Abstract  Autosomal recessive Adams-Oliver syndrome was diagnosed in three remotely related Bedouin consanguineous families. Genome-wide linkage analysis ruled out association with known Adams-Oliver syndrome genes, identifying a single-homozygosity ∼1.8-Mb novel locus common to affected individuals (LOD score 3.37). Whole-exome sequencing followed by Sanger sequencing identified only a single mutation within this locus, shared by all affected individuals and found in patients from five additional apparently unrelated Bedouin families: a 1-bp deletion mutation in a predicted alternative splice variant of EOGT, leading to a putative truncated protein. RT-PCR demonstrated that the EOGT-predicted alternative splice variant is ubiquitously expressed. EOGT encodes EGF-domain-specific O-linked N-acetylglucosamine transferase, responsible for extracellular O-GlcNAcylation of epidermal growth factor-like domain-containing proteins, and is essential for epithelial cell-matrix interactions. F-actin staining in diseased fibroblasts showed apparently intact cell cytoskeleton and morphology, suggesting the EOGT mutation acts not through perturbation of cytoskeleton but through other mechanisms yet to be elucidated.

Abstract  BACKGROUND: The continued advance of antibiotic resistance threatens the treatment and control of many infectious diseases. This is exemplified by the largest global outbreak of extensively drug-resistant (XDR) tuberculosis (TB) identified in Tugela Ferry, KwaZulu-Natal, South Africa, in 2005 that continues today. It is unclear whether the emergence of XDR-TB in KwaZulu-Natal was due to recent inadequacies in TB control in conjunction with HIV or other factors. Understanding the origins of drug resistance in this fatal outbreak of XDR will inform the control and prevention of drug-resistant TB in other settings. In this study, we used whole genome sequencing and dating analysis to determine if XDR-TB had emerged recently or had ancient antecedents.

METHODS AND FINDINGS: We performed whole genome sequencing and drug susceptibility testing on 337 clinical isolates of Mycobacterium tuberculosis collected in KwaZulu-Natal from 2008 to 2013, in addition to three historical isolates, collected from patients in the same province and including an isolate from the 2005 Tugela Ferry XDR outbreak, a multidrug-resistant (MDR) isolate from 1994, and a pansusceptible isolate from 1995. We utilized an array of whole genome comparative techniques to assess the relatedness among strains, to establish the order of acquisition of drug resistance mutations, including the timing of acquisitions leading to XDR-TB in the LAM4 spoligotype, and to calculate the number of independent evolutionary emergences of MDR and XDR. Our sequencing and analysis revealed a 50-member clone of XDR M. tuberculosis that was highly related to the Tugela Ferry XDR outbreak strain. We estimated that mutations conferring isoniazid and streptomycin resistance in this clone were acquired 50 y prior to the Tugela Ferry outbreak (katG S315T [isoniazid]; gidB 130 bp deletion [streptomycin]; 1957 [95% highest posterior density (HPD): 1937-1971]), with the subsequent emergence of MDR and XDR occurring 20 y (rpoB L452P [rifampicin]; pncA 1 bp insertion [pyrazinamide]; 1984 [95% HPD: 1974-1992]) and 10 y (rpoB D435G [rifampicin]; rrs 1400 [kanamycin]; gyrA A90V [ofloxacin]; 1995 [95% HPD: 1988-1999]) prior to the outbreak, respectively. We observed frequent de novo evolution of MDR and XDR, with 56 and nine independent evolutionary events, respectively. Isoniazid resistance evolved before rifampicin resistance 46 times, whereas rifampicin resistance evolved prior to isoniazid only twice. We identified additional putative compensatory mutations to rifampicin in this dataset. One major limitation of this study is that the conclusions with respect to ordering and timing of acquisition of mutations may not represent universal patterns of drug resistance emergence in other areas of the globe.

CONCLUSIONS: In the first whole genome-based analysis of the emergence of drug resistance among clinical isolates of M. tuberculosis, we show that the ancestral precursor of the LAM4 XDR outbreak strain in Tugela Ferry gained mutations to first-line drugs at the beginning of the antibiotic era. Subsequent accumulation of stepwise resistance mutations, occurring over decades and prior to the explosion of HIV in this region, yielded MDR and XDR, permitting the emergence of compensatory mutations. Our results suggest that drug-resistant strains circulating today reflect not only vulnerabilities of current TB control efforts but also those that date back 50 y. In drug-resistant TB, isoniazid resistance was overwhelmingly the initial resistance mutation to be acquired, which would not be detected by current rapid molecular diagnostics employed in South Africa that assess only rifampicin resistance.

