The lived experience of severe mental illness and long-term conditions: a qualitative exploration of service user, carer, and healthcare professional perspectives on self-managing co-existing mental and physical conditions.

BACKGROUND: People with severe mental illness (SMI), such as schizophrenia, have higher rates of physical long-term conditions (LTCs), poorer health outcomes, and shorter life expectancy compared with the general population. Previous research exploring SMI and diabetes highlights that people with SMI experience barriers to self-management, a key component of care in long-term conditions; however, this has not been investigated in the context of other LTCs. The aim of this study was to explore the lived experience of co-existing SMI and LTCs for service users, carers, and healthcare professionals. METHODS: A qualitative study with people with SMI and LTCs, their carers, and healthcare professionals, using semi-structured interviews, focused observations, and focus groups across the UK. Forty-one interviews and five focus groups were conducted between December 2018 and April 2019. Transcripts were coded by two authors and analysed thematically. RESULTS: Three themes were identified, 1) the precarious nature of living with SMI, 2) the circularity of life with SMI and LTCs, and 3) the constellation of support for self-management. People with co-existing SMI and LTCs often experience substantial difficulties with self-management of their health due to the competing demands of their psychiatric symptoms and treatment, social circumstances, and access to support. Multiple long-term conditions add to the burden of self-management. Social support, alongside person-centred professional care, is a key facilitator for managing health. An integrated approach to both mental and physical healthcare was suggested to meet service user and carer needs. CONCLUSION: The demands of living with SMI present a substantial barrier to self-management for multiple co-existing LTCs. It is important that people with SMI can access person-centred, tailored support for their LTCs that takes into consideration individual circumstances and priorities.

Estimated glucose disposal rate as a candidate biomarker for thrombotic biomarkers in T1D: a pooled analysis.

PURPOSE: To determine the utility of estimated glucose disposal rate (eGDR) as a candidate biomarker for thrombotic biomarkers in patients with type 1 diabetes (T1D). METHODS: We reanalysed baseline pretreatment data in a subset of patients with T1D from two previous RCTs, consisting of a panel of thrombotic markers, including fibrinogen, tissue factor (TF) activity, and plasminogen-activator inhibitor (PAI)-1, and TNFα, and clinical factors (age, T1D duration, HbA1c, insulin requirements, BMI, blood pressure, and eGDR). We employed univariate linear regression models to investigate associations between clinical parameters and eGDR with thrombotic biomarkers. RESULTS: Thirty-two patients were included [mean ± SD age 31 ± 7 years, HbA1c of 58 ± 9 mmol/mol (7.5 ± 0.8%), eGDR 7.73 ± 2.61]. eGDR negatively associated with fibrinogen (P < 0.001), PAI-1 concentrations (P = 0.005), and TF activity (P = 0.020), but not TNFα levels (P = 0.881). We identified 2 clusters of patients displaying significantly different characteristics; 56% (n = 18) were categorised as 'higher-risk', eliciting significantly higher fibrinogen (+ 1514 ± 594 µg/mL; P < 0.001), TF activity (+ 59.23 ± 9.42 pmol/mL; P < 0.001), and PAI-1 (+ 8.48 ± 1.58 pmol/dL; P < 0.001), HbA1c concentrations (+ 14.20 ± 1.04 mmol/mol; P < 0.001), age (+ 7 ± 3 years; P < 0.001), duration of diabetes (15 ± 2 years; P < 0.001), BMI (+ 7.66 ± 2.61 kg/m2; P < 0.001), and lower mean eGDR (- 3.98 ± 1.07; P < 0.001). CONCLUSIONS: Compared to BMI and insulin requirements, classical surrogates of insulin resistance, eGDR is a suitable and superior thrombotic risk indicator in T1D. TRIAL REGISTRATION: ISRCTN4081115; registered 27 June 2017.

Antiplatelet therapies in diabetes.

