IgG fractions from patients with antiphospholipid syndrome and systemic lupus erythematosus bind to platelets, but do not affect collagen-induced platelet activation.

Anti-beta-2 glycoprotein 1 (anti-ß2GP1) is an antiphospholipid antibody found in patients with antiphospholipid syndrome (APS) and systemic lupus erythematosus (SLE). Its presence commonly is associated with thrombosis; however, the mechanisms of interaction of anti-ß2GP1 antibodies and platelets remain unclear. We investigated the effects of APS and SLE patient-derived IgG fractions on collagen-mediated platelet aggregation and examined the binding of patient-derived IgG to platelets before and after activation by collagen. IgG fractions, 150, 200, 300 or 350 µg/ml, isolated from 11 patients with APS and SLE were incubated with two sets of platelet-rich plasma (PRP) in the incubation wells of an aggregometer. The first set was activated by collagen and the other set was incubated for an additional 10 min. All platelets were collected by centrifugation and fixed in cell blocks. We assessed binding of IgG to platelets using immunocytochemistry (ICC). Patient-derived IgG fractions did not affect collagen-induced platelet aggregation. ICC staining using anti-human IgG antibodies demonstrated that patient-derived IgG fractions had greater affinity for non-activated platelets than those activated by 0.75 µg/ml collagen. Patient-derived IgG fractions bound to the surface of platelets and potentially could be internalized by platelets. IgG fractions from APS and SLE patients may sensitize non-activated platelets, which could increase platelet reactivity and thrombotic risk in patients. We did not detect secondary effects of patient-derived IgG fractions.

Itch in dermatomyositis: the role of increased skin interleukin-31.

BACKGROUND: Interleukin (IL)-31 is implicated in pruritus associated with pruritic skin diseases like atopic dermatitis. Although pruritus is a prominent feature in dermatomyositis (DM), few studies have evaluated the pathogenesis of DM-associated itch. OBJECTIVES: To establish the prevalence of itch in DM, and to investigate the role of IL-31 in DM-related itch. METHODS: Pruritus and disease activity of DM were evaluated by a visual analogue scale (VAS) and the Cutaneous Disease and Activity Severity Index (CDASI), respectively. Expression of IL-31 and IL-31 receptor alpha (IL-31RA) in lesional DM, nonlesional DM and healthy control skin was evaluated by quantitative reverse-transcriptase polymerase chain reaction and immunofluorescence. Flow cytometry was performed on skin cells isolated from lesional DM skin to identify cellular sources of IL-31 in DM. RESULTS: Among 191 patients with DM, 50·8% had moderate-to-severe itch, and itch was correlated with increased cutaneous severity (r = 0·34). In patients with itchy DM, gene expression of IL31 and IL31RA in lesional skin was upregulated compared with nonlesional skin and healthy control skin. IL31 mRNA expression positively correlated with VAS itch score (r = 0·67). On immunofluorescence, immunoreactivity for IL-31 and IL-31RA was stronger in lesional skin. Flow cytometry showed that lesional DM skin contained significantly more IL-31-producing cells, and CD4+ cells were the most common cell type. Lenabasum, an emerging treatment for DM, significantly downregulated IL-31 from CpG-stimulated peripheral blood mononuclear cells. CONCLUSIONS: Increased skin IL-31 may play a role in DM-associated itch, and ongoing trials will evaluate the effects of systemic treatment on IL-31 and itch in DM.

Identification of reference miRNAs in plasma useful for the study of oestrogen-responsive miRNAs associated with acquired Protein S deficiency in pregnancy.

BACKGROUND: Accumulating evidence indicate that circulating microRNAs (miRNAs) are useful independent non-invasive biomarkers, with unique miRNA signatures defined for various pathophysiological conditions. However, there are no established universal housekeeping miRNAs for the normalisation of miRNAs in body fluids. We have previously identified an oestrogen-responsive miRNA, miR-494, in regulating the anticoagulant, Protein S, in HuH-7 liver cells. Moreover, increased thrombotic risk associated with elevated circulating oestrogen levels is frequently observed in pregnant women and oral contraceptive users. In order to identify other oestrogen-responsive miRNAs, including miR-494, that may be indicative of increased thrombotic risk in plasma, we used nanoString analysis to identify robust and stable endogenous reference miRNAs for the study of oestrogen-responsive miRNAs in plasma. RESULTS: We compared the plasma miRNA expression profile of individuals with: (1) Low circulating oestrogens (healthy men and non-pregnant women not taking oral contraceptives), (2) High circulating synthetic oestrogens, (women taking oral contraceptives) and (3) High circulating natural oestrogens (pregnant females >14 weeks gestation). From the nanoString analyses, 11 candidate reference miRNAs which exhibited high counts and not significantly differentially expressed between groups were selected for validation using realtime quantitative polymerase chain reaction (RT-qPCR) and digital droplet PCR (DDPCR) in pooled plasma samples, and the stability of their expression evaluated using NormFinder and BestKeeper algorithms. Four miRNAs (miR-25-5p, miR-188-5p, miR-222-3p and miR-520f) demonstrated detectable stable expression between groups and were further analysed by RT-qPCR in individual plasma samples, where miR-188-5p and miR-222-3p expression were identified as a stable pair of reference genes. The miRNA reference panel consisting of synthetic spike-ins cel-miR-39 and ath-miR159a, and reference miRNAs, miR-188-5p and miR-222-3p was useful in evaluating fold-change of the pregnancy-associated miRNA, miR-141-3p, between groups. CONCLUSION: The miRNA reference panel will be useful for normalising qPCR data comparing miRNA expression between men and women, non-pregnant and pregnant females, and the potential effects of endogenous and synthetic oestrogens on plasma miRNA expression.

