Protein associated with SMAD1 (PAWS1/FAM83G) is a substrate for type I bone morphogenetic protein receptors and modulates bone morphogenetic protein signalling

Bone morphogenetic proteins (BMPs) control multiple cellular processes in embryos and adult tissues. BMPs signal through the activation of type I BMP receptor kinases, which then phosphorylate SMADs 1/5/8. In the canonical pathway, this triggers the association of these SMADs with SMAD4 and their translocation to the nucleus, where they regulate gene expression. BMPs can also signal independently of SMAD4, but this pathway is poorly understood. Here, we report the discovery and characterization of PAWS1/FAM83G as a novel SMAD1 interactor. PAWS1 forms a complex with SMAD1 in a SMAD4-independent manner, and BMP signalling induces the phosphorylation of PAWS1 through BMPR1A. The phosphorylation of PAWS1 in response to BMP is essential for activation of the SMAD4-independent BMP target genes NEDD9 and ASNS. Our findings identify PAWS1 as the first non-SMAD substrate for type I BMP receptor kinases and as a novel player in the BMP pathway. We also demonstrate that PAWS1 regulates the expression of several non-BMP target genes, suggesting roles for PAWS1 beyond the BMP pathway

USP15 targets ALK3/BMPR1A for deubiquitylation to enhance bone morphogenetic protein signalling

Protein kinase ALK3/BMPR1A mediates bone morphogenetic protein (BMP) signalling through phosphorylation and activation of SMADs 1/5/8. SMAD6, a transcriptional target of BMP, negatively regulates the BMP pathway by recruiting E3 ubiquitin ligases and targeting ALK3 for ubiquitin-mediated degradation. Here, we identify a deubiquitylating enzyme USP15 as an interactor of SMAD6 and ALK3. We show that USP15 enhances BMP-induced phosphorylation of SMAD1 by interacting with and deubiquitylating ALK3. RNAi-mediated depletion of USP15 increases ALK3 K48-linked polyubiquitylation, and reduces both BMP-induced SMAD1 phosphorylation and transcription of BMP target genes. We also show that loss of USP15 expression from mouse myoblast cells inhibits BMP-induced osteoblast differentiation. Furthermore, USP15 modulates BMP-induced phosphorylation of SMAD1 and transcription during Xenopus embryogenesis

PAWS1 controls Wnt signalling through association with casein kinase 1α

The BMP and Wnt signalling pathways determine axis specification during embryonic development. Our previous work has shown that PAWS1 (also known as FAM83G) interacts with SMAD1 and modulates BMP signalling. Here, surprisingly, we show that overexpression of PAWS1 in Xenopus embryos activates Wnt signalling and causes complete axis duplication. Consistent with these observations in Xenopus, Wnt signalling is diminished in U2OS osteosarcoma cells lacking PAWS1, while BMP signalling is unaffected. We show that PAWS1 interacts and co-localises with the α isoform of casein kinase 1 (CK1), and that PAWS1 mutations incapable of binding CK1 fail both to activate Wnt signalling and to elicit axis duplication in Xenopus embryos.</p

Influenza epidemiology, vaccine coverage and vaccine effectiveness in children admitted to sentinel Australian hospitals in 2014: The influenza complications alert network (FluCAN)

The Influenza Complications Alert Network (FluCAN) is a sentinel hospital-based surveillance programme operating in all states and territories in Australia. We summarise the epidemiology of children hospitalised with laboratory-confirmed influenza in 2014 and reports on the effectiveness of inactivated trivalent inactivated vaccine (TIV) in children. In this observational study, cases were defined as children admitted with acute respiratory illness (ARI) with influenza confirmed by PCR. Controls were hospitalised children with ARI testing negative for influenza. Vaccine effectiveness (VE) was estimated as 1 minus the odds ratio of vaccination in influenza positive cases compared with test-negative controls using conditional logistic regression models. From April until October 2014, 402 children were admitted with PCR-confirmed influenza. Of these, 28% were aged < 1 year, 16% were Indigenous, and 39% had underlying conditions predisposing to severe influenza. Influenza A was detected in 90% of cases of influenza; influenza A(H1N1)pdm09 was the most frequent subtype (109/141 of subtyped cases) followed by A(H3N2) (32/141). Only 15% of children with influenza received antiviral therapy. The adjusted VE of one or more doses of TIV for preventing hospitalised influenza was estimated at 55.5% (95% confidence intervals (CI): 11.6–77.6%). Effectiveness against influenza A(H1N1)pdm09 was high (91.6%, 95% CI: 36.0–98.9%) yet appeared poor against H3N2. In summary, the 2014 southern hemisphere TIV was moderately effective against severe influenza in children. Significant VE was observed against influenza A(H1N1)pdm0

