Electrochemical characterization of a family AA10 LPMO and the impact of residues shaping the copper site on reactivity.
Research on enzymes for lignocellulose biomass degradation has progressively increased in recent years due to the interest in taking advantage of this natural resource. Among these enzymes are the lytic polysaccharide monooxygenases (LPMOs) that oxidatively depolymerize crystalline cellulose using a reactive oxygen species generated in a reduced mono‑copper active site. The copper site comprises of a highly conserved histidine-brace, providing three equatorial nitrogen ligands, whereas less conserved residues close to the copper contribute to shaping and confining the site. The catalytic copper site is exposed to the solvent and to the crystalline substrates, and as so, the influence of the copper environment on LPMO properties, including the redox potential, is of great interest. In the current work, a direct electrochemical study of an LPMO (ScLPMO10C) was conducted allowing to retrieve kinetic and thermodynamic data associated with the redox transition in the catalytic centre. Moreover, two residues that do not bind to the copper but shape the copper sites were mutated, and the properties of the mutants were compared with those of the wild-type enzyme. The direct electrochemical studies, using cyclic voltammetry, yielded redox potentials in the +200 mV range, well in line with LPMO redox potentials determined by other methods. Interestingly, while the mutations hardly affected the formal redox potential of the enzyme, they drastically affected the reactivity of the copper site and enzyme functionality.


Expression and Clinical Significance of High-Mobility Group AT-hook 2 (HMGA2) in Osteosarcoma.
Although high-mobility group AT-hook 2 (HMGA2) has been shown to have crucial roles in the pathogenesis and metastasis of various malignancies, its expression and significance in osteosarcoma remain unknown. Here we evaluate the expression, clinical prognostic value, and overall function of HMGA2 in osteosarcoma. Sixty-nine osteosarcoma patient specimens within a tissue microarray (TMA) were analyzed by immunohistochemistry for HMGA2 expression. Demographics and clinicopathological information including age, gender, tumor location, metastasis, recurrence, chemotherapy response, follow-up time, and disease status were also collected. After validation of expression, we determined whether there was a correlation between HMGA2 expression and patient clinicopathology. HMGA2 expression was also evaluated in osteosarcoma cell lines and patient tissues by Western blot, we analyzed the expression of HMGA2 in the human osteosarcoma cell lines MG63, 143B, U2OS, Saos-2, MNNG/HOS, and KHOS. HMGA2-specific siRNA and clonogenic assays were then used to determine the effect of HMGA2 inhibition on osteosarcoma cell proliferation, growth, and chemosensitivity. HMGA2 expression was elevated in the osteosarcoma patient specimens and human osteosarcoma cell lines. HMGA2 was differentially expressed in human osteosarcoma cell lines. Specifically, a relatively high expression of HMGA2 was present in KHOS, MNNG/HOS, 143B and a relatively low expression was in MG63, U2OS as well as Saos-2. HMGA2 expression is correlated with metastasis and shorter overall survival. High HMGA2 expression is an independent predictor of poor osteosarcoma prognosis. There was no significant correlation between HMGA2 expression and the age, gender, or tumor site of the patient. HMGA2 expression is predominantly within the nucleus. The expression of HMGA2 also directly correlated to neoadjuvant chemoresistance. There was a significant reduction of HMGA2 expression in the siRNA transfection group. After the use of siRNA, the proliferation of osteosarcoma cells is decreased and the chemosensitivity of osteosarcoma cells is significantly increased. Our study supports HMGA2 as a potential prognostic biomarker and therapeutic target in osteosarcoma.


Sponastrime dysplasia. A radiologic-pathologic correlation.
The 2nd family with Sponastrime Dysplasia is described. The clinical, radiologic and chondro-osseous morphology of boy and girl siblings are presented. The facial appearance is an "oriental look" with midface hypoplasia and a saddle nose. The radiological findings include the spinal changes of lordosis, osteoporosis and pear-shaped vertebrae, as well as striated metaphyses (osteopathia striata). The morphological findings suggest a disturbance in the formation of cartilage, with a defect in collagen and proteoglycans synthesis in this rare autosomal recessive skeletal dysplasia.


CD3(-)CD16(-)CD56(bright) immunoregulatory NK cells are increased in the tumor microenvironment and inversely correlate with advanced stages in patients with papillary thyroid cancer.
The innate immune system is the first line of defense and plays a key role in thyroid cancer development. The role of the tumor-infiltrating natural killer (NK) cells is becoming increasingly important in research and potential cancer therapies. NK cell subpopulations, CD3(-)CD16(+)CD56(dim) and CD3(-)CD16(-)CD56(bright), demonstrate a significant role in the tumor immuno-surveillance process. We investigated the distribution of CD3(-)CD16(+)CD56(dim) and CD3(-)CD16(-)CD56(bright) NK subpopulations in tissue and blood samples from patients with papillary thyroid cancer (PTC) and nodular goiter (NG). Twenty-eight patients with PTC, 13 patients with NG, and 50 healthy donors were included in the study. Tissue and blood samples from all patients and blood samples from healthy donors were analyzed for CD3(-)CD16(+)CD56(dim) and CD3(-)CD16(-)CD56(bright) NK cells by flow cytometry. A significant predominance of CD3(-)CD16(+)CD56(dim) cells compared to CD3(-)CD16(-)CD56(bright) NK cells was found in blood samples in all groups (p<0.0001 in PTC, NG, and healthy donors). Increased infiltration by CD3(-)CD16(-)CD56(bright) NK cells was observed in thyroid tissue of patients with PTC, as compared to CD3(-)CD16(+)CD56(dim) NK cells (p=0.046), while CD3(-)CD16(+)CD56(dim) NK cells demonstrated a higher infiltration of NG tissues. CD3(-)CD16(+)CD56(dim) NK cell tissue infiltration positively correlated with advanced stages of PTC. In contrast, the CD3(-)CD16(-)CD56(bright) NK cell population was negatively associated with tumor stage in patients with PTC. CD3(-)CD16(-)CD56(bright) NK cell infiltration seems to be associated with PTC progression. These findings contribute to a better understanding of the immune response in PTC and may lead to novel immunotherapeutic approaches in these patients.


Identification of pre-osteonectin produced by cell-free translation of fetal porcine calvarial mRNA.
The cell-free biosynthesis of the bone protein osteonectin was studied using mRNA from fetal porcine calvariae. Total RNA was extracted from the calvariae with guanidinium thiocyanate and was partially purified by precipitation with acid/ethanol. Translations were performed using the reticulocyte lysate system and were optimized with respect to mRNA concentration and K+ (70 mM) and Mg2+ (0.6 mM) concentration. Cell-free synthesized osteonectin, radiolabeled with [35S]methionine, was specifically immunoprecipitated with rabbit antiserum to porcine osteonectin and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and fluorography. When analyzed under reduced conditions, the translated protein migrated with an Mr 45,000 compared to an Mr 39,000 for cell-synthesized osteonectin. When translated in the presence of microsomal membranes, the immunoprecipitated osteonectin co-migrated with the cell-synthesized osteonectin, indicating that a signal sequence of about 45-50 amino acids (Mr 6,000) had been removed. Under nonreduced conditions the pre-osteonectin co-migrated with osteonectin (Mr 39,000) on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, suggesting that a highly folded structure is retained by disulfide bridges under denaturing conditions. The relationship between the immunoprecipitated pre-osteonectin from the cell-free translations and both the cell-synthesized and tissue-extracted osteonectin was confirmed by one-dimensional peptide mapping of Staphylococcus aureus V-8 protease digestions. The results indicate that porcine osteonectin is synthesized on polysomes in a pre-osteonectin form which is translocated vectorially into microsomal vesicles and cotranslationally processed by the removal of a signal peptide.


Norepinephrine turnover in iron deficiency: effect of two semi-purified diets.
The effects of two dietary treatments on norepinephrine turnover in iron deficiency were examined. These studies were designed to bridge the gap between previous studies of poor thermoregulation in iron deficiency which used a diet (HMW, Hubbel-Mendel-Wakeman formulation) relatively high in fat (46% of calories) and moderate in carbohydrate (46% of calories) and the more recent studies of thermogenesis in iron deficiency which use the AIN-76 recommended diet which is relatively low in fat (11% of calories) and high in carbohydrate (67% of calories). Iron deficient rats grew less well and had depressed thyroid hormone concentrations regardless of dietary treatment group. The HMW diet significantly increased norepinephrine turnover in heart in iron deficient animals relative to AIN diet but had no effect in controls. Brown adipose tissue norepinephrine turnover was threefold higher in HMW rats fed a low iron diet, and only 67 percent higher in control rats. This study demonstrates that certain modest macronutrient manipulations affect norepinephrine content and turnover more in iron deficient than controls. However, abnormalities in thyroid hormone concentrations persist in iron deficient animals regardless of these dietary treatments.


Development of PEGylated solid lipid nanoparticles of pentoxifylline for their beneficial pharmacological potential in pathological cardiac hypertrophy.
The present study was designed to develop PEGylated solid lipid nanoparticles (SLNs) of pentoxifylline (with increased t½) for the management of cardiac hypertrophy. PEGylated SLNs were prepared by melt emulsification method and characterised by zeta potential, polydispersity index, entrapment efficiency, in vitro release and pharmacokinetic profile. Hypertrophy was assessed as increase in tumour necrosis factor-alpha, mean arterial blood pressure, LV protein content and left ventricular end diastolic pressure. PEGylated SLNs (F19) with increased t½ were developed and characterised. F19 attenuates hypertrophy assessed in terms of parameters employed. Thus, PEGylated SLNs (F19) of pentoxifylline have enhanced t½ and consequently more significantly preclude cardiac hypertrophy as compared to pentoxifylline.


Different forms of administration of biotherapy 7dH in mice experimentally infected by Trypanosoma cruzi produce different effects.
To evaluate the effects of different forms of administration of the blood trypomastigotes biotherapy 7dH in mice experimentally infected with Trypanosoma cruzi. Male swiss mice were inoculated with 1400 blood trypomastigotes of the Y strain of T. cruzi and allocated into 5 treatment groups: IC (distilled water); TCBZ (benznidazole); TBA(7dH) (biotherapy 7dH 20 days after infection); TBB(7dH)7 (biotherapy 7dH seven days before infection); TBB(7dH)30 (biotherapy 7dH 30 days before infection). Parasitological parameters assessed included pre-patent and patent periods, parasitemia peak, total parasitemia, mortality and survival rates. Cure index was obtained by fresh blood examination, hemoculture and polymerase chain reaction (PCR). The TBB(7dH)7 group showed a reduction in parasitemia peak, parasitemia area under the curve and total parasitemia. TBB(7dH)30 showed a tendency to increased pre-patent and survival periods, peak parasitemia was increased without increased total parasitemia. TBA(7dH) did not present significant alterations in the parasitological parameters analyzed. Biotherapy 7dH given before infection (7 or 30 days) produces different effects suggesting modulation of the host's immune system. The effects range from reduced parasitemia to its effective increase. The use of biotherapy to treat T. cruzi infection including dose, potency and schedule deserves further investigation.