Evaluation of the reliability of the Cutaneous Dermatomyositis Disease Area and Severity Index and the Cutaneous Assessment Tool-Binary Method in juvenile dermatomyositis among paediatric dermatologists, rheumatologists and neurologists.

BACKGROUND: The Cutaneous Dermatomyositis Disease Area and Severity Index (CDASI) and Cutaneous Assessment Tool-Binary Method (CAT-BM) have been shown to be reliable and valid outcome measures to assess cutaneous disease in adult dermatomyositis (DM) and juvenile DM (JDM), respectively. OBJECTIVES: This study compared the CDASI and CAT-BM for use by paediatric dermatologists, paediatric rheumatologists and paediatric neurologists in patients with JDM. METHODS: Five paediatric dermatologists, five paediatric rheumatologists and five paediatric neurologists each evaluated 14 patients with JDM using the CDASI, CAT-BM, and skin Physician Global Assessment (PGA) scales. Inter-rater reliability, intra-rater reliability, construct validity and completion time were compared. RESULTS: Inter-rater reliability for CDASI activity and damage scores was good to moderate for paediatric dermatologists and rheumatologists, but poor for paediatric neurologists. The inter-rater reliability for CAT-BM activity scores was moderate for paediatric dermatologists and rheumatologists, but poor for paediatric neurologists and poor across all specialties for damage scores. Intra-rater reliability for the CDASI and CAT-BM activity and damage scores was moderate to excellent for paediatric dermatologists, rheumatologists and neurologists. Strong associations were found between skin PGA activity and damage scores and CDASI or CAT-BM activity and damage scores, respectively (P < 0·002). The CDASI had a mean completion time of 5·4 min compared with that for the CAT-BM of 3·1 min. CONCLUSIONS: Our data confirm the reliability of the CDASI activity and damage scores and the CAT-BM activity scores when used by paediatric dermatologists and rheumatologists in assessing JDM. Significant variation existed in the paediatric neurologists' scores.

The reliability of the Cutaneous Dermatomyositis Disease Area and Severity Index (CDASI) among dermatologists, rheumatologists and neurologists.

BACKGROUND: Previous studies have shown that skin disease in dermatomyositis (DM) is best assessed using the Cutaneous Dermatomyositis Disease Area and Severity Index (CDASI). Although the CDASI has been validated for use by dermatologists, it has not been validated for use by other physicians such as rheumatologists and neurologists, who also manage patients with DM and assess skin activity in clinical trials. OBJECTIVES: To assess the reliability of the CDASI among dermatologists, rheumatologists and neurologists. METHODS: Fifteen patients with cutaneous DM were assessed using the CDASI and the Physician Global Assessment (PGA) by five dermatologists, five rheumatologists and five neurologists. RESULTS: The mean CDASI activity scores for dermatologists, rheumatologists and neurologists were 21·0, 21·8 and 20·8, respectively. These mean scores were not different among the specialists. The CDASI damage score means for dermatologists, rheumatologists and neurologists were 5·3, 7·0 and 4·8, respectively. The mean scores between dermatologists and rheumatologists were significantly different, but the means between dermatologists and neurologists were not. The intraclass correlation coefficients (ICCs) for interrater reliability for CDASI activity and damage were good to excellent for dermatologists and rheumatologists, and moderate to excellent for neurologists. The ICCs for intrarater reliability for CDASI activity and damage were excellent for dermatologists and rheumatologists and moderate to excellent for neurologists. The PGA displayed lower interrater and intrarater reliability relative to the CDASI. CONCLUSIONS: Our results confirm the reliability of the CDASI when used by dermatologists and rheumatologists. The data for its use by neurologists were not as robust.

Evaluation of the influence of the definition of an isolated hip fracture as an exclusion criterion for trauma system benchmarking: a multicenter cohort study.

PURPOSE: To assess whether the definition of an IHF used as an exclusion criterion influences the results of trauma center benchmarking. METHODS: We conducted a multicenter retrospective cohort study with data from an integrated Canadian trauma system. The study population included all patients admitted between 1999 and 2010 to any of the 57 adult trauma centers. Seven definitions of IHF based on diagnostic codes, age, mechanism of injury, and secondary injuries, identified in a systematic review, were used. Trauma centers were benchmarked using risk-adjusted mortality estimates generated using the Trauma Risk Adjustment Model. The agreement between benchmarking results generated under different IHF definitions was evaluated with correlation coefficients on adjusted mortality estimates. Correlation coefficients >0.95 were considered to convey acceptable agreement. RESULTS: The study population consisted of 172,872 patients before exclusion of IHF and between 128,094 and 139,588 patients after exclusion. Correlation coefficients between risk-adjusted mortality estimates generated in populations including and excluding IHF varied between 0.86 and 0.90. Correlation coefficients of estimates generated under different definitions of IHF varied between 0.97 and 0.99, even when analyses were restricted to patients aged ≥65 years. CONCLUSIONS: Although the exclusion of patients with IHF has an influence on the results of trauma center benchmarking based on mortality, the definition of IHF in terms of diagnostic codes, age, mechanism of injury and secondary injury has no significant impact on benchmarking results. Results suggest that there is no need to obtain formal consensus on the definition of IHF for benchmarking activities.