Performance characteristics of five SARS-CoV-2 serological assays: Clinical utility in health-care workers.

STUDY OBJECTIVE: SARS-CoV-2, which causes coronavirus disease (COVID-19), continues to cause significant morbidity and mortality. The diagnosis of acute infection relies on reverse transcription-polymerase chain reaction (RT-PCR)-based viral detection. The objective of this study was to evaluate the optimal serological testing strategy for anti-SARS-CoV-2 antibodies which provides an important indicator of prior infection and potential short-term immunity. METHODS: The sensitivity and specificity of four different ELISA assays (Euroimmun IgG, Euroimmun NCP-IgG, Fortress and DIAsource) and one CLIA assay (Roche ELECSYS) were evaluated in 423 samples; 137 patients with confirmed RT-PCR COVID-19 infection (true positives), and 100 pre-pandemic samples collected prior to October 2019 (true negatives). A further 186 samples were collected from health-care staff and analysed by all five assays. RESULTS: The Fortress ELISA assay demonstrated the highest sensitivity and specificity followed by the Roche ECLIA assay. The highest overall sensitivity came from the assays that measured total antibody (IgM-IgG combined) and the three assays that performed the best (Fortress, Roche, Euroimmun IgG) all have different antigens as their target proteins which suggests that antigen target does not affect assay performance. In mildly symptomatic participants with either a negative RT-PCR or no RT-PCR performed, 16.76% had detectable antibodies suggesting previous infection. CONCLUSIONS: We recommend a combined testing strategy utilizing assays with different antigenic targets using the fully automated Roche ECLIA assay and confirming discordant samples with the Fortress Total Antibody ELISA assay. This study provides an important indicator of prior infection in symptomatic and asymptomatic individuals.

The aldosterone to renin ratio in the diagnosis of primary aldosteronism: Promises and challenges.

The complexity of evaluating patients for secondary treatable causes of hypertension is underappreciated. Primary aldosteronism (PA) is the most prevalent cause of secondary hypertension (3%-32% of hypertensive patients). The recent endocrine society clinical practice guideline (ESCPG), "The Management of Primary Aldosteronism: Case Detection, Diagnosis, and Treatment", differs from the previous version in the explicit recognition of PA as a major public health issue. Despite this, PA is underdiagnosed. The guidelines call on physicians to substantially ramp up the screening of hypertensive patients at risk of PA. Further, it recommends the plasma aldosterone to renin ratio (ARR), as the test of choice for screening for PA. However, the ARR is a highly variable test with reported diagnostic sensitivities and specificities ranging from 66% to 100% and 61% to 100%, respectively. Variability of the ARR can be attributed to the high degree of within-subject variation, differences in sampling protocols, laboratory assays, reporting units, the effect of medications and the population characteristics used to establish the decision thresholds. These factors render the possibility of false positive and false negative results-which have the potential to adversely impact patients. The limitations and caveats to the use of the ARR necessitate an effective clinic-laboratory interface, with specialist physician and clinical scientist collaboration for ARR result interpretation. Improvement in the diagnostic sensitivity and specificity of the ARR is predicated on harmonisation of pretesting patient preparation criteria, knowledge of the analytical methods used to derive the ratio and the method-specific threshold for PA.

Switching from abacavir to tenofovir disoproxil fumarate is associated with rises in soluble glycoprotein VI, suggesting changes in platelet-collagen interactions.

OBJECTIVES: Altered platelet function has been proposed as an underlying mechanism to explain increased risk of myocardial infarction in people living with HIV associated with use of the nucleoside reverse transcriptase inhibitor abacavir (ABC). We aimed to examine changes in platelet biomarkers in people living with HIV switching from ABC. METHODS: In a prospective, 48-week substudy of virally suppressed HIV-1-positive subjects randomized to remain on ABC/lamivudine (ABC/3TC) or switch to tenofovir disoproxil fumarate/emtricitabine, we measured soluble glycoprotein VI (sGPVI), soluble P-selectin, soluble CD40 ligand and von Willebrand factor in plasma collected over time and assessed differences using mixed effect models. RESULTS: Of 312 randomized participants, 310 were included in the analysis. Mean (SD) age 46.4 (9.3) years, 262 (85%) men and 201 (65%) white. At baseline, there was no significant between-group difference in sGPVI [tenofovir disoproxil fumarate/emtricitabine 3.75 (0.25) versus ABC/3TC 3.61 (0.22) ng/ml, P = 0.69]. Greater increases in sGPVI from baseline to week 48 occurred in those switched from ABC/3TC (effect size +0.57 ng/ml; 95% confidence interval, 0.2-0.94; P = 0.003). There was no significant baseline difference or change overtime in soluble P-selectin, soluble CD40 ligand or von Willebrand factor between groups. CONCLUSION: The significant increases in sGPVI that occur with a switch from ABC/3TC are suggestive of changes in platelet function centred on platelet/collagen interactions and potentially represent an underlying mechanism to explain increased risk of myocardial infarction with ABC.

Evaluation of cystatin C in malignancy and comparability of estimates of GFR in oncology patients.

