Automated long-term recording and analysis of neural activity in behaving animals.
Addressing how neural circuits underlie behavior is routinely done by measuring electrical activity from single neurons in experimental sessions. While such recordings yield snapshots of neural dynamics during specified tasks, they are ill-suited for tracking single-unit activity over longer timescales relevant for most developmental and learning processes, or for capturing neural dynamics across different behavioral states. Here we describe an automated platform for continuous long-term recordings of neural activity and behavior in freely moving rodents. An unsupervised algorithm identifies and tracks the activity of single units over weeks of recording, dramatically simplifying the analysis of large datasets. Months-long recordings from motor cortex and striatum made and analyzed with our system revealed remarkable stability in basic neuronal properties, such as firing rates and inter-spike interval distributions. Interneuronal correlations and the representation of different movements and behaviors were similarly stable. This establishes the feasibility of high-throughput long-term extracellular recordings in behaving animals.


Creating high-level components with a generative representation for body-brain evolution.
One of the main limitations of scalability in body-brain evolution systems is the representation chosen for encoding creatures. This paper defines a class of representations called generative representations, which are identified by their ability to reuse elements of the genotype in the translation to the phenotype. This paper presents an example of a generative representation for the concurrent evolution of the morphology and neural controller of simulated robots, and also introduces GENRE, an evolutionary system for evolving designs using this representation. Applying GENRE to the task of evolving robots for locomotion and comparing it against a non-generative (direct) representation shows that the generative representation system rapidly produces robots with significantly greater fitness. Analyzing these results shows that the generative representation system achieves better performance by capturing useful bias from the design space and by allowing viable large scale mutations in the phenotype. Generative representations thereby enable the encapsulation, coordination, and reuse of assemblies of parts.


Detection of Koi Herpesvirus (KHV) and Carp Oedema Virus (CEV) in Invasive Round Goby, Neogobius Melanostomus Pallas, 1814, from Poland and Germany.
The aim of the study was to determine the transmission potential of carp edema virus (CEV) and koi herpesvirus (KHV) introduced to Europe by the invasive round goby (Neogobius melanostomus). A total of 70 round goby specimens were collected from the Szczecin Lagoon, Poland, and locations in Germany in the third and fourth quarters of 2018. The fish were analysed to detect KHV and CEV by PCR. Six fish specimens were positive for the presence of KHV, while none of the gobies examined showed the presence of CEV. The CEV genome was detected in the goby specimens from Germany and from Poland. Considering the high pace of the spread of the round goby and its effectiveness in acquisition of new ecological niches, it should be kept out during refilling of carp ponds. Further studies should focus on experimental cohabitation of CEV-infected round gobies and specific-pathogen-free (SPF) carp to investigate the potential for active virus transfer.


Does pain mediate or moderate the relationship between physical activity and depressive symptoms in older people? Findings from The Irish Longitudinal Study on Ageing (TILDA).
Background. Depression is an increasing problem in older adults, which is exacerbated by under diagnosis and ineffective treatment options. Broadly speaking, as people age, their levels of regular physical activity (PA) decrease, while their experience of chronic pain increases. PA has been shown to be an effective, yet under-utilised, treatment for depression in this age-cohort although the influence of pain on the relationship between PA and depressive symptoms has not been considered. Methods. Secondary analysis of national data from The Irish Longitudinal Study on Ageing (TILDA, 2011) (n = 8163 participants aged 50 years and older) examined the mediating or moderating role of pain in the relationship between depressive symptoms and PA, and the impact of PA, pain and depressive symptoms on health-care utilisation. Results. Approximately 8.5% TILDA older adults were depressed. No mediating or moderating effects of pain were found in the association between PA and depressive symptoms. Higher levels of PA were found to be independently associated with lower depressive symptoms, while higher levels of pain significantly increased the likelihood of depressive symptoms supporting previous findings. Depressive symptoms and higher levels of pain were also found to significantly increase health-care utilisation. Conclusions. Consistent with previous findings in this field, both PA and pain were found to be independently associated with depressive symptoms in Irish older adults. Furthermore, pain does not play a mediating or moderating role in the relationship between PA and depressive symptoms. Continued support for ongoing initiatives in this area aimed at increasing PA in older adults as a means to improve both physical and mental well-being is advised. The absence of any synergistic effect between PA and pain suggests that clinicians and health service providers should continue to promote PA as a treatment for depression, irrespective of the pain levels of their patients.


The association of health literacy with cervical cancer prevention knowledge and health behaviors in a multiethnic cohort of women.
Our purpose was to describe the relationship between health literacy, ethnicity, and cervical cancer screening practices and to evaluate physician recognition of low literacy. We studied a prospective cohort of English-speaking patients > or =18 years (n = 529) in ambulatory women's clinics. Univariate and multivariate analyses were used to evaluate demographics, health practices and beliefs, and knowledge regarding cervical cancer screening and prevention. Physicians' assessments of patient reading skills were obtained. Low health literacy (<9th grade) was found among 40% of participants. Minority women were half as likely to know the purpose of the Papanicolaou test (9% vs 21%; P <.03) and were significantly more likely to have low literacy levels compared with white women (46% vs 15%; P <.05). Literacy was the only factor independently associated with knowledge related to cervical cancer screening (adjusted odds ratio, 2.25; 95% CI, 1.05-4.80). Physicians detected only 20% of the lowest readers. Poor health literacy was a better predictor of cervical cancer screening knowledge than ethnicity or education, yet physicians infrequently recognized low literacy. Improved physician awareness and development of low literacy interventions may improve cervical cancer screening, particularly for the most vulnerable women.


Thrombocytosis as a Rare Paraneoplastic Syndrome Occurring in Hepatocellular Carcinoma: A Case Report.
Hepatocellular carcinoma (HCC) affects more than half a million people worldwide each year. Paraneoplastic syndromes associated with HCC include erythrocytosis, hypercalcemia, hypercholesterolemia, hypoglycemia and thrombocytosis. Thrombocytosis is a rare paraneoplastic syndrome in HCC mediated by thrombopoietin (TPO) production. We report a case of thrombocytosis as a paraneoplastic syndrome in a patient with HCC and hepatitis C cirrhosis. A 56-year-old patient was evaluated with abdominal distension and pain of 1-month duration. He had a history of hepatitis C infection with liver cirrhosis, CTP (Child-Turcotte-Pugh) class C, MELD (model for end-stage liver disease) score 22, methadone dependence, alcohol abuse and depression. His physical examination was remarkable for distended abdomen with shifting dullness, palpable hepatomegaly and scleral icterus. Routine laboratory tests showed a platelet count of 754 k/µL, white blood cell count 12.4 k/µL, serum sodium level 128 mEq/L, alanine aminotransferase 93 U/L, aspartate aminotransferase 871 U/L, total serum bilirubin 4.3 mg/dL, direct serum bilirubin 2.8mg/dL and albumin 2.6 g/dL. Computed tomography of the abdomen and pelvis revealed hepatomegaly with numerous hypodensities suspicious for HCC. Abdominal paracentesis was done, serum ascites albumin gradient (SAAG) was 2.4 g/dL consistent with portal hypertension, and spontaneous bacterial peritonitis was ruled out. Magnetic resonance imaging of the liver was consistent with infiltrating HCC, portal vein thrombosis and retroperitoneal lymphadenopathy. His alpha fetoprotein (AFP) level was 79,102 ng/mL and TPO level was 126 pg/mL. JAK2 mutation was negative and no other cause of reactive thrombocytosis could be identified. One year prior to this admission, the patient was noted to have a normal platelet count and AFP level. He was not considered a candidate for liver transplantation due to ongoing substance abuse, and expired 1 month later. Thrombocytosis is a rare paraneoplastic condition seen in HCC. It is presumed to be secondary to increased production of TPO by the tumor. We observed an elevated level of TPO in our patient. Thrombocytosis in HCC is associated with a high tumor burden, portal vein thrombosis (PVT), serum AFP levels and a poor prognosis. Thrombocytosis in a cirrhotic patient should alert the presence of HCC and is associated with poor outcomes.


Single-Isocenter Frameless Volumetric Modulated Arc Radiosurgery for Multiple Intracranial Metastases.
Stereotactic radiosurgery (SRS) is a well-accepted treatment for patients with intracranial metastases, but outcomes with volumetric modulated arc radiosurgery (VMAR) are poorly described. To report our initial clinical experience applying a novel single-isocenter technique to frameless VMAR for simultaneous treatment of multiple intracranial metastases. We performed a retrospective analysis of 15 patients undergoing frameless VMAR for multiple intracranial metastases using a single, centrally located isocenter in the period 2009 and 2011. Of these, 3 patients were treated for progressive or recurrent intracranial disease. A total of 62 metastases (median, 3 per patient; range, 2-13) were treated to a median dose of 20 Gy (range, 15-30 Gy). Three patients were treated with fractionated SRS. Follow-up including clinical examination and magnetic resonance imaging (MRI) occurred every 3 months. The median follow-up for all patients was 7.1 months (range, 1.1-24.3), with 11 patients (73.3%) followed until death. For the remaining 4 patients alive at the time of analysis, the median follow-up was 19.6 months (range, 9.2-24.3). Local control at 6 and 12 months was 91.7% (95% confidence interval [CI], 84.6%-100.0%) and 81.5% (95% CI, 67.9%-100.0%), respectively. Regional failure was observed in 9 patients (60.0%), and 7 patients (46.7%) received salvage therapy. Overall survival at 6 months was 60.0% (95% CI, 40.3%-88.2%). Grade 3 or higher treatment-related toxicity was not observed. The median total treatment time was 7.2 minutes (range, 2.8-13.2 minutes). Single-isocenter, frameless VMAR for multiple intracranial metastases is a promising technique that may provide similar clinical outcomes compared with conventional radiosurgery.


Exercise deficit disorder in youth: a paradigm shift toward disease prevention and comprehensive care.
Despite the widely recognized benefits of daily play, recreation, sports, and physical education on the physical and psychosocial well-being of children and adolescents, many contemporary children and adolescents worldwide do not meet the recommendations for daily physical activity (PA). The decline in PA seems to start early in life, which leads to conditions characterized by reduced levels of PA in the pediatric population that are inconsistent with current public health recommendations. Unlike many other diseases and disorders in pediatrics, physical inactivity in youth is unique in that it currently lacks a clinical gold standard for diagnosis. This makes the diagnosis and treatment medically challenging, though no less important, as the resultant ramifications of a missed diagnosis are of significant detriment. Exercise-deficient children need to be identified early in life and treated with developmentally appropriate exercise programs designed to target movement deficiencies and physical weaknesses in a supportive environment. Without such interventions early in life, children are more likely to become resistant to our interventions later in life and consequently experience adverse health consequences. Integrative approaches that link health care professionals, pediatric exercise specialists, school administrators, community leaders, and policy makers may provide the best opportunity to promote daily PA, reinforce desirable behaviors, and educate parents about the exercise-health link.


