Magister Scientiae (Medical Bioscience) - MSc(MBS) / Exposure to endocrine-disrupting chemicals as bisphenol A (BPA) during gestation and early postnatal life is known to disrupt normal developmental processes and alter the body’s endocrine system leading to deleterious effects in the developing central nervous system (CNS). BPA is an industrial synthetic chemical commonly used in the production of a range of polymers and consumer products, despite concerns about its safety. There is therefore the need to protect the developing CNS from potential damage through the administration of neuroprotective agents. Most medicinal plants are reported to possess significant protective potential against tissue damage through different mechanisms that prevent cell death, oxidative stress, inflammation, immunodeficiency, etc. In this study, the protective effects of fermented rooibos (Aspalathus linearis) tea against the deleterious effects of BPA were investigated. Rooibos is a herbal beverage indigenous to South Africa with widely acclaimed health benefits often linked to the bioactivity of its polyphenolic compounds, especially aspalathin. The anti-allergic, cardiovascular, antioxidant and neuroprotective effects of this herb have been previously reported hence, the present study aims to investigate if regular consumption of rooibos tea during pregnancy and lactation could protect the developing brain from the deleterious effects of BPA in a Wistar rat model. A total of 40 three-month old adult female pregnant dams, with an average weight of 250g, were divided into four groups (n=10). Group 1 control rats received 9% normal saline ad libitum; group 2 rats received 400μg/kg/day BPA only; group 3 rats received 20% fermented rooibos tea as well as 400μg/kg/day BPA, while group 4 rats received ad libitum 20% fermented rooibos tea only. Offspring rats were housed in the same cages as the dams and only separated after weaning on postnatal day (PND) 21. Neurobehavioural assessment using the open field test was done on postnatal day (PND) 42 after which the final body masses were taken before the rats were decapitated under deep anaesthesia, and the desired CNS parts carefully dissected out and processed for histological, biochemical and immunohistochemical studies. The results obtained showed that there was significant impairment of neurobehavioural activity, decreased cerebral and cerebellar antioxidant enzyme activity, reduced hippocampal CA1 length, significant loss of cerebellar Purkinje cells and significant astrocyte activation demonstrated by increased glial fibrillary acidic protein (GFAP) activity in experimental rats exposed to BPA only. However, co-administration of rooibos tea significantly attenuated the BPA-induced distortions. Taken together, these findings suggest that rooibos could be a potent neuroprotective agent against BPA-induced structural, functional and biochemical alterations in the developing CNS.

Bisphenol A (BPA) is an alkylphenol compound used in plastic manufacturing, which enters the aquatic environment through wastewater treatment plants and landfill leachates. Exposure of fish to BPA results in developmental defects, decreased hatching, and increased occurrence of the egg yolk precursor protein vitellogenin in male fish. BPA is an estrogenic like compound (xenoestrogen) that can bind and activate the nuclear estrogen receptors, ERα and ERβ, causing changes in gene transcription (genomic mechanism). However, the ability of BPA to activate rapid estrogen signaling and to disrupt nongenomic physiological responses to estrogens is not known. One important nongenomic estrogen action in fish is to maintain meiotic arrest of oocytes and to prevent precocious oocyte maturation through activation of the transmembrane G-protein coupled estrogen receptor (Gper). Binding of estrogens to Gper on zebrafish oocytes results in rapid activation of epidermal growth factor receptor (Egfr) signaling and subsequent Mapk3/1 activation. We show here that BPA and three structurally-related chemicals, tetrachlorobisphenol A, tetrabromobisphenol A and nonylphenol, can mimic estrogen by activating this mechanism of meiotic arrest through Gper in