Expression of calcification and extracellular matrix genes in the cardiovascular system of the healthy domestic sheep (Ovis aries).

The maintenance of a healthy cardiovascular system requires expression of genes that contribute to essential biological activities and repression of those that are associated with functions likely to be detrimental to cardiovascular homeostasis. Vascular calcification is a major disruption to cardiovascular homeostasis, where tissues of the cardiovascular system undergo ectopic calcification and consequent dysfunction, but little is known about the expression of calcification genes in the healthy cardiovascular system. Large animal models are of increasing importance in cardiovascular disease research as they demonstrate more similar cardiovascular features (in terms of anatomy, physiology and size) to humans than do rodent species. We used RNA sequencing results from the sheep, which has been utilized extensively to examine calcification of prosthetic cardiac valves, to explore the transcriptome of the heart and cardiac valves in this large animal, in particular looking at expression of calcification and extracellular matrix genes. We then examined genes implicated in the process of vascular calcification in a wide array of cardiovascular tissues and across multiple developmental stages, using RT-qPCR. Our results demonstrate that there is a balance between genes that promote and those that suppress mineralization during development and across cardiovascular tissues. We show extensive expression of genes encoding proteins involved in formation and maintenance of the extracellular matrix in cardiovascular tissues, and high expression of hematopoietic genes in the cardiac valves. Our analysis will support future research into the functions of implicated genes in the development of valve calcification, and increase the utility of the sheep as a large animal model for understanding ectopic calcification in cardiovascular disease. This study provides a foundation to explore the transcriptome of the developing cardiovascular system and is a valuable resource for the fields of mammalian genomics and cardiovascular research

Projector-Based Embedding Eliminates Density Functional Dependence for QM/MM Calculations of Reactions in Enzymes and Solution

Combined quantum mechanics/molecular mechanics (QM/MM) methods are increasingly widely utilized in studies of reactions in enzymes and other large systems. Here, we apply a range of QM/MM methods to investigate the Claisen rearrangement of chorismate to prephenate, in solution, and in the enzyme chorismate mutase. Using projector-based embedding in a QM/MM framework, we apply treatments up to the CCSD­(T) level. We test a range of density functional QM/MM methods and QM region sizes. The results show that the calculated reaction energetics are significantly more sensitive to the choice of density functional than they are to the size of the QM region in these systems. Projector-based embedding of a wave function method in DFT reduced the 13 kcal/mol spread in barrier heights calculated at the DFT/MM level to a spread of just 0.3 kcal/mol, essentially eliminating dependence on the functional. Projector-based embedding of correlated ab initio methods provides a practical method for achieving high accuracy for energy profiles derived from DFT and DFT/MM calculations for reactions in condensed phases

2,3,4-Tribromo­thio­phene

In the title compound, C4HBr3S, there are two essentially planar mol­ecules in the asymmetric unit. In the crystal structure, bifurcated C—H⋯Br hydrogen bonds link the mol­ecules into chains. Weak Br⋯Br inter­actions [Br⋯Br = 3.634 (4)–3.691 (4) Å] then lead to undulating sheets in the bc plane

Evaluation of species-specific polyclonal antibodies to detect and differentiate between Neospora caninum and Toxoplasma gondii

Neosporosis and toxoplasmosis are major causes of abortion in livestock worldwide, leading to substantial economic losses. Detection tools are fundamental to the diagnosis and management of those diseases. Current immunohistochemistry (IHC) tests, using sera raised against whole parasite lysates, have not been able to distinguish between Toxoplasma gondii and Neospora caninum. We used T. gondii and N. caninum recombinant proteins, expressed in Escherichia coli and purified using insoluble conditions, to produce specific polyclonal rabbit antisera. We aimed to develop species-specific sera that could be used in IHC on formalin-fixed, paraffin-embedded (FFPE) tissue sections to improve the diagnosis of ruminant abortions caused by protozoa. Two polyclonal rabbit sera, raised against recombinant proteins, anti–Neospora-rNcSRS2 and anti–Toxoplasma-rTgSRS2, had specificity for the parasite they were raised against. We tested the specificity for each polyclonal serum using FFPE tissue sections known to be infected with T. gondii and N. caninum. The anti–Neospora-rNcSRS2 serum labeled specifically only N. caninum–infected tissue blocks, and the anti–Toxoplasma-rTgSRS2 serum was specific to only T. gondii–infected tissues. Moreover, tissues from 52 cattle and 19 sheep previously diagnosed by lesion profiles were tested using IHC with our polyclonal sera and PCR. The overall agreement between IHC and PCR was 90.1% for both polyclonal anti-rNcSRS2 and anti-rTgSRS2 sera. The polyclonal antisera were specific and allowed visual confirmation of protozoan parasites by IHC, but they were not as sensitive as PCR testing.</p

