Functional properties of atypical beta-adrenoceptors on the guinea pig duodenum.
In this study, we attempted to further characterize atypical beta-adrenoceptors on the guinea pig duodenum. (-)-Enantiomers of isoprenaline and noradrenaline were more potent than its (+)-enantiomers. The isomeric activity ratios ((+)/(-)) were less than those obtained in the guinea pig atria and trachea. The concentration-response curves to catecholamines ((-)-isoprenaline, (-)-noradrenaline and (-)-adrenaline), to the selective beta(3)-adrenoceptor agonist, BRL37344 ((R*, R*)-(+/-)-4-[2-[(2-(3-chlorophenyl)-2-hydroxyethyl)amino]propyl]phenoxyacetic acid sodium), and to the non-conventional partial beta(3)-adrenoceptor agonist, (+/-)-CGP12177A ((+/-)-[4-[3-[(1,1-dimethylethyl)amino]-2-hydroxypropoxy]-1,3-dihydro-2H-benzimidazol-2-one] hydrochloride), were resistant to blockade by (+/-)-pindobind, the beta-adrenoceptor alkylating agent. (-)-Noradrenaline and (-)-adrenaline were more potent than dopamine and (-)-phenylephrine, respectively. Selective beta(2)-adrenoceptor agonists possess agonistic activities at atypical beta-adrenoceptors. (+/-)-Propranolol and (+/-)-bupranolol had no agonistic effect, whereas (+/-)-alprenolol, (+/-)-pindolol, (+/-)-nadolol, (+/-)-CGP12177A and (+/-)-carteolol exhibited agonistic activities at atypical beta-adrenoceptors. These results suggest that pharmacological properties of atypical beta-adrenoceptors differ from those of conventional beta(1)- and beta(2)-adrenoceptors on the guinea pig.


Mass spectrometry-based identification of new anti-Ly and known antisynthetase autoantibodies.
To discover new and detect known antisynthetase autoantibodies (ASAs) through protein immunoprecipitation combined with gel-free liquid chromatography-tandem mass spectrometry (IP-MS). IP-MS was performed using sera of individuals showing features of antisynthetase syndrome (ASyS) without (n=5) and with (n=12) previously detected ASAs, and healthy controls (n=4). New candidate aminoacyl-tRNA-synthetase (ARS) autoantigens identified through unbiased IP-MS were confirmed by IP-western blot. A targeted IP-MS assay for various ASA specificities was developed and validated with sera of patients with known ASAs (n=16), disease controls (n=20) and healthy controls (n=25). The targeted IP-MS assay was applied in an additional cohort of patients with multiple ASyS features or isolated myositis without previously detected ASAs (n=26). Autoantibodies to cytoplasmic cysteinyl-tRNA-synthetase (CARS1) were identified by IP-MS and confirmed by western blot as a new ASA specificity, named anti-Ly, in the serum of a patient with ASyS features. Rare ASAs, such as anti-OJ, anti-Zo and anti-KS, and common ASAs could also be identified by IP-MS. A targeted IP-MS approach for ASA detection was developed and validated. Application of this method in an additional cohort identified an additional patient with anti-OJ autoantibodies that were missed by line and dot immunoassays. CARS1 is the dominant cognate ARS autoantigen of the newly discovered anti-Ly ASA specificity. Rare and common ASA specificities could be detected by both unbiased and targeted IP-MS. Unbiased and targeted IP-MS are promising methods for discovery and detection of autoantibodies, especially autoantibodies that target complex autoantigens.


A novel assay to screen siRNA libraries identifies protein kinases required for chromosome transmission.
One of the hallmarks of cancer is chromosome instability (CIN), which leads to aneuploidy, translocations, and other chromosome aberrations. However, in the vast majority of human tumors the molecular basis of CIN remains unknown, partly because not all genes controlling chromosome transmission have yet been identified. To address this question, we developed an experimental high-throughput imaging (HTI) siRNA assay that allows the identification of novel CIN genes. Our method uses a human artificial chromosome (HAC) expressing the GFP transgene. When this assay was applied to screen an siRNA library of protein kinases, we identified PINK1, TRIO, IRAK1, PNCK, and TAOK1 as potential novel genes whose knockdown induces various mitotic abnormalities and results in chromosome loss. The HAC-based assay can be applied for screening different siRNA libraries (cell cycle regulation, DNA damage response, epigenetics, and transcription factors) to identify additional genes involved in CIN. Identification of the complete spectrum of CIN genes will reveal new insights into mechanisms of chromosome segregation and may expedite the development of novel therapeutic strategies to target the CIN phenotype in cancer cells.


Effect of valsartan added to background ACE inhibitor therapy in patients with heart failure: results from Val-HeFT.
To investigate the effect of valsartan in the Valsartan-Heart Failure Trial (Val-HeFT) when added to angiotensin-converting enzyme inhibitor (ACEi) alone in patients with heart failure (HF). Subjects in Val-HeFT receiving ACEi but not beta-blocker at baseline were analysed; 1532 were assigned to valsartan and 1502 assigned to placebo. Primary outcome events (all-cause mortality, hospitalisation for adjudicated heart failure, sudden death with resuscitation and need for >4 h of parenteral therapy for worsening heart failure) were monitored. Mortality was not affected by valsartan but morbidity endpoints were significantly reduced (36.3% in placebo, 31.0% in valsartan, p=0.002) in patients receiving an ACEi but no beta-blocker. Quality of life (QOL) was significantly improved, ejection fraction (EF) significantly increased, left ventricular (LV) diameter significantly reduced and plasma B-type natriuretic peptide, norepinephrine and aldosterone levels significantly reduced with valsartan compared to placebo. The morbidity benefit was significant in patients on ACEi doses below the median (22% reduction, p=0.003) and not statistically significant in those receiving ACEi doses above the median (14% reduction, p=0.143). Valsartan reduces heart failure hospitalisations and slows LV remodelling in patients treated with an ACEi in the absence of beta-blockade, particularly in those on lower doses of ACEi.


Relevance of the protein macrodipole in the membrane-binding process. Interactions of fatty-acid binding proteins with cationic lipid membranes.
The fatty acid-binding proteins L-BABP and Rep1-NCXSQ bind to anionic lipid membranes by electrostatic interactions. According to Molecular Dynamics (MD) simulations, the interaction of the protein macrodipole with the membrane electric field is a driving force for protein binding and orientation in the interface. To further explore this hypothesis, we studied the interactions of these proteins with cationic lipid membranes. As in the case of anionic lipid membranes, we found that both proteins, carrying a negative as well as a positive net charge, were bound to the positively charged membrane. Their major axis, those connecting the bottom of the β-barrel with the α-helix portal domain, were rotated about 180 degrees as compared with their orientations in the anionic lipid membranes. Fourier transform infrared (FTIR) spectroscopy of the proteins showed that the positively charged membranes were also able to induce conformational changes with a reduction of the β-strand proportion and an increase in α-helix secondary structure. Fatty acid-binding proteins (FABPs) are involved in several cell processes, such as maintaining lipid homeostasis in cells. They transport hydrophobic molecules in aqueous medium and deliver them into lipid membranes. Therefore, the interfacial orientation and conformation, both shown herein to be electrostatically determined, have a strong correlation with the specific mechanism by which each particular FABP exerts its biological function.


Case series: Bladder clot evacuation using a prostate morcellation device.
We sought to provide a technical update on the use of a prostate morcellator device (PMD) to manage organized blood clots of the bladder following laser prostatectomy. Herein, we describe our experience in using the Wolf Piranha morcellator in managing organized bladder blood clots supplemented with a retrospective chart review of the patients in whom this procedure was performed. Six patients, all male with a mean age of 75 ± 8.9 years, had organized bladder clots following either holmium laser enucleation or photoselective vaporization of the prostate managed with a PMD. Clots were recognized based on hematuria or urinary retention a median of 3.5 days following the aforementioned procedures. Initial management was attempted with more conservative measures, including a three-way Foley catheter, followed by cystoscopy with an Ellik evacuator, or a glass Tommey syringe. Morcellation times were a mean of 10.2 ± 6.15 minutes (range 2-18). This technique was able to manage clots that were an average of 173.3 ± 115.9 cc in size. The procedure was well-tolerated. No patients experienced intraoperative or morcellator-related complications. Benign prostatic hypertrophy frequently requires surgical endoscopic management and can be complicated by hematuria and bladder blood clot formation. When these clots become organized, this can lead to urinary retention and the required management, evacuation, may be difficult. The use of a Wolf Piranha PMD is a safe, well-tolerated, and effective in evacuating organized blood clots of the bladder.


Glucocorticoid receptor polymorphisms and haplotypes and their expression in health and disease.
Cortisol is involved in many physiological processes, including immunosuppressive and anti-inflammatory actions, and therefore cortisol and its synthetic analogs are widely used to treat a large number of diseases. In glucocorticoid treatment, a large variability of clinical responses is observed. This variability may, in part, be ascribed to genetic variation in the glucocorticoid receptor (GR) gene. In this review we present a catalogue of the various single nucleotide polymorphisms (SNPs) in the glucocorticoid receptor gene and their consequences for human health and disease. Many different GR SNP association studies have been described. However, most studies come down to only a few SNPs reported with different annotations. In this review we clarified these different annotations to uniform names. Most associations between GR SNPs and phenotype have been found in body composition, metabolism, the cardiovascular system, the immune system and psychiatric illnesses. However, many associations have not been replicated (yet), and future replication studies and meta-analyses are needed. There is a substantial body of evidence for GR SNPs to have effects on clinical phenotype. However, as most SNP frequencies are low and their variation is within the range of the general population, the impact of a single SNP for health and disease in the general population is probably modest. However, in-depth studying of the molecular mechanisms of repeatedly observed clinical associations could lead to new possibilities for drug development. In particular the development of selective glucocorticoid receptor modulators holds promise.


[Therapeutic drug monitoring of 6-thioguanine nucleotides in paediatric acute lymphoblastic leukaemia: interest and limits].
6-mercaptopurine, a key drug for the treatment of acute lymphoblastic leukaemia in children, is a prodrug metabolized into 6-thioguanine (6-TGN) which are the active compounds and into methylated metabolites, primary by thiopurine S-methyltransferase enzyme (TPMT). This enzyme displays important inter subject variability linked to a genetic polymorphism: when treated with standard doses of thiopurine, TPMT-deficient and heterozygous patients are at great risk for developing severe and potentially life-threatening toxicity (hematopoietic, hepatic, mucositis...) but show a better survival rate while patients with high TPMT activity (wild type) present lower peripheral red blood cells 6-TGN concentrations and a higher risk of leukemia relapse. Genotyping remains crucial before 6-MP administration at diagnosis to identify patients with homozygous mutant TPMT genotype and therefore prevent severe and life-threatening toxicity, and to individualize therapy according to TMPT genotype. Follow-up of ALL treatment should preferentially be based on repeated determinations of intracellular active metabolites (6-thioguanine nucleotides) and methylated metabolites in addition to haematological surveillance.