zebrafish (Danio rerio). BPA bound to zebrafish Gper and inhibited spontaneous oocyte maturation (OM) of denuded oocytes. Treatment of oocytes with Actinomycin D did not block the effects of BPA, suggesting that this inhibition of OM is through a nongenomic mechanism. Incubation of oocytes with a selective GPER antibody and the specific GPER antagonist G-15 blocked the effects of BPA on OM, further suggesting that BPA inhibition of OM is through its interaction with the receptor. Various inhibitors of the EGFR pathway were utilized to determine if the inhibition of OM by BPA is mediated through this mechanism. BPA activation of the Egfr pathway resulted in Mapk3/1 (also known as Erk 1/2) phosphorylation. The results show that BPA disrupts oocyte maturation through a novel mechanism involving activation of a Gper/Egfr/Mapk3/1 pathway with potential adverse impacts on reproductive success. / text

The luteinizing hormone / choriogonadotropin receptor (LH/CG-R) is a member of theG protein-coupled receptor family. The LH/CG-R has seven transmembrane helices, threeexoloops, three cytoloops, a C-terminal tail, and an extensive N-terminal exodomain. Theexodomain is capable of binding hormone with high affinity without hormone action. Previousstudies have shown that the amino-terminal region of the LH/CG receptor contacts both subunitsof human chorionic gonadotropin (hCG). In particular, three residues (Leu20, Cys22, and Gly24)were found to be crucial for hormone binding. In this thesis work, benzoylphenylalanine (Bpa),a photoactivatable reagent, was used to continue investigating the interactions of the N-terminalregion of the LH/CG-R with hCG. Bpa has been directly incorporated at a defined position intopeptides representing amino acids 17-36 of the LH/CG-R. These peptides were radiolabeledwith 125I and used in photoaffinity labeling studies to identify and characterize the contact site(s)between the N-terminal region of the LH/CG-R and hCG. Results suggest that Cys22 is theprimary contact residue in this region. Peptide and hormone concentration dependent as well asUV duration dependent photoaffinity labeling experiments confirm that the photolabeling ofhCG by hLHR17-36(C22Bpa) is specific. Competition of labeling studies indicate that the hLHR17-36(C22Bpa) peptide is a good mimic of the wild type N-terminal portion of the receptor. In-geldigestions of photolabeled hCG ?? and photolabeled hCG ?? with CNBr indicate that the Nterminalregions of both hCG ?? and hCG ?? were photoaffinity labeled by hLHR17-36(C22Bpa).Based on the fact that the N-terminal regions of each subunit are located on the convex side ofthe heterodimer, these results provide evidence that the N-terminal portion of the receptor wrapsaround the back of hCG, contacting the convex face of the hormone.

Phenolic compounds exist in abundance in the world around us due to natural and synthetic biological processes. Phenols produced from chemical, petrol, and pharmaceutical processes can be harmful, stress-inducing exotoxins. Bisphenol-A (BPA) and butylated hydroxyanisole (BHA) are phenol-based compounds found in polycarbonate plastics and food preservatives, therefore presenting a particularly high degree of human exposure. Exposure to these compounds has been associated with numerous diseases, yet their mechanisms of action are not well understood. A quantitative evaluation of growth of yeast deletion strains was used to identify the genes that modulate response to phenol stress induced by BPA and BHA in the model organism, yeast Saccharomyces cerevisiae. Twelve genes were identified as essential for growth in the presence of both BPA and BHA. pep5 and vph2 are integral to vacuolar ATPase function, erg2 and erg24 are involved in the erg pathway, and ref2, rsc1, bdf1, tps2, hpr1, and snf12 are important in chromatin remodeling and transcription processes. Additional genes required for growth and growth-inhibiting genes were identified that were specific to BPA or BHA. Many of the genes identified in S. cerevisiae have orthologs in humans that may modulate exposure to toxic phenols in a similar manner and could provide insight into the pathways involved in phenol exposure related disease.

