[Ionic mechanism of the stimulatory effect of noradrenaline on smooth muscle cells of the pulmonary artery].
Noradrenaline (NA) in a concentration of 5 X 10(-6) M produces depolarization of smooth muscle cells of the rabbit pulmonary artery and reduction of membrane resistance followed by contraction and increased excitability of muscle cells. Experiments with repolarization of the membrane exposed to NA in normal and Ca-free Krebs solutions have shown that activation of the NA-induced contraction is mainly due to Ca++ entering the cells through NA-sensitive potential-dependent Ca-channels. The NA-induced depolarization results from an initial decrease of K-permeability of the membrane subsequent increase of the permeability of NA-sensitive potential-dependent channels for Na+ and/or Cl-, which provides further depolarization of the membrane. Depolarization ceases after becoming sufficient for activation of potential-dependent non-inactivated K-channels. Voltage clamp experiments have shown that the NA-induced increased excitability is related to a reduction of slow, particularly of fast component of outward current, whose early activation prevents the development of regenerative process of action potential generation under normal conditions.


Leptin decreases gluconeogenesis and gluconeogenic substrate availability in patients with lipodystrophy.
The effects of leptin, an adipocyte-derived hormone that signals overall energy sufficiency, can only be studied in leptin-deficient conditions. In patients with lipodystrophy, a rare disease and unique model of leptin deficiency, treatment with recombinant leptin (metreleptin) improves glycemia and decreases energy expenditure. We hypothesized that these improvements might be mediated by reduced gluconeogenesis (GNG), an energy-requiring process. To determine the effects of metreleptin on GNG and GNG substrates. Single-arm prospective study of metreleptin administration in 15 patients with lipodystrophy, nine of whom had data on GNG. (NIH, 2013-2018). Total GNG, insulin-mediated suppression of GNG, glycerol, palmitate, alanine, lactate, peripheral and hepatic insulin sensitivity, and markers of glycemia (e.g., HbA1c, glucose, fasting insulin). Metreleptin administration decreased basal GNG, increased insulin-mediated suppression of GNG, and improved insulin sensitivity and markers of glycemic control. Metreleptin reduced carbon sources for GNG, including plasma alanine and lactate, and rate of appearance (Ra) of glycerol, and decreased Ra of palmitate, a driver of GNG. Glycerol and palmitate Ra correlated with GNG prior to but not during metreleptin administration. Alanine strongly correlated with GNG both before and during metreleptin administration. Metreleptin treatment in patients with lipodystrophy reduced GNG likely through decreased availability of carbon sources for gluconeogenesis, such as alanine, lactate, and glycerol. Associations between alanine and GNG persisted after metreleptin treatment while correlations with glycerol and palmitate Ra did not persist, suggesting reduced importance of lipolysis as a driver of GNG in the leptin-replete state.


Barrier inhomogeneities at vertically stacked graphene-based heterostructures.
The integration of graphene and other atomically flat, two-dimensional materials has attracted much interest and been materialized very recently. An in-depth understanding of transport mechanisms in such heterostructures is essential. In this study, vertically stacked graphene-based heterostructure transistors were manufactured to elucidate the mechanism of electron injection at the interface. The temperature dependence of the electrical characteristics was investigated from 300 to 90 K. In a careful analysis of current-voltage characteristics, an unusual decrease in the effective Schottky barrier height and increase in the ideality factor were observed with decreasing temperature. A model of thermionic emission with a Gaussian distribution of barriers was able to precisely interpret the conduction mechanism. Furthermore, mapping of the effective Schottky barrier height is unmasked as a function of temperature and gate voltage. The results offer significant insight for the development of future layer-integration technology based on graphene-based heterostructures.


Primary Tumor Sidedness, RAS and BRAF Mutations and MSI Status as Prognostic Factors in Patients with Colorectal Liver Metastases Treated with Surgery and Thermal Ablation: Results from the Amsterdam Colorectal Liver Met Registry (AmCORE).
The aim of this study was to assess primary tumor sidedness of colorectal cancer (CRC), rat sarcoma viral oncogene homolog (RAS) and v-raf murine sarcoma viral oncogene homolog B (BRAF) mutations and microsatellite instability (MSI) status as prognostic factors predicting complications, survival outcomes, and local tumor progression (LTP) following surgery and thermal ablation in patients with colorectal liver metastases (CRLM). This Amsterdam Colorectal Liver Met Registry (AmCORE) based study included 520 patients, 774 procedures, and 2101 tumors undergoing local treatment (resection and/or thermal ablation) from 2000 to 2021. Outcomes following local treatment were analyzed for primary tumor sidedness of CRC, RAS, and BRAF mutations and MSI status. The Kaplan-Meier method was used to estimate local tumor progression-free survival (LTPFS), local control (LC), distant progression-free survival (DPFS), and overall survival (OS). Uni- and multivariable analyses were performed based on Cox proportional hazards model. The chi-square test was used to analyze complications. Complications (p = 0.485), OS (p = 0.252), LTPFS (p = 0.939), and LC (p = 0.423) was not associated with tumor-sidedness. Compared to right-sided colon cancer (CC) (reference HR 1.000), DPFS was superior for left-sided CC and rectal cancer (p = 0.018) with an HR for left-sided CC of 0.742 (95% CI, 0.596-0.923) and for RC of 0.760 (95% CI, 0.597-0.966). Regarding RAS mutations, no significant difference was found in OS (p = 0.116). DPFS (p = 0.001), LTPFS (p = 0.039), and LC (p = 0.025) were significantly lower in the RAS mutation group. Though no difference in LTPFS was found between RAS wildtype and RAS mutated CRLM following resection (p = 0.532), LTPFS was worse for RAS mutated tumors compared to RAS wildtype following thermal ablation (p = 0.037). OS was significantly lower in the BRAF mutation group (p < 0.001) and in the MSI group (p < 0.001) following local treatment, while both did not affect DPFS, LTPFS, and LC. This AmCORE based study suggests the necessity of wider margins to reduce LTP rates in patients with RAS mutated CRLM, especially for thermal ablation. Upfront knowledge regarding molecular biomarkers may contribute to improved oncological outcomes.


A case series of alopecia areata in children: impact of personal and family history of stress and autoimmunity.
The epidemiology of alopecia areata (AA) is well documented in adults but has not been studied adequately in children. To evaluate the clinical and epidemiological profile of AA in children and assess the significance of thyroid screening. One hundred and fifty-seven children (83 boys, 74 girls, aged 1-16 years) who visited our clinic with a first episode of AA from 1996 to 2000 were retrospectively studied. One hundred children served as clinical controls. The age of peak incidence of AA was 0-5 years. The youngest child was 1 year old. In the majority of the cases (131/157, 83.4%) the disease was mild or moderate (less than 50% hair loss). In 15 patients (9.5%), AA was preceded by a stressful event. Five patients had a personal history of autoimmune disease (3.2 vs. 5% of the controls, (P = not significant [NS]) while 18 patients had a personal history of atopy (11.4 vs. 18% of the controls, P = NS). Twenty-one patients had a family history of autoimmune disease other than thyroiditis (13.4 vs. 5% of the controls, P = 0.04), while 23 patients had a family history of thyroid disorder (14.6 vs. 3% of the controls, P = 0.006). In eight patients (5%) subclinical hypothyroidism of autoimmune aetiology (Hashimoto's thyroiditis) was revealed at the time of investigation. Six out of the eight patients with Hashimoto's thyroiditis had a family history of thyroid disorder, which was statistically significant when compared to AA patients without thyroiditis (P < 0.001). The severity of AA was associated with early age of onset of the disease (P = 0.02). The age of peak incidence of AA in children is 0-5 years. Children with AA have an increased family history of autoimmunity, and, among children with a first episode and short duration of AA (< 6 months), thyroid screening might be restricted in those with a positive family history of thyroid disorder. Thyroid screening should be routinely performed in all children with long-standing AA.


Direct and Indirect Links between Peer Factors and Adolescent Adjustment Difficulties.
The purpose of the current investigation was to examine the role of emotion regulation in the link between peer factors and adolescent adjustment difficulties. The sample consisted of 206 adolescents (ages 10-18 years) and parents. Peer factors (i.e., peer antisocial behavior, peer co-rumination, peer emotion regulation) and youth depressive symptoms were based on youth reports. Youth emotion regulation and antisocial behavior were assessed using parent and youth ratings. Results showed that peer antisocial behavior was directly (but not indirectly) related to youth antisocial behavior and depressive symptoms, whereas peer emotion regulation was indirectly (but not directly) related to both adolescent outcomes. In addition, peer co-rumination was indirectly related to youth antisocial behavior and directly and indirectly related to youth depressive symptoms. In general, the results indicated little evidence of moderation by adolescent age, sex, or ethnic differences. Implications for peer relationships as socialization contexts are discussed.


Osteosarcoma tissues and cell lines from patients with differing serum alkaline phosphatase concentrations display minimal differences in gene expression patterns.
Serum alkaline phosphatase (ALP) concentration is a prognostic factor for osteosarcoma in multiple studies, although its biological significance remains incompletely understood. To determine whether gene expression patterns differed in osteosarcoma from patients with differing serum ALP concentrations, microarray analysis was performed on 18 primary osteosarcoma samples and six osteosarcoma cell lines from dogs with normal and increased serum ALP concentration. No differences in gene expression patterns were noted between tumours or cell lines with differing serum ALP concentration using a gene-specific two-sample t-test. Using a more sensitive empirical Bayes procedure, defective in cullin neddylation 1 domain containing 1 (DCUN1D1) was increased in both the tissue and cell lines of the normal ALP group. Using quantitative PCR (qPCR), differences in DCUN1D1 expression between the two groups failed to reach significance. The homogeneity of gene expression patterns of osteosarcoma associated differing serum ALP concentrations are consistent with previous studies suggesting serum ALP concentration is not associated with intrinsic differences of osteosarcoma cells.


Occultifur tropicalis f.a., sp. nov., a novel cystobasidiomycetous yeast species isolated from tropical regions.
Five strains representing a single novel anamorphic yeast species were isolated from sugar cane. Two strains were from tissue (DMKU-SE38, DMKU-SE59(T)) and two from the external surface (DMKU-SP385, DMKU-SP403) of leaves collected in Thailand and the fifth (IMUFRJ 52020) from the rhizoplane of sugar cane in an organically cultivated field in Brazil. On the basis of sequence analysis of the D1/D2 region of the large subunit (LSU) rRNA gene and the internal transcribed spacer (ITS) region, they were classified as representing a single species of the genus Occultifur. The sequences of the D1/D2 region of the LSU rRNA genes and the ITS regions of the five strains were either identical or differed from each other by only one nucleotide substitution. The novel species was related most closely to Occultifur brasiliensis f.a. CBS 12687(T) but with 0.7-1.0% nucleotide substitutions (4-6 nt) in the D1/D2 region of the LSU rRNA gene and 2.5-2.7% nucleotide substitutions (14-15 nt) in the ITS region. The name Occultifur tropicalis f.a., sp. nov. is proposed. The type strain is DMKU-SE59(T) ( =BCC 61184(T) =NBRC 109696(T) =CBS 13389 (T)).


