Breaking tolerance in transgenic mice expressing the human TSH receptor A-subunit: thyroiditis, epitope spreading and adjuvant as a 'double edged sword'.

Transgenic mice with the human thyrotropin-receptor (TSHR) A-subunit targeted to the thyroid are tolerant of the transgene. In transgenics that express low A-subunit levels (Lo-expressors), regulatory T cell (Treg) depletion using anti-CD25 before immunization with adenovirus encoding the A-subunit (A-sub-Ad) breaks tolerance, inducing extensive thyroid lymphocytic infiltration, thyroid damage and antibody spreading to other thyroid proteins. In contrast, no thyroiditis develops in Hi-expressor transgenics or wild-type mice. Our present goal was to determine if thyroiditis could be induced in Hi-expressor transgenics using a more potent immunization protocol: Treg depletion, priming with Complete Freund's Adjuvant (CFA) + A-subunit protein and further Treg depletions before two boosts with A-sub-Ad. As controls, anti-CD25 treated Hi- and Lo-expressors and wild-type mice were primed with CFA+ mouse thyroglobulin (Tg) or CFA alone before A-sub-Ad boosting. Thyroiditis developed after CFA+A-subunit protein or Tg and A-sub-Ad boosting in Lo-expressor transgenics but Hi- expressors (and wild-type mice) were resistant to thyroiditis induction. Importantly, in Lo-expressors, thyroiditis was associated with the development of antibodies to the mouse TSHR downstream of the A-subunit. Unexpectedly, we observed that the effect of bacterial products on the immune system is a "double-edged sword". On the one hand, priming with CFA (mycobacteria emulsified in oil) plus A-subunit protein broke tolerance to the A-subunit in Hi-expressor transgenics leading to high TSHR antibody levels. On the other hand, prior treatment with CFA in the absence of A-subunit protein inhibited responses to subsequent immunization with A-sub-Ad. Consequently, adjuvant activity arising in vivo after bacterial infections combined with a protein autoantigen can break self-tolerance but in the absence of the autoantigen, adjuvant activity can inhibit the induction of immunity to autoantigens (like the TSHR) displaying strong self-tolerance

The NOD mouse beyond autoimmune diabetes

Autoimmune diabetes arises spontaneously in Non-Obese Diabetic (NOD) mice, and the pathophysiology of this disease shares many similarities with human type 1 diabetes. Since its generation in 1980, the NOD mouse, derived from the Cataract Shinogi strain, has represented the gold standard of spontaneous disease models, allowing to investigate autoimmune diabetes disease progression and susceptibility traits, as well as to test a wide array of potential treatments and therapies. Beyond autoimmune diabetes, NOD mice also exhibit polyautoimmunity, presenting with a low incidence of autoimmune thyroiditis and Sjögren’s syndrome. Genetic manipulation of the NOD strain has led to the generation of new mouse models facilitating the study of these and other autoimmune pathologies. For instance, following deletion of specific genes or via insertion of resistance alleles at genetic loci, NOD mice can become fully resistant to autoimmune diabetes; yet the newly generated diabetes-resistant NOD strains often show a high incidence of other autoimmune diseases. This suggests that the NOD genetic background is highly autoimmune-prone and that genetic manipulations can shift the autoimmune response from the pancreas to other organs. Overall, multiple NOD variant strains have become invaluable tools for understanding the pathophysiology of and for dissecting the genetic susceptibility of organ-specific autoimmune diseases. An interesting commonality to all autoimmune diseases developing in variant strains of the NOD mice is the presence of autoantibodies. This review will present the NOD mouse as a model for studying autoimmune diseases beyond autoimmune diabetes

Exceptional Hyperthyroidism and a Role for both Major Histocompatibility Class I and Class II Genes in a Murine Model of Graves' Disease

