Positional identification and functional analysis of genes regulating autoimmune arthritis

The major histocompatibility complex (MHC) is the most gene-dense and polymorphic region in the human genome with strong associations to many autoimmune disorders, including rheumatoid arthritis (RA). However, even the genetic association between MHC and RA was known more than 40 years ago, we still have not fully explained the functional roles of the MHC genes and identified the underlying specific polymorphisms. This thesis describes some of our research aimed for a better understanding of this topic, which can largely be divided into three parts as follows. First, we made use of a panel of MHC class II (MHC-II) congenic strains to evaluate the functional roles of MHC-II polymorphisms in arthritis. By performing an extensive genetic and functional analysis, we showed that MHC-II RT1-B (the rat orthologs of HLA-DQ) determines the onset and severity of experimental arthritis, possibly due to the amino acid variations in the P1 pocket of RT1-B. In addition, we showed that natural allelic variants in Tap2, another gene in the MHC-II region, regulates the thymic selection of CD8+ T cells. Second, in order to investigate whether other MHC genes also contribute to arthritis susceptibility, we assessed arthritis development in congenic strains mapped to other parts of the MHC region. We identified a second arthritis-regulatory QTL in the MHC class III region, that regulates not only the onset and severity, but also chronicity of arthritis. We subsequently mapped this effect to a conserved, 33-kb large haplotype Ltab-Ncr3 comprising five polymorphic genes. Interestingly, unlike other positionally-identified arthritis genes in rats, Ltab-Ncr3 regulates only adjuvant arthritis models but not autoimmunity triggered by specific tissue antigens, such as type II collagen. Furthermore, we found that gene expression and alternative splicing of the Ltab-Ncr3 genes correlate remarkably with arthritis severity and some of the gene expression differences were reproduced in a cohort of RA patients and healthy controls. Third, the MHC-II gene expression is regulated by class II transactivator (CIITA or C2TA), and in humans, genetic variation in CIITA has been associated with differential expression of MHC-II and susceptibility to autoimmune diseases. Using a congenic mouse strain with an allelic variant in the type I promoter of C2ta, we demonstrate that whereas genetic polymorphisms in C2ta promoter result in differential MHC-II expression and antigen presentation, these do not necessarily have a strong impact on autoimmune diseases such as arthritis. In summary, these studies demonstrate how the congenic approach remains powerful to conclusively identify and characterise genes regulating a complex disease like arthriti

Profiting from Dow Jones Industrial Index and Hang Seng Index using moving average and MACD optimization model

Before the internet, high-speed laptop computers, and big data became accessible and popular, academia on stock market trading concentrated on Efficient Market Hypothesis (EMH). EMH hinges on the idea that the market is efficient and there is no extra return that could be generated. With the dynamic development of the internet, big-data and computing technology, many researchers started to pay attention to Technical Analysis and its usage. Numerous academic papers claimed that technical analysis can enhance returns by using various technical tools. This paper explores in-depth the simulation model of Moving Average and Moving Average Convergence/Divergence (MACD) to come up with optimized parameters that will allow traders to profit from trading Dow Jones Industrial Index and Hang Seng Index

Hematopoiesis in the lung: from development to adulthood

Megakaryocytes (MK) are responsible for platelet biogenesis, which is thought to occur canonically in the adult bone marrow (BM) and in the fetal liver during development. However, emerging evidence highlights the lung as a previously underappreciated residence for MKs that may significantly contribute to circulating platelet mass. While a diversity of cells specific to the BM are known to promote the maturation and trafficking of MKs, little investigation into the impact of the lung niche on the development and function of MKs has been done. Here, we describe the application of single cell RNA sequencing (scRNA-Seq) coupled with histological, ploidy and flow cytometric analyses to profile primary MKs derived from syngeneic mouse lung and hematopoietic tissues. Transcriptional profiling demonstrated that lung MKs have a unique signature distinct from their hematopoietic counterparts with lung MKs displaying enrichment for maturation markers, potentially indicating a propensity for more efficient platelet production. Reciprocally, fetal lung MKs also showed the robust expression of cytokines and growth factors known to promote lung development. Lastly, lung MKs possess an enrichment profile skewed towards roles in immunity and inflammation. These findings highlight the existence of a lung-specific MK phenotype and support the notion that the lung plays an independent role in the development and functional maturation of MKs. In addition to MKs, the lung houses many resident hematopoietic cells, including hematopoietic stem and progenitor cells (HSPCs). The existence of lung HSPCs suggests that the differentiation and development of lung resident hematopoietic cells may occur in-situ. To investigate the potential role the lung has in instructing site specific hematopoiesis, we employed explant cultures of murine and human fetal lungs. This displayed adherent endothelial cells transitioning into floating hematopoietic cells, suggesting that the fetal lung is a source of hemogenic endothelial cells that have the functional capacity to undergo endothelial to hematopoietic transition (EHT) to produce HSPCs. Flow cytometric and functional assessment of fetal lung explants showed the production of HSPCs that expressed key EHT and pre-HSPC markers. Expression profiles revealed by scRNA-Seq and small molecule modulation demonstrated that fetal lung EHT is reliant on canonical EHT signaling pathways. These findings suggest that functional HECs are present in the fetal lung, thus establishing this location as a potential extramedullary site of de-novo hematopoiesis. Overall, these findings suggest that the lung may have a greater role in instructing tissue specific hematopoiesis and/or overall hematopoietic development