This report researches and analyses the sustainability perspectives of paper receipts, focusing on the economical, environmental and social aspects using a qualitative research method and by simultaneously drawing inspiration from the Sustainability Impact Assessment (SIA) model. The result section is conducted by using a literature review where a combination of previously used sources within my masters program is analysed and by reviewing articles and statistics in the search engine at the Uppsala University e-library database. The overall research purpose, to assess the sustainability of consumer receipts, is achieved by using the concept of sustainable development as the theoretical framework. This report suggests that there are some sustainability concerns with the current consumption of the traditional paper receipt, mainly in the context of environmental and social considerations. Three different alternatives to the traditional paper receipt are furthermore discussed as examples of potential routes forward. Two of the alternatives, electronic receipts and introducing a charge on paper receipts, are arguably the two most sustainable alternatives to regular paper receipts. The third alternative cannot be perceived as equally sustainable since “BPA/Bisphenol-free” receipts contain other chemicals equivalently hazardous to humans and the environment.

Forensic science is the field of study that studies crimes and crime scenes. One of the major sub-areas of forensic science is crime scene investigations (CSI). Bloodstain pattern analysis (BPA) is a part of CSI and refers to the study of bloodstain patterns. The purpose of this project is to investigate the needs of those involved in the judicial chain when using a laser scanner to reconstruct a crime scene and how to represent any uncertainties. An additional purpose is to look into what expectations bloodstain pattern analysts have on their work situation. Interviews were held with nine persons involved in the judicial chain and an online questionnaire was distributed to the bloodstain pattern analysts across Sweden. To analyze the interviews a thematic analysis was used which led to three themes being identified (benefit, desires, obstacles) with eleven sub-themes. For the questionnaire two types of data were presented, numeric and written. The numeric result displayed results such as how confident they felt doing work or if the amount of cases was too much or too little. The written result displayed results showing that BPA is cumbersome, not because it is hard to use but rather because each case is unique, and many factors have to be considered. The conclusion of this study is that needs can be met using the framework that combined uncertainties and visualization, and the questionnaire showed that the bloodstain pattern analyst are a group of people who seek knowledge and welcome new technology.

ABSTRACT Background: Developmental exposure to endocrine disrupting chemical (EDC), such as Bisphenol A (BPA) has been shown to affect the health during early stages in life. Aim: To assess whether developmental low-dose exposure to BPA alters adipocyte distribution and blood lipid levels in Fischer 344 (F344) rats. Method: Pregnant F344 rats were exposed to BPA via their drinking water from gestational day 3.5 throughout lactation. Doses given were one below the current European Food Safety Authority´s preliminary tolerable daily intake (TDI) of 4 µg BPA/kg bw/day, 0.5 [lower dose, Ld] and one equal to the previous TDI; 50 [higher dose, Hd] µg BPA/kg bw/day. Half the offspring was sacrificed at 5, and half at 52 weeks of age. Body weight was registered, and plasma lipid levels were analyzed. Inguinal white adipose tissue (iWAT) was weighed, Oil red O-stained and analyzed histologically. Results: Five-week-old Ld males and females Hd exhibited significantly higher triglyceride levels (31%, p<0.01; 41%,p<0.05, respectively) compared to control. Total cholesterol was borderline significantly increased (9%, p=0.0554) in 5-week-old Ld males, compared with control. Adipocyte cell density in the 5-week-old offspring was significantly increased; Ld female rats had increased cell density compared to control and Hd (22% and 23%,p<0.05, respectively), whereas the cell density of Hd males increased compared to Ld (29%, p<0.05). In the 52-week-old offspring blood lipid levels and iWAT cell density were not significantly affected. Conclusions: Results of the present study supports that developmental low-dose BPA exposure contributes to elevated triglyceride levels in 5-week-old animals.

