Genetic Analysis of Pituitary Thyrotrope Development

The pituitary gland produces polypeptide hormones that regulate many functions including growth, lactation, reproduction, metabolism, and the stress response. Pituitary thyrotrope cells produce the heterodimeric glycoprotein hormone thyrotropin, which is critical for stimulating thyroid gland development and production of thyroid hormone. Less is known about the drivers of thyrotrope cell fate than the other specialized cells in this organ. The transcription factor POU1F1 is critical for generation of thyrotropes, somatotropes and lactotropes, and GATA2 is critical for both thyrotropes and gonadotropes. Additional factors are likely involved in driving thyrotrope fate. SV40-immortalized cell lines have been invaluable for studying the regulation of pituitary hormone production. Here I use two established immortalized cell lines to identify epigenomic and gene expression changes that are associated with adoption of the thyrotrope fate. GHF-T1 cells represent a POU1F1-expressing progenitor which does not produce hormones, and TaT1 cells represent a thyrotrope-like line that expresses POU1F1, GATA2 and thyrotropin (TSH). I also developed a novel, genetically engineered mouse line that expresses SV40 in response to cre recombinase, and I used this line to develop novel pituitary cell lines. These cell lines can be used for transcriptome and epigenome studies to understand the development and function of the pituitary gland. I identified the transcription factors and epigenomic changes in chromatin that are associated with thyrotrope differentiation. I generated and integrated genome-wide information about DNA accessibility, histone modifications, POU1F1 binding and RNA expression data to identify regulatory elements and candidate transcriptional regulators. I identified POU1F1 binding sites that are unique to each cell line. POU1F1 binding sites are commonly associated with bZIP factor motifs in GHF-T1 cells and Helix-Turn-Helix or basic Helix-Loop-Helix motifs in TαT1 cells, suggesting classes of transcription factors that may recruit POU1F1 to unique sites. I validated enhancer function of novel elements we mapped near Cga, Pitx1, Gata2, and Tshb by transfection in TαT1 cells. Finally, I confirmed that an enhancer element near Tshb can drive expression in thyrotropes of transgenic mice and demonstrated that GATA2 enhances Tshb expression via this element. These data extend the ENCODE analysis to an organ that is critical for growth and metabolism. This information could be valuable for understanding pituitary development and disease pathogenesis. Targeted oncogenesis is the process of driving tumor formation by engineering transgenic mice that express an oncogene under the control of a cell-type specific promoter. Using CRISPR/Cas9 we inserted a cassette with coding sequences for SV40 T antigens and IRES-GFP into the Rosa26 locus, downstream from a stop sequence flanked by loxP sites: Rosa26 LSL-SV40-GFP . These mice were mated with previously established Prop1-cre and Tshb-cre transgenic lines. The majority of Rosa26 LSL-SV40-GFP/+ ; Prop1-cre and all Rosa26 LSL-SV40-GFP/+ ; Tshb-cre mice developed dwarfism and large tumors by 4 weeks. Prop1-cre-mediated activation of SV40 expression affected cell specification, reducing thyrotrope differentiation and increasing gonadotrope cell fate selection. GFP-positive cells from flow-sorted Rosa26 LSL-SV40GFP/+ LSL-SV40-GFP/+; Prop1-cre and Rosa26 ; Tshb-cre mice express PROP1 and TSH, respectively. Tumors from both of these mouse lines were adapted to growth in cell culture. I established a progenitor-like cell line (PIT-P1) that expresses Sox2 and Pitx1, and a thyrotrope-like cell line (PIT-T1) that expresses Cga and Pou1f1. These studies demonstrate the utility of the novel, Rosa26 LSL-SV40-GFP mouse line for targeted oncogenesis and development of cell lines.PHDGenetics and Genomics PhDUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/162910/1/azdaly_1.pd

A systematic review of measures of self-reported adherence to unsupervised home-based rehabilitation exercise programmes, and their psychometric properties