Cardiovascular complications remain the main cause of mortality and morbidity in diabetes. This is related to advanced vascular pathology in this population, together with an enhanced thrombotic environment. The increased risk in thrombosis is secondary to platelet hyper-reactivity and increased levels and/or altered activity of coagulation factors. The current review is focused on the role of antiplatelet agents in modulating the thrombotic milieu in diabetes and improving vascular outcome in this high-risk population. We review the latest evidence for the use of aspirin in primary vascular prevention together with long-term treatment with this agent for secondary prevention. We also discuss the effects of the various P2Y12 inhibitors, including clopidogrel, prasugrel and ticagrelor, on both short- and long-term secondary vascular prevention. Moreover, we briefly review antiplatelet therapies in special groups of people including those intolerant to aspirin, individuals with peripheral vascular disease and those with cerebrovascular pathology. The overall aim of this review is to provide the healthcare professional with a pragmatic guide for the management of thrombotic risk using established antiplatelet therapies to improve vascular outcome in persons with diabetes.

Characterizing adults with Type 2 diabetes mellitus and intellectual disability: outcomes of a case-finding study.

AIMS: To report the results of a case-finding study conducted during a feasibility trial of a supported self-management intervention for adults with mild to moderate intellectual disability and Type 2 diabetes mellitus, and to characterize the study sample in terms of diabetes control, health, and access to diabetes management services and support. METHODS: We conducted a cross-sectional case-finding study in the UK (March 2013 to June 2015), which recruited participants mainly through primary care settings. Data were obtained from medical records and during home visits. RESULTS: Of the 325 referrals, 147 eligible individuals participated. The participants' mean (sd) HbA1c concentration was 55 (15) mmol/mol [7.1 (1.4)%] and the mean (sd) BMI was 32.9 (7.9) kg/m2 , with 20% of participants having a BMI >40 kg/m2 . Self-reported frequency of physical activity was low and 79% of participants reported comorbidity, for example, cardiovascular disease, in addition to Type 2 diabetes. The majority of participants (88%) had a formal or informal supporter involved in their diabetes care, but level and consistency of support varied greatly. Post hoc exploratory analyses showed a significant association between BMI and self-reported mood, satisfaction with diet and weight. CONCLUSIONS: We found high obesity and low physical activity levels in people with intellectual disability and Type 2 diabetes. Glycaemic control was no worse than in the general Type 2 diabetes population. Increased risk of morbidity in this population is less likely to be attributable to poor glycaemic control and is probably related, at least in part, to greater prevalence of obesity and inactivity. More research, focused on weight management and increasing activity in this population, is warranted.

Vitamin D and diabetes mellitus: Causal or casual association?

The incidence of both type 2 and type 1 diabetes mellitus has been increasing worldwide. Vitamin D deficiency, or the awareness of its prevalence, has also been increasing. Vitamin D may have a role in the pathogenic mechanisms predisposing to type 2 diabetes by modulating insulin resistance and/or pancreatic ß-cell function. Vitamin D status or elements involved in its activation or transport may also be involved in the development of type 1 diabetes mellitus through immunomodulatory role . Based on these observations a potential association between vitamin D and diabetes has been hypothesized. In this review we discuss up to date evidence linking vitamin D with the development of diabetes. Moreover, the role of vitamin D supplementation in the prevention of both types of diabetes is analysed together with its role in improving glycemic control in diabetic patients. We also address the potential role of vitamin D deficiency in the development of macro- and microvascular complications in diabetes. Finally, we provide recommendation for Vitamin D therapy in diabetes in view of current evidence and highlight areas for potential future research in this area.

The Pathogenesis of Hashimoto's Thyroiditis: Further Developments in our Understanding.

Hashimoto's thyroiditis (HT) is part of a spectrum of thyroid autoimmune conditions and this review provides an update on the latest developments in the field. HT has a genetic predisposition with a number of immune-related and thyroid-specific genes conferring disease susceptibility. However, disentangling genes with protective and predisposing effect is a complex process that requires further work. The recent increase in the incidence of HT implicates environmental factors in disease pathogenesis including improved hygiene, increased dietary iodine intake, new treatment modalities and chemical agents. Additional unmodifiable predisposing factors include stress, climate, age and gender. Both cellular and humoral immunity play a role in HT pathogenesis. Defects in T regulatory cells and increased activation of follicular helper T cells may have a role in disease initiation/perpetuation. Infiltrating lymphocytes can be directly cytotoxic to thyroid follicular cells (TFC) or may affect cell viability/function indirectly through cytokine production, which alters TFC integrity and modulates their metabolic and immune function. Thyroid peroxidase and thyroglobulin antibodies are present in the majority of HT patients and help with management decisions. Antibodies against the sodium iodide symporter and pendrin are present in a minority with little known about their clinical relevance. In addition to immune cells, recent work has identified DNA fragments, generated following cell death, and micro RNA as potential factors in HT pathogenesis. Despite the large number of studies, the mechanistic pathways in HT are still not fully understood and further work is required to enhance our knowledge and identify novel preventative and therapeutic clinical targets.