The luteinizing hormone receptor (LHR) belongs to the G protein-coupled receptorfamily. It consists of two distinct domains; the N-terminal extracellular exodomain and themembrane associated endodomain which includes 7 transmembrane domains, 3 exoloops, 3cytoloops and a C-terminal tail. Sequence alignment and computer modeling suggest thepresence of Leu Rich Repeat (LRR) motifs in the exodomain. Although their structuralsimilarity is high, each LRR is not equally important for hormone binding. Ala-scanning andtruncation studies performed in our laboratory suggest that LRR2 and LRR4 appear to be themost crucial. The Ala-scanning data suggest that Leu103 and Ile105 in LRR4 are important forhormone binding. However, it is not clear whether these two residues make direct contact withhuman chorionic gonadotropin (hCG) or if they are necessary for the overall structural integrityof LRR4. In this work, the LHR peptide mimics of LRR4 were used for photoaffinity labeling todetermine whether Leu103 and Ile105 directly interact with hormone. Furthermore, LRR4peptides containing the photoactivable benzoylphenylalanine (Bpa) were used to determinewhether the LRR structure really exists in the LHR exodomain, whether LRR 4 interact withhCG, and which residues of LRR4 interact with hCG. Bpa was directly incorporated intodifferent positions of the LRR4 peptide sequence to examine the labeling ability of individualamino acids. The results suggest that LRR4, in particular the sequence of Lys101-Cys106,makes direct contact with hCG. However Leu103 and Ile105 do not interact with hCG but mayform the hydrophobic core of the LRR4 loop, which appears to be crucial for the LRR structure.Existing data suggest that glycoprotein hormones initially bind the exodomain. Thehormone/exodomain complex undergoes conformational adjustments and stimulates theendodomain of the receptor to generate hormone signals. The exoloops modulate hormonebinding and signaling; however, little is known about whether the hormone/exodomain complexcontacts the endodomain. To address this issue, we investigated whether the exoloops interactwith the hormone. First, we examined exoloop 3 that connects transmembrane domains 6 and 7which are important for signal generation. We present the first physical evidence that LHRexoloop 3 interacts with hCG.

Apesar da monofilia de Pentatomidae ter sido amplamente demonstrada, as relações infrafamiliares precisam ser esclarecidas. Por exemplo, um dos maiores táxons da família, a subfamília Pentatominae, ainda não é reconhecido como um grupo monofilético. Para resolver estes problemas é necessário, além de conhecer a diversidade do grupo, realizar estudos cladísticos nos níveis genéricos e cladísticos. Na presente Tese são descritas cinco novas espécies do gênero Chloropepla (Pentatominae): C. paveli, C. stysi, Chloropepla sp. nov. 1, Chloropepla sp. nov. 2 e Chloropepla sp. nov. 3. As espécies são caracterizadas, principalmente, por atributos da genitália de machos e fêmeas. Uma chave ampliada para a identificação das espécies do gênero é fornecida. A distribuição setentrional do grupo é expandida da Venezuela para a Costa Rica. Uma sinopse de Chloropepla também é apresentada, com uma descrição ampliada do gênero e diagnose das espécies, ambas baseadas em análise de parcimônia das 12 espécies conhecidas. Esta análise cladística confirmou a monofilia do grupo, baseada no peritrema ostiolar longo e evanescente, hypandrium amplo com projeções dorsais atingindo o X segmento, parâmeros cilíndricos, dirigidos dorsalmente, conjuntiva membranosa reduzida, quase que inteiramente obscurecida pela phallotheca. A relação filogenética resultante entre as espécies de Chloropepla foi submetida à BPA com as subregiões e províncias da região Neotropical como terminais. Esta