Reliability of administrative data for the identification of Parkinson's disease cohorts.
Parkinson's disease (PD) is a major worldwide public health problem with a prevalence that is expected to increase dramatically in the coming decades. Because administrative data are useful for epidemiologic and health service studies, we aimed to define procedural algorithms to identify PD patients (on a regional basis) using these data. We built two a priori algorithms, respecting privacy laws, with increasing theoretical specificity for PD including: (1) a hospital discharge diagnosis of PD; (2) PD-specific exemption; (3) a minimum of two separate prescriptions of an antiparkinsonian drug. The two algorithms differed for drugs included. Sensitivities were tested on an opportunistic sample of 319 PD patients from the databases of 5 regional movement disorders clinics. The estimated prevalence of PD in the sample population from Tuscany was 0.49 % for algorithm 1 and 0.28 % for algorithm 2. Algorithm 1 correctly identified 291 PD patients (sensitivity 91.2 %), and algorithm 2 identified 242 PD patients (sensitivity 75.9 %). We developed two reproducible algorithms demonstrating increasing theoretical specificity with good sensitivity in identifying PD patients based on an evaluation of administrative data. This may represent a low-cost strategy to reliably follow up a large number of PD patients as a whole for evaluating the effects of therapies, disease progression and prevalence.


Combining mathematics and empirical data to predict emergence of RNA viruses that differ in reservoir use.
RNA viruses may be particularly capable of contributing to the increasing biomedical problem of infectious disease emergence. Empirical studies and epidemiological models are informative for the understanding of evolutionary processes that promote pathogen emergence, but rarely are these approaches combined in the same study. Here, we used an epidemiology model containing observations of pathogen productivity in reservoirs, as a means to predict which pathogens should be most prone to emerge in a primary host such as humans. We employed as a model system a collection of vesicular stomatitis virus populations that had previously diverged in host-use strategy: specialists, directly selected generalists and indirectly selected (fortuitous) generalists. Using data from experiments where these viral strategists were challenged to grow on unencountered novel hosts in vitro, logistic growth models determined that the directly selected generalist viruses tended to grow best on model reservoirs. Furthermore, when we used the growth data to estimate average reproductive rate across secondary reservoirs, we showed that the combined approach could be used to estimate relative success of the differing virus strategists when encountering a primary host. Our study suggests that synergistic approaches combining epidemiological modelling with empirical data from experimental evolution may be useful for developing efforts to predict which types of pathogens pose the greatest probability of emerging in the future.


Multiplex amplification of all coding sequences within 10 cancer genes by Gene-Collector.
Herein we present Gene-Collector, a method for multiplex amplification of nucleic acids. The procedure has been employed to successfully amplify the coding sequence of 10 human cancer genes in one assay with uniform abundance of the final products. Amplification is initiated by a multiplex PCR in this case with 170 primer pairs. Each PCR product is then specifically circularized by ligation on a Collector probe capable of juxtapositioning only the perfectly matched cognate primer pairs. Any amplification artifacts typically associated with multiplex PCR derived from the use of many primer pairs such as false amplicons, primer-dimers etc. are not circularized and degraded by exonuclease treatment. Circular DNA molecules are then further enriched by randomly primed rolling circle replication. Amplification was successful for 90% of the targeted amplicons as seen by hybridization to a custom resequencing DNA micro-array. Real-time quantitative PCR revealed that 96% of the amplification products were all within 4-fold of the average abundance. Gene-Collector has utility for numerous applications such as high throughput resequencing, SNP analyses, and pathogen detection.


Stainless steel crown and pulpotomy: procedure and technique for primary molars.
The purpose of this article is to facilitate performance of the stainless steel crown and pulpotomy procedures in order to save chair time and provide a more durable and reliable final restoration for primary molars. The rationale and methods for preparing the tooth, crown selection, and placement of a stainless steel crown are included, along with alternatives for selection and technique for fabrication of more esthetic options. Medicament and tips for the pulpotomy procedure also are discussed. Information for patient education and consent regarding the procedures and their limitations is provided to complete this step-by-step guide for the clinician.


Wise Investment? Modeling Industry Profitability and Risk of Targeted Chemotherapy for Incurable Solid Cancers.
Pharmaceutical development involves substantial financial risk. This risk, rising development costs, and the promotion of continued research and development have been cited as major drivers in the progressive increase in drug prices. Currently, cost-effective analyses are being used to determine the value of treatment. However, cost-effective analyses practically function as a threshold for value and do not directly address the rationale for drug prices. We set out to create a functional model for industry price decisions and clarify the minimum acceptable profitability of new drugs. Assuming that industry should only invest in profitable ventures, we employed a linear cost-volume-profit breakeven analysis to equate initial capital investment and risk and post-drug-approval profits, where drug development represents the bulk of investment. A Markov decision analysis model was also used to define the relationships between investment events risk. A systematic literature search was performed to determine event probabilities, clinical trial costs, and total expenses as inputs into the model. Disease-specific inputs, current market size across regions, and lengths of treatment for cancer types were also included. With development of single novel chemotherapies costing from $802 to $1,042 million (2002 US dollars), pharmaceutical profits should range from $4.3 to $5.2 billion, with an expected rate of return on investment of 11% annually. However, diversification across cancer types for chemotherapy can reduce the minimum required profit to less than $3 billion. For optimal diversification, industry should study four tumor types per drug; however, nonprofit organizations could tolerate eight parallel development tracks to minimize the risk of development failure. Assuming that pharmaceutical companies hold exclusive rights for drug sales for only 5 years after market approval, the minimum required profit per drug per month per patient ranges from $294 for end-stage lung cancer to $3,231 for end-stage renal cell carcinoma. Pharmaceutical development in oncology is costly, with substantial risk, but is also highly profitable. Minimum acceptable profits per drug per month of treatment per patient vary with prevalence of disease, but they should be less than $5,000 per month of treatment in the developed world. Minimum acceptable profits may be lower for treatments with additional efficacy in the earlier stages of a tumor type. However, this type of event could not be statistically modeled.


Culturally sensitive substance abuse intervention for Hispanic and African American adolescents: empirical examples from the Alcohol Treatment Targeting Adolescents in Need (ATTAIN) Project.
This study presents preliminary analyses examining the effects of an alcohol and other drug use (AOD) intervention with minority juvenile offenders. Furthermore, the study investigates the impact of cultural factors on baseline AOD use among Hispanic and African American youth, as well as on treatment outcome. Participants were 213 juvenile offenders referred for treatment (mean age = 15.7 years), 97 of whom have completed treatment to date. The intervention was carried out in clinics placed within the neighborhoods in which the participants resided. Intervention Alcohol Treatment Targeting Adolescents in Need (ATTAIN) is a controlled clinical trial evaluating the effectiveness of a brief motivational, cognitive behavioral intervention, guided self-change (GSC). Participants are assigned randomly to the individual format of guided self-change (I-GSC), the family involved format of guided self-Change (F-GSC), choice of one of these two, or a waiting list control condition. Only participants involved in active intervention are included in the present report. Data were collected via structured face-to-face interviews. Alcohol and marijuana use measures were collected using the Time-line Follow-back interview (TLFB). There were significant reductions in alcohol and marijuana use for all ethnic groups from baseline to post-intervention. Cultural factors (discrimination, acculturation, ethnic pride and cultural mistrust) were associated with pre-intervention levels of alcohol and marijuana use. Among Hispanics, pre-intervention level of substance use were higher among foreign-born than US-born youth. Analyses conducted with the US-born Hispanic group showed that ethnic orientation and ethnic pride were associated positively with greater reductions in alcohol use. The intervention provided through ATTAIN appears to be effective with a multi-ethnic population of juvenile delinquents. Cultural factors, such as ethnic orientation and ethnic mistrust, appear to constitute amenability to treatment factors, with US-born Hispanic youth lower in acculturation responding better to the intervention.


Management of penoscrotal extramammary Paget disease: case series and review of the literature.
Extramammary Paget disease (empd) is a rare, slow-growing neoplasm, considered to be an adenocarcinoma of the apocrine glands. In men, the penoscrotal region is the most commonly affected area. The disease can present as carcinoma in situ or as invasive disease that can subsequently metastasize to lymph nodes and distant sites. Because of the rarity of empd, the medical literature available to guide management of the disease is limited, particularly in patients with metastases. In addition, metastatic disease may pose a diagnostic challenge, because invasive cancer of the genitourinary or gastrointestinal tract can occur in association with empd. In the present case series, we describe our experience in treating penoscrotal empd with multimodality therapy, and we review the existing literature concerning its diagnosis and management.


Multi-agent simulation regulated by microbe-oriented thermodynamics and kinetics equations for exploiting interspecies dynamics and evolution between methanogenesis, sulfidogenesis, hydrogenesis and exoelectrogenesis.
Multi-agent simulation (MAS) regulated by microbe-oriented thermodynamics and kinetics equations were performed for exploiting the interspecies dynamics and evolution in anaerobic respiration and bioelectrochemical systems. A newly-defined kinetically thermodynamic parameter is recognized microbes as agents in various conditions, including electron donors and acceptors, temperature, pH, etc. For verification of the MAS, the treatment of synthetic wastewater containing glucose and acetate was evaluated in four 25°C laboratory-scale reactors with different electron acceptors and cathode materials that had potential for methanogenesis, hydrogenesis, sulfidogenesis and exoelectrogenesis. Within 1000 h operation, the reactors performance and microbial structures using 16S rRNA sequencing matched with the MAS, suggesting acetoclastic exoelectrogenesis predominance (Geobacter). After 2400 h, MAS observed the co-existence of acetoclastic methanogenesis and acetoclastic and propionate exoelectrogenesis, as was reported previously. Such microbial evolution from the short-term to long-term operation likely resulted from the glucose-driven propionate. The MAS developed is applicable in a wide range of complex engineering and natural ecosystems.


