To Örebro University

Inflammation is one of the vital mechanisms through which the immune system responds to harmful stimuli. During inflammation, pro and anti-inflammatory cytokines interplay to orchestrate fine-tuned, dynamic immune responses. The cytokine interplay governs switches in the inflammatory response and dictates the propagation and development of the inflammatory response. Molecular pathways underlying the interplay are complex, and time-resolved monitoring of mediators and cytokines is necessary as a basis to study them in detail. Our understanding can be advanced by mathematical models which enable to analyze the system of interactions and their dynamical interplay in detail. We, therefore, used a mathematical modeling approach to study the interplay between prominent pro and anti-inflammatory cytokines with a focus on tumor necrosis factor (TNF) and interleukin 10 (IL-10) in lipopolysaccharide (LPS)-primed primary human monocytes. Relevant time-resolved data were generated by experimentally adding or blocking IL-10 at different time points. The model was successfully trained and could predict independent validation data and was further used to perform simulations to disentangle the role of IL-10 feedbacks during an acute inflammatory event. We used the insight to obtain a reduced predictive model including only the necessary IL-10-mediated feedbacks. Finally, the validated reduced model was used to predict early IL-10 - TNF switches in the inflammatory response. Overall, we gained detailed insights into fine-tuning of inflammatory responses in human monocytes and present a model for further use in studying the complex and dynamic process of cytokine-regulated acute inflammation.

Convergent evidence associates exposure to endocrine disrupting chemicals (EDCs) with major human diseases, even at regulation-compliant concentrations. This might be because humans are exposed to EDC mixtures, whereas chemical regulation is based on a risk assessment of individual compounds. Here, we developed a mixture-centered risk assessment strategy that integrates epidemiological and experimental evidence. We identified that exposure to an EDC mixture in early pregnancy is associated with language delay in offspring. At human-relevant concentrations, this mixture disrupted hormone-regulated and disease-relevant regulatory networks in human brain organoids and in the model organisms Xenopus leavis and Danio rerio, as well as behavioral responses. Reinterrogating epidemiological data, we found that up to 54% of the children had prenatal exposures above experimentally derived levels of concern, reaching, for the upper decile compared with the lowest decile of exposure, a 3.3 times higher risk of language delay.

Rendel F, Alfredsson CF, Bornehag CG, Sundström BE, Nånberg E. Effects of Di‐isononyl Phthalate on Neuropeptide Y Expression in Differentiating Human Neuronal Cells, Basic & Clinical Pharmacology & Toxicology 2017, Mar;120(3):318–323 (https://onlinelibrary.wiley.com/doi/full/10.1111/bcpt.12670).

The above article, published online on 13 September 2016 in Wiley Online Library (wileyonlinelibrary.com), has been retracted by agreement between the authors, the journal Editor in Chief, Kim Brøsen, the Nordic Association for the Publication of BCPT and John Wiley & Sons Ltd. The retraction has been agreed following an investigation into the accuracy of the data acquisition conducted by Karlstad University into the accuracy of the data acquisition, which concluded that manipulation had taken place during tests performed on an ELISA equipment in order to achieve a preferred result.

Phthalates are used as plasticizers in polyvinyl chloride (PVC) materials and it is known that phthalates may migrate into the surrounding environment and then become a source for human uptake. The aim of the study was to investigate whether residential PVC flooring was related to the urinary levels of phthalate metabolites determined in pregnant women. The data were from the Swedish SELMA study where sampling was conducted during the time period 2007-2010. Spot urine samples from 1674 women at the end of the first trimester were analyzed for 14 metabolites from seven phthalates and one phthalate alternative. Data on flooring material in the kitchen and the parents' bedrooms as well as potential confounders were collected by postal questionnaires at the same time as the urine samples were taken. Multiple regression modeling by least square geometric mean and weighted quantile sum regression was applied to log-transformed and creatinine-adjusted phthalate metabolite concentrations adjusted for potential confounders from questionnaire data. This study has found significantly higher urinary levels of the BBzP metabolite (MBzP) in pregnant women living in homes with PVC flooring as compared to homes with other flooring materials.

