The effects of erythrocyte alloantigen L on the avian immune response

Experiment 1 examined the alloantigen system L effects on Rous sarcomas in three B complex genotypes. The parental stock were 50% Modified Wisconsin Line 3 and 50% inbred Line 6.15- 5. B2B5 L1L2 x B2B5 L 1L2 matings produced experimental chicks. Chicks were inoculated with 20 pock-forming units (pfu) Rous sarcoma virus (RSV) at 6 weeks-of-age. Tumors were scored six times over 10 weeks postinoculation. Tumor scores were used to assign a tumor profile index (TPI) to each chicken. Results were evaluated by ANOVA. The B complex affected the responses. Separate analyses revealed L system effects ( P \u3c 0.05) only in B5B 5 chickens. Experiment 2 examined the influence of Ea-L on antibody response to SRBC and Brucella abortus (BA). The mating protocol was the same as in Experiment 1. At 4 and 11 weeks of age the experimental birds were injected intravenously with standard concentrations of SRBC and BA. Total and ME-resistant antibody titers were determined as described. Results were analyzed by ANOVA. Ea-L had an effect on total primary antibody titer to SRBC in a B5B 5 background (p \u3c 0.004) and on total (p \u3c 0.011) and ME-resistant (p \u3c 0.017) secondary titer to SRBC in a B5 B5 genotypic background. Ea-L also affected total (p \u3c 0.004) and ME-resistant (p \u3c 0.005) secondary titer to Brucella abortus in a B5 B5 background. Experiment 3 examined the effect of Ea-L on resistance and acquired immunity to E. tenella infection. The mating protocol was the same as in Experiments 1 and 2. In the resistance and susceptibility study, chicks were weighed and inoculated with 30,000 E. tenella oocysts at 6 weeks of age. 6 days post-inoculation, the birds were weighed again and assigned a cecal lesion score. In the immunity study, the challenge procedure was preceded by inoculations of 500 oocysts per day beginning at 5 weeks. Weight gain and cecal lesion scores were evaluated by ANOVA. The B complex affected lesion score in the immunity but not the resistance and susceptibility study and did not affect weight gain in either study. The L system had no effect in either study

Media Pluralism Monitor: lessons have been learned, but concern remains for Bulgarian media

Nelly Ognyanova, Professor in Media Law at Sofia University, and Orlin Spassov, Associate Professor in Media and Communication Studies at Sofia University and Executive Director of Media Democracy Foundation, explain the results of the pilot implementation in Bulgaria of the Media Pluralism Monitor, a monitoring tool which aims to assess risks to media pluralism in the EU Member States and identify threats to such pluralism

Beneficiation of indian iron ore lumps and fines by using underbed air–pulsated BATAC jigs

After setting a target of 100 MT/yr of Steel by 2012, Indian Steelmakers and Iron Ore producers are already struggling due to the depleted grade of Iron Ore available in India. The main impurities dominating the Indian Iron Ores are SiO2 and Al2O3 which should be reduced with an economical method of Beneficiation. Jigs are the earliest type of process equipments employed in mineral separation but the newly developed under bed pulsated BATAC Jig has considerable advantages over its counterparts. There are various types of BATAC Jigs available for Lump and Fines such as Lump Ore (2 products/3 products) Jig and Fine Ore (2- product) Jig. A South African iron ore producer recently commissioned a 10 Mtpa-capacity greenfield Iron Ore beneficiation plant with 2 Lump Ore BATAC Jigs and 2 Fine Ore with a combined capacity of 1240 tph at Assmang Khumani Iron Ore Mine in Northern Cape, South Africa. The Concentrate contains a Fe grade of > 66%. A further capacity expansion project to 16 Mtpa product is currently underway using 3 more BATAC jigs. The first large scale Iron Ore Jig beneficiation plant in India was commissioned in 2006 at Noamundi in the state of Jharkhand. Tata Steel is already operating a 300 tph Fine Ore BATAC Jig Iron Ore Plant there. Patnaik Minerals also followed the pattern and started constructing 100 tph Fine Ore Jigging Plant at Joda, Jharkhand. There are different combinations possible in which unit operations can be arranged which include Jigs as the heart of the beneficiation process. These various types of flow sheets provided this beneficiation method with an advantage over other unit operations. Lessons learned and best practices regarding equipment selection and operation acquired during the last 10 years are summarized to define potential BATAC Jig applications in Iron Ore. Finally, field experiences and results are analyzed to establish the best strategy to fit specific Indian Iron Ore conditions