Arabidopsis CRL4 Complexes: Surveying Chromatin States and Gene Expression.
CULLIN4 (CUL4) RING ligase (CRL4) complexes contain a CUL4 scaffold protein, associated to RBX1 and to DDB1 proteins and have traditionally been associated to protein degradation events. Through DDB1, these complexes can associate with numerous DCAF proteins, which directly interact with specific targets promoting their ubiquitination and subsequent degradation by the proteasome. A characteristic feature of the majority of DCAF proteins that associate with DDB1 is the presence of the DWD motif. DWD-containing proteins sum up to 85 in the plant model species Arabidopsis. In the last decade, numerous Arabidopsis DWD proteins have been studied and their molecular functions uncovered. Independently of whether their association with CRL4 has been confirmed or not, DWD proteins are often found as components of additional multimeric protein complexes that play key roles in essential nuclear events. For most of them, the significance of their complex partnership is still unexplored. Here, we summarize recent findings involving both confirmed and putative CRL4-associated DCAF proteins in regulating nuclei architecture remodelling, DNA damage repair, histone post-translational modification, mRNA processing and export, and ribosome biogenesis, that definitely have an impact in gene expression and de novo protein synthesis. We hypothesized that, by maintaining accurate levels of regulatory proteins through targeted degradation and transcriptional control, CRL4 complexes help to surveil nuclear processes essential for plant development and survival.


Accurate multiple network alignment through context-sensitive random walk.
Comparative network analysis can provide an effective means of analyzing large-scale biological networks and gaining novel insights into their structure and organization. Global network alignment aims to predict the best overall mapping between a given set of biological networks, thereby identifying important similarities as well as differences among the networks. It has been shown that network alignment methods can be used to detect pathways or network modules that are conserved across different networks. Until now, a number of network alignment algorithms have been proposed based on different formulations and approaches, many of them focusing on pairwise alignment. In this work, we propose a novel multiple network alignment algorithm based on a context-sensitive random walk model. The random walker employed in the proposed algorithm switches between two different modes, namely, an individual walk on a single network and a simultaneous walk on two networks. The switching decision is made in a context-sensitive manner by examining the current neighborhood, which is effective for quantitatively estimating the degree of correspondence between nodes that belong to different networks, in a manner that sensibly integrates node similarity and topological similarity. The resulting node correspondence scores are then used to predict the maximum expected accuracy (MEA) alignment of the given networks. Performance evaluation based on synthetic networks as well as real protein-protein interaction networks shows that the proposed algorithm can construct more accurate multiple network alignments compared to other leading methods.


Feasibility of Leadless Cardiac Pacing Using Injectable Magnetic Microparticles.
A noninvasive, effective approach for immediate and painless heart pacing would have invaluable implications in several clinical scenarios. Here we present a novel strategy that utilizes the well-known mechano-electric feedback of the heart to evoke cardiac pacing, while relying on magnetic microparticles as leadless mechanical stimulators. We demonstrate that after localizing intravenously-injected magnetic microparticles in the right ventricular cavity using an external electromagnet, the application of magnetic pulses generates mechanical stimulation that provokes ventricular overdrive pacing in the rat heart. This temporary pacing consistently managed to revert drug-induced bradycardia, but could only last up to several seconds in the rat model, most likely due to escape of the particles between the applied pulses using our current experimental setting. In a pig model with open chest, MEF-based pacing was induced by banging magnetic particles and has lasted for a longer time. Due to overheating of the electromagnet, we intentionally terminated the experiments after 2 min. Our results demonstrate for the first time the feasibility of external leadless temporary pacing, using injectable magnetic microparticles that are manipulated by an external electromagnet. This new approach can have important utilities in clinical settings in which immediate and painless control of cardiac rhythm is required.


Resection strategies in tumoral epilepsy: is a lesionectomy enough?
Resection strategies in patients with tumor-related epilepsy vary from lesionectomy to larger epilepsy operations with no consensus on optimal approaches. The objective of this study is to use our prior experience in the management of these patients, to derive optimal strategies for the surgical management of epilepsy related to brain tumors. A prospectively compiled database of epilepsy and tumor patients was used to identify patients who underwent surgical resection of a neoplasm but then developed epilepsy, or who presented with epilepsy and were found to harbor a brain tumor. Seizure frequency, histopathology, type of surgical resection, and outcomes were compiled. Of 235 epilepsy surgery patients and 75 low/intermediate grade glioma surgery patients, 13 (5.5%) and 21 (28%) patients, respectively, had tumoral epilepsy. Twenty-two patients were male and 18 tumors were in the left hemisphere. Tumoral epilepsy occurred predominantly in temporal (50%) and perirolandic (26.5%) locations. The etiology was WHO grade I tumors in 29%, grade II in 35%, and grade III in 33%. In the epilepsy group, following lesionectomy in three and tailored resections in the majority, seizure outcomes were Engel class I in all except one case. In the tumor group, after the initial operation seven additional resections were performed due to seizure recurrence. Outcomes in this group were Engel class 1A in 18 patients and 1B, 1C and IIA in 1 patient each. Drawing upon these data, we propose a classification of the likely reasons of failure in seizure control in patients with tumoral epilepsy. This review reiterates the concept that a complete resection of the lesion is the best approach for dealing with tumors presenting with epilepsy. Overall excellent outcomes can be accomplished following aggressive initial tumor resection, re-resection in the context of recurrence, and epilepsy style operations in selected patients with a longer history of seizures.


Inhalation anesthetics augment oxidant-induced pulmonary vasoconstriction: evidence for a membrane effect.
Inhalational anesthetics "fluidize" biologic membranes. Since arachidonate metabolism also occurs in cell membranes, anesthetic agents may modify arachidonic acid mediator production. The authors used the isolated perfused rabbit lung preparation to examine the effects of inhalational anesthetics on the production of arachidonate mediators. The oxidant tert-butyl-hydroperoxide (t-bu-OOH) is known to cause pulmonary vasoconstriction by causing increased production of thromboxane A2 (TxA2). The authors administered three anesthetics (halothane, cyclopropane, and nitrous oxide) of widely different potencies, at different dosages, each to three different groups of preparations and challenged the lungs at each anesthetic dose with t-bu-OOH. They found a dose-related augmentation of the pulmonary vasopressor response to t-bu-OOH. Preparations given t-bu-OOH alone showed no change in response over time. Lungs perfused with indomethacin (5 micrograms . ml-1 in Krebs-Henseleit buffer), ventilated with cyclopropane (2 MAC), and challenged with t-bu-OOH showed almost complete inhibition of the response to t-bu-OOH. Indomethacin at this concentration is a specific inhibitor of cyclooxygenase. The authors also have demonstrated significantly increased perfusate levels of thromboxane B2 (TxB2), the inactive metabolite of TxA2, after oxidant challenge during exposure to 2% halothane compared with TxB2 levels before halothane exposure. The authors believe that the augmented pressor response and mediator production occur because of increased substrate (arachidonic acid) availability induced by anesthetic agent.


Inflammatory bowel disease is associated with an increased risk of hospitalization for heart failure: a Danish Nationwide Cohort study.
Inflammatory bowel disease (IBD) has been linked to adverse cardiovascular events, but a relation to heart failure (HF) is uncertain. We investigated the IBD-associated risk of HF in a nationwide setting. A total of 5 436 647 Danish citizens, with no history of IBD or HF, were included on January 1, 1997, and followed up until first hospitalization for HF, death, or December 31, 2011. Of these subjects, 23 681 developed IBD for which disease activity was determined continuously throughout the study. The risk of hospitalization for HF was estimated with a Poisson regression model adjusting for comorbidity and cardiovascular pharmacotherapy as time-dependent covariates. During a mean follow-up of 11.8 years in the reference population and 6.4 years in the IBD group, hospitalization for HF occurred in 553 subjects with IBD and 171 405 in the reference population. Patients with IBD had a 37% increased risk of hospitalization for HF (incidence rate ratio, 1.37; 95% confidence interval, 1.26-1.49) compared with the reference population. IBD activity-specific analyses showed markedly increased risk of HF hospitalization during flares (incidence rate ratio, 2.54; 95% confidence interval, 2.13-3.04) and persistent activity (incidence rate ratio, 2.73; 95% confidence interval, 2.25-3.33) but not in IBD remission (incidence rate ratio, 1.04; 95% confidence interval, 0.94-1.16). In a nationwide cohort, IBD was associated with an increased risk of hospitalization for HF, and this risk was strongly correlated to periods of active disease. The mechanisms underlying this finding warrant further studies.


Analysis of safety, risk factors and pretreatment methods during rush hymenoptera venom immunotherapy.
The safety profile of venom immunotherapy is a relevant issue. We evaluated the frequency of severe adverse events (SAE), associated risk factors, retrospective comparison of pretreatment protocols including solely H1 receptor blockers and a combination of H1 and H2 receptor blockers during rush Hymenoptera venom immunotherapy. The study group comprised 118 patients. The treatment was initiated according to a 5-day rush protocol with the use of standardized venom allergens of either wasp or honeybee. During the rush induction, side effects occurred in 18 patients (15.2%), whereas SAE were present in 7 patients (5.9%). Twelve out of 18 (66.6%) developed anaphylactic reactions on the fourth day of the rush protocol, with the majority of cases at a dose of 40 or 60 microg of the venom extract (p = 0.001). The frequency of SAE was also significantly higher on the fourth day than thereafter (p = 0.0001) as well as in patients allergic to bee venom (p = 0.049). All systemic side effects were more frequent in women (p = 0.0065). However, this relation was not true when SAE were consider (p = 0.11). A higher percentage of SAE was observed in the subjects pretreated with both H1 and H2 receptor antagonists than in those pretreated with H1 blocker only (8.8 vs. 4.1%); however, the difference was not significant. Considerable severity of allergic adverse events requires particular attention to patients allergic to bee venom and during rush phase, especially when rapidly increasing doses are administered. Pretreatment with H2 blockers is debatable and warrants further investigation.