Peptide and protein modulation of local Ca2+ release events in permeabilized skeletal muscle fibers.
Local discrete elevations in myoplasmic Ca2+ (Ca2+ sparks) arise from the opening of a small group of RyRs. Summation of a large number of Ca2+ sparks gives rise to the whole cell Ca2+ transient necessary for muscle contraction, Unlike sarcoplasmic reticulum vesicle preparations and isolated single channels in artificial membranes, the study of Ca2+ sparks provides a means to understand the regulation of a small group of RyRs in the environment of a functionally intact triad and in the presence of endogenous regulatory proteins. To gain insight into the mechanisms that regulate the gating of RyRs we have utilized laser scanning confocal microscopy to measure Ca2+ sparks in permeabilized frog skeletal muscle fibers. This review summarizes our recent studies using both exogenous (ImperatoxinA and domain peptides) and endogenous (calmodulin) modulators of RyR to gain insight into the number of RyR Ca2+ release channels underlying a Ca2+ spark, how domain-domain interactions within RyR regulate the functional state of the channel as well as gating mechanisms of RyR in living muscle fibers.


Development and deployment of a rapid recombinase polymerase amplification Ebola virus detection assay in Guinea in 2015.
In the absence of a vaccine or specific treatments for Ebola virus disease (EVD), early identification of cases is crucial for the control of EVD epidemics. We evaluated a new extraction kit (SpeedXtract (SE), Qiagen) on sera and swabs in combination with an improved diagnostic reverse transcription recombinase polymerase amplification assay for the detection of Ebola virus (EBOV-RT-RPA). The performance of combined extraction and detection was best for swabs. Sensitivity and specificity of the combined SE and EBOV-RT-RPA were tested in a mobile laboratory consisting of a mobile glovebox and a Diagnostics-in-a-Suitcase powered by a battery and solar panel, deployed to Matoto Conakry, Guinea as part of the reinforced surveillance strategy in April 2015 to reach the goal of zero cases. The EBOV-RT-RPA was evaluated in comparison to two real-time PCR assays. Of 928 post-mortem swabs, 120 tested positive, and the combined SE and EBOV-RT-RPA yielded a sensitivity and specificity of 100% in reference to one real-time RT-PCR assay. Another widely used real-time RT-PCR was much less sensitive than expected. Results were provided very fast within 30 to 60 min, and the field deployment of the mobile laboratory helped improve burial management and community engagement.


Pivotal contributions of megakaryocytes to the biology of idiopathic myelofibrosis.
In order to investigate the biologic processes underlying and resulting from the megakaryocytic hyperplasia that characterizes idiopathic myelofibrosis (IMF), peripheral blood CD34+ cells isolated from patients with IMF, polycythemia vera (PV), and G-CSF-mobilized healthy volunteers were cultured in the presence of stem cell factor and thrombopoietin. IMF CD34+ cells generated 24-fold greater numbers of megakaryocytes (MKs) than normal CD34+ cells. IMF MKs were also shown to have a delayed pattern of apoptosis and to overexpress the antiapoptotic protein bcl-xL. MK hyperplasia in IMF is, therefore, likely a consequence of both the increased ability of IMF progenitor cells to generate MKs and a decreased rate of MK apoptosis. Media conditioned (CM) by CD61+ cells generated in vitro from CD34+ cells were then assayed for the levels of growth factors and proteases. Higher levels of transforming growth factor-beta (TGF-beta) and active matrix metalloproteinase-9 (MMP9) were observed in media conditioned with IMF CD61+ cells than normal or PV CD61+ cells. Both normal and IMF CD61+ cells produced similar levels of VEGF. MK-derived TGF-B and MMP-9, therefore, likely contribute to the development of many pathological epiphenomena associated with IMF.


Ric-8 proteins are molecular chaperones that direct nascent G protein α subunit membrane association.
Ric-8A (resistance to inhibitors of cholinesterase 8A) and Ric-8B are guanine nucleotide exchange factors that enhance different heterotrimeric guanine nucleotide-binding protein (G protein) signaling pathways by unknown mechanisms. Because transgenic disruption of Ric-8A or Ric-8B in mice caused early embryonic lethality, we derived viable Ric-8A- or Ric-8B-deleted embryonic stem (ES) cell lines from blastocysts of these mice. We observed pleiotropic G protein signaling defects in Ric-8A(-/-) ES cells, which resulted from reduced steady-state amounts of Gα(i), Gα(q), and Gα(13) proteins to <5% of those of wild-type cells. The amounts of Gα(s) and total Gβ protein were partially reduced in Ric-8A(-/-) cells compared to those in wild-type cells, and only the amount of Gα(s) was reduced substantially in Ric-8B(-/-) cells. The abundances of mRNAs encoding the G protein α subunits were largely unchanged by loss of Ric-8A or Ric-8B. The plasma membrane residence of G proteins persisted in the absence of Ric-8 but was markedly reduced compared to that in wild-type cells. Endogenous Gα(i) and Gα(q) were efficiently translated in Ric-8A(-/-) cells but integrated into endomembranes poorly; however, the reduced amounts of G protein α subunits that reached the membrane still bound to nascent Gβγ. Finally, Gα(i), Gα(q), and Gβ(1) proteins exhibited accelerated rates of degradation in Ric-8A(-/-) cells compared to those in wild-type cells. Together, these data suggest that Ric-8 proteins are molecular chaperones required for the initial association of nascent Gα subunits with cellular membranes.


Resting states are resting traits--an FMRI study of sex differences and menstrual cycle effects in resting state cognitive control networks.
To what degree resting state fMRI is stable or susceptible to internal mind states of the individual is currently an issue of debate. To address this issue, the present study focuses on sex differences and investigates whether resting state fMRI is stable in men and women or changes within relative short-term periods (i.e., across the menstrual cycle). Due to the fact that we recently reported menstrual cycle effects on cognitive control based on data collected during the same sessions, the current study is particularly interested in fronto-parietal resting state networks. Resting state fMRI was measured in sixteen women during three different cycle phases (menstrual, follicular, and luteal). Fifteen men underwent three sessions in corresponding time intervals. We used independent component analysis to identify four fronto-parietal networks. The results showed sex differences in two of these networks with women exhibiting higher functional connectivity in general, including the prefrontal cortex. Menstrual cycle effects on resting states were non-existent. It is concluded that sex differences in resting state fMRI might reflect sexual dimorphisms in the brain rather than transitory activating effects of sex hormones on the functional connectivity in the resting brain.


Tumour-stromal interactions in acid-mediated invasion: a mathematical model.
It is well established that the tumour microenvironment can both promote and suppress tumour growth and invasion, however, most mathematical models of invasion view the normal tissue as inhibiting tumour progression via immune modulation or spatial constraint. In particular, the production of acid by tumour cells and the subsequent creation of a low extracellular pH environment has been explored in several 'acid-mediated tumour invasion' models where the acidic environment facilitates normal cell death and permits tumour invasion. In this paper, we extend the acid-invasion model developed by Gatenby and Gawlinski (1996) to include both the competitive and cooperative interactions between tumour and normal cells, by incorporating the influence of extracellular matrix and protease production at the tumour-stroma interface. Our model predicts an optimal level of tumour acidity which produces both cell death and matrix degradation. Additionally, very aggressive tumours prevent protease production and matrix degradation by excessive normal cell destruction, leading to an acellular (but matrix filled) gap between the tumour and normal tissue, a feature seen in encapsulated tumours. These results suggest, counterintuitively, that increasing tumour acidity may, in some cases, prevent tumour invasion.


Reactions of 2,6-dimethyl-1,3-dioxane-4-ol (aldoxane) with deoxyguanosine and DNA.
In a recent study, we identified several new DNA adducts of the carcinogen acetaldehyde, including N(2)-(2,6-dimethyl-1,3-dioxan-4-yl)deoxyguanosine (N(2)-aldoxane-dG, 2). Our goal in this study was to investigate further the formation of 2 by allowing 2,6-dimethyl-1,3-dioxane-4-ol (aldoxane, 5) to react with dG and DNA. Aldoxane is readily formed by trimerization of acetaldehyde. The reaction of aldoxane with dG and DNA produced diastereomers of N(2)-aldoxane-dG (2) as observed in the reactions of acetaldehyde with dG and DNA, supporting the intermediacy of aldoxane in their formation. Unexpectedly, however, an array of other adducts was formed in these reactions, including 3-(2-deoxyribos-1-yl)-5,6,7,8-tetrahydro-8-hydroxy-6-methylpyrimido[1,2-a]purine-10(3H)one (3), 2-amino-7,8-dihydro-8-hydroxy-6-methyl-3H-pyrrolo[2,1-f]purine-4(6H)one (13), N(2)-(3-hydroxybutylidene)dG (9), N(2)-[(2-hydroxypropyl)-6-methyl-1,3-dioxane-4-yl]dG (14), and N(2)-ethylidene-dG (1). Adduct 1 was the major product and was found to be quite stable in DNA. The adducts result from a cascade of aldehydes, e.g., 2-butenal (crotonaldehyde, 12), 3-hydroxybutanal (7) and its dimer (2-hydroxypropyl)-6-methyl-1,3-dioxane-4-ol (paraldol, 6), and acetaldehyde, produced from aldoxane under the reaction conditions. The reactions of aldoxane with dG and DNA were compared with those of paraldol. The paraldol reactions gave products resulting from reactions of dG and DNA with paraldol, 3-hydroxybutanal, and crotonaldehyde (adducts 3, 13, and 9) but the products of the aldoxane and acetaldehyde reactions (adducts 1 and 2) were not observed, indicating that paraldol is more stable under the reaction conditions than is aldoxane. The results of this study provide new insights about the formation of DNA adducts from aldehydes via condensation products of the latter.