Effects of Whey Protein on Skeletal Muscle Microvascular and Mitochondrial Plasticity Following 10-Weeks of Exercise Training in Men with Type-2 Diabetes

Copyright remains with the author(s) or their institution(s).Abstract Skeletal muscle microvascular dysfunction and mitochondrial rarefaction feature in type 2 diabetes mellitus (T2DM) linked to low tissue glucose disposal rate (GDR). Exercise training and milk protein supplementation independently promote microvascular and metabolic plasticity in muscle associated with improved nutrient delivery, but combined effects are unknown. In a randomised-controlled trial, 24 men (55.6 y, SD 5.7) with T2DM ingested whey protein drinks (protein/carbohydrate/fat: 20/10/3 g; WHEY) or placebo (carbohydrate/fat: 30/3 g; CON) before/after 45 mixed-mode intense exercise sessions over 10 weeks, to study effects on insulin-stimulated (hyperinsulinemic clamp) skeletal-muscle microvascular blood flow (mBF) and perfusion (near-infrared spectroscopy), and histological, genetic, and biochemical markers (biopsy) of microvascular and mitochondrial plasticity. WHEY enhanced insulin-stimulated perfusion (WHEY-CON 5.6%; 90% CI −0.1, 11.3), while mBF was not altered (3.5%; −17.5, 24.5); perfusion, but not mBF, associated (regression) with increased GDR. Exercise training increased mitochondrial (range of means: 40%–90%) and lipid density (20%–30%), enzyme activity (20%–70%), capillary:fibre ratio (∼25%), and lowered systolic (∼4%) and diastolic (4%–5%) blood pressure, but without WHEY effects. WHEY dampened PGC1α −2.9% (90% compatibility interval: −5.7, −0.2) and NOS3 −6.4% (−1.4, −0.2) expression, but other messenger RNA (mRNA) were unclear. Skeletal muscle microvascular and mitochondrial exercise adaptations were not accentuated by whey protein ingestion in men with T2DM. ANZCTR Registration Number: ACTRN12614001197628. Novelty: • Chronic whey ingestion in T2DM with exercise altered expression of several mitochondrial and angiogenic mRNA. • Whey added no additional benefit to muscle microvascular or mitochondrial adaptations to exercise. • Insulin-stimulated perfusion increased with whey but was without impact on glucose disposal. Résumé Le dysfonctionnement microvasculaire du muscle squelettique et la raréfaction mitochondriale caractérisant le diabète de type 2 (« T2DM ») sont liés à un faible taux d’élimination du glucose tissulaire (« GDR »). L’entraînement physique et la supplémentation en protéines du lait favorisent indépendamment la plasticité microvasculaire et métabolique dans le muscle; cette plasticité est associée à une amélioration de l’apport de nutriments, mais les effets combinés sont inconnus. Dans un essai contrôlé randomisé, 24 hommes (55,6 ans, SD 5,7) aux prises avec le T2DM consomment des boissons protéinées de lactosérum (protéines / glucides / lipides: 20/10/3 g; « WHEY ») ou un placebo (glucides / lipides: 30/3 g; « CON ») avant / après 45 séances d’exercice intense en mode mixte sur 10 semaines, et ce, pour examiner les effets sur le flux sanguin microvasculaire (« mBF ») et la perfusion (spectroscopie proche infrarouge) stimulés par l’insuline (clamp hyperinsulinémique), des variables histologiques, génétiques et des marqueurs biochimiques (biopsie) de la plasticité microvasculaire et mitochondriale. WHEY améliore la perfusion stimulée par l’insuline (WHEY-CON 5,6 %; IC 90 % −0,1, 11,3), tandis que le mBF n’est pas modifié (3,5 %; −17,5, 24,5); la perfusion, mais pas le mBF, est associée (régression) à une