Parental home-based pulse oximetry monitoring for adults with intellectual disabilities at risk of serious respiratory problems including COVID-19: a brief report.

BACKGROUND: People with intellectual disabilities (ID) are at high risk of developing respiratory health issues. The COVID-19 pandemic has compounded this, with serious consequences, and for some, death. Despite home-based oxygen saturation monitoring being recommended for people with ID, there is a stark lack of evidence in the literature on its feasibility. METHOD: We conducted 3-day baseline home-based oxygen saturation monitoring, using pulse oximeters, with eight parents of nine adults with ID in Scotland. Two eligible parents also completed a further 2 weeks of monitoring, and returned an evaluation questionnaire on its feasibility. RESULTS: Baseline mean readings for eight adults with ID were within the normal range (%Sp02  ≥ 95), and for another one 94%. Fluctuations over the 3-day assessment period were experienced by six of these individuals. However, these variations were within limits which are not dangerous (lowest reading 92%), implying that parental home-based pulse oximetry monitoring is likely to be safe for adults with ID. The two parents who completed the evaluation found home-based pulse oximetry monitoring to be easy/very easy to do, and effective/very effective. CONCLUSIONS: This is the first research study, albeit with a very small sample, to report on the potential feasibility of parental home-based pulse oximetry monitoring for adults with ID. Home-based pulse oximetry monitoring appears to be safe in adults with ID at risk of developing serious respiratory problems, and not difficult for their parents to do. There is an urgent need to replicate this work, using a larger sample, to promote home-based respiratory health monitoring more widely for people with ID.

Characterizing and predicting the relationship between translaminar fracture toughness and pull-out length distributions under distinct temperatures.

The translaminar fracture toughness reflects the damage tolerance of a fibre-reinforced composite under longitudinal tension, which often governs the final failure of structures. One of the main energy-dissipation mechanisms that contributes to the translaminar toughness of composites is the fibre pull-out process. The present study aims to quantify and model the statistical distribution of fibre pull-out lengths formed on the translaminar fracture surface of composites, for the first time in the literature; this is done under different temperatures, so that the relationship between pull-out length distributions, micromechanical properties and the translaminar fracture toughness can be established. The fracture surfaces of cross-ply compact tension specimens tested under three different temperatures have been scanned through X-ray computed tomography to quantify the extent of fibre pull-out on the fracture surfaces; the distribution of pull-out lengths showed alarger average and larger variability with an increase in temperature, which also lead to an increase in translaminar fracture toughness. A similar trend has been captured by the proposed analytical model, which predicts the pull-out length distribution based on the analysis of quasi-fractal idealizations of the fracture surface, yielding an overall accuracy of more than 85%. This article is part of the theme issue 'Ageing and durability of composite materials'.

Prevalence of factors associated with edentulousness (no natural teeth) in adults with intellectual disabilities.

BACKGROUND: Poor oral health is largely preventable. Prevention includes toothbrushing and regular dental checks. Oral health has important consequences for general nutrition, chewing, communication, wider systemic disease, self-confidence and participation in society. This study investigated the prevalence of edentulousness (no natural teeth) in adults with intellectual disabilities (IDs) compared with the general population and associated factors. METHODS: An adult cohort with IDs residing in Greater Glasgow and Clyde, Scotland, underwent detailed health assessments between 2002 and 2004. Between 2004 and 2006, a subsample had an oral check. Data on edentulousness in the cohort were compared with adult participants from Greater Glasgow and Clyde in the 2008 Scottish Health Survey. Within the IDs cohort, binary logistic regression analyses investigated potential relationships between edentulousness and demographic and clinical factors. RESULTS: Five hundred sixty adults with IDs were examined [53.2% (298) male, mean age = 46.3 years, range 18-81 years] and compared with 2547 general population: edentulousness was 9% vs. 1% aged 25-34 years; 22% vs. 2% aged 35-44 years; 39% vs. 7% aged 45-54 years; 41% vs. 18% aged 55-64 years; and 76% vs. 34% aged 65-74 years. In both groups, edentulousness increased with age. After stratification for age, rates of edentulousness were consistently higher in the ID cohort. Odds ratios within age strata were not homogenous (Mantel-Haenszel test, P < 0.0001). Edentulousness was more likely in those with more severe IDs (adjusted odds ratio (AOR) = 2.36; 95% confidence interval (CI) [1.23 to 4.51]); those taking antipsychotics (AOR = 2.09; 95% CI [1.25 to 3.51]) and those living in the most deprived neighbourhoods (AOR = 2.69; 95% CI [1.11 to 6.50]). There was insufficient evidence for associations with sex, type of accommodation/support, antiepileptics, problem behaviours or autism. CONCLUSIONS: Adults with IDs have a high prevalence of edentulousness and need supported daily oral care to reduce the need for extractions. Despite previous reports on poor oral care and the move towards person-centred care, carers and care-giving organisations need greater support to implement daily oral care. Prescribers need awareness of the potentially contributory role of antipsychotics, which may relate to xerostomia.

