Functional rare and low frequency variants in BLK and BANK1 contribute to human lupus.

Systemic lupus erythematosus (SLE) is the prototypic systemic autoimmune disease. It is thought that many common variant gene loci of weak effect act additively to predispose to common autoimmune diseases, while the contribution of rare variants remains unclear. Here we describe that rare coding variants in lupus-risk genes are present in most SLE patients and healthy controls. We demonstrate the functional consequences of rare and low frequency missense variants in the interacting proteins BLK and BANK1, which are present alone, or in combination, in a substantial proportion of lupus patients. The rare variants found in patients, but not those found exclusively in controls, impair suppression of IRF5 and type-I IFN in human B cell lines and increase pathogenic lymphocytes in lupus-prone mice. Thus, rare gene variants are common in SLE and likely contribute to genetic risk.

Heterogeneity of Human Neutrophil CD177 Expression Results from CD177P1 Pseudogene Conversion.

Most humans harbor both CD177neg and CD177pos neutrophils but 1-10% of people are CD177null, placing them at risk for formation of anti-neutrophil antibodies that can cause transfusion-related acute lung injury and neonatal alloimmune neutropenia. By deep sequencing the CD177 locus, we catalogued CD177 single nucleotide variants and identified a novel stop codon in CD177null individuals arising from a single base substitution in exon 7. This is not a mutation in CD177 itself, rather the CD177null phenotype arises when exon 7 of CD177 is supplied entirely by the CD177 pseudogene (CD177P1), which appears to have resulted from allelic gene conversion. In CD177 expressing individuals the CD177 locus contains both CD177P1 and CD177 sequences. The proportion of CD177hi neutrophils in the blood is a heritable trait. Abundance of CD177hi neutrophils correlates with homozygosity for CD177 reference allele, while heterozygosity for ectopic CD177P1 gene conversion correlates with increased CD177neg neutrophils, in which both CD177P1 partially incorporated allele and paired intact CD177 allele are transcribed. Human neutrophil heterogeneity for CD177 expression arises by ectopic allelic conversion. Resolution of the genetic basis of CD177null phenotype identifies a method for screening for individuals at risk of CD177 isoimmunisation.

Classification, epidemiology and clinical subgrouping of antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis.

It is now 25 years since the first European studies on vasculitis--the anti-neutrophil cytoplasmic antibody (ANCA) standardization project. Over that period of time, there have been major developments in the classification of the vasculitides, which has permitted the conduct of high-quality epidemiology studies. Studying the epidemiology of rare diseases such as the ANCA-associated vasculitides (AAV) poses considerable challenges to epidemiologists. The first is the need for a clear definition of a case with good differentiation from similar disorders. The second is case capture. The vasculitides are rare, and therefore, a large population is required to determine the incidence and prevalence, and this poses questions of feasibility. A large population increases the risk of incomplete case detection but permits a reasonable number of cases to be collected in a practicable time frame, whereas a smaller population requires a much longer time frame to collect the necessary cases, which may also not be feasible. Statistical methods of capture-recapture analysis enable estimates to be made of the number of missing cases. The third is case ascertainment. The AAV are virtually always managed in secondary care, and therefore, hospital-based case ascertainment may be appropriate. Fourthly, the rarity of the conditions makes prospective case-control studies investigating risk factors difficult to conduct because the population size required to achieve statistical confidence is in excess of that which is readily available. Thus, much of the data on risk factors are derived from retrospective studies with inherent potential bias.

Vitamin D deficiency and risk for rheumatic diseases: an update.

PURPOSE OF REVIEW: The role of vitamin D in situations other than calcium homeostasis and bone health has become very topical. It is apparent that vitamin D has significant effects on the immune system and as such may contribute to the pathogenesis of autoimmune disease. This review examines the evidence-to-date that vitamin D has a role in immune-mediated rheumatic disorders. RECENT FINDINGS: Low vitamin D status is reported in many inflammatory rheumatic conditions. In some this extends to an association with disease activity. Vitamin D acts on a number of cells involved in both innate and acquired immunity biasing the adaptive immune system away from Th17 and Th1, towards Th2 and Tregs. Deficiency accordingly could encourage autoimmunity. Direct evidence for this plausible mechanism in specific diseases remains largely to be demonstrated. To date, there is a dearth of controlled trials of vitamin D in prophylaxis or therapy. SUMMARY: Vitamin D deficiency may well be an important factor in autoimmune rheumatic disease, including initial disease development and worsening the disease once present. This is testable and there is a pressing need for therapeutic studies.

