Multislice T1‐prepared 2D single‐shot EPI: analysis of a clinical T1 mapping method unbiased by B0 or B1 inhomogeneity

Quantitative MR imaging is as sensitive in detecting lesions as qualitative imaging, but it is potentially more specific in differentiating disease. T₁ mapping in particular might help to assess acute ischemic stroke, multiple sclerosis, epilepsy and Alzheimer's disease better. Thus, a rapid and robust clinical technique is vital. In 1990, Ordidge and colleagues developed the multislice T₁ ‐prepared two‐dimensional (2D) single‐shot echo planar imaging technique. Subsequent studies demonstrated its clinical viability, but none performed an in‐depth analysis of the strengths and advantages of this T₁ mapping method. Herein, theoretical and experimental evidence shows that the technique accounts for 2D slice profile effects and is unbiased by B₀ or B₁ inhomogeneity. This is verified explicitly by varying the linear shims, the T₁ preparation flip angle and the excitation flip angle. Furthermore, it is shown that the repetition time (and hence scan time) can be reduced without a loss of T₁ accuracy. Copyright © 2016 John Wiley & Sons, Ltd.     

On the transmit field inhomogeneity correction of relaxation‐compensated amide and NOE CEST effects at 7 T

High field MRI is beneficial for chemical exchange saturation transfer (CEST) in terms of high SNR, CNR, and chemical shift dispersion. These advantages may, however, be counter‐balanced by the increased transmit field inhomogeneity normally associated with high field MRI. The relatively high sensitivity of the CEST contrast to B₁ inhomogeneity necessitates the development of correction methods, which is essential for the clinical translation of CEST. In this work, two B₁ correction algorithms for the most studied CEST effects, amide‐CEST and nuclear Overhauser enhancement (NOE), were analyzed. Both methods rely on fitting the multi‐pool Bloch‐McConnell equations to the densely sampled CEST spectra. In the first method, the correction is achieved by using a linear B₁ correction of the calculated amide and NOE CEST effects. The second method uses the Bloch‐McConnell fit parameters and the desired B₁ amplitude to recalculate the CEST spectra, followed by the calculation of B₁‐corrected amide and NOE CEST effects. Both algorithms were systematically studied in Bloch‐McConnell equations and in human data, and compared with the earlier proposed ideal interpolation‐based B₁ correction method. In the low B₁ regime of 0.15–0.50 μT (average power), a simple linear model was sufficient to mitigate B₁ inhomogeneity effects on a par with the interpolation B₁ correction, as demonstrated by a reduced correlation of the CEST contrast with B₁ in both the simulations and the experiments.     

Estimating B1+ in the breast at 7 T using a generic template

Dynamic contrast‐enhanced MRI is the workhorse of breast MRI, where the diagnosis of lesions is largely based on the enhancement curve shape. However, this curve shape is biased by RF transmit (B₁⁺) field inhomogeneities. B₁⁺ field information is required in order to correct these. The use of a generic, coil‐specific B₁⁺ template is proposed and tested. Finite‐difference time‐domain simulations for B₁⁺ were performed for healthy female volunteers with a wide range of breast anatomies. A generic B₁⁺ template was constructed by averaging simulations based on four volunteers. Three‐dimensional B₁⁺ maps were acquired in 15 other volunteers. Root mean square error (RMSE) metrics were calculated between individual simulations and the template, and between individual measurements and the template. The agreement between the proposed template approach and a B₁⁺ mapping method was compared against the agreement between acquisition and reacquisition using the same mapping protocol. RMSE values (% of nominal flip angle) comparing individual simulations with the template were in the range 2.00‐4.01%, with mean 2.68%. RMSE values comparing individual measurements with the template were in the range8.1‐16%, with mean 11.7%. The agreement between the proposed template approach and a B₁⁺ mapping method was only slightly worse than the agreement between two consecutive acquisitions using the same mapping protocol in one volunteer: the range of agreement increased from ±16% of the nominal angle for repeated measurement to ±22% for the B₁⁺ template. With local RF transmit coils, intersubject differences in B₁⁺ fields of the breast are comparable to the accuracy of B₁⁺ mapping methods, even at 7 T. Consequently, a single generic B₁⁺ template suits subjects over a wide range of breast anatomies, eliminating the need for a time‐consuming B₁⁺ mapping protocol.     