Patient experiences and the association with organizational factors in general practice: results from the Norwegian part of the international, multi-centre, cross-sectional QUALICOPC study.
General practitioners (GPs) constitute a vital part of a strong primary health care system. We need further knowledge concerning factors that may affect the patients' experiences in their meetings with the GPs. We investigated to what degree organizational factors and GP characteristics are associated with patients' communicative experiences in a consultation. We used data from the Norwegian part of the international, multi-center study Quality and Costs of Primary Care in Europe (QUALICOPC). We included 198 Norwegian GPs and 1529 patients. The patients completed a survey concerning experiences in a consultation with a GP on the inclusion day. The GPs completed a survey regarding organizational aspects of their own practice. Main outcome measures were seven statements concerning how the patients experienced the communication with the GP during the consultation. A generalized estimating equation logistic regression model was used to identify variations in patient experiences associated with characteristics of the GPs and their practices. The patients reported overall positive experiences with their GP consultations. Patients who consulted a GP with a short patient list were less likely than patients who consulted a GP with a medium sized list to regard the GP as polite (Odds Ratio (OR) 0.2; 95 % CI 0.1-0.7), to report that the GP asked questions about their health problems (OR 0.6; 0.4-1.0) or that the GP used sufficient time (OR 0.5; CI 0.3-0.9). Patients who consulted a GP with a long patient list compared to patients who consulted a GP with a medium sized list were less likely to feel that they could cope better after the GP visit (OR 0.5; 0.3-0.9) and more likely to feel that the GP hardly looked at them while talking (OR 1.8; 1.0-3.0). No associations with patient experiences were found with the average duration of the consultations, whether the GP worked in a fee-for-service model or whether the GP was the patient's regular doctor. Norwegian patients report predominantly positive experiences when consulting a GP. Positive communication experiences are most likely to be reported when the GP has a medium sized patient list.


The 4D nucleome project.
The 4D Nucleome Network aims to develop and apply approaches to map the structure and dynamics of the human and mouse genomes in space and time with the goal of gaining deeper mechanistic insights into how the nucleus is organized and functions. The project will develop and benchmark experimental and computational approaches for measuring genome conformation and nuclear organization, and investigate how these contribute to gene regulation and other genome functions. Validated experimental technologies will be combined with biophysical approaches to generate quantitative models of spatial genome organization in different biological states, both in cell populations and in single cells.


Influenza-associated mortality in 2009 in four sentinel sites in Bangladesh.
To estimate influenza-associated mortality in Bangladesh in 2009. In four hospitals in Bangladesh, respiratory samples were collected twice a month throughout 2009 from inpatients aged < 5 years with severe pneumonia and from older inpatients with severe acute respiratory infection. The samples were tested for influenza virus ribonucleic acid (RNA) using polymerase chain reaction. The deaths in 2009 in five randomly selected unions (the smallest administrative units in Bangladesh) in each hospital's catchment area were then investigated using formal records and informal group discussions. The deaths of those who had reportedly died within 14 days of suddenly developing fever with cough and/or a sore throat were assumed to be influenza-associated. The rate of such deaths in 2009 in each of the catchment areas was then estimated from the number of apparently influenza-associated deaths in the sampled unions, the proportion of the sampled inpatients in the local hospital who tested positive for influenza virus RNA, and the estimated number of residents of the sampled unions. Of the 2500 people known to have died in 2009 in all 20 study unions, 346 (14%) reportedly had fever with cough and/or sore throat within 14 days of their deaths. The estimated mean annual influenza-associated mortality in these unions was 11 per 100,000 population: 1.5, 4.0 and 125 deaths per 100,000 among those aged < 5, 5-59 and > 59 years, respectively. The highest burden of influenza-associated mortality in Bangladesh in 2009 was among the elderly.


Tbx1 represses Mef2c gene expression and is correlated with histone 3 deacetylation of the anterior heart field enhancer.
The TBX1 gene is haploinsufficient in 22q11.2 deletion syndrome (22q11.2DS), and genetic evidence from human patients and mouse models points to a major role of this gene in the pathogenesis of this syndrome. Tbx1 can activate and repress transcription, and previous work has shown that one of its functions is to negatively modulate cardiomyocyte differentiation. Tbx1 occupies the anterior heart field (AHF) enhancer of the Mef2c gene, which encodes a key cardiac differentiation transcription factor. Here, we show that increased dosage of Tbx1 correlates with downregulation of Mef2c expression and reduced acetylation of its AHF enhancer in cultured mouse myoblasts. Consistently, 22q11.2DS-derived and in vitro-differentiated human induced pluripotent stem cells (hiPSCs) expressed higher levels of MEF2C and showed increased AHF acetylation, compared with hiPSCs from a healthy donor. Most importantly, we show that in mouse embryos, loss of Tbx1 enhances the expression of the Mef2c-AHF-Cre transgene in a specific region of the splanchnic mesoderm, and in a dosage-dependent manner, providing an in vivo correlate of our cell culture data. These results indicate that Tbx1 regulates the Mef2c AHF enhancer by inducing histone deacetylation.


Development of a Freshness Assay for Royal Jelly Based on the Temperature- and Time-Dependent Changes of Antimicrobial Effectiveness and Proteome Dynamics of Royal Jelly Proteins.
Although the antimicrobial, nutritional, and health-promoting properties of royal jelly (RJ) have been widely confirmed, the effects of storage temperature and time on RJ quality remain to be further explored. Herein, the antimicrobial and proteomic dynamics of RJ stored under different conditions were comprehensively investigated to identify consistent and sensitive markers of RJ degradation. We confirmed the negative correlation between antimicrobial properties and increased the storage temperature and duration in RJ. Using surface plasmon resonance, we showed the protein degradation-induced conformation changes in RJ, which reflected the overall variation in RJ proteins caused by the storage conditions. Further proteomic and western blotting analyses demonstrated the sensitivity and reliability of major RJ protein 4 (MRJP4) as a measure of temperature- and time-dependent RJ changes. Based on these results, we developed a colloidal gold immunoassay strip for MRJP4 detection, providing a reliable, simple, and rapid method for the evaluation of RJ freshness.


Infant vitamin d supplementation and allergic conditions in adulthood: northern Finland birth cohort 1966.
Allergen-induced secretion of Th2-type cytokines and IgE production have recently been reported to be increased in mice treated with 1,25(OH)(2)D, the active form of vitamin D. Our objective was to investigate whether vitamin D supplementation in infancy is associated with the risk of atopy, allergic rhinitis, and asthma. The Northern Finland Birth Cohort consists of all individuals in the two most northern provinces of Finland who were due to be born in 1966. Data on vitamin D supplementation during the first year of life was obtained in 1967. Current asthma and allergic rhinitis were reported at age 31 years (n = 7,648), and atopy determined by skin-prick test in a sub-sample still living in northern Finland or the Helsinki area (n = 5,007). The prevalence of atopy and allergic rhinitis at age 31 years was higher in participants who had received vitamin D supplementation regularly during the first year compared to others (OR 1.46, 95%CI 1.4-2.0, and OR 1.66, 95%CI 1.1-1.6, respectively). A similar association was observed for asthma (OR 1.35, 95%CI 0.99-1.8). These associations persisted after adjustment for a wide range of behavioral and social factors (adjusted: OR 1.33 for all, P = 0.01 for atopy, P = 0.001 for allergic rhinitis, and P = 0.08 for asthma). We observed an association between vitamin D supplementation in infancy and an increased risk of atopy and allergic rhinitis later in life. Further study is required to determine whether these observations reflect long-term effects on immune regulation or differences in unmeasured determinants of vitamin D supplementation.


Molecular and Serological Survey of the Cat-Scratch Disease Agent (Bartonella henselae) in Free-Ranging Leopardus geoffroyi and Leopardus wiedii (Carnivora: Felidae) From Pampa Biome, Brazil.
The genus Bartonella comprises emerging bacteria that affect humans and other mammals worldwide. Felids represent an important reservoir for several Bartonella species. Domestic cats are the main reservoir of Bartonella henselae, the agent of cat scratch disease (CSD). It can be transmitted directly by scratches and bites from infected cats and via cat fleas. This study aims to investigate the circulation of Bartonella spp. in free-ranging Neotropical wild felids from Southern Brazil using serological and molecular methods. In this study, 53 live-trapped free-ranging wild felids were sampled, 39 Leopardus geoffroyi and 14 Leopardus wiedii, from five municipalities in the Rio Grande, do Sul state, southern Brazil. All captured animals were clinically healthy. Two blood samples of L. geoffroyi were positive, by PCR, for the presence of B. henselae DNA. Conversely, none of L. wiedii blood samples were positive when tested using PCR. Indirect immunofluorescence assay (IFA) showed that 28% of serum samples of wild felids were reactive (seropositive) for B. henselae by immunofluorescence, with titers ranging from 64 to 256. The results presented here provide the first evidence of a Bartonella-enzootic cycle involving L. geoffroyi and L. wiedii, which may account for the spillover of the emerging zoonotic pathogen B. henselae for the indigenous fauna in Southern Brazil.


Heterotrimeric G proteins demonstrate differential sensitivity to beta-arrestin dependent desensitization.
G15 is a heterotrimeric G protein of the Gq/11 family. In this study, we describe its exceptional poor sensitivity to the general regulatory mechanism of G protein-coupled receptor (GPCR) desensitization. Enhancing beta2 adrenergic receptor desensitization by arrestin overexpression, did not affect signalling to G15. Similarly, increased levels of arrestin did not affect G15 signalling triggered by the activation of V2 vasopressin and delta opioid receptors. Furthermore, co-immunoprecipitation experiments showed that G15 alpha subunit (as opposed to Galphaq and Galphas) is recruited to a V2 vasopressin receptor mutant that is constitutively desensitized by beta-arrestin. Interestingly, co-expression of Galpha15 partially rescued cell surface localization and signalling capabilities of the same mutant receptor and reduced beta2 adrenergic receptor internalization. Taken together, these findings provide evidence for a novel mechanism whereby GPCR desensitization can be bypassed and G15 can support sustained signalling in cells chronically exposed to hormones or neurotransmitters.