análise demonstrou uma relação próxima entre as áreas amazônicas e indicou uma natureza híbrida da sub-região chaquenha. Finalmente, a relação entre oito gêneros de Pentatominae é investigada: Arvelius, Chlorocoris, Chloropepla, Eludocoris, Fecelia, Loxa, Mayrinia e Rhyncholepta. Duas análises de parcimônia foram realizadas: uma baseada somente em caracteres morfológicos e outra baseada em morfologia e em sequências ribossomais (evidencia total). Um fragmento de rDNA mitocondrial 16S e dois de rDNA nuclear 28S foram sequenciados e analisados utilindo-se o método de optimização direta. As análises apresentaram diferentes relações entre os gêneros. Contudo, algumas relações se mantêm em ambos os cladogramas: Loxa + Mayrinia + Chlorocoris (Monochrocerus), Arvelius + E. humeralis + R. humeralis and Chlorochoris (Chlorocoris) + Fecelia + Chlorochoris (Arawacoris). Os resultados obtidos enfatizam a necessidade de estudos futures sobre o uso de dados moleculares em análises genéricas e específicas em Pentatominae. Além disto, as homologias também precisam ser melhor investigas e testadas, com base em estudos cladísticos, dentro de Pentatomidae, Pentatominae e tribos. / The monophyly of Pentatomidae was already highly confirmed. However, the infra-family relationships still need to be clarified. For example, one of the largest taxa of the family, the sub-family Pentatominae is not recognized as a monophyletic group. To solve these problems is necessary to know the diversity of the group as well as to perform cladistics studies at the level of genera and species. In this thesis, five new species of the genus Chloropepla (Pentatominae) are described: C. paveli, C. stysi, Chloropepla sp. nov. 1, Chloropepla sp. nov. 2 and Chloropepla sp. nov. 3. The species are mainly characterized by features of male and female genitalia. An extended key to identification of the species of the genus is provided. The northern distribution of the group is expanded from Venezuela to Costa Rica. A synopsis of Chloropepla is also presented, with an extended description of the genus and diagnosis for the species, both based in a parsimonious analysis of the 12 know species. This cladistics analysis confirmed the monophyly of the group, based on ostiolar ruga long and evanescent, wide hypandrium, with dorsal projections flanking the segment X, parameres cylindrical, dorsally directed, membranous conjunctiva reduced, almost entirely obscured by the phallotheca. The resultant phylogenetic relationship of Chloropepla species were submitted to a BPA with the sub-regions and provinces of Neotropical region as terminals. This analysis showed a near relation among the Amazonian areas and indicated a hybrid nature of the Chacoan subregion. Finally, the relationship of eight Pentatominae genera is investigated: Arvelius, Chlorocoris, Chloropepla, Eludocoris, Fecelia, Loxa, Mayrinia and Rhyncholepta. Two parsimony analyses were performed: one based solely on morphological characters and other based on morphological plus ribosomal DNA sequences (total evidence). One fragment of 16S mitochondrial and two of 28S nuclear rDNA were sequenced and analyzed using the direct optimization method. The results of both analysis differed in the relationships of the genera. However, some relations are recovered in both cladograms: Loxa + Mayrinia + Chlorocoris (Monochrocerus), Arvelius + E. humeralis + R. humeralis and Chlorochoris (Chlorocoris) + Fecelia + Chlorochoris (Arawacoris). The results here obtained emphasize the necessity of further studies on the use of molecular data in analyses on genera and species levels in Pentatominae. Besides, the homologies also need to be better investigated and tested within Pentatomidae, Pentatominae and tribes levels, based on cladistics studies.

