CD73-generated adenosine facilitates Toxoplasma gondii differentiation to long-lived tissue cysts in the central nervous system.
Toxoplasma gondii is an obligate intracellular protozoan pathogen that traffics to the central nervous system (CNS) following invasion of its host. In the CNS, T. gondii undergoes transformation from a rapidly dividing tachyzoite to a long-lived, slow-dividing bradyzoite contained within cysts. The role of extracellular adenosine in T. gondii pathogenesis has not been previously investigated. T. gondii uses host purines such as adenosine for its energy needs, as it is unable to make its own. Here, we show that CD73(-/-) mice, which lack the ability to generate extracellular adenosine, are protected from T. gondii chronic infection, with significantly fewer cysts and reduced susceptibility to reactivation of infection in the CNS independent of host effector function. Parasite dissemination to the brain was unimpaired in CD73(-/-) hosts, suggesting that the reduced cyst number is due to impaired parasite differentiation in the CNS. Confirming this, T. gondii tachyzoites formed fewer cysts following alkaline pH stress in astrocytes isolated from CD73(-/-) mice compared with wild type, and in fibroblasts treated with a CD73 inhibitor. Cyst formation was rescued in CD73(-/-) astrocytes supplemented with adenosine, but not with adenosine receptor agonist 5'-N-ethylcarboxamidoadenosine. Furthermore, mice lacking adenosine receptors had no defect in cyst formation. Based on these findings, we conclude that CD73 expression promotes Toxoplasma bradyzoite differentiation and cyst formation by a mechanism dependent on the generation of adenosine, but independent of adenosine receptor signaling. Overall, these findings suggest that modulators of extracellular adenosine may be used to develop therapies aimed at defending against human toxoplasmosis.


Effect of added fiber on the glucose and metabolic response to a mixed meal in normal and diabetic subjects.
10 g of guar was added to a test meal given to 11 subjects, six of whom had insulin-dependent diabetes, after an overnight fast. The addition of guar to the meal produced an overall decrease in the blood glucose concentrations after the meal in both normal and diabetic subjects. The insulin secretory response in the normal subjects was less when guar was included in the meal than when it was not. In normal subjects, but not in diabetic subjects, the addition of guar also resulted in a diminished response of lactate, pyruvate, and alanine to the test meal and a blunting of the fall in serum ketones. This is likely related to the fact that the same dose of insulin was given in the diabetic subjects with both test meals. Thus, in the normal circumstances, the diminished glycemic response to the guar-containing meal resulted in diminished insulin secretion, which was not the case with the diabetic subjects.


Chimeric antibodies with specificity for tumor antigens: demonstration of in situ localization to tumors after antibody therapy.
In this study, we compare various methods for the detection of a tumor-associated target antigen and deposition of the bound therapeutic monoclonal antibody in patients enrolled in two separate trials, one involving the administration of two radiolabeled monoclonal antibodies and the other involving an unlabeled antibody. In the first trial, patients with TAG-72 expressing metastatic colon cancer scheduled for surgical intervention received radiolabeled murine and chimeric B72.3 antibody followed by radioimmune imaging and subsequent laparotomy. Normal and tumor tissues obtained at surgery were processed for routine histology, immunohistochemistry, radiometry, and autoradiography. Both anti-TAG-72 antibodies localized to known tumor sites as evidenced by radioimmune imaging. Resected tissue revealed a high tumor-to-normal radiolocalization ratio, and autoradiography demonstrated even deposition of the radiolabeled antibodies throughout the entire tumor deposit with sparing of surrounding normal tissue. In contrast, immunohistochemistry on the same sections revealed comparatively weak antigen expression and patchy antibody localization. In the second trial, patients with GD2 antigen expressing metastatic melanoma received the unlabeled chimeric anti-GD2 antibody C14.18. Immunologic detection of the GD2 antigen and C14.18 deposition was performed on biopsy section as well as on single cell suspension. FACS analysis of the single cell suspension proved more sensitive for the detection of bound antibody than immunohistochemistry, although both methods yielded comparable results for GD2 antigen expression. Our findings demonstrate that the optimal method for the detection of tumor-associated antigen and bound therapeutic antibody can vary depending upon the nature of the antibody (radiolabeled vs. unlabeled and murine vs. chimeric), fixation stability of the target antigen, and the type of pathologic material available for study.


Etiology and antibiotic resistance patterns of community-acquired urinary tract infections in J N M C Hospital Aligarh, India.
Urinary tract infections (UTIs) remain the common infections diagnosed in outpatients as well as hospitalized patients. Current knowledge on antimicrobial susceptibility pattern is essential for appropriate therapy. Extended-Spectrum beta-Lactamase (ESBL) producing bacteria may not be detected by routine disk diffusion susceptibility test, leading to inappropriate use of antibiotics and treatment failure. The aim of this study was to determine the distribution and antibiotic susceptibility patterns of bacterial strains isolated from patients with community acquired urinary tract infections (UTIs) at Aligarh hospital in India as well as identification of ESBL producers in the population of different uropathogens. Urinary isolates from symptomatic UTI cases attending to the JN Medical College and hospital at Aligarh were identified by conventional methods. Antimicrobial susceptibility testing was performed by Kirby Bauer's disc diffusion method. Isolates resistant to third generation cephalosporin were tested for ESBL production by double disk synergy test method. Of the 920 tested sample 100 samples showed growth of pathogens among which the most prevalent were E. coli (61%) followed by Klebsiella spp (22%). The majority (66.66%) of the isolates were from female while the remaining were from male. Among the gram-negative enteric bacilli high prevalence of resistance was observed against ampicillin and co-trimoxazole. Most of the isolates were resistant to 4 or more number of antibiotics. Forty two percent of isolates were detected to produce ESBL among which 34.42 % were E. coli isolates. This study revealed that E. coli was the predominant bacterial pathogen of community acquired UTIs in Aligarh, India. It also demonstrated an increasing resistance to Co-trimoxazole and production of extended spectrum beta-lactamase among UTI pathogens in the community. This study is useful for clinician in order to improve the empiric treatment.


Characterization of a DNA fragment carrying the raw starch-digesting alpha-amylase and salt-dependent alpha-amylase genes from Bacillus circulans F-2.
A 5.4 kb HindIII DNA fragment carrying the gene encoding raw starch-digesting alpha-amylase (RSDA), has been previously cloned from Bacillus circulans F-2 and expressed in Escherichia coli [Kim et al. (1990) Biochim. Biophys. Acta 1048, 2233-2238]. Interestingly, when the cell extract of E. coli harboring a plasmid carrying this fragment was incubated with 1 M NaCl, it exhibited about 10 times higher enzyme activity than when assayed without NaCl. Differential zymograms showed two different amylase activities: one for RSDA and the other for a salt-dependent alpha-amylase (SDA). Even though RSDA activity was detected without NaCl, SDA activity was detected only in high concentrations of NaCl. SDA activity was fully detected at above 1 M NaCl. Results from subcloning of the genes, fractionation analysis of cell extracts, and immunological assays clearly suggested that the two amylases are genetically distinct and that genes for both enzymes are closely linked on the 5.4 kb DNA fragment.


A serious case of anthrax associated with compartment syndrome.
Compartment syndrome linked to skin anthrax is a rare complication that may develop and it should be noted that the disease may progress in spite of medical drug treatment. Our case was a farmer who was exposed after slaughtering a dead animal, a time delay for treatment hided this history and then developed compartment syndrome. In anthrax cases with delayed treatment and aggressive progression, circulation in the extremities should be carefully noted. We believe that the cases with compartment syndrome progressing in spite of medical drug treatment may be assessed for fasciotomy as a treatment approach.


Utility of Percentage Signal Recovery and Baseline Signal in DSC-MRI Optimized for Relative CBV Measurement for Differentiating Glioblastoma, Lymphoma, Metastasis, and Meningioma.
The percentage signal recovery in non-leakage-corrected (no preload, high flip angle, intermediate TE) DSC-MR imaging is known to differ significantly for glioblastoma, metastasis, and primary CNS lymphoma. Because the percentage signal recovery is influenced by preload and pulse sequence parameters, we investigated whether the percentage signal recovery can still differentiate these common contrast-enhancing neoplasms using a DSC-MR imaging protocol designed for relative CBV accuracy (preload, intermediate flip angle, low TE). We retrospectively analyzed DSC-MR imaging of treatment-naïve, pathology-proved glioblastomas (n = 14), primary central nervous system lymphomas (n = 7), metastases (n = 20), and meningiomas (n = 13) using a protocol designed for relative CBV accuracy (a one-quarter-dose preload and single-dose bolus of gadobutrol, TR/TE = 1290/40 ms, flip angle = 60° at 1.5T). Mean percentage signal recovery, relative CBV, and normalized baseline signal intensity were compared within contrast-enhancing lesion volumes. Classification accuracy was determined by receiver operating characteristic analysis. Relative CBV best differentiated meningioma from glioblastoma and from metastasis with areas under the curve of 0.84 and 0.82, respectively. The percentage signal recovery best differentiated primary central nervous system lymphoma from metastasis with an area under the curve of 0.81. Relative CBV and percentage signal recovery were similar in differentiating primary central nervous system lymphoma from glioblastoma and from meningioma. Although neither relative CBV nor percentage signal recovery differentiated glioblastoma from metastasis, mean normalized baseline signal intensity achieved 86% sensitivity and 50% specificity. Similar to results for non-preload-based DSC-MR imaging, percentage signal recovery for one-quarter-dose preload-based, intermediate flip angle DSC-MR imaging differentiates most pair-wise comparisons of glioblastoma, metastasis, primary central nervous system lymphoma, and meningioma, except for glioblastoma versus metastasis. Differences in normalized post-preload baseline signal for glioblastoma and metastasis, reflecting a snapshot of dynamic contrast enhancement, may motivate the use of single-dose multiecho protocols permitting simultaneous quantification of DSC-MR imaging and dynamic contrast-enhanced MR imaging parameters.


The p53Pro72Arg polymorphism is associated with albuminuria among aboriginal Australians.
Albuminuria is a widely recognized marker of renal disease and cardiovascular risk. This is especially true in Aboriginal Australians living in remote communities who suffer high rates of end-stage renal disease and cardiovascular mortality. During a survey of risk factors for renal and cardiovascular disease in one such community, an association between a common polymorphism at codon 72 (Arg/Pro) of the p53 gene and markers of renal disease was sought. A cross-sectional community survey including 217 people was performed. Genotypes of the polymorphism were distributed in Hardy-Weinberg equilibrium, with p53Arg allele frequency of 0.45 (range, 0.41 to 0.50). Overall prevalence of albuminuria was high (31% microalbuminuria; 14% overt albuminuria). Urine albumin/creatinine ratio (ACR) was significantly associated with the number of p53Pro alleles (P = 0.01), and there was an interaction with tobacco smoking (P = 0.04). The p53 genotype was also associated with increasing HbA1c, but the relationship between p53 and ACR was independent of this. This is a previously unreported association. This study does not address the mechanism, but this finding, if confirmed, expands the described effects of p53 in cellular proliferation and apoptosis to include a role in the course of renal and possibly cardiovascular disease in this population.


