<p>Several investigations have described adverse effects in fish inhabiting polluted areas, though the causative, or aetiologic compounds have not been substantiated. Since environmental pollutants exist as complex mixtures, reliable risk estimations are difficult to obtain. Chemical analyses of known toxicants are not sufficient since unknown toxic compounds will not be identified. Diagnostic tools – biomarkers – will provide information on the potential toxicity, but only limited information on the aetiologic compounds. A promising strategy for pinpointing the major toxicants in environmental samples is to fractionate samples in several steps, guided by the toxicity of the isolated fractions. This enables a more precise identification of aetiologic compounds than the analysis of a non-fractionated sample.</p><p>The present thesis describes the toxicological and chemical characterization of organic pollutants in the polychlorinated biphenyl (PCB) polluted bay Örserumsviken on Sweden’s Baltic coast. Compounds in organic extracts of abiotic matrices collected in the bay were separated by degree of aromaticity prior to exposing early life-stages of rainbow trout (Oncorhynchus mykiss). The fraction containing PCBs and other dicyclic aromatic compounds (DACs) was less potentially toxic than the fraction containing polycyclic aromatic compounds (PACs). An estimated 1-5% of the total ethoxyresorufin O-deethylase (EROD) induction could be explained by PCBs, and less than 4% by the most commonly monitored PACs. Though the aetiologic compounds were not identified, a significant part of the EROD induction was isolated in a PAC-subfraction containing compounds with more than five rings and the major part of teratogenicity was isolated in a fraction containing three- and four-ringed PACs. As markers for the unidentified potent compounds in the DAC- and PAC-fractions, PCBs and polycyclic aromatic hydrocarbons (PAHs) were analyzed in feral fish eggs from the bay. Adult northern pike (Esox lucius) had elevated levels of hepatic DNA adducts, which correlated with PAH concentrations in their eggs. ΣPCB concentration in pike eggs was equal to the injected concentration in the highest dose of the DAC-fraction, which caused a 50-fold EROD induction in rainbow trout larvae. This demonstrates that the compounds present in the abiotic matrices posed a threat to all life-stages of fish in this area.</p><p>The findings in this thesis clearly reveal the limits of our knowledge regarding compounds responsible for potential toxicity in field situations. We need to place greater emphasis on the toxicological and chemical implications of our continuous release of PACs and to work for much more restrictive regulations.</p>

Aquatic sediments are of major importance for the cycling of environmental pollutants, acting as both sinks and secondary sources of contaminants to the ecosystem. Sediment-living organisms can affect the fate and transport of contaminants through activities like feeding and burrowing, collectively called bioturbation. Apart from high contaminant levels, the Baltic benthic ecosystem is affected by stressors such as eutrophication-induced anoxic conditions and invading alien species. The main objectives of this thesis were to determine the effects of bioturbation on contaminant fluxes in Baltic Sea sediments and to increase the understanding of how these other stressors act together upon contaminant fate in the benthic ecosystem. Bioturbation affected contaminants in a species-specific way. The native species Monoporeia affinis and Macoma balthica increased the incorporation of BDE-99 and Cd deposited on the sediment surface, enhancing their retention in the sediment. The invasive polychaete Marenzelleria sp. did not contribute to the incorporation of surface-deposited contaminants, however, significantly increased the release of contaminants back to the water column. Reoxygenation of anoxic laminated sediments and bioturbation by Marenzelleria increased the sediment-to-water flux of dissolved organic contaminants. When the bioturbation-driven release of PCB was compared to the release caused by physical sediment resuspension, results indicated that the continuous activities of benthic infauna can be just as, or even more, important than physical disturbance for the remobilization of sediment-bound contaminants. Bioaccumulation was significantly higher when contaminants were deposited associated to phytoplankton compared to lignin or sediment, suggesting that there are likely seasonal differences in the mobilization of contaminants in the benthic ecosystem. In summary, bioturbation is an important process influencing contaminant fate in Baltic Sea sediments, and the risk of remobilization of historically buried contaminants may increase with improved benthic redox conditions and the invasion of new deeper-digging species, such as Marenzelleria.

