Contreraslyhne0402

Direct reprogramming, also known as a trans-differentiation, is a technique to allow mature cells to be converted into other types of cells without inducing a pluripotent stage. It has been suggested as a major strategy to acquire the desired type of cells in cell-based therapies to repair damaged tissues. Studies related to switching the fate of cells through epigenetic modification have been progressing and they can bypass safety issues raised by the virus-based transfection methods. In this study, a protocol was established to directly convert fully differentiated fibroblasts into diverse mesenchymal-lineage cells, such as osteoblasts, adipocytes, chondrocytes, and ectodermal cells, including neurons, by means of DNA demethylation, immediately followed by culturing in various differentiating media. First, 24 h exposure of 5-azacytidine (5-aza-CN), a well-characterized DNA methyl transferase inhibitor, to NIH-3T3 murine fibroblast cells induced the expression of stem-cell markers, that is, increasing cell plasticity. Next, 5-aza-CN treated fibroblasts were cultured in osteogenic, adipogenic, chondrogenic, and neurogenic media with or without bone morphogenetic protein 2 for a designated period. Differentiation of each desired type of cell was verified by quantitative reverse transcriptase-polymerase chain reaction/ western blot assays for appropriate marker expression and by various staining methods, such as alkaline phosphatase/alizarin red S/oil red O/alcian blue. These proposed procedures allowed easier acquisition of the desired cells without any transgenic modification, using direct reprogramming technology, and thus may help make it more available in the clinical fields of regenerative medicine.Chronic joint pain due to loss of cartilage function, degradation of subchondral bone, and related conditions are common plights of an arthritis patient. Antioxidant compounds could solve the problems in arthritic condition. The objective of this study was to evaluate the anti-arthritic activity of D-carvone against complete Freund's adjuvant (CFA)-induced arthritis in rats. D-carvone was orally administered for 25 days at the doses of 30 and 60 mg/kg against CFA-induced arthritic rats. Changes in body weight, paw swelling, organ index, hematological parameters, oxidative stress markers, inflammatory cytokines, and histopathology were recorded. Oral treatment of D-carvone significantly improved the body weight, reduced the paw swelling, edema formation, and organ index in arthritic rats. The levels of white blood cells were reduced, red blood cells and hemoglobin levels were improved in D-carvone treated arthritic rats. Lipid peroxidation levels were lowered whereas enzymatic and non-enzymatic antioxidants were significantly elevated by D-carvone administration against arthritic rats. D-carvone significantly modulated inflammatory cytokine levels and improved the ankle joint pathology against CFA-induced arthritic inflammation. In conclusion, D-carvone proved significant anti-arthritic activity against CFA-induced arthritis in rats.In vivo animal models are limited in their ability to mimic the extremely complex systems of the human body, and there is increasing disquiet about the ethics of animal research. Many authorities in different geographical areas are considering implementing a ban on animal testing, including testing for cosmetics and pharmaceuticals. Therefore, there is a need for research into systems that can replicate the responses of laboratory animals and simulate environments similar to the human body in a laboratory. An in vitro two-dimensional cell culture model is widely used, because such a system is relatively inexpensive, easy to implement, and can gather considerable amounts of reference data. However, these models lack a real physiological extracellular environment. Recent advances in stem cell biology, tissue engineering, and microfabrication techniques have facilitated the development of various 3D cell culture models. These include multicellular spheroids, organoids, and organs-on-chips, each of which has its own advantages and limitations. Organoids are organ-specific cell clusters created by aggregating cells derived from pluripotent, adult, and cancer stem cells. Patient-derived organoids can be used as models of human disease in a culture dish. Biomimetic organ chips are models that replicate the physiological and mechanical functions of human organs. Many organoids and organ-on-a-chips have been developed for drug screening and testing, so competition for patents between countries is also intensifying. We analyzed the scientific and technological trends underlying these cutting-edge models, which are developed for use as non-animal models for testing safety and efficacy at the nonclinical stages of drug development.The mitogen-activated protein kinase (MAPK) pathway controls intestinal epithelial barrier permeability by regulating tight junctions (TJs) and epithelial cells damage. Heme oxygenase-1 (HO-1) and carbon monoxide (CO) protect the intestinal epithelial barrier function, but the molecular mechanism is not yet clarified. MAPK activation and barrier permeability were studied using monolayers of Caco-2 cells treated with tissue necrosis factor α (TNF-α) transfected with FUGW-HO-1 or pLKO.1-sh-HO-1 plasmid. Intestinal mucosal barrier permeability and MAPK activation were also investigated using carbon tetrachloride (CCl4) administration with CoPP (a HO-1 inducer), ZnPP (a HO-1 inhibitor), CO releasing molecule 2 (CORM-2), or inactived-CORM-2-treated wild-type mice and mice with HO-1 deficiency in intestinal epithelial cells. TNF-α increased epithelial TJ disruption and cleaved caspase-3 expression, induced ERK, p38, and JNK phosphorylation. In addition, HO-1 blocked TNF-α-induced increase in epithelial TJs disruption, cleaved caspase-3 expression, as well as ERK, p38, and JNK phosphorylation in an HO-1-dependent manner. CoPP and CORM-2 directly ameliorated intestinal mucosal injury, attenuated TJ disruption and cleaved caspase-3 expression, and inhibited epithelial ERK, p38, and JNK phosphorylation after chronic CCl4 injection. Conversely, ZnPP completely reversed these effects. Furthermore, mice with intestinal epithelial HO-1 deficient exhibited a robust increase in mucosal TJs disruption, cleaved caspase-3 expression, and MAPKs activation as compared to the control group mice. These data demonstrated that HO-1-dependent MAPK signaling inhibition preserves the intestinal mucosal barrier integrity by abrogating TJ dysregulation and epithelial cell damage. The differential targeting of gut HO-1-MAPK axis leads to improved intestinal disease therapy.Synthetic cathinones have become popular drugs of abuse. We describe our recent experience with two highly agitated patients following ingestion of the cathinone derivative chloromethcathinone, and cannabis. Both patients suffered from excited delirium syndromes that lasted for over 24 hours. Clinicians should be aware of this phenomenon, especially since routine toxicology screenings do not detect the presence of these agents.Pituitary apoplexy is an infrequent but life-threatening complication of pituitary adenomas. When apoplexy occurs in a hormonally active adenoma, this may induce spontaneous remission of the clinical syndrome. In these cases, clinical suspicion of Cushing's disease or acromegaly may arise at presentation, but due to spontaneous remission of active hormone production, it is not possible to biochemically confirm this diagnosis in retrospect. Resolution of clinical symptoms during follow up retrospectively suggests the diagnosis. However, we describe a patient with Cushing's disease presenting with pituitary apoplexy, who was biochemically in remission at presentation. The diagnosis could be confirmed in retrospect using hair cortisol analysis, thereby enabling clinicians to adequately anticipate remission of Cushing's disease.Bruton's tyrosine kinase (BTK) inhibitors are increasingly used in untreated and previously treated chronic lymphocytic leukaemia (CLL) patients. Invasive fungal infections (IFI) were rarely observed in patients treated for CLL in the pre-BTK era. In this article, we describe two patients with CLL who developed an IFI during treatment with the BTK inhibitor ibrutinib. The atypical presentation and the serious course of this complication are described.The syndrome of inappropriate antidiuretic hormone secretion (SIADH) is a known cause of hyponatremia, caused by excessive ADH secretion which, in turn, leads to water retention. SIADH has been associated with multiple etiologies, one of which is traumatic brain injury (TBI). Most cases of SIADH after TBI describe a course in which hyponatraemia develops several days to weeks after the trauma and then resolves within a few weeks. We demonstrate a case of SIADH after TBI, which persisted several years after initial presentation, but eventually did resolve spontaneously after five years.This case report describes a patient who presented with a debilitating hepatitis C virus-related cryoglobulinaemic vasculitis who was treated with immunosuppression and direct-acting antivirals. After returning symptoms revealed a relapse of the hepatitis C virus infection, treatment with direct-acting antivirals was repeated. Subsequently, he achieved a sustained virological response and his vasculitis subsided.Haemophagocytic lymphohistiocytosis (HLH) is a rare hyperinflammatory condition that can be triggered by infections, malignancies, or auto-immune diseases. Here, we present a patient with rapidly progressive HLH triggered by a herpes simplex virus type 2 (HSV-2) primary infection. The patient was successfully treated with intravenous high-dose acyclovir, immunoglobulins, and dexamethasone. This is the first report of HSV-2-associated HLH in an immunocompetent adult patient.