The Hierarchical Taxonomy of Psychopathology (HiTOP): A Quantitative Nosology Based on Consensus of Evidence.
Traditional diagnostic systems went beyond empirical evidence on the structure of mental health. Consequently, these diagnoses do not depict psychopathology accurately, and their validity in research and utility in clinicalpractice are therefore limited. The Hierarchical Taxonomy of Psychopathology (HiTOP) consortium proposed a model based on structural evidence. It addresses problems of diagnostic heterogeneity, comorbidity, and unreliability. We review the HiTOP model, supporting evidence, and conceptualization of psychopathology in this hierarchical dimensional framework. The system is not yet comprehensive, and we describe the processes for improving and expanding it. We summarize data on the ability of HiTOP to predict and explain etiology (genetic, environmental, and neurobiological), risk factors, outcomes, and treatment response. We describe progress in the development of HiTOP-based measures and in clinical implementation of the system. Finally, we review outstanding challenges and the research agenda. HiTOP is of practical utility already, and its ongoing development will produce a transformative map of psychopathology.


A comparison of procedures and materials for speech reception thresholds.
Ascending and descending procedures were used to obtain speech reception thresholds from 32 normal-hearing subjects whereas digits and spondee words were used as test materials. Comparisons of thresholds were made for the two procedures and materials. Results indicated that slightly lower (less SPL) speech thresholds resulted from the use of a descending procedure with both types of material. In addition, digit material was found to yield generally lower threshold values than spondees. Advantages to the use of the two procedures and materials are discussed.


Update on the pathophysiology of degenerative disc disease and new developments in treatment strategies.
Degenerative disc disease (DDD) continues to be a prevalent condition that afflicts populations on a global scale. The economic impact and decreased quality of life primarily stem from back pain and neurological deficits associated with intervertebral disc degeneration. Although much effort has been invested into understanding the etiology of DDD and its relationship to the onset of back pain, this endeavor is a work in progress. The purpose of this review is to provide focused discussion on several areas in which recent advances have been made. Specifically, we have categorized these advances into early, middle, and late phases of age-related or degenerative changes in the disc and into promising minimally invasive treatments, which aim to restore mechanical and biological functions to the disc.


Impact of lymphatic vessel invasion on survival in curative resected gastric cancer.
Lymphatic vessel invasion (LV) has been regarded as a prognostic factor in some solid tumors. The aim was to clarify the impact of lymphatic vessel invasion on survival in curative resected gastric cancer. In this retrospective study, we reviewed the records of 1,024 patients who underwent curative resection for gastric cancer. Among all of the studied patients, 285 of them (27.8%) had lymphatic vessel invasion. There were significant differences in tumor size, tumor location, depth of invasion, and lymph node metastasis (LN) between the patients with lymphatic vessel invasion and those without. The 5-year survival rates in patients were 80.1%, 59.2%, 40.9%, and 30.5% for LN-LV-, LN-LV+, LN+LV-, and LN+LV+ group, respectively. Multivariate analysis revealed that age, tumor location, the depth of invasion, and lymph node metastasis were independent prognostic factors for curative resected gastric cancer. Lymphatic vessel invasion was not an independent prognostic factor in node-positive gastric cancer; however, it was true in node-negative gastric cancer. Lymphatic vessel invasion is one of the independent prognostic factors for node-negative gastric cancer after curative resection.


The dosage compensation complex shapes the conformation of the X chromosome in Drosophila.
The dosage compensation complex (DCC) in Drosophila globally increases transcription from the X chromosome in males to compensate for its monosomy. We discovered a male-specific conformation of the X chromosome that depends on the associations of high-affinity binding sites (HAS) of the DCC. The core DCC subunits MSL1-MSL2 are responsible for this male-specific organization. Contrary to emerging concepts, we found that neither DCC assembly nor the conformation of the male X chromosome are influenced by nuclear pore components. We propose that nuclear organization of HAS is central to the faithful distribution of the DCC along the X chromosome.


Risk of surgery in patients with stricturing type of Crohn's disease at the initial diagnosis: a single center experience.
It remains uncertain which patients with stricturing-type Crohn's disease (CD) require early small bowel surgery after the initial diagnosis. We aimed to clarify clinical characteristics associated with the intervention in such condition of CD. We retrospectively evaluated the clinical course of 53 patients with CD and small bowel strictures who were initially treated with medications after the initial diagnosis. We investigated possible associations between small bowel surgery and the following: clinical factors and radiologic findings at initial diagnosis and the types of medications administered during follow-up. Twenty-eight patients (53%) required small bowel resection during a median follow-up period of 5.0 years (range, 0.5-14.3 years). The cumulative incidence rates of small bowel surgery at 2, 5, and 10 years were 26.4%, 41.0%, and 63.2%, respectively. Univariate analysis indicated that obstructive symptoms (P=0.036), long-segment stricture (P<0.0001), and prestenotic dilation (P<0.0001) on radiography were associated with small bowel surgery, and immunomodulatory (P=0.037) and biological therapy (P=0.008) were significant factors during follow-up. Multivariate analysis revealed that long-segment stricture (hazard ratio [HR], 4.25; 95% confidence interval [CI], 1.78-10.53; P=0.001) and prestenotic dilation (HR, 3.41; 95% CI, 1.24-9.62; P=0.018) on radiography showed a positive correlation with small bowel surgery, and biological therapy (HR, 0.40; 95% CI, 0.15-0.99; P=0.048) showed a negative correlation. CD patients with long-segment stricture and prestenotic dilation on radiography seem to be at a higher risk of needing small bowel surgery. For such patients, early surgical intervention might be appropriate, even at initial diagnosis.


Affinities of penicillins and cephalosporins for the penicillin-binding proteins of Escherichia coli K-12 and their antibacterial activity.
The affinities of a range of penicillins and cephalosporins for ther penicillin-binding proteins of Escherichia coli K-12 have been studied, and the results were compared with the antibacterial activity of the compounds against E. coli K-12 and an isogenic permeability mutant. Different penicillins and cephalosporins exhibited different affinities for the "essential" penicillin-binding proteins of E. coli K-12, in a manner which directly correlated with their observed effects upon bacterial morphology. Furthermore, the affinities of the compounds for their "primary" lethal penicillin-binding protein targets showed close agreement with their antibacterial activities against the permeability mutant.


Usnic acid, a lichen secondary metabolite inhibits Group A Streptococcus biofilms.
Group A Streptococci (GAS) are involved in a number of life threatening diseases and biofilm formation by these pathogens are considered as an important virulence determinant as it mediates antibiotic resistance among them. In the present study, we have explored the ability of (+)-usnic acid, a lichen secondary metabolite, as an antibiofilm agent against four serotypes of Streptococcus pyogenes causing pharyngitis. Usnic acid inhibited the biofilms of M serotypes M56, st38, M89 efficiently and the biofilm of M74 to a lesser extent. Confocal imaging of the treated samples showed that usnic acid reduced the biomass of the biofilms when compared to that of the control. Fourier Transfer Infrared (FT-IR) spectroscopy indicated that usnic acid reduced the cellular components (proteins and fatty acids) of the biofilms. Interestingly, the FT-IR spectrum further revealed that usnic acid probably acted upon the fatty acids of the biofilms as evident from the disappearance of a peak at 2,455-2,100 cm(-1) when compared to the control only in serotypes M56, st38 and M89 but not in M74. The present study shows, for the first time, that usnic acid can act as an effective antibiofilm agent against GAS.


Effect of eye testing order on automated perimetry results using the Swedish Interactive Threshold Algorithm standard 24-2.
To evaluate whether the order of eye testing affects the mean deviation (MD) or the test reliability of visual field testing using the Swedish Interactive Threshold Algorithm (SITA) standard 24-2. Consecutive patients with manifest or suspect glaucoma with 2 prior sets of SITA standard 24-2 test results performed on the right eye first were enrolled. A subsequent test was performed on the left eye first. For each eye, the MD and the test reliability indexes (> or = 20%) were compared among the 3 successive examinations. Forty-seven patients (29 women and 18 men; mean +/- SD age, 70.6 +/- 11.9 years) were enrolled. The MD +/- SD was -5.83 +/- 5.43 dB OD and -5.46 +/- 4.86 dB OS. There was no statistically significant difference in the MD or the test reliability among the 3 test results for either eye. Fixation loss was responsible for the unreliable fields in almost all cases. Among this cohort of patients experienced with automated perimetry in a glaucoma subspecialty practice, changing the order of eye testing using the SITA standard 24-2 did not have a significant effect on the MD or the test reliability. Intereye fatigue may not be clinically significant with this algorithm. Fixation loss remains a problem with the use of this algorithm.


Personal breastfeeding behavior of physician mothers is associated with their clinical breastfeeding advocacy.
Despite excellent breastfeeding initiation rates, physician mothers as a group are at risk of premature breastfeeding cessation. The main obstacles and reasons for breastfeeding cessation among physician mothers are work-related. We conducted this study to further explore physician mothers' personal infant feeding decisions and behavior as well as their clinical breastfeeding advocacy. We interviewed 80 physician mothers, mainly affiliated with the University of Florida College of Medicine (Gainesville, FL), using a questionnaire. Descriptive statistics were calculated with SPSS software version 16 (SPSS, Chicago, IL). The 80 mothers had a total of 152 children and were able to successfully initiate breastfeeding for 97% of the infants. Although maternal goal for duration of breastfeeding had been 12 months or more for 57% of the infants, only 34% of the children were actually still breastfeeding at 12 months. In 43% of cases, physician mothers stated that breastfeeding cessation was due to demands of work. Furthermore, physician mothers who reported actively promoting breastfeeding among their female patients and housestaff had significantly longer personal breastfeeding duration compared with physician mothers who denied actively promoting breastfeeding. Our findings not only emphasize the discrepancy between physician mothers' breastfeeding duration goal and their actual breastfeeding duration, but also highlight the association between their personal breastfeeding success and their own active breastfeeding advocacy. Whether this association is causal cannot be determined by the current study and can be examined further by prospective studies. Our results support developing and implementing workplace strategies and programs to promote breastfeeding duration among physician mothers returning to work.