Abstract  This present work describes the production and biochemical characterization of lipase by Candida rugosa and Geotrichum candidum in a culture supplemented with soybean molasses. After optimizing the fermentation times for both microorganisms, the effects of changing the soybean molasses concentration, the fermentative medium pH and the fermentation temperature were evaluated using the Central Composite Planning. When soybean molasses was used at a concentration of 200 g/L at 27 ± 1 °C and pH 3.5, the lipolytic activity measured in the broth was 12.3 U/mL after 12 h for C. rugosa and 11.48 U/mL after 24 h for G. candidum. The molecular masses were 38.3 kDa to G. candidum lipase and 59.7 kDa to C. rugosa lipase, determined by SDS-PAGE. The lipase from both microorganisms exhibited maximal hydrolytic activity at a temperature of 40 °C and were inhibited at pH 10.0. Using different concentration of p-nitrophenylbutyrate (p-NPB), the kinetic parameters were calculated, as follows: the Km of lipase from G. candidum was 465.44 μM and the Vmax 0.384 μmol/min; the Km and Vmax of lipase from C. rugosa were 129.21 μM and 0.034 μmol/min, respectively. Lipases activity were increased by metallic ions Mg(2+) and Na(+) and inhibited by metallic ion Cu(3+).

Abstract  Over-weight and obesity are serious problems that increase the risk not only for chronic diseases like diabetes and heart disease but also of various types of cancer. This study was conducted to evaluate cytokinesis-block micronucleus cytome (CBMN-cyt) assay parameters and plasma concentrations of 8-hydroxy-2'-deoxyguanosine (8-OHdG), and their relationship with age, body-mass index (BMI) and waist-to-hip ratio (WHR) in 83 obese, 21 over-weight and 21 normal-weight subjects. Frequencies of micronuclei (MN), nucleoplasmic bridges (NPB), nuclear buds (NBUD), and apoptotic and necrotic cells in lymphocytes of obese subjects were found to be significantly higher than those found in normal-weight and over-weight subjects (p<0.01 and p<0.05), whereas plasma concentrations of 8-OHdG in obese subjects were lower than those observed in normal-weight and over-weight subjects (p<0.05 and p<0.01, respectively). There was a negative correlation between age and frequency of necrotic cells and NDI (p<0.05), whereas there was no correlation between BMI, WHR, CBMN cyt assay parameters and plasma 8-OHdG in normal-weight subjects. In over-weight subjects, a negative correlation was observed between age and NDI (p<0.01) and a positive correlation between age and frequency of NPB (p<0.01) and between BMI and frequency of NBUD (p<0.05). In obese subjects, a negative correlation was observed between age and NDI (p<0.01) and between BMI and NDI (p<0.05), whereas no correlation was observed between WHR and CBMN-cyt assay parameters and plasma 8-OHdG. However, frequencies of MN, NPB, NBUD, apoptotic and necrotic cells in total over-weight/obese (p<0.01/p<0.05) and all subjects (p<0.01) increased with increasing BMI. The increase in genomic damage (MN, NPB and NBUD) in obese subjects and the positive correlation between genomic damage and BMI in total over-weight/obese subjects indicate that obesity increases genomic damage and may be associated with an increased risk of cancer, because an increase in MN frequency is a predictor of cancer risk.

Abstract  Osteogenesis imperfecta (OI) is a genetically heterogeneous group of disorders, characterized by abnormal bone fragility, blue sclera, deafness, joint laxity, and soft-tissue dysplasia. The purpose of this study was to elucidate the genetic or molecular basis for OI type IA in a Chinese family. We evaluated the members of a family, in which six individuals are affected with increased bone fragility and blue sclera. Results of exome sequencing revealed a novel 1-bp deletion (c.2329delG, p.A777fs) in exon 33 of the COL1A1 gene in two affected individuals, but not in a control family member without OI. The variation co-segregated with the disease in all the OI patients but not in the unaffected family members. The mutation caused a frameshift alteration after codon 777, leading to premature termination of the COL1A1 protein. Thus, our findings identified a novel frameshift deletion c.2329delG (p.A777fs) in the COL1A1 gene, which is associated with OI type IA in a Chinese family.

Abstract  BACKGROUND: Lead is a heavy toxic metal element in biological systems and is one of the major pollutants as a result of its widespread use in industries. In spite of its negative roles the coordination chemistry of Pb(II) complexes is a matter of interest. The N,N'-bidentate aromatic bases such as BPY,4-BPY and PHEN (BPY = 2,2'bipyridine, 4-BPY = 4,4'-dimethyl-2,2'-bipyridine, PHEN = 1,10-Phenanthroline) are widely used to build supramolecular architectures because of their excellent coordinating ability and large conjugated system that can easily form π-π interactions among their aromatic moieties. A series of novel Pb(II) complexes in concert with 5-CTPC, 5-BTPC (5-CTPC = 5-chlorothiophen-2-carboxylate, 5-BTPC = 5-bromothiophen-2-carboxylate) and corresponding bidentate chelating N.N' ligands have been synthesized and characterized.