B7‐H1 and CD8+ Treg: The enigmatic role of B7‐H1 in peripheral tolerance

The interaction between B7‐H1 (PD‐L1) expressed on APC with PD‐1 expressed by T cells was shown previously to result in inhibition of T‐cell activation and autoimmune diseases. A paper in this issue of the European Journal of Immunology demonstrates that DC B7‐H1 expression can in fact enhance autoimmunity, rather than suppress it. Using a model of direct injection of self antigen‐loaded DC into the CNS, the authors demonstrate that DC with intact B7‐H1 expression exacerbate CNS autoimmune disease. Importantly, the improved disease outcome in animals treated with B7‐H1^?/? DC is a result of a population of CD8⁺ Treg cells that expand at the site of autoimmune inflammation.     

A novel method for simultaneous 3D B(1) and T(1) mapping: the method of slopes (MoS)

A novel three-dimensional simultaneous B(1) and T(1) mapping method is introduced: the method of slopes (MoS). The linearity of the spoiled gradient recalled echo (SPGR) signal vs flip angle relation is exploited: B(1) mapping is achieved by a two-point extrapolation to signal null with a correction scheme while T(1) mapping uses the slopes of the SPGR signal vs flip angle curves near the origin and near the signal null. This new method improves upon the existing variable flip angle (VFA) T(1)-mapping method in that (i) consistency between B(1) and T(1) maps is ensured (ii) the sampling scheme is T(1)-independent (iii) the noise bias and singularity, associated with using a linear form for the SPGR signal equation, is eliminated by using the full equation. The method is shown to yield accurate and robust results via simulations. Initial estimates of B(1) and T(1) values are obtained from three data points via simple computations and straight line approximations. Initial estimates of B(1) values, for a range of values, are shown to be accurate due to the proposed B(1) correction scheme. The accuracy and robustness of T(1) values is achieved via a non-linear fitting algorithm which includes a fourth data point sampled at high SNR. The MoS was validated by comparing resulting B(1) and T(1) maps with those obtained using other standard methods. Finally, the ability to obtain brain B(1) and T(1) maps using the MoS was demonstrated by in vivo experiments. The MoS is expected to perform well on other motion-free anatomical regions as well.     

A theoretical and experimental comparison of different techniques for B1 mapping at very high fields

With the increasing use of ultrahigh‐field MR with multiple transmit channels, mapping of the B₁⁺ field has become a critical factor in many studies, leading to the publication of a large number of sequences for the measurement of the flip angle in recent years. In this article, the accuracy, precision and practicability of some of the most prominent of these techniques are investigated both theoretically, using error propagation computations and Monte‐Carlo simulations, and experimentally for different settings. For an exemplary experiment, which is typical for high‐field applications, the flip angle uncertainty is calculated and measured for two‐ and three‐dimensional acquisitions for techniques based on both magnitude and phase data. Simulated and measured results show good agreement. An experimental assessment of T₁ and B₀ dependence yields weak variations with these parameters for only a few of the sequences. Measurements on human scanners show crucial influences of specific absorption rate limitations, especially at ultrahigh field. Copyright © 2012 John Wiley & Sons, Ltd.     

Novel HIV‐1 clade B candidate vaccines designed for HLA‐B*5101+ patients protected mice against chimaeric ecotropic HIV‐1 challenge

Novel candidate HIV‐1 vaccines have been constructed, which are tailor‐designed for HLA‐B^*5101⁺ patients infected with HIV‐1 clade B. These vaccines employ novel immunogen HIVB‐B^*5101 derived from consensus HIV‐1 clade B Gag p17 and p24 regions coupled to two Pol‐derived B^*5101‐restricted epitopes, which are together with a third B^*5101 epitope in Gag dominant in HIV‐1‐infected long‐term non‐progressing patients. Both plasmid DNA and modified vaccinia virus Ankara (MVA) vectors supported high expression levels of the HIVB‐B^*5101 immunogen in cultured cells. Heterologous DNA prime‐recombinant MVA boost regimen induced efficiently HIV‐1‐specific CD8⁺ T‐cell responses in BALB/c mice. These vaccine‐elicited T cells were multifunctional, killed efficiently target cells in vivo, and protected mice against challenge with ecotropic HIV‐1/NL4‐3 and ecotropic HIV‐1/NDK chimaeric viruses with HIV‐1 clade B or D backbones, respectively, and ecotropic murine leukemia virus gp80 envelope, and therefore did so in the absence of anti‐HIV‐1 gp120 antibodies. These results support further development of HIVB‐B^*5101 vaccines in combined heterologous‐modality regimens. The use of allele‐specific vaccines in humans is discussed in the context of other developments in the HIV‐1 field.     