Establishment of mands following tact training as a function of reinforcer strength.
We examined some conditions under which a response acquired as a tact might facilitate the establishment of a mand. We taught 3 participants with developmental disabilities to tact the items ranked highest and lowest in a preference assessment and subsequently tested to see if the responses occurred as mands. All participants manded for the highly preferred item but rarely manded for the nonpreferred item. These results indicate that, although tact and mand functions are different, conditions can be created to facilitate transfer from the former to the latter. Implications for communication training are discussed.


Parallel RNA extraction using magnetic beads and a droplet array.
Nucleic acid extraction is a necessary step for most genomic/transcriptomic analyses, but it often requires complicated mechanisms to be integrated into a lab-on-a-chip device. Here, we present a simple, effective configuration for rapidly obtaining purified RNA from low concentration cell medium. This Total RNA Extraction Droplet Array (TREDA) utilizes an array of surface-adhering droplets to facilitate the transportation of magnetic purification beads seamlessly through individual buffer solutions without solid structures. The fabrication of TREDA chips is rapid and does not require a microfabrication facility or expertise. The process takes less than 5 minutes. When purifying mRNA from bulk marine diatom samples, its repeatability and extraction efficiency are comparable to conventional tube-based operations. We demonstrate that TREDA can extract the total mRNA of about 10 marine diatom cells, indicating that the sensitivity of TREDA approaches single-digit cell numbers.


Outcome of limb reconstruction system in open tibial diaphyseal fractures.
Management of open tibial diaphyseal fractures with bone loss is a matter of debate. The treatment options range from external fixators, nailing, ring fixators or grafting with or without plastic reconstruction. All the procedures have their own set of complications, like acute docking problems, shortening, difficulty in soft tissue management, chronic infection, increased morbidity, multiple surgeries, longer hospital stay, mal union, nonunion and higher patient dissatisfaction. We evaluated the outcome of the limb reconstruction system (LRS) in the treatment of open fractures of tibial diaphysis with bone loss as a definative mode of treatment to achieve union, as well as limb lengthening, simultaneously. Thirty open fractures of tibial diaphysis with bone loss of at least 4 cm or more with a mean age 32.5 years were treated by using the LRS after debridement. Distraction osteogenesis at rate of 1 mm/day was done away from the fracture site to maintain the limb length. On the approximation of fracture ends, the dynamized LRS was left for further 15-20 weeks and patient was mobilized with weight bearing to achieve union. Functional assessment was done by Association for the Study and Application of the Methods of Illizarov (ASAMI) criteria. Mean followup period was 15 months. The mean bone loss was 5.5 cm (range 4-9 cm). The mean duration of bone transport was 13 weeks (range 8-30 weeks) with a mean time for LRS in place was 44 weeks (range 24-51 weeks). The mean implant index was 56.4 days/cm. Mean union time was 52 weeks (range 31-60 weeks) with mean union index of 74.5 days/cm. Bony results as per the ASAMI scoring were excellent in 76% (19/25), good in 12% (3/25) and fair in 4% (1/25) with union in all except 2 patients, which showed poor results (8%) with only 2 patients having leg length discrepancy more than 2.5 cm. Functional results were excellent in 84% (21/25), good in 8% (2/25), fair in 8% (2/25). Pin tract infection was seen in 5 cases, out of which 4 being superficial, which healed to dressings and antibiotics. One patient had a deep infection which required frame removal. Limb reconstruction system proved to be an effective modality of treatment in cases of open fractures of the tibia with bone loss as definite modality of treatment for damage control as well as for achieving union and lengthening, simultaneously, with the advantage of early union with attainment of limb length, simple surgical technique, minimal invasive, high patient compliance, easy wound management, lesser hospitalization and the lower rate of complications like infection, deformity or shortening.


Direct evidence that a class II molecule and a simple globular protein generate multiple determinants.
We have examined the individual contributions of the I-A kappa alpha chain, the I-A kappa beta chain, and the foreign antigen hen egg-white lysozyme (HEL) in the formation of the determinant being recognized by the T cell receptor. As functional probes we have used (a) a panel of 10 HEL-specific T cell hybridomas, (b) a panel of antigen-presenting cells (APC) possessing mutations in either the I-A kappa alpha or I-A kappa beta chains, and (c) proteolytic fragment of HEL and related synthetic peptides. The ability of the I-A kappa beta and I-A kappa alpha mutant cell lines to present antigen to the 10 T cell hybridomas divided these T cells into six distinct groups. These HEL-specific T cells therefore appear to recognize several distinct domains on the I-A kappa molecule. The 10 T cell hybrids were then shown to recognize at least three distinct determinants on the HEL molecule, with 8 of the 10 hybrids recognizing one of two major determinants HEL(46-61) or HEL(34-45). Combining the response patterns to the panel of I-A kappa mutant APC lines with the antigen specificity revealed that the 10 T cell hybrids recognized at least eight unique determinants formed by the I-A kappa alpha chains, I-A kappa beta chains, and HEL peptides. This analysis provides direct evidence that a large number of different determinants or T cell receptor ligands can be generated from a single Ia molecule and a simple globular protein.


Long QT syndrome: beyond the causal mutation.
Congenital long QT syndrome (LQTS) is caused by single autosomal-dominant mutations in a gene encoding for a cardiac ion channel or an accessory ion channel subunit. These single mutations can cause life-threatening arrhythmias and sudden death in heterozygous mutation carriers. This recognition has been the basis for world-wide staggering numbers of subjects and families counselled for LQTS and treated based on finding (putative) disease-causing mutations. However, prophylactic treatment of patients is greatly hampered by the growing awareness that simple carriership of a mutation often fails to predict clinical outcome: many carriers never develop clinically relevant disease while others are severely affected at a young age. It is still largely elusive what determines this large variability in disease severity, where even within one pedigree, an identical mutation can cause life-threatening arrhythmias in some carriers while in other carriers no disease becomes clinically manifested. This suggests that additional factors modify the clinical manifestations of a particular disease-causing mutation. In this article, potential demographic, environmental and genetic factors are reviewed, which, in conjunction with a mutation, may modify the phenotype in LQTS, and thereby determine, at least partially, the large variability in disease severity.


Identification of potential antigenic peptides of Brucella through proteome and peptidome.
Brucellosis, caused by Brucella spp., is a major zoonotic public health threat. Although several Brucella vaccines have been demonstrated for use in animals, Brucella spp. can cause human infection and to date, there are no human-use vaccines licensed by any agency. Recently, methods in vaccine informatics have made major breakthroughs in peptide-based epitopes, opening up a new avenue of vaccine development. The purpose of this article was to identify potential antigenic peptides in Brucella by proteome and peptidome analyses. Mouse infection models were first established by injection with Brucella and spleen protein profiles were then analysed. Subsequently, the major histocompatibility complex class I or II (major histocompatibility complex [MHC]-I/II)-binding peptides in blood samples were collected by immunoprecipitation and peptides derived from Brucella proteins were identified through liquid chromatography-mass spectrometry (LC-MS/MS). These peptides were then evaluated in a variety of ways, such as in terms of conservation in Brucella and synchronicity in predicted peptides (similarity and coverage), which allowed us to more effectively measure their antigenic potential. The expression of the inflammatory cytokines IL1B and IFN-γ was significantly altered in the spleen of infected mice and some Brucella proteins, such as Muri, AcpP and GroES, were also detected. Meanwhile, in blood, 35 peptides were identified and most showed high conservation, highlighting their potential as antigen epitopes for vaccine development. In particular, we identified four proteins containing both MHC-I- and MHC-II-binding peptides including AtpA, AtpD, DnaK and BAbS19_II02030. They were also compared with the predicted peptides to estimate their reliability. The peptides we screened could bind to MHC molecules. After being stimulated with antigen T epitopes, Memory T cells can stimulate T cell activation and promote immune responses. Our results indicated that the peptides we identified may be good candidate targets for the design of subunit vaccines and these results pave the way for the study of safer vaccines against Brucella.


Formation and movement of myosin-containing structures in living fibroblasts.
Gizzard myosin, fluorescently labeled with tetramethylrhodamine iodoacetamide, was microinjected into living 3T3 fibroblasts to label myosin-containing structures. The fluorophore was located predominantly on the heavy chain near the COOH terminus of the S1 head and on the 17-kD light chain. After microinjection of a tracer amount into living 3T3 cells, the fluorescent myosin showed a distribution identical to that revealed by immunofluorescence with antimyosin antibodies. Injected myosin became localized in small beads, which were found along large stress fibers, along fine fibers, and in a poorly organized form near the lamellipodia. De novo assembly of beads was observed continuously within or near the lamellipodia, suggesting that myosin molecules may undergo a constant cycling between polymerized and unpolymerized states. The nascent structures then moved away from lamellipodia and became organized into linear arrays. Similar movement was also observed for beads already associated with linear structures, and may represent a continuous flux of myosin structures. The dynamic reorganization of myosin may play an important role in cell movement and polarity.


Temperature measurements in normal and tumor tissue of dogs undergoing whole body hyperthermia.
Temperature was measured in the left ventricle, aorta, liver, brain, lung, bone marrow, kidney, and spontaneous solid tumors in dogs undergoing whole body hyperthermia in a radiant heat device. Rectal temperature was found to be a satisfactory indicator of systemic arterial temperature during plateau temperature conditions but rectal temperature underestimated arterial temperature during heating and overestimated it during cooling. Lung temperature, based on small airway temperature, was the same as rectal temperature during plateau temperature conditions. Liver and brain temperatures were slightly higher (0.1-0.2 degree C) than rectal temperature during the plateau phase. During plateau temperature conditions, kidney temperature measurements were higher than rectal temperature when one site/kidney was measured but were lower than rectal temperature when two sites/kidney were measured suggesting invasive thermometry may have affected measured temperature values. Tibial marrow temperature was greater than rectal temperature during heating but fell below rectal temperature during plateau temperature conditions by as much as 1.3 degree C. Femoral marrow temperature was below rectal during heating but gradually exceeded it during steady state conditions, by 0.1-0.4 degree C. Temperature in solid tumors was variable, sometimes exceeding (0.6 degree C) and sometimes being less (1.8 degree C) than rectal temperature.