Dominant and sensitive control of oxidative flux by the ATP-ADP carrier in human skeletal muscle mitochondria: Effect of lysine acetylation.

The adenine nucleotide translocase (ANT) of the mitochondrial inner membrane exchanges ADP for ATP. Mitochondria were isolated from human vastus lateralis muscle (n = 9). Carboxyatractyloside titration of O2 consumption rate (Jo) at clamped [ADP] of 21 µM gave ANT abundance of 0.97 ±â€¯0.14 nmol ANT/mg and a flux control coefficient of 82% ±â€¯6%. Flux control fell to 1% ±â€¯1% at saturating (2 mM) [ADP]. The KmADP for Jo was 32.4 ±â€¯1.8 µM. In terms of the free (-3) ADP anion this KmADP was 12.0 ±â€¯0.7 µM. A novel luciferase-based assay for ATP production gave KmADP of 13.1 ±â€¯1.9 µM in the absence of ATP competition. The free anion KmADP in this case was 2.0 ±â€¯0.3 µM. Targeted proteomic analyses showed significant acetylation of ANT Lysine23 and that ANT1 was the most abundant isoform. Acetylation of Lysine23 correlated positively with KmADP, r = 0.74, P = 0.022. The findings underscore the central role played by ANT in the control of oxidative phosphorylation, particularly at the energy phosphate levels associated with low ATP demand. As predicted by molecular dynamic modeling, ANT Lysine23 acetylation decreased the apparent affinity of ADP for ANT binding.

Lycra splinting garments for adults with intellectual disabilities who fall due to gait or balance issues: a feasibility study.

BACKGROUND: Adults with intellectual disabilities (IDs) experience high rates of falls and have high rates of gait or balance issues which contribute to falls. Lycra splinting garments (LSGs) have potential to improve gait or balance, but they have never before been tested with adults with IDs who fall due to gait or balance issues. The aim of this study was to test in adults with IDs, the feasibility of using LSGs to improve movement and function and reduce falls, whilst also exploring usability and likely compliance. METHOD: A convenience sample of nine adults with IDs wore tailored LSGs over a 6-week assessment period. Laboratory-based foot clearance, balance, and gait measures were collected pre- and post-LSG-wear. Falls charts and questionnaires on usability and likely compliance were also completed. RESULTS: Seven participants experienced a reduction in falls during their six weeks of LSG wear; most notably in the group of five participants who wore lycra splinting socks, compared with only two in the group of four who wore lycra splinting shorts or leggings only. CONCLUSION: Lycra splinting socks are likely to bring about positive outcomes for adults with IDs who fall due to gait/balance issues on an individual case by case basis over time, but further research is required to test this hypothesis under randomised controlled trial conditions. Potential benefits of more intrusive LSGs are outweighed by reported problems with usability and compliance.

Characterization of Immune System Cell Subsets in Fixed Tissues from Alpine Chamois (Rupicapra rupicapra).

Immune system cell subsets in lymph nodes and spleen from alpine chamois (Rupicapra rupicapra subspecies rupicapra) living in the Italian Alps were characterized immunohistochemically. Seven primary antibodies (against human CD3, CD79αcy, CD68, or ovine CD4, CD8, CD21 and γδ T-cell receptor [TCR] epitopes) were tested on tissues fixed either in formalin or in zinc salts (ZS) and cross-reactivity with chamois immune cell epitopes was shown. ZS fixation allowed wider identification of immune cells, without the need for antigen retrieval. CD4(+) and CD21(+) cells were labelled only in ZS-fixed tissues. Reagents specific for human CD3, CD79 and CD68 antigens successfully detected chamois immune cells, both in ZS-fixed and formalin-fixed tissues. The reactivity and distribution of immune cells in lymph nodes and spleen were similar to those described in other domestic and wild ruminants. Results from this study may allow future investigation of the immune response and pathogenesis of diseases in the chamois.