Functional and molecular characterization of B cell-responsive Vδ1+ γδ T cells

Cells expressing the Vδ1⁺ gene segment are a minor γδ T cell population in human peripheral blood but predominate in epithelia and (inflamed) tissues. The characteristic dendritic-like morphology of these γδ T cells is consistent with their putative immune surveillance role in epithelia. Their function, however, remains unknown. We and others previously reported that a subset of Vδ1⁺ γδ T cells proliferates after stimulation with Epstein-Barr virus (EBV)-transformed B lymphoblastoid cell lines (LCL), but not with fresh peripheral blood-derived B cells. These responses were independent of the type of T cell receptor (TcR) γ chain co-expressed with the Vδ1 chain. The in vivo relevance of this LCL-mediated activation as well as the nature of the stimulatory ligand on the LCL is not well established. In this study, we tested the proliferative response of Vδ1⁺ LCL-responsive T cells against non-EBV-transformed B cells, activated through CD40 by murine EL4 B5 cells, and to a panel of B cell lines differing in the expression of EBV nuclear antigen proteins and adhesion/co-stimulatory molecules. The role of the Epstein-Barr virus-derived antigen in the induction of this response could be excluded as the activated (non-EBV-transformed) peripheral blood B cells were also able to induce a proliferative response in the LCL-responsive Vδ1⁺ T cells. Therefore, the stimulatory ligand on B cells is of cellular rather than of viral origin, and its expression is up-regulated upon activation of B cells. The expression of B7 and CD39 molecules on the surface of activated B cells appeared to be crucial since antibodies to these structures could block the induction of proliferation of the Vδ1⁺ T cells. Finally, we investigated the diversity of the responding Vδ1⁺ γδ T cell clones by sequence analysis of the TcRδ junctional regions. No restricted V-D-J sequences were found among the LCL-responsive Vδ1⁺ T cell clones, arguing strongly against a mono- or oligoclonal Vδ1⁺ γδ T cell response to LCL. These findings may explain the presence of polyclonally activated Vδ1⁺ T cells in inflamed tissues where activated B cells are often present.     

Differential presentation of endogenous and exogenous hepatitis B surface antigens influences priming of CD8+ T cells in an epitope‐specific manner

Little is known about whether presentation of endogenous and exogenous hepatitis B virus (HBV) surface antigens on APCs targeted by vaccination and/or virus‐harboring hepatocytes influences de novo priming of CD8⁺ T cells. We showed that surface antigen‐expressing transfectants exclusively display a K^b/S190 epitope, whereas cells pulsed with recombinant surface particles (rSPs) exclusively present a K^b/S208 epitope to CD8⁺ T cells. The differential presentation of these epitopes largely reflects the selective, but not exclusive, priming of K^b/S190‐ and K^b/S208‐specific T cells in C57BL/6 mice by endogenous/DNA‐ or exogenous/protein‐based vaccines, respectively. Silencing the K^b/S190 epitope (K^b/S190_V194F) in antigen‐expressing vectors rescued the presentation of the K^b/S208 epitope in stable transfectants and significantly enhanced priming of K^b/S208‐specific T cells in C57BL/6 mice. A K^b/S190‐mediated immunodominance operating in surface antigen‐expressing cells, but not in rSP‐pulsed cells, led to an efficient suppression in the presentation of the K^b/S208 epitope and a consequent decrease in the priming of K^b/S208‐specific T cells. This K^b/S190‐mediated immunodominance also operated in 1.4HBV‐S^mut transgenic (tg) hepatocytes selectively expressing endogenous surface antigens and allowed priming of K^b/S208‐ but not K^b/S190‐specific T cells in 1.4HBV‐S^mut tg mice. However, IFN‐γ⁺ K^b/S208‐specific T cells could not inhibit HBV replication in the liver of 1.4HBV‐S^mut tg mice. These results have practical implications for the design of T‐cell‐stimulating therapeutic vaccines.     