Inn1 and Cyk3 regulate chitin synthase during cytokinesis in budding yeasts.
The chitin synthase that makes the primary septum during cell division in budding yeasts is an important therapeutic target with an unknown activation mechanism. We previously found that the C2-domain of the Saccharomyces cerevisiae Inn1 protein plays an essential but uncharacterised role at the cleavage site during cytokinesis. By combining a novel degron allele of INN1 with a point mutation in the C2-domain, we screened for mutations in other genes that suppress the resulting defect in cell division. In this way, we identified 22 dominant mutations of CHS2 (chitin synthase II) that map to two neighbouring sites in the catalytic domain. Chs2 in isolated cell membranes is normally nearly inactive (unless protease treatment is used to bypass inhibition); however, the dominant suppressor allele Chs2-V377I has enhanced activity in vitro. We show that Inn1 associates with Chs2 in yeast cell extracts. It also interacts in a yeast two-hybrid assay with the N-terminal 65% of Chs2, which contains the catalytic domain. In addition to compensating for mutations in the Inn1 C2-domain, the dominant CHS2 alleles suppress cytokinesis defects produced by the lack of the Cyk3 protein. Our data support a model in which the C2-domain of Inn1 acts in conjunction with Cyk3 to regulate the catalytic domain of Chs2 during cytokinesis. These findings suggest novel approaches for developing future drugs against important fungal pathogens.


Telehealth use in Australian primary healthcare during COVID-19: a cross-sectional descriptive survey.
This study aimed to investigate Australian primary healthcare professionals' experiences of the rapid upscaling of telehealth during COVID-19. A cross-sectional survey. Two hundred and seventeen general practitioners, nurses and allied health professionals employed in primary healthcare settings across Australia were recruited via social media and professional organisations. An online survey was disseminated between December 2020 and March 2021. The survey comprised items about individual demographics, experiences of delivering telehealth consultations, perceived quality of telehealth consultations and future perceptions of telehealth. Telephone was the most widely used method of providing telehealth, with less than 50% of participants using a combination of telephone and video. Key barriers to telehealth use related to the inability to undertake physical examination or physical intervention. Telehealth was perceived to improve access to healthcare for some vulnerable groups and those living in rural settings, but reduced access for people from non-English-speaking backgrounds. Quality of telehealth care was considered mostly or somewhat the same as care provided face-to-face, with actual or perceived negative outcomes related to missed or delayed diagnosis. Overwhelmingly, participants wanted telehealth to continue with guaranteed ongoing funding. Some 43.7% of participants identified the need to further improve telehealth models of care. The rapid shift to telehealth has facilitated ongoing care during the COVID-19 pandemic. However, further work is required to better understand how telehealth can be best harnessed to add value to service delivery in usual care.


DNA polymerase II is encoded by the DNA damage-inducible dinA gene of Escherichia coli.
The structural gene for DNA polymerase II was cloned by using a synthetic inosine-containing oligonucleotide probe corresponding to 11 amino acids, which were determined by sequencing the amino terminus of the purified protein. The labeled oligonucleotide hybridized specifically to the lambda clone 7H9 from the Kohara collection as well as to plasmid pGW511 containing the SOS-regulated dinA gene. Approximately 1400 base pairs of dinA sequence were determined. The predicted amino-terminal sequence of dinA demonstrated that this gene encoded DNA polymerase II. Sequence analysis of the upstream region localized a LexA binding site overlapping the -35 region of the dinA promoter, and this promoter element was found to be only two nucleotides downstream from the 3' end of the araD gene. These results demonstrate that the gene order is thr-dinA (pol II)-ara-leu on the Escherichia coli chromosome and that the DNA polymerase II structural gene is transcribed in the same direction as the araBAD operon. Based on the analysis of the predicted protein, we have identified a sequence motif Asp-Xaa-Xaa-Ser-Leu-Tyr-Pro-Ser in DNA polymerase II that is highly conserved among a diverse group of DNA polymerases, which include those from humans, yeast, Herpes and vaccinia viruses, and phages T4 and PRD1. The demonstration that DNA polymerase II is a component of the SOS response in E. coli suggests that it plays an important role in DNA repair and/or mutagenesis.


Weight Change Nomograms for the First Month After Birth.
Clinicians expect newborns to surpass birth weight by age 10 to 14 days, yet few studies have examined the natural history of weight change in the weeks after birth. We sought to determine the distribution of weight loss and subsequent regain during the first month, the proportion not surpassing birth weight by 14 and 21 days, and whether findings differed by delivery mode. For 161 471 singleton neonates delivered at ≥36 weeks' gestation at Kaiser Permanente Northern California Medical Centers between 2009 and 2013 and weighing 2000 to 5000 g at birth, we extracted daily weights from inpatient electronic records and weights from outpatient visits in the first month. Quantile regression appropriate for repeated measures was used to estimate percentiles of weight change as a function of time after birth, stratified by delivery mode. After exclusions, weight data were analyzed from 143 889 newborns (76% born vaginally). Based on percentile estimates, 50% of newborns were at or above birth weight at 9 and 10 days after vaginal and cesarean delivery, respectively. Among those delivered vaginally, 14% and 5% were not back to birth weight by 14 and 21 days, respectively. For those delivered by cesarean, 24% and 8% were not back to birth weight by 14 and 21 days, respectively. It is not uncommon for newborns to be below birth weight 10 to 14 days after delivery. A larger percentage of newborns delivered by cesarean had yet to regain birth weight at every time point through 1 month.


Deficient neurogenesis in forebrain-specific presenilin-1 knockout mice is associated with reduced clearance of hippocampal memory traces.
To examine the in vivo function of presenilin-1 (PS1), we selectively deleted the PS1 gene in excitatory neurons of the adult mouse forebrain. These conditional knockout mice were viable and grew normally, but they exhibited a pronounced deficiency in enrichment-induced neurogenesis in the dentate gyrus. This reduction in neurogenesis did not result in appreciable learning deficits, indicating that addition of new neurons is not required for memory formation. However, our postlearning enrichment experiments lead us to postulate that adult dentate neurogenesis may play a role in the periodic clearance of outdated hippocampal memory traces after cortical memory consolidation, thereby ensuring that the hippocampus is continuously available to process new memories. A chronic, abnormal clearance process in the hippocampus may conceivably lead to memory disorders in the mammalian brain.


Blood and alveolar lymphocyte subsets in pulmonary cytomegalovirus infection after lung transplantation.
Cytomegalovirus (CMV) pneumonitis has been shown to be associated with lymphocytic alveolitis after lung transplantation. In the present study, we investigated a series of bronchoalveolar (BAL) and blood samples, collected in the absence of rejection or acute infectious episodes. in order -1: to evaluate intra-alveolar cell population changes concomitant with CMV replication and -2: to reappraise the value of cell population analysis in the management of patients after lung transplantation. We used flow cytometry to investigate modifications of lymphocyte subpopulations related to pulmonary cytomegalovirus infections in blood and BAL samples from a series of 13 lung transplant recipients. After exclusion of samples obtained during pulmonary rejection, bronchiolitis obliterans or acute bacterial infection, 48 blood and BAL samples were retained for analysis: 17 were CMV positive by shell-vial assay and 31 were CMV negative in blood and BAL. Our results demonstrate that pulmonary CMV infection is associated with a significant increase in the total lymphocyte population in BAL samples, but with minor modifications of the various lymphocyte subpopulations and a significantly higher absolute number of B lymphocytes in blood samples. Cytomegalovirus pulmonary infection is accompanied by only minor changes in BAL lymphocyte subpopulations. The study of BAL lymphocyte subpopulations therefore appears to be of limited clinical value in the diagnosis of pulmonary CMV infection. However, increased blood B-lymphocytes seems to be a clinical feature associated with CMV infection.


Migfilin and its binding partners: from cell biology to human diseases.
Links between the plasma membrane and the actin cytoskeleton are essential for maintaining tissue integrity and for controlling cell morphology and behavior. Studies over the past several decades have identified dozens of components of such junctions. One of the most recently identified is migfilin, a widely expressed protein consisting of an N-terminal filamin-binding domain, a central proline-rich domain and three C-terminal LIM domains. Migfilin is recruited to cell-matrix contacts in response to adhesion and colocalizes with beta-catenin at cell-cell junctions in epithelial and endothelial cells. Migfilin also travels from the cytoplasm into the nucleus, a process that is regulated by RNA splicing and calcium signaling. Through interactions with multiple binding partners, including Mig-2, filamin and VASP, migfilin links the cell adhesion structures to the actin cytoskeleton. It regulates actin remodeling, cell morphology and motility. In nuclei, migfilin interacts with the cardiac transcriptional factor CSX/NKX2-5 and promotes cardiomyocyte differentiation. It probably functions as a key regulator both at cell adhesion sites and nuclei, coordinating multiple cellular processes, and is implicated in the pathogenesis of several human diseases.


Comparative effectiveness and harms of intravitreal antivascular endothelial growth factor agents for three retinal conditions: a systematic review and meta-analysis.
Intravitreal antivascular endothelial growth factor (VEGF) agents are widely used to treat ocular conditions but the benefits and harms of these treatments are uncertain. We conducted a systematic review to compare the effects of aflibercept, bevacizumab and ranibizumab on best-corrected visual acuity (BCVA) changes, quality of life and ocular or systemic adverse events in patients with neovascular age-related macular degeneration (NVAMD), diabetic macular oedema (DME) and central or branch retinal vein occlusion (RVO). We searched published and unpublished literature sources to February 2017 for randomised controlled trials and cohort or modelling studies reporting comparative costs in the USA. Two reviewers extracted data and graded the strength of the evidence using established methods. Of 17 included trials, none reported a clinically important difference (≥ 5 letters) in visual acuity gains between agents. Nine trials provide high-strength evidence of no difference between bevacizumab and ranibizumab for NVAMD. Three trials provide moderate-strength evidence of no difference between bevacizumab and ranibizumab for DME. There was low-strength evidence of similar effects between aflibercept and ranibizumab for NVAMD, aflibercept and bevacizumab for RVO and all three agents for DME. There was insufficient evidence to compare bevacizumab and ranibizumab for RVO. Rates of ocular adverse events were low, and systemic harms were generally similar between groups, although 1 DME trial reported more arterial thrombotic events with ranibizumab versus aflibercept. Overall, no agent had a clear advantage over another for effectiveness or safety. Aflibercept and ranibizumab were significantly less cost-effective than repackaged bevacizumab in two trials. Systematic review registration number: CRD42016034076.