Differences in Virulence Between Bovine-Derived Clinical Isolates of Pasteurella multocida Serotype A from the UK and the USA in a Model of Bovine Pneumonic Pasteurellosis.

The time of onset and subsequent degree and progression of clinical signs, bacterial colonization and tissue pathology during experimental disease induced by intratracheal inoculation of either a UK or USA isolate of Pasteurella multocida serotype A recovered from clinical cases of bovine pneumonia were determined. Calves aged 8 weeks were challenged with 300 ml phosphate buffered saline (PBS) alone (group 1, n = 3, negative control) or containing 7.1 × 10(8) colony forming units (cfu) of UK isolate (group 2, n = 8) or 5.8 × 10(8) cfu of USA isolate (group 3, n = 8). Bronchoalveolar lavage (BAL) at 0, 1 and 4 days post challenge (dpc) and at the time of necropsy examination (7-8 dpc) showed no significant differences between groups 2 and 3 in bacterial numbers recovered. No P. multocida were recovered from group 1 animals. No clinical disease was present in group 1 calves and in group 3 was limited to scour in 1 calf at 1 dpc. All calves in group 2 had reduced food intake at 4-5 dpc, five had periods of dullness, three a mild nasal discharge at 1 dpc, four had mild to substantial respiratory stridor and one was killed at 6 dpc for humane reasons. Rectal temperatures remained about 39°C in group 1 calves, but increased in P. multocida-challenged calves to 40-41°C within 8-12 h of challenge. Significantly (P = 0.01) greater percentages of lung surface area were consolidated in group 2 (mean ± SD, 21 ± 10.1) compared with group 3 (7 ± 8.6) calves. Significantly more extensive and severe histological lesions were present in the lung lobes (P = 0.006) and lymph nodes (P = 0.02) of group 2 compared with group 3 calves. Pleurisy was present in group 2 calves only and no pathology was present in group 1. Pulsed-field gel electrophoresis (PFGE) produced 11 (group 2, UK isolate) or 10 (group 3, USA isolate) bands with differences in banding patterns. Results overall showed that two isolates, distinct geographically and genetically (by PFGE), caused pneumonic pasteurellosis in a single host with significantly different severity of pathology. This information is relevant to the development of novel vaccine control and interpretation of diagnostic results.

Structural Development, Cellular Differentiation and Proliferation of the Respiratory Epithelium in the Bovine Fetal Lung.

Fetal bovine lung samples of 11 different gestational ages were assigned to a classical developmental stage based on histological morphology. Immunohistochemistry was used to characterize the morphology of forming airways, proliferation rate of airway epithelium and the presence of epithelial cell types (i.e. ciliated cells, club cells, neuroepithelial cells (NECs) and type II pneumocytes). Typical structural organization of pseudoglandular (84-98 days gestational age [DGA]), canalicular (154-168 DGA) and alveolar (224-266 DGA) stages was recognized. In addition, transitional pseudoglandular-canalicular (112-126 DGA) and canalicular-saccular (182 DGA) morphologies were present. The embryonic stage was not observed. A significantly (P <0.05) higher proliferation rate of pulmonary epithelium, on average 5.5% and 4.4% in bronchi and bronchioles, respectively, was present in the transitional pseudoglandular-canalicular phase (112-126 DGA) compared with all other phases, while from 8 weeks before term (224-266 DGA) proliferation had almost ceased. The first epithelial cells identified by specific marker proteins in the earliest samples available for study (84 DGA) were ciliated cells and NECs. Club cells were present initially at 112 DGA and type II pneumocytes at 224 DGA. At the latest time points (224-226 DGA) these latter cell types were still present at a much lower percentage compared with adult cattle. This study characterized bovine fetal lung development by histological morphology and cellular composition of the respiratory epithelium and suggests that the apparent structural anatomical maturity of the bovine lung at term is not matched by functional maturity of the respiratory epithelium.