Chemotherapy (CT)-induced neutropenia and febrile neutropenia (FN) can lead to changes in the treatment plan, potentially worsening the cancer outcome. This study evaluated the effect of the glycopegylated granulocyte-colony stimulating factor lipegfilgrastim, used as primary (PP) or secondary prophylaxis (SP), on treatment modifications in adult patients receiving cytotoxic CT with or without biological/targeted therapy (BT) for solid and haematological tumours.

This phase 4, prospective, observational study was conducted in eight centres in the Netherlands, in 2015-2017. Other study objectives were to characterise the population of cancer patients receiving lipegfilgrastim, to evaluate the incidence of CT-induced neutropenic events, and to assess safety.

Of 142 patients, 73.94% had breast cancer and 55.63% received CT in the adjuvant setting. Most patients received lipegfilgrastim as PP (74.65%) and were at low (34.51%) or high risk (39.44%) of FN. CT dose delays were recorded for 22.64% and 36.11% of patients receiving lipegfilgrastim for PP and SP, respectively. CT dose reductions were recorded for 2.11% of patients; no CT dose omissions and one BT dose omission occurred. FN and grade III/IV neutropenia were reported for 5.63% and 9.86% of patients, respectively; associated hospitalisations were rare. The most frequently lipegfilgrastimrelated adverse events (AE) were myalgia, bone pain, and back pain. Serious AEs (55) were reported for 30 (21.13%) patients. There were two deaths, unrelated to lipegfilgrastim administration.

Administration of lipegfilgrastim in routine clinical practice in the Netherlands results in limited CT/BT dose modifications and low incidence of neutropenic events, with no new safety concerns.

Administration of lipegfilgrastim in routine clinical practice in the Netherlands results in limited CT/BT dose modifications and low incidence of neutropenic events, with no new safety concerns.

The study aimed to look at alterations in expression of matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) and their potential use as biomarkers in the pathogensis of SLE.

SLE patients (n = 41) and healthy controls (n = 50) were recruited. Quantitative RT-PCR/ELISA assays were performed for expression of MMP and TIMP mRNA in whole blood and PBMC; and corresponding serum protein levels. Intracellular levels of MMP-2 and MMP-9 proteins were analysed by flow cytometry.

Based on SLEDAI scores patients were grouped into active (SLEDAI ≥ 10) and inactive cases (SLEDAI < 10). In active cases, MMP-2 expression significantly increased and TIMP-2 expression was decreased (p < 0.0001) both at serum secretion (p = 0.0003) and mRNA (p < 0.0001) levels as compared to inactive cases. MMP-9 and TIMP-1 showed significantly reduced serum secretion and mRNA expression (p < 0.0001) in active cases as compared to inactive cases. Intracellular concentration of MMP-9 was reported to be higher in neutrophils, while MMP-2 was mainly found in lymphocytes of SLE patients as compared to controls. MMP/TIMP ratio profile was altered as SLE disease progresses.