Ethnic inequalities in health-related quality of life among older adults in England: secondary analysis of a national cross-sectional survey.
The population of older adults (ie, those aged ≥55 years) in England is becoming increasingly ethnically diverse. Previous reports indicate that ethnic inequalities in health exist among older adults, but information is limited by the paucity of data from small minority ethnic groups. This study aimed to analyse inequalities in health-related quality of life (HRQoL) and five determinants of health in older adults across all ethnic groups in England. In this cross-sectional study, we analysed data from five waves (July 1, 2014, to April 7, 2017) of the nationally representative English General Practice Patient Survey (GPPS). Study participants were adults aged 55 years or older who were registered with general practices in England. We used regression models (age-adjusted and stratified by gender) to estimate the association between ethnicity and HRQoL, measured by use of the EQ-5D-5L index and its domains (mobility, self-care, usual activities, pain or discomfort, and anxiety or depression). We also estimated associations between ethnicity and five determinants of health (presence of long-term conditions or multimorbidity, experience of primary care, degree of support from local services, patient self-confidence in managing own health, and degree of area-level social deprivation). We examined robustness to differential handling of missing data, alternative EQ-5D-5L value sets, and differences in area-level social deprivation. There were 1 416 793 GPPS respondents aged 55 years and older. 1 394 361 (98·4%) respondents had complete data on ethnicity and gender and were included in our analysis. Of these, 152 710 (11·0%) self-identified as belonging to minority ethnic groups. HRQoL was worse for men or women, or both, in 15 (88·2%) of 17 minority ethnic groups than the White British ethnic group. In both men and women, inequalities were widest for Gypsy or Irish Traveller (linear regression coefficient -0·192 [95% CI -0·318 to -0·066] in men; -0·264 [-0·354 to -0·173] in women), Bangladeshi (-0·111 [-0·136 to -0·087] in men; -0·209 [-0·235 to -0·184] in women), Pakistani (-0·084 [-0·096 to -0·073] in men; -0·206 [-0·219 to -0·193] in women), and Arab (-0·061 [-0·086 to -0·035] in men; -0·145 [-0·180 to -0·110] in women) ethnic groups, with magnitudes generally greater for women than men. Differentials tended to be widest for the self-care EQ-5D-5L domain. Ethnic inequalities in HRQoL were accompanied by increased prevalence of long-term conditions or multimorbidity, poor experiences of primary care, insufficient support from local services, low patient self-confidence in managing their own health, and high area-level social deprivation, compared with the White British group. We found evidence of wide ethnic inequalities in HRQoL and five determinants of health for older adults in England. Outcomes varied between minority ethnic groups, highlighting heterogeneity in the direction and magnitude of associations. We recommend further research to understand the drivers of inequalities, together with policy changes to improve equity of socioeconomic opportunity and access to services for older adults from minority ethnic groups. University of Manchester and National Institute for Health Research.


Puerarin alleviates delayed-type hypersensitivity via cytokine inhibition by modulating Th1/Th2 balance.
Puerarin has long been used as a traditional Chinese medicine, which possesses various physiological properties, including anti-inflammation, anti-oxidative, anti-diabetic and anti-cancer activities. The aim of the current study was to investigate the effect of puerarin on delayed-type hypersensitivity (DTH) induced by ovalbumin (OVA) in mice and explore its underlying mechanisms. The results showed that puerarin significantly attenuated DTH, resulting from a decrement in footpad swelling, reduction in inflammatory cell as well as a decline in anti-OVA IgG in serum. In the homogenized supernatant of footpad tissues, the classic Th1-cytokines, including interferon (IFN)-γ was suppressed following puerarin treatment. Furthermore, a high dose of puerarin inhibited interleukin (IL)-4 production, the classic Th2-cytokine. The concanavalin A stimulation and MTT assays indicated a suppressive effect of puerarin on Th1 response via decreasing IFN-γ production in OVA-primed lymphocytes. Detailed studies revealed that puerarin modulated the Th1/Th2 balance in DTH responses, attributing to lower T-bet/GATA binding protein-3 mRNA and protein level ratios, which led to the shift change of IFN-γ/IL-4 with puerarin treatment. These findings demonstrate that puerarin alleviated inflammation in DTH triggered by OVA application via curbing inflammatory cytokines by modulating the Th1/Th2 balance. These results suggest that puerarin may be an alternative therapeutic option for the treatment of DTH.


Acute traumatic coagulopathy: pathophysiology and resuscitation.
Acute Traumatic Coagulopathy occurs immediately after massive trauma when shock, hypoperfusion, and vascular damage are present. Mechanisms for this acute coagulopathy include activation of protein C, endothelial glycocalyx disruption, depletion of fibrinogen, and platelet dysfunction. Hypothermia and acidaemia amplify the endogenous coagulopathy and often accompany trauma. These multifactorial processes lead to decreased clot strength, autoheparinization, and hyperfibrinolysis. Furthermore, the effects of aggressive crystalloid administration, haemodilution from inappropriate blood product transfusion, and prolonged surgical times may worsen clinical outcomes. We review normal coagulation using the cell-based model of haemostasis and the pathophysiology of acute traumatic coagulopathy. Developed trauma systems reduce mortality, highlighting critical goals for the trauma patient in different phases of care. Once patients reach a trauma hospital, certain triggers reliably indicate when they require massive transfusion and specialized trauma care. These triggers include base deficit, international normalized radio (INR), systolic arterial pressure, haemoglobin concentration, and temperature. Early identification for massive transfusion is critically important, as exsanguination in the first few hours of trauma is a leading cause of death. To combat derangements caused by massive haemorrhage, damage control resuscitation is a technique that addresses each antagonist to normal haemostasis. Components of damage control resuscitation include damage control surgery, permissive hypotension, limited crystalloid administration, haemostatic resuscitation, and correction of hyperfibrinolysis.


Integrative features of the yeast phosphoproteome and protein-protein interaction map.
Following recent advances in high-throughput mass spectrometry (MS)-based proteomics, the numbers of identified phosphoproteins and their phosphosites have greatly increased in a wide variety of organisms. Although a critical role of phosphorylation is control of protein signaling, our understanding of the phosphoproteome remains limited. Here, we report unexpected, large-scale connections revealed between the phosphoproteome and protein interactome by integrative data-mining of yeast multi-omics data. First, new phosphoproteome data on yeast cells were obtained by MS-based proteomics and unified with publicly available yeast phosphoproteome data. This revealed that nearly 60% of ∼6,000 yeast genes encode phosphoproteins. We mapped these unified phosphoproteome data on a yeast protein-protein interaction (PPI) network with other yeast multi-omics datasets containing information about proteome abundance, proteome disorders, literature-derived signaling reactomes, and in vitro substratomes of kinases. In the phospho-PPI, phosphoproteins had more interacting partners than nonphosphoproteins, implying that a large fraction of intracellular protein interaction patterns (including those of protein complex formation) is affected by reversible and alternative phosphorylation reactions. Although highly abundant or unstructured proteins have a high chance of both interacting with other proteins and being phosphorylated within cells, the difference between the number counts of interacting partners of phosphoproteins and nonphosphoproteins was significant independently of protein abundance and disorder level. Moreover, analysis of the phospho-PPI and yeast signaling reactome data suggested that co-phosphorylation of interacting proteins by single kinases is common within cells. These multi-omics analyses illuminate how wide-ranging intracellular phosphorylation events and the diversity of physical protein interactions are largely affected by each other.


Parental transmission of type 2 diabetes: the Framingham Offspring Study.
Study of parental transmission of diabetes provides insight into the relative contributions of underlying maternal and paternal influences. We estimated risk for type 2 diabetes and milder degrees of glucose intolerance associated with parental diabetes among subjects of the population-based Framingham Offspring Study, in which participants are primarily Caucasian and at relatively low risk for diabetes and for which both parental and offspring phenotypes were ascertained by direct examination. Parental diabetes, assessed over 40 years of biennial follow-up, was defined by use of hypoglycemic drug therapy or a casual plasma glucose level > or = 11.1 mmol/l at any examination. Offspring glucose tolerance, assessed over 20 years of quadrennial follow-up, was defined by fasting plasma glucose levels > or = 7.8 mmol/l at any two examinations, use of hypoglycemic drug therapy at any examination, or with a 75-g oral glucose tolerance test (1980 World Health Organization criteria) at the most recent examination. We calculated odds ratios (ORs) and 95% CIs for offspring glucose tolerance status using generalized estimating equations to account for differential correlations within and between families. The 2,527 offspring came from 1,303 nuclear families, of which 77.6% had two or more siblings per family and in which the prevalence of parental diabetes was 24.6%. The mean offspring age was 54 years (range 26-82), 53% were women, 8.6% had diabetes, 11.4% had impaired glucose tolerance, 76.3% had no parental diabetes, 10.5% had maternal diabetes, 11.5% had paternal diabetes, and 1.7% had bilineal diabetes. Relative to individuals without parental diabetes, the age-adjusted ORs (95% CI) for offspring type 2 diabetes or abnormal glucose tolerance (fasting plasma glucose > or = 6.1 mmol/l or 2-h postchallenge glucose tolerance > or = 7.8 mmol/l) among individuals with maternal diabetes were 3.4 (2.3-4.9) and 2.7 (2.0-3.7), respectively; among individuals with paternal diabetes were 3.5 (2.3-5.2) and 1.7 (1.2-2.4), respectively; and among individuals with bilineal diabetes were 6.1 (2.9-13.0) and 5.2 (2.6-10.5), respectively. Although maternal and paternal diabetes conferred equivalent risk for offspring type 2 diabetes, offspring with maternal diabetes were slightly more likely to have abnormal glucose tolerance compared with those with paternal diabetes (OR 1.6, 95% CI 1.1-2.4). Offspring with maternal diabetes and an age of onset of <50 years had marked increased risk for both type 2 diabetes (9.7, 4.3-22.0) and abnormal glucose tolerance (9.0, 4.2-19.7). We conclude that risk ratios for offspring type 2 diabetes are consistent with a simple additive risk model, where risk when both parents are affected equals the sum of risk when either parent is affected. For maternal diabetes to confer excess risk for mild but not overt glucose intolerance, offspring of diabetic fathers may transit abnormal to impaired glucose tolerance relatively quickly, or diabetic mothers may transmit risk for a mild slowly progressive form of abnormal glucose tolerance in addition to overt diabetes. Very high risk for abnormal glucose homeostasis among offspring with young age-of-onset maternal diabetes is consistent with hypotheses that perinatal exposures increase diabetes risk. Given equivalent risk ratios for type 2 diabetes, fathers may transmit unique paternal genetic factors of similar strength to maternal environmental factors.


