New MR sequences in daily practice: susceptibility weighted imaging. A pictorial essay

Background Susceptibility-weighted imaging (SWI) is a relatively new magnetic resonance (MR) technique that exploits the magnetic susceptibility differences of various tissues, such as blood, iron and calcification, as a new source of contrast enhancement. This pictorial review is aimed at illustrating and discussing its main clinical applications. Methods SWI is based on high-resolution, threedimensional (3D), fully velocity-compensated gradientecho sequences using both magnitude and phase images. A phase mask obtained from the MR phase images is multiplied with magnitude images in order to increase the visualisation of the smaller veins and other sources of susceptibility effects, which are displayed at best after postprocessing of the 3D dataset with the minimal intensity projection (minIP) algorithm. Results SWI is very useful in detecting cerebral microbleeds in ageing and occult low-flow vascular malformations, in characterising brain tumours and degenerative diseases of the brain, and in recognizing calcifications in various pathological conditions. The phase images are especially useful in differentiating between paramagnetic susceptibility effects of blood and diamagnetic effects of calcium. SWI can also be used to evaluate changes in iron content in different neurodegenerative disorders. Conclusion SWI is useful in differentiating and characterising diverse brain disorders

Harmonization guidelines for HLA-peptide multimer assays derived from results of a large scale international proficiency panel of the Cancer Vaccine Consortium

PURPOSE: The Cancer Vaccine Consortium of the Cancer Research Institute (CVC-CRI) conducted a multicenter HLA-peptide multimer proficiency panel (MPP) with a group of 27 laboratories to assess the performance of the assay. EXPERIMENTAL DESIGN: Participants used commercially available HLA-peptide multimers and a well characterized common source of peripheral blood mononuclear cells (PBMC). The frequency of CD8+ T cells specific for two HLA-A2-restricted model antigens was measured by flow cytometry. The panel design allowed for participants to use their preferred staining reagents and locally established protocols for both cell labeling, data acquisition and analysis. RESULTS: We observed significant differences in both the performance characteristics of the assay and the reported frequencies of specific T cells across laboratories. These results emphasize the need to identify the critical variables important for the observed variability to allow for harmonization of the technique across institutions. CONCLUSIONS: Three key recommendations emerged that would likely reduce assay variability and thus move toward harmonizing of this assay. (1) Use of more than two colors for the staining (2) collect at least 100,000 CD8 T cells, and (3) use of a background control sample to appropriately set the analytical gates. We also provide more insight into the limitations of the assay and identified additional protocol steps that potentially impact the quality of data generated and therefore should serve as primary targets for systematic analysis in future panels. Finally, we propose initial guidelines for harmonizing assay performance which include the introduction of standard operating protocols to allow for adequate training of technical staff and auditing of test analysis procedures

The Role of Recombination for the Coevolutionary Dynamics of HIV and the Immune Response

The evolutionary implications of recombination in HIV remain not fully understood. A plausible effect could be an enhancement of immune escape from cytotoxic T lymphocytes (CTLs). In order to test this hypothesis, we constructed a population dynamic model of immune escape in HIV and examined the viral-immune dynamics with and without recombination. Our model shows that recombination (i) increases the genetic diversity of the viral population, (ii) accelerates the emergence of escape mutations with and without compensatory mutations, and (iii) accelerates the acquisition of immune escape mutations in the early stage of viral infection. We see a particularly strong impact of recombination in systems with broad, non-immunodominant CTL responses. Overall, our study argues for the importance of recombination in HIV in allowing the virus to adapt to changing selective pressures as imposed by the immune system and shows that the effect of recombination depends on the immunodominance pattern of effector T cell responses

Comparison of total parathyroidectomy without autotransplantation and without thymectomy versus total parathyroidectomy with autotransplantation and with thymectomy for secondary hyperparathyroidism: TOPAR PILOT-Trial