Findings suggest disturbed MMP and TIMP levels have a role in the pathogenesis of SLE.

Findings suggest disturbed MMP and TIMP levels have a role in the pathogenesis of SLE.

Organisation of the emergency department (ED) is gaining attention due to an increased demand on emergency services, leading to crowding and influencing the quality of care. It is known that the organisation of acute care influences the performance of the ED. In the Netherlands, the organisation of EDs differs between hospitals. However, detailed information about the various organisational structures is lacking. This study aims to determine the organisational structures and the different roles and responsibilities of internists and emergency physicians (EPs) in the EDs.

We performed a nationwide observational study between January 2018 and February 2019. All hospitals with an ED in the Netherlands were identified, contacted, and surveyed. Requested information was retrieved from internists and complemented with local administrative hospital data.

76 out of 89 EDs responded to the questionnaire (84%); 93% of EDs were operational 24/7. A registered acute internist was present at 47 locations (62%) and an EP at 60 EDs (79%). At 10 locations (13.2%), internists reported not being physically present at the ED. Supervision and working agreements between EPs and internists differed between the hospitals. Collaboration between EPs and internists was graded satisfactory (7.4/10).

This is the first study providing a detailed overview of the ED organisation in the Netherlands regarding internal medicine patients. This organisation differs in terms of staffing, presence of EPs and internists, and working agreements. The influence of the various organisational structures of EDs on quality of acute care should be the subject of future research.

This is the first study providing a detailed overview of the ED organisation in the Netherlands regarding internal medicine patients. This organisation differs in terms of staffing, presence of EPs and internists, and working agreements. The influence of the various organisational structures of EDs on quality of acute care should be the subject of future research.

Frailty screening in the emergency department may identify frail patients at risk for adverse outcomes. This study investigated if the Dutch Safety Management Program (VMS) screener predicts outcomes in older patients in the emergency department.

In this prospective cohort study, patients aged 70 years or older presenting to the emergency department were recruited on workdays between 1000 AM and 700 PM from May 2017 until August 2017. Patients were screened in four domains activities of daily living, malnutrition, risk of delirium, and risk of falling. After 90 days of follow up, mortality, functional decline, living situation, falls, readmission to the emergency department, and readmission to the hospital were recorded. VMS was studied using the total VMS score as a predictor with ROC curve analysis, and using a cut-off point to divide patients into frail and non-frail groups to calculate positive predictive value (PPV) and negative predictive value (NPV).

A total of 249 patients were included. Higher se medical outcomes. This could be useful to determine which patients should undergo additional screening.A 28-year-old female patient was admitted to our hospital with severe dyspnoea and hypoxemia due to methaemoglobinaemia caused by dapsone. The patient recovered completely after repeated infusions of methylene blue and cessation of dapsone. However, 12 days after cessation of dapsone, the patient was readmitted due to recurrence of symptoms based on a relapse of methaemoglobinaemia. Toxicological analysis revealed a toxic dapsone level at readmission and no other explanation for methaemoglobinaemia. Several possible mechanisms as explanation for the recurrence of methaemoglobinaemia are listed and additional tests were performed. In addition to supportive care, treatment consisted of methylene blue; furthermore, cimetidine and ascorbic acid were added. An overview of the pathophysiology, diagnostics, treatment, and possible explanations for this relapse of methaemoglobinaemia caused by dapsone are given. This case shows the importance of considering the possibility of a late rebound methaemoglobinaemia after discontinuation of dapsone.

Hypertension (HT) is a chronic condition associated with serious complications. In the present cross-sectional study, we aimed to analyse factors that contribute to blood pressure control in subjects with HT.

Subjects with HT admitted to outpatient internal medicine clinics of the institution were enrolled in the study. According to the Joint National Committee (JNC) VIII criteria, subjects with a mean blood pressure above target levels were defined as poorly-controlled hypertensive patients and others were grouped as well-controlled hypertensive patients. Clinical and laboratory parameters were compared between study groups.

Smokers were more prevalent in the poorly-controlled HT group compared to the well-controlled HT group (p = 0.001). The number of patients who adhered to dietary and exercise recommendations were greater in well-controlled HT group than poorly-controlled HT group (p < 0.001 for both). The rate of combined therapy was greater in well-controlled HT group compared to poorly-controlled HT group (p = 0.04).

We suggest that, in addition to dietary and exercise recommendations and smoking cessation, treatment with combination therapy could be better in reaching blood pressure targets in patients with HT.

We suggest that, in addition to dietary and exercise recommendations and smoking cessation, treatment with combination therapy could be better in reaching blood pressure targets in patients with HT.Clinical management of renal artery stenosis has seen a major shift, after randomised clinical trials have shown no group benefit of endovascular intervention relative to optimal medical control. However, the inclusion criteria of these trials have been criticised for focusing on a subset of patients with atherosclerotic renal artery stenosis where intervention was unlikely to be beneficial. Moreover, new imaging and computational techniques have become available, which have the potential to improve identification of patients that will respond to interventional treatment. This review addresses the challenges associated with clinical decision making in patients with renal artery stenosis. Opportunities for novel diagnostic techniques to improve patient selection are discussed, along with ongoing Dutch studies and network initiatives that investigate these strategies.A soluble form of the interleukin-2 receptor (sIL-2R) is secreted upon T-cell activation. Increased blood levels of sIL-2R occur in a variety of immunological diseases. Although the biological function of sIL-2R is incompletely understood, both in health and disease, sIL-2R serum measurements are commonly conducted in clinical practice as it may help to facilitate diagnosis of specific immune-mediated diseases, such as haemophagocytic lymphohistiocytosis and sarcoidosis. In these, and in other immune-diseases, sIL-2R levels may be used as a biomarker to monitor/predict disease activity and treatment response. In this review, we will give a brief overview of the biology of the IL-2/IL-2R system and will subsequently discuss the clinical utility of sIL-2R measurement, especially in the context of haemophagocytic lymphohistiocytosis, sarcoidosis, rheumatoid arthritis, systemic lupus erythematosus, juvenile idiopathic arthritis, adult-onset Still's disease, ANCA-associated vasculitis, and IgG4-related disease.