Perspective of Continuous Glucose Monitoring-Based Interventions at the Various Stages of Type 2 Diabetes.

Continuous glucose monitoring (CGM) is now advocated for the clinical management of individuals with type 1 diabetes (T1D). However, this glucose monitoring strategy is not routinely used in type 2 diabetes (T2D), given the large population, significant cost implications and relatively limited supporting evidence. T2D is a more heterogenous condition compared with T1D with various glucose lowering therapies that do not necessarily require CGM to ensure within target glucose levels. While all individuals with T2D may benefit from CGM at certain time points, the whole T2D population does not necessarily require this technology continuously, which should be prioritized based on patient benefit and cost effectiveness. In this pragmatic opinion piece, we describe the rationale and evidence for CGM use in different subgroups of individuals with T2d, divided according to the stage of the condition, glycemic therapies, presence of diabetes complications, or associated co-morbidities. We discuss a total of 16 T2D subgroups and provide a clinical view on CGM use in each, based on current evidence while also highlighting areas of knowledge gaps. This work provides health care professionals with a simple guide to CGM use in different T2D groups and gives suggestion for future studies to justify expansion of this technology.

Polycystic ovary syndrome has no independent effect on vascular, inflammatory or thrombotic markers when matched for obesity.

INTRODUCTION: Previous studies investigating cardiovascular (CV) risk in obese women with polycystic ovary syndrome (PCOS) have been potentially confounded by not adequately accounting for body weight. OBJECTIVE: To assess if PCOS increases CV risk independently in young obese women by examining carotid intima-media wall thickness (cIMT) and platelet function. DESIGN: A case-control study comparing women with PCOS (n = 21) to age (32·8 ± 7·2 vs 33·5 ± 6·7 years), and weight (100·9 ± 16·7 vs 99·3 ± 14·7 kg)-matched controls (n = 19). Platelet function was examined by flow cytometry, clot structure and fibrinolysis by turbidimetric assays and endothelial function by ELISA and post ischaemic reactive hyperaemia. RESULTS: The PCOS group had higher testosterone 1·2 ± 0·3 vs 0·9 ± 0·3 nmol/l (P = 0·01), HOMA-IR 2·5 ± 1·7 vs 1·7 ± 1·0 (P = 0·08), impaired glucose regulation 33·3% vs 5·3% (P = 0·02), and urinary isoprostane 16·0 ± 4·4 vs 11·8 ± 7·1 ng/ml (P = 0·04) compared to controls. Mean cIMT 0·5 ± 0·05 vs 0·48 ± 0·06 mm (P = 0·36), and basal platelet surface expression (percentage of positive cells) of P-selectin 0·52 ± 0·3 vs 0·43 ± 0·23 (P = 0·40) and fibrinogen binding 0·97 ± 0·4 vs 0·83 ± 0·3 (P = 0·48) did not significantly differ between the PCOS and control groups respectively. Furthermore, platelets sensitivity to stimulation with adenosine-5'-diphosphate or inhibition with prostacyclin, clot structure and fibrinolytic efficiency ex vivo, endothelial reactive hyperaemic index (RHI), inflammation (hsCRP) and adhesion markers (sE-selectin, sP-selectin, sVCAM-1 and sICAM-1) were not significantly different between the two groups. CONCLUSIONS: PCOS appeared not to independently increase atherothrombotic risk when matched for obesity. It is likely that any excess CV risk in young obese women with PCOS can either be attributed to obesity or is not yet apparent at this early stage of the condition.

A novel mechanism for hypofibrinolysis in diabetes: the role of complement C3.