RELIGION AND CHURCH IN THE USTASHA IDEOLOGY

Vjersko je pitanje za ustaše bilo jednako važno kao i nacionalni problemi u Nezavisnoj Državi Hrvatskoj. Vjera je bila jedna od glavnih različitosti između Hrvata i Srba. Stoga su ustaše često poistovjećivali katolicizam s hrvatskom nacijom i nacionalizmom, te je razumljivo da je vjera bila vrlo važna u njihovoj ideologiji. Nakon raspada Jugoslavije u travnju 1941., glavni cilj nove vlade bio je stvoriti jednu ne samo samostalnu nego i katoličku državu, u kojoj bi Crkva i vjera dobile svoje pravo mjesto. Naime, po prvi put nakon 1918. katolicizam je u hrvatskim zemljama postao tolerirana, tj. institucionalna, vjera. U svojim ideološkim djelima nacionalisti su stavljali naglasak na veliku ulogu Crkve u NDH te se redom pozitivno izjašnjavali glede takvog stanja. Razlog tomu bila je tradicionalna privrženost Crkve hrvatskoj državotvornoj ideji, kao i izniman utjecaj crkvenih institucija u hrvatskom puku, uloga koju su te institucije imale tijekom hrvatske povijesti, kao i tradicionalna opozicija pravoslavlju i javno proklamirani antikomunizam. Stoga su ustaše u Crkvi vidjeli svoga prirodnog saveznika, dovoljno moćnoga da se oslone na njegovu pomoć u svojim težnjama za stvaranjem hrvatske katoličke države. Ipak, u svojim odnosima prema Katoličkoj crkvi ustaše nisu bili ekstremni te je njihov pokret u osnovi ostao građanski i nacionalistički. Iako su ustaše promovirali vlastitu opredijeljenost katoličkoj vjeri, to je ipak u većoj mjeri bilo samo izjašnjavanje o njihovu hrvatskom kulturnom identitetu i glavni razlikovni čimbenik prema pravoslavlju, a ne istinsko vjersko izjašnjavanje. Naprotiv, ustaše uglavnom nisu bili zainteresirani za pitanja vjerskih dogmi i teoloških raspri. Propagirali su katolicizam ne zbog toga što su ga smatrali »jedinom pravom i iskonskom vjerom« na svijetu, nego jer im je katolicizam mogao pomoći u vlastitom nacionalnom samoodređenju u etnički i vjerski izmiješanim hrvatskim zemljama. Upravo stoga nove hrvatske vlasti nisu pokrenule »križarski rat« protiv islama, nego su ga, naprotiv, tolerirale jer se vjerovalo da savez s muslimanima Bosne i Hercegovine može biti od taktičke važnosti. S druge strane, pravoslavlje se našlo na udaru vlasti NDH, upravo zbog svoje nacionalne pozadine – tako da je glavni udar novih vlasti bio usmjeren protiv srbizma. Budući je u NDH živjelo blizu dva milijuna Srba, pitanje pravoslavlja i Pravoslavne crkve imalo je vrlo važnu ulogu u ustaškome ideološkom sustavu. Hrvatski su se nacionalisti trudili braniti svoj »tvrdi« stav i beskompromisnost prema Pravoslavnoj crkvi u Hrvatskoj, ističući da je tu riječ o »nacionalnoj« Crkvi (instituciji) koja je igrala važnu ulogu u formiranju tadašnjega srpskog nacionalizma. A budući da nisu mogli tolerirati buntovne aktivnosti srpske manjine, niti osporiti utjecaj njihovih crkvenih institucija, ustaše su odlučili jednostavno ih otrgnuti od službenoga srpskog utjecaja na način da dio srpske manjine prevede