This study aimed to develop and validate a brief practitioner-friendly health literacy screening tool, called Rapid Estimate of Inadequate Health Literacy (REIHL), that estimates patients' health literacy inadequacy in demanding clinical settings.

This is a methodological study of 304 community-dwelling older adults recruited from one community health centre and five district elderly community centres. Logistic regression models were used to identify the coefficients of the REIHL score's significant factors. Receiver operating characteristic (ROC) curve analysis was then used to assess the REIHL's sensitivity and specificity. Path analysis was employed to examine the REIHL's criterion validity with the Chinese Health Literacy Scale for Chronic Care and concurrent validity with self-rated health scale and the Geriatric Depression Scale-15.

The REIHL has scores ranging from 0 to 23. It had 76.9% agreement with the Chinese Health Literacy Scale for Chronic Care. The area under the ROC curve for predicting health literacy inadequacy was 0.82 (95% confidence interval=0.78-0.87, P<0.001). The ROC curve of the REIHL showed that scores ≥11 had a sensitivity of 77.8% and specificity of 75.6% for predicting health literacy inadequacy. The path analysis model showed excellent fit (Chi squared [2, 304] 0.16, P=0.92, comparative fit index 1.00, root mean square error of approximation <0.001, 90% confidence interval=0.00-0.04), indicating that the REIHL has good criterion and concurrent validity.

The newly developed REIHL is a practical tool for estimating older adults' inadequate health literacy in clinical care settings.

The newly developed REIHL is a practical tool for estimating older adults' inadequate health literacy in clinical care settings.Cell polarity is essential for various asymmetric cellular events, and the partitioning defective (PAR) protein PAR3 (encoded by PARD3 in mammals) plays a unique role as a cellular landmark to establish polarity. In epithelial cells, PAR3 localizes at the subapical border, such as the tight junction in vertebrates, and functions as an apical determinant. Although we know a great deal about the regulators of PAR3 localization, how PAR3 is concentrated and localized to a specific membrane domain remains an important question to be clarified. In this study, we demonstrate that ASPP2 (also known as TP53BP2), which controls PAR3 localization, links PAR3 and protein phosphatase 1 (PP1). The ASPP2-PP1 complex dephosphorylates a novel phosphorylation site, Ser852, of PAR3. Furthermore, Ser852- or Ser889-unphosphorylatable PAR3 mutants form protein clusters, and ectopically localize to the lateral membrane. Concomitance of clustering and ectopic localization suggests that PAR3 localization is a consequence of local clustering. We also demonstrate that unphosphorylatable forms of PAR3 exhibited a low molecular turnover and failed to coordinate rapid reconstruction of the tight junction, supporting that both the phosphorylated and dephosphorylated states are essential for the functional integrity of PAR3.Accurate measurements of cell morphology and behaviour are fundamentally important for understanding how disease, molecules and drugs affect cell function in vivo Here, by using muscle stem cell (muSC) responses to injury in zebrafish as our biological paradigm, we established a 'ground truth' for muSC behaviour. This revealed that segmentation and tracking algorithms from commonly used programs are error-prone, leading us to develop a fast semi-automated image analysis pipeline that allows user-defined parameters for segmentation and correction of cell tracking. Cell Tracking Profiler (CTP) is a package that runs two existing programs, HK Means and Phagosight within the Icy image analysis suite, to enable user-managed cell tracking from 3D time-lapse datasets to provide measures of cell shape and movement. We demonstrate how CTP can be used to reveal changes to cell behaviour of muSCs in response to manipulation of the cell cytoskeleton by small-molecule inhibitors. CTP and the associated tools we have developed for analysis of outputs thus provide a powerful framework for analysing complex cell behaviour in vivo from 4D datasets that are not amenable to straightforward analysis.Endocytosis of caveolae has previously been implicated in the repair of plasma membrane wounds. Here, we show that caveolin-1-deficient fibroblasts lacking caveolae upregulate a tubular endocytic pathway and have a reduced capacity to reseal after permeabilization with pore-forming toxins compared with wild-type cells. Silencing endophilin-A2 expression inhibited fission of endocytic tubules and further reduced plasma membrane repair in cells lacking caveolin-1, supporting a role for tubular endocytosis as an alternative pathway for the removal of membrane lesions. Endophilin-A2 was visualized in association with cholera toxin B-containing endosomes and was recruited to recently formed intracellular vacuoles containing Trypanosoma cruzi, a parasite that utilizes the plasma membrane wounding repair pathway to invade host cells. Endophilin-A2 deficiency inhibited T. cruzi invasion, and fibroblasts deficient in both caveolin-1 and endophilin-A2 did not survive prolonged exposure to the parasites. These findings reveal a novel crosstalk between caveolin-1 and endophilin-A2 in the regulation of clathrin-independent endocytosis and plasma membrane repair, a process that is subverted by T. cruzi parasites for cell invasion.The function of microRNAs (miRNAs) can be cell autonomous or communicated to other cell types and has been implicated in diverse biological processes. We previously demonstrated that miR-517a-3p (miR-517a), a highly expressed member of the chromosome 19 miRNA cluster (C19MC) that is transcribed almost exclusively in human trophoblasts, attenuates viral replication via induction of autophagy in non-trophoblastic recipient cells. However, the molecular mechanisms underlying these effects remain unknown. Here, we identified unc-13 homolog D (UNC13D) as a direct, autophagy-related gene target of miR-517a, leading to repression of UNC13D. In line with the antiviral activity of miR-517a, silencing UNC13D suppressed replication of vesicular stomatitis virus (VSV), whereas overexpression of UNC13D increased VSV levels, suggesting a role for UNC13D silencing in the antiviral activity of miR-517a. We also found that miR-517a activated NF-κB signaling in HEK-293XL cells expressing TLR8, but the effect was not specific to C19MC miRNA. Taken together, our results define mechanistic pathways that link C19MC miRNA with inhibition of viral replication.