MCP‐1/CCR2 interactions direct migration of peripheral B and T lymphocytes to the thymus during acute infectious/inflammatory processes

Mature lymphocyte immigration into the thymus has been documented in mouse, rat, and pig models, and highly increases when cells acquire an activated phenotype. Entrance of peripheral B and T cells into the thymus has been described in healthy and pathological situations. However, it has not been proposed that leukocyte recirculation to the thymus could be a common feature occurring during the early phase of a Th1 inflammatory/infectious process when a large number of peripheral cells acquire an activated phenotype and the cellularity of the thymus is seriously compromised. The data we present here demonstrate that in well‐established Th1 models triggered by different types of immunogens, for example, LPS treatment (a bacterial product), Candida albicans infection (a fungus), and after Trypanosoma cruzi infection (a parasite), a large number of mature peripheral B and T cells enter the thymus. This effect is dependent on, but not exclusive of, the available space in the thymus. Our data also demonstrate that MCP‐1/CCR2 (where MCP‐1 is monocyte chemoattractant protein‐1) interaction is responsible for the infiltration of peripheral cells to the thymus in these Th1‐inflammatory/infectious situations. Finally, systemic expression of IL‐12 and IL‐18 produced during the inflammatory process is ultimately responsible for these migratory events.     

amamutdb.no: A Relational Database for MAN2B1 Allelic Variants that Compiles Genotypes,Clinical Phenotypes,and Biochemical and Structural Data of Mutant MAN2B1 in α‐Mannosidosis

α‐Mannosidosis is an autosomal recessive lysosomal storage disorder caused by mutations in the MAN2B1 gene, encoding lysosomal α‐mannosidase. The disorder is characterized by a range of clinical phenotypes of which the major manifestations are mental impairment, hearing impairment, skeletal changes, and immunodeficiency. Here, we report an α‐mannosidosis mutation database, amamutdb.no, which has been constructed as a publicly accessible online resource for recording and analyzing MAN2B1 variants ( http://amamutdb.no ). Our aim has been to offer structured and relational information on MAN2B1 mutations and genotypes along with associated clinical phenotypes. Classifying missense mutations, as pathogenic or benign, is a challenge. Therefore, they have been given special attention as we have compiled all available data that relate to their biochemical, functional, and structural properties. The α‐mannosidosis mutation database is comprehensive and relational in the sense that information can be retrieved and compiled across datasets; hence, it will facilitate diagnostics and increase our understanding of the clinical and molecular aspects of α‐mannosidosis. We believe that the amamutdb.no structure and architecture will be applicable for the development of databases for any monogenic disorder.     

Design and construction of a new recombinant fusion protein (2b2t+EPC1) and its assessment for serodiagnosis of cystic echinococcosis

The immunodiagnostic tests for cystic echinococcosis (CE) are mostly serological tests based on ELISA that use hydatid cyst antigens for primary screening because of its simple preparation and availability. The challenge to develop new serological methods (as compared to those based on the hydatid cyst fluid antigens) to meet the gold standard remains. Appropriate sources of antigenic material are necessary for application to improve the efficacy of immunodiagnostic tests at a population level. In the current study, a fusion protein containing the coding sequence of antigen B2t and two sequences of EPC1 antigen with some modifications was reconstructed. Using bioinformatics tools, these sequences were joined together by applying the sequence of a rigid α‐helix‐forming linker to obtain an appropriate structure of a fusion protein. Synthetic recombinant fusion protein was expressed using pET28a as a vector and evaluated by indirect ELISA test for sera from patients with hepatic CE and other parasitic infections. The sensitivity of the fusion protein was lower (88.46%) than the available ELISA kit (96.15%). However, the differences in sensitivity were not statistically significant as compared to the recombinant fusion peptide with the commercial kit (p = 0.269). The specificity of the recombinant fusion protein (95.45%) was not significantly lower than the commercial kit (96.59%; p = 1.000). Moreover, surprisingly there was no difference in the cross‐reactivity values of performance between the recombinant‐ELISA and commercial kit. The positive and negative predictive values of the recombinant antigen were achieved as 92% and 93.33%, respectively, while for the commercial kit, they were obtained as 94.33% and 97.70%, respectively. In conclusion, as an early evaluation of these antigens the performance of our recombinant fusion protein in ELISA is relatively promising. Although, it seemed that this peptide with specific antigenic epitopes might be more appropriate for the serological evaluation of CE by use of bioinformatics tools, our findings showed that cross‐reactions and a negative reaction could occur in clinical performance. This fusion protein may have utility for diagnosis in humans, but further evaluation is needed using the WHO ultrasound classification for CE.     