Several investigations have described adverse effects in fish inhabiting polluted areas, though the causative, or aetiologic compounds have not been substantiated. Since environmental pollutants exist as complex mixtures, reliable risk estimations are difficult to obtain. Chemical analyses of known toxicants are not sufficient since unknown toxic compounds will not be identified. Diagnostic tools – biomarkers – will provide information on the potential toxicity, but only limited information on the aetiologic compounds. A promising strategy for pinpointing the major toxicants in environmental samples is to fractionate samples in several steps, guided by the toxicity of the isolated fractions. This enables a more precise identification of aetiologic compounds than the analysis of a non-fractionated sample. The present thesis describes the toxicological and chemical characterization of organic pollutants in the polychlorinated biphenyl (PCB) polluted bay Örserumsviken on Sweden’s Baltic coast. Compounds in organic extracts of abiotic matrices collected in the bay were separated by degree of aromaticity prior to exposing early life-stages of rainbow trout (Oncorhynchus mykiss). The fraction containing PCBs and other dicyclic aromatic compounds (DACs) was less potentially toxic than the fraction containing polycyclic aromatic compounds (PACs). An estimated 1-5% of the total ethoxyresorufin O-deethylase (EROD) induction could be explained by PCBs, and less than 4% by the most commonly monitored PACs. Though the aetiologic compounds were not identified, a significant part of the EROD induction was isolated in a PAC-subfraction containing compounds with more than five rings and the major part of teratogenicity was isolated in a fraction containing three- and four-ringed PACs. As markers for the unidentified potent compounds in the DAC- and PAC-fractions, PCBs and polycyclic aromatic hydrocarbons (PAHs) were analyzed in feral fish eggs from the bay. Adult northern pike (Esox lucius) had elevated levels of hepatic DNA adducts, which correlated with PAH concentrations in their eggs. ΣPCB concentration in pike eggs was equal to the injected concentration in the highest dose of the DAC-fraction, which caused a 50-fold EROD induction in rainbow trout larvae. This demonstrates that the compounds present in the abiotic matrices posed a threat to all life-stages of fish in this area. The findings in this thesis clearly reveal the limits of our knowledge regarding compounds responsible for potential toxicity in field situations. We need to place greater emphasis on the toxicological and chemical implications of our continuous release of PACs and to work for much more restrictive regulations.

There is a great concern about children’s health as the developing brain in foetuses and children is much more vulnerable to injury caused by different classes of chemicals than the adult brain. This vulnerability is partly due to the fact that the adult brain is well protected against chemicals by the blood brain barrier (BBB) and children have increased absorption rates and diminished ability to detoxify many exogenous compounds, in comparison to that of adults. Moreover, the development of the central nervous system (CNS) is a very complex process involving several different important events, e.g. proliferation, migration and differentiation of cells. These events are occurring within a strictly controlled time frame and therefore create different windows of vulnerability. Furthermore, the brain consists of numerous different cell types (neuronal, glial and endothelial cells) that have specific functions. The development of each cell type occurs within a specific time window and is therefore susceptible to environmental disturbances at different time periods. Evidence indicates that exposure to industrial chemicals, pesticides or drugs, contributes to the increasing incidence of neurodevelopment disorders. However, due to lack of studies only a few industrial chemicals have been identified as developmental neurotoxicants so far. The current developmental neurotoxicity (DNT) guidelines (OECD TG 426 and US EPA 712-C-98-239) are based entirely on in vivo studies that are time consuming, complex, costly and not suitable for the testing of a high number of chemicals. Applying alternative approaches such as in silico, in vitro and non-mammalian models as a part of an integrated test strategy, could speed up the process of DNT evaluation and reduce and refine animal usage. Both in vitro and non-mammalian test systems offer the possibility of providing an early screening for a large number of chemicals, and could be particularly useful in characterising the compound-induced mechanism of toxicity of various developmental processes. This thesis has characterised two primary neuronal cultures (cerebellar granule cells (CGCs) and cortical neuronal cultures) and identified them as relevant models for DNT testing, since the key processes of brain development are present, such as cell proliferation, migration and neuronal/glial differentiation. Furthermore, two emerging technologies (gene expression and electrical activity) have been evaluated and were identified as promising tools for in vitro DNT assessment. In combination with other assays they could be included into a DNT intelligent testing strategy to speed up the process of DNT evaluation mainly by prioritising chemicals with DNT potential for further testing. / The work of this thesis was performed at ECVAM, European Commission, Italy.At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 2: In press. Paper 3: In progress. Paper 4: In progress.