Ras-Raf-Arf signaling critically depends on the Dmp1 transcription factor.
Dmp1 prevents tumor formation by activating the Arf-p53 pathway. In cultured primary cells, the Dmp1 promoter was efficiently activated by oncogenic Ha-Ras(V12), but not by overexpressed c-Myc or E2F-1. Dmp1 promoter activation by Ras(V12) depended on Raf-MEK-ERK signaling. Induction of p19(Arf) and p21(Cip1) by oncogenic Raf was compromised in Dmp1-null cells, which were resistant to Raf-mediated premature senescence. A Ras(V12)-responsive element was mapped to the 5' leader sequence of the murine Dmp1 promoter, where endogenous Fos and Jun family proteins bind. Dmp1 promoter activation by Ras(V12) was strikingly impaired in c-Jun as well as in JunB knock-down cells, suggesting the critical role of Jun proteins in the activation of the Dmp1 promoter. A Ras(V12)-responsive element was mapped to the unique Dmp1/Ets site on the Arf promoter, where endogenous Dmp1 proteins bind upon oncogenic Raf activation. Therefore, activation of Arf by Ras/Raf signaling is indirectly mediated by Dmp1, explaining why Dmp1-null primary cells are highly susceptible to Ras-induced transformation. Our data indicate the presence of the novel Jun-Dmp1 pathway that directly links oncogenic Ras-Raf signaling and p19(Arf), independent of the classical cyclin D1/Cdk4-Rb-E2F pathway.


[Role of Slit-Robo signaling in the proliferation of human mucoepidermoid carcinoma Mc3 cells].
To study the role of Slit-Robo signaling in the proliferation of human mucoepidermoid carcinoma Mc3 cells. We measured the expressions of Slit2 and Robo1 proteins in human mucoepidermoid carcinoma Mc3 cells using immunohistochemistry and flow cytometry. To test the role of Slit-Robo signaling in the proliferation of the cells, we treated the cells with the monoclonal antibody R5 against Robo1 receptor extracellular domain and observed the changes in the cell proliferation by cell counting and colonogenic assay; the expression of proliferating cell nuclear antigen (PCNA) in the cells following the treatment was measured with flow cytometry. Treatment of Mc3 cells with the monoclonal antibody R5 caused significantly suppressed cell growth and proliferation and obviously lowered the expression of PCNA. Slit-Robo signaling can suppress the proliferation of Mc3 cells possibly by up-regulating the expression of PCNA.


Cigarette smoking and incident chronic kidney disease: a systematic review.
Several studies have examined the role of cigarette smoking in the development of renal disease in human populations. However, there have been no systematic reviews on the evidence linking smoking with incident renal disease. We performed an evidence-based evaluation of peer-reviewed research published during 1966-2005, from a search of five databases, including Ovid MEDLINE and EMBASE. Of the 28 studies that were reviewed, 11 were excluded from the final analysis due to poor methodological quality (n = 6), no reported risk estimate for the association between smoking and kidney disease (n = 3), inability to find a Japanese translator (n = 1), and duplicate cohort (n = 1). Seventeen studies were included in the final analysis; seven studies found an overall significant association between smoking and incident chronic kidney disease, and three studies found a significantly increased risk of chronic kidney disease in current smokers that was gender and/or dose related. An increased risk of developing chronic kidney disease among smokers was significantly associated with male gender (relative risk 2.4, 95% confidence interval 1.2-4.5), >20 cigarettes smoked/day (odds ratio 1.51, 95% confidence interval 1.06-2.15, and relative risk 2.3, 95% confidence interval 1.2-4.3), and smoking >40 years (odds ratio 1.45, 95% confidence interval 1.00-2.09). A pooled estimate of the relative risk (meta-analysis) was deemed inappropriate due to the heterogeneity in methodologies utilized by the different studies. This comprehensive review reveals overall evidence for current cigarette smoking as a risk factor for incident chronic kidney disease. Further investigation is needed to more carefully examine the strength of the association between cigarette smoking and incident kidney disease.


Genetic Variation and Structure in Contrasting Geographic Distributions: Widespread Versus Restricted Black-Tailed Prairie Dogs (Subgenus Cynomys).
Species of restricted distribution are considered more vulnerable to extinction because of low levels of genetic variation relative to widespread taxa. Species of the subgenus Cynomys are an excellent system to compare genetic variation and degree of genetic structure in contrasting geographic distributions. We assessed levels of genetic variation, genetic structure, and genetic differentiation in widespread Cynomys ludovicianus and restricted C. mexicanus using 1997bp from the cytochrome b and control region (n = 223 C. ludovicianus; 77 C. mexicanus), and 10 nuclear microsatellite loci (n = 207 and 78, respectively). Genetic variation for both species was high, and genetic structure in the widespread species was higher than in the restricted species. C. mexicanus showed values of genetic variation, genetic structure, and genetic differentiation similar to C. ludovicianus at smaller geographic scales. Results suggest the presence of at least 2 historical refuges for C. ludovicianus and that the Sierra Madre Occidental represents a barrier to gene flow. Chihuahua and New Mexico possess high levels of genetic diversity and should be protected, while Sonora should be treated as an independent management unit. For C. mexicanus, connectivity among colonies is very important and habitat fragmentation and habitat loss should be mitigated to maintain gene flow.


Bioterrorism and the people: how to vaccinate a city against panic.
Bioterrorism policy discussions and response planning efforts have tended to discount the capacity of the public to participate in the response to an act of bioterrorism, or they have assumed that local populations would impede an effective response. Fears of mass panic and social disorder underlie this bias. Although it is not known how the population will react to an unprecedented act of bioterrorism, experience with natural and technological disasters and disease outbreaks indicates a pattern of generally effective and adaptive collective action. Failure to involve the public as a key partner in the medical and public-health response could hamper effective management of an epidemic and increase the likelihood of social disruption. Ultimately, actions taken by nonprofessional individuals and groups could have the greatest influence on the outcome of a bioterrorism event. Five guidelines for integrating the public into bioterrorism response planning are proposed: (1) treat the public as a capable ally in the response to an epidemic, (2) enlist civic organizations in practical public health activities, (3) anticipate the need for home-based patient care and infection control, (4) invest in public outreach and communication strategies, and (5) ensure that planning reflects the values and priorities of affected populations.


Vaping Associated Pulmonary Injury (VAPI) with superimposed Mycoplasma pneumoniae infection.
There is growing evidence that vaping has the potential to cause adverse health effects. Vaping is affecting the younger and healthier population which is a public concern. Mycoplasma pneumoniae pneumonia is a benign condition and is usually underdiagnosed and is managed in an outpatient setting. Here we present a case of fulminant MPP in a young adult probably associated with VAPI. A 24-year-old woman presented to our hospital for severe hypoxic respiratory failure needing intubation and intensive care unit admission. She had a history for vaping for 2 years prior to presentation. She had fever and an elevated white count. Her Chest X-Ray and CT scan of the chest were consistent with bilateral predominantly lower lobe patchy opacities. She had mildly elevated serum LDH and Urine toxicology screen was positive for THC. Serum IgM Mycoplasma level was positive and her BAL fluid analysis showed lipid-laden macrophages. She was diagnosed as a probable case of VAPI per CDC guidelines with superimposed fulminant MPP. Vaping is known to increase the risk of viral and bacterial pneumonia by compromising the respiratory local immune response. Vaping also causes lipoid pneumonia where the alveoli are filled with lipid-laden macrophages with surrounding inflammation. We hypothesize that this patient had fulminant MPP in the setting of background VAPI. The association between vaping and MPP infection has not been established in the literature and this is the first documented report to establish a link between e-cigarettes and fulminant MPP. Further research is needed to confirm this association.


Bacterial, fungal and parasitic contamination of cockroaches in public hospitals of Hamadan, Iran.
To determine the possible role of cockroaches in dissemination of medically important microorganisms, a study was carried out in public hospitals and residential areas of Hamadan city, west of Iran. Bacteria, fungi and parasites of medical importance were isolated and identified. The total number of Blattella germanica collected from hospitals were 133 as the case group. The cockroaches collected from residential areas were 45 as the control group. A total of 178 cockroaches were collected, over a period of two years (133 from hospitals; and 45 from residential areas) in Hamadan. Medically important microorganisms were isolated from external and internal surfaces using standard methods. In the case group, 130 out of 133 (98%) Blattella germanica showed contamination with high bacterial load (more than 1 x 10(3)) whereas only 2 out of 45 (4.45%) cockroaches of the control group were carrying medically important bacteria with high bacterial load. Bacteriological examinations revealed that almost all test cockroaches had at least one of the following microorganisms either in their body surface or digestive tract. Enterobacter (22.6%), Klebsiela (21%), Enterococcus (17.3%), Staphylococcus (16.5%), Esherichia coli and Streptococcus (8.3%), Pseudomonas (3%), and also Shigella, Haemophilus and group A beta-hemolytic Streptococcus each less than 1%. In addition the results showed (74.4.%) of test cockroaches harboured fungi-Candida (48.9%), Mucor (10.5%), Aspergillus niger (7.5%), Rhizopus (4.5) and also Penicillium and Aspergillus fumigans each 1.5%. Some parasitic worms of medical importance were also isolated from the test cockroaches, but carriage rates were low. The data from this study emphasise the importance of cockroaches as potential vectors of medically important microorganisms such as pathogenic bacteria and fungi in hospital environments.


Cytotoxicity analysis of pre- and post-hurricane harvey soil samples collected from greater houston bayous.
Rapid urbanization, anthropogenic pollution and frequent flooding events are affecting the soil and water quality along the streams and bayous of Houston. Soil acts as sink and reservoir of heavy metals and nutrients affecting human and animal health. The objectives of the study are 1) to analyze the effects of the metal and nutrient concentration of bayou flood plain surface soil samples on the gut cell cytotoxicity and 2) to evaluate the spatial and temporal difference in soil contamination on cell viability of colon cancer (HT-29) and normal colon epithelial (CCD 841 CoN) cell lines. To evaluate soil contamination between pre- and post-hurricane (Summer and Fall) conditions in six Bayous (Brays, Buffalo, Halls, Hunting, Greens and White Oak Bayous) of Harris County, Texas, in vitro bioassay analysis was applied to soil extracts. The MTT assay determined that, with increase in concentration of Bayou soil from 12.5% to 100%, the viability of CCD 841 CoN and HT-29 cells decreased significantly, across all sampling locations during both summer and fall seasons. Among all the bayous, the viability of CCD 841 CoN cells in summer and fall followed the pattern of White Oak > Greens > Halls > Brays Bayou, where the viability of cells exposed to White Oak soils was 3-4 times higher than cells exposed to Brays Bayou soil at 100% soil concentration. The viability of HT-29 cells in both seasons followed the pattern of Greens > White Oak > Halls > Brays Bayou, where the viability of cells exposed to Greens Bayou soil was more than 3-4 times higher than the cells exposed to Brays Bayou soil at 100% concentration. The higher concentration of metals and nutrients such as P, Zn, Cd, and Cu might have contributed to the significant cell lethality in Brays Bayou samples compared to other locations.