Apesar da monofilia de Pentatomidae ter sido amplamente demonstrada, as relações infrafamiliares precisam ser esclarecidas. Por exemplo, um dos maiores táxons da família, a subfamília Pentatominae, ainda não é reconhecido como um grupo monofilético. Para resolver estes problemas é necessário, além de conhecer a diversidade do grupo, realizar estudos cladísticos nos níveis genéricos e cladísticos. Na presente Tese são descritas cinco novas espécies do gênero Chloropepla (Pentatominae): C. paveli, C. stysi, Chloropepla sp. nov. 1, Chloropepla sp. nov. 2 e Chloropepla sp. nov. 3. As espécies são caracterizadas, principalmente, por atributos da genitália de machos e fêmeas. Uma chave ampliada para a identificação das espécies do gênero é fornecida. A distribuição setentrional do grupo é expandida da Venezuela para a Costa Rica. Uma sinopse de Chloropepla também é apresentada, com uma descrição ampliada do gênero e diagnose das espécies, ambas baseadas em análise de parcimônia das 12 espécies conhecidas. Esta análise cladística confirmou a monofilia do grupo, baseada no peritrema ostiolar longo e evanescente, hypandrium amplo com projeções dorsais atingindo o X segmento, parâmeros cilíndricos, dirigidos dorsalmente, conjuntiva membranosa reduzida, quase que inteiramente obscurecida pela phallotheca. A relação filogenética resultante entre as espécies de Chloropepla foi submetida à BPA com as subregiões e províncias da região Neotropical como terminais. Esta análise demonstrou uma relação próxima entre as áreas amazônicas e indicou uma natureza híbrida da sub-região chaquenha. Finalmente, a relação entre oito gêneros de Pentatominae é investigada: Arvelius, Chlorocoris, Chloropepla, Eludocoris, Fecelia, Loxa, Mayrinia e Rhyncholepta. Duas análises de parcimônia foram realizadas: uma baseada somente em caracteres morfológicos e outra baseada em morfologia e em sequências ribossomais (evidencia total). Um fragmento de rDNA mitocondrial 16S e dois de rDNA nuclear 28S foram sequenciados e analisados utilindo-se o método de optimização direta. As análises apresentaram diferentes relações entre os gêneros. Contudo, algumas relações se mantêm em ambos os cladogramas: Loxa + Mayrinia + Chlorocoris (Monochrocerus), Arvelius + E. humeralis + R. humeralis and Chlorochoris (Chlorocoris) + Fecelia + Chlorochoris (Arawacoris). Os resultados obtidos enfatizam a necessidade de estudos futures sobre o uso de dados moleculares em análises genéricas e específicas em Pentatominae. Além disto, as homologias também precisam ser melhor investigas e testadas, com base em estudos cladísticos, dentro de Pentatomidae, Pentatominae e tribos. / The monophyly of Pentatomidae was already highly confirmed. However, the infra-family relationships still need to be clarified. For example, one of the largest taxa of the family, the sub-family Pentatominae is not recognized as a monophyletic group. To solve these problems is necessary to know the diversity of the group as well as to perform cladistics studies at the level of genera and species. In this thesis, five new species of the genus Chloropepla (Pentatominae) are described: C. paveli, C. stysi, Chloropepla sp. nov. 1, Chloropepla sp. nov. 2 and Chloropepla sp. nov. 3. The species are mainly characterized by features of male and female genitalia. An extended key to identification of the species of the genus is provided. The northern distribution of the group is expanded from Venezuela to Costa Rica. A synopsis of Chloropepla is also presented, with an extended description of the genus and diagnosis for the species, both based in a parsimonious analysis of the 12 know species. This cladistics analysis confirmed the monophyly of the group, based on ostiolar ruga long and evanescent, wide hypandrium, with dorsal projections flanking the segment X, parameres cylindrical, dorsally directed, membranous conjunctiva reduced, almost entirely obscured by the phallotheca. The resultant phylogenetic relationship of Chloropepla species were submitted to a BPA with the sub-regions and provinces of Neotropical region as terminals. This analysis showed a near relation among the Amazonian areas and indicated a hybrid nature of the Chacoan subregion. Finally, the relationship of eight Pentatominae genera is investigated: Arvelius, Chlorocoris, Chloropepla, Eludocoris, Fecelia, Loxa, Mayrinia and Rhyncholepta. Two parsimony analyses were performed: one based solely on morphological characters and other based on morphological plus ribosomal DNA sequences (total evidence). One fragment of 16S mitochondrial and two of 28S nuclear rDNA were sequenced and analyzed using the direct optimization method. The results of both analysis differed in the relationships of the genera. However, some relations are recovered in both cladograms: Loxa + Mayrinia + Chlorocoris (Monochrocerus), Arvelius + E. humeralis + R. humeralis and Chlorochoris (Chlorocoris) + Fecelia + Chlorochoris (Arawacoris). The results here obtained emphasize the necessity of further studies on the use of molecular data in analyses on genera and species levels in Pentatominae. Besides, the homologies also need to be better investigated and tested within Pentatomidae, Pentatominae and tribes levels, based on cladistics studies.