Background: The general population is exposed to phthalates, a group of chemicals with strong evidence for endocrine disrupting properties, commonly used in a large number of consumer products. Based on published research and evidence compiled by environmental agencies, certain phthalate applications and products have become restricted, leading to an increasing number of “new generation compounds” coming onto the market during recent years replacing older phthalates. Some examples of such newer compounds are di-iso-nonyl phthalate (DiNP), di-iso-decyl phthalate (DiDP), and most recently di-isononyl-cyclohexane-1,2-dicarboxylate (DiNCH).

Objectives: In order to evaluate temporal trends in phthalate exposure, first trimester urinary biomarkers of phthalates were measured in the Swedish SELMA study over a period of 2.5 years (2007–2010).

Methods: We collected first morning void urine samples around week 10 of pregnancy from 1651 pregnant women. Spot samples were analyzed for 13 phthalate metabolites and one phthalate replacement and least square geometric mean (LSGM) levels of the metabolites were compared between the sampling years when adjusted for potential confounders.

Results: All 14 metabolites were detectable in more than 99% of the SELMA subjects. The levels were generally comparable to other studies, but the SELMA subjects showed slightly higher exposure to butyl-benzyl phthalate (BBzP) and di-butyl phthalate (DBP). Di-ethyl-hexyl phthalate (DEHP) metabolites levels decreased while DiNP, DiDP/di-2-propylheptyl phthalate (DPHP), and DiNCH metabolites levels increased during the sampling period.

Conclusions: Urinary metabolite levels of the older phthalates and more recently introduced phthalate replacement compound changed during the short sampling period in this Swedish pregnancy cohort. Our results indicate that replacement of phthalates can make an impact on human exposure to these chemicals. During this particularly vulnerable stage of life, phthalate exposures are of particular concern as the impacts, though not immediately noticeable, may increase the risk for health effects later in life.

Neuropeptide Y (NPY) is an abundant neuropeptide in the mammalian brain important for behavioural consequences of stress and energy metabolism. We have addressed possible effects of the phthalate DiNP on NPY expression in human SH‐SY5Y cells, a neuronal in vitro differentiation model. Pico‐ to nanomolar doses of DiNP and its metabolite MiNP resulted in decreased NPY mRNA and peptide expression in retinoid‐differentiated cells. Thus, dys‐regulated NPY may be an adverse outcome for exposure to low doses of DiNP in human beings.

Ellagic acid has previously been reported to induce reduced proliferation and activation of apoptosis in several tumor cell lines including our own previous data from non-differentiated human neuroblastoma SH-SY5Y cells. The aim of this study was now to investigate if in vitro differentiation with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate or the vitamin A derivative all-trans retinoic acid altered the sensitivity to ellagic acid in SH-SY5Y cells. The methods used were cell counting and LDH-assay for evaluation of cell number and cell death, flow cytometric analysis of SubG(1)-and TUNEL-analysis for apoptosis and western blot for expression of apoptosis-associated proteins. In vitro differentiation was shown to reduce the sensitivity to ellagic acid with respect to cell detachment, loss of viability and activation of apoptosis. The protective effect was phenotype-specific and most prominent in all-trans retinoic acid-differentiated cultures. Differentiation-dependent up-regulation of Bcl-2 and integrin expression is introduced as possible protective mechanisms. The presented data also point to a positive correlation between proliferative activity and sensitivity to ellagic-acid-induced cell detachment. In conclusion, the presented data emphasize the need to consider degree of neuronal differentiation and phenotype of neuroblastoma cells when discussing a potential pharmaceutical application of ellagic acid in tumor treatment.