Analysis of Diagnostic Modalities in Hospital-admitted Patients Evaluated for COVID-19.
To assess the diagnostic performance of reverse transcriptase polymerase chain reaction (RT-PCR), low-dose chest computed tomography (CT), and serological testing, alone and in combinations, as well as routine inflammatory markers in patients evaluated for COVID-19 during the first wave in early 2020. We retrospectively analyzed data of all patients who were admitted to the emergency department due to fever and/or respiratory symptoms. CT scans were rated using the COVID-19 Reporting and Data System (CO-RADS) suspicion score. True disease status (COVID-19 - positive vs. negative) was adjudicated by two independent clinicians. Receiver-operating characteristic curves and areas under the curves were calculated for inflammatory markers. Sensitivities and specificities were calculated for RT-PCR, CT, and serology alone, as well as the combinations of RT-PCR+CT, RT-PCR+serology, CT+serology, and all three modalities. Of 221 patients with a median age of 72 years, 113 were classified as COVID-19 positive. Among 180 patients from which data on CT and RT-PCR were available, RT-PCR had the highest sensitivity to detect COVID-19 (0.87; 95%CI=0.78-0.93). Notably, the addition of CT in the analysis increased sensitivity to 0.89 (95%CI=0.8-0.94), but lowered specificity from 1 (95%CI=0.96-1) to 0.9 (95%CI=0.83-0.95). The combination of RT-PCR, CT and serology (n=60 patients with complete dataset) yielded a sensitivity of 0.83 (95%CI=0.61-0.94) and specificity of 0.86 (95%CI=0.72-0.93). RT-PCR was the best single test in patients evaluated for COVID-19. Conversely, the routine performance of chest CT adds little sensitivity and decreases specificity.


The environmental impact of recombinant bovine somatotropin (rbST) use in dairy production.
The environmental impact of using recombinant bovine somatotropin (rbST) in dairy production was examined on an individual cow, industry-scale adoption, and overall production system basis. An average 2006 U.S. milk yield of 28.9 kg per day was used, with a daily response to rbST supplementation of 4.5 kg per cow. Rations were formulated and both resource inputs (feedstuffs, fertilizers, and fuels) and waste outputs (nutrient excretion and greenhouse gas emissions) calculated. The wider environmental impact of production systems was assessed via acidification (AP), eutrophication (EP), and global warming (GWP) potentials. From a producer perspective, rbST supplementation improved individual cow production, with reductions in nutrient input and waste output per unit of milk produced. From an industry perspective, supplementing one million cows with rbST reduced feedstuff and water use, cropland area, N and P excretion, greenhouse gas emissions, and fossil fuel use compared with an equivalent milk production from unsupplemented cows. Meeting future U.S. milk requirements from cows supplemented with rbST conferred the lowest AP, EP, and GWP, with intermediate values for conventional management and the highest environmental impact resulting from organic production. Overall, rbST appears to represent a valuable management tool for use in dairy production to improve productive efficiency and to have less negative effects on the environment than conventional dairying.


Norfloxacin derivatives as DNA gyrase and urease inhibitors: synthesis, biological evaluation and molecular docking.
Background: DNA gyrase and urease enzymes are important targets for the treatment of gastroenteritis, appendicitis, tuberculosis, urinary tract infections and Crohn's disease. Materials & methods: Esterification of norfloxacin was performed to enhance DNA gyrase and urease enzyme inhibition potential. Structure elucidation and chemical characterization were done through spectral (1H NMR, Fourier transform IR, 13C NMR) and carbon, hydrogen, nitrogen and sulfur analysis along with molecular docking. Results & conclusion: The majority of derivatives exhibited significant results but the 3e derivative showed maximum bactericidal, DPPH scavenging (96%), DNA gyrase and urease enzyme inhibitory activity with IC50 of 0.15 ± 0.24 and 1.14 ± 0.11 μM respectively which was further supported by molecular docking studies. So, the active derivatives can serve as a lead compound for the treatment of various pathological conditions.


Studies of native plants in Taiwan for lectin content.
The lectin content of some native plants in Taiwan was studied. The majority of the plants possessing lectin belonged to Leguminosae. Five lectins which are readily available were further studied for their hemagglutination titers, specificity, susceptibility to heat and proteolytic enzymes. The lectins of Ricinus communis had hemagglutination titer of 1:512 which was the highest among the five tested. All lectins were heat labile, destroyed by heating at 100 degrees C for 30 min, but resistant to trypsin and pepsin. The resistance was not due to the presence of any inhibitor in the crude extract, but possibly due to the special configuration of the lectin molecule or other unknow reasons. Among the five lectins only the lectin of Phaseolus vulgaris could not be inhibited by any of the carbohydrates tested. It seems that the lectin of Phaseolus vulgaris is nonspecific, or not specific for ABO blood group, or specific for other blood groups. The inhibiting activity of carbohydrates seems to be specific. The configuration of carbohydrates at carbon atom 2 and/or 4, the type of linkage (alpha or beta), or the carbon number where the linkage is formed seem to be important to the potency of carbohydrates to inhibit the activity of lectin.


High-resolution X-ray CT for 3D petrography of ferruginous sandstone for an investigation of building stone decay.
Diestian ferruginous sandstone has been used as the dominant building stone for monuments in the Hageland, a natural landscape in east-central Belgium. Like all rocks, this stone type is sensitive to weathering. Case hardening was observed in combination with blackening of the exterior parts of the dressed stones. To determine the 3D petrography and to identify the structural differences between the exterior and interior parts, X-ray computed tomography was used in combination with more traditional research techniques like optical microscopy and scanning electron microscopy. The 3D characterization of the ferruginous sandstone was performed with a high-resolution X-ray CT scanner (www.ugct.ugent.be) in combination with the flexible 3D analysis software Morpho+, which provides the necessary petrophysical parameters of the scanned samples in 3D. Besides providing the required 3D parameters like porosity, pore-size distribution, grain size, grain orientation, and surface analysis, the results of the 3D analysis can also be visualized, which enables to understand and interpret the analysis results in a straightforward way. The complementarities between high-quality X-ray CT images and flexible 3D software and its relation with the more traditional microscopical research techniques are opening up new gateways in the study of weathering processes of natural building stones.


Perception of facial esthetics: a comparison of similar class II cases treated with attempted growth modification or later orthognathic surgery.
In recent years, there has been considerable interest in the esthetic outcomes of various types of Class II treatment. In this study, it was the authors' intent to determine the esthetic outcomes for 60 Class II division 1 patients: 28 patients treated during the active growth phase with an activator and fixed appliances and 32 patients treated at the completion of growth with fixed appliances and by orthognathic surgery. Using a visual analogue scale, a mixed panel of 14 judges scored the pre- and posttreatment attractiveness of these patients from frontal and lateral facial photographs. Statistical analysis by two-sample t-tests indicated that, on average, esthetic scores improved with treatment, regardless of the treatment modality. There was, however, considerable individual variation in the degree of improvement, even to the point that there was a decline in esthetics for some patients. Despite somewhat different modes of treatment, it was found that neither the average pre- and posttreatment esthetic scores nor the change in esthetic score with treatment was significantly different for the two groups. Although clinical planning decisions should still be made on an individual basis, the findings of this study suggest that the perceived esthetic outcome in many Class II division 1 patients may well be just as favorable, regardless of whether they are managed early during the growth phase or later, at the completion of growth by orthognathic surgery.


Turbulence-induced channel crosstalk in an orbital angular momentum-multiplexed free-space optical link.
A multichannel free-space optical (FSO) communication system based on orbital angular momentum (OAM)-carrying beams is studied. We numerically analyze the effects of atmospheric turbulence on the system and find that turbulence induces attenuation and crosstalk among channels. Based on a model in which the constituent channels are binary symmetric and crosstalk is a Gaussian noise source, we find optimal sets of OAM states at each turbulence condition studied and determine the aggregate capacity of the multichannel system at those conditions. OAM-multiplexed FSO systems that operate in the weak turbulence regime are found to offer good performance. We verify that the aggregate capacity decreases as the turbulence increases. A per-channel bit-error rate evaluation is presented to show the uneven effects of crosstalk on the constituent channels.


Performance of the M11 and M11-L density functionals for calculations of electronic excitation energies by adiabatic time-dependent density functional theory.
Adiabatic time-dependent density functional theory is a powerful method for calculating electronic excitation energies of complex systems, but the quality of the results depends on the choice of approximate density functional. In this article we test two promising new density functionals, M11 and M11-L, against databases of 214 diverse electronic excitation energies, and we compare the results to those for 16 other density functionals of various kinds and to time-dependent Hartree-Fock. Charge transfer excitations are well known to be the hardest challenge for TDDFT. M11 is a long-range-corrected hybrid meta-GGA, and it shows better performance for charge transfer excitations than any of the other functionals except M06-HF, which is a specialized functional that does not do well for valence excitations. Several other long-range-corrected hybrid functionals also do well, and we especially recommend M11, ωB97X, and M06-2X for general spectroscopic applications because they do exceptionally well on ground-state properties as well as excitation energies. Local functionals are preferred for many applications to extended systems because of their significant cost advantage for large systems. M11-L is a dual-range local functional and-unlike all previous local functionals-it has good performance for Rydberg states as well as for valence states. Thus it is highly recommended for excitation energy calculations on extended systems.