To explore associations between occupational exposure to four specific organic solvents, respectively, and female breast cancer, including subtypes.

Using the Danish Cancer Registry, we identified 38 375 women under age 70 years with primary breast cancer. Five randomly selected breast-cancer-free controls per case matched on year of birth were retrieved from the Danish Civil Registration System . A nationwide pension fund was used to retrieve full employment history, and exposure to 1,1,1-trichloroethane, trichloroethylene (TCE), benzene and toluene was assessed using a job exposure matrix. ORs were estimated using conditional logistic regression with adjustment for reproductive factors and socioeconomic status.

Overall results indicated no noteworthy associations between the specific organic solvents and breast cancer before and after age 50 years, except for a small increased risk after age 50 in women exposed to TCE (OR=1.15, 95% CI 0.97-1.36). After age 50 years, exposure to TCE was associated with a small increased risk in women with over 20 years of latency (OR=1.26, 95% CI 1.02-1.56). Further, an increased risk of oestrogen receptor positive (ER+) tumours was also observed (OR=1.21, 95% CI 1.01-1.47), and high cumulative exposure and longer latency also increased the risk of this subtype.

This study provides limited evidence supporting the association between occupational exposure to each of the four organic solvents and breast cancer. The risk of ER+ breast tumours after age 50 years may be increased in women with TCE exposure, and this possible association therefore needs further attention in future studies.

