would induce long-term hyperactivity in the HPA-axis of rats that had become vulnerable by prenatal stress, and thereby change reactivity in the ASR. Prenatal stress was achieved by maternal gestational exposure to Chronic Mild Stress (CMS). At age 3 months, the offspring were blood sampled by a stressful procedure, and subsequently tested in the acoustic startle paradigm. Prenatal CMS strongly reduced prepulse inhibition (PPI) whereas postnatal blood sampling under restraint generally increased PPI. Our data demonstrate interplay between pre- and postnatal stressful events, but also that this interaction is complex and could influence the interplay between PPI and basal startle. Our results suggest that circumstances dating back to early development may have implications for adult life behavior, and based on this we propose a new theory of a threshold in the induction of a stress response in the ASR test, which influences whether the PPI or basal startle response will be affected.

The risk of infertility among hairdressers. Five-year follow-up of female hairdressers in a Danish national registry.

BACKGROUND: One in seven married couples is involuntarily infertile. Several chemical exposures in the work environment have been hypothesized to affect female reproduction, and some are present in products used in hairdressing and related trades. Recent Swedish findings indicate that employment in hairdressing poses a risk for female reproductive function. This study examined the possible association between work as a hairdresser and subsequent hospital contact due to female infertility. METHODS: A cohort of all women in Denmark aged 20-44 years on 1 January 1998 (baseline) and registered as economically active hairdressers, according to national registers, was formed to calculate age-standardized risk ratios (RRs) for hospital contacts due to female infertility during a 5-year follow-up period. Hairdressers were compared to a standard population, that is, all economically active women in Denmark aged 20-44 years at baseline, and to women working as shop assistants. RESULTS: Sixty-eight cases of hospital contact due to female infertility were observed among the female hairdressers. On the basis of the standard population, the expected number was 73.27, which gives an observed RR of 0.928 (95% CI: 0.72-1.18). Hairdressers and shop assistants exhibited similar rates of hospital contact due to female infertility (1.01; 95% CI: 0.77-1.29). CONCLUSION: The findings are not corroborating the hypothesis that hairdressers are at increased risk of infertility, but small risks in the entire group or high risks in small subgroups may not be detected by the study.

Developmental toxicity of toluene in male rats: effects on semen quality, testis morphology, and apoptotic neurodegeneration.

In one study, pregnant Wistar rats were exposed to 1200 ppm toluene by inhalation 6 h a day from gestational day (GD) 7 to postnatal day (PND) 18. Sperm analysis was performed in the adult male offspring at PND 110 by using computer-assisted sperm analysis. Toluene did not affect the semen quality of exposed rats. In another study, pregnant rats were exposed to 1800 ppm from GD 7 to GD 20, and the male offspring were killed at PND 11, 21 or 90. Paired testes weight, histopathology and immunoexpression of vimentin in Sertoli cells were used as markers of testis toxicity. In the brain, the number of apoptotic cells in the hippocampus and cerebellum were counted after visualisation by means of the TUNEL assay. Mean body weight in pups of exposed dams was lower than in pups from control litters. This decrease was still statistically significant at PND 11, but at PND 21 and 90 the body weight of toluene-exposed males tended to approach that of the controls. Absolute and relative testes weights were reduced in all three age groups, although not to a statistically significant degree. Histopathological examinations of the testis and immuno-expression of vimentin did not reveal any differences between toluene-exposed animals and control animals. In the hippocampus, almost no apoptosis was observed in any age group, and there were no differences in apoptotic neurodegeneration between male rats exposed to 1800 ppm and control animals at PND 11, 21 or 90. Generally, a marked increase in number of apoptotic cells was observed in cerebellar granule cells at PND 21 compared with the other age groups. Toluene induced a statistically significant increase in the number of apoptotic cells in the cerebellar granule layer at PND 21. The mean was increased from 37 in the control group to 71 in the toluene-exposed group. Thus, the granular cell layer in cerebellum is a highly relevant tissue with which to study toluene-induced apoptosis, because of the continuous migration of neurons and high frequency of neuronal apoptosis during the weaning period. In summary, it is concluded, that neither pre- and postnatal exposure to 1200 ppm toluene nor prenatal exposure to 1800 ppm induced significant effects on the reproductive parameters investigated. However, prenatal exposure to 1800 ppm toluene did increase neuronal apoptosis in the cerebellum of weaned male rats, possibly by delaying postnatal migration of granule cells to their final destination, or by toluene-induced retardation of generalised fetal growth.