<p>Abstract</p> <p>Background</p> <p>Secondary hyperparathyroidism (sHPT) is common in patients with chronic renal failure. Despite the initiation of new therapeutic agents, several patients will require parathyroidectomy (PTX). Total PTX with autotransplantation of parathyroid tissue (TPTX+AT) and subtotal parathyroidectomy (SPTX) are currently considered as standard surgical procedures in the treatment of sHPT. Recurrencerates after TPTX+AT or SPTX are between 10% and 12% (median follow up: 36 months).</p> <p>Recent retrospective studies demonstrated a lower rate of recurrent sHPT of 0–4% after PTX without autotransplantation and thymectomy (TPTX) with no higher morbidity when compared to the standard procedures. The observed superiority of TPTX is flawed due to different definitions of outcomes, varying follow up periods and different surgical treatment strategies (with and without thymectomy).</p> <p>Methods/Design</p> <p>Patients with sHPT (intact parathyroid hormone > 10 times above the upper limit of normal) on long term dialysis (>12 months) will be randomized either to TPTX or TPTX+AT and followed for 36 months. Outcome parameters are recurrence rates of sHPT, frequencies of reoperations due to refractory hypoparathyroidism or recurrent/persistent hyperparathyroidism, postoperative morbidity and mortality and quality of life. 50 patients per group will be randomized in order to obtain relevant frequencies of outcome parameters that will form the basis for a large scale confirmatory multicentred randomized controlled trial.</p> <p>Discussion</p> <p>sHPT is a disease with a high incidence in patients with chronic renal failure. Even a small difference in outcomes will be of clinical relevance. To assess sufficient data about the rate of recurrent sHPT after both methods, a multicentred, randomized controlled trial (MRCT) under standardized conditions is mandatory.</p> <p>Due to the existing uncertainties the calculated number of patients necessary in each treatment arm (n > 4000) makes it impossible to perform this study as a confirmatory trial. Therefore estimates of different outcomes are performed using a pilot MRCT comparing 50 versus 50 randomized patients in order to establish a hypothesis that can be tested thereafter.</p> <p>If TPTX proves to have a lower rate of recurrent sHPT, no relevant disadvantages and no higher morbidity than TPTX+AT, current surgical practice may be changed.</p> <p>Trial registration</p> <p>International Standard Randomized Controlled Trial Number Registration (ISRCTN86202793)</p

Sequential Broadening of CTL Responses in Early HIV-1 Infection Is Associated with Viral Escape

BACKGROUND: Antigen-specific CTL responses are thought to play a central role in containment of HIV-1 infection, but no consistent correlation has been found between the magnitude and/or breadth of response and viral load changes during disease progression. METHODS AND FINDINGS: We undertook a detailed investigation of longitudinal CTL responses and HIV-1 evolution beginning with primary infection in 11 untreated HLA-A2 positive individuals. A subset of patients developed broad responses, which selected for consensus B epitope variants in Gag, Pol, and Nef, suggesting CTL-induced adaptation of HIV-1 at the population level. The patients who developed viral escape mutations and broad autologous CTL responses over time had a significantly higher increase in viral load during the first year of infection compared to those who did not develop viral escape mutations. CONCLUSIONS: A continuous dynamic development of CTL responses was associated with viral escape from temporarily effective immune responses. Our results suggest that broad CTL responses often represent footprints left by viral CTL escape rather than effective immune control, and help explain earlier findings that fail to show an association between breadth of CTL responses and viral load. Our results also demonstrate that CTL pressures help to maintain certain elements of consensus viral sequence, which likely represent viral escape from common HLA-restricted CTL responses. The ability of HIV to evolve to escape CTL responses restricted by a common HLA type highlights the challenges posed to development of an effective CTL-based vaccine

The Efficiency of the Human CD8+ T Cell Response: How Should We Quantify It, What Determines It, and Does It Matter?

Multidisciplinary techniques, in particular the combination of theoretical and experimental immunology, can address questions about human immunity that cannot be answered by other means. From the turnover of virus-infected cells in vivo, to rates of thymic production and HLA class I epitope prediction, theoretical techniques provide a unique insight to supplement experimental approaches. Here we present our opinion, with examples, of some of the ways in which mathematics has contributed in our field of interest: the efficiency of the human CD8+ T cell response to persistent viruses

The impact of viral mutations on recognition by SARS-CoV-2 specific T cells.

We identify amino acid variants within dominant SARS-CoV-2 T cell epitopes by interrogating global sequence data. Several variants within nucleocapsid and ORF3a epitopes have arisen independently in multiple lineages and result in loss of recognition by epitope-specific T cells assessed by IFN-γ and cytotoxic killing assays. Complete loss of T cell responsiveness was seen due to Q213K in the A∗01:01-restricted CD8+ ORF3a epitope FTSDYYQLY207-215; due to P13L, P13S, and P13T in the B∗27:05-restricted CD8+ nucleocapsid epitope QRNAPRITF9-17; and due to T362I and P365S in the A∗03:01/A∗11:01-restricted CD8+ nucleocapsid epitope KTFPPTEPK361-369. CD8+ T cell lines unable to recognize variant epitopes have diverse T cell receptor repertoires. These data demonstrate the potential for T cell evasion and highlight the need for ongoing surveillance for variants capable of escaping T cell as well as humoral immunity.This work is supported by the UK Medical Research Council (MRC); Chinese Academy of Medical Sciences(CAMS) Innovation Fund for Medical Sciences (CIFMS), China; National Institute for Health Research (NIHR)Oxford Biomedical Research Centre, and UK Researchand Innovation (UKRI)/NIHR through the UK Coro-navirus Immunology Consortium (UK-CIC). Sequencing of SARS-CoV-2 samples and collation of data wasundertaken by the COG-UK CONSORTIUM. COG-UK is supported by funding from the Medical ResearchCouncil (MRC) part of UK Research & Innovation (UKRI),the National Institute of Health Research (NIHR),and Genome Research Limited, operating as the Wellcome Sanger Institute. T.I.d.S. is supported by a Well-come Trust Intermediate Clinical Fellowship (110058/Z/15/Z). L.T. is supported by the Wellcome Trust(grant number 205228/Z/16/Z) and by theUniversity of Liverpool Centre for Excellence in Infectious DiseaseResearch (CEIDR). S.D. is funded by an NIHR GlobalResearch Professorship (NIHR300791). L.T. and S.C.M.are also supported by the U.S. Food and Drug Administration Medical Countermeasures Initiative contract75F40120C00085 and the National Institute for Health Research Health Protection Research Unit (HPRU) inEmerging and Zoonotic Infections (NIHR200907) at University of Liverpool inpartnership with Public HealthEngland (PHE), in collaboration with Liverpool School of Tropical Medicine and the University of Oxford.L.T. is based at the University of Liverpool. M.D.P. is funded by the NIHR Sheffield Biomedical ResearchCentre (BRC – IS-BRC-1215-20017). ISARIC4C is supported by the MRC (grant no MC_PC_19059). J.C.K.is a Wellcome Investigator (WT204969/Z/16/Z) and supported by NIHR Oxford Biomedical Research Centreand CIFMS. The views expressed are those of the authors and not necessarily those of the NIHR or MRC