Neuroprotective effect of nerolidol against neuroinflammation and oxidative stress induced by rotenone.
Parkinson disease (PD) is a movement disorder affecting 1 % of people over the age of 60. The etiology of the disease is unknown; however, accumulating evidence suggests that mitochondrial defects, oxidative stress, and neuroinflammation play important roles in developing the disease. Current medications for PD can only improve its symptoms, but are unable to halt its progressive nature. Although many therapeutic approaches are available, new drugs are urgently needed for the treatment of PD. Thus, the present study was undertaken to investigate the neuroprotective potential of nerolidol, a sesquiterpene alcohol, on a rotenone-induced experimental model of PD, where male Wistar rats intraperitoneally received rotenone (ROT) at a dose of 2.5 mg/kg of body weight once daily for 4 weeks. Nerolidol, which has antioxidant and anti-inflammatory properties, was injected intraperitoneally at 50 mg/kg of body weight, once daily for 4 weeks, and at 30 min prior to ROT administration. ROT administration significantly reduced the activities of antioxidant enzymes superoxide dismutase (SOD) and catalase (CAT), and the level of the antioxidant tripeptide glutathione (GSH). Moreover, ROT increased the levels of the lipid peroxidation product malondialdehyde (MDA), proinflammatory cytokines (IL-1β, IL-6, and TNF-α), and inflammatory mediators (COX-2 and iNOS) in rat brain tissues. Immunostaining of brain tissue sections revealed a significant increase in the number of activated astrocytes (GFAP) and microglia (Iba-1), along with the concomitant loss of dopamine (DA) neurons in the substantia nigra pars compacta and dopaminergic nerve fibers in the striatum of ROT-treated rats. As expected, nerolidol supplementation to ROT-injected rats significantly increased the level of SOD, CAT, and GSH, and decreased the level of MDA. Nerolidol also inhibited the release of proinflammatory cytokines and inflammatory mediators. Finally, nerolidol treatment prevented ROT-induced glial cell activation and the loss of dopaminergic neurons and nerve fibers, and ultimately attenuated ROT-induced dopaminergic neurodegeneration. Our findings are the first to show that the neuroprotective effect of nerolidol is mediated through its anti-oxidant and anti-inflammatory activities, which strongly supports its therapeutic potential for the treatment of PD.


Residue proximity information and protein model discrimination using saturation-suppressor mutagenesis.
Identification of residue-residue contacts from primary sequence can be used to guide protein structure prediction. Using Escherichia coli CcdB as the test case, we describe an experimental method termed saturation-suppressor mutagenesis to acquire residue contact information. In this methodology, for each of five inactive CcdB mutants, exhaustive screens for suppressors were performed. Proximal suppressors were accurately discriminated from distal suppressors based on their phenotypes when present as single mutants. Experimentally identified putative proximal pairs formed spatial constraints to recover >98% of native-like models of CcdB from a decoy dataset. Suppressor methodology was also applied to the integral membrane protein, diacylglycerol kinase A where the structures determined by X-ray crystallography and NMR were significantly different. Suppressor as well as sequence co-variation data clearly point to the X-ray structure being the functional one adopted in vivo. The methodology is applicable to any macromolecular system for which a convenient phenotypic assay exists.


Diagnosis of Acute Gastroenteritis with Immunochromatography and Effectiveness of Rotavirus Vaccine in a Japanese Clinic.
Despite the well known effectiveness of two licensed live attenuated oral rotavirus (RV)-vaccines, Rotarix and RotaTeq, constant monitoring of vaccine effectiveness (VE) is essential considering the evolving power and reassortment capability of RVs. In this study, we detected RV, norovirus (NV) and adenovirus (AV) infections using immunochromatography (IC)-based kits in children with acute gastroenteritis (AGE) who attended a pediatric clinic in Kiryu city, Gunma, Japan during June, 2014-September, 2018. VEs were determined using a test-negative study design. Among 1658 AGE-children, RV, NV and AV were detected in 96 (5.8 %), 146 (8.8 %) and 46 (2.8 %) children, respectively. Interestingly, the distributions of infections were found to be associated with age and sex. Namely, RV infections were significantly higher in female (P=0.02) and in the 19-30 month age group children, while NV and AV infections predominated in the 13-24 month and 7-18 month age groups, respectively. The disease severity for RV and NV infections remained similar and significantly higher than that of AV infections. The VE of RV-vaccines was 49.8 % (95 % CI: 22.7 to 67.3 %) against all RV infections, which was increased up to 67.2 % (95 % CI: 35.3 to 83.4 %) against severe RV infections. RV-vaccinated children experienced less severe symptoms in RV-infections while non-RV AGE remained less serious for both RV-vaccinated and unvaccinated children. Finally, the prevalence of RV infection remained minimized (≤5.4 %) in this population since 2015. Thus, this study provided important information on distribution of major AGEs in young children and exhibited the effective role of RV vaccines in post-vaccine era.


Neural tube defects, their implications and solutions in Muslim society.
Neural Tube Defects (NTDs) are serious congenital abnormalities and most of them are incompatible with life. The extremely debilitating quality of life, if one survives, calls for actions to prevent such sufferings. Experts agree on the role of Folic Acid in primary prevention of NTDs, yet, despite best efforts, the use of Folic Acid has reduced NTDs by only 50%. These cases too can be prevented by employing secondary preventive measures. These involve timely interruption of pregnancy -- a decision which, in addition to a medical judgment, is based on ethics, social, cultural and Muslim religious value systems in Pakistan. Indeed, it is a complex issue but empathic understanding and strong co-ordination, once established between different disciplines, can help parents to decide and opt for necessary secondary prevention by interruption of malformed foetus within the given time frame mandated by medical and religious authorities.


Changes in phenotypic gene expression in rat mandibular condylar cartilage cells during long-term culture.
Gene expression patterns have been investigated in prolonged cultures of rat mandibular condylar cartilage (MCC) cells to examine the possibility of culture-induced phenotypic changes. MCC cells were isolated from newborn rats and grown in the presence of 10% fetal bovine serum (FBS) and basic fibroblast growth factor (bFGF). MCC cells were passaged and cultured in the presence of 10% FBS and bFGF until confluent. After confluence, the medium was changed to that supplemented with 10% FBS, ascorbate, and beta-glycerophosphate (day 1). Mineralization and gene expression of MCC cells have been investigated. Mineralization, visualized by staining with Alizarin red, was observed to begin at day 13 in culture, and increased up to day 22 in culture, which was the length of this study. Type II collagen and aggrecan mRNAs were highly expressed at the start of culture (day 1-4) and decreased in a time-dependent manner. Type I collagen, alkaline phosphatase, and osteopontin mRNAs expressed biphasic patterns that peaked at the start of culture and the beginning of mineralization (day 13-16). Osteonectin mRNA was expressed throughout the culture period. Osteocalcin mRNA was expressed before the beginning of mineralization peaking at day 7. These observations suggest that the gene expression patterns of MCC cells can be categorized into two different periods in prolonged culture: maturation (day 1-10) and mineralization (day 13-22). The day culture system of MCC represents a new model system in which the differentiation of embryonic MCC cells can be examined.


Disturbance of calcium metabolism by anticonvulsant drugs.
A survey of calcium metabolism in epileptic patients in a residential centre showed a subnormal serum calcium level in 22.5% of patients and a raised alkaline phosphatase in 29%. Hypocalcaemia was related to high dosage of anticonvulsant drugs, to multiple drug therapy, and to the use of individual anticonvulsant drugs in the following order, with decreasing order of importance: pheneturide, primidone, phenytoin, phenobarbitone. Subnormal serum calcium levels occurred more commonly in patients with a raised liver alkaline phosphatase isoenzyme than in those whose phosphatase was mainly of bone origin.Preliminary results of treatment with calciferol suggested that the disturbance of calcium metabolism was the result of vitamin D deficiency. It is possible that anticonvulsant drugs accelerate the breakdown of vitamin D by liver enzyme induction.


Pediatric Metabolic and Bariatric Surgery: Evidence, Barriers, and Best Practices.
Severe obesity among youth is an "epidemic within an epidemic" and portends a shortened life expectancy for today's children compared with those of their parents' generation. Severe obesity has outpaced less severe forms of childhood obesity in prevalence, and it disproportionately affects adolescents. Emerging evidence has linked severe obesity to the development and progression of multiple comorbid states, including increased cardiometabolic risk resulting in end-organ damage in adulthood. Lifestyle modification treatment has achieved moderate short-term success among young children and those with less severe forms of obesity, but no studies to date demonstrate significant and durable weight loss among youth with severe obesity. Metabolic and bariatric surgery has emerged as an important treatment for adults with severe obesity and, more recently, has been shown to be a safe and effective strategy for groups of youth with severe obesity. However, current data suggest that youth with severe obesity may not have adequate access to metabolic and bariatric surgery, especially among underserved populations. This report outlines the current evidence regarding adolescent bariatric surgery, provides recommendations for practitioners and policy makers, and serves as a companion to an accompanying technical report, "Metabolic and Bariatric Surgery for Pediatric Patients With Severe Obesity," which provides details and supporting evidence.


Ocular motor measures of cognitive dysfunction in multiple sclerosis I: inhibitory control.
Our ability to control and inhibit behaviours that are inappropriate, unsafe, or no longer required is crucial for functioning successfully in complex environments. Here, we investigated whether a series of ocular motor (OM) inhibition tasks could dissociate deficits in patients with multiple sclerosis (MS), including patients with only a probable diagnosis (clinically isolated syndrome: CIS), from healthy individuals as well as a function of increasing disease duration. 25 patients with CIS, 25 early clinically definite MS patients (CDMS: ≤7 years of diagnosis), 24 late CDMS patients (>7 years from diagnosis), and 25 healthy controls participated. All participants completed a series of classic OM inhibition tasks [antisaccade (AS) task, memory-guided (MG) task, endogenous cue task], and a neuropsychological inhibition task [paced auditory serial addition test (PASAT)]. Clinical disability was characterised in CDMS patients using the Expanded Disability Severity Scale (EDSS). OM (latency and error) and PASAT performance were compared between patient groups and controls, as well as a function of disease duration. For CDMS patients only, results were correlated with EDSS score. All patient groups made more errors than controls on all OM tasks; error rate did not increase with increasing disease duration. In contrast, saccade latency (MG and endogenous cue tasks) was found to worsen with increasing disease duration. PASAT performance did not discriminate patient groups or disease duration. The EDSS did not correlate with any measure. These OM measures appear to dissociate deficit between patients at different disease durations. This suggests their utility as a measure of progression from the earliest inception of the disease.