AIMS/HYPOTHESIS: Impaired fibrin clot lysis is a key abnormality in diabetes and complement C3 is one protein identified in blood clots. This work investigates the mechanistic pathways linking C3 and hypofibrinolysis in diabetes using ex vivo/in vitro studies. METHODS: Fibrinolysis and C3 plasma levels were determined in type 1 diabetic patients and healthy controls, and the effects of glycaemia investigated. C3 incorporation into fibrin clots and modulation of fibrinolysis were analysed by ELISA, immunoblotting, turbidimetric assays and electron and confocal microscopy. RESULTS: Clot lysis time was longer in diabetic children than in controls (599 ± 18 and 516 ± 12 s respectively; p < 0.01), C3 levels were higher in diabetic children (0.55 ± 0.02 and 0.43 ± 0.02 g/l respectively; p < 0.01) and both were affected by improving glycaemia. An interaction between C3 and fibrin was confirmed by the presence of lower protein levels in sera compared with corresponding plasma and C3 detection in plasma clots by immunoblot. In a purified system, C3 was associated with thinner fibrin fibres and more prolongation of lysis time of clots made from fibrinogen from diabetic participants compared with controls (244 ± 64 and 92 ± 23 s respectively; p < 0.05). Confocal microscopy showed higher C3 incorporation into diabetic clots compared with controls, and fully formed clot lysis was prolonged by 764 ± 76 and 428 ± 105 s respectively (p < 0.05). Differences in lysis, comparing diabetes and controls, were not related to altered plasmin generation or C3-fibrinogen binding assessed by plasmon resonance. CONCLUSIONS/INTERPRETATION: C3 incorporation into clots from diabetic fibrinogen is enhanced and adversely affects fibrinolysis. This may be one novel mechanism for compromised clot lysis in diabetes, potentially offering a new therapeutic target.

Oral 11ß-HSD1 inhibitor AZD4017 improves wound healing and skin integrity in adults with type 2 diabetes mellitus: a pilot randomized controlled trial.

BACKGROUND: Chronic wounds (e.g. diabetic foot ulcers) reduce the quality of life, yet treatments remain limited. Glucocorticoids (activated by the enzyme 11ß-hydroxysteroid dehydrogenase type 1, 11ß-HSD1) impair wound healing. OBJECTIVES: Efficacy, safety, and feasibility of 11ß-HSD1 inhibition for skin function and wound healing. DESIGN: Investigator-initiated, double-blind, randomized, placebo-controlled, parallel-group phase 2b pilot trial. METHODS: Single-center secondary care setting. Adults with type 2 diabetes mellitus without foot ulcers were administered 400 mg oral 11ß-HSD1 inhibitor AZD4017 (n = 14) or placebo (n = 14) bi-daily for 35 days. Participants underwent 3-mm full-thickness punch skin biopsies at baseline and on day 28; wound healing was monitored after 2 and 7 days. Computer-generated 1:1 randomization was pharmacy-administered. Analysis was descriptive and focused on CI estimation. Of the 36 participants screened, 28 were randomized. RESULTS: Exploratory proof-of-concept efficacy analysis suggested AZD4017 did not inhibit 24-h ex vivoskin 11ß-HSD1 activity (primary outcome; difference in percentage conversion per 24 h 1.1% (90% CI: -3.4 to 5.5) but reduced systemic 11ß-HSD1 activity by 87% (69-104%). Wound diameter was 34% (7-63%) smaller with AZD4017 at day 2, and 48% (12-85%) smaller after repeat wounding at day 30. AZD4017 improved epidermal integrity but modestly impaired barrier function. Minimal adverse events were comparable to placebo. Recruitment rate, retention, and data completeness were 2.9/month, 27/28, and 95.3%, respectively. CONCLUSION: A phase 2 trial is feasible, and preliminary proof-of-concept data suggests AZD4017 warrants further investigation in conditions of delayed healing, for example in diabetic foot ulcers. SIGNIFICANCE STATEMENT: Stress hormone activation by the enzyme 11ß-HSD type 1 impairs skin function (e.g. integrity) and delays wound healing in animal models of diabetes, but effects in human skin were previously unknown. Skin function was evaluated in response to treatment with a 11ß-HSD type 1 inhibitor (AZD4017), or placebo, in people with type 2 diabetes. Importantly, AZD4017 was safe and well tolerated. This first-in-human randomized, controlled, clinical trial found novel evidence that 11ß-HSD type 1 regulates skin function in humans, including improved wound healing, epidermal integrity, and increased water loss. Results warrant further studies in conditions of impaired wound healing, for example, diabetic foot ulcers to evaluate 11ß-HSD type 1 as a novel therapeutic target forchronic wounds.