augmentation du GDR. L’entraînement à l’exercice augmente la densité mitochondriale (gamme de moyennes: 40-90 %) et lipidique (20−30 %), l’activité enzymatique (20−70 %), le ratio capillaire: fibre (∼25 %) et diminue les pressions systolique (∼4 %) et diastolique (4−5 %), mais sans effets de WHEY. WHEY amortit l’expression de PGC1α −2,9 % (intervalle de compatibilité de 90 % : −5,7, −0,2) et NOS3 −6,4 % (−1,4, −0,2), mais les autres ARN messager (ARNm) ne sont pas clairs. Les adaptations microvasculaires et mitochondriales des muscles squelettiques causées par l’entraînement physique ne sont pas accentuées par la consommation de protéines de lactosérum chez les hommes aux prises avec le T2DM. Numéro d’enregistrement ANXCTR : ACTRN12614001197628. [Traduit par la Rédaction] Les nouveautés: • La consommation prolongée de lactosérum en présence de T2DM combinée à l’entraînement physique modifie l’expression de plusieurs ARNm mitochondriaux et angiogéniques. • Le lactosérum n’ajoute aucun avantage supplémentaire aux adaptations microvasculaires ou mitochondriales musculaires à l’exercice physique. • La perfusion stimulée par l’insuline augmente avec le lactosérum mais n’a pas d’impact sur l’élimination du glucose.falseGaffney, Kim Lucero, Adam Macartney-Coxson, Donia Clapham, Jane Whitfield, Patricia Palmer, Barry Wakefield, StJohn Faulkner, James Stoner, Lee Rowlands, David Stephen eng Canada Appl Physiol Nutr Metab. 2021 Feb 16. doi: 10.1139/apnm-2020-0943. Skeletal muscle microvascular dysfunction and mitochondrial rarefaction feature in type-2 diabetes mellitus (T2DM) linked to low tissue glucose disposal rate (GDR). Exercise training and milk protein supplementation independently promote microvascular and metabolic plasticity in muscle associated with improved nutrient delivery, but combined effects are unknown. In a randomised-controlled trial, 24 men (55.6 y, SD5.7) with T2DM ingested whey protein drinks (protein/carbohydrate/fat: 20/10/3 g; WHEY) or placebo (carbohydrate/fat: 30/3 g; CON) before/after 45 mixed-mode intense exercise sessions over 10 weeks, to study effects on insulin-stimulated (hyperinsulinemic clamp) skeletal-muscle microvascular blood flow (mBF) and perfusion (near-infrared spectroscopy), and histological, genetic, and biochemical markers (biopsy) of microvascular and mitochondrial plasticity. WHEY enhanced insulin-stimulated perfusion (WHEY-CON 5.6%; 90%CI -0.1, 11.3), while mBF was not altered (3.5%; -17.5, 24.5); perfusion, but not mBF, associated (regression) with increased GDR. Exercise training increased mitochondrial (range of means: 40-90%) and lipid density (20-30%), enzyme activity (20-70%), capillary:fiber ratio (~25%), and lowered systolic (~4%) and diastolic (4-5%) blood pressure, but without WHEY effects. WHEY dampened PGC1alpha -2.9% (90%CI -5.7, -0.2) and NOS3 -6.4% (-1.4, -0.2) expression, but other mRNA were unclear. Skeletal muscle microvascular and mitochondrial exercise adaptations were not accentuated by whey protein ingestion in men with T2DM. Clinical Trial Registration Number: ACTRN12614001197628 Novelty Bullets: * Chronic whey ingestion in T2DM with exercise altered expression of several mitochondrial and angiogenic mRNA. * Whey added no additional benefit to muscle microvascular or mitochondrial adaptations to exercise. * Insulin-stimulated perfusion increased with whey but was without impact on glucose disposal

Population‐based cohort study of outcomes following cholecystectomy for benign gallbladder diseases