Oleaster oil (Olea europaea sylvestris) effects on the efficiency of the reproductive system of diet-induced obese male Wistar rats.
Obesity is the result of oxidative stress (OS), which appears to be linked to male infertility. Oleaster oil has multiple therapeutic virtues due to its richness in natural bioactive compounds. This study aimed to evaluate the effect of Algerian oleaster oil on the reproductive system effectiveness of diet-induced obese male Wistar rats. The oil was extracted with an oleodoseur from the oleaster fruits. Rats were fed for 28 weeks two diets: a normal diet (NC group, n=8) and a high-fat, high-sucrose diet (HFHS group, n=32). The obese rats in the HFHS group were then split into three groups at random: the HFHS group, which continued to receive the HFHS diet; the HFHS L-OO group; and the HFHS H-OO group, which received for 6 weeks, respectively, in combination with the HFHS diet, oleaster oil at 1.5 and 3 mL/100 g body weight (BW). Then, the animals were sacrificed. Semen and testes were removed for semen analysis, OS, and histopathological evaluation. The oleaster oil diet induces an improvement in sperm count and viability and a decrease in the percentage of fragmented sperm DNA compared to the group fed the HFHS diet, preserving the testes against OS with a highly significant decrease in testicular MDA. This beneficial effect of oleaster oil on the reproductive efficiency of rats fed the HFHS diet was dose-dependent. The results showed that the oil decreased body weight, prevented OS and improved male reproductive indicators in HFHS-fed animals. The potential positive effects of oleaster oil on fertility biomarkers may be attributed to its ability to mitigate OS.


Rapid Identification of Tropilaelaps Mite (Mesostigmata: Laelapidae) Species Using a COI Barcode-HRM.
Tropilaelaps mite (Mesostigmata: Laelapidae) is an ectoparasite of bees present, to date, only on the Asian continent. In the context of the threat of Tropilaelaps's introduction into new regions, accurate, rapid, and sensitive detection of the Tropilaelaps spp. is essential. In the present study, we developed a novel molecular method for bee mite's identification, which consists of a new real-time PCR method. A high-resolution melting analysis (HRM) was then performed on the amplified products to differentiate the species. PCR amplification was applied on the cytochrome c oxidase subunit I gene (580 bp). Short fragments from the most variable regions of this gene were identified in silico to amplify and discriminate among the four Tropilaelaps species. Four reference plasmids were synthesized to characterize species by well-distinguished melting curves. The method was then validated in terms of its specificity and sensitivity using a panel of 12 specimens. The results showed that an HRM method can be applied for the intended objective: for rapid and simultaneous identification of Tropilaelaps species. To our knowledge, this study reports the first direct HRM assay developed for the genome of a bee mite, specific for Tropilaelaps species. This COI barcode-HRM technique could be a promising tool for mite species identification.


[Channel-forming protein from a Na, K-ATPase preparation from porcine renal medulla].
Protein inducing formation of the monovalent cation selective channels on bilayer lipid membrane (BLM) was isolated by ethanol extraction from the great subunits of Na+, K+ ATPase or pork kidney microsomes. Conductance of the single channels in the presence of 2 microgram/ml of protein, 0.016 A KC1 and 0.08 M NaCl is equal to 40 p. The membrane potential of a ten-fold potassium ion gradient is close to a theoretical one. Ouabain (5 X 10(-5) M-1 X 10(4) M) inhibits the formation of the channels. KCl (0.03-0.08 M) eliminates the ouabain effect.


Exposure of beta L-crystallin to oxidizing free radicals enhances its susceptibility to transglutaminase activity.
1. The effects of free radicals on the capacity of beta L-crystallin to act as a substrate for the enzyme transglutaminase were investigated. 2. beta L-Crystallin was exposed to defined radical species that were generated radiolytically, and transglutaminase activity, using the modified protein as substrate, was subsequently measured by monitoring the incorporation of [14C]putrescine. 3. Exposure of beta L-crystallin to hydroxyl radicals, thymine peroxyl radicals and acetone peroxyl radicals at concentrations of up to 135 microM increased the capacity of the protein to incorporate putrescine. With higher concentrations of these radicals this capacity of beta L-crystallin to act as a transglutaminase substrate declined to control levels or lower. 4. Superoxide radicals were inactive in this regard; hydroperoxyl radicals were active only at high concentrations. 5. It has previously been suggested that changes in the crystallins that occur during aging and with cataract may be due to oxidative reactions and to transglutaminase activity. This study suggests that these phenomena may be considered together rather than separately.


Selected mitochondrial DNA landscapes activate the SIRT3 axis of the UPRmt to promote metastasis.
By causing mitochondrial DNA (mtDNA) mutations and oxidation of mitochondrial proteins, reactive oxygen species (ROS) leads to perturbations in mitochondrial proteostasis. Several studies have linked mtDNA mutations to metastasis of cancer cells but the nature of the mtDNA species involved remains unclear. Our data suggests that no common mtDNA mutation identifies metastatic cells; rather the metastatic potential of several ROS-generating mutations is largely determined by their mtDNA genomic landscapes, which can act either as an enhancer or repressor of metastasis. However, mtDNA landscapes of all metastatic cells are characterized by activation of the SIRT/FOXO/SOD2 axis of the mitochondrial unfolded protein response (UPRmt). The UPRmt promotes a complex transcription program ultimately increasing mitochondrial integrity and fitness in response to oxidative proteotoxic stress. Using SOD2 as a surrogate marker of the UPRmt, we found that in primary breast cancers, SOD2 is significantly increased in metastatic lesions. We propose that the ability of selected mtDNA species to activate the UPRmt is a process that is exploited by cancer cells to maintain mitochondrial fitness and facilitate metastasis.


Effect of acetaminophen on the leukocyte-labeling efficiency of indium oxine In 111.
The effect of acetaminophen on the labeling efficiency of leukocytes with indium oxine In 111 was studied. A blood sample was obtained from eight healthy men before and after they received acetaminophen 650 mg every four hours for 24 hours. After dividing the plasma from each sample into three portions, leukocytes were separated and labeled with indium oxine In 111. In an in vitro study, 200 ml of blood was obtained from one of the men, and the plasma was separated into four portions. Acetaminophen in 95% ethanol was added to three of the plasma fractions to produce acetaminophen concentrations of 4, 20, and 100 micrograms/ml; ethanol was added to the fourth fraction as a control. Each plasma fraction was then subdivided into three aliquots, and leukocytes were labeled as in the in vivo study. Mean leukocyte labeling efficiencies in both studies were calculated from the ratios of leukocyte radioactivity to initial radioactivity in the samples, expressed as percentages. Leukocyte labeling efficiencies before acetaminophen administration ranged from 79 to 85%; after administration, labeling efficiencies ranged from 70 to 87%. No significant differences in mean labeling efficiency before and after acetaminophen administration were noted in any of the subjects. Leukocyte labeling efficiencies in all in vitro plasma fractions were reduced, ranging from 54 to 63%, but no significant differences in labeling efficiency between any of the plasma fractions were found. Using the labeling procedures in this study, exposure of leukocytes from healthy men to acetaminophen in vivo or in vitro does not affect labeling efficiency with indium oxine In 111.


The effect of the Fanconi anemia polypeptide, FAC, upon p53 induction and G2 checkpoint regulation.
Fanconi anemia (FA) is an autosomal recessive disease marked by developmental defects, bone marrow failure, and cancer susceptibility. FA cells are hypersensitive to DNA cross-linking and alkylating agents and accumulate in the G2 phase of the cell cycle in response to these agents. FA cells also display genomic instability, suggesting a possible defect in the p53 pathway. To test the effect of heterologous expression of FAC cDNA on drug-induced cytotoxicity, G2 accumulation, and p53 induction in FA cells, we compared two isogenic FA cell lines: HSC536N (mock), a FA type C cell line sensitive to mitomycin C (MMC), and the same cell line transfected (corrected) with wild-type FAC cDNA (HSC536N [+FAC]). HSC536N (+FAC) cells showed a 30-fold increase in resistance to MMC concentration. Similarly, increases in resistance were observed following exposure to cisplatin, carboplatin, and cyclophosphamide. In addition, HSC536N (+FAC) cells showed a twofold lower G2 accumulation following MMC treatment. To analyze the possible interaction of FAC with the p53 pathway, we analyzed p53 induction in mock and corrected cell lines following exposure to MMC. HSC536N (mock) cells induced p53 at lower MMC concentrations than HSC536N (corrected). Caffeine, a known G2 checkpoint inhibitor, not only inhibited G2 accumulation seen in both cell lines but also caused the resistant HSC536N (+FAC) to become as sensitive to MMC as HSC536N (mock) cell line. We conclude that the FAC protein has a specific cytoprotective effect and may function as a cell cycle regulator of the G2 phase of the cell cycle.


Co-transplantation of human fetal thymus, bone and CD34(+) cells into young adult immunodeficient NOD/SCID IL2Rγ(null) mice optimizes humanized mice that mount adaptive antibody responses.
Both the thymus (T) and bone (B) are necessary hematopoietic niches in adult humans. We previously showed that co-transplantation of human fetal T and B tissues into neonatal immunodeficient NOD/SCID IL2Rγ(null) (NSG, N) mice facilitated hematopoiesis. However, transplantation into neonatal mice resulted in high frequency of early death, making it unrealistic for repetitive experiments. In this study, young adult N mice were pre-engrafted with T and B, T alone, B alone or no tissues. The animals were irradiated and injected with autologous fetal liver (FL)-derived CD34(+) cells (34). The resultant mice were TB34N, T34N, B34N and 34N, respectively, and challenged with T cell dependent antigens (Ags). The humanized TB34N mice showed best performance of these mouse models in many aspects resembling the adult human Ag-experienced spleen. The TB34N mice exhibited better hematopoietic reconstitution; balanced development of T- and B-cell, and common progenitor cells; follicular lymphoid structures with a functional germinal center (GC) enriched with follicular dendritic cells (FDCs) and plasma cells (PCs); secretion of hIgG in the sera in response to Ags at comparable levels to those of human; derivations of hIgG mAb-secreting hybridoma clones. Collectively, the humanized TB34N mice could develop an adaptive immunity that was capable of producing Ag-specific hIgG at a significant level via class switching. This unprecedented TB34N platform in humanized mice would be useful in dissecting human immunity, for generating human Abs and clinical applications.


[The hemodynamic reactions of hypertension patients taking the calcium antagonists finoptin and korinfar].
As many as 98 patients suffering from essential hypertension (EH) were examined for the effects of verapamil and nifedipine. The hemodynamics and heart contractility were estimated by means of echocardiography on an "Aloka D-280" outfit (Japan). The following parameters of the cardio- and hemodynamics were computed: the stroke and minute blood volumes, cardiac index, mean velocity of ventricular evacuation, performance of the left heart, energy consumption by the myocardium, arterial pressure (systolic, diastolic and mean), total peripheral vascular resistance, ejection fraction, and the rate of circulatory shortening of myocardial fibers. Analysis of the effects of verapamil and nifedipine on the cardio- and hemodynamics in EH patients has demonstrated that in addition to their unidirectional action manifested by a steady hypotensive effect, the drugs under consideration produced an action of different directions as regards certain essential parameters in a considerable number of the patients.