Effects of R-(+)- and S-(-)-limonene on the respiratory tract in mice.

The effects of airborne R-(+)- and S-(-)- limonene were studied in conscious BALB/c mice by continuous monitoring respiratory rate (f), tidal volume (VT) and mid-expiratory flow rate (VD) during an exposure period of 30 min. Both enantiomers decreasedf from a trigeminal reflex, i.e., due to sensory irritation. The exposure concentration decreasing f by 50% (RD50) in the first 10 min of the exposure period was estimated to be 1,076 ppm for R-(+)-limonene and 1,467 ppm for S-(-)-limonene. Results for sensory irritation of R-(+)-limonene in BALB/c mice and humans are in close agreement. The reported sensory irritation threshold is above 80 ppm in humans while the no-observed-effect level was estimated to be 100 ppm in mice. The enantiomers were devoid of pulmonary irritation or general anesthetic effects with R-(+)-limonene < or =1,599 ppm and S-(-)-limonene < or =2,421 ppm. R-(+)-limonene did not influence VT below 629 ppm. S-(-)-limonene increased VT above 1,900 ppm. Both enantiomers induced a mild bronchoconstrictive effect above 1,000 ppm.

Developmental neurotoxicity after toluene inhalation exposure in rats.

Rats were exposed to 1200 ppm or 0 ppm toluene (CAS 108-88-3) for 6 h per day from day 7 of pregnancy until day 18 postnatally. Developmental and neurobehavioral effects in the offspring were investigated using a test battery including assessment of functions similar to those in the proposed OECD TG for Developmental Neurotoxicity Study, i.e., physical development, reflex ontogeny, motor function, motor activity, sensory function, and learning and memory. The exposure did not cause maternal toxicity or decreased viability of the offspring. Lower birth weight, delayed ontogeny of reflexes, and increased motor activity in the open field was registered in the exposed offspring. Impaired cognitive function was revealed in the exposed female offspring at the age of 3.5 months, i.e., they used more time to locate the hidden platform in the Morris water maze after platform relocation. The difference was not related to poorer swimming capabilities, because swim speeds were similar to control values. The results show that exposure to 1200 ppm toluene during brain development caused long-lasting developmental neurotoxicity in rats.

Acute airway effects of formaldehyde and ozone in BALB/c mice.

1. Concentration and time-effect relationships of formaldehyde and ozone on the airways were investigated in BALB/c mice. The effects were obtained by continuous monitoring of the respiratory rate, tidal volume, expiratory flow rate, time of inspiration, time of expiration, and respiratory patterns. 2. With concentrations up to 4 p.p.m., formaldehyde showed mainly sensory irritation effects of the upper airways that decrease the respiratory rate from a trigeminal reflex. The no-effect level (NOEL) was about 0.3 p.p.m. This value is close to the human NOEL, which is about 0.08 p.p.m. 3. Ozone caused rapid, shallow breathing in BALB/c mice. Later on, the respiratory rate decreased due to another vagal response that indicated an incipient lung oedema. The NOEL in mice was about 1 p.p.m. during 30 min of ozone exposure. No major effect occurs in resting humans at about 0.4 p.p.m. 4. Thus, the upper airway irritant, formaldehyde, and the deep lung irritant, ozone, showed the same types of respiratory effects in humans and in BALB/c mice. Also, the sensitivity was nearly identical. Continuous monitoring of respiratory effects in BALB/c mice, therefore, may be a valuable method for the study of effects of other environmental pollutants, which, however, should be confirmed in further studies.

Effects of prenatal exposure to toluene on postnatal development and behavior in rats.

Development and neurobehavioral effects of prenatal exposure to toluene (CAS 108-88-3) were studied after exposing pregnant rats (Mol:WIST) to 1800 ppm of the solvent for 6 h daily on days 7-20 of gestation. Body weights of exposed offspring were lower until day 10 after parturition. Neurobehavioral evaluation of the pups revealed no effects on motor function (rotarod), activity level (open field), acoustic startle, and prepulse inhibition. Measurements of hearing function using auditory brain stem response revealed small effects in male-exposed offspring. Performance in a Morris water maze during initial learning gave some indications of impaired cognitive functions, which was confirmed during further testing, especially in reversal and new learning. Effects on cognitive functions seemed most marked in female offspring.