Background The aim was to describe the management of benign gallbladder disease and identify characteristics associated with all‐cause 30‐day readmissions and complications in a prospective population‐based cohort. Methods Data were collected on consecutive patients undergoing cholecystectomy in acute UK and Irish hospitals between 1 March and 1 May 2014. Potential explanatory variables influencing all‐cause 30‐day readmissions and complications were analysed by means of multilevel, multivariable logistic regression modelling using a two‐level hierarchical structure with patients (level 1) nested within hospitals (level 2). Results Data were collected on 8909 patients undergoing cholecystectomy from 167 hospitals. Some 1451 cholecystectomies (16·3 per cent) were performed as an emergency, 4165 (46·8 per cent) as elective operations, and 3293 patients (37·0 per cent) had had at least one previous emergency admission, but had surgery on a delayed basis. The readmission and complication rates at 30 days were 7·1 per cent (633 of 8909) and 10·8 per cent (962 of 8909) respectively. Both readmissions and complications were independently associated with increasing ASA fitness grade, duration of surgery, and increasing numbers of emergency admissions with gallbladder disease before cholecystectomy. No identifiable hospital characteristics were linked to readmissions and complications. Conclusion Readmissions and complications following cholecystectomy are common and associated with patient and disease characteristics

A RP-HPLC-UV method for the dual detection of fluconazole and clobetasol propionate and application to a model dual drug delivery hydrogel

Advanced drug delivery systems have become widely investigated to improve the efficacy of treatments for several diseases. These devices offer improved efficient, sustained, and targeted delivery which improves patient compliance, quality of life and minimises potential systemic side effects. As these therapeutic devices have advanced there is a potential for the development of products which deliver multiple drugs for simultaneous treatment of diseases. Given the interest in these dual-delivery devices it follows that new analytical methods need to be developed to detect and quantify different analytes during device development and validation. Here, for the first time, a reverse-phase high performance liquid chromatography (RP-HPLC) method is validated, utilising UV detection, for the dual detection of fluconazole and clobetasol propionate. The method is tested on a dual loaded model implant material intended as mucosal patches for the direct treatment of lichen planus and associated fungal infections. The method described here exhibited specificity and robustness with accurate and precise results. Good linearity was obtained between 0.25 and 2.5 mg mL−1 for fluconazole and 5 and 50 μg mL−1 for clobetasol propionate, with an R2 value of 0.9999 for the dual detection of fluconazole and clobetasol propionate. The developed method demonstrated selectivity and the solution containing both fluconazole and clobetasol propionate remained stable over a range of storage temperatures for up to 28 days. Within this validation study, the protocol was applied to a relevant dual loaded film showing the suitability of the method in studying drug release characteristics. The method described here also has a broader applicability for analysis and quantification of in vitro and in vivo drug release studies

Serological testing of blood donors to characterise the impact of COVID-19 in Melbourne, Australia, 2020.

Rapidly identifying and isolating people with acute SARS-CoV-2 infection has been a core strategy to contain COVID-19 in Australia, but a proportion of infections go undetected. We estimated SARS-CoV-2 specific antibody prevalence (seroprevalence) among blood donors in metropolitan Melbourne following a COVID-19 outbreak in the city between June and September 2020. The aim was to determine the extent of infection spread and whether seroprevalence varied demographically in proportion to reported cases of infection. The design involved stratified sampling of residual specimens from blood donors (aged 20-69 years) in three postcode groups defined by low (7 cases/1,000 population) COVID-19 incidence based on case notification data. All specimens were tested using the Wantai SARS-CoV-2 total antibody assay. Seroprevalence was estimated with adjustment for test sensitivity and specificity for the Melbourne metropolitan blood donor and residential populations, using multilevel regression and poststratification. Overall, 4,799 specimens were collected between 23 November and 17 December 2020. Seroprevalence for blood donors was 0.87% (90% credible interval: 0.25-1.49%). The highest estimates, of 1.13% (0.25-2.15%) and 1.11% (0.28-1.95%), respectively, were observed among donors living in the lowest socioeconomic areas (Quintiles 1 and 2) and lowest at 0.69% (0.14-1.39%) among donors living in the highest socioeconomic areas (Quintile 5). When extrapolated to the Melbourne residential population, overall seroprevalence was 0.90% (0.26-1.51%), with estimates by demography groups similar to those for the blood donors. The results suggest a lack of extensive community transmission and good COVID-19 case ascertainment based on routine testing during Victoria's second epidemic wave. Residual blood donor samples provide a practical epidemiological tool for estimating seroprevalence and information on population patterns of infection, against which the effectiveness of ongoing responses to the pandemic can be assessed