Dermatitis herpetiformis and celiac disease associated with Addison's disease.
We treated two patients with dermatitis herpetiformis and Addison's disease, and one patient with celiac disease without the rash, but with Addison's disease and juvenile diabetes. In two of the patients, the concomitant diseases also included a thyroid disease. The predisposing factors to the multiple endocrine disorders in these patients with gluten-sensitive skin and/or small intestinal disease remained unknown. Two of the three patients had HLA-B8, none was known to have affected relatives, and the Addison's disease appeared before, at the same time, or after the patients contracted dermatitis herpetiformis or celiac disease.


A study of the incidence of porcine taeniasis and other intestinal parasites in a rural community of South Western Nigeria.
In the course of an investigation into the incidence of porcine taeniasis among humans and and pigs which live in close contact with each other in a rural town of South-Western Nigeria, no incidence of the parasite was found. Ascaris, Strongyle (hookworm in case of humans) and Trichuris were the helminths encountered while Entamoeba histolytica and Entamoeba coli were the common protozoan cysts. Polyparasitism was common and a 12 year old boy had Salmonella infection concurrently with Ascaris and hookworm. Analysis of hospital records of the town for ten years (1972-81) showed only 2 cases out of 1665 patients diagnosed as positive for Taenia infection through faecal examination. Abattoir records of incidence of bovine and porcine cysticercosis in various towns of Oyo State also showed very low incidence in these animals. It is concluded that the rural town of Erua and the Oyo state in which it is locates are not endemic for porcine and bovine taeniasis.


Impaired glycolytic metabolism causes chondrocyte hypertrophy-like changes via promotion of phospho-Smad1/5/8 translocation into nucleus.
Hypertrophy-like changes are often observed in chondrocytes during the development of osteoarthritis (OA). These changes play a crucial part in the OA-associated cartilage degradation and osteophyte formation. However, the pathogenesis leading to such changes is still unknown. In this study, we investigated the mechanism by which these hypertrophy-like changes are induced from the viewpoint of impaired glycolytic metabolism. The effect of sodium fluoride (NaF) on glycolytic metabolism of cultured chondrocytes was confirmed by measurement of intracellular adenosine triphosphate (ATP) production. Translocation of phosphorylated Smad1/5/8 to the nucleus was evaluated by subcellular fractionation and Western blotting. Chondrocyte hypertrophy-like changes were investigated by real-time RT-PCR and Western blot analysis of differentiation markers. ATP production was dose-dependently decreased by NaF in the human chondrocytic cell line HCS-2/8. In addition, both chondrocyte proliferation and differentiation were inhibited, whereas cell death was promoted by treatment with NaF. Interestingly, combinational treatment with NaF and lactate enhanced translocation of phospho-Smad1/5/8 to the nucleus, as well as gene expression of ALP, VEGF, COL10a1, and matrix metalloproteinase13 (MMP13), which were the markers of late mature and hypertrophic chondrocytes. Furthermore, the production of type X collagen and activation of MMP9 were also promoted under the same conditions. These findings suggest that decreased ATP production by NaF promotes hypertrophy-like changes via activation of phospho-Smad1/5/8 in the presence of lactate. Novel metabolic aspects of OA pathogenesis are indicated herein.


Histone post-translational modifications in Silene latifolia X and Y chromosomes suggest a mammal-like dosage compensation system.
Silene latifolia is a model organism to study evolutionary young heteromorphic sex chromosome evolution in plants. Previous research indicates a Y-allele gene degeneration and a dosage compensation system already operating. Here, we propose an epigenetic approach based on analysis of several histone post-translational modifications (PTMs) to find the first epigenetic hints of the X:Y sex chromosome system regulation in S. latifolia. Through chromatin immunoprecipitation we interrogated six genes from X and Y alleles. Several histone PTMS linked to DNA methylation and transcriptional repression (H3K27me3, H3K23me, H3K9me2 and H3K9me3) and to transcriptional activation (H3K4me3 and H4K5, 8, 12, 16ac) were used. DNA enrichment (Immunoprecipitated DNA/input DNA) was analyzed and showed three main results: (i) promoters of the Y allele are associated with heterochromatin marks, (ii) promoters of the X allele in males are associated with activation of transcription marks and finally, (iii) promoters of X alleles in females are associated with active and repressive marks. Our finding indicates a transcription activation of X allele and transcription repression of Y allele in males. In females we found a possible differential regulation (up X1, down X2) of each female X allele. These results agree with the mammal-like epigenetic dosage compensation regulation.


Diagnostic accuracy of extended biopsies for the staging of microfocal prostate cancers in autopsy specimen.
Clinically insignificant prostate cancers may be predicted when biopsies show a microfocal cancer (MiFC). However, at least one-third of MiFC are underestimated by biopsies. The aim of this study was to evaluate the staging accuracy of different biopsy regimen showing a MiFC. We performed 18 biopsy cores on 164 autopsy prostates. Six cores were taken from the mid-peripheral zone (MPZ), 6 from the lateral PZ (LPZ) and 6 from the central zone (CZ). We tested seven different biopsy regimens by distinguishing the MPZ, LPZ or CZ biopsies either separately or associated with each other. Of the cancers detected by biopsies, we selected those showing a MiFC and compared our findings with whole mount analysis. The positive predictive value of a MiFC referred to how often, when needle biopsies showed a MiFC, there was a clinically insignificant cancer on whole mount prostate analysis. We found that the positive predictive value of a MiFC on 6 or 12 biopsy cores was similar irrespective of biopsy location (P approximately 1). On MPZ, MPZ plus LPZ and all 18 biopsies, it was 40, 70 and 87%, respectively (P<0.1). Tumor volume of cancers showing a MiFC on MPZ biopsies was significantly higher than those showing a MiFC on MPZ plus LPZ, or all 18 biopsies (P<0.05). These results show that performing additional cores in case of MiFC on sextant biopsies may help differentiating significant from insignificant cancers.


[Hebephilia as a sexual disorder].
The term "hebephilia" describes the sexual preference for the body scheme of pubescent minors (Tanner stages 2 and 3). For most clinicians the definition of hebephilia as a sexual disorder is not obvious. In all assessed males included in the Prevention Project Dunkelfeld at the Institute for Sexual Medicine at the Charité between 2005 and 2011, who met the inclusion criteria and showed no evidence for exclusion criteria (n=222), the existence of a hebephilia was examined. Approximately two thirds of the present sample (n=153) showed responsiveness for the body scheme of pubescent minors. Of these, only 15% were exclusively attracted by the pubescent body scheme and 85% were cases of mixed types. Concerning the clinical aspects of the sexual preference disorder, about 95% reported child sexual abuse and/or having used child abusive images at least once in their lifetime. Additionally, hebephiles reported a higher level of clinical/psychological stress and more distinct personality characteristics in relation to a comparative sample. In terms of risk factors, hebephiles showed more offense-supportive attitudes compared to a male comparative sample. Hebephilia is a sexual disorder, but cannot be independently coded in the presently valid classification systems (DSM-IV-TR and ICD-10). Plans to separately include this in future in the DSM-5 represent an important step from a sexological point of view.


Evaluation of sufentanil as a supplement to anaesthesia in lengthy spinal surgery.
The efficacy of sufentanil citrate as a supplement to anaesthesia was studied in 31 patients undergoing lengthy spinal surgery (mean duration 150 min). Anaesthesia was induced with 1 microgram/kg sufentanil (bolus) and 2 mg/kg thiopental, followed by 1.5 mg/kg succinylcholine to facilitate intubation, all given intravenously; after intubation, administration of N2O:O2 (70:30) was started. Further doses of sufentanil (10-25 micrograms) were given, on average 2.3 times, depending on the length of the operation, systolic blood pressure, pulse rate and other autonomous signs; the mean total amount of sufentanil administered was 126.3 micrograms. Our findings showed that sufentanil provided good cardiovascular stability and rapid recovery. More specifically mean arterial blood pressure and heart rate were significantly reduced 1 min after intubation (P < or = 0.001). Thereafter mean arterial blood pressure increased, returning to preanaesthesia values and remaining constant while heart rate remained constant throughout the operation at values similar to the value immediately after induction of anaesthesia. Recovery was rapid, awakening time was 6 min or less in 50% of patients and 10 min or less in 90% of patients, while the corresponding values for response time were 8 and 12 min, for extubation time, 9 and 13 min, and for orientation time, 15 and 19 min. Two patients experienced nausea and vomiting and one patient mild chest-wall rigidity. Finally, respiratory depression was not observed in any of the patients.


Use of the Stroop phenomenon as a diagnostic tool for malingering.
To assess a computerised version of the Stroop test for detection of malingering of cognitive deficit. Sixty subjects were assessed using this test. Twenty had cognitive deficits due to brain damage of miscellaneous aetiologies. Ten were healthy, not acquainted with the test, and were asked to simulate cognitive impairment. Another 10 simulators were psychology students trained in the use of the test. Twenty healthy subjects served as controls. Results were analysed for reaction time, error percentage, and the Stroop effect. There was a significant difference in reaction time among groups, showing a direct relation of age among control subjects, and also longer reaction time in patients with brain damage than in controls. Controls and patients with brain damage showed a clear Stroop effect. Simulators had a significantly prolonged reaction time, increased error percentage, and inverted or absent Stroop effect. This alteration of the Stroop effect is never present in organic cognitive deficits and seems to be a characteristic pattern of feigning, independently of knowledge of the test. This technique is recommended as a valuable tool to detect feigned cognitive impairment.


Transcanal Transpromontorial Acoustic Neuroma Surgery: Results and Facial Nerve Outcomes.
Recently, the transcanal approach for the removal of acoustic neuromas has been introduced. Facial nerve (FN) preservation is one of the main challenges of this kind of surgery. To describe our experience in the surgical treatment of acoustic neuromas, focusing on the functional results of FN preservation after a transcanal approach. A retrospective chart review was carried out on clinical data and videos from operations on 49 patients who underwent surgery with a totally transcanal exclusive endoscopic approach for Koos stage I-II lesions, or an enlarged transcanal transpromontorial approach for Koos stage II-III tumors, between March 2012 and February 2017. Patients and tumor characteristics, clinical manifestations, radiologic features, audiological results, FN outcomes (according to the House-Brackmann [HB] grading system) and complications were evaluated. Tumors were classified according to the Koos grading system. The age of the patients (34 females and 15 males) ranged from 27 to 77 years (mean age: 54.9 yr). Preoperative diagnosis was "vestibular schwannoma" in all patients. At the last follow-up (range 1-60 mo, mean 13.9 mo), 42 of 49 showed grade I HB FN function, 5 of 49 grade II HB, and 2 of 49 grade III HB. Overall, in 95.9%, FN function was preserved (grade I-II HB) with stable results at follow-up; in 4.1% of cases, FN function was reduced, but not worse than grade III. The transcanal approach represents a feasible, minimally invasive, and conservative technique for the management of acoustic neuromas of the internal auditory canal.