Risk and ethical concerns of hunting male elephant: behavioural and physiological assays of the remaining elephants.
Hunting of male African elephants may pose ethical and risk concerns, particularly given their status as a charismatic species of high touristic value, yet which are capable of both killing people and damaging infrastructure. We quantified the effect of hunts of male elephants on (1) risk of attack or damage (11 hunts), and (2) behavioural (movement dynamics) and physiological (stress hormone metabolite concentrations) responses (4 hunts) in Pilanesberg National Park. For eleven hunts, there were no subsequent attacks on people or infrastructure, and elephants did not break out of the fenced reserve. For three focal hunts, there was an initial flight response by bulls present at the hunting site, but their movements stabilised the day after the hunt event. Animals not present at the hunt (both bulls and herds) did not show movement responses. Physiologically, hunting elephant bulls increased faecal stress hormone levels (corticosterone metabolites) in both those bulls that were present at the hunts (for up to four days post-hunt) and in the broader bull and breeding herd population (for up to one month post-hunt). As all responses were relatively minor, hunting male elephants is ethically acceptable when considering effects on the remaining elephant population; however bulls should be hunted when alone. Hunting is feasible in relatively small enclosed reserves without major risk of attack, damage, or breakout. Physiological stress assays were more effective than behavioural responses in detecting effects of human intervention. Similar studies should evaluate intervention consequences, inform and improve best practice, and should be widely applied by management agencies.


Real-life use of vitamin D3-fortified bread and milk during a winter season: the effects of CYP2R1 and GC genes on 25-hydroxyvitamin D concentrations in Danish families, the VitmaD study.
Common genetic variants rs10741657 and rs10766197 in CYP2R1 and rs4588 and rs842999 in GC and a combined genetic risk score (GRS) of these four variants influence late summer 25-hydroxyvitamin D (25(OH)D) concentrations. The objectives were to identify those who are most at risk of developing low vitamin D status during winter and to assess whether vitamin D3-fortified bread and milk will increase 25(OH)D concentrations in those with genetically determined low 25(OH)D concentrations at late summer. We used data from the VitmaD study. Participants were allocated to either vitamin D3-fortified bread and milk or non-fortified bread and milk during winter. In the fortification group, CYP2R1 (rs10741657) and GC (rs4588 and rs842999) were statistically significantly associated with winter 25(OH)D concentrations and CYP2R1 (rs10766197) was borderline significant. There was a negative linear trend between 25(OH)D concentrations and carriage of 0-8 risk alleles (p < 0.0001). No association was found for the control group (p = 0.1428). There was a significant positive linear relationship between different quintiles of total vitamin D intake and the increase in 25(OH)D concentrations among carriers of 0-2 (p = 0.0012), 3 (p = 0.0001), 4 (p = 0.0118) or 5 (p = 0.0029) risk alleles, but not among carriers of 6-8 risk alleles (p = 0.1051). Carriers of a high GRS were more prone to be vitamin D deficient compared to carriers of a low GRS. Furthermore, rs4588-AA carriers have a low but very stable 25(OH)D concentration, and interestingly, also low PTH level.


Olfactometric Comparison of the Volatile Insecticide, Metofluthrin, Through Behavioral Responses of Aedes albopictus (Diptera: Culicidae).
Testing behavioral response to insecticidal volatiles requires modifications to the existing designs of olfactometers. To create a testing apparatus in which there is no chemical memory to confound tests, we detail the technical aspects of a new tool with design influences from other olfactometry tools. In addition, this new tool was used to evaluate a novel formulation of metofluthrin for use as an outdoor residual treatment. After sourcing materials to prioritize glass and metal construction, a modular wind tunnel was developed that hybridizes wind tunnel and olfactometer specifications. Volatile contaminants were removed by strong ultraviolet light within the chamber before and between trials. Repellent trials were conducted with an experimental formulation of metofluthrin and a commercial formulation of esfenvalerate, prallethrin, and piperonyl butoxide (Onslaught Fast Cap) against Aedes albopictus (Skuse). Toxicant vapors were delivered with attractants from a lure with behavioral responses scored 20 min post-exposure. Upwind attraction to the attractant lure and the Onslaught Fast Cap plus lure resulted in 90 and 75% capture, respectively. In contrast, metofluthrin vapors resulted in less than 10% attraction, while also causing repellency, disorientation, knockdown, and mortality effects. Our findings demonstrated that an inert modular wind tunnel was functional for mitigating toxic secondary exposures of spatial repellents amidst complex behavioral analysis in mosquitoes. The resulting observations with formulated metofluthrin positively reinforce the merit of transitioning metofluthrin into expanded roles in mosquito management.


Tumour-specific amplitude-modulated radiofrequency electromagnetic fields induce differentiation of hepatocellular carcinoma via targeting Cav3.2 T-type voltage-gated calcium channels and Ca2+ influx.
Administration of amplitude modulated 27·12 MHz radiofrequency electromagnetic fields (AM RF EMF) by means of a spoon-shaped applicator placed on the patient's tongue is a newly approved treatment for advanced hepatocellular carcinoma (HCC). The mechanism of action of tumour-specific AM RF EMF is largely unknown. Whole body and organ-specific human dosimetry analyses were performed. Mice carrying human HCC xenografts were exposed to AM RF EMF using a small animal AM RF EMF exposure system replicating human dosimetry and exposure time. We performed histological analysis of tumours following exposure to AM RF EMF. Using an agnostic genomic approach, we characterized the mechanism of action of AM RF EMF. Intrabuccal administration results in systemic delivery of athermal AM RF EMF from head to toe at levels lower than those generated by cell phones held close to the body. Tumour shrinkage results from differentiation of HCC cells into quiescent cells with spindle morphology. AM RF EMF targeted antiproliferative effects and cancer stem cell inhibiting effects are mediated by Ca2+ influx through Cav3·2 T-type voltage-gated calcium channels (CACNA1H) resulting in increased intracellular calcium concentration within HCC cells only. Intrabuccally-administered AM RF EMF is a systemic therapy that selectively block the growth of HCC cells. AM RF EMF pronounced inhibitory effects on cancer stem cells may explain the exceptionally long responses observed in several patients with advanced HCC. FUND: Research reported in this publication was supported by the National Cancer Institute's Cancer Centre Support Grant award number P30CA012197 issued to the Wake Forest Baptist Comprehensive Cancer Centre (BP) and by funds from the Charles L. Spurr Professorship Fund (BP). DWG is supported by R01 AA016852 and P50 AA026117.


Otitis Media Practice During the COVID-19 Pandemic.
The global coronavirus disease-2019 (COVID-19) pandemic has changed the prevalence and management of many pediatric infectious diseases, including acute otitis media (AOM). Coronaviruses are a group of RNA viruses that cause respiratory tract infections in humans. Before the COVID-19 pandemic, coronavirus serotypes OC43, 229E, HKU1, and NL63 were infrequently detected in middle ear fluid (MEF) specimens and nasopharyngeal aspirates in children with AOM during the 1990s and 2000s and were associated with a mild course of the disease. At times when CoV was detected in OM cases, the overall viral load was relatively low. The new severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) is the causative pathogen responsible for the eruption of the COVID-19 global pandemic. Following the pandemic declaration in many countries and by the World Health Organization in March 2020, preventive proactive measures were imposed to limit COVID-19. These included social distancing; lockdowns; closure of workplaces; kindergartens and schools; increased hygiene; use of antiseptics and alcohol-based gels; frequent temperature measurements and wearing masks. These measures were not the only ones taken, as hospitals and clinics tried to minimize treating non-urgent medical referrals such as OM, and elective surgical procedures were canceled, such as ventilating tube insertion (VTI). These changes and regulations altered the way OM is practiced during the COVID-19 pandemic. Advents in technology allowed a vast use of telemedicine technologies for OM, however, the accuracy of AOM diagnosis in those encounters was in doubt, and antibiotic prescription rates were still reported to be high. There was an overall decrease in AOM episodes and admissions rates and with high spontaneous resolution rates of MEF in children, and a reduction in VTI surgeries. Despite an initial fear regarding viral shedding during myringotomy, the procedure was shown to be safe. Special draping techniques for otologic surgery were suggested. Other aspects of OM practice included the presentation of adult patients with AOM who tested positive for SARS-2-CoV and its detection in MEF samples in living patients and in the mucosa of the middle ear and mastoid in post-mortem specimens.


Expression and splicing patterns of human papillomavirus type-16 mRNAs in pre-cancerous lesions and carcinomas of the cervix, in human keratinocytes immortalized by HPV 16, and in cell lines established from cervical cancers.
We have analysed the splicing patterns of human papillomavirus (HPV) type-16 mRNAs in a human epithelial cell line immortalized by HPV 16 (HPKII), in cell lines established from cervical carcinomas (SiHa and CaSki) and in pre-invasive and invasive carcinomas of the cervix. The presence of mRNA species previously described, which could encode the E6, E6I, E6II, E6III, E7, E2, E2C, E4, E5 and L1 proteins, was determined, using the RNA polymerase chain reaction (PCR) technique with primers that flank unique splice sites. The state of the viral DNA in the tumor biopsies was established by Southern blot analysis. The various HPV 16 transcripts could be detected in cell lines and in tumor biopsies. The size of the RNA PCR products were in agreement with the previously mapped splice sites. The full range of transcripts was revealed in the HPKII cell line and in a number of pre-invasive carcinomas. Messenger RNAs which could encode the E6III, E4 and E5 proteins were most prevalent in all types of tumor. The overall results of DNA and RNA analyses in cell lines and tumor specimens indicate that (1) expression of either of the early or late transcripts studied is not specifically related to (a) tumor stage or (b) the physical state of the viral genome; and (2) alterations in the splicing patterns of HPV 16 transcripts may not be involved in tumor progression.


[A case of poorly differentiated adenocarcinoma of the rectum with disseminated carcinomatosis of the bone marrow successfully treated with mFOLFOX-6/bevacizumab].
A 45-year-old woman, complaining of back pain and bloody stool was given a diagnosis of poorly differentiated adenocarcinoma of the rectum with disseminated carcinomatosis to bone marrow and disseminated intravascular coagulation syndrome (DIC). We started chemotherapy using mFOLFOX-6 with simultaneous DIC treatment. After we confirmed that DIC had improved following 2 courses of mFOLFOX-6, bevacizumab was added to mFOLFOX-6. Laboratory studies revealed a serum CEA level of 314.4ng/ml, which improved to 4.6ng/ml after a total of 6 courses of chemotherapy. Colonoscopy findings showed almost normal rectal mucosa after a total of 10 courses of chemotherapy. Outpatient treatment was started after 5 courses of chemotherapy, and was continuing according to schedule at 7 months from the onset of this disease.