Autoimmunity in primary antibody deficiency is associated with protein tyrosine phosphatase nonreceptor type 22 (PTPN22).

BACKGROUND: The 1858T allele of protein tyrosine phosphatase nonreceptor type 22 (PTPN22; R620W) exhibits one of the strongest and most consistent associations with sporadic autoimmune disease. Although autoimmunity is common in patients with primary antibody deficiency (PAD), it remains unknown whether its pathogenesis is similar when it arises in this context compared with in immunocompetent patients. OBJECTIVE: We set out to determine whether the 1858T allele of PTPN22 was associated with PAD or with autoimmunity in the context of PAD. METHODS: We genotyped rs2476601 (g.1858C>T), a single nucleotide polymorphism encoding substitution of arginine for tryptophan in PTPN22 (R620W), in 193 patients with PAD and 148 control subjects from an Australian cohort. We also performed a subgroup analysis according to the presence of autoimmunity and B-cell phenotypes. RESULTS: C/T and T/T PTPN22 genotypes were more common in patients with PAD than in the matched control subjects (C/T, 18.1% vs 9.5%; T/T, 1.04% vs 0.6%). The T allele was associated with an increased risk of PAD relative to control subjects (odds ratio, 2.10; 95% CI, 1.11-4.00). The distribution of genotypes in control subjects was similar to those reported previously and did not deviate significantly from Hardy-Weinberg equilibrium. We found a strong association between the 1858T allele and PAD with coexistent autoimmune diseases. In patients with PAD and autoimmunity, 16 (43.2%) of 37 had at least one T allele of PTPN22 compared with 27 (17.3%) of 156 with the C/C genotype (P=.0014; odds ratio, 3.64; 95% CI, 1.68-7.88). We found no evidence that this effect was mediated by enrichment of CD21low B cells. CONCLUSION: The 1858T PTPN22 allele is strongly associated with autoimmunity in patients with PAD.

Unusually indolent MPO-ANCA: associated vasculitis-report of two cases.

Vasculitis associated with antineutrophil cytoplasmic antibodies specific to myeloperoxidase generally presents as a life- and organ-threatening disease that evolves over several months. It is a syndrome in which prompt diagnosis and therapy are important both in terms of short-term survival and long-term organ damage. Two cases with quite a different course, sustained and indolent with limited progression over many years, are described in this report. They are compared to cases in the literature. Indolent cases of granulomatous polyangiitis associated with antineutrophil cytoplasmic antibodies against proteinase-3 are well recognised, but these two cases of microscopic polyangiitis are almost unique.

Analysis of complement expression in light-induced retinal degeneration: synthesis and deposition of C3 by microglia/macrophages is associated with focal photoreceptor degeneration.

PURPOSE: To investigate the expression and localization of complement system mRNA and protein in a light-induced model of progressive retinal degeneration. METHODS: Sprague-Dawley (SD) rats were exposed to 1000 lux of bright continuous light (BCL) for up to 24 hours. At time points during (1-24 hours) and after (3 and 7 days) exposure, the animals were euthanatized and the retinas processed. Differential expression of complement genes at 24 hours of exposure was assessed using microarray analysis. Expression of complement genes was validated by quantitative PCR, and expression of selected genes was investigated during and after BCL exposure. Photoreceptor apoptosis was assessed using TUNEL and C3 was further investigated by spatiotemporal analysis using in situ hybridization and immunohistochemistry. RESULTS: Exposure to 24 hours of BCL induced differential expression of a suite of complement system genes, including classic and lectin components, regulators, and receptors. C1qr1, MCP, Daf1, and C1qTNF6 all modulated in concert with photoreceptor death and AP-1 expression, which reached a peak at 24 hours exposure. C1s and C4a reached peak expression at 3 days after exposure, while expression of C3, C3ar1, and C5r1 were maximum at 7 days after exposure. C3 mRNA was detected in ED1- and IBA1-positive microglia/macrophages, in the retinal vessels and optic nerve head and in the subretinal space, particularly at the margins of the emerging lesion. CONCLUSIONS: The data indicate that BCL induces the prolonged expression of a range of complement genes and show that microglia/macrophages synthesize C3 and deposit it in the ONL after BCL injury. These findings have relevance to the role of complement in progressive retinal degeneration, including atrophic AMD.