Effects of aspirin on clot structure and fibrinolysis using a novel in vitro cellular system.

OBJECTIVES: The purpose of this study was to investigate the direct effects of aspirin on fibrin structure/function. METHODS AND RESULTS: Chinese Hamster Ovary cell lines stably transfected with fibrinogen were grown in the absence (0) and presence of increasing concentrations of aspirin. Fibrinogen was purified from the media using affinity chromatography, and clots were made from recombinant protein. Mean final turbidity [OD(+/-SEM)] was 0.083(+/-0.03), 0.093(+/-0.002), 0.101(+/-0.005), and 0.125(+/-0.003) in clots made from 0, 1, 10, and 100 mg/L aspirin-treated fibrinogen, respectively (P<0.05). Permeability coefficient (Ks cm2 x 10(-8)) was 1.68(+/-0.29) and 4.13(+/-0.33) comparing fibrinogen produced from cells grown with 0 mg/L and 100 mg/L aspirin respectively (P<0.05). Scanning electron microscopy confirmed a looser clot structure and increased fiber thickness of clots made from aspirin-treated fibrinogen, whereas rheometer studies showed a significant 30% reduction in clot rigidity. Fibrinolysis was quicker in clots made from aspirin-treated fibrinogen. Ex vivo studies in 3 normal volunteers given 150 mg aspirin daily for 1 week demonstrated similar changes in clot structure/function. CONCLUSION: Aspirin directly altered clot structure resulting in the formation of clots with thicker fibers and bigger pores, which are easier to lyse. This study clearly demonstrates an alternative mode of action for aspirin, which should be considered in studies evaluating the biochemical efficacy of this agent.

Maturity onset diabetes of the young and fibrin-related thrombosis risk.

BACKGROUND: Fibrin network characteristics determine predisposition to cardiovascular disease (CVD). Individuals with type 1 (T1DM) and type 2 diabetes mellitus (T2DM) have higher risk of CVD and display deranged fibrin network structure. Those with maturity onset diabetes of the young (MODY) may also be at increased risk but their fibrin clot properties have not been studied. METHODS: Plasma clots properties from 13 individuals with HNF1A-MODY, 12 matched-individuals with T2DM and 12 with T1DM were studied using a validated turbidimetric assay and confocal microscopy. Plasma levels of fibrinogen, plasminogen activator inhibitor-1, complement C3 and C-reactive protein were also measured. RESULTS: MODY clot maximum absorbance was 0.37 ± 0.03 AU, similar to T1DM (0.32 ± 0.03 AU; p = 0.26), but lower than T2DM (0.49 ± 0.03 AU; p = 0.02), with confocal microscopy confirming structural differences. Clot lysis time in MODY was similar to T1DM (456 ± 50 and 402 ± 20 s, respectively; p = 0.09) but shorter than T2DM (588 ± 58 s; p = 0.006). Comparing inflammatory/thrombotic proteins in HNF1A-MODY and T2DM, C3 levels were lower in MODY than T2DM (0.58 ± 0.09 and 0.80 ± 0.1 mg/ml, respectively; p < 0.01). CONCLUSIONS: HNF1A-MODY fibrin network alterations are at least as pronounced as in T1DM but less thrombotic than T2DM clots. Differences in fibrin clot characteristics comparing HNF1A-MODY and T2DM may, in part, relate to lower C3 levels.

Techniques to quantify TSH receptor antibodies.

The presence of antibodies to TSH receptor (TSHR) is the hallmark of Graves disease (GD). These antibodies mimic the action of TSH, resulting in TSHR stimulation and hyperthyroidism, and have been associated with GD-associated extrathyroidal manifestations. TSH binding inhibition assays and bioassays for measurement of TSHR antibody levels have been used for clinical and research purposes. In the former, inhibition of TSH binding to purified or recombinant TSHR by a patient's immunoglobulins is measured by radioactive or chemiluminescent techniques. In the latter, cyclic AMP production is measured by use of radioimmunoassays or chemiluminescent methods in cells natively or artificially expressing TSHR. In this Review, the different techniques used for the detection of antibodies to TSHR are discussed, together with the clinical applications of antibody measurement, including diagnosis of GD and Graves ophthalmopathy. Prediction of relapse after medical treatment and the clinical course of Graves ophthalmopathy are also addressed.