Recombinant nucleases CEL I from celery and SP I from spinach for mutation detection

<p>Abstract</p> <p>Background</p> <p>The detection of unknown mutations is important in research and medicine. For this purpose, a mismatch-specific endonuclease CEL I from celery has been established as a useful tool in high throughput projects. Previously, CEL I-like activities were described only in a variety of plants and could not be expressed in an active form in bacteria.</p> <p>Results</p> <p>We describe expression of active recombinant plant mismatch endonucleases and modification of their activities. We also report the cloning of a CEL I ortholog from <it>Spinacia oleracea </it>(spinach) which we termed SP I nuclease. Active CEL I and SP I nucleases were expressed as C-terminal hexahistidine fusions and affinity purified from the cell culture media. Both recombinant enzymes were active in mutation detection in <it>BRCA1 </it>gene of patient-derived DNA. Native SP nuclease purified from spinach is unable to incise at single-nucleotide substitutions and loops containing a guanine nucleotide, but the recombinant SP I nuclease can cut at these sites.</p> <p>Conclusion</p> <p>The insect cell-expressed CEL I orthologs may not be identical to their native counterparts purified from plant tissues. The present expression system should facilitate further development of CEL I-based mutation detection technologies.</p

Anti-Reflection Coating for Nitrogen-Vacancy Optical Measurements in Diamond

We realize anti-reflection (AR) coatings for optical excitation and fluorescence measurements of nitrogen-vacancy (NV) color centers in bulk diamond by depositing quarter-wavelength thick silica layers on the diamondsurface. These AR coatings improve NV-diamond optical measurements by reducing optical reflection at the diamond-air interface from ≈17% to ≈2%, which allows more effective NV optical excitation and more efficient detection of NV fluorescence. We also show that diamondAR coatings eliminate standing-wave interference patterns of excitation laser intensity within bulk diamond, and thereby greatly reduce spatial variations in NV fluorescence, which can degrade spatially resolved magnetic field sensing using NV centers.Engineering and Applied SciencesPhysic

Boarding schools : A longitudinal examination of Australian Indigenous and non-Indigenous boarders’ and non-boarders’ wellbeing

Improving educational outcomes for Indigenous Australian students is a key strategy to helping Indigenous people reach their full potential. This has resulted in well-intentioned efforts by Australian educators and governments to ensure Indigenous children have positive school experiences. However, Indigenous students still lag behind their non-Indigenous counterparts in educational outcomes. This is particularly so for Indigenous students living in rural and remote parts of Australia where educational opportunities are limited, especially in high school. One solution to this problem has been to enrol these students in boarding schools in urban and metropolitan centres. While research on the success of boarding schools for Indigenous students is scarce, what little that does exist is not encouraging. The focus of this research was to examine the effects of boarding for Indigenous (n = 11) and non-Indigenous students’ (n = 158) wellbeing (N = 1423) in two large private boys’ schools. Participating students aged 12–18 years old completed a survey measuring wellbeing constructs on two occasions, 12 months apart. Non-Indigenous boys were generally higher in wellbeing compared with Indigenous boys. There was also evidence of improved social wellbeing beyond that of non-Indigenous boarders over time. Overall, while evidence of merit was weak, boarding schools may benefit their Indigenous students’ development in social wellbeing

Hidden negative linear compressibility in lithium L-tartrate†

Development of artificial muscles, next-generation pressure sensors and precision optics relies on advances in materials with anomalous mechanical properties. Negative linear compressibility, NLC, is one such rare, counterintuitive phenomenon, in which a material expands along one axis under hydrostatic pressure. Both classical and recent NLC materials face a pay-off between the active pressure range and magnitude of NLC, and in the vast majority of cases the NLC effect decreases with pressure. By decoupling the mechanical behaviour of building units for the first time in a winerack framework containing two different strut types, we show that lithium L-tartrate exhibits NLC with a maximum value, Kmax = -21 TPa^-1, and an overall NLC capacity, χNLC = 5.1 %, that are comparable to the most exceptional materials to date. Furthermore, the contributions from molecular strut compression and angle opening interplay to give rise to so-called “hidden” negative linear compressibility, in which NLC is absent at ambient pressure, switched on at 2 GPa and sustained up to the limit of our experiment, 5.5 GPa. Analysis of the changes in crystal structure using variable-pressure synchrotron X-ray diffraction reveals new chemical and geometrical design rules to assist the discovery of other materials with exciting hidden anomalous mechanical properties

Does a loaded warm-up influence jump asymmetry and badminton-specific change of direction performance?