Chromatin Immunoprecipitation (ChIP) to Study the Transcriptional Regulatory Network that Controls Iron Homeostasis in Arabidopsis thaliana.
In plants, gene expression is orchestrated by thousands of transcription factors (TFs). For instance, a large set of bHLH TFs are involved in the regulation of iron homeostasis in Arabidopsis thaliana. The identification of the direct target genes of TFs through uncovering the interaction between the TFs and cis-regulatory elements has become an essential step toward a comprehensive understanding of the iron homeostasis transcriptional regulatory network in Arabidopsis. Chromatin immunoprecipitation (ChIP) followed by qRT-PCR (ChIP-qPCR), sequencing (ChIP-seq), or chip hybridization (ChIP-chip) is a robust tool to investigate protein-DNA interactions in plants in a physiological context. The procedure generally includes six steps: DNA-protein crosslink, isolation of nuclei, shearing of chromatin, immunoprecipitation, DNA purification, and qRT-PCR analyses. In this protocol, we describe guidelines, experimental setup, and conditions for ChIP experiment in Arabidopsis. This protocol focuses on seedlings grown in control and iron deficiency conditions, but can readily be adapted for use with other Arabidopsis tissues or samples. In addition, the protocol could also be applied to perform ChIP-chip or ChIP-seq experiments.


RUNX1/ETO regulates reactive oxygen species (ROS) levels in t(8,21) acute myeloid leukaemia via FLT3 and RAC1.
Reactive oxygen species (ROS) homeostasis is crucial for leukaemogenesisand deregulation would hamper leukaemic progression. Although the regulatory effects of RUNX1/ETO has been extensively studied, its underlying molecular mechanims in ROS production in t(8,21) AML is yet to be fully elucidated. Here, we report that RUNX1/ETO could directly control FLT3 by occupying several DNA elements on FLT3 locus. The possible hijacking mechanism by RUNX1/ETO over FLT3 mediated ROS modulation in AML t(8;21) was made apparent when suppression of RUNX1/ETO led to decrement in ROS levels and the direct oxidative marker FOXO3 but not in FLT3 and RAC1 suppressed t(8,21) AML cell line Furthermore, nuclear import of RUNX1/ETO was aberrated following RUNX1/ETO and RAC1 suppression suggesting association in ROS control. A different picture was depicted in non t(8;21) cells where suppression of RAC1 and FLT3 led to decreased levels of FOXO3a and ROS. Results alltogether indicate a possible dysregulation of ROS levels by RUNX1/ETO in t(8,21) AML.


Abnormal frontostriatal connectivity and serotonin function in gambling disorder: A preliminary exploratory study.
The neurobiological mechanisms of gambling disorder are not yet fully characterized, limiting the development of treatments. Defects in frontostriatal connections have been shown to play a major role in substance use disorders, but data on behavioral addictions, such as gambling disorder, are scarce. The aim of this study was to 1) investigate whether gambling disorder is associated with abnormal frontostriatal connectivity and 2) characterize the key neurotransmitter systems underlying the connectivity abnormalities. Fifteen individuals with gambling disorder and 17 matched healthy controls were studied with resting-state functional connectivity MRI and three brain positron emission tomography scans, investigating dopamine (18F-FDOPA), opioid (11C-carfentanil) and serotonin (11C-MADAM) function. Frontostriatal connectivity was investigated using striatal seed-to-voxel connectivity and compared between the groups. Neurotransmitter systems underlying the identified connectivity differences were investigated using region-of-interest and voxelwise approaches. Individuals with gambling disorder showed loss of functional connectivity between the right nucleus accumbens (NAcc) and a region in the right dorsolateral prefrontal cortex (DLPFC) (PFWE <0.05). Similarly, there was a significant Group x right NAcc interaction in right DLPFC 11C-MADAM binding (p = 0.03) but not in 18F-FDOPA uptake or 11C-carfentanil binding. This was confirmed in voxelwise analyses showing a widespread Group x right NAcc interaction in the prefrontal cortex 11C-MADAM binding (PFWE <0.05). Right NAcc 11C-MADAM binding potential correlated with attentional impulsivity in individuals with gambling disorder (r = -0.73, p = 0.005). Gambling disorder is associated with right hemisphere abnormal frontostriatal connectivity and serotonergic function. These findings will contribute to understanding the neurobiological mechanism and may help identify potential treatment targets for gambling disorder.


[Diabetes increases short- and long-term mortality after transcatheter aortic valve implantation (TAVI)].
Long-term mortality after transcatheter aortic valve implantation (TAVI) in elderly patients with abundant comorbidities is considerable. We aimed to determine the impact of diabetes on short- and long-term mortality after TAVI. Our study includes 300 consecutive patients (mean age, 82 ± 5 years) who underwent TAVI (158 transapical, 142 transfemoral procedures). All patients were followed by regular telephone contacts. 36% suffered from diabetes. Diabetes could be identified as significant predictor of short- and long-term mortality after TAVI. In diabetic patients, 30-day-mortality was 2,5 fold elevated (18.3% vs. 7.3%, p = 0.004). Furthermore, they were at significantly higher risk of peri-interventional stroke (p = 0.04), stage 3 acute kidney injury (p = 0.003), and prolonged ventilation (p = 0.01). Even after successful TAVI and discharge from hospital, long-term mortality was significantly elevated in diabetic patients (56% vs. 30%, p < 0.0001). Of note, 25% of diabetic vs. only 8% of non-diabetic patients died from cardiac causes during follow-up, suggesting that TAVI is not able to reduce cardiac-related mortality risk in diabetic patients to the same extent as in non-diabetics. Diabetes represents a powerful predictor of adverse early and late outcome after TAVI. These findings should be incorporated into the assessment of the risk-to-benefit ratio of TAVI in diabetic patients.


LncRNA HOTAIR promotes α-synuclein aggregation and apoptosis of SH-SY5Y cells by regulating miR-221-3p in Parkinson's disease.
Parkinson's disease (PD) is a common neurodegenerative disease. Here, the purpose of the study was to explore the function of long non-coding RNA (lncRNA) HOX transcript antisense RNA (HOTAIR) in PD and its underlying mechanism. An in vivo 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-hydrochloride (MPTP)-induced mouse model of PD was generated and the SH-SY5Y cells were treated with MPP + to induce neuronal damage in vitro. Quantitative real-time polymerase chain reaction (QRT-PCR) and Western blot were used to detect the expression of HOTAIR, miR-221-3p, α-synuclein and apoptosis-related genes. MTT, flow cytometry and TUNEL assay was used to detect cell viability and apoptosis, respectively. The levels of inflammatory cytokines TNF-α,IL-1β and IL-6 were detected by ELISA assay. The levels of lactate dehydrogenase (LDH), reactive oxygen species (ROS), and superoxide dismutase (SOD) were determined using the appropriate assay kits. The interactions between miR-221-3p and HOTAIR or α-synuclein were determined by dual luciferase assay and RNA binding protein immunoprecipitation (RIP). Co-localization of HOTAIR and miR-221-3p was also proved by immunofluorescence staining. The results showed that HOTAIR was highly expressed, while miR-221-3p expression was decreased in PD model in vivo and in vitro. In SH-SY5Y cells treated with MPP+, the knockdown of HOTAIR increased cell viability and reduced cell apoptosis, the secretion of inflammatory cytokines and oxidative stress reaction, while HOTAIR overexpression led to opposite effects. Furthermore, HOTAIR sponged miR-221-3p which directly targeted α-synuclein and thus regulated the expression of α-synuclein. Meanwhile, inhibiting miR-221-3p could partially reverse the neuroprotective effects of HOTAIR knockdown. In conclusion, HOTAIR attenuated the injury of SH-SY5Y cells induced by MPP+ via miR-221-3p/α-synuclein axis, suggesting the potential therapeutic value of HOTAIR in PD.


The changing role of infection prevention practice as documented by the Certification Board of Infection Control and Epidemiology practice analysis survey.
The Certification Board of Infection Control and Epidemiology (CBIC) appointed an advisory committee to conduct a practice analysis (PA) of infection prevention and control professionals to identify current practices in place. The PA was also used to determine the need for an advanced practice examination. Results of the PA would assist in the development of a revised certification examination. Nine thousand five hundred ninety (9590) e-mail surveys were distributed to infection control professionals (ICPs) in the United States and Canada as well as to a subsample of ICPs in Saudi Arabia. Decision rules and criteria were applied to each identified task in the PA. A total of 1304 responses were available for analysis, representing a response rate of approximately 21.8%. The majority of the respondents was certified in infection control, had a background as a registered nurse, worked in a community hospital, and had from 2 to 6 years experience, a notable change from the 2001 survey. Six major categories with 147 tasks were identified. Based on the 2005 PA results, research tasks were combined with education, which is consistent with the 2001 PA findings. The PA reflects current changes in the practice of infection prevention/control and epidemiology in the United States and Canada. The need for a separate advanced practice examination was not indicated from the survey results. The test specifications accepted for adoption by the CBIC will be used to build all examination forms for the certification program.


Evaluation of multiple parameters of HIV-1 replication cycle in testing of AIDS drugs in vitro.
Evaluation of the activities of antiretroviral agents and an immunoregulatory compound has been made using two models of HIV-1 infection and three measurements of virus expression. Acute infection of Jurkat cells or chronic/inducible infection in U1.1 cells was monitored at multiple time points after drug treatment. The 50% effective concentrations (EC50) of the HIV-1 inhibitors suramin, 3'-azido-3'-deoxythymidine (AZT), and 2',3'-dideoxycytidine, as measured by HIV-1 RNA hybridization in Jurkat cells two days after infection, were comparable to EC50 values obtained in parallel measurements of extracellular p24 levels and percent HIV-1 IF-positive cells. However, these measurements diverged: at seven days after infection the EC50 of AZT was greater than 10 microM when intracellular HIV-1 RNA was assayed, 0.2 microM by IF, and 0.03 microM by p24 assay. Human thymic humoral factor displayed no direct inductive activity in chronic HIV-1 infection in U1.1 cells, while phorbol ester and lymphocyte supernatants induced all parameters. These observations warrant care when interpreting results of only a single assay and suggest that definitive assay of HIV-1 infection requires measurements of multiple parameters of virus expression.


Origin and nature of correlations in the Ia feedback pathway of the muscle control system.
Mammalian primary muscle spindle endings receive two inputs from fusimotor fibres and length changes of their extrafusal environment. Both inputs are also powerful noise sources disturbing the discharge patterns of the receptors. In this paper emphasis is laid on the extrafusal component. It is shown that extrafusal muscle activity can be viewed as a stochastic process which, acting as a common source, can correlate discharge patterns of adjacent muscle spindles. Some quantitative characteristics of these correlations and their relation to the underlying mechanical events are investigated in some detail in order to provide data for more theoretical considerations on their possible physiological importance.