Comparison between Intravenous Prostaglandin E2 and Extraamniotic Prostaglandin F2alpha Instillation for Termination in Second-Trimester Pregnancy.
> Objective: To evaluate the efficacy and safety of two different methods, intravenous prostaglandin E2 and extraamniotic prostaglandin F2alpha instillation, in second-trimester pregnancy termination. Methods: We designed a prospective randomized longitudinal study. 130 consecutive patients with various indications for second-trimester pregnancy termination were recruited. Patients were managed randomly with either intravenous continuous prostaglandin E2 infusion or extraamniotic prostaglandin F2alpha instillation. Laminaria were inserted in patients with unfavorable cervixes. The instillation-abortion time, success rate within 24 hours, dosage of both medications, and side effects were recorded and analyzed. Results: There was a significantly shorter instillation-abortion time (11.85 +/- 9.65 versus 22.18 +/- 16.83 hours, P < 0.001), higher complete abortion rate (71.91% versus 41.5%, P = 0.013), and a higher rate of successful abortion within 24 hours (87.6% versus 56.1%, P < 0.001) in patients treated with intravenous prostaglandin E2 than in those with extraamniotic prostaglandin F2alpha. Conclusions: Prostaglandin E2 had a higher rate of successful abortion and fewer side effects than prostaglandin F2alpha. This implies that intravenous prostaglandin E2 might be a better choice for second-trimester pregnancy termination compared with extraamniotic prostaglandin F2alpha.


Aqua lymphatic therapy in women who suffer from breast cancer treatment-related lymphedema: a randomized controlled study.
Lymphedema is an adverse effect of breast cancer surgery. Aqua lymphatic therapy (ALT) is a novel treatment for limb volume reduction. The aim of this study was to examine whether ALT is a safe method and whether there are differences in adherence, limb volume, and quality of life between women who perform only self-management treatment and women who participate as well in ALT. Design of the study was single-blind randomized clinical trial. The setting was in a hydrotherapy pool, 1.2 m depth, and a temperature of 32-33 degrees capital ES, Cyrillic. Forty-eight women (56 +/- 10 years), with a 12.8% lymphedema relative volume, participated in the study. The control group was instructed to perform the self-management treatment. The study group joined a weekly session of ALT for 3 months in addition to the self-management therapy. Adherence was assessed by a self-reported diary, limb volume by a water displacement device, quality of life by the Upper Limb Lymphedema Questionnaire (ULL27), prior to, and after the intervention period. There was no episode of arm infection or aggravation in limb volume during the study period. ALT had a positive, statistically and clinically significant immediate effect on limb volume but no long-term effect was noted. The adherence rate to ALT was significantly higher than the adherence to self-management therapy. QOL improved in the study group. ALT was found to be a safe method, with high adherence, in treating women who suffer from mild to moderate lymphedema. A significant immediate and insignificant long-term effect on limb volume was noted.


Phase I Study of the Anti-CD22 Antibody-Drug Conjugate Pinatuzumab Vedotin with/without Rituximab in Patients with Relapsed/Refractory B-cell Non-Hodgkin Lymphoma.
Purpose: Pinatuzumab vedotin is an antibody-drug conjugate with the potent antimicrotubule agent monomethyl auristatin E (MMAE) conjugated to an anti-CD22 antibody via a protease-cleavable linker. This phase I study determined its recommended phase II dose (RP2D) and evaluated its safety, tolerability, and antitumor activity alone and with rituximab in relapsed/refractory (r/r) non-Hodgkin lymphoma (NHL) and chronic lymphocytic leukemia (CLL).Experimental Design: Patients received escalating doses of pinatuzumab vedotin every 21 days. Clinical activity at the RP2D alone or with rituximab was evaluated in r/r diffuse large B-cell lymphoma (DLBCL) and r/r indolent NHL (iNHL) patients.Results: Seventy-five patients received single-agent pinatuzumab vedotin. The RP2D was 2.4 mg/kg, based on dose-limiting toxicities (DLT) of grade 4 neutropenia >7 days in 1 of 3 patients and grade 4 neutropenia <7 days in 2 of 3 patients treated at 3.2 mg/kg (maximum assessed dose). No DLTs occurred at 2.4 mg/kg. At the RP2D, neutropenia was the most common grade ≥3 adverse event. Peripheral neuropathy-related grade ≥2 adverse events most frequently resulted in treatment discontinuation. Rituximab cotreatment did not impact safety, tolerability, or pharmacokinetics of pinatuzumab vedotin. Unconjugated MMAE exposure was much lower than antibody-conjugated MMAE exposure, without accumulation with repeat dosing. At the RP2D, objective responses were observed in DLBCL (9/25) and iNHL (7/14) patients; 2 of 8 patients treated with pinatuzumab vedotin (RP2D) and rituximab had complete responses. CLL patients showed no objective responses.Conclusions: The RP2D of pinatuzumab vedotin alone and with rituximab was 2.4 mg/kg, which was well tolerated, with encouraging clinical activity in r/r NHL. Clin Cancer Res; 23(5); 1167-76. ©2016 AACR.


Macrophages in dermal disease progression of phospholipase D4-deficient Fleckvieh calves.
A new gene defect in Fleckvieh calves leads to a syndrome with partial phenotype overlap with bovine hereditary zinc deficiency. A mutation in a gene encoding phospholipase D4 (PLD4), an endosomal exonuclease, causes the disorder. In mice, PLD4 activity indirectly regulates the Toll-like receptor 9 (TLR9) pathway via degradation of microbial DNA. PLD4 absence thus results in visceral macrophage activation comparable to human macrophage activation syndrome. In this study, disease progression and the role of macrophages in affected calves were monitored clinically, clinicopathologically, and histologically over time. Breeding data identified 73 risk matings of heterozygous carriers resulting in 54 potentially PLD4-deficient calves born on farms. PLD4 status was examined via 5'-exonuclease assay, detecting 6 calves carrying the defect. These were purchased and monitored daily until final necropsy. The calves developed progressive skin lesions starting with small scaling areas terminating in severe crusting dermatitis, especially in areas with mechanical exposure. Histological and immunohistochemical analyses indicated that macrophages with cytoplasmic vacuolation increased considerably in skin sections obtained weekly during the disease course. Macrophage increase correlated with increased dermal lesion severity. Macrophage activation was confirmed by prominent phagocytic activity in the superficial dermis using electron microscopy. Dermal mRNA abundance of CCL2 and CCL3 measured by quantitative polymerase chain reaction verified macrophage activation. Further increase in mRNA of downstream molecule MyD88 and cytokine IL12b connected bovine PLD4 deficiency to increased TLR9 pathway activation. In contrast to human macrophage activation syndrome, the main feature of bovine PLD4 deficiency was local disease in organs with contact to microbial DNA (skin, intestine, lungs).


Cost-effectiveness analysis of the use of high-flow oxygen through nasal cannula in intensive care units in NHS England.
To estimate the cost-effectiveness of Nasal High Flow (NHF) in the intensive care unit (ICU) compared with standard oxygen or non-invasive ventilation (NIV) from a UK NHS perspective. Three cost-effectiveness models were developed to reflect scenarios of NHF use: first-line therapy (pre-intubation model); post-extubation in low-risk, and high-risk patients. All models used randomized control trial data on the incidence of intubation/re-intubation, events leading to intubation/re-intubation, mortality and complications. NHS reference costs were primarily used. Sensitivity analyses were conducted. When used as first-line therapy, Optiflow™ NHF gives an estimated cost-saving of £469 per patient compared with standard oxygen and £611 versus NIV. NHF cost-savings for high severity sub-group were £727 versus standard oxygen, and £1,011 versus NIV. For low-risk post-intubation patients, NHF generates estimated cost-saving of £156 versus standard oxygen. NHF decreases the number of re-intubations required in these scenarios. Results were robust in most sensitivity analyses. For high-risk post-intubation patients, NHF cost-savings were £104 versus NIV. NHF results in a non-significant increase in re-intubations required. However, reduction in respiratory failure offsets this. For patients in ICU who are at risk of intubation or re-intubation, NHF cannula is likely to be cost-saving.


Altered Plasma Proteins in Myogenous Temporomandibular Disorders.
The aims of this study were (1) to compare the levels and interactions of several plasma proteins in patients with myogenous temporomandibular disorders (TMDM) compared to healthy and pain-free controls, (2) to compare the levels and interactions in two TMDM subgroups, myalgia (MYA) and myofascial pain (MFP), and (3) to explore associations between the proteins and clinical data. Thirty-nine patients with TMDM (MFP, n = 25, MYA, n = 14), diagnosed according to the diagnostic criteria for TMD (DC/TMD), aged 38 years, and sex-matched pain-free controls completed an extended DC/TMD Axis II questionnaire and the plasma concentration of 87 biomarkers were analyzed. Nine proteins separated TMDM from controls (p = 0.0174) and 12 proteins separated MYA from MFP (p = 0.019). Pain duration, characteristic pain intensity, pain catastrophizing, perceived stress, and insomnia severity were significantly associated with protein markers (p < 0.001 to p < 0.022). In conclusion, several plasma proteins were upregulated in TMDM and either upregulated or downregulated in MYA compared to MFP. Some proteins in TMDM were associated with pain variables, sleep disturbance, and emotional function. These results show that systemic differences in protein expression exist in patients with TMDM and that altered levels of specific plasma proteins are associated with different clinical variables.


Synovial fluid mononuclear cells provide an environment for long-term survival of antibody-secreting cells and promote the spontaneous production of anti-citrullinated protein antibodies.
In rheumatoid arthritis (RA), observations point to a crucial role for (autoreactive) B cells in disease pathogenesis. Here, we studied whether cells from the synovial environment impact on the longevity of autoreactive B cell responses against citrullinated antigens. Synovial fluid mononuclear cells and peripheral blood mononuclear cells (SFMC/PBMC) were obtained from patients with established RA and assessed for the presence of B cell subpopulations. Cells spontaneously secreting anti-citrullinated protein antibodies (ACPA-IgG) directly ex vivo were detected by antigen-specific Enzyme-Linked ImmunoSpot (ELISpot) assay. SFMC and PBMC were cultured to assess the degree of spontaneous ACPA-IgG secretion. Cells surviving for several weeks were characterised by carboxyfluorescein succinimidyl ester (CFSE) labelling and Ki-67 staining. Cells spontaneously secreting ACPA-IgG were readily detectable in peripheral blood and synovial fluid (SF) of patients with ACPA-positive RA. SFMC showed an up to 200-fold increase in ex vivo ACPA-IgG secretion compared with PBMC despite lower numbers of B cells in SFMC. ELISpot confirmed the presence of spontaneously ACPA-IgG-secreting cells, accounting for up to 50% (median 12%) of all IgG-secreting cells in SF. ACPA-IgG secretion was remarkably stable in SFMC cultures, maintained upon depletion of the CD20+ B cell compartment and detectable for several months. CFSE labelling and Ki-67 staining confirmed the long-term survival of non-dividing plasma cells (PCs). This study demonstrates a high frequency of differentiated, spontaneously ACPA-IgG-secreting cells in SF. These cells are supported by SFMC for prolonged survival and autoantibody secretion, demonstrating that the synovial compartment is equipped to function as inflammatory niche for PC survival.