Molecular heterogeneity and CXorf67 alterations in posterior fossa group A (PFA) ependymomas

Of nine ependymoma molecular groups detected by DNA methylation profiling, the posterior fossa type A (PFA) is most prevalent. We used DNA methylation profiling to look for further molecular heterogeneity among 675 PFA ependymomas. Two major subgroups, PFA-1 and PFA-2, and nine minor subtypes were discovered. Transcriptome profiling suggested a distinct histogenesis for PFA-1 and PFA-2, but their clinical parameters were similar. In contrast, PFA subtypes differed with respect to age at diagnosis, gender ratio, outcome, and frequencies of genetic alterations. One subtype, PFA-1c, was enriched for 1q gain and had a relatively poor outcome, while patients with PFA-2c ependymomas showed an overall survival at 5 years of > 90%. Unlike other ependymomas, PFA-2c tumors express high levels of OTX2, a potential biomarker for this ependymoma subtype with a good prognosis. We also discovered recurrent mutations among PFA ependymomas. H3 K27M mutations were present in 4.2%, occurring only in PFA-1 tumors, and missense mutations in an uncharacterized gene, CXorf67, were found in 9.4% of PFA ependymomas, but not in other groups. We detected high levels of wildtype or mutant CXorf67 expression in all PFA subtypes except PFA-1f, which is enriched for H3 K27M mutations. PFA ependymomas are characterized by lack of H3 K27 trimethylation (H3 K27-me3), and we tested the hypothesis that CXorf67 binds to PRC2 and can modulate levels of H3 K27-me3. Immunoprecipitation/mass spectrometry detected EZH2, SUZ12, and EED, core components of the PRC2 complex, bound to CXorf67 in the Daoy cell line, which shows high levels of CXorf67 and no expression of H3 K27-me3. Enforced reduction of CXorf67 in Daoy cells restored H3 K27-me3 levels, while enforced expression of CXorf67 in HEK293T and neural stem cells reduced H3 K27-me3 levels. Our data suggest that heterogeneity among PFA ependymomas could have clinicopathologic utility and that CXorf67 may have a functional role in these tumors

Quantification of dengue virus specific T cell responses and correlation with viral load and clinical disease severity in acute dengue infection

Background In order to understand the role of dengue virus (DENV) specific T cell responses that associate with protection, we studied their frequency and phenotype in relation to clinical disease severity and resolution of viraemia in a large cohort of patients with varying severity of acute dengue infection. Methodology/Principal findings Using ex vivo IFNγ ELISpot assays we determined the frequency of dengue viral peptide (DENV)-NS3, NS1 and NS5 responsive T cells in 74 adult patients with acute dengue infection and examined the association of responsive T cell frequency with the extent of viraemia and clinical disease severity. We found that total DENV-specific and DENV-NS3-specific T cell responses, were higher in patients with dengue fever (DF), when compared to those with dengue haemorrhagic fever (DHF). In addition, those with DF had significantly higher (p = 0.02) DENV-specific T cell responses on day 4 of infection compared to those who subsequently developed DHF. DENV peptide specific T cell responses inversely correlated with the degree of viraemia, which was most significant for DENV-NS3 specific T cell responses (Spearman’s r = -0.47, p = 0.0003). The frequency of T cell responses to NS1, NS5 and pooled DENV peptides, correlated with the degree of thrombocytopenia but had no association with levels of liver transaminases. In contrast, total DENV-IgG inversely correlated with the degree of thrombocytopenia and levels of liver transaminases. Conclusions/Significance Early appearance of DENV-specific T cell IFNγ responses before the onset of plasma leakage, appears to associate with milder clinical disease and resolution of viraemia, suggesting a protective role in acute dengue infection