Variations in dissipation rate, microbial function and antibiotic resistance due to repeated introductions of manure containing sulfadiazine and chlortetracycline to soil.
Antibiotic persistence following five successive treatments of sulfadiazine (SDZ) and chlortetracycline (CTC), alone and in combination, in manure-amended soil was studied under laboratory conditions. The resulting effects on soil respiration and enzyme activities as well as pollution-induced community tolerance, were also examined. A trend of initial suppression followed by recovery was observed in the dissipation rates of SDZ or CTC during the antibiotic treatments, and combined treatment with both antibiotics did not alter the respective dissipation rates significantly. Soil respiration activity with SDZ and/or CTC treatments was inhibited during the initial two treatments; however, the activity thereafter recovered to or exceeded the level of the individual manure treatment. Initially, soil urease and dehydrogenase activities were not affected; however, after the fifth treatment, these activities were significantly stimulated in the CTC individual and combined treatments compared with their activities in the individual manure treatment. Bacterial community tolerance to SDZ and CTC in manure-amended soil increased significantly (p⩽0.05) with antibiotic treatment frequency.


Metals doped carbon nanotubes and carbon nanocages (Co2-CNT(8, 0) and Sc2-C78) as catalysts of ORR in fuel cells.
The nano structures (CNT(8, 0) and C78) are doped with two Co and Sc metals in order to obtain the suitable catalysts to process the ORR. The effects of different solvents on ORR on Co-C-C-Co-CNT(8, 0) and Sc-C78 are examined in benzene, methanol and water by polarizable continuum model. The ORR on Co-C-C-Co-CNT(8, 0) and Sc-C-C-Sc-C78 are processed by two paths as follow: *OOH-Co-C-C-Co-CNT(8, 0) → *OH-Co-C-C-Co-CNT(8, 0)-*OH → *OH-Co-C-C-Co-CNT(8, 0)→ *Co-C-C-Co-CNT(8, 0) reactions and *OOH-Co-C-C-Co-CNT(8, 0) → *OH-Co-C-C-Co-CNT(8, 0)-*OH → *OH-Co-C-C-Co-CNT(8, 0)→ *Co-C-C-Co-CNT(8, 0) reactions. Results shown that the Co-C-C-Co-CNT(8, 0) has higher potential than Sc-C-C-Sc-C78 for ORR in benzene, methanol and water. Over potential on Co-C-C-Co-CNT(8, 0) and Sc-C-C-Sc-C78 are lower than various metals based nano catalysts.


Intra- and interobserver reproducibility of left ventricular mechanical dyssynchrony assessment by real time three-dimensional echocardiography.
Real time three-dimensional (3D) echocardiography allows the assessment of left ventricular (LV) mechanical dyssynchrony and may be useful in predicting response to cardiac resynchronization therapy. However, reproducibility of 3D dyssynchrony in past reports varied widely. We evaluated intra- and interobserver reproducibility of parameters of LV mechanical dyssynchrony by 3D echocardiography and explored the impact of image quality as a possible source of variability. LV volumes and ejection fraction (LV EF) were measured by 3D echocardiography in 90 subjects. LV mechanical dyssynchrony was calculated as the standard deviation of the time-to-minimum segmental volume (Tmsv) over 16 (SDI16) and 12 (SDI12) segments and as the maximum time-difference in Tmsv over 16 (Dif16), 12 (Dif12), and 2 (DifS-L) segments. Opposing wall delay in peak myocardial systolic velocity (S-L delay) by tissue Doppler imaging (TDI) was also evaluated. Feasibility of 3D measurements was 88.9%. Intra- and interobserver intraclass correlation coefficients were excellent for LV volumes, LV EF, and S-L delay (all ≥ 0.90), very good for SDI16 (0.83 and 0.85), moderate to good for SDI12, Dif16, Dif12, and DifS-L (0.51-0.81). No systematic bias was present between readings for 3D dyssynchrony, but limits of agreement resulted fairly large for most parameters with the exception of SDI16, which showed reproducibility slightly lower than TDI. Suboptimal image quality was a significant source of variability in 3D-dyssynchrony assessment. The 3D assessment of LV volumes and dyssynchrony was feasible in our community-based cohort. SDI16 was the most reproducible among 3D-dyssynchrony parameters and the least affected by image quality.


Mediastinal mass following successful chemotherapy for ovary dysgerminoma: benign process or disease relapse? A case report.
Ovarian dysgerminoma is a rare tumor that affects adolescent girls and young women. Due to its high radio-chemosensitivity, prognosis is normally excellent. Relapses occur in less than 20% of early stage disease, but are more frequent in advanced disease. It is known that some benign mediastinal processes may mimic tumor relapse, particularly in young patients. This is the case of physiologic thymic hyperplasia, which occurs as a rebound phenomenon after chemotherapy in young women with ovarian dysgerminoma. Until now, no cases of dysgerminoma with benign mediastinal mass have been published. A young woman with bulky ovarian dysgerminoma, who obtained complete disease remission after chemotherapy, subsequently developed a mediastinal mass which was initially confused with a mediastinal relapse. CT scan features (close thymic location, homogeneous hypodensity, absence of infiltration of mediastinal structures) and subsequent PET/CT scan (homogeneous glucose uptake and a typical inverted V morphology) supported the diagnosis of thymic hyperplasia. No further invasive procedures were performed. 34 months from the diagnosis the patient is in good physical condition with no signs of relapse. Our case underlines the importance of knowing the age- and treatment-related incidence of physiologic thymic hyperplasia in young women with ovarian dysgerminoma in order to reduce the potential pitfalls and to avoid unnecessary invasive diagnostic procedures.


A fetal skeletal muscle actin mRNA in the mouse and its identity with cardiac actin mRNA.
We compare a recombinant cDNA plasmid (pAF81) complementary to a fetal skeletal muscle actin mRNA with a plasmid (pAM91) complementary to the actin mRNA expressed in adult skeletal muscle. The two mRNAs are significantly diverged in silent nucleotide positions; they are coexpressed in fetal skeletal muscle, and in differentiating muscle cell cultures their accumulation begins coordinately. The sequence of pAF81 shows that the amino acid sequence of mouse fetal skeletal muscle actin is almost identical to that of adult bovine cardiac actin. Hybridization of pAF81 to RNA from different mouse tissues shows that fetal skeletal muscle actin mRNA is very homologous or identical to fetal and adult cardiac actin mRNA. Only one gene homologous to pAF81 is detected on blots of restricted mouse DNA. We conclude that this gene must be expressed both in fetal skeletal muscle and in fetal heart. Whereas mRNA transcribed from this gene is the major actin mRNA species in adult heart, it is present in low amounts, if at all, in adult skeletal muscle.


The calciuria of increased fixed acid production in humans: evidence against a role for parathyroid hormone and 1,25(OH)2-vitamin D.
We measured mineral and acid balances, serum iPTH, urinary cAMP/creatinine, and plasma concentrations of 25OHD and 1,25(OH)2D in 7 healthy adults during control conditions and during increased fixed acid production achieved either by the administration of NH4Cl (N = 3) or by increased dietary protein intake (N = 4). When acid production was increased, the subjects were in positive acid balance and negative Ca balance because of increased urinary Ca excretion. Serum iPTH fell slightly but urinary cAMP and the plasma levels of vitamin D metabolites did not change. W conclude that the accelerated skeletal and urinary losses of Ca that occur when fixed acid production is increased are not contributed to nor compensated for by the parathyroid-vitamin D endocrine systems.


Clinical evaluation of the effect of Zataria multiflora Boiss and Trachyspermum copticum (L.) on the patients with irritable bowel syndrome.
Irritable Bowel Syndrome (IBS) is the most common gastrointestinal dysfunction characterized by abdominal pain or discomfort, as well as changes in bowel movements and stool shape. Given the global trend towards the increased use of complementary and alternative medicine (CAM), the high prevalence of IBS, the lack of a standard treatment for all IBS subtypes, and patients' willingness to use CAM treatments, investigations into CAM treatments are needed. Accordingly, the present study aimed to investigate the effect of a mix of two herbal medicines (Zataria-Trachyspermum, ZT) on the clinical symptoms of patients with IBS. The present study was performed on 3 groups including the positive control, experimental, and placebo groups. The sample size was calculated as 150 participants. Fifty patients were assigned to one of three parallel groups (ZT capsule, placebo, and mebeverine capsule) by block randomization. All three groups were treated for 4 weeks. The patients were monitored in the follow-up stage for 2 additional weeks. After the fourth week of intervention, symptoms of pain, bloating, and reflux showed a significant decrease in the ZT group compared to the placebo and mebeverine groups (P <0.05). Moreover, the participants in the ZT group reported a significant decrease in fatigue compared to the other two groups (P <0.05). This study confirmed the positive effect of ZT on IBS symptoms, especially pain, bloating, constipation, and bowel movements.


Nonsustained ventricular tachycardia in heart failure with preserved ejection fraction.
Ventricular tachycardia (VT) is a common arrhythmia in heart failure with reduced ejection fraction but its incidence, predictors, and significance have not been determined in heart failure with preserved ejection fraction (HFpEF). We performed a retrospective review of arrhythmias in two cohorts of patients with an HFpEF diagnosis. Patients in cohort 1 (n = 40) underwent routine arrhythmia surveillance with a 14-day ambulatory electrocardiogram (ECG) monitor. Patients in cohort 2 (n = 85) had cardiac pacemakers and underwent routine device interrogations. In cohort 1, 13 patients (32.5%) had one or more episodes of nonsustained VT (NSVT) on ambulatory ECG. In cohort 2, 38 patients (44.7%) had NSVT on cardiac pacemaker interrogations. During a median (interquartile range) follow-up of 3.0 (1.6 to 5.1) years, 15 (12%) patients died (20% of patients with NSVT versus 6.8% of those without NSVT; P = .03). In logistic regression analysis, NSVT was associated with a 3.4-fold higher odds of death (95% confidence interval 1.08 to 10.53; P = .04) in HFpEF. In conclusion, patients with HFpEF have a relatively high, and possibly underappreciated, burden of NSVT, which confers a higher risk of mortality. The frequent episodes of NSVT in these patients may provide insight into the mechanism of sudden cardiac death in HFpEF.


Adult testicular granulosa cell tumour: A rare case report.
Adult Granulosa cell tumours (AGT) are a rarely reported subtype of testicular cancer with an unpredictable clinical course. We report the case of a 45 year old man who was diagnosed with a 15mm testicular adult granulosa cell tumour with no metastatic disease at presentation. A radical inguinal orchidectomy was performed with histology demonstrating no invasion of the tunica vaginalis, epididymis, spermatic cord and no lymphovascular invasion. No further treatment was undertaken due to good prognostic factors. A follow up CT Thorax, Abdomen, Pelvis at 4 months demonstrated no disease progression.