This study provides limited evidence supporting the association between occupational exposure to each of the four organic solvents and breast cancer. The risk of ER+ breast tumours after age 50 years may be increased in women with TCE exposure, and this possible association therefore needs further attention in future studies.Roux-en-Y gastric bypass (RYGB) is one of the most commonly performed weight-loss procedures, but how severe obesity and RYGB affects circulating HDL-associated microRNAs (miRNAs) remains unclear. Here, we aim to investigate how HDL-associated miRNAs are regulated in severe obesity and how weight loss after RYGB surgery affects HDL-miRNAs. Plasma HDL were isolated from patients with severe obesity (n=53) before, 6 and 12 months after RYGB by immunoprecipitation using goat anti-human apoA-I microbeads. HDL were also isolated from 18 healthy participants. miRNAs were extracted from isolated HDL and levels of miR-24, miR-126, miR-222 and miR-223 were determined by TaqMan miRNA assays. We found that HDL-associated miR-126, miR-222 and miR-223 levels, but not miR-24 levels, were significantly higher in patients with severe obesity when compared with healthy controls. There were significant increases in HDL-associated miR-24, miR-222 and miR-223 at 12 months after RYGB. Additionally, cholesterol efflux capacity and paraoxonase (PON1) activity were increased and intracellular adhesion molecule-1 (ICAM-1) levels decreased. The increases in HDL-associated miR-24 and miR-223 were positively correlated with increase in cholesterol efflux capacity (r=0.326, P=0.027 and r=0.349, P=0.017 respectively). An inverse correlation was observed between HDL-associated miR-223 and ICAM-1 at baseline. Together, these findings show that HDL-associated miRNAs are differentially regulated in healthy versus patients with severe obesity and are altered after RYGB. These findings provide insights into how miRNAs are regulated in obesity before and after weight reduction, and may lead to the development of novel treatment strategies for obesity and related metabolic disorders.In any given organism, approximately one-third of all proteins have a yet-unknown function. A widely distributed domain of unknown function is DUF1127. Approximately 17,000 proteins with such an arginine-rich domain are found in 4,000 bacteria. Most of them are single-domain proteins, and a large fraction qualifies as small proteins with fewer than 50 amino acids. We systematically identified and characterized the seven DUF1127 members of the plant pathogen Agrobacterium tumefaciens They all give rise to authentic proteins and are differentially expressed as shown at the RNA and protein levels. The seven proteins fall into two subclasses on the basis of their length, sequence, and reciprocal regulation by the LysR-type transcription factor LsrB. The absence of all three short DUF1127 proteins caused a striking phenotype in later growth phases and increased cell aggregation and biofilm formation. Protein profiling and transcriptome sequencing (RNA-seq) analysis of the wild type and triple mutant revealed a laracquisition, cellular metabolism, and biofilm formation. The study shows that small proteins have important previously missed biological functions. How small basic proteins can have such a broad impact is a fascinating prospect of future research.K+ and NO3- are the major forms of potassium and nitrogen that are absorbed by the roots of most terrestrial plants. In this study, we observed that a close relationship between NO3- and K+ in Arabidopsis (Arabidopsis thaliana) is mediated by NITRATE TRANSPORTER1.1 (NRT1.1). The nrt1.1 knockout mutants showed disturbed K+ uptake and root-to-shoot allocation, and were characterized by growth arrest under K+-limiting conditions. The K+ uptake and root-to-shoot allocation of these mutants were partially recovered by expressing NRT1.1 in the root epidermis-cortex and central vasculature using SULFATE TRANSPORTER1;2 and PHOSPHATE1 promoters, respectively. Two-way analysis of variance based on the K+ contents in nrt1.1-1/K + transporter1, nrt1.1-1/high-affinity K + transporter5-3, nrt1.1-1/K + uptake permease7, and nrt1.1-1/stelar K + outward rectifier-2 double mutants and the corresponding single mutants and wild-type plants revealed physiological interactions between NRT1.1 and K+ channels/transporters located in the root epidermis-cortex and central vasculature. Further study revealed that these K+ uptake-related interactions are dependent on an H+-consuming mechanism associated with the H+/NO3- symport mediated by NRT1.1. Collectively, these data indicate that patterns of NRT1.1 expression in the root epidermis-cortex and central vasculature are coordinated with K+ channels/transporters to improve K+ uptake and root-to-shoot allocation, respectively, which in turn ensures better growth under K+-limiting conditions.Light is indispensable for the anthocyanin accumulation of red pear (Pyrus pyrifolia). Anthocyanin biosynthesis is catalyzed by a series of enzymes encoded by structural genes, which are regulated by a MYB-basic/helix-loop-helix-WD repeat (MYB-bHLH-WDR [MBW]) complex. The bHLH proteins of subgroup (SG) IIIf are believed to be involved in the regulation of anthocyanin accumulation. In this study, we revealed that pear PpbHLH64, which belongs to SGIIIb, positively regulates anthocyanin biosynthesis and is regulated by light at the transcriptional and posttranslational levels. Specifically, an exposure to light induced PpbHLH64 expression and anthocyanin accumulation in pear fruit and calli. Under light conditions, pear calli overexpressing PpbHLH64 exhibited enhanced red coloration, whereas the anthocyanin accumulation decreased in the PpbHLH64-RNA interference calli. Additionally, the transient overexpression of PpbHLH64 in pear fruit peel increased anthocyanin accumulation, whereas the virus-induced gene silencing of PpbHLH64 had the opposite effect. Further analyses indicated that PpbHLH64 is a transcriptional activator that directly binds to the promoter of UDP-GLUCOSEFLAVONOID 3-O-GLYCOSYLTRANFERASE to upregulate expression. Moreover, PpbHLH64 interacted with PpMYB10, but not with PpMYB114, to form an MBW complex that significantly induces the accumulation of anthocyanins. Furthermore, PpbHLH64 was targeted by CONSTITUTIVE PHOTOMORPHOGENIC1 in darkness for subsequent degradation by the 26S proteasome. A genetic analysis indicated that PpbHLH64 functions downstream of B-BOX18, a component of the light signal transduction pathway. However, we were unable to detect the direct interaction between PpbHLH64 and PpBBX18. The characterization of PpbHLH64 in this study highlights the importance of SGIIIb bHLH proteins for light-induced anthocyanin accumulation.A "smart canopy" ideotype has been proposed with leaves being upright at the top and more horizontal toward the bottom of the plant to maximize light interception and conversion efficiencies, and thus increasing yield. The genetic control of leaf angle has, to date, been studied on one or two leaves, or data have been merged from multiple leaves to generate average values. This approach has limited our understanding of the diversity of leaf angles across layers and their genetic control. Genome-wide association studies and quantitative trait loci mapping studies in sorghum (Sorghum bicolor) were performed using layer-specific angle data collected manually and via high-throughput phenotyping strategies. The observed distribution of angles in indoor and field settings is opposite to the ideotype. Several genomic regions were associated with leaf angle within layers or across the canopy. The expression of the brassinosteroid-related transcription factor BZR1/BES1 and the auxin-transporter Dwarf3 were found to be highly correlated with the distribution of angles at different layers. The application of a brassinosteroid biosynthesis inhibitor could not revert the undesirable overall angle distribution. These discoveries demonstrate that the exploitation of layer-specific quantitative trait loci/genes will be instrumental to reversing the natural angle distribution in sorghum according to the "smart canopy" ideotype.Eukaryotic flagella undertake different beat types as necessary for different functions; for example, the Leishmania parasite flagellum undergoes a symmetric tip-to-base beat for forward swimming and an asymmetric base-to-tip beat to rotate the cell. In multi-ciliated tissues or organisms, the asymmetric beats are coordinated, leading to movement of the cell, organism or surrounding fluid. This coordination involves a polarisation of power stroke direction. Here, we asked whether the asymmetric beat of the single Leishmania flagellum also has a fixed polarisation. We developed high frame rate dual-colour fluorescence microscopy to visualise flagellar-associated structures in live swimming cells. This showed that the asymmetric Leishmania beat is polarised, with power strokes only occurring in one direction relative to the asymmetric flagellar machinery. Polarisation of bending was retained in deletion mutants whose flagella cannot beat but have a static bend. Furthermore, deletion mutants for proteins required for asymmetric extra-axonemal and rootlet-like flagellum-associated structures also retained normal polarisation. Leishmania beat polarisation therefore likely arises from either the nine-fold rotational symmetry of the axoneme structure or is due to differences between the outer doublet decorations.Integrin adhesions are a structurally and functionally diverse family of transmembrane, multi-protein complexes that link the intracellular cytoskeleton to the extracellular matrix (ECM). The different members of this family, including focal adhesions (FAs), focal complexes, fibrillar adhesions, podosomes and invadopodia, contain many shared scaffolding and signaling 'adhesome' components, as well as distinct molecules that perform specific functions, unique to each adhesion form. In this Hypothesis, we address the pivotal roles of mechanical forces, generated by local actin polymerization or actomyosin-based contractility, in the formation, maturation and functionality of two members of the integrin adhesions family, namely FAs and invadopodia, which display distinct structures and functional properties. FAs are robust and stable ECM contacts, associated with contractile stress fibers, while invadopodia are invasive adhesions that degrade the underlying matrix and penetrate into it. We discuss here the mechanisms, whereby these two types of adhesion utilize a similar molecular machinery to drive very different - often opposing cellular activities, and hypothesize that early stages of FAs and invadopodia assembly use similar biomechanical principles, whereas maturation of the two structures, and their 'adhesive' and 'invasive' functionalities require distinct sources of biomechanical reinforcement.