Host-pathogen dynamics of squirrelpox virus infection in red squirrels (Sciurus vulgaris).

To improve our understanding of squirrelpox virus (SQPV) infection in the susceptible host, three red squirrels were challenged with wild-type SQPV via scarification of the hind-limb skin. All squirrels seroconverted to the infection by the end of the experiment (17 days post-challenge). Challenged animals suffered disease characterised by the development of multiple skin and oral lesions with rapid progression of skin lesions at the infection site by day 10 post-challenge. No internal pathological changes were found at post-mortem examination. A novel SQPV Taqman(®) Real-time PCR detected viral DNA from multiple organs, with the largest amounts consistently associated with the primary and secondary skin and oral lesions where viral replication was most likely occurring. Immunohistochemistry clearly detected viral antigen in the stratified squamous epithelium of the epidermis, tongue and the oropharyngeal mucosa-associated lymphoid tissue and was consistently associated with histological changes resulting from viral replication. The lack of internal pathological changes and the detection of relatively low levels of viral DNA when compared with primary and secondary skin lesions argue against systemic disease, although systemic spread of the virus cannot be ruled out. This study allowed a comprehensive investigation of the clinical manifestation and progression of SQPV infection with a quantitative and qualitative analysis of virus dissemination and shedding. These findings suggest two separate routes of SQPV transmission under natural conditions, with both skin and saliva playing key roles in infected red squirrels.

The provision of aids and adaptations, risk assessments, and incident reporting and recording procedures in relation to injury prevention for adults with intellectual disabilities: cohort study.

BACKGROUND: Adults with intellectual disabilities (IDs) experience a higher incidence of injury, compared with the general population. The aim of this study was to investigate the provision of aids and adaptations, residential service providers' individual risk assessments and training in these, and injury incident recording and reporting procedures, in relation to injury prevention. METHOD: Interviews were conducted with a community-based cohort of adults with IDs (n = 511) who live in Greater Glasgow, Scotland, UK and their key carer (n = 446). They were asked about their aids and adaptations at home, and paid carers (n = 228) were asked about individual risk assessments, their training, and incident recording and reporting procedures. RESULTS: Four hundred and twelve (80.6%) of the adults with IDs had at least one aid or adaptation at home to help prevent injury. However, a proportion who might benefit, were not in receipt of them, and surprisingly few had temperature controlled hot water or a bath thermometer in place to help prevent burns/scalds, or kitchen safety equipment to prevent burns/scalds from electric kettles or irons. Fifty-four (23.7%) of the paid carers were not aware of the adult they supported having had any risk assessments, and only 142 (57.9%) had received any training on risk assessments. Considerable variation in incident recording and reporting procedures was evident. CONCLUSION: More work is needed to better understand, and more fully incorporate, best practice injury prevention measures into routine support planning for adults with IDs within a positive risk-taking and risk reduction framework.

Pathogenesis of scrapie in ARQ/ARQ sheep after subcutaneous infection: effect of lymphadenectomy and immune cell subset changes in relation to prion protein accumulation.

It is well established that the infectious agent of scrapie can replicate in the lymphoreticular system (LRS). However, the effects of removal of LRS target tissues on the pathogenesis of the infection and the accumulation of disease-associated prion protein (PrP(d)) in LRS tissues on specific immune cell subsets are poorly understood aspects. To address these questions 16 ARQ/ARQ sheep were subcutaneously inoculated in the drainage area of the prefemoral lymph node with brain homogenate derived from Suffolk sheep naturally infected with scrapie. Fourteen sheep were then subjected to either early (14-17 days post-inoculation [dpi]) or late (175-201 dpi) lymphadenectomy and culled at preclinical or clinical stages of infection. Neither late nor even early lymphadenectomy prevented infection or had any effect on the accumulation of PrP(d) in the LRS or CNS suggesting a rapid organic dissemination of the infectious agent after inoculation. Lymph nodes from eight scrapie inoculated sheep selected on the basis of the amount of PrP(d) in their LRS tissues (negative, low or high) were examined for six different immune cell markers. The PrP(d) negative lymph nodes from two sheep with no evidence of scrapie infection showed lower numbers of cluster of determination (CD) 21 positive cells than PrP(d) positive nodes, irrespective of their location (hind leg or head). However, quantitative differences in the expression of this marker were not detected when comparing lymph nodes with low and high levels of PrP(d) accumulation, suggesting that proliferation of CD21 positive cells is related to scrapie infection, but not directly linked to the magnitude of PrP(d) accumulation. An additional observation of the study was that sheep that were methionin-threonine at codon 112 of the prion protein gene showed lower attack rates than methionine homozygotes (67% and 100%, respectively) and also generally lower levels of PrP(d) accumulation in the LRS and brain and increased survival times, suggesting an influence of such polymorphism in the susceptibility to scrapie.