Heparanase in primary human osteoblasts.

Heparanase (HPSE) is known to be involved in fracture repair in mice, but its presence and function in human bone formation remains unclear. Our aim was to determine the expression of HPSE in human bone forming osteoblasts and to better understand its role in osteogenesis. HPSE protein expression and enzymatic activity were demonstrated in osteoblasts isolated from trabecular bone specimens of patients with osteoporosis (OP) and from healthy subjects, although the levels differed markedly. Thus, low levels of HPSE expression were observed in osteoporotic osteoblasts, including in the nucleus compared to those from healthy subjects. Notably, HPSE gene expression was associated with alkaline phosphatase (ALP) activity, the bone turnover marker. Gene profile studies demonstrated that osteogenic genes were downregulated in osteoporotic osteoblasts. We further exposed osteoblasts to exogenous HPSE and found that the level of histone H3 phosphorylation was increased. We provide evidence, for the first time, demonstrating that HPSE expresses and functions in human osteoblasts. Our data suggest that previously undescribed function of HPSE-mediated osteoblastogenesis through regulation of osteogenic gene expression and histone H3 modification. HPSE upregulation may be a novel therapeutic approach in the prevention and treatment of OP.

Expansion of circulating T cells resembling follicular helper T cells is a fixed phenotype that identifies a subset of severe systemic lupus erythematosus.

OBJECTIVE: In the sanroque mouse model of lupus, pathologic germinal centers (GCs) arise due to increased numbers of follicular helper T (Tfh) cells, resulting in high-affinity anti-double-stranded DNA antibodies that cause end-organ inflammation, such as glomerulonephritis. The purpose of this study was to examine the hypothesis that this pathway could account for a subset of patients with systemic lupus erythematosus (SLE). METHODS: An expansion of Tfh cells is a causal, and therefore consistent, component of the sanroque mouse phenotype. We validated the enumeration of circulating T cells resembling Tfh cells as a biomarker of this expansion in sanroque mice, and we performed a comprehensive comparison of the surface phenotype of circulating and tonsillar Tfh cells in humans. This circulating biomarker was enumerated in SLE patients (n = 46), Sjögren's syndrome patients (n = 17), and healthy controls (n = 48) and was correlated with disease activity and end-organ involvement. RESULTS: In sanroque mice, circulating Tfh cells increased in proportion to their GC counterparts, making circulating Tfh cells a feasible human biomarker of this novel mechanism of breakdown in GC tolerance. In a subset of SLE patients (14 of 46), but in none of the controls, the levels of circulating Tfh cells (defined as circulating CXCR5+CD4+ cells with high expression of Tfh-associated molecules, such as inducible T cell costimulator or programmed death 1) were increased. This cellular phenotype did not vary with time, disease activity, or treatment, but it did correlate with the diversity and titers of autoantibodies and with the severity of end-organ involvement. CONCLUSION: These findings in SLE patients are consistent with the autoimmune mechanism in sanroque mice and identify Tfh effector molecules as possible therapeutic targets in a recognizable subset of patients with SLE.

Antineutrophil cytoplasmic antibody-associated vasculitides: could geographic patterns be explained by ambient ultraviolet radiation?