Variation of serum hyaluronan with activity in individuals with knee osteoarthritis.
Serum hyaluronan (HA) was evaluated for diurnal variation in participants with osteoarthritis (OA) of the knee. Twenty participants with radiographic OA of at least one knee were admitted overnight to the General Clinical Research Center for serial serum sampling. Serum was obtained between 6:00 p.m. and 8:00 p.m. on Day 1 (T3) after a day of normal activity. During the night of bed rest, participants remained supine for a minimum of 5 h between the hours of 3:00 a.m. and 8:00 a.m. Blood was drawn prior to arising from bed (T0), and 1h (T1) and 4 h (T2) after arising and performing usual morning activities, including eating breakfast. During the morning, participants were encouraged to remain physically active and were not permitted to sit for more than 30 min at a time. Serum HA was measured by enzyme-linked immunosorbent assay. Results were analyzed using non-parametric Freidman's test with Dunn's post-hoc Multiple Comparison test. Serum levels of HA increased significantly from T0 to T1 (P < 0.01). There were no other significant changes in serum HA levels observed between any of the other time points. Although a rise in serum HA with activity and eating has been demonstrated previously in individuals with rheumatoid arthritis, this is the first study to demonstrate a similar rise in individuals with OA. These results suggest that serum sampling for HA in OA clinical trials should be performed more than 1h after arising in the morning and at least 1h after breaking an overnight fast.


Juvenile myelomonocytic leukemia displays mutations in components of the RAS pathway and the PRC2 network.
Juvenile myelomonocytic leukemia (JMML) is a rare and severe myelodysplastic and myeloproliferative neoplasm of early childhood initiated by germline or somatic RAS-activating mutations. Genetic profiling and whole-exome sequencing of a large JMML cohort (118 and 30 cases, respectively) uncovered additional genetic abnormalities in 56 cases (47%). Somatic events were rare (0.38 events/Mb/case) and restricted to sporadic (49/78; 63%) or neurofibromatosis type 1 (NF1)-associated (8/8; 100%) JMML cases. Multiple concomitant genetic hits targeting the RAS pathway were identified in 13 of 78 cases (17%), disproving the concept of mutually exclusive RAS pathway mutations and defining new pathways activated in JMML involving phosphoinositide 3-kinase (PI3K) and the mTORC2 complex through RAC2 mutation. Furthermore, this study highlights PRC2 loss (26/78; 33% of sporadic JMML cases) that switches the methylation/acetylation status of lysine 27 of histone H3 in JMML cases with altered RAS and PRC2 pathways. Finally, the association between JMML outcome and mutational profile suggests a dose-dependent effect for RAS pathway activation, distinguishing very aggressive JMML rapidly progressing to acute myeloid leukemia.


The surface electrocardiogram in complete intra-his heart block.
In some cases, the surface ECG may be useful in predicting that the level of heart block is intra-His rather than supra-His. In five patients with third degree heart block the level of block was documented to be within the His bundle. In all five, the surface ECG showed an escape rhythm with QRS complexes of less than 0.10s. duration. The initial QRS forces were abnormal in that "septal" q waves were not present in leads 1 and V6. Two of the five patients had occasional conducted beats with normal "septal" q waves. Thus, heart block associated with an escape rhythm of narrow QRS morphology but abnormal septal depolarization may indicate that the level of the block is intra-His rather than supra-His.


Continuing education for registered nurses in the Highlands and Western Isles of Scotland.
This paper focuses on an educational approach used in the Highlands and Western Isles of Scotland in the continuing education of registered nurses. It includes a rationale for the educational approach used by this author and her colleagues, and a discussion of the approach itself. The discussion includes reference to concepts borrowed from learning theorists that underpin the approach, and case study material to illustrate how it works in practice. It is argued that the educational approach in this paper can potentially lead to meaningful learning. Meaningful learning is defined in this context. Finally, the outcomes of the educational approach are highlighted, with a discussion of its value to educators elsewhere.


Distribution of genotypes of F-specific RNA bacteriophages in human and non-human sources of faecal pollution in South Africa and Spain.
To assess whether the distribution of genotypes of F-specific RNA bacteriophages reflects faecal pollution of human and animal origin in water environments. Stool samples, animal feedlot waste slurries and a wide variety of faecally polluted waters were studied in South Africa and Spain. Genotyping was performed by plaque and spot hybridization with genotype-specific probes. Only genotypes II and III were detected in human stool. Animal faeces contained predominantly, but not exclusively, genotypes I and IV. Raw hospital and municipal sewage contained mostly genotypes II and III, whereas genotypes I and II prevailed in settled sewage, secondary treated sewage and non-point diffuse effluents from developing communities. Abattoir wastewaters contained mostly genotypes I and IV. No differences were observed between the distribution of genotypes in Spain and South Africa. Although the association of genotypes II and III with human excreta and I and IV with animal excreta was statistically significant, the results suggest that the association cannot be used for absolute distinction between faecal pollution of human and animal origin. This study contributes greatly to understanding the usefulness of genotypes of F-specific RNA bacteriophages in source tracking of faecal wastes.


A phenomenological neural network powered by the National Wastewater Surveillance System for estimation of silent COVID-19 infections.
Although wastewater-based epidemiology (WBE) has emerged as an inexpensive and non-intrusive method in contrast to clinical testing to track public health at community levels, there is a lack of structured interpretative criteria to translate the SARS-CoV-2 concentrations in wastewater to COVID-19 infection cases. The difficulties lie in the uncertainties of the amount of virus shed by an infected individual to wastewater as documented in clinical studies. This situation is even worse considering the existence of a population of silent infections and many other confounding factors. In this research, a quantitative framework of a phenomenological neural network (PNN) was developed to compute silent infections. The PNN was trained using the WBE data from the National Wastewater Surveillance System (NWSS) - a program launched by the CDC of the United States in 2020. It is found that the PNN excelled with superior interpretability and reduced overfitting. A big-data perspective on virus shedding by an infected population revealed more deterministic virus-shedding dynamics compared to the clinical studies perspective on virus shedding by an infected individual. With such characteristics employed as the theoretical basis for the estimation of the silent infections, a ratio of silent to reported infections was found to be 5.7 as the national median during the studied period. The study also noted the influence of temperature, sewershed population, and per-capita flow rates on the computation of silent infections. It is expected that the proposed framework in this work would facilitate public health actions guided by the SARS-CoV-2 concentrations in wastewater. In case of a new wave emergence or a new virus disease outbreak like COVID-19, the PNN powered by the NWSS would outline consolidated and systematic information that would enable rapid deployment of public health actions.


Effect of the interaction between fibronectin and VLA-4 on the proliferation of human B cells, especially a novel human B-cell line, OPM-3.
Very late antigen (VLA)-4 integrin has been suggested to play an important role in haemopoiesis. However, little is known concerning the roles of the fibronectin (FN)/VLA-4 interaction in the proliferation of human B cells. In this study we investigated the effect of immobilized FN on the proliferation of various B-cell lines, including a newly-established B-cell line, OPM-3, and human tonsillar B cells, that primarily express VLA-4 but not VLA-5. Immobilized FN significantly promoted the proliferation of OPM-3 cells and normal B cells via VLA-4. The cross-linking of beta1 integrins of OPM-3 cells resulted in the phosphorylation of the focal adhesion kinase (FAK) associated 90 kD protein, an increase in FAK-associated kinase activity, and the phosphorylation of Raf-1. Furthermore, the MEK1 inhibitor, PD98059, inhibited the FN-promoted proliferation of OPM-3 cells. These results demonstrate that the FN/VLA-4 interaction transmits the growth signal(s) which may be mediated by Ras pathway in OPM-3 cells, and suggest that OPM-3 cells may be of great value in studying the roles of the FN/VLA-4 interaction in human B-cell growth.


Meso-scale testing and development of test procedures to maintain mass balance.
The Conrad Blucher Institute for Surveying and Science (Texas A&M University--Corpus Christi) has conducted numerous petroleum experiments at the Shoreline Environmental Research Facility (Corpus Christi, Texas, USA). The meso-scale facility has multiple wave tanks, permitting some control in experimental design of the investigations, but allowing for real-world conditions. This paper outlines the evolution of a materials balance approach in conducting petroleum experiments at the facility. The first attempt at a materials balance was during a 1998 study on the fate/effects of dispersant use on crude oil. Both water column and beach sediment samples were collected. For the materials balance, the defined environmental compartments for oil accumulation were sediments, water column, and the water surface, while the discharge from the tanks was presumed to be the primary sink. The "lessons learned" included a need to quantify oil adhesion to the tank surfaces. This was resolved by adhering strips of the polymer tank lining to the tank sides that could be later removed and extracted for oil. Also, a protocol was needed to quantify any floating oil on the water surface. A water surface (oil slick) quantification protocol was developed, involving the use of solid-phase extraction disks. This protocol was first tested during a shoreline cleaner experiment, and later refined in subsequent dispersant effectiveness studies. The effectiveness tests were designed to simulate shallow embayments which created the need for additional adjustments in the tanks. Since dispersant efficacy is largely affected by hydrodynamics, it was necessary to scale the hydrodynamic conditions of the tanks to those expected in our prototype system (Corpus Christi Bay, Texas). The use of a scaled model permits the experiment to be reproduced and/or evaluated under different conditions. To minimize wave reflection in the tank, a parabolic wave dissipater was built. In terms of materials balance, this design reduced available surface area as a sink for oil adsorption.


Draft Genome Sequence of Clostridium aceticum DSM 1496, a Potential Butanol Producer through Syngas Fermentation.
Clostridium aceticum DSM 1496 is a Gram-negative anaerobic chemolithoautotrophic acetogenic bacterium that is capable of producing commodity chemicals from syngas fermentation. In this study, we report the draft genome sequence of the C. aceticum DSM 1496 strain (4.16 Mb) to elucidate the syngas fermentation metabolic pathway.


The incidence of acute renal failure and high mortality rate in elderly patients hospitalized with community acquired pneumonia.
Community acquired pneumonia is associated with high mortality and health care costs, especially in old age. The clinical presentation of pneumonia in the elderly may be asymptomatic or atypical. One of the known complication is an acute kidney injury. The purpose of our study was to estimate the incidence of this complication in elderly patients hospitalized with pneumonia in our geriatric hospital. From a group of 180 elderly patients hospitalized with community-acquired pneumonia 34.4 % developed acute kidney injury. In this group, 51.6 % of patients died compared to 14.4 % in the group of patients without acute kidney injury (p < 0.001). The lower level of e-GFR was significantly associated with mortality (p < 0.001): out of seven patients with e-GFR level of 15-29 mg/mmol, five patients died (71.4 %). Elderly patients with community-acquired pneumonia suffering acute kidney injury experienced worse in-hospital outcomes; mortality rate was significantly higher in our study. We found a relationship between low level of e-GFR and mortality. Clinicians should be alert for early detection and prevention of kidney injury in patients admitted with pneumonia.