Autoimmune hyperthyroidism, Graves' disease, can be induced by immunizing susceptible strains of mice with adenovirus encoding the human thyrotropin receptor (TSHR) or its A-subunit. Studies in two small families of recombinant inbred strains showed that susceptibility to developing TSHR antibodies (measured by TSH binding inhibition, TBI) was linked to the MHC region whereas genes on different chromosomes contributed to hyperthyroidism. We have now investigated TSHR antibody production and hyperthyroidism induced by TSHR A-subunit adenovirus immunization of a larger family of strains (26 of the AXB and BXA strains). Analysis of the combined AXB and BXA families provided unexpected insight into several aspects of Graves' disease. First, extreme thyroid hyperplasia and hyperthyroidism in one remarkable strain, BXA13, reflected an inability to generate non-functional TSHR antibodies measured by ELISA. Although neutral TSHR antibodies have been detected in Graves' sera, pathogenic, functional TSHR antibodies in Graves' patients are undetectable by ELISA. Therefore, this strain immunized with A-subunit-adenovirus that generates only functional TSHR antibodies may provide an improved model for studies of induced Graves' disease. Second, our combined analysis of linkage data from this and previous work strengthens the evidence that gene variants in the immunoglobulin heavy chain V region contribute to generating thyroid stimulating antibodies. Third, a broad region that encompasses the MHC region on mouse chomosome 17 is linked to the development of TSHR antibodies (measured by TBI). Most importantly, unlike other strains, TBI linkage in the AXB and BXA families to MHC class I and class II genes provides an explanation for the unresolved class I/class II difference in humans

Comparison of the sensitivity of the UKCAT and A levels to sociodemographic characteristics: a national study

Background: The UK Clinical Aptitude Test (UKCAT) was introduced to facilitate widening participation in medical and dental education in the UK by providing universities with a continuous variable to aid selection; one that might be less sensitive to the sociodemographic background of candidates compared to traditional measures of educational attainment. Initial research suggested that males, candidates from more advantaged socioeconomic backgrounds and those who attended independent or grammar schools performed better on the test. The introduction of the A* grade at A level permits more detailed analysis of the relationship between UKCAT scores, secondary educational attainment and sociodemographic variables. Thus, our aim was to further assess whether the UKCAT is likely to add incremental value over A level (predicted or actual) attainment in the selection process. Methods: Data relating to UKCAT and A level performance from 8,180 candidates applying to medicine in 2009 who had complete information relating to six key sociodemographic variables were analysed. A series of regression analyses were conducted in order to evaluate the ability of sociodemographic status to predict performance on two outcome measures: A level ‘best of three’ tariff score; and the UKCAT scores. Results: In this sample A level attainment was independently and positively predicted by four sociodemographic variables (independent/grammar schooling, White ethnicity, age and professional social class background). These variables also independently and positively predicted UKCAT scores. There was a suggestion that UKCAT scores were less sensitive to educational background compared to A level attainment. In contrast to A level attainment, UKCAT score was independently and positively predicted by having English as a first language and male sex. Conclusions: Our findings are consistent with a previous report; most of the sociodemographic factors that predict A level attainment also predict UKCAT performance. However, compared to A levels, males and those speaking English as a first language perform better on UKCAT. Our findings suggest that UKCAT scores may be more influenced by sex and less sensitive to school type compared to A levels. These factors must be considered by institutions utilising the UKCAT as a component of the medical and dental school selection process

Novel Information on the Epitope of an Inverse Agonist Monoclonal Antibody Provides Insight into the Structure of the TSH Receptor

The TSH receptor (TSHR) comprises an extracellular leucine-rich domain (LRD) linked by a hinge region to the transmembrane domain (TMD). Insight into the orientation of these components to each other is required for understanding how ligands activate the receptor. We previously identified residue E251 at the LRD-hinge junction as contributing to coupling TSH binding with receptor activation. However, a single residue cannot stabilize the LRD-hinge unit. Therefore, based on the LRD crystal structure we selected for study four other potential LRD-hinge interface charged residues. Alanine substitutions of individual residues K244, E247, K250 and R255 (as well as previously known E251A) did not affect TSH binding or function. However, the cumulative mutation of these residues in varying permutations, primarily K250A and R255A when associated with E251A, partially uncoupled TSH binding and function. These data suggest that these three residues, spatially very close to each other at the LRD base, interact with the hinge region. Unexpectedly and most important, monoclonal antibody CS-17, a TSHR inverse agonist whose epitope straddles the LRD-hinge, was found to interact with residues K244 and E247 at the base of the convex LRD surface. These observations, together with the functional data, exclude residues K244 and E247 from the TSHR LRD-hinge interface. Further, for CS-17 accessibility to K244 and E247, the concave surface of the TSHR LRD must be tilted forwards towards the hinge region and plasma membrane. Overall, these data provide insight into the mechanism by which ligands either activate the TSHR or suppress its constitutive activity