Aspirin therapy is associated with less compact fibrin networks and enhanced fibrinolysis in patients with abdominal aortic aneurysm.

OBJECTIVE: Thrombotic changes in fibrin networks contribute to increased cardiovascular risk in patients with abdominal aortic aneurysm (AAA). Given that aspirin modulates the fibrin network, we aimed to determine if aspirin therapy is associated with changes in ex-vivo fibrin clot characteristics in AAA patients and also conducted an exploratory analysis of 5-year mortality in these individuals. METHODS: We recruited 145 male patients, divided into controls (aortic diameter < 3 cm, n = 49), AAA not taking aspirin (AAA-Asp, n = 50) and AAA on 75 mg day(-1) aspirin (AAA+Asp, n = 46), matched for aneurysm size. Characteristics of clots made from plasma and plasma-purified fibrinogen were investigated using turbidimetric analysis, permeation studies, and confocal and electron microscopy. Plasma fibrinogen, D-dimer and inflammatory marker levels were also measured. RESULTS: Maximum absorbance (MA) of plasma clots from controls was lower than that of AAA patients not on aspirin (AAA-Asp) at 0.30 ± 0.01 and 0.38 ± 0.02 au, respectively (P = 0.002), whereas aspirin-treated subjects had MA similar to controls (0.31 ± 0.02 P = 0.9). Plasma clot lysis time displayed an identical pattern at 482 ± 15, 597 ± 24 and 517 ± 27 s for control, AAA-Asp and AAA+Asp (P = 0.001 and P = 0.8). The lysis time of clots made from purified fibrinogen of AAA-Asp was longer than that of AAA+Asp patients (756 ± 47 and 592 ± 52 s, respectively; P = 0.041). Permeation studies and confocal and electron microscopy showed increased clot density in AAA-Asp compared with the AAA+Asp group. Mortality in AAA-Asp and AAA+Asp was similar, despite increased cardiovascular risk in the latter group, and both exhibited higher mortality than controls. CONCLUSION: Aspirin improves fibrin clot characteristics in patients with AAA, which may have important clinical implications.

Detection of IL-12, IL-13, and IL-15 messenger ribonucleic acid in the thyroid of patients with autoimmune thyroid disease.

IL-12, IL-13, and IL-15 are important cytokines that have not been studied previously in human autoimmune thyroid diseases. By applying RT-PCR on RNA extracted from tissue samples, we have investigated in vivo gene expression of these cytokines in multinodular goiter (MNG), Graves' disease (GD), and Hashimoto's thyroiditis (HT). In addition, in vitro studies were carried out using the transformed human thyroid cell line, HT-ori3, and primary thyroid cell cultures derived from patients with GD. These cells were used either unstimulated or stimulated for 12 h with TSH, IL-1, or interferon-gamma (IFN-gamma). IL-12 p40 gene expression was identified in 2 of 10 MNG samples, 6 of 12 GD, and 3 of 4 HT. HT-ori3 and primary thyroid cell cultures were positive for IL-12 p40 messenger RNA (mRNA) expression in both unstimulated and stimulated cell cultures. IL-13 mRNA was expressed in 2 MNG, 9 GD, and 1 HT sample. Both HT-ori3 and primary thyroid cultures expressed IL-13 after TSH, IL-1, or IFN-gamma stimulation; unstimulated primary cultures of thyroid cells, however, did not express IL-13. IL-15 gene expression was detected in 8 MNG, 8 GD, and 4 HT samples. HT-ori3 and primary thyroid cell cultures, stimulated with TSH, IL-1, or IFN-gamma, showed expression of this cytokine. Unstimulated cells showed only a weak expression. Our results indicate that the cytokine patterns in the various diseases studied are heterogeneous; that thyroid cells can express IL-12, IL-13, and IL-15 mRNA in culture, particularly after TSH, IL-1, or IFN-gamma stimulation; and that IL-15 is expressed in most of the tissue samples studied.

Detection of binding and blocking autoantibodies to the human sodium-iodide symporter in patients with autoimmune thyroid disease.