Glucose Signaling Is Connected to Chromosome Segregation Through Protein Kinase A Phosphorylation of the Dam1 Kinetochore Subunit in Saccharomycescerevisiae.
The Dam1 complex is an essential component of the outer kinetochore that mediates attachments between spindle microtubules and chromosomes. Dam1p, a subunit of the Dam1 complex, binds to microtubules and is regulated by Aurora B/Ipl1p phosphorylation. We find that overexpression of cAMP-dependent protein kinase (PKA) catalytic subunits (i.e., TPK1, TPK2, TPK3) is lethal in DAM1 mutants and increases the rate of chromosome loss in wild-type cells. Replacing an evolutionarily conserved PKA site (S31) in Dam1p with a nonphosphorylatable alanine suppressed the high-copy PKA dosage lethality in dam1-1 Consistent with Dam1p as a target of PKA, we find that in vitro PKA can directly phosphorylate S31 in Dam1p and we observed phosphorylation of S31 in Dam1p purified from asynchronously growing yeast cells. Cells carrying high-copy TPK2 or a Dam1p phospho-mimetic S31D mutant displayed a reduction in Dam1p localization at the kinetochore, suggesting that PKA phosphorylation plays a role in assembly and/or stability of the Dam1 complex. Furthermore, we observed spindle defects associated with S31 phosphorylation. Finally, we find that phosphorylation of Dam1p on S31 is reduced when glucose is limiting as well as during α-factor arrest, conditions that inhibit PKA activity. These observations suggest that the PKA site of Dam1p participates in regulating kinetochore activity. While PKA is a well-established effector of glucose signaling, our work shows for the first time that glucose-dependent PKA activity has an important function in chromosome segregation.


Molecular cloning and genetic analysis of the suppressor-of-white-apricot locus from Drosophila melanogaster.
We report genetic and molecular analysis of the suppressor-of-white-apricot [su(wa)] locus, one of several retrotransposon insertion allele-specific suppressor loci in Drosophila melanogaster. First, we isolated and characterized eight new mutations allelic to the original su(wa)1 mutation. These studies demonstrated that su(wa) mutations allelic to su(wa)1 affected a conventional D. melanogaster complementation group. Second, we cloned the chromosomal region containing the su(wa) complementation group by P element transposon tagging. The ca. 14-kilobase region surrounding the su(wa) complementation group contained five distinct transcription units, each with a different developmentally programmed pattern of expression. Third, we used a modified procedure for P-mediated gene transfer to identify the transcription unit corresponding to su(wa) by gene transfer. Fourth, we found that the presumptive su(wa) transcription unit produced a family of transcripts (ranging from ca. 3.5 to ca. 5.2 kilobases) in all developmental stages, tissue fractions, and cell lines we examined, suggesting that the gene is universally expressed.


Epidemiology of Work-Related Neck Pain Among Spine Surgeons.
Cross-sectional survey. This study aimed to investigate work-related neck pain among AO spine surgeons in different regions by estimating its prevalence, predictors, consequences, and management methods. A cross-sectional survey of 411 spine surgeon members of AO spine was conducted during March-May 2021, using the Modified Nordic Questionnaire and the Neck Disability Index. Data on neck pain experience during the last 12 months and its consequences and risk factors were collected. Logistic regression analysis was done to identify significant predictors of neck pain. Significance was set at P < .05. The 1-year neck pain was experienced by 66.7% of surgeons. According to the Neck Disability Index, more than one-half (52.8%) experienced disability due to neck pain of mild (45.5%), moderate (6.5%), and severe (.8%) grades. Neck pain was responsible for stopping work in 17.5% of surgeons, with a median of 3.5 (IQR, 2-7.8) days off work. One-half of the participants (56.3%) were treated by medical care, 31.5% by physiotherapy, and 16.5% requested rest days and sick leave. Physical stress (P < .001) and non-exercising (P = .04) were the significant predictors of neck pain. The 12-month prevalence of neck pain was high among spine surgeons, with an impact on activities of daily living, mainly of a mild degree, reported by one-half of surgeons. Physical stress was the only significant predictor, while sports practice was a protective factor against neck pain. Medication was the primary management adopted-an increased focus on pain prevention through improved workplace ergonomics and sports activity programs is recommended.


[Rearing of female cattle twins using complete feed pellets in comparison to straw pellets plus concentrate mixture. 1. Weight gain performance and feed expenditure as well as rumen fermentation conditions during the 4th-12th months of life].
The investigations with young female cattle (twin cattle, n = 30) the applicability of straw as sole roughage after the end of the suckling period was tested. Completely (I) and partly (II) pelleted rations of dried feed with a straw component of 65% each in the total rations were used. Feed intake, development of body weight, feed expenditure as well as indexes of rumen fermentation and of the concentration of some metabolism parameters in the blood serum were measured. In the complete test period (between 5th and 12th months of life, 243 days) there were not any significant differences between the groups in all registrations. Average daily weight gains of 570 g were achieved with an overall high dry matter (2.6 kg/100 kg live weight) and roughage (straw) intake (1.6 kg dry matter/100 kg live weight). The standard expenditure of energy and protein given for the stage of development was differentiatedly undercut. The energy expenditure being below the standard (ca. 10%), the total energy expenditure (kEFUcattle) was by 20 (II) and 25% (I) below standard expenditure. Energy expenditure aimed at weight gain reached very satisfactory values with 3.68 (I) and 3.79 kEFUcattle/kg weight gain (II). Protein expenditure fell short of the standard for the development stage by 14 (II) and 20% (I) resp. All parameters ascertained of the rumen and intermediate metabolism were in the physiologic standard range, as is characteristic of feeding regimes with a relatively high quota of roughage.


Replication interference between human papillomavirus types 16 and 18 mediated by heterologous E1 helicases.
Co-infection of multiple genotypes of human papillomavirus (HPV) is commonly observed among women with abnormal cervical cytology, but how different HPVs interact with each other in the same cell is not clearly understood. A previous study using cultured keratinocytes revealed that genome replication of one HPV type is inhibited by co-existence of the genome of another HPV type, suggesting that replication interference occurs between different HPV types when co-infected; however, molecular mechanisms underlying inter-type replication interference have not been fully explored. Replication interference between two most prevalent HPV types, HPV16 and HPV18, was examined in HPV-negative C33A cervical carcinoma cells co-transfected with genomes of HPV16 and HPV18 together with expression plasmids for E1/E2 of both types. Levels of HPV16/18 genome replication were measured by quantitative real-time PCR. Physical interaction between HPV16/18 E1s was assessed by co-immunoprecipitation assays in the cell lysates. The replication of HPV16 and HPV18 genomes was suppressed by co-expression of E1/E2 of heterologous types. The interference was mediated by the heterologous E1, but not E2. The oligomerization domain of HPV16 E1 was essential for HPV18 replication inhibition, whereas the helicase domain was dispensable. HPV16 E1 co-precipitated with HPV18 E1 in the cell lysates, and an HPV16 E1 mutant Y379A, which bound to HPV18 E1 less efficiently, failed to inhibit HPV18 replication. Co-infection of a single cell with both HPV16 and HPV18 results in replication interference between them, and physical interaction between the heterologous E1s is responsible for the interference. Heterooligomers composed of HPV16/18 E1s may lack the ability to support HPV genome replication.


Molecular mechanisms and pathways involved in bovine embryonic genome activation and their regulation by alternative in vivo and in vitro culture conditions.
Understanding gene expression patterns in response to altered environmental conditions at different time points of the preimplantation period would improve our knowledge on regulation of embryonic development. Here we aimed to examine the effect of alternative in vivo and in vitro culture conditions at the time of major embryonic genome activation (EGA) on the development and transcriptome profile of bovine blastocysts. Four different blastocyst groups were produced under alternative in vivo and in vitro culture conditions before or after major EGA. Completely in vitro- and in vivo-produced blastocysts were used as controls. We compared gene expression patterns between each blastocyst group and in vivo blastocyst control group using EmbryoGENE's bovine microarray. The data showed that changing culture conditions from in vivo to in vitro or vice versa, either before or after the time of major EGA, had no effect on the developmental rates; however, in vitro conditions during that time critically influenced the transcriptome of the blastocysts produced. The source of oocyte had a critical effect on developmental rates and the ability of the embryo to react to changing culture conditions. Ontological classification highlighted a marked contrast in expression patterns for lipid metabolism and oxidative stress response between blastocysts generated in vivo versus in vitro, with opposite trends. Molecular mechanisms and pathways that are influenced by altered culture conditions during EGA were defined. These results will help in the development of new strategies to modify culture conditions at this critical stage to enhance the development of competent blastocysts.


Fatty Oil of Descurainia Sophia Nanoparticles Improve Monocrotaline-Induced Pulmonary Hypertension in Rats Through PLC/IP3R/Ca2+ Signaling Pathway.
Fatty oil of Descurainia Sophia (OIL) has poor stability and low solubility, which limits its pharmacological effects. We hypothesized that fatty oil nanoparticles (OIL-NPs) could overcome this limitation. The protective effect of OIL-NPs against monocrotaline-induced lung injury in rats was studied. We prepared OIL-NPs by wrapping fatty oil with polylactic-polyglycolide nanoparticles (PLGA-NPs) and conducted in vivo and in vitro experiments to explore its anti-pulmonary hypertension (PH) effect. In vitro, we induced malignant proliferation of pulmonary artery smooth muscle cells (RPASMC) using anoxic chambers, and studied the effects of OIL-NPs on the malignant proliferation of RPASMC cells and phospholipase C (PLC)/inositol triphosphate receptor (IP3R)/Ca2+ signal pathways. In vivo, we used small animal echocardiography, flow cytometry, immunohistochemistry, western blotting (WB), polymerase chain reaction (PCR) and metabolomics to explore the effects of OIL-NPs on the heart and lung pathological damage and PLC/IP3R/Ca2+ signal pathway of pulmonary hypertension rats. We prepared fatty into OIL-NPs. In vitro, OIL-NPs could improve the mitochondrial function and inhibit the malignant proliferation of RPASMC cells by inhibiting the PLC/IP3R/Ca2+signal pathway. In vivo, OIL-NPs could reduce the pulmonary artery pressure of rats and alleviate the pathological injury and inflammatory reaction of heart and lung by inhibiting the PLC/IP3R/Ca2+ signal pathway. OIL-NPs have anti-pulmonary hypertension effect, and the mechanism may be related to the inhibition of PLC/IP3R/Ca2+signal pathway.


Fecal Impaction With Multisystem Organ Involvement.
Fecal impactions are a common complaint in the emergency department (ED) population. The potential for significant derangement in physiologic processes of other organ systems is often underappreciated. A 19-year-old male, previously healthy, presented to the ED at our institution with complaint of abdominal pain, which was found to be secondary to severe fecal impaction. In the search for alternative diagnoses, imaging was performed, which revealed effects on multiple other organ systems. This case illustrates the secondary effects of a severe fecal impaction. The emergency physician must be aware of these consequences, as the opportunity to review labs and imaging is not often provided in the standard workup of these patients.