Correlation between increased AP-1NGF binding activity and induction of nerve growth factor transcription by multiple signal transduction pathways in C6-2B glioma cells.
Transcription mechanisms regulating nerve growth factor (NGF) gene expression in the CNS are yet to be thoroughly understood. We have used C6-2B rat glioma cells to characterize the signal transduction pathways that contribute to transcriptional and posttranscriptional regulation of NGF mRNA. Because the NGF promoter contains an AP-1 consensus sequence, we have investigated whether increases in AP-1 binding activity correlate with enhanced NGF mRNA expression. Gel mobility shift assays using an oligonucleotide homologous to the AP-1 responsive element of the rat NGF gene (AP-1NGF) revealed that 12-O-tetradecanoyl phorbol-13-acetate (TPA) and, to a lesser extent, isoproterenol (ISO) and thapsigargin, a microsomal Ca(2+)-ATPase inhibitor, stimulated binding to AP-1NGF within 2 h. All of these stimuli increased NGF mRNA levels within 3 h. Cycloheximide pretreatment blocked the TPA and ISO-mediated binding to AP-1NGF suggesting that de novo synthesis of c-Fos/c-Jun may be required for the transcriptional regulation of NGF gene. Nuclear run-on assays and NGF mRNA decay studies revealed that TPA increases NGF transcription whereas ISO affects both transcription and mRNA stabilization. We propose that (i) different signal transduction mechanisms regulate the expression of the NGF gene in cells derived from the CNS, and (ii) both mRNA transcription and stability account for the cAMP-mediated increase in NGF mRNA levels.


Sexual Compulsivity Scale, Compulsive Sexual Behavior Inventory, and Hypersexual Disorder Screening Inventory: Translation, Adaptation, and Validation for Use in Brazil.
Epidemiological, behavioral, and clinical data on sexual compulsivity in Brazil are very limited. This study sought to adapt and validate the Sexual Compulsivity Scale (SCS), the 22-item version of the Compulsive Sexual Behavior Inventory (CSBI-22), and the Hypersexual Disorder Screening Inventory (HDSI) for use in Brazil. A total of 153 participants underwent psychiatric assessment and completed self-reported measures. The adaptation process of the instruments from English to Portuguese followed the guidelines of the International Society for Pharmacoeconomics and Outcomes Research. The reliability and validity of the HDSI criteria were evaluated and the construct validity of all measures was examined. For the SCS and HDSI, factor analysis revealed one factor for each measure. For the CSBI-22, four factors were retained although we only calculated the scores of two factors (control and violence). All scores had good internal consistency (alpha >.75), presented high temporal stability (>.76), discriminated between patients and controls, and presented strong (ρ > .81) correlations with the Sexual Addiction Screening Test (except for the violence domain = .40) and moderate correlations with the Impulsive Sensation Seeking domain of the Zuckerman Kuhlman Personality Questionnaire (ρ between .43 and .55). The sensitivity of the HDSI was 71.93 % and the specificity was 100 %. All measures showed very good psychometric properties. The SCS, the HDSI, and the control domain of the CSBI-22 seemed to measure theoretically similar constructs, as they were highly correlated (ρ > .85). The findings support the conceptualization of hypersexuality as a cluster of problematic symptoms that are highly consistent across a variety of measures.


SWI/SNF chromatin remodeling subunit Smarca4/BRG1 is essential for female fertility†.
Mammalian folliculogenesis is a complex process that involves the regulation of chromatin structure for gene expression and oocyte meiotic resumption. The SWI/SNF complex is a chromatin remodeler using either Brahma-regulated gene 1 (BRG1) or BRM (encoded by Smarca4 and Smarca2, respectively) as its catalytic subunit. SMARCA4 loss of expression is associated with a rare type of ovarian cancer; however, its function during folliculogenesis remains poorly understood. In this study, we describe the phenotype of BRG1 mutant mice to better understand its role in female fertility. Although no tumor emerged from BRG1 mutant mice, conditional depletion of Brg1 in the granulosa cells (GCs) of Brg1fl/fl;Amhr2-Cre mice caused sterility, whereas conditional depletion of Brg1 in the oocytes of Brg1fl/fl;Gdf9-Cre mice resulted in subfertility. Recovery of cumulus-oocyte complexes after natural mating or superovulation showed no significant difference in the Brg1fl/fl;Amhr2-Cre mutant mice and significantly fewer oocytes in the Brg1fl/fl;Gdf9-Cre mutant mice compared with controls, which may account for the subfertility. Interestingly, the evaluation of oocyte developmental competence by in vitro culture of retrieved two-cell embryos indicated that oocytes originating from the Brg1fl/fl;Amhr2-Cre mice did not reach the blastocyst stage and had higher rates of mitotic defects, including micronuclei. Together, these results indicate that BRG1 plays an important role in female fertility by regulating granulosa and oocyte functions during follicle growth and is needed for the acquisition of oocyte developmental competence.


[Experimental herpetic infection of the CNS with predominant involvement of the spinal cord].
Clinical, morphological, and virological characteristics of a model of herpes infection of the CNS with predominant involvement of the spinal cord, an acute ascending myelitis with primary demyelinization, are presented. The development of spinal cord lesions after HSV inoculation into the spleen was observed in different species of laboratory animals: mice, hamsters, guinea pigs, and rabbits; some animals (30%-70%) developed generalized infection with the involvement of the brain. The frequency of the spinal cord involvement was found to depend on the age of the animal and the dose of the virus-containing inoculum. Differences in the clinical and morphological manifestations of herpes infection were due to different mechanisms of herpes simplex virus spread in the body: either centripetally on the axons of the visceral nerves innervating the spleen into the thoracal cord, or hematogenic disseminations with virus penetration across the hematoencephalic barrier. The total duration of the disease was 3-12 days depending on the animal species.


Effect of growth factors on Escherichia coli alpha-hemolysin-induced mediator release from human inflammatory cells: involvement of the signal transduction pathway.
Previously, we have shown that Escherichia coli alpha-hemolysin represents a potent stimulus for inflammatory mediator release (O2- release, beta-glucuronidase release, and leukotriene generation) from human polymorphonuclear granulocytes (PMN) as well as for histamine release from a human lymphocyte-monocyte-basophil cell suspension (LMB). In contrast, the E. coli alpha-hemolysin leads to a downregulation of cytokine release (interleukin 6 [IL-6], tumor necrosis factor alpha, and IL-1 beta) from human LMB. This study was undertaken (i) to analyze the priming efficacy of growth factors (granulocyte-macrophage colony-stimulating factor [GM-CSF] and granulocyte CSF [G-CSF]) on inflammatory mediator release from human PMN and LMB challenged with hemolysin-producing E. coli bacteria as well as with cell-free E. coli alpha-hemolysin and (ii) to identify major components involved in GM-CSF and G-CSF priming. GM-CSF pretreatment led to an increased chemiluminescence response from human PMN by up to 100%, leukotriene B4 generation was enhanced up to fivefold, and histamine release from human LMB increased from 45% +/- 15% to 75% +/- 5% (mean +/- standard distribution) of the total histamine content. G-CSF priming induced an increase in the chemiluminescence response by up to 50% +/- 5% from human PMN and an increase in histamine release from human LMB by 20% +/- 5%. The growth factors, GM-CSF and G-CSF, modulated neither beta-glucuronidase release from human PMN nor IL-8 release from human PMN and LMB challenged with the E. coli alpha-hemolysin. GM-CSF and G-CSF pretreatment increased the fluoride (NaF)-induced chemiluminescence response by up to 10-fold; the serine/threonine phosphatase inhibitor okadaic acid inhibited GM-CSF- and G-CSF-induced priming. NaF-induced histamine release was enhanced up to 60 and 30% by GM-CSF and G-CSF priming, respectively. GM-CSF and G-CSF pretreatment did not modulate phorbol 12-myristate 13-acetate-induced chemiluminescence response or histamine release. GM-CSF by itself induced an increase in 5-lipoxygenase-specific mRNA expression within 5 min. Our results indicate that (i) GM-CSF and G-CSF interact with inflammatory cells via distinct cellular signalling, (ii) the signal transduction pathway is dependent on the cellular mediator, and (iii) the use of growth factors may be a potent tool to influence the clinical outcome in infectious diseases.


Correlation of desmoglein 1 and 3 immunohistochemistry with autoantibody levels and clinical severity in pemphigus.
Pemphigus is a chronic potentially fatal autoimmune bullous disorder. Pemphigus vulgaris (PV) and pemphigus foliaceus (PF) are the two common subtypes. PV is the most common and aggressive type characterized by oral mucosal erosions and cutaneous lesions. PF presents with blisters on the scalp, face, and upper trunk, and spares the mucosae. Direct immunofluorescence (DIF) is the gold standard for diagnosis. Immunohistochemistry (IHC) is an emerging alternate diagnostic tool. In this study, our objectives were to identify the staining patterns of desmoglein 1 (dsg 1) and desmoglein 3 (dsg 3) IHC and to correlate the same with autoantibody levels and clinical severity in patients with PV and PF. Forty-nine clinically, histologically, and DIF-confirmed cases of pemphigus were included in the study. The IHC patterns were scored from 0 to 3+ with 3+ dsg 1 IHC exhibiting intense membranous staining in the upper layers of the epidermis and 3+ dsg 3 IHC showing intense basal layer staining. Enzyme-linked immunosorbent assay (ELISA) for anti-dsg 1 and 3 antibodies was performed in 38 cases where serum samples were available. The pemphigus disease activity index system was utilized for clinical scoring. A 0 to 1+ score was observed for dsg 1 IHC in 100% of PF cases. A score of 0 to 1+ was observed for dsg 3 IHC in 97.3% of PV cases. One hundred percent of cases with PF and 83.9% of patients with PV tested positive for ELISA anti-dsg 1 and 3 antibody titers, respectively. Anti-dsg 1 and 3 ELISA titers significantly correlated with the dsg 1 and dsg 3 IHC scores. The mucosal scores showed a significant association with both dsg 1 and 3 IHC (p < 0.001). The cutaneous scores showed a significant association with the dsg 3 IHC (p < 0.001). The IHC patterns for dsg 1 and 3 proved reliable in giving concordant results with the ELISA antibody titers and clinical severity.