Importance of the monthly biochemical evaluation to identify patients on hemodialysis with malnutrition.
Malnutrition is a frequent complication in patients on hemodialysis and early diagnosis is important to reduce the morbidity and mortality of treatment. To investigate the usefulness of biochemical tests performed monthly in order to identify hemodialysis patients with malnutrition. The nutritional status of 252 patients was evaluated by objective and subjective global assessment, and the patients classified as malnourished and not malnourished. Then, during 4 consecutive months, serum creatinine, phosphorus, urea pre-and post-dialysis and Kt/V were recorded for calculation of averages. After analysis of these variables by the ROC curve we calculated the sensitivity and specificity of these parameters to identify patients with malnutrition. In ROC curve, the area under the curve for urea was 0.683, 0.71 for phosphorus, 0.724 for Kt/V and 0.765 for creatinine. For values of urea ≤ 90 mg/dL, phosphorus ≤ 4.2 mg/dL, Kt/V ≥ 1.6 and creatinine ≤ 6.5 mg/dL, the specificity ranged between 80% and 88% and sensitivity between 26% and 51%. The negative predictive value ranged between 90% and 92% and positive predictive value between 23% and 32%. The association of two or more of these índices did not change significantly these values. Our results suggest that serum urea ≤ 90 mg/dL, creatinine ≤ 6.5 mg/dL, phosphorus ≤ 4.2 mg/dL, and Kt/V ≥ 1.6 can be used for screening patients with malnutrition. However, using these cutoffs the parameters tend to overestimate the number of patients with malnutrition.


The Influence of End-of-Life Care on Organ Donor Potential.
Many patients with acute devastating brain injury die outside intensive care units and could go unrecognized as potential organ donors. We conducted a prospective observational study in seven hospitals in the Netherlands to define the number of unrecognized potential organ donors outside intensive care units, and to identify the effect that end-of-life care has on organ donor potential. Records of all patients who died between January 2013 and March 2014 were reviewed. Patients were included if they died within 72 h after hospital admission outside the intensive care unit due to devastating brain injury, and fulfilled the criteria for organ donation. Physicians of included patients were interviewed using a standardized questionnaire regarding logistics and medical decisions related to end-of-life care. Of the 5170 patients screened, we found 72 additional potential organ donors outside intensive care units. Initiation of end-of-life care in acute settings and lack of knowledge and experience in organ donation practices outside intensive care units can result in under-recognition of potential donors equivalent to 11-34% of the total pool of organ donors. Collaboration with the intensive care unit and adjusting the end-of-life path in these patients is required to increase the likelihood of organ donation.


Temperature regulates splicing efficiency of the cold-inducible RNA-binding protein gene Cirbp.
In mammals, body temperature fluctuates diurnally around a mean value of 36°C-37°C. Despite the small differences between minimal and maximal values, body temperature rhythms can drive robust cycles in gene expression in cultured cells and, likely, animals. Here we studied the mechanisms responsible for the temperature-dependent expression of cold-inducible RNA-binding protein (CIRBP). In NIH3T3 fibroblasts exposed to simulated mouse body temperature cycles, Cirbp mRNA oscillates about threefold in abundance, as it does in mouse livers. This daily mRNA accumulation cycle is directly controlled by temperature oscillations and does not depend on the cells' circadian clocks. Here we show that the temperature-dependent accumulation of Cirbp mRNA is controlled primarily by the regulation of splicing efficiency, defined as the fraction of Cirbp pre-mRNA processed into mature mRNA. As revealed by genome-wide "approach to steady-state" kinetics, this post-transcriptional mechanism is widespread in the temperature-dependent control of gene expression.


Extra-anal mucosectomy: laparascopic-assisted endorectal pull-through using a prolapsing technique.
The mucosectomy of the aganglionic rectal mucosa has been performed transabdominally in the Soave procedure. Recently, mucosectomy with transanal approach was reported both in the laparoscopic and the open surgical procedure. However, the operative view and working field are restricted because the dissection has to be done in the anal canal. The authors report an innovative approach, an extra-anal mucosectomy, to perform mucosectomy in the Soave procedure for Hirschspung's disease in combination with the laparoscopic-assisted prolapsing technique. Four patients underwent laparoscopic surgery for Hirschsprung's disease with extra-anal mucosectomy between 1995 and 1997. One 10-mm and 5-mm ports were used throughout the operation. The rectum was pulled out through the anal canal to create a rectal prolapse and was divided outside the anal canal. The mucosectomy was performed in the everted rectal mucosa outside the anal canal to the level of dentate line. The colon was sutured to the anal mucosa 2 mm above the dentate line. Mean operative time was 3 hours. The present technique made the whole mucosectomy possible under direct observation. The extra-anal mucosectomy in conjunction with a laparoscopic-assisted prolapsing technique seems to be a safe and reliable modality in the surgical treatment of Hirschsprung's disease.


In vitro inhibition of adherence of Streptococcus mutans strains by nonadherent mutants of S. mutans 6715.
Four nonadherent mutants from Streptococcus mutans 6715 mutant UAB66 (serotype g) with similar phenotypes were shown to inhibit the adherence of adherence-proficient S. mutans serotypes c and g strains. One mutant, UAB108, was shown to inhibit adherence by wild-type strains representing serotypes a, d, and e as well. This inhibition of adherence was seen with pairs of strains grown in partially defined (PD) medium supplemented with 1% sucrose in both microtiter plates and glass tubes. The inhibiting factor was present in culture supernatant fluids of inhibiting strains grown in PD medium plus 1% sucrose and was heat stable. Ethanol precipitation of culture supernatant fluids of these strains yielded a water-soluble polymer which effectively inhibited the adherence of UAB66. This polymer, isolated from UAB108, was also shown to inhibit the adherence of UAB66 at lower concentrations than that needed to inhibit adherence with dextran T10. Partially purified glucosyltransferase, isolated from the culture supernatant fluids of glucose-grown UAB108, produced a water-soluble glucan which was shown to inhibit the adherence of UAB66 as well. The methods developed permit rapid screening for strains or mutants of strains or both that inhibit adherence or plaque formation or both by wild-type strains of S. mutans.


CRISPR-Cas12a delivery by DNA-mediated bioresponsive editing for cholesterol regulation.
CRISPR-Cas12a represents an efficient tool for genome editing in addition to the extensively investigated CRISPR-Cas9. However, development of efficient nonviral delivery system for CRISPR-Cas12a remains challenging. Here, we demonstrate a DNA nanoclew (NC)-based carrier for delivery of Cas12a/CRISPR RNA (crRNA) ribonucleoprotein (RNP) toward regulating serum cholesterol levels. The DNA NC could efficiently load the Cas12a/crRNA RNP through complementation between the DNA NC and the crRNA. Addition of a cationic polymer layer condensed the DNA-templated core and allowed further coating of a charge reversal polymer layer, which makes the assembly negatively charged under a physiological pH but reverts to positive charge under an acidic environment. When Pcsk9 was selected as the target gene because of its important role in regulating the level of serum cholesterol, efficient Pcsk9 disruption was observed in vivo (~48%), significantly reducing the expression of PCSK9 and gaining the therapeutic benefit of cholesterol control (~45% of cholesterol reduction).


mGluR4-positive allosteric modulation as potential treatment for Parkinson's disease.
Although Parkinson's disease was first diagnosed nearly 200 years ago, its effective treatment still remains elusive for most of those diagnosed. The gold standard of treatment for most patients is 3,4-dihydroxy-L-phenylalanine. This drug works for most individuals early in the disease; however, resistant symptoms start to emerge after several years of treatment. There has been increased interest in finding novel therapies to help Parkinson's disease patients. Such strategies may have the benefit of not only treating the symptomatic issues of the disorder, but might also offer promise in protecting dopaminergic neurons from further degeneration. One such target that is now receiving much attention from the scientific community is the metabotropic glutamate receptor mGluR4. In this article, we briefly review Parkinson's disease and then recent work in the mGluR area, with a focus on the efforts being made toward finding and optimizing novel mGluR4 positive allosteric modulators (PAMs). Preclinically in rodent models, mGluR4 activation has offered much promise as a novel treatment of Parkinson's disease. Additionally, the specific use of PAMs, rather than direct-acting agonists at the orthosteric glutamate site, continues to be validated as a viable treatment option for this target. It is anticipated that continued progress in this area will further our understanding of the potential of mGluR4 modulation as a novel symptomatic and potentially disease-modifying treatment for Parkinson's disease.


The Efficacy of Pulsed Dye Laser Treatment for Acne Vulgaris: A Systemic Review and Meta-Analysis.
Previous reports have shown that pulsed dye laser (PDL) is promising for the treatment of acne; however, results provide conflicting evidence. To determine the efficacy of PDL in treating acne vulgaris. A systematic review and meta-analysis of studies published before March 31, 2020 was conducted. Randomized controlled trials and case series were analyzed. The meta-analysis estimated the standardized mean difference (SMD) in acne severity score changes in patients treated by PDL versus control and also the SMD of the acne severity scores and comedone counts changes before and after PDL treatment. Eleven studies were included in this systematic review. Six studies were included in the meta-analysis. Pulsed dye laser treatment was not found to be superior to the control group in treating acne vulgaris (SMD: -0.285; 95% confidence interval [CI], -0.886 to 0.317). However, single-arm studies revealed a significantly improved acne severity score after PDL therapy (SMD, -1.321; 95% CI, -2.057 to -0.586), especially when a multiple-session treatment and a longer pulse duration were employed. The comedone counts were significantly decreased after PDL therapy (SMD, -0.596; 95% CI, -1.137 to -0.054). When treatment consisted of 4 or more sessions or longer pulse duration, PDL could significantly decrease the acne severity score.


A Canadian guideline on the use of next-generation sequencing in oncology.
Rapid advancements in next-generation sequencing (ngs) technology have created an unprecedented opportunity to decipher the molecular profile of tumours to more effectively prevent, diagnose, and treat cancer. Oncologists now have the option to order molecular tests that can guide treatment decisions. However, to date, most oncologists have received limited training in genomics, and they are now faced with the challenge of understanding how such tests and their interpretation align with patient management. Guidance on how to effectively use ngs technology is therefore needed to aid oncologists in applying the results of genomic tests. The Canadian guideline presented here describes best practices and unmet needs related to ngs-based testing for somatic variants in oncology, including clinical application, assay and sample selection, bioinformatics and interpretation of reports performed by laboratories, patient communication, and clinical trials.