Radiotherapy plus immune checkpoint inhibitor in prostate cancer.
The immune checkpoint inhibitor (ICI) is a promising strategy for treating cancer. However, the efficiency of ICI monotherapy is limited, which could be mainly attributed to the tumor microenvironment of the "cold" tumor. Prostate cancer, a type of "cold" cancer, is the most common cancer affecting men's health. Radiotherapy is regarded as one of the most effective prostate cancer treatments. In the era of immune therapy, the enhanced antigen presentation and immune cell infiltration caused by radiotherapy might boost the therapeutic efficacy of ICI. Here, the rationale of radiotherapy combined with ICI was reviewed. Also, the scheme of radiotherapy combined with immune checkpoint blockades was suggested as a potential option to improve the outcome of patients with prostate cancer.


Monitoring PRRSV-1 in suckling piglets in an endemic herd using reverse transcriptase quantitative real time polymerase chain reaction: comparison of the rate of detection in serum and oral fluid samples and evaluation of pooling.
Defining shedding and exposure status for PRRSV is essential in herd stabilisation protocols and weaning-age pigs is a key subpopulation. Oral fluid (OF) sampling is a welfare-friendly and cost saving promising alternative to blood sampling. The first objective of our study was to compare the rate of detection of PRRSV-1 in individual serum sample, individual OF sample, litter-based OF sample, collected the day before weaning. The second objective was to evaluate the interest of pooling samples. The study was performed on a 210-sows, PRRSV-1 exposed, with confirmed shedding, non-vaccinated against PRRSV, herd. 80 litters were sampled and 26 were viropositive and therefore included. The rate of detection of PRRSV-1 with RT-qrtPCR in blood samples, iOF and cOF was 67, 23 and 77%, respectively. The Ct values from RT-qrtPCR on collective OF were statistically lower if the serum of the piglet of the litter was positive. The lower the Cycle threshold (Ct) value of RT-qrtPCR on collective OF, the higher the probability that the serum sampled in the same litter was positive. Ability to detect PRRSV RNA after pooling was 67% for sera and 58% for cOF. The rate of detection of PRRSV-1 was about the same in cOF and blood samples. Virus sequencing, if required, should be performed on individual serum samples. The smaller the Ct of a cOF sample from a litter, the greater the likelihood that the serum sample from a piglet of that litter is positive.A cost-effective and representative sampling protocol to monitor sow herds stabilisation of a sow batch could be: to collect both cOF and one serum sample per litter; to perform firstly RT-qrtPCR on pooled cOF; in case of negative results to consider the batch negative; in case of positive results in a unvaccinated herd or a killed vaccine vaccinated one to consider the batch positive; in case of positive result in a herd vaccinated with a modified live vaccine serum samples of litters with positive cOF should be tested for sequencing (selecting the litters with the lowest Ct for cOF).


Imaging voltage and brain chemistry with genetically encoded sensors and modulators.
Neurons and glia are functionally organized into circuits and higher-order structures that allow the precise information processing required for complex behaviors. To better understand the structure and function of the brain, we must understand synaptic connectivity, action potential generation and propagation, as well as well-orchestrated molecular signaling. Recently, dramatically improved sensors for voltage, intracellular calcium, and neurotransmitters/modulators, combined with advanced microscopy provide new opportunities for in vivo dissection of cellular and circuit activity in awake, behaving animals. This review focuses on the current trends in genetically encoded sensors for molecules and cellular events and their potential applicability to the study of nervous system in health and disease.


Transcription factors as tools for metabolic engineering in plants.
The functions of an increasing number of plant transcription factors are being elucidated, and many of these factors have been found to impact flux through metabolic pathways. Because transcription factors, as opposed to most structural genes, tend to control multiple pathway steps, they have emerged as powerful tools for the manipulation of complex metabolic pathways in plants. The review describes the highlights of recent experiments that have targeted transcription factors that control plant metabolic pathways, and discusses their potential as tools for metabolic engineering.


Relatives' experiences of the hospitalisation of older people with COVID-19: A qualitative interview study.
During the COVID-19 pandemic, visitors were restricted from hospitals, separating them from hospitalised friends and family to reduce the infection risk. The objective was to explore how relatives of older people acutely admitted to hospital with COVID-19 experienced being a relative, and how they felt about their contact with health care professionals (HCPs) when visitor restrictions prevented their physical presence in the ward. This study employed a qualitative design. We used individual qualitative semi-structured interviews and the participants were relatives of acutely admitted older people from three COVID-19 wards in Denmark. A total of 18 relatives participated, 14 female and 4 male, aged between 45 and 83 years. The analysis was guided by Graneheim and Lundman's qualitative content analysis. The analysis derived the following three themes: (1) the importance of trust in a period of uncertainty; (2) the meaning of contact with HCPs, and (3) active but at a distance-a balancing act. The participants' feelings of uncertainty were prominent. The unknown nature of the disease and the unusual situation challenged relatives' trust in HCPs and the health care system. The findings highlight relatives' stress when the possibilities for visiting are restricted and the importance of trust in, and the relationship with HCPs. This study can strengthen HCPs' understanding of relatives' situation when older people are hospitalised during and after the COVID-19 pandemic.


Stability properties of sperm whale oxymyoglobin.
Sperm whale oxymyoglobin was isolated directly from muscle and was examined for its stability properties over the wide range of pH 5-13 in 0.1 M buffer at 25 degrees C. The remarkable pH dependence for the autoxidation rate was analyzed using the kinetic equation derived in terms of nucleophilic displacement processes of O-2 from oxymyoglobin by the entering water molecule or hydroxyl ion with the iron resulting in the ferric form. Most of the autoxidation reaction of the oxymyoglobin can be best explained by the proton-catalyzed processes involving the distal histidine as the catalytic residue. The kinetic equation could also be used as an interesting diagnostic probe into differences in the heme reactivity and the heme environment of different types of oxymyoglobin from other sources.


A combined analysis of data to identify predictive factors for spermatogenesis in men with hypogonadotropic hypogonadism treated with recombinant human follicle-stimulating hormone and human chorionic gonadotropin.
To compare the efficacy and safety of recombinant human FSH (r-hFSH) and hCG treatment for male hypogonadotropic hypogonadism (HH) in different populations and to identify characteristics predictive of spermatogenesis. A combined analysis of data from four clinical trials. Phase III, open-label, noncomparative studies with similar designs conducted in Australia, Europe, Japan, and the United States. One hundred men with complete idiopathic or acquired HH. Pretreatment with hCG for 3-6 months, followed by combination therapy with hCG and r-hFSH (150 IU three times weekly) for up to 18 months. Doses of r-hFSH were adjusted according to spermatozoa count until the maximum dose was reached. The primary efficacy endpoint was a spermatozoa concentration of >or=1.5 x 10(6)/mL. A total of 81 men remained azoospermic but achieved normal serum T concentrations after hCG pretreatment. Of these, 68 (84.0%) achieved spermatogenesis and 56 (69.1%) achieved spermatozoa concentrations >or=1.5 x 10(6)/mL. Large baseline mean testicular volume, low body mass index, and advanced sexual maturity were predictors of good response to therapy. Similar treatment responses were observed across different study populations. R-hFSH (combined with hCG) is effective for the restoration of fertility in the majority of men with HH.


Endoscopic injection sclerotherapy of esophageal varices. Considerations on the first 85 patients.
Eighty-five consecutive cases of esophageal varices treated by endoscopic sclerotherapy are analyzed retrospectively. Most of these cases (84.7%) were done in emergency in patients (60) with poor liver function. Bleeding was stopped in 94.4% of cases but 22% of them had at least one rebleeding episode. Only a few and minor complications were observed. The safety and the immediate good results obtained by the procedure recommend it as a primary measure in bleeding esophageal varices. Moreover the actual role of endoscopic sclerotherapy as an elective or prophylactic procedure in the treatment of hemorrhage from esophageal varices, is discussed.


Partnerships in primary care in Australia: network structure, dynamics and sustainability.
Partnerships represent a prescriptive form of network governance, based on the idea of cooperation. This article has four aims. The first is to describe why network governance and partnerships are important now, and what one particular example - Primary Care Partnerships - is addressing. The second is to analyse the network structure of two of these partnerships, and the third is to examine network dynamics. The fourth aim is to explore relationships and sustainability over the longer term. Two government-funded and steered partnerships, which were established to increase coordination between primary care services in Victoria, Australia, were examined. Annual interviews at three points in time between 2002 and 2005 were used to explore relationships between organizations within these two partnerships. The structure of two different communication networks, based on contacts for work and contacts for strategic information, were examined using social network analysis. Tracing network structures over time highlighted partnership dynamics. The network structures changed over the three years of the study, but an important constant was the continuing centrality of the independent staff employed to manage the partnerships. Over the longer term, it seems to be more important to fund independent partnership staff, rather than people who connect partnerships to the funding agency. If partnerships are seen as valuable in improving service coordination and health outcomes, then long term rather than just start-up funding support is required.


Multi-biomarker responses to pesticides in an agricultural population from Central Brazil.
We evaluated farmworkers exposed to pesticides and individuals with no history of occupational exposure to pesticides. It was performed the comet assay to evaluate DNA damage. The immunophenotyping of TCD4+ lymphocyte subpopulations in peripheral blood was performed by flow cytometry. The single nucleotide polymorphisms (SNPs) in PON1, XRCC1, IL6, IL6R, TNF-α, and MIR137 genes were evaluated by real-time PCR. The exposed group was composed mostly by males (69.44%), with direct exposure to pesticides (56%) and with an average age range of 46 ± 13.89 years, being that 58.3% of farmworkers directly exposed to pesticides and reported the full use of personal protective equipment (PPE). DNA damage was greater in the exposed group (p < 0.05), reinforced by the use of PPE to denote a lower degree of DNA damage (p = 0.002). In this context, in the exposed group, we demonstrated that the use of PPE, age, gender and intoxication events were the variables that most contributed to increase DNA damage (p < 0.0001). Besides, the exposed group showed a significant increase in the subpopulations of T lymphocytes CD3+CD4+ (p < 0.05) and CD3+CD4+CD25+ (p < 0.0001) and a significant decrease in CD3+CD4+CD25-FOXP3+ (p < 0.05). SNPs in the TNF-α (rs361525) gene presented a difference in the genotype distribution between the groups (p = 0.002). The genotype distribution of TNF-α (rs361525) was also positively correlated with the DNA damage of the exposed group (r = 0.19; p = 0.01), demonstrating a higher risk of DNA damage in the farmworkers presenting the A mutated allele. Our findings demonstrate that pesticides can exert various deleterious effects on human health by damaging the DNA as well as by influencing the immune system in the case of both direct or indirect exposure and these issues are associated to age, gender, intoxication and the nonuse of PPE.