Efficacy of Mouth Rinses and Nasal Spray in the Inactivation of SARS-CoV-2: A Systematic Review and Meta-Analysis of In Vitro and In Vivo Studies.
Severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) is a global and evolving pandemic associated with heavy health and financial burdens. Considering the oral cavity as the major reservoir for SARS-CoV-2, a systematic review and meta-analysis were conducted to assess the efficacy of mouth rinses and nasal sprays in reducing the salivary viral load of SARS-CoV-2. All in vivo and in vitro studies that assessed the virucidal efficacy of mouth rinses and nasal sprays against SARS-CoV-2 and were published in the English language from December 2019 to April 2022 were considered for analyses. Special Medical Subject Headings terms were used to search Pubmed, Scopus, Embase Ovid, and Web of Science databases. The toxicological data reliability assessment tool (ToxRToool) was used to assess the quality of the included studies. Thirty-three studies (11 in vivo and 22 in vitro) were deemed eligible for inclusion in this analysis. Results of the pooled data showed that povidone-iodine is the most efficacious intervention in vivo in terms of reducing the SARS-CoV-2 salivary viral load, followed by chlorhexidine. The mean difference in the viral load was 86% and 72%, respectively. Similarly, povidone-iodine was associated with the highest log10 reduction value (LRV) in vitro, followed by cetylpyridinium chloride, (LRV = 2.938 (p < 0.0005) and LRV = 2.907 (p = 0.009), respectively). Povidone-iodine-based oral and nasal preparations showed favourable results in terms of reducing SARS-CoV-2 viral loads both in vivo and in vitro. Considering the limited number of patients in vivo, further studies among larger cohorts are recommended.


Ovarian ossification associated with endometriosis.
Bone formation in the ovary, with the exception of developing in the setting of mature cystic teratoma, is exceedingly rare. A 46-year-old woman with a history of endometriosis and chronic pelvic pain underwent total abdominal hysterectomy and bilateral salpingo-oophorectomy. A 3 cm solid heavily calcified mass with a stony hard consistency was detected within the right ovary. Microscopic examination revealed extensive calcification of the right ovarian stroma with formation of abundant mature bone, adjacent to small foci of endometriosis. Endometriosis can be associated with ovarian ossification, forming an extensively calcified adnexal mass. Conservative treatment with close follow-up may be adequate in patients with a history of endometriosis who present with a small heavily calcified ovarian mass and wish to preserve their fertility.


[Study of the properties of E. coli of serological group 03 isolated in acute intestinal diseases].
The authors studied the properties of 115 strains of E. coli of serological group 03 isolated from 49 children and adults with acute intestinal disturbances. The majority of the children (82.9%) were aged under one year. Results of the study of the antigenic structure and biochemical properties permitted to differentiate the strains isolated into 3 serological types, with the prevalence of strains of type O3 : K2 (L) : H2 (78.3%), and 8 biochemical variants. The majority of the strains possessed hemolytic properties. Strains of serological group O3 were isolated repeatedly from the patients during the sickness, whereas none were revealed in examination of 132 healthy children and adults. The data obtained permitted to consider these microbes to be possible causative agents of intestinal coliinfection, and to refer them to the enteropathogenic category.


Assessment of a short hypnosis in a paediatric operating room in reducing postoperative pain and anxiety: A randomised study.
To assess the impact of a short hypnotic session on postoperative anxiety and pain in major orthopaedic surgery. Despite specific information given before a scheduled paediatric surgery, perioperative anxiety can become important. Randomised Clinical Study. The study is an open single-centre randomised clinical study comparing a "control" group versus a "hypnosis" group receiving a short hypnosis pre-induction session as additional experimental analgesic procedure. The primary endpoint was the postoperative anxiety, blindly assessed using a visual analogue scale. The study involved 120 children (age 10-18 years). The results showed no difference between control group versus hypnosis group. Twenty-four hours after surgery (Day+1), the patient's anxiety score was not different between control and hypnosis groups (median [Q1-Q3]: 1 [0; 3] vs. 0 [0; 3], respectively, p = .17). Each group experienced a significant decrease in anxiety level between the day before surgery (Day-1) and the day after surgery (Day+1) (median ([Q1-Q3]) difference of the anxiety score: 2 [4; 0] and 2 [4; 0], respectively, p < .0001 in each group). The postoperative pain scores were low and not different between groups (median [Q1-Q3]: 2 [0; 3] in control group vs. 3 [1; 3] in hypnosis group, p = .57). This randomised study on a short hypnosis session performed in the operating room prior to a major surgery showed no difference in postoperative anxiety and pain levels. The decrease in anxiety and pain levels may be due to the addition of nurse pre-operative interviews and optimisation in communication in the operating room. As postoperative anxiety level was low in both control and hypnosis groups, nurse pre-operative interviews and nurse training in hypnosis may contribute to the optimisation of global management and decrease the postoperative anxiety level.


Euphorbia characias as bioenergy crop: a study of variations in energy value components according to phenology and water status.
Euphorbia characias has drawn much attention as a potential bioenergy crop given its considerable amount of latex, rich in hydrocarbon-like compounds, and its ability to grow in large areas of semiarid lands. Compositions of major constituents with an energy value have been determined for the three phenological stages of this plant (preflowering, flowering, and postflowering) and different irrigation treatments. Metabolites from both nonpolar and polar extracts have been identified and quantified by GC-MS, GC-FID, HPLC-ELSD, and UPLC-PDA-MS. The results highlight that the end of the flowering period is the optimal harvesting time to maximize the yields of E. characias as a potential energy crop. The total water requirements to obtain the maximum yields of hexane- and methanol-extractables were determined for its annual development cycle.


Id2 promotes tumor cell migration and invasion through transcriptional repression of semaphorin 3F.
Id proteins (Id1 to Id4) are helix-loop-helix transcription factors that promote metastasis. It was found that Semaphorin 3F (SEMA3F), a potent inhibitor of metastasis, was repressed by Id2. High metastatic human tumor cell lines had relatively high amounts of Id2 and low SEMA3F levels compared with their low metastatic counterparts. No correlation between metastatic potential and expression of the other Id family members was observed. Furthermore, ectopic expression of Id2 in low metastatic tumor cells downregulated SEMA3F and, as a consequence, enhanced their ability to migrate and invade, two requisite steps of metastasis in vivo. Id2 overexpression was driven by the c-myc oncoprotein. SEMA3F was a direct target gene of the E47/Id2 pathway. Two E-box sites, which bind E protein transcription factors including E47, were identified in the promoter region of the SEMA3F gene. E47 directly activated SEMA3F promoter activity and expression and promoted SEMA3F biological activities, including filamentous actin depolymerization, inactivation of RhoA, and inhibition of cell migration. Silencing of SEMA3F inhibited the E47-induced SEMA3F expression and biological activities, confirming that these E47-induced effects were SEMA3F dependent. E47 did not induce expression of the other members of the SEMA3 family. Id2, a dominant-negative inhibitor of E proteins, abrogated the E47-induced SEMA3F expression and biological activities. Thus, high metastatic tumor cells overexpress c-myc, leading to upregulation of Id2 expression; the aberrantly elevated amount of Id2 represses SEMA3F expression and, as a consequence, enhances the ability of tumor cells to migrate and invade.


Foot volume increase on nifedipine is not prevented by pretreatment with diuretics.
Despite their natriuretic effects, dihydropyridine calcium-channel blockers (CCBs) often induce ankle oedema, probably due to vasodilation in the dependent legs. Since concomitant administration of frusemide does not prevent the acute increase in foot volume on nifedipine, we investigated whether diuretic pretreatment attenuates foot swelling on CCBs. In four separate experiments, 10 healthy volunteers received: (i) 20 mg of nifedipine without active pretreatment (pretreatment with placebo only); (ii) 20 mg of nifedipine after 5 days' treatment with amiloride 5 mg twice daily; (iii) 20 mg of nifedipine after 5 days' treatment with chlorthalidone 50 mg once daily; and (iv) no active drugs (pretreatment with placebo and placebo in place of nifedipine) as the control. Foot volumes were measured using an accurate water displacement technique (intra-individual coefficient of variance 0.27%). Amiloride and chlorthalidone pretreatment induced marked volume depletion, with a 2-3% reduction in body weight, a 5-10% increase in haematocrit and a 14-23% increase in plasma colloid osmotic pressure. In addition, the mean +/- SEM foot volume after both chlorthalidone (1282 +/- 37 ml) and amiloride (1289 +/- 40 ml) was lower than without pretreatment (1315 +/- 38 ml) (P < 0.05). Neither amiloride nor chlorthalidone significantly influenced the acute increase in foot volume on nifedipine. However, due to pretreatment effects, the foot volume after nifedipine was higher (P < 0.05) without pretreatment (1356 +/- 36 ml) than after amiloride (1318 +/- 38 ml) or chlorthalidone (1319 +/- 37 ml). Amiloride significantly attenuated the natriuretic effect of nifedipine, whereas chlorthalidone prevented the nifedipine-induced rise in colloid osmotic pressure and haematocrit. Diuretic pretreatment and the concomitant volume depletion did not prevent acute foot swelling on nifedipine, although the absolute foot volume remained lower after such pretreatment. Therefore diuretics mitigate the oedema of CCBs, but do not directly interfere with oedema formation.


Changes in the Practice of Coronary Revascularization between 2006 and 2010 in the Republic of Korea.
Evidence suggests that technological innovations and reimbursement schemes of the National Health Insurance Service may have impacted the management of coronary artery disease. Thus, we investigated changes in the practice patterns of coronary revascularization. Revascularization and in-hospital mortality among Koreans ≥20 years old were identified from medical claims filed between 2006 and 2010. The age- and sex-standardized procedure rate per 100,000 person-years was calculated directly from the distribution of the 2008 Korean population. The coronary revascularization rate increased from 116.1 (95% confidence interval, 114.9-117.2) in 2006 to 131.0 (129.9-132.1) in 2010. Compared to the rate ratios in 2006, the rate ratios for percutaneous coronary intervention (PCI) and coronary artery bypass graft (CABG) surgery in 2010 were 1.16 (1.15-1.17) and 0.80 (0.76-0.84), respectively. Among patients who received PCI, the percentage with drug-eluting stents increased from 89.1% in 2006 to 93.0% in 2010. In-hospital mortality rates from PCI significantly increased during the study period (p=0.03), whereas those from CABG significantly decreased (p=0.01). The in-hospital mortality rates for PCI and CABG were higher in elderly and female patients and at the lowest-volume hospitals. The annual volume of coronary revascularization continuously increased between 2006 and 2010 in Korea, although this trend differed according to procedure type. A high percentage of drug-eluting stent procedures and a high rate of in-hospital mortality at low-volume hospitals were noted.