Ethnic inequality within the elderly population in utilizing healthcare services.
The accessibility of minority ethnic groups to healthcare services is challenging in many societies, most especially among the elderly population. Elderly individuals from minority groups have been found to have lower levels of utilizing healthcare services, including preventive care, intensive hospital care, advanced technological procedures and rehabilitation. Universal health coverage is incapable of addressing all of healthcare's access inequities and there is a need to assess the overall outcomes, including mortality rates over time, functionality of discharged patients, quality of life and/or unplanned readmissions that may indicate low quality hospital discharge processes. There is a need to investigate the impact of perceived trust/distrust in the healthcare system of elderly patients from minority ethnicities on their willingness to consume medical services.To ensure equity in service provision, there is a need to examine whether medical providers, even unconsciously, prioritize vital services, such as rehabilitation services to populations that share similar social backgrounds. An essential measure is enhancement of health literacy at all levels, from the individual to policy-makers and strategic adoption of health literacy programs that encompass all ethnicities, considering their respective needs, norms and expectations.Ethnic equality in accessing medical services is crucial in view of the numerous migrants and asylum-seekers who look for refuge in varied societies globally. Such populations are perceived as faring worse in healthcare quality of care, and this highlights the need to adapt the healthcare systems to the varied health behaviors, contextual factors, language barriers, lower health literacy levels and limited access to timely care. Improving equity and access to medical care is dependent on enhanced health literacy; policies that consider diverse needs of majority and minority groups; and advanced research. Concurrent implementation of these measures will be well aligned with the global strive to promote the Sustainable Development Goals (SDGs).


A high prevalence of blaOXA-48 in Klebsiella (Raoultella) ornithinolytica and related species in hospital wastewater in South West England.
Klebsiella species occupy a wide range of environmental and animal niches, and occasionally cause opportunistic infections that are resistant to multiple antibiotics. In particular, Klebsiella pneumoniae (Kpne) has gained notoriety as a major nosocomial pathogen, due principally to the rise in non-susceptibility to carbapenems and other beta-lactam antibiotics. Whilst it has been proposed that the urban water cycle facilitates transmission of pathogens between clinical settings and the environment, the level of risk posed by resistant Klebsiella strains in hospital wastewater remains unclear. We used whole genome sequencing (WGS) to compare Klebsiella species in contemporaneous samples of wastewater from an English hospital and influent to the associated wastewater treatment plant (WWTP). As we aimed to characterize representative samples of Klebsiella communities, we did not actively select for antibiotic resistance (other than for ampicillin), nor for specific Klebsiella species. Two species, Kpne and K. (Raoultella) ornithinolytica (Korn), were of equal dominance in the hospital wastewater, and four other Klebsiella species were present in low abundance in this sample. In contrast, despite being the species most closely associated with healthcare settings, Kpne was the dominant species within the WWTP influent. In total, 29 % of all isolates harboured the blaOXA-48 gene on a pOXA-48-like plasmid, and these isolates were almost exclusively recovered from the hospital wastewater. This gene was far more common in Korn (68 % of isolates) than in Kpne (3.4 % of isolates). In general plasmid-borne, but not chromosomal, resistance genes were significantly enriched in the hospital wastewater sample. These data implicate hospital wastewater as an important reservoir for antibiotic-resistant Klebsiella, and point to an unsuspected role of species within the Raoultella group in the maintenance and dissemination of plasmid-borne blaOXA-48. This article contains data hosted by Microreact.


Serum magnesium concentrations in atrial fibrillation.
Serum magnesium and digoxin levels were obtained in 13 nontoxic patients with atrial fibrillation due to chronic rheumatic heart disease receiving digoxin for the control of ventricular rate and heart failure. Fairly good correlations were made between serum digoxin levels and ventricular rates. Hypomagnesemia was quite common (7 out of 13) and mean magnesium serum levels were significantly lowered in total as well in 7 hypomagnesemic patients, as compared to in healthy controls. Magnesium sulphate was successfully used in patients with magnesium deficiency to control the ventricular rates.


Valence framing effects on moral judgments: A meta-analysis.
Valence framing effects occur when participants make different choices or judgments depending on whether the options are described in terms of their positive outcomes (e.g. lives saved) or their negative outcomes (e.g. lives lost). When such framing effects occur in the domain of moral judgments, they have been taken to cast doubt on the reliability of moral judgments and raise questions about the extent to which these moral judgments are self-evident or justified in themselves. One important factor in this debate is the magnitude and variability of the extent to which differences in framing presentation impact moral judgments. Although moral framing effects have been studied by psychologists, the overall strength of these effects pooled across published studies is not yet known. Here we conducted a meta-analysis of 109 published articles (contributing a total of 146 unique experiments with 49,564 participants) involving valence framing effects on moral judgments and found a moderate effect (d = 0.50) among between-subjects designs as well as several moderator variables. While we find evidence for publication bias, statistically accounting for publication bias attenuates, but does not eliminate, this effect (d = 0.22). This suggests that the magnitude of valence framing effects on moral decisions is small, yet significant when accounting for publication bias.


Curcumin inhibits VEGF-mediated angiogenesis in human intestinal microvascular endothelial cells through COX-2 and MAPK inhibition.
Angiogenesis, the growth of new blood vessels, is a critical homeostatic mechanism which regulates vascular populations in response to physiological requirements and pathophysiological demand, including chronic inflammation and cancer. The importance of angiogenesis in gastrointestinal chronic inflammation and cancer has been defined, as antiangiogenic therapy has demonstrated benefit in models of inflammatory bowel disease and colon cancer treatment. Curcumin is a natural product undergoing evaluation for the treatment of chronic inflammation, including inflammatory bowel disease (IBD). The effect of curcumin on human intestinal angiogenesis is not defined. The antiangiogenic effect of curcumin on in vitro angiogenesis was examined using primary cultures of human intestinal microvascular endothelial cells (HIMECs), stimulated with vascular endothelial growth factor (VEGF). Curcumin inhibited proliferation, cell migration and tube formation in HIMECs induced by VEGF. Activation of HIMECs by VEGF resulted in enhanced expression of cyclo-oxygenase-2 (COX-2) mRNA, protein and prostaglandin E(2) (PGE(2)) production. Pretreatment of HIMECs with 10 microM curcumin as well as 1 microM NS398, a selective inhibitor of COX-2, resulted in inhibition of COX-2 at the mRNA and protein level and PGE(2) production. Similarly COX-2 expression in HIMECs was significantly inhibited by Jun N-terminal kinase (JNK; SP600125) and p38 mitogen-activated protein kinase (MAPK; SB203580) inhibitors and was reduced by p44/42 MAPK inhibitor (PD098059). Taken together, these data demonstrate an important role for COX-2 in the regulation of angiogenesis in HIMECs via MAPKs. Moreover, curcumin inhibits microvascular endothelial cell angiogenesis through inhibition of COX-2 expression and PGE(2) production, suggesting that this natural product possesses antiangiogenic properties, which warrants further investigation as adjuvant treatment of IBD and cancer.


Prosthetic valve endocarditis: an overview.
Infection of an intracardiac prosthesis, the incidence of which is about 2.5% among patients having undergone valve replacement, is a serious complication with considerable morbidity and mortality. Early prosthetic valve endocarditis (PVE), with an onset within 60 days of valve replacement, accounts for approximately one-third of all cases, while the remaining two-thirds, occur more than two months postoperatively (late prosthetic valve endocarditis). Prosthetic valve endocarditis is most commonly caused by Staphylococcus epidermidis, less frequently by viridans streptococci, Staphylococcus aureus, and gram-negative bacilli. The most likely pathogenetic mechanisms in prosthetic valve endocarditis are intraoperative contamination and postoperative infections at extracardiac sites. Prominent clinical features include fever, new or changing heart murmurs, leukocytosis, anemia and hematuria. The etiologic microorganism can be isolated in more than 90% of all cases. Patients with proven prosthetic valve endocarditis should be examined daily to detect signs of congestive heart failure and changes in murmurs; electrocardiographic monitoring is essential for documentation of arrhythmias. With limitations, echocardiography, especially two-dimensional, may help to demonstrate vegetations or valvular dehiscence. Cinefluoroscopy may reveal loosening or dehiscence of the sewing ring or impaired motion of a radio-opaque poppet due to thrombus or vegetation. Cardiac catheterization, not always necessary even when surgical intervention is anticipated, may provide valuable information on the degree of dysfunction, multiple valve involvement, left ventricular function and extent of concomitant coronary artery disease. In patients with mechanical valves, prosthetic valve endocarditis may be associated with a high incidence of valve ring and myocardial abscesses; the reported frequency of valve ring abscesses is lower with porcine heterografts. Infections on mechanical valves characteristically localize to the sewing ring with subsequent detachment of the prosthesis and valvular incompetence; infections on porcine heterografts tend to localize to the cusps, leading to valvular incompetence because of leaflet destruction. Large vegetations may result in functional stenosis. Over the last ten years the overall mortality of prosthetic valve endocarditis was 53.8%; 73.6% in early and 43% in late prosthetic valve endocarditis. More recently, however, the survival rate appears to be improving. In general, the mortality associated with prosthetic valve endocarditis caused by fungi and Staphylococcus aureus is highest and that of streptococci lowest.(ABSTRACT TRUNCATED AT 400 WORDS)


Polymer ligand exchange to control stabilization and compatibilization of nanocrystals.
We demonstrate polymer ligand exchange to be an efficient method to control steric stabilization and compatibilization of nanocrystals. A rational design of polymer binding groups and ligand exchange conditions allows to attach polymer brushes with grafting densities >1 nm(-2) to inorganic nanocrystals for nearly any nanocrystal/polymer combination using only a few types of binding groups. We demonstrate the potential of the method as an alternative to established grafting-from and grafting-to routes in considerably increasing the stabilization of inorganic nanocrystals in solution, to prepare completely miscible polymer nanocomposites with a controllable distance between nanoparticles, and to induce and control aggregation into percolation networks in polymeric matrices for a variety of different nanocrystal/polymer combinations. A dense attachment of very short polymer ligands is possible enabling to prepare ordered nanoparticle monolayers with a distance or pitch of only 7.2 nm, corresponding to a potential magnetic storage density of 12.4 Tb/in(2). Not only end-functionalized homopolymers, but also commercially available copolymers with functional comonomers can be used for stable ligand exchange, demonstrating the versatility and broad potential of the method.


