Longitudinal expression profiling of CD4+ and CD8+ cells in patients with active to quiescent giant cell arteritis.

BACKGROUND: Giant cell arteritis (GCA) is the most common form of vasculitis affecting elderly people. It is one of the few true ophthalmic emergencies but symptoms and signs are variable thereby making it a challenging disease to diagnose. A temporal artery biopsy is the gold standard to confirm GCA, but there are currently no specific biochemical markers to aid diagnosis. We aimed to identify a less invasive method to confirm the diagnosis of GCA, as well as to ascertain clinically relevant predictive biomarkers by studying the transcriptome of purified peripheral CD4+ and CD8+ T lymphocytes in patients with GCA. METHODS: We recruited 16 patients with histological evidence of GCA at the Royal Victorian Eye and Ear Hospital, Melbourne, Australia, and aimed to collect blood samples at six time points: acute phase, 2-3 weeks, 6-8 weeks, 3 months, 6 months and 12 months after clinical diagnosis. CD4+ and CD8+ T-cells were positively selected at each time point through magnetic-assisted cell sorting. RNA was extracted from all 195 collected samples for subsequent RNA sequencing. The expression profiles of patients were compared to those of 16 age-matched controls. RESULTS: Over the 12-month study period, polynomial modelling analyses identified 179 and 4 statistically significant transcripts with altered expression profiles (FDR < 0.05) between cases and controls in CD4+ and CD8+ populations, respectively. In CD8+ cells, two transcripts remained differentially expressed after 12 months; SGTB, associated with neuronal apoptosis, and FCGR3A, associatied with Takayasu arteritis. We detected genes that correlate with both symptoms and biochemical markers used for predicting long-term prognosis. 15 genes were shared across 3 phenotypes in CD4 and 16 across CD8 cells. In CD8, IL32 was common to 5 phenotypes including Polymyalgia Rheumatica, bilateral blindness and death within 12 months. CONCLUSIONS: This is the first longitudinal gene expression study undertaken to identify robust transcriptomic biomarkers of GCA. Our results show cell type-specific transcript expression profiles, novel gene-phenotype associations, and uncover important biological pathways for this disease. In the acute phase, the gene-phenotype relationships we have identified could provide insight to potential disease severity and as such guide in initiating appropriate patient management.

Geo-epidemiology of temporal artery biopsy-positive giant cell arteritis in Australia and New Zealand: is there a seasonal influence?

OBJECTIVE: Previous studies, although inconclusive, have suggested possible associations of environmental risk factors with the development of giant cell arteritis (GCA). We aim to investigate seasonal influence on the incidence of GCA across Australia and New Zealand. METHODS: In establishing an international study to investigate the molecular aetiology of GCA, archived temporal artery biopsy (TAB) specimens primarily from Australia and New Zealand were obtained. Demographic details including age, sex and date of TAB were collected from collaborating pathology departments. The season in which GCA was diagnosed was determined and compared with previous reports investigating the association between environmental risk factors and GCA. RESULTS: Our study comprises data from 2224 TAB-positive patients with GCA; 2099 of which were from patients in Australia and New Zealand. The mean age at time of diagnosis was 76.4 years of age. The female-to-male ratio was 2.2:1. We noted equal distribution of the incidence rate across all four seasons (530-580 cases diagnosed every quarter). Statistical analysis of seasonal variation by Poisson regression and cosinor methods showed no incidence preponderance across seasons. Our results do not support a seasonal component contributing to the onset of disease. Our literature search identifies no consistent environmental risk factor in association with GCA. CONCLUSION: This is the largest GCA data set reported outside of Europe. Our results demonstrate equal distribution of the incidence rate across all four seasons. In contrast to some earlier reports, we did not identify evidence of a seasonal component contributing to the onset of disease.

Mitochondrial replacement in an iPSC model of Leber's hereditary optic neuropathy.

Cybrid technology was used to replace Leber hereditary optic neuropathy (LHON) causing mitochondrial DNA (mtDNA) mutations from patient-specific fibroblasts with wildtype mtDNA, and mutation-free induced pluripotent stem cells (iPSCs) were generated subsequently. Retinal ganglion cell (RGC) differentiation demonstrates increased cell death in LHON-RGCs and can be rescued in cybrid corrected RGCs.

Development of a Modular Automated System for Maintenance and Differentiation of Adherent Human Pluripotent Stem Cells.