Genome-wide SNP and haplotype analyses reveal a rich history underlying dog domestication.
Advances in genome technology have facilitated a new understanding of the historical and genetic processes crucial to rapid phenotypic evolution under domestication. To understand the process of dog diversification better, we conducted an extensive genome-wide survey of more than 48,000 single nucleotide polymorphisms in dogs and their wild progenitor, the grey wolf. Here we show that dog breeds share a higher proportion of multi-locus haplotypes unique to grey wolves from the Middle East, indicating that they are a dominant source of genetic diversity for dogs rather than wolves from east Asia, as suggested by mitochondrial DNA sequence data. Furthermore, we find a surprising correspondence between genetic and phenotypic/functional breed groupings but there are exceptions that suggest phenotypic diversification depended in part on the repeated crossing of individuals with novel phenotypes. Our results show that Middle Eastern wolves were a critical source of genome diversity, although interbreeding with local wolf populations clearly occurred elsewhere in the early history of specific lineages. More recently, the evolution of modern dog breeds seems to have been an iterative process that drew on a limited genetic toolkit to create remarkable phenotypic diversity.


Radioembolization for hepatocellular carcinoma.
Radioembolization is a form of brachytherapy in which intra-arterially injected (90)Y-loaded microspheres serve as sources for internal radiation purposes. It produces average disease control rates above 80% and is usually very well tolerated. Main complications do not result from the microembolic effect, even in patients with portal vein occlusion, but rather from an excessive irradiation of non-target tissues including the liver. All the evidence that support the use of radioembolization in HCC is based on retrospective series or non-controlled prospective studies. However, reliable data can be obtained from the literature, particularly since the recent publication of large series accounting for nearly 700 patients. When compared to the standard of care for the intermediate and advanced stages (transarterial embolization and sorafenib), radioembolization consistently provides similar survival rates. Two indications seem particularly appealing in the boundaries of these stages for first-line radioembolization. First, the treatment of patients straddling between the intermediate and advanced stages (intermediate patients with bulky or bilobar disease that are considered poor candidates for TACE, and advanced patients with solitary tumors invading a segmental or lobar branch of the portal vein). Second, the treatment of patients that are slightly above the criteria for resection, ablation or transplantation, for which downstaging could open the door for a radical approach. Radioembolization can also be used to treat patients progressing to TACE or sorafenib. With a number of clinical trials underway, the available evidence shows that it adds a significant value to the therapeutic weaponry against HCC of tertiary care centers dealing with this major cancer problem.


Follicular lymphoma of the thyroid and the role of core needle biopsy.
Primary thyroid lymphoma (PTL) is a rare malignancy, accounting for less than 5% of all thyroid neoplasms. The follicular subtype is even more rare, accounting for approximately 10% of all PTL cases. We report a case of a 64-year-old woman, who presented with a rapidly growing goitre with mass effect and B symptoms. She had a history of Hashimoto's thyroiditis and her thyroid ultrasound revealed diffuse goitre with a dominant nodule (56 × 63 × 60 mm) within the right thyroid lobe. Ultrasound-guided percutaneous fine-needle aspiration of the right thyroid nodule was classified as benign, according to Bethesda System, with lymphocytic thyroiditis. A CT scan of the neck showed diffuse enlargement of the thyroid gland extending towards the anterior mediastinum with tracheal deviation and lymphadenopathy within levels VII and right II-IV. The core needle biopsy of the right thyroid nodule revealed a follicular non-Hodgkin's B cell lymphoma with a Ki67 of 60%. According to the Ann Arbor staging system, she was at stage IIIE. She underwent chemotherapy with R-CHOP (rituximab, cyclophosphamide, doxorubicin, vincristine, prednisone) with remarkable clinical improvement and is currently in remission 2 years after the diagnosis. PTL is an extremely rare malignancy that usually arises in a lymphocytic thyroiditis background, presenting as a rapidly enlarging goitre, which can lead to compressive symptoms or airway comprise. Primary thyroid lymphoma (PTL) is a rare malignancy, accounting for less than 5% of thyroid neoplasms. PTL should be suspected when a patient presents with a rapidly enlarging goitre, especially in the setting of Hashimoto's thyroiditis. Fine-needle aspiration has a limited capacity for PTL diagnosis due to similar cytomorphological features of lymphoma with thyroiditis. Therefore, in case of clinical suspicion and if fine needle aspiration fails to diagnose PTL, a tissue biopsy should be performed. Treatment is dependent on both the stage and histology of PTL. Chemotherapy and local radiotherapy remain the mainstay treatment for PTL.


Functionalized nanoporous silica for the removal of heavy metals from biological systems: adsorption and application.
Surface-functionalized nanoporous silica, often referred to as self-assembled monolayers on mesoporous supports (SAMMS), has previously demonstrated the ability to serve as very effective heavy metal sorbents in a range of aquatic and environmental systems, suggesting that they may be advantageously utilized for biomedical applications such as chelation therapy. Herein we evaluate surface chemistries for heavy metal capture from biological fluids, various facets of the materials' biocompatibility, and the suitability of these materials as potential therapeutics. Of the materials tested, thiol-functionalized SAMMS proved most capable of removing selected heavy metals from biological solutions (i.e., blood, urine, etc.) Consequentially, thiol-functionalized SAMMS was further analyzed to assess the material's performance under a number of different biologically relevant conditions (i.e., variable pH and ionic strength) to gauge any potentially negative effects resulting from interaction with the sorbent, such as cellular toxicity or the removal of essential minerals. Additionally, cellular uptake studies demonstrated no cell membrane permeation by the silica-based materials generally highlighting their ability to remain cellularly inert and thus nontoxic. The results show that organic ligand functionalized nanoporous silica could be a valuable material for a range of detoxification therapies and potentially other biomedical applications.


Experiments in computing: a survey.
Experiments play a central role in science. The role of experiments in computing is, however, unclear. Questions about the relevance of experiments in computing attracted little attention until the 1980s. As the discipline then saw a push towards experimental computer science, a variety of technically, theoretically, and empirically oriented views on experiments emerged. As a consequence of those debates, today's computing fields use experiments and experiment terminology in a variety of ways. This paper analyzes experimentation debates in computing. It presents five ways in which debaters have conceptualized experiments in computing: feasibility experiment, trial experiment, field experiment, comparison experiment, and controlled experiment. This paper has three aims: to clarify experiment terminology in computing; to contribute to disciplinary self-understanding of computing; and, due to computing's centrality in other fields, to promote understanding of experiments in modern science in general.


Epidemiological data from the COVID-19 outbreak, real-time case information.
Cases of a novel coronavirus were first reported in Wuhan, Hubei province, China, in December 2019 and have since spread across the world. Epidemiological studies have indicated human-to-human transmission in China and elsewhere. To aid the analysis and tracking of the COVID-19 epidemic we collected and curated individual-level data from national, provincial, and municipal health reports, as well as additional information from online reports. All data are geo-coded and, where available, include symptoms, key dates (date of onset, admission, and confirmation), and travel history. The generation of detailed, real-time, and robust data for emerging disease outbreaks is important and can help to generate robust evidence that will support and inform public health decision making.


Isolation of the Schizosaccharomyces pombe RAD54 homologue, rhp54+, a gene involved in the repair of radiation damage and replication fidelity.
The RAD54 gene of Saccharomyces cerevisiae encodes a putative helicase, which is involved in the recombinational repair of DNA damage. The RAD54 homologue of the fission yeast Schizosaccharomyces pombe, rhp54+, was isolated by using the RAD54 gene as a heterologous probe. The gene is predicted to encode a protein of 852 amino acids. The overall homology between the mutual proteins of the two species is 67% with 51% identical amino acids and 16% similar amino acids. A rhp54 deletion mutant is very sensitive to both ionizing radiation and UV. Fluorescence microscopy of the rhp54 mutant cells revealed that a large portion of the cells are elongated and occasionally contain aberrant nuclei. In addition, FACS analysis showed an increased DNA content in comparison with wild-type cells. Through a minichromosome-loss assay it was shown that the rhp54 deletion mutant has a very high level of chromosome loss. Furthermore, the rhp54 mutation in either a rad17 or a cdc2.3w mutant background (where the S-phase/mitosis checkpoint is absent) shows a significant reduction in viability. It is hypothesized that the rhp54+ gene is involved in the recombinational repair of UV and X-ray damage and plays a role in the processing of replication-specific lesions.


EnGens: a computational framework for generation and analysis of representative protein conformational ensembles.
Proteins are dynamic macromolecules that perform vital functions in cells. A protein structure determines its function, but this structure is not static, as proteins change their conformation to achieve various functions. Understanding the conformational landscapes of proteins is essential to understand their mechanism of action. Sets of carefully chosen conformations can summarize such complex landscapes and provide better insights into protein function than single conformations. We refer to these sets as representative conformational ensembles. Recent advances in computational methods have led to an increase in number of available structural datasets spanning conformational landscapes. However, extracting representative conformational ensembles from such datasets is not an easy task and many methods have been developed to tackle it. Our new approach, EnGens (short for ensemble generation), collects these methods into a unified framework for generating and analyzing protein conformational ensembles. In this work we: (1) provide an overview of existing methods and tools for protein structural ensemble generation and analysis; (2) unify existing approaches in an open-source Python package, and a portable Docker image, providing interactive visualizations within a Jupyter Notebook pipeline; (3) test our pipeline on a few canonical examples found in the literature. Representative ensembles produced by EnGens can be used for many downstream tasks such as protein-ligand ensemble docking, Markov state modeling of protein dynamics and analysis of the effect of single-point mutations.


Association between changes in eating and drinking behaviors and respiratory tract disease in newly arrived calves at a feedlot.
To investigate eating and drinking behaviors and their association with bovine respiratory disease complex (BRDC) and to evaluate methods of diagnosing BRDC. 170 newly arrived calves at a feedlot. Eating and drinking behaviors of calves were recorded at a feedlot. Calves with clinical signs of BRDC were removed from their pen and classified retrospectively as sick or not sick on the basis of results of physical and hematologic examinations. Pulmonary lesions of all calves were assessed at slaughter. Calves that were sick had significantly greater frequency and duration of drinking 4 to 5 days after arrival than calves that were not sick. Sick calves had significantly lower frequency and duration of eating and drinking 11 to 27 days after arrival but had significantly greater frequency of eating 28 to 57 days after arrival than calves that were not sick. Calves at slaughter that had a higher percentage of lung tissue with pneumonic lesions had significantly lower frequency and duration of eating 11 to 27 days after arrival but had significantly higher frequency and duration of eating 28 to 57 days after arrival. Agreement for calves being sick and having severe pulmonary lesions at slaughter was adequate. Agreement for calves being removed and having pulmonary lesions at slaughter was low. Eating and drinking behaviors were associated with signs of BRDC, but there was not an obvious predictive association between signs of BRDC in calves and eating and drinking behaviors. Fair to poor agreement was observed between antemortem and postmortem disease classification.