The aryl hydrocarbon receptor (AhR) is a ligand dependent transcription factor ubiquitously expressed in mammalian cells. It is a genetically ancient protein mostly known for binding the extremely toxic contaminant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). Binding to the AhR explains the variety of toxic responses of TCDD as well as the induction of several drug metabolizing enzymes. Induction of cytochrome P4501A1 (CYP1A1) is the most well characterized of the AhR regulated responses. The physiological functions of AhR and the endogenous ligand(s) for the receptor are under investigation but are not yet unraveled. Several tryptophan (TRP) derived indol-containing compounds have been reported to possess AhR affinity/CYP1A1 inducing capacity and TRP mediates CYP1A1 induction by UV light. The TRP photoproduct, 6-formylindolo[3,2-b]carbazole (FICZ) has the highest AhR affinity described so far and it causes a rapid and transient induction of the CYP1A1 gene in human cells. A number of reports on constitutive CYP1A1 activity in cultured cells is therefore most likely explained by the presence of TRP-derived AhR ligands in cell culture media. The aims of the studies were to investigate the impact of FICZ and FICZ metabolism on CYP1A1 gene regulation, to explore the metabolic fate of FICZ and to identify whether normal laboratory light could lead to formation of FICZ and thereby contribute to earlier observed CYP1A1 inducing effects by cell culture media. Metabolic studies using fractions of Aroclor-induced and non-induced rat liver and human liver as well as heterologously expressed enzymes revealed that FICZ can be efficiently metabolized by the CYP enzymes 1A1 and 1A2 and by an unknown cytosolic enzyme, to a number of hydroxylated and other oxidized metabolites. All of the hitherto identified 11 hydroxylated metabolites of FICZ are prone to conjugation reactions by glucuronosyltranferases and sulfotransferases. The metabolites formed by human enzymes are primarily sulfated. Thus, the sulfated metabolites of FICZ will be crucial in the future analyzes of FICZ formation in vivo. FICZ was identified to be formed, not only by UV illumination, but also by normal laboratory light. The constitutive CYP1A1 activity was significantly induced through the formation of several TRP related photoproducts in light-exposed medium. One of these photoproducts was identified as FICZ. Thus, the TRP photoproduct, FICZ, fits into a model in which FICZ auto-regulates the expression of induced enzymes. It is hypothesized that FICZ might function as a chemical messenger that activates AhR in response to light and might be one of several possible endogenous AhR ligands.

Mental disorders, like anxiety and depression, are a major health problem and can appear early in life. Developing new compounds for anxiety and other mental disorders is desirable and requires good animal models. Two widely used tests to evaluate fear and anxious behavior in rodents are the open field (OF) and elevated plus maze (EPM) tests. The models are simple and it has been argued that different environments give different behavior profiles and measure different kind of behavior. The multivariate concentric square fieldTM (MCSF) is a new test that gives the rodent opportunity to visit different environments and to evaluate more parameters in one test. Validation of the tests is mainly done with adult animals. The aim with this study is to study adolescent rat behavior in the OF, OF with start box, EPM and MCSF tests and to see if there is any difference in running trials in the morning or in the afternoon. A total of 48 adolescent male Wistar rats were divided into 12 rats per group. The groups were OF and MCSF, OF with start box and MCSF, EPM and MCSF, and repeated testing in MCSF. Each session was 20 minutes and there was one week between the two tests. Three rats where run in the morning and three in the afternoon. Statistical analysis did show significant difference in some parameters in the comparison between running trials in the morning or in the afternoon. Repeated testing in the MCSF yielded differences in trial two compared to trial one. No significant difference was found in the trend analysis. The results show individual differences and with larger groups results may have been more liable. In this study some differences were found between morning and afternoon groups. Lots of data have been generated and there are many opportunities to use the same data for additional analyses.

Many blood cell mechanisms in the human body are working all the time to maintain haemostasis in the blood vessels. Once a wound arises platelets are alerted via different substances to cover the wound and prevent loss of blood. Most of the times these mechanisms do stop the blood, and further heal the wound. During other circumstances the platelet-covering continues to form a thrombus, preventing the blood to flow and instead causes myocardial infarction or stroke. There are several risk factors triggering development of circulatory diseases such as obesity, lack of exercise, smoking, infection and stress. This thesis describes the interaction between the two platelet thrombin receptors PAR1 and PAR4, together with the interaction of the oral pathogen Porphyromonas gingivalis (with thrombin-like gingipains), and the cross talk with the stress hormone epinephrine and its α2A adrenergic receptor. Until now PAR1 is thought to be the most important thrombin receptor due to its high affinity for thrombin. From a phylogenetical and patophysiological point of view there must be a reason why platelets express two different thrombin receptors. Today PAR4 is considered less important, but this thesis implies that PAR4 plays an important role in platelet signaling and haemostasis. The results show that bacteria pre-stimulated platelets, followed by epinephrine gives a strong and full aggregation and calcium mobilization, in both aspirinated and non-aspirinated human platelets. The amount of bacteria does not itself, or epinephrine alone give aggregation or calcium mobilization. This mechanism is dependent on both Rgp type gingipain released from P. gingivalis, and PARs in an interaction with the α2A adrenergic receptor. Further, results reveal that PAR4 interacts and cross talks with the platelet α2A-adrenergic receptor in aspirinated platelets. Neither of the two platelet purinergic P2Y-receptors (P2Y12 and P2Y1) contribute to this action, but the purinergic P2X1 does. In aggregation studies a low dose of PAR4 activating peptide (AP), but not PAR1-AP, followed by epinephrine results in a strong aggregation and in a calcium mobilization. ATP secretion measurements did reveal that ATP was released during epinephrine stimulation, which indicate that ATP and P2X1 have a key role in this event. By blocking P2X1 both aggregation and calcium mobilization were abolished, but not by blocking P2Y12 and P2Y1. Inhibition of PI3-kinase, both epinephrine-induced calcium mobilization and aggregation were significant reduced. In non-aspirinated platelets PAR1 synergizes with the α2A adrenergic receptor and P2X1. In conclusion, this thesis suggests that PAR4 plays an intriguing and important role in platelets with inactived cyclooxygenase 1.  The results described in this thesis contribute to an increased knowledge of the platelet thrombin receptors.