Patient-specific induced pluripotent stem cells (iPSCs) have tremendous potential for development of regenerative medicine, disease modeling, and drug discovery. However, the processes of reprogramming, maintenance, and differentiation are labor intensive and subject to intertechnician variability. To address these issues, we established and optimized protocols to allow for the automated maintenance of reprogrammed somatic cells into iPSCs to enable the large-scale culture and passaging of human pluripotent stem cells (PSCs) using a customized TECAN Freedom EVO. Generation of iPSCs was performed offline by nucleofection followed by selection of TRA-1-60-positive cells using a Miltenyi MultiMACS24 Separator. Pluripotency markers were assessed to confirm pluripotency of the generated iPSCs. Passaging was performed using an enzyme-free dissociation method. Proof of concept of differentiation was obtained by differentiating human PSCs into cells of the retinal lineage. Key advantages of this automated approach are the ability to increase sample size, reduce variability during reprogramming or differentiation, and enable medium- to high-throughput analysis of human PSCs and derivatives. These techniques will become increasingly important with the emergence of clinical trials using stem cells.

Giant cell arteritis: ophthalmic manifestations of a systemic disease.

BACKGROUND: Giant cell arteritis (GCA) is a systemic granulomatous vasculitis, primarily affecting medium-large arteries. It has a predilection for the aorta and its major branches, including the carotid and vertebral arteries. Ophthalmic artery involvement frequently leads to irreversible visual loss, and therefore GCA is one of the few true ophthalmic emergencies. GCA, although classified as a large vessel vasculitis, is known to affect smaller-sized vessels, resulting in a multiplicity of signs in the eye, some of which are often missed. PURPOSE: We set out to highlight some of the less frequently observed clinical signs, which may provide clues to clinically diagnosing GCA in patients presenting with non-classical features and inconclusive inflammatory markers. METHODS: We review the literature and describe the diverse ocular features and some of the systemic findings that can be associated with GCA. RESULTS: Although the most common ocular manifestation of GCA is anterior ischaemic optic neuropathy, the clinical presentation of GCA can vary dramatically. In the absence of obvious ocular involvement, more subtle ophthalmic signs of anterior segment ischaemia, such as hypotony and anisocoria, may be present at the time of initial clinical examination. CONCLUSION: There are no specific biomarkers for disease to date; therefore, pertinent history and clinical examination can guide towards diagnosis in the acute setting. The diagnostic process is not always straightforward, yet appropriate and prompt diagnosis is critical to enable timely intervention and prevent significant morbidity.

Participant understanding and recall of informed consent for induced pluripotent stem cell biobanking.

The ability to generate human induced pluripotent stem cells (iPSCs) has opened new avenues for human disease modelling and therapy. The aim of our study was to determine research participants' understanding of the information given when donating skin biopsies for the generation of patient-specific iPSCs. A customised 35-item questionnaire based on previous iPSC consent guidelines was sent to participants who had previously donated samples for iPSC research. The questionnaire asked pertinent demographic details, participants' motivation to take part in iPSC research and their attitudes towards related ethical issues. 234 participants were contacted with 141 (60.3 %) complete responses received. The median duration between recruitment and follow-up questioning was 313 days (range 10-573 days). The majority of participants (n = 129, 91.5 %) believed they understood what a stem cell was; however, only 22 (16.1 %) correctly answered questions related to basic stem cell properties. We found no statistically significant difference in responses from participants with different levels of education, or those with a health sciences background. The poor understanding amongst participants of iPSC research is unlikely to be unique to our study and may impact future research if not improved. As such, there is a need to develop an easily understood yet comprehensive consent process to ensure ongoing ethical progress of iPSC biobanking.

PTPN22 R620W minor allele is a genetic risk factor for giant cell arteritis.

Giant cell arteritis (GCA) is one of the commonest forms of vasculitis in the elderly, and may result in blindness and stroke. The pathogenesis of GCA is not understood, although environmental, infectious and genetic risk factors are implicated. One gene of interest is PTPN22, encoding lymphoid protein tyrosine phosphatase (Lyp), expressed exclusively in immune cells, which is proposed to be an 'archetypal non-HLA autoimmunity gene'. The minor allele of a functional PTPN22 single nucleotide polymorphism (rs2476601, R620W), which disrupts an interaction motif in the protein, was originally reported to be associated with biopsy-proven GCA in Spanish patients, with supporting data from three replicate Northern European studies. Recently, this observation was extended with additional patients and controls, and studies encompassing European, Scandinavian, UK and American patients. The aim of our study was to determine the association between PTPN22 rs2476601 (R620W) and biopsy-proven GCA in an Australian case cohort.