RESULTS: Five new Pb (II) complexes [Pb(BPY)(5-CTPC)2] (1), [Pb(4-BPY)(5-CTPC)2] (2), [Pb2(PHEN)2(5-CTPC)4] (3), [Pb(4-BPY)(5-BTPC)2] (4) and [Pb2(PHEN)2(5-BTPC)2(ACE)2] (5) have been synthesized. Even though in all these complexes the molar ratio of Pb, carboxylate, N,N-chelating ligand are the same (1:2:1), there is a significant structural diversity. These complexes have been characterised and investigated by elemental analysis, IR, 1H-NMR,13C-NMR, TGA, and photoluminescence studies. Single crystal X-ray diffraction studies reveal that complexes (1, 2) and (4) are mononuclear while (3 and 5) are dinuclear in nature which may result from the chelating nature of the ligands, various coordination modes of the carboxylates, and the coordination geometry of the Pb(II) ions.

CONCLUSIONS: The observation of structures 2,4 and 3,5 show the structural changes made just chloro/bromo substituent of the thiophene ring. A detailed packing analysis has been undertaken to delineate the role of valuable non covalent interactions like X…π, H…X, (X = Cl/Br). A quadruple hydrogen bond linking the monomeric units and generating a supramolecular architecture is observed in (1). The metal bite unit comprised of PbN2C2 (i.e. Pb-N-C-C-N-Pb) is the repeating unit in all the five complexes and they have almost same geometrical parameters. This metal bite has been identified as the self assembly unit in complexes.

Abstract  The use of fixed combinations in acne vulgaris (acne) is very common, however comparative clinical trial data are limited. Cutaneous tolerability can influence patient compliance, and concerns about skin irritation with topical acne treatments have lead to a number of comparative split-face studies. Recently, a new fixed combination product was introduced (clin 1.0%-BP 3.75% gel) that was shown to be effective in reducing both inflammatory and noninflammatory lesions in moderate to severe acne. Here, we assess the tolerability of clin 1.0%-BP 3.75% gel compared with adap 0.1%-BP 2.5% gel in healthy volunteers with no apparent facial redness or dryness over 21-days, using a split-face methodology. Especially over the first two weeks of treatment, clin 1.0%-BP 3.75% gel was more tolerable than adap 0.1%-BP 2.5% gel, with statistically significant differences in cumulative change from baseline starting as early as day 8 (dryness) and day 9 (erythema), and composite index on days 8-12 and 16. Transepidermal water loss was less with clin 1.0%-BP 3.75% gel, although the difference was not statistically significant.

Abstract  A substrain of mice originating from the CF#1 strain (an outbred colony) reared at Osaka Prefecture University (CF#1/lr mice) develops cataracts beginning at 4 weeks of age. Affected mice were fully viable and fertile and developed cataracts by 14 weeks of age. Histologically, CF#1/lr mice showed vacuolation of the lens cortex, swollen lens fibers, lens rupture and nuclear extrusion. To elucidate the mode of inheritance, we analyzed heterozygous mutant hybrids generated from CF#1/lr mice and wild-type BALB/c mice. None of the heterozygous mutants were affected, and the ratio of affected to unaffected mice was 1:3 among the offspring of the heterozygous mutants. For the initial genome-wide screening and further mapping, we used affected progeny of CF#1/lr × (CF#1/lr × BALB/c) mice. We concluded that the cataracts in CF#1/lr mice are inherited through an autosomal recessive mutation and that the mutant gene is located on mouse chromosome 3 between D3Mit79 and D3Mit216. In this region, we identified 8 genes associated with ocular disease. All 8 genes were sequenced and a novel point mutation (1 bp insertion of cytosine) in exon 7 of the Bcar3 gene was identified. This mutation produced a premature stop codon and a truncated protein. In conclusion, we have identified the first spontaneous mutation in the Bcar3 gene associated with lens extrusion cataracts. This novel cataract model may provide further knowledge of the molecular biology of cataractogenesis and the function of the BCAR3 protein.

Abstract  BACKGROUND: Ultra violet (UV) filters with known or suspected endocrine disrupting properties are widely used in sunscreens and other personal care products, clothing, food packaging and many other consumer products. Danish kindergarten children have sunscreens applied daily during summer to prevent skin burns.

OBJECTIVES: To estimate the assumed contribution of sunscreens to the total exposure to UV filters, we measured the urinary excretion of UV filters during summer and winter in kindergarten children.