The sodium iodide symporter (NIS) is a novel autoantigen in autoimmune thyroid disease (ATD). A recent study has described the development of a bioassay for human (h) NIS antibody detection, but this will not detect antibodies that bind the symporter without modulating its activity. Therefore, the establishment of a binding assay is of importance to determine the overall prevalence of hNIS antibodies in ATD patients. An in vitro transcription and translation system was used to produce [35S]-labeled hNIS. The radiolabeled ligand reacted specifically in immunoprecipitation experiments with rabbit antiserum raised against a peptide fragment of hNIS. Subsequently, the reactivity of control and ATD sera to translated [35S]hNIS was determined using RIAs. A significant difference in the frequency of hNIS antibody-positive sera was found when patients with either Graves' disease (GD) or autoimmune hypothyroidism (AH) were compared with normal controls (P = 0.01 and P = 0.03, respectively). Of 49 GD and 29 AH sera tested, 11 (22%) and 7 (24%), respectively, were found to contain hNIS antibodies. Differences were also significant when the antibody-binding indices of the control group of sera were compared with those of both the GD and the AH patient sera (P < 0.0001 and P = 0.001, respectively). In contrast, sera from 10 patients with Addison's disease and 10 patients with vitiligo (without associated ATD) were all negative for antibody reactivity to the symporter. No differences were detected when the antibody binding indices of either the Addison's disease or the vitiligo sera were compared with those of the normal sera group (P = 0.9 and P = 0.6, respectively). Eight of the 11 (73%) GD and 3 of the 7 (43%) AH sera, which were positive for hNIS antibodies in the immunoprecipitation assay, were also found to inhibit iodide uptake in hNIS-transfected CHO-K1 cells, suggesting the existence of antibodies in some serum samples that bind to the symporter without modulating its function. Overall, a significant correlation was found between the iodide uptake inhibition and the binding assays for hNIS antibody detection (r = 0.49, P < 0.0001). In summary, we have developed a specific and quantitative assay for the detection of hNIS binding antibodies in sera of patients with ATD. This system offers the advantage of studying antibody reactivity against conformational epitopes and will be useful in understanding the role of NIS autoreactivity in the pathogenesis of ATD.

Tissue iodine content and serum-mediated 125I uptake-blocking activity in breast cancer.

In the thyroid, active transport of iodide is under control of the TSH-dependent Na+/I- symporter (NIS), whereas in the breast such control is less well understood. In this study, NIS expression was demonstrated by RT-PCR in 2 of 2 fibroadenomata and 6 of 7 breast carcinoma messenger ribonucleic acid isolates. In addition, mean total tissue iodine levels of 80.9 +/- 9.5 ng I/mg protein in 23 benign tumors (fibroadenomata) were significantly higher than those in 19 breast cancers taken from either the tumor (18.2 +/- 4.6 ng I/mg) or morphologically normal tissue taken from within the tumor-bearing breast (31.8 +/- 4.9 ng I/mg; P < 0.05 in each case). Inhibition of 125I uptake into NIS-transfected CHO cells was observed in serum from 20 of 105 (19.0%) breast carcinoma, 8 of 49 (16.3%) benign breast disease, and 27 of 86 (31.4%) Graves' patients, but in only 1 of 33 (3.0%) age-matched female controls. IgG purified from serum of patients showing positive 125I uptake inhibition also inhibited iodide uptake, suggesting that such inhibition was antibody mediated. 125I uptake inhibition was significantly associated with thyroid peroxidase antibody positivity (P < 0.05) in sera from breast cancer patients, but not in those with benign breast disease, once again suggesting an association between thyroid autoimmunity and breast carcinoma.

The modulation of the human sodium iodide symporter activity by Graves' disease sera.