Familial bilateral combined hamartoma of retina and retinal pigment epithelium associated with neurofibromatosis 1.
We report a family of three siblings followed between 2005 and 2011 with bilateral combined hamartoma of the retina and retinal pigment epithelium, with the age of diagnosis ranging from 7 to 13 years. The main reason for consultation was reduction of vision and squint. The diagnosis was determined based on the clinical findings on fundus examination: increased pigmentation at the macula with slightly elevated, gray-white macular lesion, tortuosity of perimacular blood vessels and glial epiretinal membrane. The elder brother was found to have left posterior subcapsular cataract. He was also confirmed to have neurofibromatosis type 1, the youngest sister fit in the diagnostic criteria for neurofibromatosis type 1, while the middle sister was presumed to have neurofibromatosis type 1. Follow-up showed stability of the retinal lesion in the three cases, with the progression to develop right posterior subcapsular cataract in the elder sister. This report is aimed to demonstrate that the occurrence of bilateral combined hamartoma of the retina and retinal pigment epithelium could raise the possibility of associated neurofibromatosis.


Clinical tests for confirming tracheal intubation or excluding oesophageal intubation: a diagnostic test accuracy systematic review and meta-analysis.
Unrecognised oesophageal intubation causes preventable serious harm to patients undergoing tracheal intubation. When capnography is unavailable or doubted, clinicians still use clinical findings to confirm tracheal intubation, or exclude oesophageal intubation, and false reassurance from clinical examination is a recurring theme in fatal cases of unrecognised oesophageal intubation. We conducted a systematic review and meta-analysis of the diagnostic accuracy of five clinical examination tests and the oesophageal detector device when used to confirm tracheal intubation. We searched four databases for studies reporting index clinical tests against a reference standard, from inception to 28 February 2023. We included 49 studies involving 10,654 participants. Methodological quality was overall moderate to high. We looked at misting (three studies, 115 participants); lung auscultation (three studies, 217 participants); combined lung and epigastric auscultation (four studies, 506 participants); the oesophageal detector device (25 studies, 3024 participants); 'hang-up' (two non-human studies); and chest rise (one non-human study). The reference standards used were capnography (22 studies); direct vision (10 studies); and bronchoscopy (three studies). When used to confirm tracheal intubation, misting has a false positive rate (95%CI) of 0.69 (0.43-0.87); lung auscultation 0.14 (0.08-0.23); five-point auscultation 0.18 (0.08-0.36); and the oesophageal detector device 0.05 (0.02-0.09). Tests to exclude events that invariably lead to severe damage or death must have a negligible false positive rate. Misting or auscultation have too high a false positive rate to reliably exclude oesophageal intubation and there is insufficient evidence to support the use of 'hang-up' or chest rise. The oesophageal detector device may be considered where other more reliable means are not available, though waveform capnography remains the reference standard for confirmation of tracheal intubation.


Indian herb 'Sanjeevani' (Selaginella bryopteris) can promote growth and protect against heat shock and apoptotic activities of ultra violet and oxidative stress.
Selaginella bryopteris is a lithophyte with remarkable ressurection capabilities. It is full of medicinal properties, hence also known as 'Sanjeevani' (one that infuses life). For lack of credible scientific evidence the plant is not in active use as a medicinal herb. We provide scientific evidence for why S. bryopteris is known as 'Sanjeevani'. The aqueous extract of S. bryopteris possesses growth-promoting activity as well as protective action against stress-induced cell death in a number of experimental cell systems including mammalian cells. Treatment of the cells in culture with 10% aqueous extract enhanced cell growth by about 41% in Sf9 cells and 78% in mammalian cells. Pre-treatment of cells with the Selaginella extract (SE) (1-2.5%) protected against oxidative stress (H2O2) -induced cell death. The killing potential of ultra violet (UV) was also significantly reduced when the cells were pre-treated with SE for 1 h. Thermal radiation suppressed cell growth by about 50%. Pre-treatment of cells with SE for 1 h afforded complete protection against heat-induced growth suppression. SE may possess anti-stress and antioxidant activities that could be responsible for the observed effects. Chemical analysis shows that SE contains hexoses and proteins. Taken together, S. bryopteris extract may help in stress-induced complications including those due to heat shock.


Environmentally friendly dyeing of wool and silk fabrics with natural colorant of turnsole (Chrozophora tinctoria L.).
With the increasing demand for environmentally friendly dyes, natural dye has become an attractive option replacing toxic synthetic dyes. Within the framework of this research, the natural dye of Chrozophora tinctoria L. was extracted using the microwave-assisted extraction method under different operating conditions. The influence of different factors influencing the extraction process (such as pH, extraction time, and microwave power) on the extraction efficiency was investigated by means of a multilevel three-factor design. The suggested model has a high coefficient of determination, indicating high predictability and good approximation. Optimal microwave extraction process conditions were obtained at a value of microwave power equal to 650 W, at a pH of 5 during a period of 3 min. Dyed wool and silk fabrics showed very good washing and light fastness properties.


Immunophenotypic characterization of lymphoid cell infiltrates in vitiligo.
The pathogenesis of vitiligo is still controversial. The purpose of this study was to gain insight into the nature of lymphoid cells infiltrating depigmented areas of skin in vitiligo. Immunochemical procedures were carried out in biopsies from 20 patients with active lesions to search for cells expressing CD1a, CD2, CD3, CD4, CD5, CD8, CD20, CD25, CD30, CD56, CD68 and CD79a. Results indicate that early lesions are infiltrated mainly by dendritic cells, whereas older lesions display significantly lower proportions of these cells and increased percentages of mature T cells. This finding might suggest that the autoimmune reactivity towards melanocyte antigens might be T cell-dependent and antigen-driven. It is possible that a non-immune offence of melanocytes is responsible for the exposure of intracellular antigens, while autoreactivity might be a secondary, self-perpetuating mechanism.


Effectiveness of combined microfocused ultrasound with visualization and subdermal calcium hydroxyapatite injections for the management of brachial skin laxity.
There is no publication to date on the combined use of microfocused ultrasound with visualization (MFU-V) and calcium hydroxylapatite (CaHA) for brachial skin laxity. To assess the effectiveness of combining MFU-V with diluted/hyperdiluted CaHA in a single session for treating brachial skin laxity. Female subjects who had skin laxity in the brachial regions and who desired non-surgical intervention were enrolled into this prospective, single-arm pilot study. MFU-V (Ultherapy® , Merz North America, Inc. Raleigh, N.C.) was applied using the 4.0 MHz-4.5 mm and 7.0 MHz-3.0 mm depth transducers, followed by subdermal injections of diluted (1:1)/hyperdiluted (1:2) CaHA (Radiesse® , Merz North America, Inc). Subjects were followed for six months after treatment. Objective biophysical skin assessments were conducted using a cutometer (Cutometer® Dual 580 MPA; Courage & Khazaka, Cologne, Germany). Subjective assessments included the arm visual analogue scale (VAS), global aesthetic improvement scale (GAIS), and subject global satisfaction scale. Twelve subjects participated in the study. The mean R0 reading (measure of skin firmness) progressively improved from 0.515 mm at baseline to 0.433 mm at 24 weeks (p < 0.05 for 12 and 24 weeks). The mean R2 reading (measure of skin elasticity) and mean arm VAS improved significantly from baseline at all visits (p < 0.05 for all). The majority of subjects at each visit showed improved arm appearance and were satisfied with their treatment. Both procedures were well-tolerated. Combined use of MFU-V with diluted/hyperdiluted CaHA demonstrates significant improvements in both objective and subjective measures of brachial skin laxity.


Calcitonin gene-related peptide II, substance P and vasoactive intestinal peptide in plasma and synovial fluid from patients with inflammatory joint disease.
Immunoreactive plasma and synovial fluid concentrations of calcitonin gene-related peptide II (CGRP II), substance P and vasoactive intestinal peptide (VIP) were measured in patients with osteoarthritis, gout and rheumatoid arthritis. Significantly higher levels of CGRP II and substance P-like immunoreactivity levels in synovial fluid were found in gout as well as CGRP II, substance P and VIP-like immunoreactivities in rheumatoid arthritis when compared to those in osteoarthritis. Plasma CGRP II, substance P and VIP-like immunoreactivity levels showed no significant differences among patients in the three different groups of arthritis. Our results suggest that these neuropeptides released from peripheral nerve endings into the synovial cavity probably play a pathogenic role in human joint inflammation.


Tradeoffs between air pollution mitigation and meteorological response in India.
To curb the staggering health burden attributed to air pollution, the sustainable solution for India would be to reduce emissions in future. Here we project ambient fine particulate matter (PM2.5) exposure in India for the year 2030 under two contrasting air pollution emission pathways for two different climate scenarios based on Representative Concentration Pathways (RCP4.5 and RCP8.5). All-India average PM2.5 is expected to increase from 41.4 ± 26.5 μg m-3 in 2010 to 61.1 ± 40.8 and 58.2 ± 37.5 μg m-3 in 2030 under RCP8.5 and RCP4.5 scenarios, respectively if India follows the current legislation (baseline) emission pathway. In contrast, ambient PM2.5 in 2030 would be 40.2 ± 27.5 (for RCP8.5) and 39.2 ± 25.4 (for RCP4.5) μg m-3 following the short-lived climate pollutant (SLCP) mitigation emission pathway. We find that the lower PM2.5 in the mitigation pathway (34.2% and 32.6%, respectively for RCP8.5 and RCP4.5 relative to the baseline emission pathway) would come at a cost of 0.3-0.5 °C additional warming due to the direct impact of aerosols. The premature mortality burden attributable to ambient PM2.5 exposure is expected to rise from 2010 to 2030, but 381,790 (5-95% confidence interval, CI 275,620-514,600) deaths can be averted following the mitigation emission pathway relative to the baseline emission pathway. Therefore, we conclude that given the expected large health benefit, the mitigation emission pathway is a reasonable tradeoff for India despite the meteorological response. However, India needs to act more aggressively as the World Health Organization (WHO) annual air quality guideline (10 µg m-3) would remain far off.


Purification and characterization of thymidylate synthetase from rat regenerating liver.
Thymidylate synthetase (EC 18.104.22.168) from rat regenerating liver has been purified over 5000-fold to apparent homogeneity by a procedure involving two affinity methods. Molecular weight of the native enzyme was found to be about 68,000, as determined by gel filtration. Electrophoresis in polyacrylamide gels containing sodium dodecyl sulfate yielded a single band of molecular weight of 35,000, suggesting that thymidylate synthetase is a dimer of very similar or identical subunits. The Michaelis constants for deoxyuridylate (dUMP) and (+/-)L-5,10-methylenetetrahydrofolate are 6.8 microM and 65 microM, respectively. Reaction kinetics and product inhibition studies reveal the enzymatic mechanism to be ordered sequential. 5-Fluoro-dUMP, halogenated analog of the nucleotide substrate is a competitive inhibitor of the enzyme, with an apparent Ki value of 5 nM. Amethopterin, analog of the cofactor is also a competitive inhibitor with an apparent Ki value of 23 microM.