METHODS: Spot- and first morning urines were collected during a summer and a winter day in 2013. A total of 266 urine samples were collected from 55 children and were analysed for content of benzophenone (BP), benzophenone-1 (BP-1), benzophenone-2 (BP-2), benzophenone-3 (BP-3), 5-chloro-2-hydroxybenzophenone (BP-7), 4-methyl-benzophenone (4-MBP), 4-hydroxybenzophenone (4-HBP), 3-(4-methylbenzylidene)-camphor (4-MBC), and 3-benzylidene camphor (3-BC) by LC-MS/MS.

RESULTS: Of the analysed UV filters, the children excreted predominantly BP-1, BP-3 and 4-HBP. The urine levels were significantly higher in summer samples compared to winter samples, however exposure during winter was still evident. Furthermore, children with the highest concentrations of UV filters in summer urines also tended to be among those with the highest winter levels.

CONCLUSION: Exposures to UV filters during summertime can partly be explained by the intended use of UV filters in sunscreens, which is considered to be beneficial for children during outdoor activities. However, exposure to UV filters all year round together with large inter-individual variation indicate that children's exposure to UV filters also comes from other consumer items, presumably highly influenced by the general lifestyle of an individual child: this is completely unintended, without benefit, and potentially harmful.

Abstract  UNLABELLED: Mulibrey nanism (MUL) is a rare autosomal recessive disorder with severe primordial growth retardation and multiorgan involvement, caused by mutations in TRIM37. Early clinical detection is important since more than 50 % of the patients develop congestive heart failure. We report a 12-year-old patient who presented in infancy with severe growth retardation, dysmorphic features, and cleft palate. Clinical diagnosis of MUL was established at the age of 5 years. Postmortem, molecular diagnostic confirmed MUL as a novel 1-bp deletion (c.1233delA) in exon 14 of the TRIM37 coding region. Cardiac examination at the age of 6 years revealed constrictive pericarditis with significant elevation of atrial filling pressures, consecutive hepatomegaly, and protein loosing enteropathy. Since the parents refused pericardectomy, surgery was delayed until the age of 12 years, when congestive heart failure deteriorated. Despite pericardectomy, the boy died from persistent right heart failure.

CONCLUSION: Our report underlines the necessity of early clinical diagnosis of Mulibrey nanism. Careful cardiologic examination is required to detect constrictive pericarditis, which is a major factor of mortality in these patients. Pericardectomy should be performed early, to avoid sequelae of persisting congestive heart failure.

Abstract  Nucleoli in mammalian oocytes and zygotes, sometimes referred to as nucleolus precursor bodies (NPBs), are compact and morphologically different from nucleoli in somatic cells. We applied a unique NPB analyzing method "enucleolation" technique to zygotes to remove the NPBs. It has been reported that oocyte NPBs are essential for embryonic development; in their absence, the oocytes complete maturation and can be fertilized, but no nucleoli are formed in the zygotes and embryos, leading to developmental failure. However, we found that when NPBs were removed from zygotes, the zygotes developed successfully to live-born pups. These results indicated that oocyte NPBs are essential for embryonic development, but zygote NPBs are not. In addition, the enucleolated zygotes formed somatic-type nucleoli during early embryonic development, demonstrating that somatic-type nucleoli do not originate from zygote NPBs. We summarize our recent investigation on NPBs, and provide additional comments and findings.

Abstract  BACKGROUND AND AIMS: Patients with established coronary artery disease (CAD) are likely to receive a combination of aspirin, a statin, and blood pressure (BP)-lowering agents. Combining these pharmacologic agents into a cardiovascular combination pill, such as a polypill, could be considered to reduce prescription gaps and nonadherence in high-risk patients. We aimed to evaluate the effect of the concomitant use of aspirin, a statin, and BP-lowering agent(s) in patients with CAD on vascular morbidity and mortality in current clinical practice in an observational study to provide insights in the combination pill concept related to feasibility and applicability.

METHODS: In total, 2,706 patients with CAD enrolled in the Second Manifestations of ARTerial disease study were followed for the occurrence of a subsequent vascular event (ie, myocardial infarction, ischemic cerebrovascular accident, vascular death) and all-cause mortality. The relationship between combination therapy and cardiovascular events and all-cause mortality was assessed using Cox proportional hazards regression models to calculate hazards ratios (HRs) with a 95% CI. Both covariate and propensity score adjusting methods were used to reduce confounding by indication.

RESULTS: A combination of aspirin, a statin, and ≥1 BP-lowering agent(s) was used by 67% of the patients. During a median of 5.0 years (interquartile range 2.4-10.2 years), 347 vascular events occurred and 162 patients died. Combination therapy with aspirin, statin, and ≥1 BP-lowering agent was associated with a lower risk of myocardial infarction (HR 0.68, 95% CI 0.49-0.96), ischemic cerebrovascular accident (HR 0.37, 95% CI 0.16-0.84), composite vascular end point (HR 0.66, 95% CI 0.49-0.88), vascular mortality (HR 0.53, 95% CI 0.33-0.85), and all-cause mortality (HR 0.69, 95% CI 0.49-0.96) compared with the absence of combination therapy, after adjusting for confounding covariates in a propensity score. The use of 1 or only 2 components of combination therapy was associated with a higher risk for cardiovascular events compared with the combined use of aspirin, a statin, and ≥1 BP-lowering agent(s).