To test the hypothesis that there was a temporal change in the retinal microstructure after decompression surgery for chiasmal compression, the 1-year longitudinal changes in the inner and outer retinal thickness after decompression surgery were analyzed using spectral-domain optical coherence tomography (SD-OCT) with linear mixed-effects models.

SD-OCT was obtained from 87 eyes with chiasmal compression and compared to 100 healthy controls. The preoperative and 1-year postoperative longitudinal changes in the retinal layer thickness were measured. The thickness of each of the following retinal layers was analyzed the macular retinal nerve fiber layer (RNFL), the ganglion cell layer (GCL), the inner plexiform layer (IPL), the inner nuclear layer, the outer plexiform layer, the outer nuclear layer, and the photoreceptor layer.

The RNFL, GCL, and IPL showed thinning at a rate of 1.068 μm/y (95% confidence interval [CI], 0.523, 1.613), 1.189 μm/y (95% CI 0.452, 1.925), and 1.177 μm/y (95% CI 0.645, 1.709), respectively, after decompression surgery. The preoperative thickness of the intraretinal layer was associated with postoperative visual field recovery (RNFL, odds ratio [OR] 1.221, 95% CI 1.058, 1.410; GCL, OR 1.133, 95% CI 1.024, 1.254; and IPL, OR 1.174, 95% CI 1.002, 1.376).

The changes in retinal microstructure persisted and progressed in eyes with chiasmal compression after decompression surgery. The findings provide insight into the biological and anatomical sequelae following chiasmal compression. The preoperative thickness of the inner retinal layers was associated with postoperative visual field recovery.

The changes in retinal microstructure persisted and progressed in eyes with chiasmal compression after decompression surgery. The findings provide insight into the biological and anatomical sequelae following chiasmal compression. The preoperative thickness of the inner retinal layers was associated with postoperative visual field recovery.

To investigate whether certain patient, acute care, or primary care factors are associated with medication initiation and discontinuation in the community after stroke or TIA.

This is a retrospective cohort study using prospective data on adult patients with first-ever acute stroke/TIA from the Australian Stroke Clinical Registry (April 2010 to June 2014), linked with nationwide medication dispensing and Medicare claims data. Medication users were those with ≥1 dispensing in the year postdischarge. Discontinuation was assessed among medication users and defined as having no medication supply for ≥90 days in the year postdischarge. Multivariable competing risks regression, accounting for death during the observation period, was conducted to investigate factors associated with time to medication discontinuation.

Among 17,980 registry patients with stroke/TIA, 91.4% were linked to administrative datasets. Of these, 9,817 adults with first-ever stroke/TIA were included (45.4% female, 47.6% aged ≥75 years, aIA are not optimal, with many survivors discontinuing treatment within 1 year postdischarge. Improving postdischarge care for patients with stroke/TIA is needed to minimize unwarranted discontinuation.

To determine whether hemisphere involvement and infarct location on the Alberta Stroke Program CT Score (ASPECTS) template should serve as predictors of 90-day clinical outcome in patients with acute ischemic stroke with pretreatment diffusion-weighted imaging (DWI)-ASPECTS 0-5 treated with mechanical thrombectomy (MT).

We analyzed data of all consecutive patients included in the Endovascular Treatment in Ischemic Stroke registry between January 1, 2012, and August 31, 2018, who presented with a pretreatment DWI-ASPECTS 0-5 and underwent MT. Multivariable analyses were performed in order to identify the role of infarct location and hemisphere involvement on good outcome defined by a modified Rankin Scale (mRS) score 0-2 at 90 days and on the whole distribution of mRS (shift analysis).

A total of 344 patients with a DWI-ASPECTS 0-5 (median 4, IQR 3-5) were included. Neither infarct location nor hemisphere involvement was found to be an independent predictor of good outcome. Involvement of the M6 region istroke management options and neurologic recovery for use of caregivers in the postacute phase.

To investigate the association between protease-activated receptor-1 (PAR-1) gene

polymorphisms and efficacy of clopidogrel for minor stroke or TIA.

Three single nucleotide polymorphisms (

[681G>A, rs4244285],

*

[636G>A, rs4986893], and

[IVSn-14 A/T, rs168753]) were genotyped among 2,924 patients randomized to clopidogrel plus aspirin (n = 1,461) or aspirin alone (n = 1,463). The primary efficacy outcome was new stroke (ischemic or hemorrhagic) and the safety outcome was any bleeding.