Circadian regulation of genetic and hormonal risk factors of cardiovascular disease in women.
Cardiovascular disease is the leading cause of morbidity and mortality worldwide. However, sex differences can impact differently the etiology and outcome of cardiovascular disease when comparing men and women. Women have unique genetic and hormonal risk factors that can be associated with the development of cardiovascular diseases. Furthermore, certain phenotypes of cardiovascular diseases are more prevalent to women. Molecular clocks control circadian rhythms of different physiological systems in our body, including the cardiovascular system. Increased evidence in recent years points to a link between cardiovascular disease and regulation by circadian rhythms. However, the difference between circadian regulation of cardiovascular disease in women and men is poorly understood. In this review, we highlight the recent advances in circadian-regulated cardiovascular diseases with a specific focus on the pathogenesis of heart disease in women. Understanding circadian-regulated pathways and sex-specific differences between men and women may contribute to better diagnosis and development of sex-targeted interventions to better treat cardiovascular diseases.


Glucan enhances survival in an intraabdominal infection model.
The immunomodulator glucan exists in two forms, particulate (glucan-P) and soluble (glucan-F). Both preparations of glucan, either alone or in combination with antibiotic therapy, were evaluated for their ability to augment survival in rats following cecal ligation and puncture (CL/P). Adult male rats were infused once daily for 5 consecutive days with either glucan-P (10 mg/kg), glucan-F (10 mg/kg), or 5% (w/v) dextrose in water. Three days later all rats underwent CL/P. Postoperatively, the rats received (a) no therapy, (b) saline (1 ml subcutaneously every 12 hr) or (c) ampicillin (33 mg/kg subcutaneously every 12 hr) for 7 days. Without any associated pre-or postoperative treatment, CL/P was associated with an 85% 7-day mortality. Neither glucan preparation alone significantly altered this mortality. Administering ampicillin postoperatively decreased the mortality to 53% (P less than 0.001 vs untreated controls). When postoperative ampicillin therapy was combined with preoperative glucan treatment, the mortality was reduced even further (26% for glucan-P, 21% for glucan-F; P less than 0.02 vs ampicillin-treated controls). We conclude from these results that (i) neither glucan preparation alone effectively enhances survival following CL/P when using the doses and administration schedule employed herein, (ii) both glucan-P and glucan-F do act synergistically with antibiotics to enhance survival in this rat model of polymicrobial sepsis, and (iii) in this particular model, nontoxic glucan-F is as efficacious as glucan-P.


Aryl-functionalised α,α'-Trehalose 6,6'-Glycolipid Induces Mincle-independent Pyroptotic Cell Death.
α,α'-Trehalose 6,6'-glycolipids have long been known for their immunostimulatory properties. The adjuvanticity of α,α'-trehalose 6,6'-glycolipids is mediated by signalling through the macrophage inducible C-type lectin (Mincle) and the induction of an inflammatory response. Herein, we present an aryl-functionalised trehalose glycolipid, AF-2, that leads to the release of cytokines and chemokines, including IL-6, MIP-2 and TNF-α, in a Mincle-dependent manner. Furthermore, plate-coated AF-2 also leads to the Mincle-independent production of IL-1β, which is unprecedented for this class of glycolipid. Upon investigation into the mode of action of plate-coated AF-2, it was observed that the treatment of WT and Mincle-/- bone marrow derived macrophages (BMDM), murine RAW264.7 cells, and human monocytes with AF-2 led to lytic cell death, as evidenced using Sytox Green and lactate dehydrogenase assays, and confocal and scanning electron microscopy. The requirement for functional Gasdermin D and Caspase-1 for IL-1β production and cell death by AF-2 confirmed pyroptosis as the mode of action of AF-2. The inhibition of NLRP3 and K+ efflux reduced AF-2 mediated IL-1β production and cell death, and allowed us to conclude that AF-2 leads to Capase-1 dependent NLRP3 inflammasome-mediated cell death. The unique mode of action of plate-coated AF-2 was surprising and highlights how the physical presentation of Mincle ligands can lead to dramatically different immunological outcomes.


Serial electroconvulsive Seizure alters dendritic complexity and promotes cellular proliferation in the mouse dentate gyrus; a role for Egr3.
Despite being one of the safest, most effective treatments for severe mood disorders, the therapeutic mechanisms of electroconvulsive therapy remain unknown. Electroconvulsive seizure (ECS) induces rapid, high-level expression of immediate early genes (IEGs) and brain-derived neurotrophic factor (BDNF), in addition to stimulation of neurogenesis and dendritic remodeling of dentate gyrus (DG) neurons. We have previously shown that this upregulation of BDNF fails to occur in the hippocampus of mice lacking the IEG Egr3. Since BDNF influences neurogenesis and dendritic remodeling, we hypothesized that Egr3-/- mice will exhibit deficits in neurogenesis and dendritic remodeling in response to ECS. To test this hypothesis, we examined dendritic remodeling and cellular proliferation in the DG of Egr3-/- and wild-type mice following repeated ECS. Mice received 10 daily ECSs. Dendritic morphology was examined in Golgi-Cox-stained tissue and cellular proliferation was analyzed through bromodeoxyuridine (BrdU) immunohistochemistry and confocal imaging. Serial ECS in mice results in dendritic remodeling, increased spine density, and cellular proliferation in the DG. Loss of Egr3 alters the dendritic remodeling induced by serial ECS but does not change the number of dendritic spines or cellular proliferation consequences of ECS. Egr3 influences the dendritic remodeling induced by ECS but is not required for ECS-induced proliferation of hippocampal DG cells.


The effect of complexity on the discrimination of oriented bars by the honeybee (Apis mellifera).
Visual discrimination of black bars by honeybees was studied in a Y-choice apparatus with fixed vertical patterns at constant range. The problem is to discover how bees remember different degrees of complexity of the orientation cue. Previous conclusions with parallel gratings and single bars disagree. With broad bars versus orthogonal bars, the bees learn the orientation cue if the bars are centred at the same place, but they learn the position cue in the vertical direction when the bars are at different places on the two targets. With several bars on each target, the bees learn their orientation and positions. As fixed patterns increase in complexity, the bees follow a simple rule, to look only at the range of places where the cues were displayed. The frame of reference is disrupted when a black spot is added to the training pattern. There is abundant evidence that the bees do not re-assemble the pattern or learn shapes. The filters that detect the position and orientation cues are coarsely tuned, so that they respond in a graded way, but the memory of the range of directions of the cue, as seen from the point of choice, is more exact.


A role of p38 mitogen-activated protein kinase in adenosine A₁ receptor-mediated synaptic depotentiation in area CA1 of the rat hippocampus.
Although long-term potentiation (LTP) of synaptic strength is very persistent, current studies have provided evidence that various manipulations or pharmacological treatment when applied shortly after LTP induction can reverse it. This kind of reversal of synaptic strength is termed as depotentiation and may have a function to increase the flexibility and storage capacity of neuronal networks. Our previous studies have demonstrated that an increase in extracellular levels of adenosine and subsequent activation of adenosine A₁ receptors are important for the induction of depotentiation; however, the signaling downstream of adenosine A₁ receptors to mediate depotentiation induction remains elusive. We confirm that depotentiation induced by low-frequency stimulation (LFS) (2 Hz, 10 min, 1200 pulses) was dependent on adenosine A₁ receptor activation, because it was mimicked by bath-applied adenosine A₁ receptor agonist N⁶-cyclopentyladenosine (CPA) and was inhibited by the selective adenosine A₁ receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX). Pretreatment of the hippocampal slices with the selective p38 mitogen-activated protein kinase (MAPK) inhibitors, 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl]-5-(4-pyrudyl)-1H-imidazole (SB203580) or trans-1-(4-hydroxycyclohexyl)-4-(fluorophenyl)-5-(2-methoxypyrimidin-4-yl)imidazole (SB239063), prevented the induction of depotentiation by LFS and CPA. In agreement with electrophysiological observation, both LFS- and CPA-induced depotentiation are associated with an increase in p38 MAPK activation, which are blocked by DPCPX or SB203580 application. These results suggest that activation of adenosine A₁ receptor and in turn triggering p38 MAPK signaling may contribute to the LFS-induced depotentiation at hippocampal CA1 synapses.


A rickettsia infection from Saudi Arabia.
We here present a young patient who developed fever, headache, maculopapular skin rash and an eschar 3 days following a tick bite. Serology for Rickettsia was consistent with acute spotted fever group (SFG) infection. He had a dramatic response to doxycycline. Vectors for rickettsia transmission are shown in a number of studies from Saudi Arabia while human serological studies from Oman revealed that these infections are common among the rural population. Surprisingly, despite the existence of potential vectors, this disease is rarely reported from the Arab Gulf countries. To the best of our knowledge this is the first case report of SFG acquired within Saudi Arabia. SFG should be considered in the differential diagnosis of febrile illness and exposure history.


Imaging in Pediatric Obstructive Jaundice.
Cholestatic jaundice characterized by elevated conjugated bilirubin can be due to multitude of factors in neonates and childhood. Extrahepatic biliary atresia (EHBA), choledochal cyst, neonatal hepatitis, cytomegalovirus (CMV), and biliary plug are some of the common causes in neonate and early infancy. Causes in late infancy and childhood comprises viral hepatitis, choledochal cyst, cholelithiasis, worm infestation, and biliary compression secondary to extrinsic causes (node, collection, tumor). Some serious disorders like biliary atresia must be considered with the emphasis on early diagnosis of treatable causes. In the modern era, with multiple diagnostic modalities available including high-resolution ultrasonography, magnetic resonance imaging (MRI), CT scan, and nuclear imaging [hepatobiliary iminodiacetic acid (HIDA) scan], rapid diagnosis can be made in many surgically treatable cases. The authors will discuss the imaging modality available with advantages, disadvantages, and common indications of each modality, and overview of obstructive jaundice discussing the wide spectrum of causes in neonates and late childhood. Combining available knowledge with careful and meticulous search can help narrow down the diagnosis and initiate prompt treatment.