CONCLUSION: Two-thirds of the patients with CAD use a combination of aspirin, a statin, and ≥1 BP-lowering agent(s), components of a cardiovascular fixed-dose combination pill. Combination therapy with these agents is associated with a lower risk of vascular events and total mortality. Although treatment effect in observational studies should be interpreted with caution, the results of this study support supposed benefits from combination therapy. However, the effect of fixed-dose combination pill on clinical outcome needs to be demonstrated in randomized clinical trials.

Abstract  PURPOSE: To identiy the disease causing mutation in a Chinese family presenting with early-onset cataract and dental anomalies.

MATERIALS AND METHODS: A specific Hereditary Eye Disease Enrichment Panel (HEDEP) (personalized customization by MyGenostics, Baltimore, MD) based on targeted exome capture technology was used to collect the protein coding regions of 30 early-onset cataract associated genes, and high throughput sequencing was done with Illumina HiSeq 2000 platform. The identified variant was confirmed with Sanger sequencing.

RESULTS: A novel deletion in exon 4 (c.852delG) of NHS gene was identified; the identified 1 bp deletion altered the reading frame and was predicted to result in a premature stop codon after the addition of twelve novel amino acid (p.S285PfsX13). This mutation co-segregated in affected males and obligate female carriers, but was absent in 100 matched controls.

CONCLUSIONS: Our findings broaden the spectrum of NHS mutations causing Nance-Horan syndrome and phenotypic spectrum of the disease in Chinese patients.

Abstract  We performed genome-wide analyses to identify genomic loci that interact with sodium to influence blood pressure (BP) using single-marker-based (1 and 2 df joint tests) and gene-based tests among 1876 Chinese participants of the Genetic Epidemiology Network of Salt-Sensitivity (GenSalt) study. Among GenSalt participants, the average of 3 urine samples was used to estimate sodium excretion. Nine BP measurements were taken using a random zero sphygmomanometer. A total of 2.05 million single-nucleotide polymorphisms were imputed using Affymetrix 6.0 genotype data and the Chinese Han of Beijing and Japanese of Tokyo HapMap reference panel. Promising findings (P<1.00×10(-4)) from GenSalt were evaluated for replication among 775 Chinese participants of the Multi-Ethnic Study of Atherosclerosis (MESA). Single-nucleotide polymorphism and gene-based results were meta-analyzed across the GenSalt and MESA studies to determine genome-wide significance. The 1 df tests identified interactions for UST rs13211840 on diastolic BP (P=3.13×10(-9)). The 2 df tests additionally identified associations for CLGN rs2567241 (P=3.90×10(-12)) and LOC105369882 rs11104632 (P=4.51×10(-8)) with systolic BP. The CLGN variant rs2567241 was also associated with diastolic BP (P=3.11×10(-22)) and mean arterial pressure (P=2.86×10(-15)). Genome-wide gene-based analysis identified MKNK1 (P=6.70×10(-7)), C2orf80 (P<1.00×10(-12)), EPHA6 (P=2.88×10(-7)), SCOC-AS1 (P=4.35×10(-14)), SCOC (P=6.46×10(-11)), CLGN (P=3.68×10(-13)), MGAT4D (P=4.73×10(-11)), ARHGAP42 (P≤1.00×10(-12)), CASP4 (P=1.31×10(-8)), and LINC01478 (P=6.75×10(-10)) that were associated with at least 1 BP phenotype. In summary, we identified 8 novel and 1 previously reported BP loci through the examination of single-nucleotide polymorphism and gene-based interactions with sodium.

Abstract  Patients spend the vast majority of their hospital stay in an unmonitored bed where various mobility factors can impact patient safety and quality. Specifically, bed positioning and a patient's related mobility in that bed can have a profound impact on risks such as pneumonias, blood clots, bed ulcers and falls. This issue has been exacerbated as the nurse-per-bed (NPB) ratio has decreased in recent years. To help assess these risks, it is critical to monitor a hospital bed's positional status (BPS). Two bed positional statuses, bed height (BH) and bed chair angle (BCA), are of critical interests for bed monitoring. In this paper, we develop a bed positional status detection system using a single Microsoft Kinect. Experimental results show that we are able to achieve 94.5% and 93.0% overall accuracy of the estimated BCA and BH in a simulated patient's room environment.