The transport of iodide into the thyroid, catalyzed by the Na+/I- symporter (NIS), is the initial and rate-limiting step in the formation of thyroid hormones. To study the basic characteristics of the human (h) NIS, we have established a Chinese hamster ovary cell line stably expressing the hNIS (CHO-NIS9). In agreement with previous work on the rat NIS, iodide uptake in these cells was initiated within 2 min of the addition of 131I, reaching a plateau after 30 min. Both perchlorate and thiocyanate inhibited iodide uptake in a dose-dependent manner, with inhibition evident at concentrations of 0.01 and 0.1 micromol/L, respectively, and reaching complete inhibition at 20 micromol/L and 500 micromol/L, respectively. Ouabain, which blocks the activity of the Na+/K+ adenosine triphosphatase, also inhibited iodide uptake in a dose-dependent manner, starting at concentrations of 100 micromol/L and reaching maximum inhibition at 1600 micromol/L, indicating that iodide uptake in these cells is sodium dependent. CHO-NIS9 cells were further used to study 88 sera from patients with Graves' disease, for iodide uptake inhibitory activity, which were compared with sera from 31 controls. Significant iodide uptake inhibition was taken as any inhibition in excess of the mean + 3 SD of the results with the control sera. On this basis, 27 (30.7%) of the Graves' sera, but none of the controls, inhibited iodide uptake in CHO-NIS9. IgGs from these patients also inhibited iodide uptake, indicating that this inhibitory activity was antibody mediated. In summary, we have established a CHO cell line stably expressing the hNIS and shown that antibodies in GD sera can inhibit iodide uptake in these cells. This further emphasizes the role of NIS as a novel autoantigen in thyroid immunity.

The sodium iodide symporter gene and its regulation by cytokines found in autoimmunity.

Iodide concentration by the thyroid gland, an essential step for thyroid hormone synthesis, is mediated by the Na+/I- symporter (NIS). To identify factors that may regulate this process, we have studied NIS gene expression in the Fisher rat thyroid cell line (FRTL-5) by a semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) technique. Increasing concentrations of bovine TSH (0.1, 1, 10, 50 and 100 mU/l), with or without tumour necrosis factor-alpha (TNF alpha), interferon-gamma (IFN gamma) or interleukin-1 alpha (IL-1 alpha) were added to FRTL-5 cells previously deprived of TSH for a minimum of 5 days. RNA was extracted and samples were studied for NIS expression. TSH enhanced NIS mRNA expression in a dose-dependent manner, with induction evident at 0.1 mU/l, reaching a peak at 50 mU/l, an effect detected after 6 h of stimulation, but not in the first 2 h. Both TNF alpha and, to a lesser extent, IL-1 alpha inhibited basal and TSH-induced NIS expression. High concentrations of IFN gamma also downregulated TSH-stimulated NIS mRNA expression. Using the same technique, we also investigated NIS mRNA tissue distribution in two male and one female Wistar rats. High levels of NIS expression were detected in the thyroid, stomach, and mammary gland, lower levels were found in the intestine, adipose tissue and liver, borderline levels were expressed in the salivary gland, and no expression was detected in the kidneys. In summary, we have shown that TSH upregulates rat NIS gene expression in vitro, and this induction can be modulated by cytokines. Analysis of the distribution of rat NIS mRNA ex vivo demonstrated variable levels of NIS transcription in different tissue samples.

Once versus divided daily dosing with delayed-release mesalazine: a study of tissue drug concentrations and standard pharmacokinetic parameters.

BACKGROUND: Delayed-release mesalazine is traditionally taken as three divided doses. However, it is well-recognized that dosing frequency has a significant impact on compliance and that once daily dosing is preferable. METHODS: We measured serum, urinary, faecal and rectal tissue concentrations of 5-aminosalicylic acid and N-acetyl 5-aminosalicylic acid in 24 healthy volunteers following dosing with delayed-release mesalazine, 1.2 g or 2.4 g daily, given as either a single daily dose at 08:00 hours or in three divided doses at 08:00, 13:00 and 18:00 hours. RESULTS: Urinary and faecal excretion and rectal tissue concentrations of 5-aminosalicylic acid and N-acetyl 5-aminosalicylic acid were similar following single or divided daily dosing, at both doses studied. Peak serum concentrations were found at 06:00-09:00 following divided dosing and at 17:00-20:00 following once daily dosing. However, peak and trough serum levels and serum area under curve values (AUC) were similar with both regimens and at both doses. CONCLUSIONS: Urinary, faecal and rectal tissue concentrations are similar following single or divided daily dosing. Minor differences in serum levels were apparent but maximum, minimum and AUC values were similar. Clinical trials should examine the efficacy and toxicity of once daily dosing in patients with ulcerative colitis.