Automated Spectroscopic Tissue Classification in Colorectal Surgery.
In colorectal surgery, detecting ureters and mesenteric arteries is of utmost importance to prevent iatrogenic injury and to facilitate intraoperative decision making. A tool enabling ureter- and artery-specific image enhancement within (and possibly through) surrounding adipose tissue would facilitate this need, especially during laparoscopy. To evaluate the potential of hyperspectral imaging in colorectal surgery, we explored spectral tissue signatures using single-spot diffuse reflectance spectroscopy (DRS). As hyperspectral cameras with silicon (Si) and indium gallium arsenide (InGaAs) sensor chips are becoming available, we investigated spectral distinctive features for both sensor ranges. In vivo wide-band (wavelength range 350-1830 nm) DRS was performed during open colorectal surgery. From the recorded spectra, 36 features were extracted at predefined wavelengths: 18 gradients and 18 amplitude differences. For classification of respectively ureter and artery in relation to surrounding adipose tissue, the best distinctive feature was selected using binary logistic regression for Si- and InGaAs-sensor spectral ranges separately. Classification performance was evaluated by leave-one-out cross-validation. In 10 consecutive patients, 253 spectra were recorded on 53 tissue sites (including colon, adipose tissue, muscle, artery, vein, ureter). Classification of ureter versus adipose tissue revealed accuracy of 100% for both Si range and InGaAs range. Classification of artery versus surrounding adipose tissue revealed accuracies of 95% (Si) and 89% (InGaAs). Intraoperative DRS showed that Si and InGaAs sensors are equally suited for automated classification of ureter versus surrounding adipose tissue. Si sensors seem better suited for classifying artery versus mesenteric adipose tissue. Progress toward hyperspectral imaging within this field is promising.


Characterization of miR-206 Promoter and Its Association with Birthweight in Chicken.
miRNAs have been widely investigated in terms of cell proliferation and differentiation. However, little is known about their effects on bird growth. Here we characterized the promoter of miR-206 in chicken and found that the preferable promoter was located in 1200 bp upstream of pri-miR-206. In this region, many key transcription factors, including MyoD, c-Myb, CEBPα/β, AP-4, RAP1, Brn2, GATA-1/2/3, E47, Sn, upstream stimulatory factor (USF) and CdxA, were predicted to bind and interact with miR-206 promoter. Overexpression of MyoD sharply increased miR-206 expression in both fibroblast and myoblast cells, and also the regulation in the myoblast cells was much stronger, indicating that miR-206 was regulated by MyoD combined with other muscle specific transcriptional factors. Aiming to further investigate the relationship between miR-206 mutation and transcriptional expression, total of 23 SNPs were identified in the two distinct bird lines by sequencing. Interestingly, the motif bound by MyoD was individually destroyed by G-to-C mutation located at 419 bp upstream of miR-206 precursor. Co-transfecting MyoD and miR-206 promoter in DF-1 cells, the luciferase activity of promoter containing homozygous GG types was significantly higher than CC ones (p < 0.05). Thus, this mutation caused low expression of miR-206. Consistently, eight variants including G-419C mutation exhibited a great effect on birthweight through maker-trait association analysis in F2 population (p < 0.05). Additionally, the regulation of miR-206 on embryo muscle mass mainly by increasing MyoG and muscle creatine kinase (MCK) expression (p < 0.05) with little change in MyoD, TMEM8C and myosin heavy chain (MHC). In conclusion, our findings provide a novel mutation destroying the promoter activity of miR-206 in birds and shed new light to understand the regulation mechanism of miR-206 on the embryonic muscle growth.


Analysis of the genetic diversity of Lonicera japonica Thumb. using inter-simple sequence repeat markers.
Inter-simple sequence repeats (ISSRs) were used to analyze the genetic diversity of 21 accessions obtained from four provinces in China, Shandong, Henan, Hebei, and Sichuan. A total of 272 scored bands were generated using the eight primers previously screened across 21 accessions, of which 267 were polymorphic (98.16%). Genetic similarity coefficients varied from 0.4816 to 0.9118, with an average of 0.6337. The UPGMA dendrogram grouped 21 accessions into two main clusters. Cluster A comprised four Lonicera macranthoides Hand. Mazz. accessions, of which J10 was found to be from Sichuan, and J17, J18, and J19 were found to be from Shandong. Cluster B comprised 17 Lonicera japonica Thumb. accessions, divided into the wild accession J16 and the other 16 cultivars. The results of the principal component analysis were comparable to the cluster analysis. Therefore, the ISSR markers could be effectively used to distinguish interspecific and intraspecific variations, which may facilitate identification of Lonicera japonica cultivars for planting, medicinal use, and germplasm conservation.


[Urinary tract infection in institutionalized elderly patients. Incidence of bacterial resistance and risk factors].
The goal of the present work was to study urinary tract infections (UTI) in a sociosanitary center in order to characterize etiology, define sensitivity profiles, and study associations between these profiles and various clinical parameters. A prospective study of all urinary tract infections at the caring unit of a geriatric sociosanitary center was carried out. Antibiograms were performed on all patients in which a UTI was suspected, and the association between the isolated organism or E. coli resistance with various clinical parameters (gender, bed confinement, recurrence, incontinence, neural impairment, basic daily life activities and drug use) was studied using a multivariate logistic regression analysis. Escherichia coli and Proteus mirabilis are the most commonly isolated organisms (51% and 15%, respectively). The former is associated with bed confinement and the female gender. The latter shows an inverse relation with recurrence in 90 days. Resistance to the antibiotics studied correlates with a number of clinical parameters, although associations vary for each antimicrobial agent. Previous use of antibiotics and recurrence of infection have no influence on resistance. Geriatric patients' characteristics may well account for some of the results encountered. However, the development of resistance in the center may be related to spreading from direct contact between patients. The relationship between resistance and drug use may be affected by the external administration, rather than in-center administration, of drugs.


Prevalence of autoimmune thyroiditis in patients with polycystic ovary syndrome.
Hypothyroidism is associated with pregnancy complications both for the mother and progeny and it should be considered in reproductive age. Thyroid autoimmunity is stated to be the main cause of hypothyroidism in iodine sufficient areas. Polycystic ovary syndrome (PCOS) is known as the most common endocrine disorder affecting women in reproductive age. Early diagnosis and treatment of hypothyroidism in PCOS may reduce the rate of infertility and pregnancy-related morbidity. In the present study we evaluated thyroid autoimmunity in PCOS patients. Over a period of 12 months, 78 patients with PCOS were recruited to this case-control study. Three hundred and fifty age-matched women were studied as a control group. PCOS was defined according to the revised 2003 Rotterdam criteria. Thyroid size was estimated by inspection and palpation. Serum thyroid stimulating hormone (TSH), anti-thyroperoxidase antibody (anti-TPO Ab), and anti-thyroglobulin antibody (anti-Tg Ab) were measured. The mean ± SD of serum anti-TPO Ab in PCOS patients and subjects in the control group was 216 ± 428 and 131 ± 364 IU/mL, respectively (P = 0.04). Prevalence of goiter in PCOS patients was higher than that in control subjects (62.3 vs. 35.7%, P = 0.0001). Serum TSH and anti-Tg Ab in PCOS patients and control subjects did not differ significantly. In this case-control study, anti-thyroid antibodies and goiter prevalence were significantly higher in PCOS patients. These data suggest that thyroid exam and evaluation of thyroid function and autoimmunity should be considered in such patients.


The fastest runner on artificial legs: different limbs, similar function?
The recent competitive successes of a bilateral, transtibial amputee sprint runner who races with modern running prostheses has triggered an international controversy regarding the relative function provided by his artificial limbs. Here, we conducted three tests of functional similarity between this amputee sprinter and competitive male runners with intact limbs: the metabolic cost of running, sprinting endurance, and running mechanics. Metabolic and mechanical data, respectively, were acquired via indirect calorimetry and ground reaction force measurements during constant-speed, level treadmill running. First, we found that the mean gross metabolic cost of transport of our amputee sprint subject (174.9 ml O(2)*kg(-1)*km(-1); speeds: 2.5-4.1 m/s) was only 3.8% lower than mean values for intact-limb elite distance runners and 6.7% lower than for subelite distance runners but 17% lower than for intact-limb 400-m specialists [210.6 (SD 13.2) ml O(2)*kg(-1)*km(-1)]. Second, the speeds that our amputee sprinter maintained for six all-out, constant-speed trials to failure (speeds: 6.6-10.8 m/s; durations: 2-90 s) were within 2.2 (SD 0.6)% of those predicted for intact-limb sprinters. Third, at sprinting speeds of 8.0, 9.0, and 10.0 m/s, our amputee subject had longer foot-ground contact times [+14.7 (SD 4.2)%], shorter aerial [-26.4 (SD 9.9)%] and swing times [-15.2 (SD 6.9)%], and lower stance-averaged vertical forces [-19.3 (SD 3.1)%] than intact-limb sprinters [top speeds = 10.8 vs. 10.8 (SD 0.6) m/s]. We conclude that running on modern, lower-limb sprinting prostheses appears to be physiologically similar but mechanically different from running with intact limbs.


Expression of the human poliovirus receptor/CD155 gene is activated by sonic hedgehog.
The human poliovirus receptor/CD155 is a transmembrane glycoprotein belonging to the immunoglobulin superfamily. The ectodomain of CD155 mediates cell attachment to the extracellular matrix molecule vitronectin, while its intracellular domain interacts with the dynein light chain Tctex-1. CD155 is a primate-restricted gene that is expressed during development in mesenchymal tissues and ventrally derived structures within the CNS. Its function in adults is as yet unknown, but significantly, CD155 is aberrantly expressed in neuroectodermal tumors. We show that the expression of CD155 mRNA is up-regulated when human Ntera2 cells are treated with purified Sonic hedgehog (Shh) protein. Reporter gene expression driven by the CD155 core promoter is activated by Shh in transient co-transfection assays. Analysis of the CD155 core promoter indicates that an intact GLI binding site is required for Shh activation. In addition, overexpression of Gli1 or Gli3 potently activates reporter gene expression driven by the CD155 core promoter. These data identify the CD155 gene as a transcriptional target of Shh, a finding that has significance for the normal function of CD155 during development and the expression of CD155 in neuroectodermal tumors.