[Anatomo-stereologic characteristics of the mucous membrane of the major duodenal papilla].
In 55 men and 60 women who had had not any pathology in their biliary-pancreato-duodenal system during life, microrelief of the mucous membrane in the major duodenal papilla has been studied by means of light and scanning electron microscopy. It is defined by variants of fusion of the common bile duct and the pancreatic duct in the papilla. Three main variants of the ducts fusion are defined and, respectively, three main varieties of the structure of the mucous membrane folds. When the ducts flow into the same orifice separately, the mucous membrane relief is even, parvigranular and there are no fold-valves. In the common major duodenal papilla, the architectonics of the mucous membrane resembles the valve that acts as a block-outlet mechanism.


Bradykinin-induced chemotaxis of human gliomas requires the activation of KCa3.1 and ClC-3.
Previous reports demonstrate that cell migration in the nervous system is associated with stereotypic changes in intracellular calcium concentration ([Ca(2+)](i)), yet the target of these changes are essentially unknown. We examined chemotactic migration/invasion of human gliomas to study how [Ca(2+)](i) regulates cellular movement and to identify downstream targets. Gliomas are primary brain cancers that spread exclusively within the brain, frequently migrating along blood vessels to which they are chemotactically attracted by bradykinin. Using simultaneous fura-2 Ca(2+) imaging and amphotericin B perforated patch-clamp electrophysiology, we find that bradykinin raises [Ca(2+)](i) and induces a biphasic voltage response. This voltage response is mediated by the coordinated activation of Ca(2+)-dependent, TRAM-34-sensitive K(Ca)3.1 channels, and Ca(2+)-dependent, 4,4'-diisothiocyanato-stilbene-2,2'-disulfonic acid (DIDS)-sensitive and gluconate-sensitive Cl(-) channels. A significant portion of these Cl(-) currents can be attributed to Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) activation of ClC-3, a voltage-gated Cl(-) channel/transporter, because pharmacological inhibition of CaMKII or shRNA-mediated knockdown of ClC-3 inhibited Ca(2+)-activated Cl(-) currents. Western blots show that K(Ca)3.1 and ClC-3 are expressed in tissue samples obtained from patients diagnosed with grade IV gliomas. Both K(Ca)3.1 and ClC-3 colocalize to the invading processes of glioma cells. Importantly, inhibition of either channel abrogates bradykinin-induced chemotaxis and reduces tumor expansion in mouse brain slices in situ. These channels should be further explored as future targets for anti-invasive drugs. Furthermore, these data elucidate a novel mechanism placing cation and anion channels downstream of ligand-mediated [Ca(2+)](i) increases, which likely play similar roles in other migratory cells in the nervous system.


Environmental determinants of extinction selectivity in the fossil record.
The causes of mass extinctions and the nature of biological selectivity during extinction events remain central questions in palaeobiology. Although many different environmental perturbations have been invoked as extinction mechanisms, it has long been recognized that fluctuations in sea level coincide with many episodes of biotic turnover. Recent work supports the hypothesis that changes in the areas of epicontinental seas have influenced the macroevolution of marine animals, but the extent to which differential environmental turnover has contributed to extinction selectivity remains unknown. Here I use a new compilation of the temporal durations of sedimentary rock packages to show that carbonate and terrigenous clastic marine shelf environments have different spatio-temporal dynamics and that these dynamics predict patterns of genus-level extinction, extinction selectivity and diversity among Sepkoski's Palaeozoic and modern evolutionary faunae. These results do not preclude a role for biological interactions or unusual physical events as drivers of macroevolution, but they do suggest that the turnover of marine shelf habitats and correlated environmental changes have been consistent determinants of extinction, extinction selectivity and the shifting composition of the marine biota during the Phanerozoic eon.


A case of homozygous familial hypercholesterolemia with focal segmental glomerulosclerosis.
Familial hypercholesterolemia (FH) is a common autosomal dominant inherited disorder characterized by increased levels of circulating plasma low-density lipoprotein cholesterol (LDL-C), tendon xanthomas, and premature atherosclerotic cardiovascular disease. Homozygous FH occurs in only one in a million people. Focal segmental glomerulosclerosis (FSGS) is clinically characterized by proteinuria, which is marked in the majority of cases and accompanied by nephrotic syndrome, high incidence of hypertension, and progression to renal failure. To our knowledge, we herein report for the first time a case of homozygous FH associated with FSGS. A seven-and-a-half-year-old boy was referred to our hospital due to cutaneous xanthomata and growth retardation. He had multiple nodular yellowish cutaneous xanthomatous lesions each 1 cm in size over his knees and sacral region. Laboratory data included cholesterol level of 1,050 mg/dl, low density lipoprotein cholesterol (LDL-C) 951 mg/dl, high-density lipoprotein cholesterol (HDL-C) 29 mg/dl, triglycerides 168 mg/dl, total protein 6.3 g/dl, and albumin 3.2 g/dl. Urinary protein excretion was 78 mg/m(2) per hour. A percutaneous renal biopsy was performed, and histological findings showed FSGS. Treatment with cholestyramine and atorvastatin was unsuccessful in terms of lowering lipids, and he was placed on weekly sessions of plasmapheresis. Total cholesterol was reduced from 1,050 mg/dl to 223 mg/dl, LDL-C from 951 mg/dl to 171 mg/dl, and urinary protein excretion from 78 mg/m(2) per hour to 42 mg/m(2) per hour after eight sessions of plasmapheresis. It is our belief that plasmapheresis is a treatment of choice in patients with FSGS associated with FH.


Diagnostic Value of Brain Calcifications in Adult-Onset Leukoencephalopathy with Axonal Spheroids and Pigmented Glia.
Adult-onset leukoencephalopathy with axonal spheroids and pigmented glia is a rare neurodegenerative disease resulting from mutations in the colony stimulating factor 1 receptor gene. Accurate diagnosis can be difficult because the associated clinical and MR imaging findings are nonspecific. We present 9 cases with intracranial calcifications distributed in 2 brain regions: the frontal white matter adjacent to the anterior horns of the lateral ventricles and the parietal subcortical white matter. Thin-section (1-mm) CT scans are particularly helpful in detection due to the small size of the calcifications. These calcifications had a symmetric "stepping stone appearance" in the frontal pericallosal regions, which was clearly visible on reconstructed sagittal CT images. Intrafamilial variability was seen in 2 of the families, and calcifications were seen at birth in a single individual. These characteristic calcification patterns may assist in making a correct diagnosis and may contribute to understanding of the pathogenesis of leukoencephalopathy.


Structural barriers to timely initiation of antiretroviral treatment in Vietnam: findings from six outpatient clinics.
In Vietnam, premature mortality due to AIDS-related conditions is commonly associated with late initiation to antiretroviral therapy (ART). This study explores reasons for late ART initiation among people living with HIV (PLHIV) from the perspectives of health care providers and PLHIV. The study was undertaken in six clinics from five provinces in Vietnam. Baseline CD4 counts were collected from patient records and grouped into three categories: very late initiators (≤100 cells/mm(3) CD4), late initiators (100-200 cells/mm(3)) and timely initiators (200-350 cells/mm(3)). Thirty in-depth interviews with patients who started ART and 15 focus group discussions with HIV service providers were conducted and thematic analysis of the content performed. Of 934 patients, 62% started ART very late and 11% initiated timely treatment. The proportion of patients for whom a CD4 count was obtained within six months of their HIV diagnosis ranged from 22% to 72%. The proportion of patients referred to ART clinics by voluntary testing and counselling centres ranged from 1% to 35%. Structural barriers to timely ART initiation were poor linkage between HIV testing and HIV care and treatment services, lack of patient confidentiality and a shortage of HIV/AIDS specialists. If Vietnam's treatment practice is to align with WHO recommendations then the connection between voluntary counselling and testing service and ART clinics must be improved. Expansion and decentralization of HIV/AIDS services to allow implementation at the community level increased task sharing between doctors and nurses to overcome limited human resources, and improved patient confidentiality are likely to increase timely access to HIV treatment services for more patients.


Biphasic effects of moderate drinking on prolactin during lactation.
Contrary to the popular lore that encourages women to drink alcohol as an aid to lactation, we previously showed that alcohol consumption disrupted lactational performance and the hormonal milieu of the lactating mother in the short term. Thirteen lactating women participated in a 4-session, double-blind, 2 x 2 within-subject study to test several hypotheses related to the effects of alcohol on prolactin (PRL) responses and milk yield over time. The two within-subject factors were beverage condition (control or 0.4 g/kg dose of alcohol) and pumping condition (pumping occurred at fixed intervals once or twice during the 5.3-hour session). Plasma PRL, blood alcohol concentrations (BAC), and milk yield were measured. Alcohol consumption increased basal PRL levels (p < 0.0001) and modified the PRL response to pumping (p < 0.0001) but the directionality of the response depended on when pumping occurred along the BAC curve. Pumping enhanced PRL response when it occurred during the ascending BAC limb but blunted the response when it occurred during the descending limb, providing evidence that the effects were transient and of a biphasic nature. The slower the alcohol was metabolized, the greater the relative PRL response to breast pumping (p < 0.05). The dynamics of the PRL response between pumping sessions was also altered if women drank. If women pumped within the hour after drinking alcohol, the PRL response during the next pumping some 1.5 hours later, was delayed by a few minutes. Milk yield was significantly lower after drinking alcohol but such deficits were not significantly related to PRL or the speed at which alcohol was eliminated. Effects of alcohol on suckling-induced PRL were biphasic in nature, but could not explain the deficits in lactational performance. Such findings provide further evidence that the dynamic changes in neuroendocrine state are integrally involved in alcohol's effects over time and underscore the complexity of lactation.


COVID-19 lung CT image segmentation using deep learning methods: U-Net versus SegNet.
Currently, there is an urgent need for efficient tools to assess the diagnosis of COVID-19 patients. In this paper, we present feasible solutions for detecting and labeling infected tissues on CT lung images of such patients. Two structurally-different deep learning techniques, SegNet and U-NET, are investigated for semantically segmenting infected tissue regions in CT lung images. We propose to use two known deep learning networks, SegNet and U-NET, for image tissue classification. SegNet is characterized as a scene segmentation network and U-NET as a medical segmentation tool. Both networks were exploited as binary segmentors to discriminate between infected and healthy lung tissue, also as multi-class segmentors to learn the infection type on the lung. Each network is trained using seventy-two data images, validated on ten images, and tested against the left eighteen images. Several statistical scores are calculated for the results and tabulated accordingly. The results show the superior ability of SegNet in classifying infected/non-infected tissues compared to the other methods (with 0.95 mean accuracy), while the U-NET shows better results as a multi-class segmentor (with 0.91 mean accuracy). Semantically segmenting CT scan images of COVID-19 patients is a crucial goal because it would not only assist in disease diagnosis, also help in quantifying the severity of the illness, and hence, prioritize the population treatment accordingly. We propose computer-based techniques that prove to be reliable as detectors for infected tissue in lung CT scans. The availability of such a method in today's pandemic would help automate, prioritize, fasten, and broaden the treatment of COVID-19 patients globally.