Abstract  BACKGROUND: Reelin plays a pivotal role in neurodevelopment and in post-natal synaptic plasticity and has been implicated in the pathogenesis of autism spectrum disorder (ASD). The reelin (RELN) gene expression is significantly decreased in ASD, both in the brain and peripherally. Methylation at the RELN gene promoter is largely triggered at puberty, and hypermethylation has been found in post-mortem brains of schizophrenic and bipolar patients.

METHODS: In this study, we assessed RELN gene methylation status in post-mortem temporocortical tissue samples (BA41/42 or 22) of six pairs of post-puberal individuals with ASD and typically developing subjects, matched for sex (male:female, M:F = 5:1), age, and post-mortem interval.

RESULTS: ASD patients display a significantly higher number of methylated CpG islands and heavier methylation in the 5' region of the RELN gene promoter, spanning from -458 to -223 bp, whereas controls have more methylated CpG positions and greater extent of methylation at the 3' promoter region, spanning from -222 to +1 bp. The most upstream promoter region (-458 to -364 bp) is methylated only in ASD brains, while the most downstream region (-131 to +1 bp) is methylated exclusively in control brains. Within this general framework, three different methylation patterns are discernible, each correlated with different extents of reduction in reelin gene expression among ASD individuals compared to controls.

CONCLUSIONS: The methylation pattern is different in ASD and control post-mortem brains. ASD-specific CpG positions, located in the most upstream gene promoter region, may exert a functional role potentially conferring ASD risk by blunting RELN gene expression.

Abstract  H1.1 and H1.4 bind tightly to both short DNA oligomers and to CT-DNA (Ka≈1×10(7)). Binding is accompanied by an unfavorable enthalpy change (∆H≈+22 kcal/mol) and a favorable entropy change (-T∆S≈-30 kcal/mol). The Tm for the H1.4/CT-DNA complex is increased by 9 °C over the Tm for the free DNA. H1.4 titrations of the DNA oligomers yield stoichiometries (H1/DNA) of 0.64, 0.96, 1.29, and 2.04 for 24, 36, 48, and 72-bp DNA oligomers. The stoichiometries are consistent with a binding site size of 37±1 bp. CT-DNA titration data are consistent with binding site sizes of 32 bp for H1.1 and 36 bp for H1.4. The heat capacity changes, ΔCp, for formation of the H1.1 and H1.4/CT-DNA complexes are -160 cal mol(-1) K(-1) and -192 cal mol(-1)K(-1) respectively. The large negative ΔCp values indicate the loss of water from the protein DNA interface in the complex.

Abstract  BACKGROUND/AIM: Calcium acetate/magnesium carbonate (CaMg) is a recent phosphate binder that has been shown to have protective cardiovascular (CV) effects in animal models. The aim of this study was to evaluate the relationship between CaMg therapy and CV risk markers like pulse pressure (PP), left ventricular mass index (LVMI) and valvular calcifications compared to sevelamer or no phosphate binder (NPB) therapy in chronic hemodialysis (HD) patients.

METHODS: We performed a 48-month prospective study in 138 HD patients under hemodiafiltration with a dialysate Mg concentration of 0.5 mmol/l. Patients underwent treatment with CaMg or sevelamer for at least 36 months or NPB therapy. Demographic, clinical, biochemical and echocardiographic parameters were evaluated at baseline and after a 48-month period.

RESULTS: At the end of the study, patients who had taken CaMg showed a significant reduction in PP (p < 0.001), LVMI (p = 0.003), aortic (p = 0.004) and mitral valve calcifications (p = 0.03) compared with NPB patients. Patients under CaMg showed a significant reduction of PP (p < 0.001), LVMI (p = 0.01) and aortic valve calcifications (p = 0.02) compared to sevelamer patients. In a multivariable analysis, CaMg therapy was negatively associated with progression of LVMI (p = 0.02) and aortic valve calcifications (p = 0.01). Patients under CaMg showed higher serum Mg levels (0.93 ± 0.14 mmol/l) compared to patients under sevelamer (0.87 ± 0.13) or NPB patients (0.82 ± 0.12; p < 0.001).

CONCLUSIONS: In prevalent HD patients, the use of CaMg over 48 months was associated with a reduction of PP and LVMI and with a stabilization of aortic valve calcifications. These protective and promising results of this new phosphate binder need to be confirmed in randomized controlled studies.

Abstract  BACKGROUND: Ocimum L. of family Lamiaceae is a well known genus for its ethnobotanical, medicinal and aromatic properties, which are attributed to innumerable phenylpropanoid and terpenoid compounds produced by the plant. To enrich genomic resources for understanding various pathways, de novo transcriptome sequencing of two important species, O. sanctum and O. basilicum, was carried out by Illumina paired-end sequencing.