Depression är den näst mest kostsamma sjukdomen för samhället efter hjärt-kärlsjukdom, främst på grund av långa sjukskrivningsperioder. Sjukdomen kan uppstå när som helst från sex månaders ålder, men prevalensen ökar med åldern. Det finns ett antal stressrelaterade faktorer som skulle kunna leda till depression, så som stor sorg, verbala eller fysiska övergrepp samt en svår barndom. Vad som orsakar sjukdomen är ännu inte helt känt, men det finns teorier att halterna av serotonin och noradrenalin är lägre hos deprimerade personer. Behandling som används är olika former av samtalsterapi, men även läkemedel så som selektiva serotoninåterupptagshämmare (SSRI). Det finns teorier som sammankopplar användandet av SSRI med självmord, framförallt hos personer ≤19 år. Syftet med detta litteraturarbete var att undersöka om SSRI preparat har någon effekt på depression hos barn och ungdomar och om de är säkra eller kan få allvarliga konsekvenser så som självmord. Sökningar i PubMed gjordes för att hitta relevanta artiklar. Fem av de åtta inkluderade studierna rapporterade olika effekter och säkerhet hos olika SSRI preparat bland barn och ungdomar, jämfört med placebo. Två andra studier undersökte förekomsten av suicidalitet till följd av läkemedlen. Den sista studien jämförde toxikologiska data från Rättsmedicinalverket med receptregistret på antidepressiva läkemedel från  Socialstyrelsen. Endast två av de fem studerade preparaten (fluoxetin och citalopram) hade en bättre effekt än placebo i hela populationen och ytterligare ett (sertralin) hade bättre effekt hos ungdomar. Det begicks inga självmord i studierna.    De studier som har granskats i detta arbete tyder på att olika SSRI preparat har olika bra effekt samt olika säkerhetsprofiler. Det sågs inget tydligt samband mellan behandlingen och självmord, men en något förhöjd risk för suicidalitet.

No description available.

Botulinumtoxin: För och emot dess användning i skönhetsbranschen Bakgrund: Botulinumtoxin (BTX) är ett kraftfullt neurotoxin som produceras av den grampositiva och anaeroba bakterien Clostridium botulinum. BTX används i medicinskt syfte vid behandling av till exempel kronisk migrän, spasticitet efter stroke, och urinläckage vid nervskada. Utöver det används BTX inom skönhetsbranschens som då injiceras i mycket låga doser under huden i muskler för att minska rynkor i olika ansiktsregioner Syfte: Användningen och lämpligheten inom skönhetsbranschen har ifrågasatts , och syftet med detta arbeta har därför varit att utvärdera och ta ställning till om BTX fortsättningsvis bör användas i kosmetiskt syfte. Metod: Litteraturstudie där vetenskapliga artiklar söktes i databasen PubMed med kombinationer av sökord, artiklar som sedan kunde användas för att besvara den aktuella frågeställningen. Resultat: Olika studier har visats att BTX är ett effektivt och säkert toxin vid skönhetsbehandling, men flera studier visade också att BTX-relaterade biverkningar uppkom efter skönhetsbehandling. Slutsats: BTX bör inte fortsätta användas för skönhetsbehandling trots positiva effekter mot rynkor i ansiktet som visades i studierna. Injektion med BTX kan bidra till att kunder i skönhetssalonger blir svårbedömda patienter vid besök hos läkare i primärvården som inte förstår att de problem patienten söker för beror på BTX-inducerade biverkningar efter skönhetsbehandling med BTX.