OBJECTIVES: Creatinine is the biomarker of choice for use in estimates of kidney function in oncology patients. However as non-renal factors such as muscle mass can influence creatinine concentrations, we evaluated cystatin C as an alternative biomarker and its incorporation in GFR estimating formulae in an oncology setting. Measured GFR is infrequently undertaken in adult clinical practice with the consequent reliance on calculated GFR for patient assessment. DESIGN AND METHODS: Cystatin C and creatinine concentrations were evaluated from 134 oncology patients prior to commencing chemotherapeutic cycles. Estimates of creatinine clearance (Cockroft-Gault) and GFR (using Hoek, Jonsson, MDRD and CKD-EPI) were evaluated. Cystatin C-based GFR estimates (using CKD-EPI CysC and CKD-EPI SCr/CysC) were compared with the creatinine-based GFR estimates (CG, MDRD and CKD-EPI SCr) within the GFR ranges of 60-89, 45-59 and ≤44 mL/min/1.73 m2. RESULTS: Cystatin C concentrations were significantly higher in oncology patients both prior to commencing chemotherapy (F: P<0.01 and M: P<0.0001) and during cycles of treatment (F: P<0.0001 and M: P<0.01) when compared with a reference population. Cystatin C concentrations also increased significantly during chemotherapy (P<0.0001) in a subset of female patients evaluated. Poor agreement (average 42%) was demonstrated between CKD-EPI CysC and creatinine-based GFR estimates within the investigated GFR ranges, with improved agreement (average 55%) when using the combined CKD-EPI SCr/CysC formula. CONCLUSIONS: This study demonstrated a malignancy and treatment-mediated effect on cystatin C measures, which may confound its clinical utility in estimating GFR in oncology patients.

Classical activation of microglia in CD200-deficient mice is a consequence of blood brain barrier permeability and infiltration of peripheral cells.

The interaction between CD200, expressed on several cell types, and its receptor CD200R, expressed on cells of the myeloid lineage, has been shown to be an important factor in modulating inflammation in macrophage function in several conditions including colitis and arthritis. More recently its modulatory effect on microglial activation has been identified and CD200-deficiency has been associated with increased microglial activation accompanied by increased production of inflammatory cytokines. The response of glia prepared from CD200-deficient mice to stimuli like lipopolysaccharide (LPS) is markedly greater than the response of cells prepared from wildtype mice and, consistent with this, is the recent observation that expression of Toll-like receptor (TLR)4 and signalling through NFκB are increased in microglia prepared from CD200-deficient mice. Here we show that glia from CD200-deficient mice are also more responsive to interferon-γ (IFNγ) which triggers classical activation of microglia. We investigated the effects of CD200-deficiency in vivo and report that there is an increase in expression of several markers of microglial activation including tumor necrosis factor (TNF)-α, which is a hallmark of classically-activated microglia. These changes are accompanied by increased IFNγ, and the evidence suggests that this is produced by infiltrating cells including T cells and macrophages. We propose that these cells enter the brain as a consequence of increased blood brain barrier (BBB) permeability in CD200-deficient mice and that infiltration is assisted by increased expression of the chemokines, monocyte chemotactic protein-1 (MCP-1), IFNγ-induced protein-10 (IP-10) and RANTES. This may have implications in neurodegenerative diseases where BBB permeability is compromised.

Glial Activation in AßPP/PS1 Mice is Associated with Infiltration of IFNγ-Producing Cells.

Whereas the classical histological hallmarks of Alzheimer's disease (AD) are deposition of amyloid-containing plaques and development of neurofibrillary tangles, there is also clear evidence of inflammatory changes accompanied by the presence of activated microglia and astrocytosis. However, at this time, it remains uncertain whether inflammatory changes contribute to pathogenesis of the disease or if they are secondary to deposition of amyloid-ß or other pathological changes. A greater understanding of the sequence of events would clearly improve development of strategies to delay progression of the disease. There is a realistic expectation that advances in imaging technology may provide the key to uncovering this sequence. In this study, we employed non-invasive imaging techniques to examine changes in tissue state in hippocampus and cortex of transgenic mice which overexpress amyloid-ß protein precursor and presenilin 1 and show that the observed increase in T1 relaxation time was associated with astrogliosis while the decrease in T2 relaxation time was associated with microglial activation. We explored the possibility that interferon-γ might trigger glial activation and demonstrate a genotype-related infiltration of macrophages and natural killer cells, which release interferon-γ. The evidence suggests that IFNγ triggers glial activation and expression of proinflammatory cytokines, and these changes, in turn, contribute to the decrease in long-term potentiation.

Long term potentiation is impaired in membrane glycoprotein CD200-deficient mice: a role for Toll-like receptor activation.

The membrane glycoprotein CD200 is expressed on several cell types, including neurons, whereas expression of its receptor, CD200R, is restricted principally to cells of the myeloid lineage, including microglia. The interaction between CD200 and CD200R maintains microglia and macrophages in a quiescent state; therefore, CD200-deficient mice express an inflammatory phenotype exhibiting increased macrophage or microglial activation in models of arthritis, encephalitis, and uveoretinitis. Here, we report that lipopolysaccharide (LPS) and Pam(3)CysSerLys(4) exerted more profound effects on release of the proinflammatory cytokines, interleukin (IL)-1ß, IL-6, and tumor necrosis factor-α (TNFα), in glia prepared from CD200(-/-) mice compared with wild type mice. This effect is explained by the loss of CD200 on astrocytes, which modulates microglial activation. Expression of Toll-like receptors 4 and 2 (TLR4 and -2) was increased in glia prepared from CD200(-/-) mice, and the evidence indicates that microglial activation, assessed by the increased numbers of CD11b(+) cells that stained positively for both MHCII and CD40, was enhanced in CD200(-/-) mice compared with wild type mice. These neuroinflammatory changes were associated with impaired long term potentiation (LTP) in CA1 of hippocampal slices prepared from CD200(-/-) mice. One possible explanation for this is the increase in TNFα in hippocampal tissue prepared from CD200(-/-) mice because TNFα application inhibited LTP in CA1. Significantly, LPS and Pam(3)CysSerLys(4), at concentrations that did not affect LTP in wild type mice, inhibited LTP in slices prepared from CD200(-/-) mice, probably due to the accompanying increase in TLR2 and TLR4. Thus, the neuroinflammatory changes that result from CD200 deficiency have a negative impact on synaptic plasticity.