Overall, 859 (29.4%) were AA homozygotes, 1,479 (50.6%) were AT heterozygotes, and 586 (20.0%) were TT homozygotes for

IVSn-14 polymorphisms; 1,716 (58.7%) were carriers of at least 1

loss-of-function allele (*2 or *3). Compared with aspirin alone, patients with clopidogrel-aspirin treatment had a low risk of new stroke in patients with AT genotype (7.6% vs 11.3%; hazard ratio [HR], 0.63; 95% confidence interval [CI], 0.44-0.89) and TT genotype (5.8% vs 11.6%; HR, 0.46; 95% CI, 0.25-0.82) but not in carriers of the AA genotype (10.8% vs 11.6%; HR, 0.95; 95% CI, 0.63-1.44) (

= 0.03 for interaction). The association between

IVSn-14 A/T polymorphism and clopidogrel response was present regardless of the carrier status of the

loss-of-function alleles. The

IVSn-14 genotypes were not associated with the risk of any bleeding for clopidogrel-aspirin treatment (

= 0.66 for interaction).

Among patients with minor ischemic stroke or TIA who were receiving clopidogrel and aspirin, those carrying the

IVSn-14 T allele had a lower rate of recurrent stroke than those who were not.

NCT00979589.

NCT00979589.

To evaluate the relationship between circulating phenylalanine and brain function as well as neuropsychiatric symptoms in adults with phenylketonuria.

In this prospective cross-sectional study, early-treated patients with phenylketonuria older than 30 years and age- and sex-matched controls were included. Extensive neurologic evaluation, neuropsychological and behavioral testing, sensory and motor evoked potentials, and MRI were performed. CSF concentrations of neurodegenerative markers were evaluated in addition in a subset of 10 patients.

Nineteen patients with phenylketonuria (median age 41 years) with different phenylalanine levels (median 873 μmol/L) entered the study. They showed higher prevalence of neurologic symptoms, cognitive and behavioral abnormalities, autonomic dysfunction, alterations in neurophysiologic measures, and atrophy in putamen and right thalamus compared to controls. In CSF, patients with phenylketonuria exhibited higher β-amyloid 1-42 (

= 0.003), total tau (

< 0.001), and phosphorylated tau (

= 0.032) levels compared to controls. Plasma phenylalanine levels highly correlated with the number of failed neuropsychological tests (

= 0.64,

= 0.003), neuropsychiatric symptoms (

= 0.73,

< 001), motor evoked potential latency (

= 0.48,

= 0.030), and parietal lobe atrophy.

Our study provides strong evidence for a correlation between phenylalanine levels and clinical, neuropsychological, neurophysiologic, biochemical, and imaging alterations in adult patients with phenylketonuria.

Our study provides strong evidence for a correlation between phenylalanine levels and clinical, neuropsychological, neurophysiologic, biochemical, and imaging alterations in adult patients with phenylketonuria.

To determine the associations between amyloid-PET, tau-PET, and atrophy with the behavioral/dysexecutive presentation of Alzheimer disease (AD), how these differ from amnestic AD, and how they correlate to clinical symptoms.

We assessed 15 patients with behavioral/dysexecutive AD recruited from a tertiary care memory clinic, all of whom had biologically defined AD. They were compared with 25 patients with disease severity- and age-matched amnestic AD and a group of 131 cognitively unimpaired (CU) elderly individuals. All participants were evaluated with amyloid-PET with [

F]AZD4694, tau-PET with [

F]MK6240, MRI, and neuropsychological testing.

Voxelwise contrasts identified patterns of frontal cortical tau aggregation in behavioral/dysexecutive AD, with peaks in medial prefrontal, anterior cingulate, and frontal insular cortices in contrast to amnestic AD. No differences were observed in the distribution of amyloid-PET or atrophy as determined by voxel-based morphometry. Voxelwise area under the receiver operating characteristic curve analyses revealed that tau-PET uptake in the medial prefrontal, anterior cingulate, and frontal insular cortices were best able to differentiate between behavioral/dysexecutive and amnestic AD (area under the curve 0.87). Voxelwise regressions demonstrated relationships between frontal cortical tau load and degree of executive dysfunction.

Our results provide evidence of frontal cortical involvement of tau pathology in behavioral/dysexecutive AD and highlight the need for consensus clinical criteria in this syndrome.

Our results provide evidence of frontal cortical involvement of tau pathology in behavioral/dysexecutive AD and highlight the need for consensus clinical criteria in this syndrome.Increased regulatory T cells (Treg) after radiotherapy have been reported, but the mechanisms of their induction remain incompletely understood. TGFβ is known to foster Treg differentiation within tumors and is activated following radiotherapy. Thus, we hypothesized that TGFβ blockade would result in decreased Tregs within the irradiated tumor microenvironment. We found increased Tregs in the tumors of mice treated with focal radiotherapy and TGFβ blockade. This increase was mediated by upregulation of another TGFβ family member, activin A. In vitro, activin A secretion was increased following irradiation of mouse and human breast cancer cells, and its expression was further enhanced upon TGFβ blockade. In vivo, dual blockade of activin A and TGFβ was required to decrease intratumoral Tregs in the context of radiotherapy. This resulted in an increase in CD8+ T-cell priming and was associated with a reduced tumor recurrence rate. Combination of immune checkpoint inhibitors with the dual blockade of activin A and TGFβ led to the development of tumor-specific memory responses in irradiated breast cancer. Supporting the translational value of activin A targeting to reduce Treg-mediated immunosuppression, retrospective analysis of a public dataset of patients with breast cancer revealed a positive correlation between activin A gene expression and Treg abundance. Overall, these results shed light on an immune escape mechanism driven by activin A and suggest that dual targeting of activin A and TGFβ may be required to optimally unleash radiation-induced antitumor immunity against breast cancer.