RESULTS: The sequence assembly resulted in 69117 and 130043 transcripts with an average length of 1646 ± 1210.1 bp and 1363 ± 1139.3 bp for O. sanctum and O. basilicum, respectively. Out of the total transcripts, 59648 (86.30%) and 105470 (81.10%) from O. sanctum and O. basilicum, and respectively were annotated by uniprot blastx against Arabidopsis, rice and lamiaceae. KEGG analysis identified 501 and 952 transcripts from O. sanctum and O. basilicum, respectively, related to secondary metabolism with higher percentage of transcripts for biosynthesis of terpenoids in O. sanctum and phenylpropanoids in O. basilicum. Higher digital gene expression in O. basilicum was validated through qPCR and correlated to higher essential oil content and chromosome number (O. sanctum, 2n = 16; and O. basilicum, 2n = 48). Several CYP450 (26) and TF (40) families were identified having probable roles in primary and secondary metabolism. Also SSR and SNP markers were identified in the transcriptomes of both species with many SSRs linked to phenylpropanoid and terpenoid pathway genes.

CONCLUSION: This is the first report of a comparative transcriptome analysis of Ocimum species and can be utilized to characterize genes related to secondary metabolism, their regulation, and breeding special chemotypes with unique essential oil composition in Ocimum.

Abstract  OBJECTIVE: To characterize parasomnia behaviors on arousal from NREM sleep in Parkinson's Disease (PD) and Multiple System Atrophy (MSA).

METHODS: From 30 patients with PD, Dementia with Lewy Bodies/Dementia associated with PD, or MSA undergoing nocturnal video-polysomnography for presumed dream enactment behavior, we were able to select 2 PD and 2 MSA patients featuring NREM Parasomnia Behviors (NPBs). We identified episodes during which the subjects seemed to enact dreams or presumed dream-like mentation (NPB arousals) versus episodes with physiological movements (no-NPB arousals). A time-frequency analysis (Morlet Wavelet Transform) of the scalp EEG signals around each NPB and no- NPB arousal onset was performed, and the amplitudes of the spectral frequencies were compared between NPB and no-NPB arousals.

RESULTS: 19 NPBs were identified, 12 of which consisting of 'elementary' NPBs while 7 resembling confusional arousals. With quantitative EEG analysis, we found an amplitude reduction in the 5-6 Hz band 40 seconds before NPBs arousal as compared to no-NPB arousals at F4 and C4 derivations (p<0.01).

CONCLUSIONS: Many PD and MSA patients feature various NREM sleep-related behaviors, with clinical and electrophysiological differences and similarities with arousal parasomnias in the general population.

SIGNIFICANCE: This study help bring to attention an overlooked phenomenon in neurodegenerative diseases.

Abstract  Niemann-Pick C disease (NPCD) is a rare autosomal recessive neurovisceral disorder with a heterogeneous clinical presentation. Cholestan-3β,5α,6β-triol and 7-ketocholesterol have been proposed as biomarkers for the screening of NPCD. In this work, we assessed oxysterols levels in a cohort of Italian patients affected by NPCD and analyzed the obtained results in the context of the clinical, biochemical and molecular data. In addition, a group of patients affected by Niemann-Pick B disease (NPBD) were also analyzed. NPC patients presented levels of both oxysterols way above the cut off value, except for 5 siblings presenting the variant biochemical phenotype who displayed levels of 3β,5α,6β-triol below or just above the cut-off value; 2 of them presented also normal levels of 7-KC. Both oxysterols were extremely high in a patient presenting the neonatal systemic lethal phenotype. All NPB patients showed increased oxysterols levels. In conclusion, the reported LC-MS/MS assay provides a robust non-invasive screening tool for NPCD. However, false negative results can be obtained in patients expressing the variant biochemical phenotype. These data strengthen the concept that the results should always be interpreted in the context of the patients' clinical picture and filipin staining and/or genetic studies might still be undertaken in patients with normal levels of oxysterols if symptoms are highly suggestive of NPCD. Both oxysterols are significantly elevated in NPB patients; thus a differential diagnosis should always be performed in patients presenting isolated hepatosplenomegaly, a common clinical sign of both NPCD and NPBD.

Abstract  Neuropeptide B/W receptor-1 (NPBWR1) and NPBWR2 had been known as orphan receptors GPR7 and 8, respectively. Endogenous peptide ligands of these receptors, neuropeptide B (NPB) and neuropeptide W (NPW), were identified in 2002 and 2003 (Fujii et al., 2002; Brezillon et al., 2003; Tanaka et al., 2003). These peptides have been implicated in regulation of feeding behavior, energy homeostasis, neuroendocrine function, and modulating inflammatory pain. In addition, strong and discrete expression of their receptors in the extended amygdala and bed nucleus of the stria terminalis suggests a potential role in regulating stress responses, emotion, anxiety, and fear. Recent studies of NPB/NPW using both pharmacological and phenotypic analyses of genetically engineered mice as well as a human study support this hypothesis.