Heterogeneity of Human Research Ethics Committees and Research Governance Offices across Australia: An observational study.

BACKGROUND: Conducting ethically grounded research is a fundamental facet of all investigations. Nevertheless, the administrative burdens of current ethics review are substantial, and calls have been made for a reduction in research waste. AIMS: To describe the heterogeneity in administration and documentation required by Human Research Ethics Committees (HRECs) and Research Governance Offices (RGOs) across Australia. METHODS: In establishing a nationwide study to investigate the molecular aetiology of Giant Cell Arteritis (GCA), for which archived pathological specimens from around Australia are being recruited, we identified variation across separate HREC and RGO requirements. Submission paperwork and correspondence from each collaborating site and its representative office for research were reviewed. This data was interrogated to evaluate differences in current guidelines. RESULTS: Twenty-five pathology departments across seven Australian States collaborated in this study. All states, except Victoria, employed a single ethics review model. There was discrepancy amongst HRECs as to which application process applied to our study: seven requested completion of a "National Ethics Application Form" and three a "Low Negligible Risk" form. Noticeable differences in guidelines included whether electronic submission was sufficient. There was variability in the total number of documents submitted (range five to 22) and panel review turnaround time (range nine to 136 days). CONCLUSION: We demonstrate the challenges and illustrate the heavy workload involved in receiving widespread ethics and governance approval across Australia. We highlight the need to simplify, homogenise, and nationalise human ethics for non-clinical trial studies. Reducing unnecessary administration will enable investigators to achieve research aims more efficiently.

Projected worldwide disease burden from giant cell arteritis by 2050.

OBJECTIVE: To estimate and project the number of people affected worldwide by giant cell arteritis (GCA) by 2050. Modeling the number of people visually impaired as a result of this disease will help establish the projected morbidity and resource burden. METHODS: A systematic literature review up to December 2013 was conducted using PubMed and ISI Web of Science. Studies reporting an incidence rate for GCA were used to model disease incident cases at regional and national levels. United Nations Population Prospect data were used for population projections. Morbidity burden was established through rates of visual impairment. The associated financial implications were calculated for the United States. RESULTS: The number of incident cases of GCA will increase secondary to an aging population. By 2050, more than 3 million people will have been diagnosed with GCA in Europe, North America, and Oceania. About 500,000 people will be visually impaired. By 2050, in the United States alone, the estimated cost from visual impairment due to GCA will exceed US$76 billion. Inpatient care for patients with active GCA will total about US$1 billion. Management of steroid-related adverse events will increase costs further, with steroid-induced fractures estimated to total US$6 billion by 2050. CONCLUSION: Projecting disease burden for GCA on a global scale allows for optimization of healthcare planning and prioritization of research domains. Additional population-based studies are required to more accurately project worldwide disease burden. Our work highlights the future global disease burden of GCA, and illustrates the associated financial implications.

A diagnostic challenge: chronic myelomonocytic leukaemia and recurrent anterior ischaemic optic neuropathy.

We report the first case of ischaemic optic neuropathy (ION) linked to chronic myelomonocytic leukaemia (CMML) and its associated vasculitis. We discuss the link between CMML and vasculitis and the evidence suggesting it can cause anterior ischaemic optic neuropathy through a vasculitic process. We highlight the difficulty and delay in diagnosis as the use of steroids masked an underlying systemic process. Recurrent ION and raised inflammatory markers should raise suspicion of vasculitis. Together with an elevated monocyte but low platelet count, CMML should be considered.

Heart block following topical latanoprost treatment.

The authors report a case of second degree heart block associated with topical latanoprost treatment. The authors discuss the possibility of a causative effect as the cessation of this treatment resulted in improvement of the arrhythmia. The authors highlight previous reports and research in humans and animals which demonstrate an association of arrhythmias with prostaglandin analogues. This report draws attention to the possibility that an extremely commonly prescribed topical drug may trigger arrhythmias in susceptible individuals. It is important that prescribers are aware of this possible side-effect.