na katoličanstvo. Tako su Srbe koji su živjeli u Hrvatskoj jednostavno proglasili »pravoslavnim Hrvatima« koje je velikosrpska propaganda stoljećima srbizirala. Tako posložene činjenice uvjetovale su da su ustaše na prisilne konverzije Srba na katoličanstvo gledali kao na nacionalno, a ne vjersko pitanje u NDH. U osnovi vlasti NDH svoje su napade usmjeravali uglavnom protiv Srpske pravoslavne crkve i njezinih aktivnosti na teritoriju NDH. Stoga je razumljivo da su ustaške vlasti, glede pitanja pravoslavlja, mogle tolerirati postojanje samo Hrvatske pravoslavne Crkve na teritoriju NDH, te je HPC iza 1942. postala jedina crkvena organizacija za vjernike pravoslavne vjere u NDH, bez obzira na njihovu etničku pripadnost. No ustaše nisu imali negativan stav samo prema pravoslavlju nego i prema unijatima (tj. grkokatolicima), kao i prema starokatolicima na teritoriju NDH. Osim toga i židovstvo je bilo službeno stigmatizirano u onodobnoj ustaškoj ideologiji. Židovi i Romi bili su optuženi za ubojstvo Isusa Krista i stoga okarakterizirani kao neprijatelji Katoličke crkve i širitelji komunističke ateističke ideologije u čitavom svijetu. No hrvatski nacionalisti zadržali su pozitivan stav prema islamu. Kako bi se što učinkovitije suprotstavili pravoslavlju u Bosni i Hercegovini, ustaše su promovirali ideju da su Hrvati i muslimani pripadnici jedne nacije – i to hrvatske. Nadalje, tvrdili su da je tu riječ o »braći po krvi i jeziku«. Iako su među muslimanima postojale dvojbe glede ustaškoga proklamiranog katolicizma i odnosa prema islamu, ustaše su javno zagovarali vjersku i građansku jednakost katolika i muslimana u NDH. Ipak valja napomenuti da te simpatije prema islamu nisu bile rezultat iskrenih osjećaja Hrvata katolika prema muslimanima, nego je tu bila riječ o specifi čnim političkim interesima NDH u Bosni i Hercegovini. Stoga je za razumijevanje vjerske politike novih vlasti u Hrvatskoj u razdoblju od 1941. do 1945. nužno poznavati njihove nacionalističke i političke ciljeve s kojima je vjerska politika bila u uskoj vezi.The article deals with the problem regarding the position of religion and Church in the Ustasha ideological system, which is insufficiently explored in contemporary historiography. Between Catholicism and the Croatian nationalism existed a strong historical connection. It was explicitly expressed in the extreme forms of the Croatian nationalism, such as in the Ustashas movement. Nationalism and Catholicism were not only two basic, but also closely tied ideas incorporated in their ideological system. This topic is partially elaborated in historiography, but still there is no a single monograph dedicated to it. Particularly disputable problems in historical literature are also those about the role of the Catholic Church in the Independent State of Croatia (ISC) and its relationship towards Ustashas’ nationalism, and those about relationship between Vatican and the Croatian authorities in the period between 1941 and 1945