Phase synchrony reveals organization in human atrial fibrillation.
It remains unclear if human atrial fibrillation (AF) is spatially nonhierarchical or exhibits a hierarchy of organization sustained by sources. We utilize activation times obtained at discrete locations during AF to compute the phase synchrony between tissue regions, to examine underlying spatial dynamics throughout both atria. We construct a binary synchronization network and show that this network can accurately define regions of coherence in coarse-grained in silico data. Specifically, domains controlled by spiral waves exhibit regions of high phase synchrony. We then apply this analysis to clinical data from patients experiencing cardiac arrhythmias using multielectrode catheters to simultaneously record from a majority of both atria. We show that pharmaceutical intervention with ibutilide organizes activation by increasing the size of the synchronized domain in AF and quantify the increase in temporal organization when arrhythmia changes from fibrillation to tachycardia. Finally, in recordings from 24 patients in AF we show that the level of synchrony is spatially broad with some patients showing large spatially contiguous regions of synchronization, while in others synchrony is localized to small pockets. Using computer simulations, we show that this distribution is inconsistent with distributions obtained from simulations that mimic multiwavelet reentry but is consistent with mechanisms in which one or more spatially conserved spiral waves is surrounded by tissue in which activation is disorganized.


Comparison of clinical manifestation of dengue fever in Bangladesh: an observation over a decade.
The clinical presentation of dengue fever had been observed to change with time since its first outbreak in 2000 in Bangladesh. This report showed the clinical presentation of the 2019 outbreak in Bangladesh along with its comparison to previous outbreaks witnessed in this region. This hospital-based cross-sectional study was conducted in one of the largest tertiary care hospitals in Dhaka city. A total of 553 laboratory-confirmed and 194 probable dengue cases were interviewed. The clinical manifestation of the confirmed cases of the current outbreak was compared with three of the outbreak reports retrieved from the databases. R version 3.6.3 was used for data analysis. Among the confirmed cases, two-thirds were male (63.2%) and the average age was 27(± 11) years. Positive tests for NS1 and IgM were present in 99.6% (n = 525/527) and 82.6% (n = 38/46) of the cases, respectively. Thrombocytopenia was present in 66.1% of cases. Fever (100%) was common for all. Gastrointestinal (GIT) features, including abdominal pain (86.5%), anorexia and/or vomiting (69.6%), and Diarrhea (> 3 motions/day) (26.2%) were more frequent than typical rash and other pain symptoms. Hypotension was present in approximately a quarter of patients (25%). GIT features (anorexia, nausea, and/or vomiting) and hypotension were more common among adult participants while bleeding manifestation (melena and vaginal bleeding, p = 0.009 & 0.032) was more frequent in pediatric patients. Compared to outbreaks of 2008, 2016, and 2018, increasing trends in GIT symptoms e.g. anorexia, abdominal pain, and diarrhea were observed. While a negative trend in hemorrhagic manifestations (skin rash, melena, and conjunctival hemorrhage/hemorrhagic sclera) and arthralgia/joint pain were found. The present outbreak was noticeably characterized by GIT symptoms and hypotension in addition to the typical clinical features like rash and pain symptoms. An increasing trend in GIT features and decreasing trend in hemorrhagic manifestations was noted over the last decade of dengue outbreaks.


The role of a cytosolic superoxide dismutase in barley-pathogen interactions.
Reactive oxygen species (ROS), including superoxide ( O2·-/ HO2·) and hydrogen peroxide (H2 O2 ), are differentially produced during resistance responses to biotrophic pathogens and during susceptible responses to necrotrophic and hemi-biotrophic pathogens. Superoxide dismutase (SOD) is responsible for the catalysis of the dismutation of O2·-/ HO2· to H2 O2 , regulating the redox status of plant cells. Increased SOD activity has been correlated previously with resistance in barley to the hemi-biotrophic pathogen Pyrenophora teres f. teres (Ptt, the causal agent of the net form of net blotch disease), but the role of individual isoforms of SOD has not been studied. A cytosolic CuZnSOD, HvCSD1, was isolated from barley and characterized as being expressed in tissue from different developmental stages. HvCSD1 was up-regulated during the interaction with Ptt and to a greater extent during the resistance response. Net blotch disease symptoms and fungal growth were not as pronounced in transgenic HvCSD1 knockdown lines in a susceptible background (cv. Golden Promise), when compared with wild-type plants, suggesting that cytosolic O2·-/ HO2· contributes to the signalling required to induce a defence response to Ptt. There was no effect of HvCSD1 knockdown on infection by the hemi-biotrophic rice blast pathogen Magnaporthe oryzae or the biotrophic powdery mildew pathogen Blumeria graminis f. sp. hordei, but HvCSD1 also played a role in the regulation of lesion development by methyl viologen. Together, these results suggest that HvCSD1 could be important in the maintenance of the cytosolic redox status and in the differential regulation of responses to pathogens with different lifestyles.


Characterization of guinea pig pulmonary neurokinin type 1 receptors using a novel antagonist ligand, [3H]FK888.
We have characterized the binding of a novel radioligand, [3H] FK888, to neurokinin (NK)1 receptors in guinea pig lung membranes and localized its binding in guinea pig lung sections by autoradiography. Lung membranes were incubated with [3H] FK888 at 25 degrees and the assays were terminated by rapid filtration; nonspecific binding was defined as binding in the presence of 1 microM concentrations of the nonpeptide NK1-selective antagonist CP-96,345. Kinetic analysis showed that specific binding of [3H] FK888 (approximately 70% of total binding) was rapid, reaching a plateau by 20 min, and that binding was reversed by addition of 1 microM CP-96,345, giving a kinetic Kd of 0.46 nM. Binding of [3H] FK888 was saturable at approximately 1 nM, and equilibrium binding analysis gave a Kd of 0.32 +/- 0.03 nM and a Bmax of 46.9 +/- 7.1 fmol/mg of protein (four experiments). In competition studies, substance P, CP-96,345, and FK888 competed for [3H] FK888 binding, but NKA, NKB, and NK2-selective antagonists such as SR48968 and L-659,877 did not. Guanosine-5'-O-(3-thio)triphosphate significantly shifted the competition curve for substance P competition against [3H]FK888 binding to a lower affinity state, confirming that NK1 receptors are coupled to a G protein. Autoradiographic mapping in cryostat sections of lung showed that [3H]FK888 binding was dense over smooth muscle of all airways, with moderate binding over epithelium of bronchi and bronchioles as well as submucosal glands of trachea. No significant labeling of blood vessels was observed. [3H]FK888 binds to NK1 receptors in guinea pig lung and may be a useful tool for studying the expression and regulation of NK1 receptors.


RAB13 mRNA compartmentalisation spatially orients tissue morphogenesis.
Polarised targeting of diverse mRNAs to cellular protrusions is a hallmark of cell migration. Although a widespread phenomenon, definitive functions for endogenous targeted mRNAs and their relevance to modulation of in vivo tissue dynamics remain elusive. Here, using single-molecule analysis, gene editing and zebrafish live-cell imaging, we report that mRNA polarisation acts as a molecular compass that orients motile cell polarity and spatially directs tissue movement. Clustering of protrusion-derived RNAseq datasets defined a core 192-nt localisation element underpinning precise mRNA targeting to sites of filopodia formation. Such targeting of the small GTPase RAB13 generated tight spatial coupling of mRNA localisation, translation and protein activity, achieving precise subcellular compartmentalisation of RAB13 protein function to create a polarised domain of filopodia extension. Consequently, genomic excision of this localisation element and perturbation of RAB13 mRNA targeting-but not translation-depolarised filopodia dynamics in motile endothelial cells and induced mispatterning of blood vessels in zebrafish. Hence, mRNA polarisation, not expression, is the primary determinant of the site of RAB13 action, preventing ectopic functionality at inappropriate subcellular loci and orienting tissue morphogenesis.


Efficient short interference RNA delivery to tumor cells using a combination of octaarginine, GALA and tumor-specific, cleavable polyethylene glycol system.
We recently developed a multifunctional envelope-type nano device (MEND) for efficient nucleic acid delivery. Here, we report on the development of an octaarigine (R8)-modified MEND encapsulating small interfering RNA (siRNA) with a tumor-specific, cleavable, polyethylene glycol (PEG)-lipid (PPD). We first determined the optimal concentration of R8 and pH-sensitive fusogenic peptide (GALA) on the lipid envelope of MEND (R8/GALA-MEND). Then, we examined the combination of optimized R8/GALA-MEND with a PEG-lipid. When a conventional PEG-lipid was used, the R8/GALA-MEND failed to knockdown expression of the target gene. On the other hand, PPD-modified R8/GALA-MEND exhibited efficient silencing activity to the level of the PEG-unmodified R8/GALA-MEND. In addition, we compared a R8/GALA-MEND with a MEND composed of 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) that is a conventional cationic lipid used as a lipoplex component. The knockdown ability of the R8/GALA-MEND was much higher than that of the DOTAP-based MEND at the dose that is commonly employed in in vitro siRNA transfection. These results demonstrate that the R8/GALA-MEND is a promising delivery system for the transfer of siRNA to tumor cells.


Analysis of the relationship between the KRAS G12V oncogene and the Hippo effector YAP1 in embryonal rhabdomyosarcoma.
Persistent hyperactivity of the Hippo effector YAP in activated satellite cells is sufficient to cause embryonal rhabdomyosarcoma (ERMS) in mice. In humans, YAP is abundant and nuclear in the majority of ERMS cases, and high YAP expression is associated with poor survival. However, YAP1 is rarely mutated in human ERMS. Instead, the most common mutations in ERMS are oncogenic RAS mutations. First, to compare YAP1 S127A and KRAS G12V-driven rhabdomyosarcomas, we re-analysed gene expression microarray datasets from mouse rhabdomyosarcomas caused by these genes. This revealed that only 20% of the up or downregulated genes are identical, suggesting substantial differences in gene expression between YAP and KRAS-driven rhabdomyosarcomas. As oncogenic RAS has been linked to YAP in other types of cancer, we also tested whether KRAS G12V alone or in combination with loss of p53 and p16 activates YAP in myoblasts. We found that neither KRAS G12V alone nor KRAS G12V combined with loss of p53 and p16 activated Yap or Yap/Taz-Tead1-4 transcriptional activity in C2C12 myoblasts or U57810 cells. In conclusion, whilst oncogenic KRAS mutation might activate Yap in other cell types, we could find no evidence for this in myoblasts because the expression of KRAS G12V expression did not change Yap/Taz activity in myoblasts and there was a limited overlap in gene expression between KRAS G12V and YAP1 S127A-driven tumours.