N-Methacryloyl-4-(piperidin-1-yl)-1,8-naphthalimide

In the title compound, C21H20N2O3, the naphthalimide unit is almost planar (r.m.s. deviation for the 15 non-H atoms = 0.059 Å). The carboximide N atom and the five C atoms of the 2-methyl­prop-2-enoyl substituent also lie in a plane (r.m.s. deviation = 0.009 Å), which subtends an angle of 84.34 (7)° to the naphthalamide plane. This orients the =CH2 group of the vinyl fragment towards the naphthalimide rings, giving the mol­ecule an extended configuration. The piperidine ring adopts a chair conformation and there is evidence for some delocalization between the naphthalene and piperidine units, the C—Npip bond length being 1.404 (4) Å. In the crystal structure, π–π contacts with centroid–centroid distances of 3.5351 (18) and 3.7794 (18) Å supported by C—H⋯O hydrogen bonds link adjacent mol­ecules in a head-to-tail fashion, forming dimers. These are further stabilized by other C—H⋯O contacts of varying strength, which stack the mol­ecules down the b axis

Faecal immunochemical tests (FIT) versus colonoscopy for surveillance after screening and polypectomy: a diagnostic accuracy and cost-effectiveness study.

OBJECTIVE: The English Bowel Cancer Screening Programme (BCSP) recommends 3 yearly colonoscopy surveillance for patients at intermediate risk of colorectal cancer (CRC) postpolypectomy (those with three to four small adenomas or one ≥10 mm). We investigated whether faecal immunochemical tests (FITs) could reduce surveillance burden on patients and endoscopy services. DESIGN: Intermediate-risk patients (60-72 years) recommended 3 yearly surveillance were recruited within the BCSP (January 2012-December 2013). FITs were offered at 1, 2 and 3 years postpolypectomy. Invitees consenting and returning a year 1 FIT were included. Participants testing positive (haemoglobin ≥40 µg/g) at years one or two were offered colonoscopy early; all others were offered colonoscopy at 3 years. Diagnostic accuracy for CRC and advanced adenomas (AAs) was estimated considering multiple tests and thresholds. We calculated incremental costs per additional AA and CRC detected by colonoscopy versus FIT surveillance. RESULTS: 74% (5938/8009) of invitees were included in our study having participated at year 1. Of these, 97% returned FITs at years 2 and 3. Three-year cumulative positivity was 13% at the 40 µg/g haemoglobin threshold and 29% at 10 µg/g. 29 participants were diagnosed with CRC and 446 with AAs. Three-year programme sensitivities for CRC and AAs were, respectively, 59% and 33% at 40 µg/g, and 72% and 57% at 10 µg/g. Incremental costs per additional AA and CRC detected by colonoscopy versus FIT (40 µg/g) surveillance were £7354 and £180 778, respectively. CONCLUSIONS: Replacing 3 yearly colonoscopy surveillance in intermediate-risk patients with annual FIT could reduce colonoscopies by 71%, significantly cut costs but could miss 30%-40% of CRCs and 40%-70% of AAs. TRIAL REGISTRATION NUMBER: ISRCTN18040196; Results

4,5-Dihydro­cyclo­penta­[b]thio­phen-6-one

The title compound, C7H6OS, crystallizes with two similar mol­ecules, 1 and 2, in the asymmetric unit. Both mol­ecules are essentially planar with r.m.s. deviations of 0.0193 and 0.0107 Å for the planes through the nine non-H atoms of mol­ecules 1 and 2, respectively. The thio­phene and 4,5-dihydro­cyclo­penta­dienone rings are inclined at 2.40 (13)° in 1 and 0.64 (13)° in 2. In the crystal structure π–π [3.6542 (17) Å] and C—H⋯π contacts stack the mol­ecules into columns in an inverse fashion along the b axis. An extensive series of C—H⋯O hydrogen bonds links the columns, generating an extended network structure

O- vs. N-protonation of 1-dimethylaminonaphthalene-8-ketones: formation of a peri N–C bond or a hydrogen bond to the pi-electron density of a carbonyl group

X-ray crystallography and solid-state NMR measurements show that protonation of a series of 1-dimethylaminonaphthalene-8-ketones leads either to O protonation with formation of a long N–C bond (1.637–1.669 Å) between peri groups, or to N protonation and formation of a hydrogen bond to the π surface of the carbonyl group, the latter occurring for the larger ketone groups (C(O)R, R = t-butyl and phenyl). Solid state 15N MAS NMR studies clearly differentiate the two series, with the former yielding significantly more deshielded resonances. This is accurately corroborated by DFT calculation of the relevant chemical shift parameters. In the parent ketones X-ray crystallography shows that the nitrogen lone pair is directed towards the carbonyl group in all cases