BACKGROUND: Adherence is an important factor contributing to the effectiveness of exercise-based rehabilitation. However, there appears to be a lack of reliable, validated measures to assess self-reported adherence to prescribed but unsupervised home-based rehabilitation exercises. OBJECTIVES: A systematic review was conducted to establish what measures were available and to evaluate their psychometric properties. DATA SOURCES: MEDLINE, EMBASE, PsycINFO CINAHL (June 2013) and the Cochrane library were searched (September 2013). Reference lists from articles meeting the inclusion criteria were checked to ensure all relevant papers were included. STUDY SELECTION: To be included articles had to be available in English; use a self-report measure of adherence in relation to a prescribed but unsupervised home-based exercise or physical rehabilitation programme; involve participants over the age of 18. All health conditions and clinical populations were included. DATA EXTRACTION: Descriptive data reported were collated on a data extraction sheet. The measures were evaluated in terms of eight psychometric quality criteria. RESULTS: 58 studies were included, reporting 61 different measures including 29 questionnaires, 29 logs, two visual analogue scales and one tally counter. Only two measures scored positively for one psychometric property (content validity). The majority of measures had no reported validity or reliability testing. CONCLUSIONS: The results expose a gap in the literature for well-developed measures that capture self-reported adherence to prescribed but unsupervised home-based rehabilitation exercises

Biomechanical characterization of swimmers with physical disabilities

The evaluation of swimming technique is one of the main aspects to be considered in any training program, with biomechanics being an important source of knowledge. It was our objective to characterize the biomechanical parameters (SL and SF) relating them to the swimming velocity (v) at different intensities and to analyze within each swimming stroke cycle the intra-cyclic velocity variation (IVV) in a group of motor disabled swimmers. Eight disabled male swimmers (25.83 ± 2.93 years old, 72.45 ± 9.26 kg body mass and 1.79 ± 0.11 m of height) of the following functional classes: S6 (n = 1), S8 (n = 2) and S9 (n = 5) participated in this study. Swimmers were evaluated in the kinematic parameters v, stroke frequency (SF) and stroke length (SL) along with an incremental protocol of 6 x 200 m in the the crawl stroke. Data were registered in each step at the distances of 100 and 175 m. With increasing velocity, the mean values of SL decreased while the mean values of SF increased. To achieve higher swimming velocities, swimmers compensated the lack of the propulsive segment increasing SF to increase swimming speed. For the mean values of IVV at 100m distance, a decrease between the first and second levels, followed by a tendency to stabilize from the 2nd to the 6th level is presented. For the 175 m distance, there was a decrease in IVV with an increase in swimming velocity. Stroke frequency is directly related to the magnitude of IVV, which directly influences swimming performance

Genetics of combined pituitary hormone deficiency: Roadmap into the genome era

The genetic basis for combined pituitary hormone deficiency (CPHD) is complex, involving 30 genes in a variety of syndromic and nonsyndromic presentations. Molecular diagnosis of this disorder is valuable for predicting disease progression, avoiding unnecessary surgery, and family planning. Weexpect that the application of high throughput sequencing will uncover additional contributing genes and eventually become a valuable tool for molecular diagnosis. For example, in the last 3 years, six new genes have been implicated in CPHD using whole-exome sequencing. In this review, we present a historical perspective on gene discovery for CPHD and predict approaches that may facilitate future gene identification projects conducted by clinicians and basic scientists. Guidelines for systematic reporting of genetic variants and assigning causality are emerging. We apply these guidelines retrospectively to reports of the genetic basis of CPHD and summarize modes of inheritance and penetrance for each of the known genes. In recent years, there have been great improvements in databases of genetic information for diverse populations. Some issues remain that make molecular diagnosis challenging in some cases. These include the inherent genetic complexity of this disorder, technical challenges like uneven coverage, differing results from variant calling and interpretation pipelines, the number of tolerated genetic alterations, and imperfect methods for predicting pathogenicity.Wediscuss approaches for future research in the genetics of CPHD.Fil: Fang, Qing. University of Michigan; Estados UnidosFil: George, Akima S.. University of Michigan; Estados UnidosFil: Brinkmeier, Michelle L.. University of Michigan; Estados UnidosFil: Mortensen, Amanda H.. University of Michigan; Estados UnidosFil: Gergics, Peter. University of Michigan; Estados UnidosFil: Cheung, Leonard Y.M.. University of Michigan; Estados UnidosFil: Daly, Alexandre Z.. University of Michigan; Estados UnidosFil: Ajmal, Adnan. University of Michigan; Estados UnidosFil: Pérez Millán, María Inés. University of Michigan; Estados Unidos. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Instituto de Investigaciones Biomédicas. Universidad de Buenos Aires. Facultad de Medicina. Instituto de Investigaciones Biomédicas; ArgentinaFil: Bilge Ozel, A.. University of Michigan; Estados UnidosFil: Kitzman, Jacob. University of Michigan; Estados UnidosFil: Mills, Ryan E.. University of Michigan; Estados UnidosFil: Li, Jun Z.. University of Michigan; Estados UnidosFil: Camper, Sally. University of Michigan; Estados Unido