Measurement of intrasynaptosomal free calcium by using the fluorescent indicator quin-2.
The recently synthesized calcium indicator quin -2 was incorporated into synaptosomes from guinea-pig cerebral cortex following uptake and internal hydrolysis of quin -2 tetra-acetoxymethyl ester. Incubation in physiological media containing 1 mM- or 2 mM-CaCl2 led to equilibrium cytosolic ionized calcium concentrations of 85 +/- 10 nM and 205 +/- 5 nM respectively (mean +/- S.E.M. from eight and eighteen preparations respectively). Cytosolic Ca2+ was elevated following increases in external Ca2+ concentration, plasma membrane depolarization, mitochondrial inhibition, calcium ionophore addition or replacement of external sodium by lithium. Preliminary experiments were performed to assess changes in cytosolic Ca2+ accompanying the release of the neurotransmitter acetylcholine.


Basic apoptotic mechanisms of lead toxicity in human leukemia (HL-60) cells.
Lead exposure represents a medical and public health emergency, especially in children consuming high amounts of lead-contaminated flake paints. It may also cause hematological effects to people of all ages. Recent studies in our laboratory have indicated that apoptosis may be associated with the lead-induced oxidative stress and DNA damage. However, the mechanisms underlying its effect on lymphocytes are still largely unknown. Therefore, the aim of the present study was to investigate the apoptotic mechanisms of lead nitrate [Pb(NO(3))(2)] using HL-60 cells as a test model. HL-60 cells were treated with different concentrations of Pb(NO(3))(2) for 24 h prior to cell viability assay and flow cytometry assessment. The results obtained from the trypan blue exclusion test indicated that at very low concentration, Pb(NO(3))(2) has no effect on the viability of HL-60 cells. A significant (p < 0.05) decrease in cell viability was observed when exposed to high level of Pb(NO(3))(2). Data generated from the flow cytometric assessment indicated that Pb(NO(3))(2) exposure significantly (p < 0.05) increased the proportion of annexin V positive cells (apoptotic cells) compared to the control. Pb(NO(3))(2) induced apoptosis of HL-60 cells was associated with the activation of caspase-3. In summary, these studies demonstrated that Pb(NO(3))(2) represents an apoptosis-inducing agent in HL-60 promyelocytic leukemia cells and its apoptotic mechanism functions, at least in part via, induction of phosphatidylserine externalization and caspase-3 activation.


Distribution and abundance of the salmonid parasite Parvicapsula minibicornis (Myxozoa) in the Klamath River basin (Oregon-California, U.S.A.).
The distribution and abundance of the myxosporean parasite Parvicapsula minibicornis in the Klamath River mirrored that of Ceratomyxa shasta, with which it shares both its vertebrate and invertebrate host. Assay of fish held at sentinel sites and water samples collected from those sites showed that parasite prevalence was highest below Iron Gate dam, which is the barrier to anadromous salmon passage. Above this barrier parasite levels fluctuated, with the parasite detected in the free-flowing river reaches between reservoirs. This was consistent with infection prevalence in the polychaete host, Manayunkia speciosa, which was greater than 1% only in populations tested below Iron Gate dam. Although a low prevalence of infection was detected in juvenile out-migrant fish in the Trinity River, the tributaries tested did not appear to be a significant source of the parasite to the mainstem despite the presence of large numbers of infected adult salmon that migrate and spawn there. Rainbow trout became infected during sentinel exposure, which expands the host range for P. minibicornis and suggests that wild rainbow trout populations are a reservoir for infection, especially above Iron Gate dam. High parasite prevalence in the lower Klamath River is likely a combined effect of high spore input from heavily infected, spawned adult salmon and the proximity to dense populations of polychaetes.


An intraspecific appraisal of the social intelligence hypothesis.
The prevailing hypotheses for the evolution of cognition focus on either the demands associated with group living (the social intelligence hypothesis (SIH)) or ecological challenges such as finding food. Comparative studies testing these hypotheses have generated highly conflicting results; consequently, our understanding of the drivers of cognitive evolution remains limited. To understand how selection shapes cognition, research must incorporate an intraspecific approach, focusing on the causes and consequences of individual variation in cognition. Here, we review the findings of recent intraspecific cognitive research to investigate the predictions of the SIH. Extensive evidence from our own research on Australian magpies (Cracticus tibicen dorsalis), and a number of other taxa, suggests that individuals in larger social groups exhibit elevated cognitive performance and, in some cases, elevated reproductive fitness. Not only do these findings demonstrate how the social environment has the potential to shape cognitive evolution, but crucially, they demonstrate the importance of considering both genetic and developmental factors when attempting to explain the causes of cognitive variation.This article is part of the theme issue 'Causes and consequences of individual differences in cognitive abilities'.


Association of GSTTI and GSTM1 variants with acute myeloid leukemia risk.
We aimed to investigate the relationships between polymorphisms of the glutathione S-transferases (GSTs) GSTM1, GSTTI, and GSTP1 and the risk of developing acute myeloid leukemia (AML). A total of 206 AML cases and 231 controls were collected for our study. The genotyping of GSTs (GSTM1, GSTTI, and GSTP1) was based upon the duplex polymerase chain reaction with the confronting two-pair primer (PCR-CTPP) method. Individuals carrying null GSTTI and GSTM1 genotypes had a 1.52- and 1.78-fold increased risk of developing acute leukemia, respectively, compared to non-null genotype carriers (P < 0.05). A high risk was observed in those carrying a combination of null genotypes of GSTM1 and GSTTI with GSTP1-Val allele genotypes when compared with those carrying wild-type genotypes, with an odds ratio (95% confidence interval) of 3.62 (1.53-8.82) (P < 0.05). These findings indicate that genetic variants of GSTTI and GSTM1 significantly increase the risk of developing AML. Our study offers important insights into the molecular etiology of AML.


Red fluorescent genetically encoded Ca2+ indicators for use in mitochondria and endoplasmic reticulum.
Ca2+ is a key intermediary in a variety of signalling pathways and undergoes dynamic changes in its cytoplasmic concentration due to release from stores within the endoplasmic reticulum (ER) and influx from the extracellular environment. In addition to regulating cytoplasmic Ca2+ signals, these responses also affect the concentration of Ca2+ within the ER and mitochondria. Single fluorescent protein-based Ca2+ indicators, such as the GCaMP series based on GFP, are powerful tools for imaging changes in the concentration of Ca2+ associated with intracellular signalling pathways. Most GCaMP-type indicators have dissociation constants (Kd) for Ca2+ in the high nanomolar to low micromolar range and are therefore optimal for measuring cytoplasmic [Ca2+], but poorly suited for use in mitochondria and ER where [Ca2+] can reach concentrations of several hundred micromolar. We now report GCaMP-type low-affinity red fluorescent genetically encoded Ca2+ indicators for optical imaging (LAR-GECO), engineered to have Kd values of 24 μM (LAR-GECO1) and 12 μM (LAR-GECO1.2). We demonstrate that these indicators can be used to image mitochondrial and ER Ca2+ dynamics in several cell types. In addition, we perform two-colour imaging of intracellular Ca2+ dynamics in cells expressing both cytoplasmic GCaMP and ER-targeted LAR-GECO1. The development of these low-affinity intensiometric red fluorescent Ca2+ indicators enables monitoring of ER and mitochondrial Ca2+ in combination with GFP-based reporters.


Cellular senescence in tissue repair: every cloud has a silver lining.
Cellular senescence, a form of stable cell cycle arrest induced by cellular stress, constitutes a major factor leading to the promotion of pathologies and physiological decays that take place during ageing. However, in recent years evidence has started to emerge supporting a positive role for senescent cells in various physiological processes, from embryonic development to tissue injury responses such as wound healing and tissue repair. Here, we provide an overview of cellular senescence, its negative as well as positive outcomes, with a focus on its impact on tissue repair. Furthermore, we discuss the possibility that cell senescence could contribute to the regeneration of complex structures and explore recent findings with respect to their potential for therapeutic application.


TEAD4 establishes the energy homeostasis essential for blastocoel formation.
It has been suggested that during mouse preimplantation development, the zygotically expressed transcription factor TEAD4 is essential for specification of the trophectoderm lineage required for producing a blastocyst. Here we show that blastocysts can form without TEAD4 but that TEAD4 is required to prevent oxidative stress when blastocoel formation is accompanied by increased oxidative phosphorylation that leads to the production of reactive oxygen species (ROS). Both two-cell and eight-cell Tead4(-/-) embryos developed into blastocysts when cultured under conditions that alleviate oxidative stress, and Tead4(-/-) blastocysts that formed under these conditions expressed trophectoderm-associated genes. Therefore, TEAD4 is not required for specification of the trophectoderm lineage. Once the trophectoderm was specified, Tead4 was not essential for either proliferation or differentiation of trophoblast cells in culture. However, ablation of Tead4 in trophoblast cells resulted in reduced mitochondrial membrane potential. Moreover, Tead4 suppressed ROS in embryos and embryonic fibroblasts. Finally, ectopically expressed TEAD4 protein could localize to the mitochondria as well as to the nucleus, a property not shared by other members of the TEAD family. These results reveal that TEAD4 plays a crucial role in maintaining energy homeostasis during preimplantation development.


Displacement of a native by an alien bumblebee: lower pollinator efficiency overcome by overwhelmingly higher visitation frequency.
Biological invasions might constitute a major threat to mutualisms. Introduced pollinators might competitively displace their native counterparts, which in turn affects the pollination of native plants, if native and alien visitors differ in pollinator effectiveness. Since its invasion in 1994 into south-west Argentina, the introduced European bumblebee Bombus ruderatus has continuously increased in abundance, along with a simultaneous decrease in the abundance of the native Bombus dahlbomii. The latter is the only native bumblebee species of the temperate forests of southern South America, and the main pollinator of the endemic herb Alstroemeria aurea. In order to evaluate the impact of the ongoing displacement of the native by the alien bumblebee, we compared the pollinator effectiveness (i.e., the combination of pollinator efficiency per visit and visitation frequency) between both bumblebee species, as well as related pollinator traits that might account for potential differences in pollinator efficiency. Native Bombus dahlbomii, which has a larger body and spent more time per flower, was the more efficient pollinator compared to Bombus ruderatus, both in terms of quantity and quality of pollen deposited per visit. However, Bombus ruderatus was a much more frequent flower visitor than Bombus dahlbomii. As a consequence, Bombus ruderatus is nowadays a more effective pollinator of A. aurea than its native congener. Despite the lack of evidence of an increase in seed set at the population level, comparisons with historical records of Bombus dahlbomii abundances prior to Bombus ruderatus' invasion suggest that the overall pollination intensity of A. aurea might in fact have risen as a consequence of this invasion. Field experiments like these, that incorporate the natural variation in abundance of native and alien species, are powerful means to demonstrate that the consequences of invasions are more complex than previous manipulated and controlled experiments have suggested.