Selective progesterone receptor modulator (SPRM) action on the human endometrium

INTRODUCTION: Selective Progesterone Receptor Modulators (SPRMs), of which Ulipristal Acetate (UPA) is a class member, have shown utility for gynaecological therapies including the symptom of heavy menstrual bleeding (HMB). UPA exhibits stronger antagonistic than agonistic action on the progesterone receptor (PR). During the normal menstrual cycle oestradiol stimulates endometrial proliferation and progesterone stimulates endometrial cell differentiation and opposes oestradiol-induced proliferation. SPRMs such as UPA, with antagonistic action to the PR, create the potential for unopposed endometrial proliferation alongside an endocrine environment which continues to expose the endometrium to circulating oestradiol. Contrary to expectation, SPRM UPA decreases endometrial cell proliferation as assessed by cell proliferation marker Ki-67. This observation presents an unexplained paradox that requires deeper understanding of the mechanism of action of SPRM UPA within the endometrium. HYPOTHESIS AND OBJECTIVES: The anti-proliferative effect of SPRMs is dependent on cell-cell communication between the two main endometrial cell types - stromal and epithelial cells. To test this hypothesis, four objectives were set: 1. Identify the SPRM-associated changes related to cell cycle in whole tissue endometrial transcriptome before, during, and after SPRM administration. 2. Determine the effect of SPRM on selected genes and proteins involved in cell cycle and progesterone-dependent downstream signalling in isolated human endometrial stromal cells 3. Establish endometrial epithelial organoid cultures and determine the effect of SPRM on selected genes and proteins involved in cell cycle and progesterone-dependent downstream signalling. 4. Develop a co-culture model with human endometrial stromal cells and epithelial organoids grown in a novel 3-dimensional (3D) gel microenvironment to determine the effect of SPRM upon cell cycle and progesterone-dependent downstream signalling. METHODS AND RESULTS: To address Objective 1, RNA-Sequencing and pathway analysis were employed. Over 2000 differentially expressed genes (DEG) were identified when comparing pre-UPA-treated samples to UPA-treated samples of proliferative phase endometrium tissue biopsies. DEG were involved in suppression of canonical pathways of the oestrogen-mediated S-phase entry and cell-cycle progression. Genes encoding for cell cycle regulators cyclins A, B and E, E2F2, and CDC25A were also significantly downregulated when validation experiments were conducted using standard RT-qPCR. Notably, there were no enriched pathways between pre-treatment and post-UPA treated endometrium both in proliferative and secretory phase samples, indicating the altered gene expression was reversible. To address Objectives 2 and 3, a protocol for isolation of endometrial stromal and epithelial cells was established and optimised. Endometrial epithelial cells were grown in Matrigel to form epithelial organoids. Using standard immunostaining and RT-qPCR protocols, the progesterone receptor was identified in both cell cultures and evidence of progesterone action was established by the detection of elevated mRNA levels of the progesterone-dependent genes Prolactin and IGFBP1. Despite this, no SPRM UPA-dependent (progesterone antagonist) effect was evident in either stromal cell or epithelial organoid cultures for genes and proteins of the cell cycle (Cyclin A1, Cyclin E2, E2F2, CDC25A), progesterone, androgen, glucocorticoid, and oestrogen receptors, nor for progesterone-dependent HOXA10 and FOXO1. Reverse Phase Protein Array (RPPA) using 120 pre-selected antibodies did not reveal UPA-dependent changes at the post-translational level in either stromal cell or epithelial organoid cultures. To address Objective 4, isolated endometrial stromal cells and epithelial organoids were incorporated into either Matrigel or functionalised cross-linked polymer of polyethylene glycol (PEG) gel. RT-qPCR did not reveal SPRM-associated changes in either Matrigel or PEG co-cultures. In contrast, RPPA results showed dose-dependent differential abundance of protein and phosphorylated epitopes in response to SPRM treatment in PEG co-cultures only. The SPRM UPA modulated proteins that were identified represent known mediators of cell cycle, MAPK/AKT, and chromatin regulation pathways. SUMMARY: In these studies, endometrial epithelial organoid, and stromal-epithelial co-culture models have been established. Through RNA sequencing studies in whole endometrial tissue exposed to the SPRM, UPA, novel targets of SPRM mechanism of action were identified (Cyclin A1, Cyclin E2, E2F2, and CDC25A). These targets were tested in isolated monoculture stromal cell, epithelial organoid, and stromal-epithelial co-culture models. Results from isolated stromal and organoid cultures did not replicate observations in whole endometrial biopsies. Only co-cultures of isolated endometrial stromal cells with epithelial organoids in PEG gel matrix demonstrated comparable results to observations seen in intact endometrium, confirming the hypothesis that the anti-proliferative effect of SPRMs is dependent on cell-cell communication. Furthermore, an appropriate choice of 3D scaffold for in vitro experiments is critical for determining the functional response of primary cells to exogenous treatments

Investigating the relationship between SORCS2 and SORLA and DNA double-stranded break formation, as a possible mechanism in neuropsychiatric disorders