OBJECTIVE: This ecological study describes and quantifies the association between ambient ultraviolet (UV) radiation levels, including daily winter vitamin D effective UV radiation levels and the incidence of the 3 antineutrophil cytoplasmic antibody-associated vasculitides (AAVs): Wegener's granulomatosis (WG), microscopic polyangiitis (MPA), and Churg-Strauss syndrome (CSS). Latitudinal variation in occurrence of the AAVs, especially WG, has been previously reported. For other autoimmune diseases such as multiple sclerosis and type 1 diabetes mellitus, inverse associations with latitude are hypothesized to indicate a causative role for low UV radiation exposure, possibly acting via vitamin D status. METHODS: Published epidemiologic studies provided data on incident cases, total population of study regions, age-specific incidence rates, and study location. From these data and online age-specific population data, we calculated crude incidence rates, the expected number of cases (to control for possible age confounding), and measures of ambient UV radiation. Negative binomial regression models were used to calculate the incidence rate ratio (IRR) for a 1,000 joules/m(2) increase in ambient UV radiation. RESULTS: The incidence of WG and CSS increased with increasing latitude and decreasing ambient UV radiation, with a stronger and more consistent effect across different UV radiation measures for WG, e.g., for average daily ambient clear sky erythemal UV radiation (WG: IRR 0.64 [95% confidence interval (95% CI) 0.44-0.94], P = 0.02; CSS: IRR 0.67 [95% CI 0.43-1.05], P = 0.08; MPA: IRR 1.16 [95% CI 0.92-1.47], P = 0.22). There was no apparent latitudinal variation in MPA incidence. CONCLUSION: Our findings are consistent with a protective immunomodulatory effect of ambient UV radiation on the onset of WG and CSS. We discuss possible mechanisms, including the effect of vitamin D on the immune system.

Development of basic medical sciences in a new medical school with an integrated curriculum: the ANU experience.

BACKGROUND: The development of a basic medical science curriculum in a new medical school with a problem-based focus in Australia has been subject to a number of constraints. We describe the process and early evaluation. AIM: To describe the development of a basic medical science curriculum in an Australian medical school with a problem-based curriculum. METHODS: We describe the process we used for curriculum development and the benefits and constraints that arose from pre-existing strong biomedical science on the Australian National University (ANU) campus. We outline methods we used to inform our curriculum content and report on accreditation and early internal evaluation. RESULTS: Australian medical schools design their curriculum within a relatively restrictive framework put forward by a national accreditation system. The curriculum achieved accreditation from the external accrediting agency, but early student evaluation has been mixed. CONCLUSION: Although our internal faculty evaluation and external review by the accrediting agency has supported the view that this aspect of the curriculum has performed reasonably well, student feedback is mixed and further evaluation is needed and adjustments probably warranted.

The parent-reported prevalence and management of peanut and nut allergy in school children in the Australian Capital Territory.

AIM: To describe parent-reported prevalence and management of peanut and nut allergy in school entrant children. METHOD: A population-based, cross-sectional study in the Australian National Capital. RESULTS: Out of 3851 children, parents reported 127 had a strong allergic reaction to peanuts and 19 to other nuts ever. Nut allergy ever prevalence was 3.8% (95% confidence interval 3.2-4.4%), and of peanut allergy ever 3.3% (2.8-3.9%). Children with nut allergy were more likely to have a general practitioner (odds ratio 2.64, 1.16-6.03), hay fever (3.78, 2.67-5.36), eczema (4.54, 3.15-6.56) and wheeze in the last 12 months (3.19, 2.22-4.59) and have been breastfed (2.68, 1.26-5.77) than those who did not. At follow up of 109 children with parent-reported allergy (75% response), 70% had diagnostic test-confirmed sensitisation, 32% had been prescribed an adrenalin autoinjector (6% had used one) and 46% were not eating peanut. Increasing severity of reported symptoms following consumption of peanut was associated with an increasing likelihood of recommended management. Based on parent report, the projected estimated diagnostic test-confirmed prevalence of peanut sensitisation was 2.4% (1.9%, 3.0%) for the entire sample. CONCLUSION: Among a highly representative sample of children at school entry, 1 in 30 parents reported their child to have a strong allergic reaction to nuts and over 1 in 50 are estimated to have diagnostic test-confirmed peanut sensitisation, based on parent report.