Angiotensin-(1–7) and the G Protein-Coupled Receptor Mas Are Key Players in Renal Inflammation

Angiotensin (Ang) II mediates pathophysiologial changes in the kidney. Ang-(1–7) by interacting with the G protein-coupled receptor Mas may also have important biological activities

Anti-Müllerian Hormone Serum Concentrations of Women with Germline BRCA1 or 2 BRCA2 Mutations

Study funding/competing interest(s): kConFab is supported by a grant from the Australian National Breast Cancer Foundation, and previously by the National Health and Medical Research Council (NHMRC), the Queensland Cancer Fund, the Cancer Councils of New South Wales, Victoria, Tasmania and South Australia, and the Cancer Foundation of Western Australia. KAP is an Australian National Breast Cancer Foundation Practitioner Fellow. JLH is a NHMRC Senior Principal Research Fellow. MH is a NHMRC Practitioner Fellow. RA reports personal fees from Roche Diagnostics & Beckman Coulter outside the submitted work and CS reports other from Melbourne IVF outside the submitted work. The remaining authors have nothing to declare and no conflicts of interest.Study question: Do women with BRCA1 and BRCA2 mutations have reduced ovarian reserve, as measured by circulating anti-müllerian hormone (AMH) concentration?  Summary answer: Women with a germline mutation in BRCA1 have reduced ovarian reserve as measured by AMH.  What is known already: The DNA repair enzymes encoded by BRCA1 and BRCA2 are implicated in reproductive aging. Circulating AMH is a biomarker of ovarian reserve and hence reproductive lifespan.  Study design, size, duration: Cross-sectional study of AMH concentrations of 693 women at the time of enrolment into the Kathleen Cuningham Foundation Consortium for research into Familial Breast Cancer (kConFab) cohort study (recruitment from 19/08/1997 until 18/9/2012). AMH was measured on stored plasma samples between November 2014 and January 2015 using an electrochemiluminescence immunoassay platform.  Participants/materials, setting, methods: Eligible women were from families segregating BRCA1 or BRCA2 mutations and had known mutation status. Participants were aged 25 to 45 years, had no personal history of cancer, retained both ovaries and were not pregnant or breastfeeding at the time of plasma storage. Circulating AMH was measured for 172 carriers and 216 non-carriers from families carrying BRCA1 mutations, and 147 carriers and 158 non-carriers from families carrying BRCA2 mutations. Associations between plasma AMH concentration and carrier status were tested by linear regression, adjusted for age at plasma storage, oral contraceptive use, body mass index and cigarette smoking.  Main results and the role of chance: Mean AMH concentration was negatively associated with age (P < 0.001). Mutation carriers were younger at blood draw than non-carriers (P ≤ 0.031). BRCA1 mutation carriers had, on average, 25% (95% CI: 5% - 41%, P = 0.02) lower AMH concentrations than non-carriers and were more likely to have AMH concentrations in the lowest quartile for age (OR 1.84, 95% CI: 1.11-303, P=0.02). There was no evidence of an association between AMH concentration and BRCA2 mutation status (P = 0.94).  Limitations, reasons for caution: The clinical implications of the lower AMH concentrations seen in BRCA1 mutation carriers cannot be assessed by this study design.  Wider implications of the findings: Women with a germline mutation in BRCA1 may have reduced ovarian reserve. This is consistent with other smaller studies in the literature and has potential implications for fertility and reproductive lifespan. Publisher PDFPeer reviewe

The Influence of Number and Timing of Pregnancies on Breast Cancer Risk for Women With BRCA1 or BRCA2 Mutations