Seven strains of mice as potential models of bovine pasteurellosis following intranasal challenge with a bovine pneumonic strain of Pasteurella multocida A:3; comparisons of disease and pathological outcomes.

The gram-negative bacterium Pasteurella multocida causes pneumonic and systemic pasteurellosis in bovids for which vaccines are either unavailable or inadequate. The work assessed whether an intranasal P. multocida challenge in mice might provide a model of infection for future vaccine development work. Clinical, pathological and biochemical responses were compared in seven strains of mice challenged with a virulent bovine pneumonic isolate of P. multocida A:3. Six mouse strains (Porton, CD-1, BALB/c, VM, C57BL/10 and C57BL/6) developed clinical signs of pneumonic disease and variable pneumonic lesions 41-70 h post-infection. In contrast, mouse strain RIII became septicaemic within 36 h post-infection. Concentrations of plasma acute phase proteins and serum lipopolysaccharide increased in all mice after infection, and the main or interaction effect of mouse strain and infection status was statistically significant (P<0.05). Responses in C57BL/10 mice showed close similarity to bovine pneumonic and in RIII mice to bovine systemic pasteurellosis.

Evaluation of a new rapid anti-HbF FITC assay, Trillium QuikQuant, for detection and quantitation of foetomaternal haemorrhage.

INTRODUCTION: This study evaluated a new product for flow cytometric foetomaternal haemorrhage (FMH) quantitation, Trillium QuikQuant anti-HbF FITC kit (Trillium Diagnostics, USA), in comparison with the Millipore anti-HbF FITC method. METHODS: Blood from 67 antenatal or postpartum RhD-negative women together with 108 samples spiked with 0.2%, 0.4%, 0.6%, 2%, 4%, 5%, 6%, 8% and 10% of foetal red cells were analysed by both methods. RESULTS: Results for both methods were linear to 10% foetal red cells. Analysis using the Wilcoxon test found that there was no difference between the paired results of Trillium QuikQuant and the expected values of the spiked samples (P = 0.48). In contrast, Millipore anti-HbF FITC gave statistically higher results compared with both the expected values and Trillium QuikQuant anti-HbF FITC (P = 0.0003 and P < 0.0001, respectively). The Bland-Altman plot for Trillium QuikQuant showed a mean difference of only 0.06% below the expected values, whilst Millipore was 0.13% above. The analysis time with Trillium QuikQuant was approximately 40 min, requiring significantly less hands-on time than the Millipore method that required an additional four wash steps. CONCLUSION: Trillium QuikQuant is a new precise, accurate and rapid flow cytometric kit method for the quantitation of FMH in both the antenatal and postpartum period.

The first report of otarine herpesvirus-1-associated urogenital carcinoma in a South American fur seal (Arctocephalus australis).

Otarine herpesvirus (OtHV)-1-associated urogenital carcinoma has been well documented in the California sea lion (Zalophus californianus, CSL), but this is the first report of this tumour in a captive South American fur seal (Arctocephalus australis, SAFS). The gross and microscopical morphology of the tumour in the SAFS was identical to that described previously in CSLs and the tumour in the present case had metastasized within the urogenital tract and draining lymph nodes and to the lungs and one kidney. Immunohistochemistry revealed intra- and extracytoplasmic labelling of herpesvirus antigen in the cells of the tumour tissue and transitional epithelium of the urethra. OtHV-1 nucleic acids were detected within tumour tissue and from a urogenital swab by polymerase chain reaction. The ranges of these two species of pinniped do not overlap normally in the wild, suggesting that transmission of OtHV-1 probably occurred in captivity. This confirmed susceptibility of the SAFS to the development of OtHV-1-associated urogenital carcinoma suggests that all species of Otariidae should be screened for OtHV-1 infection prior to movement within and between zoological collections.