Suppression of hepatocyte proliferation by hepatocyte nuclear factor 4α in adult mice.
Hepatocyte nuclear factor 4α (HNF4α) regulates genes involved in lipid and bile acid synthesis, gluconeogenesis, amino acid metabolism, and blood coagulation. In addition to its metabolic role, HNF4α is critical for hepatocyte differentiation, and loss of HNF4α is associated with hepatocellular carcinoma. The hepatocyte-specific Hnf4a knock-out mouse develops severe hepatomegaly and steatosis resulting in premature death, thereby limiting studies of the role of this transcription factor in the adult animal. In addition, gene compensation may complicate analysis of the phenotype of these mice. To overcome these issues, an acute Hnf4a knock-out mouse model was generated through use of the tamoxifen-inducible ErT2cre coupled to the serum albumin gene promoter. Microarray expression analysis revealed up-regulation of genes associated with proliferation and cell cycle control only in the acute liver-specific Hnf4α-null mouse. BrdU and ki67 staining confirmed extensive hepatocyte proliferation in this model. Proliferation was associated with induction of the hepatomitogen Bmp7 as well as reduced basal apoptotic activity. The p53/p63 apoptosis effector gene Perp was further identified as a direct HNF4α target gene. These data suggest that HNF4α maintains hepatocyte differentiation in the adult healthy liver, and its loss may directly contribute to hepatocellular carcinoma development, thus indicating this factor as a possible liver tumor suppressor gene.


Processivity of the single-headed kinesin KIF1A through biased binding to tubulin.
Conventional isoforms of the motor protein kinesin behave functionally not as 'single molecules' but as 'two molecules' paired. This dimeric structure poses a barrier to solving its mechanism. To overcome this problem, we used an unconventional kinesin KIF1A (refs 5, 6) as a model molecule. KIF1A moves processively as an independent monomer, and can also work synergistically as a functional dimer. Here we show, by measuring its movement with an optical trapping system, that a single ATP hydrolysis triggers a single stepping movement of a single KIF1A monomer. The step size is distributed stochastically around multiples of 8 nm with a gaussian-like envelope and a standard deviation of 15 nm. On average, the step is directional to the microtubule's plus-end against a load force of up to 0.15 pN. As the source for this directional movement, we show that KIF1A moves to the microtubule's plus-end by approximately 3 nm on average on binding to the microtubule, presumably by preferential binding to tubulin on the plus-end side. We propose a simple physical formulation to explain the movement of KIF1A.


Differences in Quality of Life in Children Across the Spectrum of Congenital Heart Disease.
To create complexity groups based upon a patient's cardiac medical history and to test for group differences in health-related quality of life (HRQOL). Patients 8-18 years with congenital heart disease (CHD) and parent-proxies from the Pediatric Cardiac Quality of Life Inventory (PCQLI) Testing Study were included. Outcome variables included PCQLI Total, Disease Impact, and Psychosocial Impact scores. Using a patient's medical history (cardiac, neurologic, psychological, and cognitive diagnosis), latent class analysis (LCA) was used to create CHD complexity groups. Covariates included demographics and burden of illness (number of: school weeks missed, physician visits in the past year, and daily medications). Generalized estimation equations tested for differences in burden of illness and patient and parent-proxy PCQLI scores. Using 1482 CHD patients (60% male; 84% white; age 12.3 ± 3.0 years), latent class analysis (LCA) estimates showed 4 distinct CHD complexity groups (Mild, Moderate 1, Moderate 2, and Severe). Increasing CHD complexity was associated with increased risk of learning disorders, seizures, mental health problems, and history of stroke. Greater CHD complexity was associated with greater burden of illness (P < .01) and lower patient- and parent-reported PCQLI scores (P < .001). LCA identified 4 congenital heart disease (CHD) complexity groupings. Increasing CHD complexity was associated with higher burden of illness and worse patient- and parent-reported HRQOL.


Comment on "Food wanting is mediated by transient activation of dopaminergic signaling in the honey bee brain".
Huang et al. (1) make an exciting claim about a human-like dopamine-regulated neuromodulatory mechanism underlying food-seeking behavior in honey bees. Their claim is based on experiments designed to measure brain biogenic amine levels and manipulate receptor activity. We have concerns that need to be addressed before broad acceptance of their results and the interpretation provided.


Selection of a chemically defined medium for culturing adult newt forelimb regenerates.
Three commercially available tissue-culture media were evaluated for their ability to support continued growth and differentiation of 14-day regenerates of adult newt forelimbs. Serums, embryo extracts, egg ultrafiltrates, and antimicrobial agents were avoided in this analysis. The hormones insulin and L-thyroxine were added to these chemically defined media to enhance continued cellular metabolism and growth. The optimum conditions appeared to be cultivated at 25 degrees C (pH 7.2 to 7.4) in media osmotically adjusted to conditions approximating amphibian blood values (i.e. 225 m0SM for 199, 244 m0SM for CMRL-1066, and 262 m0SM FOR L-15).


Weekly cisplatin and radiotherapy for low risk, locoregionally advanced human papillomavirus-positive oropharyngeal squamous cell carcinoma.
There is interest in different treatment strategies, including deintensification in good prognosis human papillomavirus-positive (HPV(+)) oropharyngeal squamous cell carcinoma (SCC). We reviewed our experience with weekly cisplatin in low-risk, locoregionally advanced HPV(+) oropharyngeal SCC since late 2009. Data from patients with low-risk HPV(+) oropharyngeal SCC treated with weekly cisplatin (40 mg/m(2) ) and 70 Gy radiotherapy were collected. Low risk was defined as stage III to IV oropharyngeal SCC excluding T1-2N1, T4 or N3 disease, or N2b to N2c disease in patients with >10 pack-year smoking history. Of 31 patients, the median age was 56 years (range, 41-69 years). All patients completed 70 Gy radiotherapy within 51 days and 84% completed at least 5 cycles of cisplatin. Grade 3 mucositis occurred in 22 patients (71%) and grade 3 febrile neutropenia in 6 patients (19%). No patients required enteral feeding at 12 months. The median follow-up was 30 months (range, 21-57 months) with no recurrences or deaths. Concurrent weekly cisplatin is relatively well-tolerated and associated with excellent disease control in low-risk, locoregionally advanced HPV(+) oropharyngeal SCC. © 2015 Wiley Periodicals, Inc. Head Neck 38: E1117-E1121, 2016.


Absorption, distribution, metabolism and excretion of 14C-vatiquinone in rats, dogs, and human subjects.
Vatiquinone is a potent inhibitor of 15-lipoxygenase and is in clinical development for the treatment of mitochondrial diseases and other disorders characterised by high levels of oxidative stress and dysregulation of energy metabolism.In rats, 14C-vatiquinone-derived radioactivity was quickly and widely distributed throughout the body and cleared from most tissues by 24 h post-dose following a single oral dose of 14C-vatiquinone.Following oral administration, 94% of dose was recovered within seven days in rats, approximately 61% of dose was recovered within seven days in dogs and approximately 93% of dose was recovered within nine days in human subjects (IND 119220). Faecal excretion was the major route (>56% dose) in all species; urinary excretion was minimal in rats and dogs (<3% dose) but was higher in humans (∼ 22% dose).Following oral administration, vatiquinone was the dominant circulating component in rats and dogs but was minor in human subjects. There were no plasma metabolites that were more than 10% of total drug related exposures in all species.Following oral administration, vatiquinone was not detectable in urine but was the most prominent component in faeces in rats, dogs, and humans.


A Parent's Journey: Incorporating Principles of Palliative Care into Practice for Children with Chronic Neurologic Diseases.
Rather than in conflict or in competition with the curative model of care, pediatric palliative care is a complementary and transdisciplinary approach used to optimize medical care for children with complex medical conditions. It provides care to the whole child, including physical, mental, and spiritual dimensions, in addition to support for the family. Through the voice of a parent, the following case-based discussion demonstrates how the fundamentals of palliative care medicine, when instituted early in the course of disease, can assist parents and families with shared medical decision making, ultimately improving the quality of life for children with life-limiting illnesses. Pediatric neurologists, as subspecialists who provide medical care for children with chronic and complex conditions, should consider invoking the principles of palliative care early in the course of a disease process, either through applying general facets or, if available, through consultation with a specialty palliative care service.


Acute Unloading Effects of Sildenafil Enhance Right Ventricular-Pulmonary Artery Coupling in Heart Failure.
Phosphodiesterase-5A inhibitors (PDE5i) are sometimes used in patients with advanced heart failure with reduced ejection fraction before heart transplant or left ventricular assist device implantation to decrease right ventricular (RV) afterload and mitigate the risk of right heart failure. Conflicting evidence exists regarding the impact of these drugs on RV contractility. The aim of this study was to explore the acute effects of PDE5i on ventricular-vascular coupling and load-independent RV contractility. Twenty-two patients underwent right heart catheterization and gated equilibrium blood pool single photon emission computed tomography, before and after 20 mg intravenous sildenafil. Single photon emission computed tomography and right heart catheterization-derived data were used to calculate RV loading and contractility. PDE5i induced a decrease in the right atrial pressure (-43%), pulmonary artery (PA) mean pressure (-26%), and PA wedge pressure (PAWP; -23%), with favorable reductions in pulmonary vascular resistance (-41%) and PA elastance (-40%), and increased cardiac output (+13%) (all P < 0.01). The RV ejection fraction increased with sildenafil (+20%), with no change of RV contractility (P = 0.74), indicating that the improvement in the RV ejection fraction was related to enhanced RV-PA coupling (r = 0.59, P = 0.004) by a decrease in the ventricular load. RV diastolic compliance increased with sildenafil. The decrease in the PAWP correlated with RV end-diastolic volume decrease; no relationship was observed with the change in LV transmural pressure, suggesting decreased pericardial constraint. Acute PDE5i administration has profound RV afterload-reducing effects, improves the RVEF, decreases RV volumes, and decreases the PAWP, predominantly through relief of pericardial constraint, without effects on RV chamber contractility. These findings support further study of PDE5i in protection of RV function in advanced heart failure with reduced ejection fraction who are at risk of RV failure.


The appearance of long-latency responses to a conditioned signal in the cortex is explained by strengthening of collateral connections between pyramidal neurons.
Experimental analysis and computer simulation of the neurophysiological processes underlying the "stable and local electrophysiological expression of conditioned reflexes" in the cerebral cortex, a phenomenon discovered in Asratyan's laboratory in the 1960s, showed that the long-latency components of cortical evoked potentials to a conditioned signal correspond to the late phases of the responses of motor cortex neurons, which are analogous to and probably generated by the same mechanism as long-latency epileptiform reactions of neurons in the epileptogenic cortex. Late long-latency components are generated via activation of NMDA receptors in the collateral connections between pyramidal neurons. The delay in the generation of responses depends on the initial activation of GABA(A) receptors and the slow kinetics of the current through NMDA channels. The appearance of late components as a result of training is explained by increases in the efficiency of collateral excitatory connections between pyramidal neurons.


High-altitude Illnesses and Air Travel: Pediatric Considerations.
Healthy children may present acute mountain sickness (AMS) within a few hours after arrival at high altitudes. In few cases, serious complications may occur, including high-altitude pulmonary edema and rarely high-altitude cerebral edema. Those with preexisting conditions especially involving hypoxia and pulmonary hypertension shall not risk travelling to high altitudes. Newborn from low altitude mothers may have prolonged time to complete postnatal adaptation. The number of children and adolescents traveling on commercial aircrafts is growing, and this poses a need for their treating physicians to be aware of the potential risks of hypoxia while air traveling.