Immunogenicity and safety of intradermal versus intramuscular route of influenza immunization in infants less than 6 months of age: a randomized controlled trial.
We aimed to explore intradermal influenza vaccination in infants <6 months. One hundred twenty-six infants 2-3 months of age were randomized to receive either two doses, 1 month apart, of 0.25 ml of a trivalent inactivated influenza vaccine (7.5 microg of hemagglutinin per strain) via the intramuscular (IM) route or 0.1 ml of the same vaccine (3 microg of hemagglutinin per strain) via the intradermal (ID) route. The vaccine was well tolerated. Only four infants had hemagglutination inhibition (HAI) titer <40 against > or =1 vaccine-covered antigen pre-vaccination. There was no difference in fold-rise of HAI titer response between those in the IM or ID group. We documented maintenance of HAI titers above seroprotective levels against all three vaccine antigens in 97.6% of subjects regardless of vaccination methods over a time of waning maternal antibodies.


Considerations on Experimental Design and Data Analysis of Chromatin Immunoprecipitation Experiments.
Arguably one of the most valuable techniques to study chromatin organization, ChIP is the method of choice to map the contacts established between proteins and genomic DNA. Ever since its inception, more than 30 years ago, ChIP has been constantly evolving, improving, and expanding its capabilities and reach. Despite its widespread use by many laboratories across a wide variety of disciplines, ChIP assays can be sometimes challenging to design, and are often sensitive to variations in practical implementation.In this chapter, we provide a general overview of the ChIP method and its most common variations, with a special focus on ChIP-seq. We try to address some of the most important aspects that need to be taken into account in order to design and perform experiments that generate the most reproducible, high-quality data. Some of the main topics covered include the use of properly characterized antibodies, alternatives to chromatin preparation, the need for proper controls, and some recommendations about ChIP-seq data analysis.


[Suitability of Russian surnames as a "quasi-genetic" marker].
The possible usefulness of Russian surnames as a quasi-genetic marker in the population study is analysed. Two approaches for the populations of different hierarchical levels are used. The coefficients of correlations between the matrices of genetical (based on gene frequencies in the populations) and quasi-genetical (based on surnames frequencies in the populations) distances were determined. The mean correlation coefficient is 0.315 +/- 0.15. Genetical and quasi-genetical distances and the level of gamete exchange between populations are connected, being reversely dependent. The mean correlation coefficients are: -0.63 +/- 0.13 and -0.23 +/- 0.16. The correlation coefficient for matrices of quasi-genetical and geographical distances was 0.39 +/- 0.08, this being -0.42 +/- 0.08 for matrices of quasi-genetical distance and geometrical mean of population dimensions. It is shown that surnames can replace genetic markers in description of the population genetic structure and in the study of the processes influencing this structure.


Detecting conserved secondary structures in RNA molecules using constrained structural alignment.
Constrained sequence alignment has been studied extensively in the past. Different forms of constraints have been investigated, where a constraint can be a subsequence, a regular expression, or a probability matrix of symbols and positions. However, constrained structural alignment has been investigated to a much lesser extent. In this paper, we present an efficient method for constrained structural alignment and apply the method to detecting conserved secondary structures, or structural motifs, in a set of RNA molecules. The proposed method combines both sequence and structural information of RNAs to find an optimal local alignment between two RNA secondary structures, one of which is a query and the other is a subject structure in the given set. The method allows a biologist to annotate conserved regions, or constraints, in the query RNA structure and incorporates these regions into the alignment process to obtain biologically more meaningful alignment scores. A statistical measure is developed to assess the significance of the scores. Experimental results based on detecting internal ribosome entry sites in the RNA molecules of hepatitis C virus and Trypanosoma brucei demonstrate the effectiveness of the proposed method and its superiority over existing techniques.


Impaired angiogenesis following hind-limb ischemia in diabetes mellitus mice.
To investigate vascular endothelial growth factor (VEGF) and its signaling pathway spontaneous response in type 2 diabetes mellitus (T2DM) mice to surgery-induced hind-limb ischemia. Sixty mice were randomly divided into two groups, one was fed with normal chow as control, and another was fed with high fat diet to induce T2DM. Fourteen weeks later, mice were surgically induced to hind-limb ischemia. Blood flow restoration was monitored with laser Doppler perfusion imaging. Tibialis anterior muscle was collected after 3 days of hind-limb ischemia. VEGF mRNA and protein expressions were analyzed using real-time PCR and ELISA; expressions of VEGF downstream signal molecules and receptors were analyzed using Western blotting and RT-PCR, respectively. Perfusion recovery 10, 20, 30 days after ischemia was significantly attenuated in T2DM compared with control group (P < 0.05). T2DM impaired VEGF signaling pathway although VEGF levels increased in T2DM group. After ischemia, T2DM group had a comparable increase in VEGF expression compared with control group, but still had an impaired VEGF signaling pathway. VEGF signaling pathway is abnormal in T2DM mice, although VEGF had a response to the ischemic stimulation.


Functional morphology of the copulatory system of box crabs with long second gonopods (Calappidae, Eubrachyura, Decapoda, Crustacea).
Male True Crabs use two pairs of gonopods to deliver mating products during copulation. Commonly, the second pair is shorter than the first pair, and most research to date has focused on species with short second gonopods. We investigated male and female copulatory organs in Calappula saussurei and Calappa pelii, two species of box crabs (Calappidae) with second gonopods which are longer than the first pair. Scanning electron microscopy and histological cross sectioning show that the female copulatory system is unique in several aspects: the genital duct is part concave and part simple type. The seminal receptacle is divided into two chambers, a ventral chamber of ectodermal and mesodermal origin, and a dorsal chamber of ectodermal origin. This dorsal chamber is the location of spermatophore reception during copulation. A sperm plug closes the dorsal chamber off. We propose that long second gonopods deliver male mating products directly into the dorsal chamber. To date, spermatophore reception has been associated with the mesodermal tissue of the seminal receptacle. The copulatory system of box crabs with long second gonopods shows novel deviations from this general pattern.


MUC4 (sialomucin complex) expression in salivary gland tumors and squamous cell carcinoma of the upper aerodigestive tract.
This study investigates MUC4 expression in normal squamous epithelia and squamous cell carcinoma (SCC) of the upper aerodigestive tract (UADT), and in salivary gland neoplasms. MUC4 antigens in tumor and adjacent normal tissue are localized by immunocytochemical studies. Fresh frozen tissues from surgical resection specimens are further analyzed by Western blot. MUC4 is identified by immunocytochemical staining throughout the normal UADT mucosa, in 34 of 40 primary UADT SCC, and in 11 of 12 metastatic cervical lymph nodes. A trend toward decreased MUC4 staining in moderately and poorly differentiated tumors is noted. Immunoblots show MUC4 in 4 of 5 SCC analyzed. Immunocytochemical staining of MUC4 in 13 major and minor salivary gland neoplasms reveal variable staining of normal and neoplastic tissue. MUC4 is demonstrated in immunoblots of normal parotid tissue and in the single parotid malignancy analyzed, but is not demonstrated in one minor salivary gland malignancy. These findings characterize normal UADT mucosal and salivary MUC4 expression, and MUC4 expression in SCC of the UADT and in salivary gland tumors. Correlation of MUC4 expression with clinical outcomes may establish MUC4 as a potential molecular prognostic marker for these tumors.


Perioperative complications after noncardiac surgery in patients with insertion of second-generation drug-eluting stents.
The perioperative outcomes of noncardiac surgery in patients who have received second-generation drug-eluting stents (DESs) have received limited study. We reviewed the medical records of 1,748 consecutive patients who received DES at our institution (1,789 procedures) from January 1, 2009, to July 1, 2012, to determine the outcomes of subsequent noncardiac surgery. During a median follow-up of 43 months, 221 patients underwent 345 noncardiac surgeries (138 low risk, 130 intermediate risk, and 77 high risk), of which 278 were in patients with previous second-generation DES implantation. The incidence of noncardiac surgery in patients with previous second-generation DES implantation was 4.5% at 1 year, 11.6% at 2 years, and 15.2% at 3 years. The mean time from stent implantation to surgery was 21±12.9 months. Mean age was 66±8 years, 99% were men, and 11% had a perioperative complication, including 5.8% major bleeding, 2.5% acute kidney injury, 2.2% major adverse cardiac event, and 1.4% stroke. Perioperative stent thrombosis occurred in 2 patients (0.7%, 95% confidence interval 0.2% to 2.6%): 1 patient had received a DES 14 months before surgery and had stent thrombosis on the day of surgery and the other had a DES implanted 21 months before surgery and developed stent thrombosis the day after surgery. In conclusion, the incidence of perioperative complications with noncardiac surgery after second-generation DES implantation was 11% and consisted mainly of bleeding (5.8%). The incidence of definite stent thrombosis was 0.7%.


Structural organization of a major neuronal G protein regulator, the RGS7-Gβ5-R7BP complex.
Signaling by the G-protein-coupled receptors (GPCRs) plays fundamental role in a vast number of essential physiological functions. Precise control of GPCR signaling requires action of regulators of G protein signaling (RGS) proteins that deactivate heterotrimeric G proteins. RGS proteins are elaborately regulated and comprise multiple domains and subunits, yet structural organization of these assemblies is poorly understood. Here, we report a crystal structure and dynamics analyses of the multisubunit complex of RGS7, a major regulator of neuronal signaling with key roles in controlling a number of drug target GPCRs and links to neuropsychiatric disease, metabolism, and cancer. The crystal structure in combination with molecular dynamics and mass spectrometry analyses reveals unique organizational features of the complex and long-range conformational changes imposed by its constituent subunits during allosteric modulation. Notably, several intermolecular interfaces in the complex work in synergy to provide coordinated modulation of this key GPCR regulator.