Diisononyl phthalate (DINP) is a plasticizer that is frequently used as a substitute for other plasticizers whose use is prohibited in certain products. In vivo studies on the neurotoxicity of DINP are however, limited. This work aims to investigate whether DINP causes neurobehavioral changes in mice and to provide useful advice on preventing the occurrence of these adverse effects. Behavioral analysis showed that oral administration of 20 or 200â mg/kg/day DINP led to mouse cognitive deficits and anxiety. Brain histopathological observations, immunohistochemistry assays (cysteine-aspartic acid protease 3 [caspase-3], glial fibrillary acidic protein [GFAP]), oxidative stress assessments (reactive oxygen species [ROS], glutathione [GSH], superoxide dismutase [SOD] activities, 8-hydroxy-2-deoxyguanosine [8-OH-dG] and DNA-protein crosslinks [DPC]), and assessment of inflammation (tumor necrosis factor alpha [TNF-Crossed D sign°[ and interleukin-1 beta [IL-1β]) of mouse brains showed that there were histopathological alterations in the brain and increased levels of oxidative stress, and inflammation for these same groups. However, some of these effects were blocked by administration of melatonin (50â mg/kg/day). Down-regulation of oxidative stress was proposed to explain the neuroprotective effects of melatonin. The data suggests that DINP could cause cognitive deficits and anxiety in mice, and that melatonin could be used to avoid these adverse effects.

Di(2-ethylhexyl) phthalate (DEHP) is a plasticizer commonly used in PVC that may leach into the environment, and has been shown to adversely affect the health of humans and animals. We undertook a study to ascertain the neurotoxicity of DEHP in Kunming mice. This study included three rounds of testing. In the first round, Kunming mice were exposed to different concentrations of DEHP (0, 5, 50, 500 mg kg(-1) per day) after which their cognitive ability was assessed using the Morris water maze (MWM) test. The reactive oxygen species (ROS) content in tissue and the malondialdehyde (MDA) content of brains were also measured. In the second round, vitamin E (50 mg kg(-1) per day) was given daily as an anti-oxidant via the intragastric route. Cognitive deficits and locomotor activity, as well as ROS and MDA contents were tested employing the same methods. In the third round, the depressive mood of mice after DEHP exposure (500 mg kg(-1) per day) was measured using the open field test, the tail suspension test, and the forced swim test. The main findings of this study include: (1) a statistical association exists between DEHP oral exposure and spatial learning (DEHP 500 mg kg(-1) per day) and memory (DEHP 50 mg kg(-1) per day) dysfunction as ascertained by an MWM test of Kunming mice. (2) A statistical association was also found between DEHP oral exposure (50 and 500 mg kg(-1) per day) and oxidative stress (ROS and MDA) of mouse brain tissue. (3) Co-administration of vitamin E (50 mg kg(-1) per day) diminishes the elevation of ROS and MDA induced by DEHP (50 mg kg(-1) per day) from significant levels to non-significant levels. (4) Co-administration of vitamin E (50 mg kg(-1) per day) protects against mouse memory dysfunction induced by DEHP (50 mg kg(-1) per day) from being significant to being not significant. (5) In the 5 mg kg(-1) per day DEHP exposure groups, oxidative stress in brain tissue, and neurobehavioral changes were not found. (6) High dose DEHP exposure (500 mg kg(-1) per day) may induce behavioral despair in mice. Conclusions: These data suggest that DEHP is neurotoxic with regard to cognitive ability and locomotor activity.

The polyphenol ellagic acid is found in many natural food sources and has been proposed as a candidate compound for clinical applications due to its anti-oxidative capacity and as a potential anti-tumorigenic compound. The objective of the present study was to evaluate the sensitivity to and possible apoptosis mechanism induced by ellagic acid in neuronal tumor cells. As a model the human neuroblastoma SH-SY5Y cell line was used. The methods applied were bright field and phase contrast microscopy, XTT- and LDH-assays, western blot, and flow cytometric analysis of DNA degradation and mitochondrial membrane potential. Ellagic acid treatment was found to induce a reduction in cell number preceded by alterations of the mitochondrial membrane potential and activation of caspase-9 and -3, DNA-fragmentation and cell death by apoptosis. The apoptotic cell death studied was not due to anoikis since it was significant in the adherent fraction of the cells. We conclude that ellagic acid induces dose- and time-dependent apoptosis, at least partly by the mitochondrial pathway, in an embryonal neuronal tumor cell system. This finding is in agreement with previously reported data on adult carcinoma cells thus suggesting a more general effect of ellagic acid on tumor cells.