Abstract  BACKGROUND: Hypertension management strategies have traditionally focused on "tailored therapy" and "stepped-care" approaches. These tend to be costly and time consuming and often fail to achieve adequate blood pressure (BP) control. The TRIUMPH study aims to investigate the effectiveness, cost-effectiveness, and acceptability of early use of a 3-in-1 BP-lowering pill ("Triple Pill") compared with usual care for the management of hypertension.

METHODS: The prospective, open, randomized controlled clinical trial (n = 700) will compare Triple Pill-based strategy to usual care among individuals with persistent mild-to-moderate hypertension (systolic BP >140 mm Hg and/or diastolic BP >90 mm Hg, or systolic BP >130 mm Hg and/or diastolic BP >80 mm Hg in patients with diabetes or chronic kidney disease) on no or minimal drug therapy. The study will be conducted within approximately 20 hospital-based clinics in India. Participants will be randomized to the Triple Pill (initially strength 1-telmisartan 20 mg, amlodipine 2.5 mg, hydrochlorothiazide 6.25 mg, with the option of subsequent titration to strength 2-telmisartan 40 mg, amlodipine 5 mg, hydrochlorothiazide 12.5 mg) or continued usual care. Participants will be followed up for 6 months. The primary outcome is the proportion of participants achieving target BP at the end follow-up.

CONCLUSION: This study will determine whether early use of a low-dose triple combination therapy has the potential to address some of the challenges in hypertension control through earlier achievement of BP control, better adherence, and fewer adverse effects, in the context of less intensive clinical follow-up.

Abstract  PURPOSE: In male patients, the pudendal block was applied only in rare cases as a therapy of neuralgia of the pudendal nerve. We compared pudendal nerve block (NPB) and combined spinal-epidural anesthesia (CSE) in order to perform a pain-free high-dose-rate (HDR) brachytherapy in a former pilot study in 2010. Regarding this background, in the present study, we only performed the bilateral perineal infiltration of the pudendal nerve.

METHODS: In 25 patients (71.8 ± 4.18 years) suffering from a high-risk prostate carcinoma, we performed the HDR-brachytherapy with the NPB. The perioperative compatibility, the subjective feeling (German school marks principle 1-6), subjective pain (VAS 1-10) and the early postoperative course (mobility, complications) were examined.

RESULTS: All patients preferred the NPB. There was no change of anesthesia form necessary. The expense time of NPB was 10.68 ± 2.34 min. The hollow needles (mean 24, range 13-27) for the HDR-brachytherapy remained on average 79.92 ± 12.41 min. During and postoperative, pain feeling was between 1.4 ± 1.08 and 1.08 ± 1.00. A transurethral 22 French Foley catheter was left in place for 6 h. All patients felt the bladder catheter as annoying, but they considered postoperative mobility as more important as complete lack of pain. The subjective feeling was described as 2.28 ± 0.74. Any side effects or complications did not appear.

CONCLUSIONS: Bilateral NPB is a safe and effective analgesic option in HDR-brachytherapy and can replace CSE. It offers the advantage of almost no impaired mobility of the patient and can be performed by the urologist himself. Using transrectal ultrasound guidance, the method can be learned quickly.

Abstract  Chromatin remodeling complexes are known to modify chemical marks on histones or to induce conformational changes in the chromatin in order to regulate transcription. De novo dominant mutations in different members of the SWI/SNF chromatin remodeling complex have recently been described in individuals with Coffin-Siris (CSS) and Nicolaides-Baraitser (NCBRS) syndromes. Using a combination of whole-exome sequencing, NGS-based sequencing of 23 SWI/SNF complex genes, and molecular karyotyping in 46 previously undescribed individuals with CSS and NCBRS, we identified a de novo 1-bp deletion (c.677delG, p.Gly226Glufs*53) and a de novo missense mutation (c.914G>T, p.Cys305Phe) in PHF6 in two individuals diagnosed with CSS. PHF6 interacts with the nucleosome remodeling and deacetylation (NuRD) complex implicating dysfunction of a second chromatin remodeling complex in the pathogenesis of CSS-like phenotypes. Altogether, we identified mutations in 60% of the studied individuals (28/46), located in the genes ARID1A, ARID1B, SMARCB1, SMARCE1, SMARCA2, and PHF6. We show that mutations in ARID1B are the main cause of CSS, accounting for 76% of identified mutations. ARID1B and SMARCB1 mutations were also found in individuals with the initial diagnosis of NCBRS. These individuals apparently belong to a small subset who display an intermediate CSS/NCBRS phenotype. Our proposed genotype-phenotype correlations are important for molecular screening strategies.