Inflammatory myositis complicating hypocomplementemic urticarial vasculitis despite on-going immunosuppression.

We present a patient with previously diagnosed hypocomplementemic urticarial vasculitis syndrome, with skin, lung, and renal involvement, who presented with congestive cardiac failure. During the course of her hospitalization, she was also found to have profound proximal muscle weakness in both upper and lower limbs associated with raised creatinine kinase levels. A muscle biopsy was performed, which demonstrated evidence of an inflammatory myositis with vasculitis, which had returned despite on-going immunosuppression. This occurrence of a new autoimmune disease may well be an example of the "waste disposal" hypothesis.

Cerebrovascular disease associated with antiphospholipid antibodies: more questions than answers.

Neurological syndromes occur in a significant number of patients with antiphospholipid antibodies. The optimal management for these patients however remains uncertain. Our study is a descriptive analysis looking retrospectively at 45 patients who presented to the principal tertiary referral centre in the Australian Capital Territory, with either cerebral arterial or venous thrombosis for which there was no obvious cause for their presentation when initially reviewed. The diagnosis was based on the clinical findings made by one of three neurologists attached to our centre. Radiological findings and the presence of either IgM or IgG anticardiolipin antibodies, IgG anti-beta-2 glycoprotein 1 antibodies or a lupus anticoagulant were then documented. In this group of patients three subgroups were identified:1. Individuals that fulfilled the Sapporo Classification Criteria2. Individuals with transiently positive antiphospholipid antibodies and3. Individuals with persistently low positive antiphospholipid antibodies. The most interesting of these three groups are those individuals with transiently positive antiphospholipid antibodies. A potential cause for presentation was identified in only one patient of this group with documented infective endocarditis and bacteraemia. Comparison with the other two groups suggested that there was little in terms of clinical presentation, radiological findings or intercurrent risk factors for thrombotic disease to distinguish between them. With disappearance of antiphospholipid antibodies, the individuals within this group have not had further thrombotic events. Our observations emphasise the problems that continue to exist in relation to the occurrence of cerebrovascular disease in the context of antiphospholipid antibodies and the optimal management of these stratified groups. Our findings also raise an as yet unanswered question as to the signficance of these transiently positive antiphospholipid antibodies. In the absence of significant intercurrent risk factors our findings would suggest that in the group we describe that they are likely to be of clinical significance.

Flexner's ethical oversight reprised? Contemporary medical education and the health impacts of corporate globalisation.

BACKGROUND: Abraham Flexner's famous reports of 1910 and 1912, Medical Education in the United States and Canada and Medical Education in Europe, were written to assist the development of a positive response in university curricula to a revolution in understanding about the scientific foundations of clinical medicine. Flexner pointed out many deficiencies in medical education that retain contemporary resonance. Generally underemphasised in Flexner's reports, however, were recommendations promoting a firm understanding of and commitment to medical ethics as a basis of medical professionalism. Indeed, Flexner's praise for the scholastic basic of German medical education appeared somewhat ironic when the ethical inadequacies of prominent Nazi doctors were revealed at the Nuremberg Trials. CORPORATE GLOBALISATION AND ITS IMPACT ON MEDICAL PROFESSIONALISM: This article suggests that contemporary medical educators, like Flexner, may be at risk of inadequately addressing a major challenge to evolving medical professionalism. Medical ethics, health law and even the international right to health are now increasingly emphasised in medical curricula. The same cannot be said, however, of lobbying principles arising from the structures of corporate globalisation, although these are rapidly becoming an even more dominant force in shaping medical practice around the globe. Conclusion Today it is the normative tension between medical ethics, health law and international human rights on the one hand and the lobbying principles and strategies of corporate globalisation that must urgently become the focus of major recommendations for reshaping the teaching of medical professionalism. Suggestions are made as to how this might practically be achieved.

Controversies in the antiphospholipid syndrome and stroke.