A two-directional approach to pyrrolizidines: total syntheses and biological evaluation of alkaloid cis-223B and (+/-)-xenovenine

Total syntheses of alkaloid cis-223B and xenovenine are reported in 3 and 4 steps respectively using a two-directional synthesis/triple reductive amination strategy, and their neurotoxic properties assessed

Should formula for infants provide arachidonic acid along with DHA? A position paper of the European Academy of Paediatrics and the Child Health Foundation

Recently adopted regulatory standards on infant and follow-on formula for the European Union stipulate that from 2021 onwards, all such products marketed in the European Union must contain 20-50 mg/100 kcal of omega-3 docosahexaenoic acid (DHA), which is equivalent to about 0.5-1 % of fatty acids and thus higher than typically found in human milk and current infant formula products, without the need to also include omega-6 arachidonic acid (ARA). This novel concept of infant formula composition has given rise to concern and controversy since there is no accountable evidence on the suitability and safety in healthy infants. Therefore, international experts in the field of infant nutrition were invited to review the state of scientific research on DHA and ARA, and to discuss the questions arising from the new European regulatory standards. Based on the available information, we recommend that infant and follow-on formula should provide both DHA and ARA. The DHA should equal at least the mean content in human milk globally (0.3 % of fatty acids) but preferably reach a level of 0.5 % of fatty acids. While optimal ARA intake levels remain to be defined, we strongly recommend that ARA should be provided along with DHA. At levels of DHA in infant formula up to about 0.64%, ARA contents should at least equal the DHA contents. Further well-designed clinical studies should evaluate the optimal intakes of DHA and ARA in infants at different ages based on relevant outcome

Time-resolved, defect-hosted, trace element mobility in deformed Witwatersrand pyrite

© 2018 China University of Geosciences (Beijing) and Peking University The Pb isotopic composition of rocks is widely used to constrain the sources and mobility of melts and hydrothermal fluids in the Earth's crust. In many cases, the Pb isotopic composition appears to represent mixing of multiple Pb reservoirs. However, the nature, scale and mechanisms responsible for isotopic mixing are not well known. Additionally, the trace element composition of sulphide minerals are routinely used in ore deposit research, mineral exploration and environmental studies, though little is known about element mobility in sulphides during metamorphism and deformation. To investigate the mechanisms of trace element mobility in a deformed Witwatersrand pyrite (FeS2), we have combined electron backscatter diffraction (EBSD) and atom probe microscopy (APM). The results indicate that the pyrite microstructural features record widely different Pb isotopic compositions, covering the entire range of previously published sulphide Pb compositions from the Witwatersrand basin. We show that entangled dislocations record enhanced Pb, Sb, Ni, Tl and Cu composition likely due to entrapment and short-circuit diffusion in dislocation cores. These dislocations preserve the Pb isotopic composition of the pyrite at the time of growth (~3 Ga) and show that dislocation intersections, likely to be common in deforming minerals, limit trace element mobility. In contrast, Pb, As, Ni, Co, Sb and Bi decorate a high-angle grain boundary which formed soon after crystallisation by sub-grain rotation recrystallization. Pb isotopic composition within this boundary indicates the addition of externally-derived Pb and trace elements during greenschist metamorphism at ~2 Ga. Our results show that discrete Pb reservoirs are nanometric in scale, and illustrate that grain boundaries may remain open systems for trace element mobility over 1 billion years after their formation

Hunt for new phenomena using large jet multiplicities and missing transverse momentum with ATLAS in 4.7 fb−1 of s√=7TeV proton-proton collisions

Results are presented of a search for new particles decaying to large numbers of jets in association with missing transverse momentum, using 4.7 fb−1 of pp collision data at s√=7TeV collected by the ATLAS experiment at the Large Hadron Collider in 2011. The event selection requires missing transverse momentum, no isolated electrons or muons, and from ≥6 to ≥9 jets. No evidence is found for physics beyond the Standard Model. The results are interpreted in the context of a MSUGRA/CMSSM supersymmetric model, where, for large universal scalar mass m 0, gluino masses smaller than 840 GeV are excluded at the 95% confidence level, extending previously published limits. Within a simplified model containing only a gluino octet and a neutralino, gluino masses smaller than 870 GeV are similarly excluded for neutralino masses below 100 GeV