Constrained use of CCR5 on CD4+ lymphocytes by R5X4 HIV-1: Efficiency of Env-CCR5 interactions and low CCR5 expression determine a range of restricted CCR5-mediated entry

R5X4 HIV-1 has impaired utilization of CCR5 on primary CD4+ lymphocytes but the mechanisms responsible are not well defined. Using a panel of diverse R5X4 Envs we identified a spectrum of CCR5 use on CD4+ lymphocytes. Greater lymphocyte CCR5 use correlated with relative resistance to CCR5 mAbs and small molecule antagonists. Increasing CCR5 expression on lymphocytes increased the proportion of entry mediated by CCR5 for all R5X4 isolates except 89.6. In cell lines with regulated CCR5 expression, strains with greater lymphocyte CCR5 use better exploited limiting levels of CCR5. Introduction of an R306S mutation in the 89.6 V3 domain enhanced its utilization of CCR5 at low levels and switched its preference to CCR5 for lymphocyte entry. Thus, the degree to which R5X4 HIV-1 use primary lymphocyte CCR5 is determined by low CCR5 expression coupled with variations in the efficiency of Env-CCR5 interactions, which is in part governed by V3 sequences.     

Oestrogen treatment of experimental autoimmune encephalomyelitis requires 17β‐oestradiol‐receptor‐positive B cells that up‐regulate PD‐1 on CD4+ Foxp3+ regulatory T cells

It is now well accepted that sex hormones have immunoregulatory activity and may prevent exacerbations in multiple sclerosis during pregnancy. Our previous studies demonstrated that oestrogen (17β-oestradiol; E₂) protection against experimental autoimmune encephalomyelitis (EAE) is mediated mainly through oestrogen receptor-α (ERα) and the membrane receptor G-protein-coupled receptor 30 (GPR30) and is abrogated in the absence of B cells and the co-inhibitory receptor, Programmed Death-1 (PD-1). To critically evaluate the cell source of the E₂ and PD-1 co-inhibitory pathways in EAE regulation, we assessed the requirement for ERs on transferred B cells and downstream effects on expression of PD-1/PD-ligand on CD4⁺ Foxp3⁺ regulatory T (Treg) cells in B-cell-replenished, E₂-treated B-cell-deficient (μMT^−/−) mice with EAE. The results clearly demonstrated involvement of ERα and GPR30 on transferred B cells that mediated the protective E₂ treatment effect on EAE and further showed an E₂-mediated B-cell-dependent up-regulation of PD-1 on CD4⁺ Foxp3⁺ Treg cells. These findings identify regulatory B-cell populations as key players in potentiating Treg-cell activity during E₂-mediated protection against EAE.     

Plasmacytoid dendritic cells and type 1 interferon promote peripheral expansion of forkhead box protein 3+ regulatory T cells specific for the ubiquitous RNA‐binding nuclear antigen La/Sjögren's syndrome (SS)‐B

RNA‐binding nuclear antigens are a major class of self‐antigen to which immune tolerance is lost in rheumatic diseases. Serological tolerance to one such antigen, La/Sjögren's syndrome (SS)‐B (La), is controlled by CD4⁺ T cells. This study investigated peripheral tolerance to human La (hLa) by tracking the fate of hLa‐specific CD4⁺ T cells expressing the transgenic (Tg) 3B5.8 T cell receptor (TCR) after adoptive transfer into lymphocyte‐replete recipient mice expressing hLa as a neo‐self‐antigen. After initial antigen‐specific cell division, hLa‐specific donor CD4⁺ T cells expressed forkhead box protein 3 (FoxP3). Donor cells retrieved from hLa Tg recipients displayed impaired proliferation and secreted interleukin (IL)?10 in vitro in response to antigenic stimulation. Transfer of highly purified FoxP3‐negative donor cells demonstrated that accumulation of hLa‐specific regulatory T cells (T_reg) was due primarily to expansion of small numbers of donor T_reg. Depletion of recipient plasmacytoid dendritic cells (pDC), but not B cells, severely hampered the accumulation of FoxP3⁺ donor T_reg in hLa Tg recipients. Recipient pDC expressed tolerogenic markers and higher levels of co‐stimulatory and co‐inhibitory molecules than B cells. Adoptive transfer of hLa peptide‐loaded pDC into mice lacking expression of hLa recapitulated the accumulation of hLa‐specific T_reg. Blockade of the type 1 interferon (IFN) receptor in hLa Tg recipients of hLa‐specific T cells impaired FoxP3⁺ donor T cell accumulation. Therefore, peripheral expansion of T_reg specific for an RNA‐binding nuclear antigen is mediated by antigen‐presenting pDC in a type 1 IFN‐dependent manner. These results reveal a regulatory function of pDC in controlling autoreactivity to RNA‐binding nuclear antigens.