Binding of human serum albumin to PEGylated liposomes: insights into binding numbers and dynamics by fluorescence correlation spectroscopy.
Liposomes for medical applications are often administered by intravenous injection. Once in the bloodstream, the liposomes are covered with a "protein corona", which impacts the behavior and eventual fate of the liposomes. Currently, many aspects of the liposomal protein corona are not well understood. For example, there is generally a lack of knowledge about the liposome binding affinities and dynamics of common types of blood plasma proteins. Fluorescence correlation spectroscopy (FCS) is a powerful experimental technique that potentially can provide such knowledge. In this study, we have used FCS to investigate the binding of human serum albumin (HSA) to standard types of PEGylated fluid-phase liposomes (consisting of DOPC and DOPE-PEG2k) and PEGylated gel-phase liposomes (consisting of DSPC and DSPE-PEG2k) with various PEG chain surface densities. We detected no significant binding of HSA to the PEGylated fluid-phase liposomes. In contrast, we found that HSA bound tightly to the PEGylated gel-phase liposomes, although only a low number of HSA molecules could be accommodated per liposome. Overall, we believe that our data provides a useful benchmark for other researchers interested in studying the liposomal protein corona.


Determination of the structural parameters of an incommensurate phase from first principles: the case of Sc-II.
We propose a procedure to accurately describe the structural parameters of an incommensurate phase using ab initio methods by approximating it with a set of analogous commensurate supercells. We apply this approach to obtain the structural parameters of the Sc-II phase, which has recently been identified as a complex incommensurate structure similar to Sr-V. The calculated incommensurate ratio gamma, lattice parameters, and Wyckoff positions of Sc-II are in excellent agreement with the available experimental data. Our results show that gamma increases with pressure up to 60 GPa approaching but never reaching the commensurate value 4/3. Hence calculations do not confirm the prediction made based on the reanalyzing of experimental data. When pressure exceeds 70 GPa, gamma shows a sharp decrease that might be considered as the precursor of a new structural phase transition.


Associations of Insomnia Symptoms With Cognition in Persons With Heart Failure.
Although cognitive impairment is common among persons with heart failure and negatively impacts self-care, hospitalization, and mortality, the associations between cognitive impairment and insomnia symptoms are not clearly understood. The purpose of this study was to explore these associations and examine if they are maintained after adjusting for relevant sociodemographic, clinical, and lifestyle factors. Guided by the Neurocognitive model of insomnia and sleep and the self-care conceptual model, a cross-sectional data analysis using parametric testing was conducted on the Health and Retirement Study wave 2016. Difficulty initiating sleep and early morning awakening, but not difficulty maintaining sleep were significantly associated with poorer cognitive performance in the bivariate and multivariate analysis. Our results are suggestive of different phenotypes of insomnia symptoms that may have different associations with cognition in persons with heart failure. Further research using objective measurements of insomnia symptoms and detailed neuropsychiatric testing of cognition is needed to confirm this conclusion.


Proteomics and Metabolomics for AKI Diagnosis.
Acute kidney injury (AKI) is a severe and frequent condition in hospitalized patients. Currently, no efficient therapy of AKI is available. Therefore, efforts focus on early prevention and potentially early initiation of renal replacement therapy to improve the outcome in AKI. The detection of AKI in hospitalized patients implies the need for early, accurate, robust, and easily accessible biomarkers of AKI evolution and outcome prediction because only a narrow window exists to implement the earlier-described measures. Even more challenging is the multifactorial origin of AKI and the fact that the changes of molecular expression induced by AKI are difficult to distinguish from those of the diseases associated or causing AKI as shock or sepsis. During the past decade, a considerable number of protein biomarkers for AKI have been described and we expect from recent advances in the field of omics technologies that this number will increase further in the future and be extended to other sorts of biomolecules, such as RNAs, lipids, and metabolites. However, most of these biomarkers are poorly defined by their AKI-associated molecular context. In this review, we describe the state-of-the-art tissue and biofluid proteomic and metabolomic technologies and new bioinformatics approaches for proteomic and metabolomic pathway and molecular interaction analysis. In the second part of the review, we focus on AKI-associated proteomic and metabolomic biomarkers and briefly outline their pathophysiological context in AKI.


Diagnostic test accuracy of self-reported screening instruments in identifying frailty in community-dwelling older people: A systematic review.
Against a backdrop of aging populations worldwide, it has become increasingly important to identify frailty screening instruments suitable for community settings. Self-reported and/or administered instruments might offer significant simplicity and efficiency advantages over clinician-administered instruments, but their comparative diagnostic test accuracy has yet to be systematically examined. The aim of this systematic review was to determine the diagnostic test accuracy of self-reported and/or self-administered frailty screening instruments against two widely accepted frailty reference standards (the frailty phenotype and the Frailty Index) within community-dwelling older adult populations. We carried out a systematic search of the Embase, CINAHL, MEDLINE, PubMed, Web of Science, PEDro, PsycINFO, ProQuest Dissertations, Open Grey and GreyLit databases up to April 2017 (with an updated search carried out over May-July 2018) to identify studies reporting comparison of self-reported and/or self-administered frailty screening instruments against an appropriate reference standard, with a minimum sensitivity threshold of 80% and specificity threshold of 60%. We identified 24 studies that met our selection criteria. Four self-reported screening instruments across three studies met minimum sensitivity and specificity thresholds. However, in most cases, study design considerations limited the reliability and generalizability of the results. Additionally, meta-analysis was not carried out, because no more than three studies were available for any of the unique combinations of index tests and reference standards. Although the present study has shown that a number of self-reported frailty screening instruments reported sensitivity and specificity within a desirable range for community application, additional diagnostic test accuracy studies are required. Geriatr Gerontol Int 2020; 20: 14-24.


Reoperation for hyperparathyroidism in multiple endocrine neoplasia type 1.
Patients with multiple endocrine neoplasia type 1 and hyperparathyroidism often undergo multiple operations because of inadequate initial surgery, presence of supernumerary and ectopic glands, regrowth of remnant glands, or autograft hyperfunction. Management of this patient population is complex. From January 1975 to December 2000 we performed 94 reoperative parathyroidectomies consisting of 79 neck reexplorations, 12 autograft removals, and 3 median sternotomies in 75 patients. Data were gathered by retrospective chart review and follow-up telephone interviews. Excluding autograft excision, reoperative surgery was successful (normocalcemia longer than 6 months) in 91%; autograft removal was successful in only 58%. With a median follow-up of 59 months, 64% of patients are currently free from hypercalcemia, and this outcome was not influenced by the total number of glands resected. The median time to recurrent hypercalcemia was 125 months. Thirty patients received an autograft after reoperation. The complication rate for all reoperations was 12%, including permanent recurrent laryngeal nerve injury in 2 patients (2.1%). Reoperative parathyroidectomy in patients with multiple endocrine neoplasia type 1 was safe and successful in the majority of patients; however, recurrent hyperparathyroidism is likely to develop in most individuals beyond 10 years of follow-up. The total number of glands accounted for after reoperation is not associated with successful outcome.


Estrogen receptor alpha-negative and progesterone receptor-positive breast cancer: lab error or real entity?
Aretrospective study comparing the estrogen receptor (ER) alpha subtype and progesterone receptor (PR) profile of breast carcinomas amongst 1625 cases over 2.5 years was carried out. Strictly speaking it is generally believed that breast carcinomas can biochemically express PR only if they are ER-positive. However, a few ERalpha-PR+ cases do exist paradoxically. This class of tumors was the focus of our study in which we looked at the possible reasons for such an immunophenotype and compared it with a group of ERalpha+PR+ breast carcinomas. An internationally recognized immunohistochemical method employing monoclonal antibodies against estrogen and progesterone receptors was used. Correlations with established risk factors i.e. menopausal status, grade, tumor size and lymph node status were analyzed for our study group (ERalpha-PR+) and compared with a control (ERalpha+PR+). Out of the total 1625 cases, 29.91% (486) were ERalpha+PR+, 5.11% (83) were ERalpha+PR-, 56.86% (924) were ERalpha-PR- and 8.12% (132) were ERalpha-PR+. Patients' age was significantly lower in the ERalpha-PR+ group (P=0.002). Statistical analysis of the grading between the two study groups revealed no significant difference (P=0.091), although the ERalpha-PR+ group contained significantly more poorly differentiated tumors than the ERalpha+PR+ one (P=0.032). Tumor size was also significantly larger in the ERalpha-PR+ than in the ERalpha+PR+ group (P=0.046). The frequency of lymph node metastases was independent of receptor profile. In conclusion, our study group does exhibit characteristics which are suggestive of a distinct breast cancer phenotype (ERalpha-PR+) with a different etiology and prognosis.


Low bone mass may not be the only cause of skeletal fragility in osteoporosis.
Skeletal fragility in postmenopausal osteoporosis is not due solely to reduction in bone mass. This fact explains some of the overlap in bone mineral measurements observed between patients who are fracturing and age- and sex-matched normals who are not. Changes in skeletal architecture and bone remodeling occur with age which can account for some of the fragility. These changes are exaggerated in patients with postmenopausal osteoporosis who are suffering spine fractures. Three abnormalities have been described by histomorphometric methods which can account for skeletal fragility out of proportion to the degree of bone loss. They are: (i) loss of trabecular connectivity such that vertical weight-bearing bars lose their cross-attachments with each other, thus becoming susceptible to buckling; (ii) inefficient and prolonged microdamage repair due to periods of pause in the formation phase of remodeling; and (iii) accumulation of unrepaired microdamage in unremodeled bone tissue in the central part of trabeculae due to reduced osteon wall thickness coupled with maintenance of trabecular thickness. Recognition of these abnormalities should broaden our approach to the study of skeletal fragility in the syndrome of postmenopausal osteoporosis.


Using developmental trajectories to examine verbal and visuospatial short-term memory development in children and adolescents with Williams and Down syndromes.
Williams (WS) and Down (DS) syndromes have been associated with specifically compromised short-term memory (STM) subsystems. Individuals with WS have shown impairments in visuospatial STM, while individuals with DS have often shown problems with the recall of verbal material. However, studies have not usually compared the development of STM skills in these domains, in these populations. The present study employed a cross-sectional developmental trajectories approach, plotting verbal and visuospatial STM performance against more general cognitive and chronological development, to investigate how the domain-specific skills of individuals with WS and DS may change as development progresses, as well as whether the difference between STM skill domains increases, in either group, as development progresses. Typically developing children, of broadly similar cognitive ability to the clinical groups, were also included. Planned between- and within-group comparisons were carried out. Individuals with WS and DS both showed the domain-specific STM weaknesses in overall performance that were expected based on the respective cognitive profiles. However, skills in both groups developed, according to general cognitive development, at similar rates to those of the TD group. In addition, no significant developmental divergence between STM domains was observed in either clinical group according to mental age or chronological age, although the general pattern of findings indicated that the influence of the latter variable across STM domains, particularly in WS, might merit further investigation.