International audienceBACKGROUND:Full-term pregnancy (FTP) is associated with a reduced breast cancer (BC) risk over time, but women are at increased BC risk in the immediate years following an FTP. No large prospective studies, however, have examined whether the number and timing of pregnancies are associated with BC risk for BRCA1 and BRCA2 mutation carriers.METHODS:Using weighted and time-varying Cox proportional hazards models, we investigated whether reproductive events are associated with BC risk for mutation carriers using a retrospective cohort (5707 BRCA1 and 3525 BRCA2 mutation carriers) and a prospective cohort (2276 BRCA1 and 1610 BRCA2 mutation carriers), separately for each cohort and the combined prospective and retrospective cohort.RESULTS:For BRCA1 mutation carriers, there was no overall association with parity compared with nulliparity (combined hazard ratio [HRc] = 0.99, 95% confidence interval [CI] = 0.83 to 1.18). Relative to being uniparous, an increased number of FTPs was associated with decreased BC risk (HRc = 0.79, 95% CI = 0.69 to 0.91; HRc = 0.70, 95% CI = 0.59 to 0.82; HRc = 0.50, 95% CI = 0.40 to 0.63, for 2, 3, and ≥4 FTPs, respectively, P trend < .0001) and increasing duration of breastfeeding was associated with decreased BC risk (combined cohort P trend = .0003). Relative to being nulliparous, uniparous BRCA1 mutation carriers were at increased BC risk in the prospective analysis (prospective hazard ration [HRp] = 1.69, 95% CI = 1.09 to 2.62). For BRCA2 mutation carriers, being parous was associated with a 30% increase in BC risk (HRc = 1.33, 95% CI = 1.05 to 1.69), and there was no apparent decrease in risk associated with multiparity except for having at least 4 FTPs vs. 1 FTP (HRc = 0.72, 95% CI = 0.54 to 0.98).CONCLUSIONS:These findings suggest differential associations with parity between BRCA1 and BRCA2 mutation carriers with higher risk for uniparous BRCA1 carriers and parous BRCA2 carriers

Worldwide trends in diabetes since 1980: a pooled analysis of 751 population-based studies with 4.4 million participants

BACKGROUND: One of the global targets for non-communicable diseases is to halt, by 2025, the rise in the age-standardised adult prevalence of diabetes at its 2010 levels. We aimed to estimate worldwide trends in diabetes, how likely it is for countries to achieve the global target, and how changes in prevalence, together with population growth and ageing, are affecting the number of adults with diabetes. METHODS: We pooled data from population-based studies that had collected data on diabetes through measurement of its biomarkers. We used a Bayesian hierarchical model to estimate trends in diabetes prevalence—defined as fasting plasma glucose of 7·0 mmol/L or higher, or history of diagnosis with diabetes, or use of insulin or oral hypoglycaemic drugs—in 200 countries and territories in 21 regions, by sex and from 1980 to 2014. We also calculated the posterior probability of meeting the global diabetes target if post-2000 trends continue. FINDINGS: We used data from 751 studies including 4 372 000 adults from 146 of the 200 countries we make estimates for. Global age-standardised diabetes prevalence increased from 4·3% (95% credible interval 2·4–7·0) in 1980 to 9·0% (7·2–11·1) in 2014 in men, and from 5·0% (2·9–7·9) to 7·9% (6·4–9·7) in women. The number of adults with diabetes in the world increased from 108 million in 1980 to 422 million in 2014 (28·5% due to the rise in prevalence, 39·7% due to population growth and ageing, and 31·8% due to interaction of these two factors). Age-standardised adult diabetes prevalence in 2014 was lowest in northwestern Europe, and highest in Polynesia and Micronesia, at nearly 25%, followed by Melanesia and the Middle East and north Africa. Between 1980 and 2014 there was little change in age-standardised diabetes prevalence in adult women in continental western Europe, although crude prevalence rose because of ageing of the population. By contrast, age-standardised adult prevalence rose by 15 percentage points in men and women in Polynesia and Micronesia. In 2014, American Samoa had the highest national prevalence of diabetes (>30% in both sexes), with age-standardised adult prevalence also higher than 25% in some other islands in Polynesia and Micronesia. If post-2000 trends continue, the probability of meeting the global target of halting the rise in the prevalence of diabetes by 2025 at the 2010 level worldwide is lower than 1% for men and is 1% for women. Only nine countries for men and 29 countries for women, mostly in western Europe, have a 50% or higher probability of meeting the global target. INTERPRETATION: Since 1980, age-standardised diabetes prevalence in adults has increased, or at best remained unchanged, in every country. Together with population growth and ageing, this rise has led to a near quadrupling of the number of adults with diabetes worldwide. The burden of diabetes, both in terms of prevalence and number of adults affected, has increased faster in low-income and middle-income countries than in high-income countries. FUNDING: Wellcome Trust