Many clinicians providing care and advice to patients with antiphospholipid syndrome (APS), where the principal clinical manifestation is stroke, do so in the setting of an evidence base of mixed quality. Indeed, systematic studies have not particularly helped the practising clinician as they have been characterised by variable criteria used to select subjects, making it impossible to extrapolate to typical clinic patients. This has left us with a number of key questions, each of which attracts controversy in terms of patient management. In this review, these are posed as a series of questions with the answer, or lack of one, considered after each question. The review draws attention to the important questions that require answers for current primary and secondary prevention, as well as treatment of APS and stroke, and suggests a series of studies that are needed to clarify these issues.

Mechanisms of accelerated immune-mediated diabetes resulting from islet beta cell expression of a Fas ligand transgene.

Nonobese diabetic (NOD) mice transgenic for Fas ligand (FasL) on islet beta cells (HIPFasL mice) exhibit an accelerated diabetes distinct from the normal autoimmune diabetes of NOD mice. This study was undertaken to define the mechanism underlying accelerated diabetes development in HIPFasL mice. It was found that diabetes in HIPFasL mice is dependent on the NOD genetic background, as HIPFasL does not cause diabetes when crossed into other mice strains and is lymphocyte dependent, as it does not develop in HIPFasL(SCID) mice. Diabetes development in NOD(SCID) recipients of diabetic HIPFasL splenocytes is slower than when using splenocytes from diabetic NOD mice. Beta cells from HIPFasL mice are more susceptible to cytokine-induced apoptosis than wild-type NOD beta cells, and this can be blocked with anti-FasL Ab. HIPFasL islets are more rapidly destroyed than wild-type islets when transplanted into nondiabetic NOD mice. This confirms that FasL(+) islets do not obtain immune privilege, and instead NOD beta cells constitutively expressing FasL are more susceptible to apoptosis induced by Fas-FasL interaction. These findings are consistent with the accelerated diabetes of young HIPFasL mice being a different disease process from the autoimmune diabetes of wild-type NOD mice. The data support a mechanism by which cytokines produced by the insulitis lesion mediate up-regulation of beta cell Fas expression, resulting in suicide or fratricide of HIPFasL beta cells that overexpress FasL.

Which clinical subgroups within the spectrum of child asthma are attributable to atopy?

STUDY OBJECTIVES: The contribution of atopy to childhood asthma has been debated. We aimed to examine the relationship between atopy and asthma, taking into account differences in respiratory symptoms and disease severity. DESIGN: A cross-sectional asthma survey involving the following: (1) a population sample of 758 (81% of eligible) school children aged 8 to 10 years from randomly selected schools in the Australian Capital Territory in 1999, and (2) a hospital-based sample of 78 (70% of eligible) children attending the hospital for asthma. Skin-prick test results to 10 common aeroallergens were available on 722 children and 77 children, respectively. Baseline spirometry was obtained on a subset of school children (n = 515, 78% of eligible). RESULTS: The association between atopy and wheeze by wheeze frequency over the past year was as follows: no episodes (odds ratio [OR], 1.00 [reference]), 1 to 3 episodes (OR, 3.27; 95% confidence interval [CI], 2.15 to 4.97), 4 to 12 episodes (OR, 3.44; 95% CI, 1.75 to 6.75), and > 12 episodes (OR, 8.70; 95% CI, 3.07 to 24.55), with a higher population attributable fraction (PAF) for > 12 episodes (75%) than 1 to 3 episodes (49%). Atopy was moderately related to asthma ever (OR, 2.09; 95% CI, 1.52 to 2.85; PAF, 33%) but strongly related to 1999 hospital attendance for asthma (OR, 16.95; 95% CI, 6.76 to 42.48; PAF, 89%). Adjustment for child age, gas heater use, and maternal smoking near the child did not materially alter these findings. CONCLUSIONS: The clinical features of frequent wheeze or hospital asthma attendance are largely attributable to atopy, but infrequent wheeze or a history of asthma ever are not. Atopic children are overrepresented in the severe range of the asthma spectrum.