SORCS2 and SORLA are members of the Vps10p-domain receptor family, which comprises five multifunctional neuronal receptors. All family members are involved in intracellular sorting and trafficking of various neurotrophic factors and their precursor forms, as well transmembrane receptors and synaptic proteins. This gene family has been implicated in a broad range of cellular processes, including neuronal health, differentiation and synaptic plasticity. Importantly for this thesis, SORLA and SORCS2 have been shown to play a role (SORLA in vivo and in vitro; SORCS2 in vitro) in the processing of the amyloid precursor protein (APP) and thus amyloid β (Aβ) production. Genetic and functional studies provide further evidence for the involvement of both receptors, alongside other family members, in cognition and various brain disorders. DNA damage and compromised DNA repair have been implicated in brain aging, psychiatric and neurodegenerative disorders. Recently, a specific type of DNA damage- DNA double- stranded break (DSB) formation, has been implicated in both Alzheimer’s disease (AD) and physiological brain activity, i.e. learning and memory processes. In mice, exploration of a novel environment led to a transient increase in DSB formation in the dentate gyrus (an area of the brain important for learning and memory). The breaks were repaired within 24 hours. Subsequently, it was shown that these breaks are generated by Topoisomerase II β (TopoIIβ) and are required for the expression of early-response genes. In mice, pathological Aβ levels were associated with increased neuronal DSB formation and repair deficits. The effect of Aβ on DSB formation and repair was attributed to aberrant network activity and dysregulated NMDA receptor signalling. Given the link between sortilins and neuropsychiatric illness, as well as their role in APP processing and, recently, NMDA receptor trafficking (SorCS2), my project aimed to address the relationship between the two genes and DSB formation. I examined levels of DNA DSBs in WT and Sorcs2-/- mice before and after exploration of a novel environment. Following exploratory activity, WT mice showed increased numbers of DSBs, which were repaired within 24 hours. Meanwhile, I detected elevated levels of DSBs at baseline in the dentate gyrus of Sorcs2-/- mice. Moreover, compared to the corresponding WT mice, the Sorcs2-/- mice exhibited an altered response to the novel environment, as they failed to acquire new breaks. To explore possible mechanisms that could explain this phenotype, I knocked out SORCS2 in the human dopaminergic neuronal cell line LUHMES using CRISPR/Cas9 genome editing. I did not observe any difference in the extent of DSB formation in untreated SORCS2 knockout (KO) clones, compared to WT controls grown simultaneously. However, SORCS2 KO neurons showed elevated levels of DSBs following treatment with etoposide- a compound that prevents the re-ligation of TopoIIβ-induced DSBs. In addition, knocking out SORCS2 was also associated with reduced neuronal viability. I next explored possible mechanisms underlying the observed increase in etoposide induced DSBs in the SORCS2 KO clones. In line with previous work, treatment with NMDA led to a significant increase in the number of DSBs in WT LUHMES neurons. I did not, however, observe any difference in the extent of DSB formation in NMDAtreated SORCS2 KO clones compared to WT controls. This experiment was performed on a small number of WT and KO clones and the results may thus reflect lack of power to detect a difference. However, if this lack of difference was confirmed in subsequent experiments, this would suggest that the increase in etoposide-induced DSBs in the SORCS2 KO clones is unlikely to be due to dysregulated NMDA signalling. Meanwhile, I was unable to measure Aβ42 levels reliably in LUHMES. Thus, the possible role of Aβ in the increased DSB formation observed in etoposide-treated SORCS2 KO clones cannot be excluded. Surprisingly, mouse primary hippocampal neurons derived from Sorcs2-/- pups did not show any difference in DSB levels with or without treatment with etoposide. However, this experiment was performed on a limited number of hippocampal cultures and thus the results obtained may not reflect a true lack of difference. The possible discrepancy between the results in the etoposide-treated primary hippocampal and LUHMES dopaminergic neurons, but more importantly the role of SorCS2 in dopaminergic signalling, led me to investigate whether knocking out SORCS2 alters extracellular dopamine levels, as a potential mechanism underlying the increased DSB formation. ELISA analysis showed no difference in dopamine release between SORCS2 KO and WT LUHMES neurons. Due to technical difficulties and time constraints, I was unable to assess the levels of DSB formation in Sorl1-/- mice or primary hippocampal neurons derived from them. Obtaining SORL1 KO LUHMES cell lines using CRISPR/Cas9 genome editing also proved to be challenging. However, I generated mutant cell lines carrying the rare SORL1 missense mutation, G508S, which is associated with early onset AD. Despite being located at a splice site, introducing the G508S mutation in LUHMES did not affect splicing. However, it led to reduced SORL1 mRNA, but not protein, levels. In conclusion, the work completed in this project constitutes the first evidence for a potential role of the sortilins in DSB formation. Future work investigating this link further might provide us with valuable knowledge on the cellular mechanisms underlying neurodegenerative and psychiatric conditions