Purpose: Previously, it has been shown that loaded warm-up (LWU) can improve change of direction speed (CODS) in professional badminton players. However, the effect of asymmetry on CODS in badminton players and the influence of LWU on asymmetry has not been examined. Methods: Twenty-one amateur badminton players (age: 29.5 ± 8.4; playing experience: 8.4 ± 4.2 years) completed two trials. In the first, they performed a control warm-up (CWU). In the second, they performed the same warm-up but with three exercises loaded with a weight vest (LWU). Following both warm-ups, players completed single leg jump (SLCMJ) and badminton-specific CODS tests. Results: No significant differences between CWU and LWU were observed for CODS, SLCMJ or SLCMJ asymmetry. However, small effect sizes suggested faster CODS (mean difference: -5%; d = -0.32) and lower asymmetries (mean difference: -3%; d = -0.39) following LWU. Five players (24%) experienced CODS improvements greater than the minimum detectable change whilst two (10%) responded negatively. Asymmetry was not correlated with CODS following CWU (ρ = 0.079; p = 0.733) but was negatively associated with CODS after LWU (ρ = -0.491; p = 0.035). Conclusion: LWU may prove a strategy to trial on an individual basis but generic recommendations should not be applied

New Classes of Alanine Racemase Inhibitors Identified by High-Throughput Screening Show Antimicrobial Activity against Mycobacterium tuberculosis

In an effort to discover new drugs to treat tuberculosis (TB) we chose alanine racemase as the target of our drug discovery efforts. In Mycobacterium tuberculosis, the causative agent of TB, alanine racemase plays an essential role in cell wall synthesis as it racemizes L-alanine into D-alanine, a key building block in the biosynthesis of peptidoglycan. Good antimicrobial effects have been achieved by inhibition of this enzyme with suicide substrates, but the clinical utility of this class of inhibitors is limited due to their lack of target specificity and toxicity. Therefore, inhibitors that are not substrate analogs and that act through different mechanisms of enzyme inhibition are necessary for therapeutic development for this drug target.To obtain non-substrate alanine racemase inhibitors, we developed a high-throughput screening platform and screened 53,000 small molecule compounds for enzyme-specific inhibitors. We examined the 'hits' for structural novelty, antimicrobial activity against M. tuberculosis, general cellular cytotoxicity, and mechanism of enzyme inhibition. We identified seventeen novel non-substrate alanine racemase inhibitors that are structurally different than any currently known enzyme inhibitors. Seven of these are active against M. tuberculosis and minimally cytotoxic against mammalian cells.This study highlights the feasibility of obtaining novel alanine racemase inhibitor lead compounds by high-throughput screening for development of new anti-TB agents

Alteration of Differentiation Potentials by Modulating GATA Transcription Factors in Murine Embryonic Stem Cells

Background. Mouse embryonic stem (ES) cells can be differentiated in vitro by aggregation and/or retinoic acid (RA) treatment. The principal differentiation lineage in vitro is extraembryonic primitive endoderm. Dab2, Laminin, GATA4, GATA5, and GATA6 are expressed in embryonic primitive endoderm and play critical roles in its lineage commitment. Results. We found that in the absence of GATA4 or GATA5, RA-induced primitive endoderm differentiation of ES cells was reduced. GATA4 (−/−) ES cells express higher level of GATA5, GATA6, and hepatocyte nuclear factor 4 alpha marker of visceral endoderm lineage. GATA5 (−/−) ES cells express higher level of alpha fetoprotein marker of early liver development. GATA6 (−/−) ES cells express higher level of GATA5 as well as mesoderm and cardiomyocyte markers which are collagen III alpha-1 and tropomyosin1 alpha. Thus, deletion of GATA6 precluded endoderm differentiation but promoted mesoderm lineages. Conclusions. GATA4, GATA5, and GATA6 each convey a unique gene expression pattern and influences ES cell differentiation. We showed that ES cells can be directed to avoid differentiating into primitive endoderm and to adopt unique lineages in vitro by modulating GATA factors. The finding offers a potential approach to produce desirable cell types from ES cells, useful for regenerative cell therapy