Iron studies and red cell transfusion in cardiothoracic and orthopaedic surgical patients: a retrospective audit at a tertiary hospital.

BACKGROUND AND OBJECTIVES: Preoperative diagnosis and treatment of anaemia are important to minimize adverse postoperative outcomes. This audit reviewed red cell transfusion practice, degree of anaemia, iron deficiency anaemia (IDA) and chronic disease or anaemia of inflammation (AI) in cardiothoracic and orthopaedic surgical patients who had available iron studies. MATERIALS AND METHODS: A total of 178 consecutive cardiothoracic and orthopaedic surgical patients with available iron studies were retrospectively reviewed. RESULTS: Of patients, 36·5% had preoperative iron studies. However, 63·2% males and 45·3% females with postoperative iron studies presented with anaemia; 38·5% patients with preoperative iron studies had AI; 21·5% IDA; 23·1% normal. For patients with iron studies requested within the first two postoperative intervals (≤ 5 days and 6 ≤ 10 days) 73·8% and 63·6%, respectively, had AI; few had classical IDA or were normal, and 51·5% patients transfused postsurgery had a discharge Hb ≥ 110 g/l. Restricting the discharge Hb to 90 or 100 g/l may have eliminated postsurgical transfusion in 14·8-42·6% patients. CONCLUSION: Iron studies were more commonly requested postoperatively despite many being anaemic at admission. A higher proportion of patients with postoperative iron studies had AI, and few had classical IDA or normal iron parameters, suggesting a transient inflammatory effect of surgery. This may mask underlying IDA or normal iron parameters and affect treatment. Preadmission assessment, including iron status, should be emphasized allowing diagnosis and correction of presurgical anaemia with treatment modalities other than red cell transfusion. In the postsurgical setting, consideration of a restrictive transfusion regimen sufficient to alleviate a patient's clinical symptoms would ensure that this valuable resource is appropriately used.

Immunophenotype of cells within cervine rectoanal mucosa-associated lymphoid tissue and mesenteric lymph nodes.

Rectoanal mucosa-associated lymphoid tissue (RAMALT) is a part of the lymphoid system that can be sampled easily in live animals, especially ruminants. RAMALT biopsy is useful for the diagnosis of transmissible spongiform encephalopathies, including scrapie in sheep and goats and chronic wasting disease (CWD) in cervids. Diagnosis is reliant on detection of abnormal prion protein (PrP(d)), which is associated with lymphoid follicles. For enzyme linked immunosorbent assays (ELISAs) detecting PrP(d) it is necessary to ensure that lymphoid follicles are present in biopsy samples to avoid false-negative results. Monoclonal antibodies known to recognize specific immune cell subsets present in lymphoid tissues of sheep were tested for cross-reactivity with cervine RAMALT and mesenteric lymph nodes (MLNs) preserved in zinc salts fixative. The distribution of cells expressing CD3, CD4, CD79, CD21 and class II molecules of the major histocompatibility complex was determined in these tissues. Cells of each immunophenotype had similar distributions in RAMALT and MLNs and these distributions were similar to those reported previously for sheep and cattle. The identification and validation of cervine lymphoid follicle cell markers (CD79 and CD21) may allow reduction in false-negative results during diagnosis of CWD by ELISA.

Transfusion-related acute lung injury in a neutropenic patient.

Transfusion-related acute lung injury (TRALI) is a leading cause of transfusion related morbidity and mortality. Current concepts regarding the pathogenesis of this disorder imply a "two-hit" model in which neutrophils are sequestered in the pulmonary capillary bed, and subsequently activated by substances in the transfused blood product. We report a case of TRALI in a patient with neutropenia and discuss the possible factors contributing to the respiratory symptoms in this patient. We also emphasise the importance of recognising mild cases of TRALI in order to investigate the implicated donor/s appropriately, and to minimise the risk for more severe episodes in other patients.

Global IRS-1 phosphorylation analysis in insulin resistance.