In vitro selection of oligonucleotides that bind double-stranded DNA in the presence of triplex-stabilizing agents.
A SELEX approach has been developed in order to select oligonucleotides that bind double-stranded DNA in the presence of a triplex-stabilizing agent, and was applied to a target sequence containing an oligopurine-oligopyrimidine stretch. After only seven rounds of selection, the process led to the identification of oligonucleotides that were able to form triple helices within the antiparallel motif. Inspection of the selected sequences revealed that, contrary to GC base pair which were always recognized by guanines, recognition of AT base pair could be achieved by either adenine or thymine, depending on the sequence context. While thymines are strongly preferred for several positions, some others can accommodate the presence of adenines. These results contribute to set the rules for designing oligonucleotides that form stable triple helices in the presence of triplex-stabilizing agents at physiological pH. They set the basis for further experiments regarding extension of potential target sequences for triple-helix formation or recognition of ligand-DNA complexes.


A global plastid phylogeny of the fern genus Asplenium (Aspleniaceae).
The infrageneric relationships and taxonomy of the largest fern genus, Asplenium (Aspleniaceae), have remained poorly understood. Previous studies have focused mainly on specific species complexes involving a few or dozens of species only, or have achieved a large taxon sampling but only one plastid marker was used. In the present study, DNA sequences from six plastid markers (atpB, rbcL, rps4, rps4-trnS, trnL and trnL-F) of 1030 accessions (616 of them newly sequenced here) representing c. 420 species of Asplenium (60% of estimated species diversity), 16 species of Hymenasplenium, three Diplaziopsidaceae, and four Rhachidosoraceae were used to produce the largest genus-level phylogeny yet for ferns. Our major results include: (i) Asplenium as broadly circumscribed is monophyletic based on our inclusion of representatives of 32 of 38 named segregate genera; (ii) 11 major clades in Asplenium are identified, and their relationships are mostly well-resolved and strongly supported; (iii) numerous species, unsampled in previous studies, suggest new relationships and numerous cryptic species and species complexes in Asplenium; and (iv) the accrued molecular evidence provides an essential foundation for further investigations of complex patterns of geographical diversification, speciation and reticulate evolution in this family.


High-Level dCas9 Expression Induces Abnormal Cell Morphology in Escherichia coli.
Along with functional advances in the use of CRISPR/Cas9 for genome editing, endonuclease-deficient Cas9 (dCas9) has provided a versatile molecular tool for exploring gene functions. In principle, differences in cell phenotypes that result from the RNA-guided modulation of transcription levels by dCas9 are critical for inferring with gene function; however, the effect of intracellular dCas9 expression on bacterial morphology has not been systematically elucidated. Here, we observed unexpected morphological changes in Escherichia coli mediated by dCas9, which were then characterized using RNA sequencing (RNA-Seq) and chromatin immunoprecipitation sequencing (ChIP-Seq). Growth rates were severely decreased, to approximately 50% of those of wild type cells, depending on the expression levels of dCas9. Cell shape was changed to abnormal filamentous morphology, indicating that dCas9 affects bacterial cell division. RNA-Seq revealed that 574 genes were differentially transcribed in the presence of high expression levels of dCas9. Genes associated with cell division were upregulated, which was consistent with the observed atypical morphologies. In contrast, 221 genes were downregulated, and these mostly encoded proteins located in the cell membrane. Further, ChIP-Seq results showed that dCas9 directly binds upstream of 37 genes without single-guide RNA, including fimA, which encodes bacterial fimbriae. These results support the fact that dCas9 has critical effects on cell division as well as inner and outer membrane structure. Thus, to precisely understand gene functions using dCas9-driven transcriptional modulation, the regulation of intracellular levels of dCas9 is pivotal to avoid unexpected morphological changes in E. coli.


An Automated Detection System of Drug-Drug Interactions from Electronic Patient Records Using Big Data Analytics.
The aim of the study was to build a proof-of-concept demonstratrating that big data technology could improve drug safety monitoring in a hospital and could help pharmacovigilance professionals to make data-driven targeted hypotheses on adverse drug events (ADEs) due to drug-drug interactions (DDI). We developed a DDI automatic detection system based on treatment data and laboratory tests from the electronic health records stored in the clinical data warehouse of Rennes academic hospital. We also used OrientDb, a graph database to store informations from five drug knowledge databases and Spark to perform analysis of potential interactions betweens drugs taken by hospitalized patients. Then, we developed a machine learning model to identify the patients in whom an ADE might have occurred because of a DDI. The DDI detection system worked efficiently and computation time was manageable. The system could be routinely employed for monitoring.


Hepatoprotection by the farnesoid X receptor agonist GW4064 in rat models of intra- and extrahepatic cholestasis.
Farnesoid X receptor (FXR) is a bile acid-activated transcription factor that is a member of the nuclear hormone receptor superfamily. Fxr-null mice exhibit a phenotype similar to Byler disease, an inherited cholestatic liver disorder. In the liver, activation of FXR induces transcription of transporter genes involved in promoting bile acid clearance and represses genes involved in bile acid biosynthesis. We investigated whether the synthetic FXR agonist GW4064 could protect against cholestatic liver damage in rat models of extrahepatic and intrahepatic cholestasis. In the bile duct-ligation and alpha-naphthylisothiocyanate models of cholestasis, GW4064 treatment resulted in significant reductions in serum alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase, as well as other markers of liver damage. Rats that received GW4064 treatment also had decreased incidence and extent of necrosis, decreased inflammatory cell infiltration, and decreased bile duct proliferation. Analysis of gene expression in livers from GW4064-treated cholestatic rats revealed decreased expression of bile acid biosynthetic genes and increased expression of genes involved in bile acid transport, including the phospholipid flippase MDR2. The hepatoprotection seen in these animal models by the synthetic FXR agonist suggests FXR agonists may be useful in the treatment of cholestatic liver disease.


Novel variants of CD44 arising from alternative splicing: changes in the CD44 alternative splicing pattern of MCF-7 breast carcinoma cells treated with hyaluronidase.
CD44 is a cell-surface glycoprotein postulated to play a role in a variety of biological processes, including lymphocyte homing and tumor-cell metastasis. Several isoforms of CD44 have been identified in human cells, and the genesis of some of these isoforms has been attributed to alternative splicing. In the study presented here we amplified three novel transcript variants of CD44 from human cell lines using a reverse transcriptase-polymerase chain reaction strategy. Two of the novel isoforms differed from previously described CD44 isoforms as a result of alternative splicing that occurred at previously reported splice junctions. The third novel CD44 isoform was generated from a previously unreported alternative splice junction near the 5' end of the open reading frame. Southern blot analysis of genomic DNA revealed that these novel isoforms and all of the previously described CD44 isoforms arose from alternative splicing. The capability of cells to modify their CD44 alternative splicing pattern was demonstrated in MCF-7 cells, which altered their CD44-isoform expression pattern in response to treatment with hyaluronidase. A better understanding of mechanisms regulating CD44 alternative splicing may provide insights into diverse processes, including tumor-cell metastasis and lymphocyte homing.


Pulse wave amplitude is associated with brachial artery diameter: implications for gender differences in microvascular function.
The ratio of pulse wave amplitude (PWA) during reactive hyperemia compared to baseline as measured by peripheral arterial tonometry (PAT) is a non-invasive measure of microvascular endothelial function referred to as the pulse wave amplitude reactive hyperemia index (PWA-RHI). Whether upstream conduit vessel structure may affect downstream resistance vessel PWA has not been clearly examined. We tested the hypothesis that digital PWA is influenced by brachial artery diameter (BAD) and that this association would influence comparison of PWA-RHI between genders. Measures of vascular structure and microvascular function were carried out in 115 patients varying in cardiovascular risk profiles (average age 57 years, male n = 79, CAD n = 43). PWA was assessed using plethysmography at baseline and following 5 minutes of brachial artery occlusion. BAD was assessed using high-resolution ultrasonography. Results : There was a negative association between BAD and PWA-RHI ( r = -0.34, p < 0.05). Women had greater PWA-RHI and smaller BAD compared with men (p < 0.05). When co-varying for BAD, there were no longer gender differences in PWA-RHI. Moreover, when a sub-group of men and women without CAD (n = 40), matched for BAD, were examined, there were no gender differences in PWA-RHI. In conclusion, PWA-RHI obtained from PAT is associated with BAD. Studies examining gender differences in microvascular endothelial function with PAT may need to correct for BAD as a potential confounder.


Sequence variations in the human Kir6.2 gene, a subunit of the beta-cell ATP-sensitive K-channel: no association with NIDDM in while Caucasian subjects or evidence of abnormal function when expressed in vitro.
The ATP-sensitive K-channel plays a central role in insulin release from pancreatic beta cells. This channel consists of two subunits: a sulphonyl-urea receptor, SUR1, and an inwardly rectifying K-channel subunit, Kir6.2. We screened 135 white Caucasian patients with non-insulin-dependent diabetes mellitus (NIDDM) and 90 non-diabetic subjects for mutations in the Kir6.2 gene by single-stranded conformational polymorphism (SSCP) analysis. We identified one silent mutation (A190A) and four missense mutations (E23K, L270V, I337V and S385C) in normal and diabetic individuals. In a single diabetic subject, we identified a two-amino acid insertion (380KP). We also screened 39 Afro-Caribbean diabetic subjects and identified one additional missense (L355P) and one more silent (S363S) mutation. The E23K and I337V variants were completely linked. The common variants (E23K, 1337V and L270V) were found with similar frequency in diabetic and normal subjects. Diabetic subjects with the variants responded normally to sulphonylurea therapy. When mutant Kir6.2 subunits were coexpressed with SUR1 in Xenopus oocytes, there was no difference in the sensitivity of the whole-cell currents to metabolic inhibition or to the sulphonylurea tolbutamide. We therefore conclude that mutations in Kir6.2 are unlikely to be a major cause of NIDDM.