AIMS/HYPOTHESIS: IRS-1 serine phosphorylation is often elevated in insulin resistance models, but confirmation in vivo in humans is lacking. We therefore analysed IRS-1 phosphorylation in human muscle in vivo. METHODS: We used HPLC-electrospray ionisation (ESI)-MS/MS to quantify IRS-1 phosphorylation basally and after insulin infusion in vastus lateralis muscle from lean healthy, obese non-diabetic and type 2 diabetic volunteers. RESULTS: Basal Ser323 phosphorylation was increased in type 2 diabetic patients (2.1 ± 0.43, p ≤ 0.05, fold change vs lean controls). Thr495 phosphorylation was decreased in type 2 diabetic patients (p ≤ 0.05). Insulin increased IRS-1 phosphorylation at Ser527 (1.4 ± 0.17, p ≤ 0.01, fold change, 60 min after insulin infusion vs basal) and Ser531 (1.3 ± 0.16, p ≤ 0.01, fold change, 60 min after insulin infusion vs basal) in the lean controls and suppressed phosphorylation at Ser348 (0.56 ± 0.11, p ≤ 0.01, fold change, 240 min after insulin infusion vs basal), Thr446 (0.64 ± 0.16, p ≤ 0.05, fold change, 60 min after insulin infusion vs basal), Ser1100 (0.77 ± 0.22, p ≤ 0.05, fold change, 240 min after insulin infusion vs basal) and Ser1142 (1.3 ± 0.2, p ≤ 0.05, fold change, 60 min after insulin infusion vs basal). CONCLUSIONS/INTERPRETATION: We conclude that, unlike some aspects of insulin signalling, the ability of insulin to increase or suppress certain IRS-1 phosphorylation sites is intact in insulin resistance. However, some IRS-1 phosphorylation sites do not respond to insulin, whereas other Ser/Thr phosphorylation sites are either increased or decreased in insulin resistance.

Increased abundance of the adaptor protein containing pleckstrin homology domain, phosphotyrosine binding domain and leucine zipper motif (APPL1) in patients with obesity and type 2 diabetes: evidence for altered adiponectin signalling.

AIMS/HYPOTHESIS: The adiponectin signalling pathway is largely unknown, but recently the adaptor protein containing pleckstrin homology domain, phosphotyrosine binding domain and leucine zipper motif (APPL1), has been shown to interact directly with adiponectin receptor (ADIPOR)1. APPL1 is present in C2C12 myoblasts and mouse skeletal muscle, but its presence in human skeletal muscle has not been investigated. METHODS: Samples from type 2 diabetic, and lean and non-diabetic obese participants were analysed by: immunoprecipitation and western blot; HPLC-electrospray ionisation (ESI)-mass spectrometry (MS) analysis; peak area analysis by MS; HPLC-ESI-MS/MS/MS analysis; and RT-PCR analysis of APPL1 mRNA. RESULTS: Immunoprecipitation and western blot indicated a band specific to APPL1. Tryptic digestion and HPLC-ESI-MS analysis of whole-muscle homogenate APPL1 unambiguously identified APPL1 with 56% sequence coverage. Peak area analysis by MS validated western blot results, showing APPL1 levels to be significantly increased in type 2 diabetic and obese as compared with lean participants. Targeted phosphopeptide analysis by HPLC-ESI-MS/MS/MS showed that APPL1 was phosphorylated specifically on Ser(401). APPL1 mRNA expression was significantly increased in obese and type 2 diabetic participants as compared with lean participants. After bariatric surgery in morbidly obese participants with subsequent weight loss, skeletal muscle APPL1 abundance was significantly reduced (p < 0.05) in association with an increase in plasma adiponectin (p < 0.01), increased levels of ADIPOR1 (p < 0.05) and increased muscle AMP-activated protein kinase (AMPK) phosphorylation (p < 0.05). CONCLUSIONS/INTERPRETATION: APPL1 abundance is significantly higher in type 2 diabetic muscle; APPL1 is phosphorylated in vivo on Ser(401). Improvements in hyperglycaemia and hypoadiponectinaemia following weight loss are associated with reduced skeletal muscle APPL1, and increased plasma adiponectin levels and muscle AMPK phosphorylation.