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.     

Accurate,precise, simultaneous myocardial T1 and T2 mapping using a radial sequence with inversion recovery and T2 preparation

We propose a simultaneous myocardial T1 and T2 mapping technique using a radial sequence with inversion recovery and T2 preparation, which achieves high accuracy and precision, with T1 and T2 reproducibility similar to the Modified Look‐Locker Inversion recovery (MOLLI) sequence and the conventional bright blood T2 mapping technique, respectively. The sequence was developed by incorporating gold angle radial fast low angle shot (FLASH) readout combined with an inversion pulse and T2prep pulses. The extended Bloch equation simulation with slice profile correction (BLESSPC) algorithm was proposed to reconstruct T1 and T2 maps at the same time in a few seconds, while maintaining good T1 and T2 estimation accuracy. Accuracy and precision were compared among the proposed technique, MOLLI and conventional T2 mapping techniques using phantom studies, 10 healthy volunteers and three patients. In phantom studies, the proposed technique was more accurate than MOLLI (P < 0.05) while achieving similar precision (P = 0.3) in T1 estimation, and was more accurate (P < 0.05) and precise (P < 0.001) than conventional T2 mapping (two‐parameter fitting) in T2 estimation. In vivo, the proposed technique achieved significantly higher T1 values (P < 0.001) and similar reproducibility (P = 0.3) compared with MOLLI, with significantly lower T2 values (P < 0.001) and similar reproducibility (P = 0.6) compared with the conventional T2 mapping technique. Thus, the proposed radial T1‐T2 mapping technique allows for accurate, precise, simultaneous myocardial T1 and T2 mapping in an 11‐heartbeat single breath‐hold acquisition.     

Optimal echo times for multi‐gradient echo‐based B0 field‐mapping

B₀ field maps are used ubiquitously in neuroimaging, in disciplines ranging from magnetic resonance spectroscopy to temperature mapping and susceptibility‐weighted imaging. Most B₀ maps are acquired using standard gradient‐echo–based vendor‐provided sequences, often comprised of two echoes spaced a few milliseconds apart. Herein, we analyze the optimal spacing of echo times, defined as those maximizing precision—minimizing the standard deviation—for a fixed total acquisition time. Field estimation is carried out using a weighted least squares estimator. The standard deviation is shown to be approximately inversely proportional to the total acquisition time, suggesting a law of diminishing returns, whereby substantial gains are obtained up to a certain point, with little improvement beyond that point. Validations are provided in a phantom and a group of volunteers. Multi‐gradient echo sequences are readily available on all manufacturer platforms, making our recommendations straightforward to implement on any modern scanner.     

Imaging of the region of the osteochondral junction (OCJ) using a 3D adiabatic inversion recovery prepared ultrashort echo time cones (3D IR‐UTE‐cones) sequence at 3 T

The purpose of this study is to develop a 3D adiabatic inversion recovery prepared ultrashort echo time Cones (3D IR‐UTE‐Cones) sequence for high resolution and contrast imaging of the region of osteochondral junction (OCJ) of human knee joint using a clinical 3 T scanner. A feasibility study on direct imaging of the OCJ region was performed on a human patellar cartilage sample and on eight cadaveric knee joints using T₁‐weighted, proton density (PD)‐weighted and short‐T₂‐weighted 3D IR‐UTE‐Cones sequences. Contrast to noise ratio was measured to evaluate the effectiveness of the 3D IR‐UTE‐Cones sequences for selective imaging of the OCJ region. Computed tomography imaging was performed in parallel for the cadaveric knee joints. The optimized T₁‐weighted 3D IR‐UTE‐Cones sequence was used to image the knee joints of eight healthy volunteers and six patients with osteoarthritis (OA) to evaluate morphological changes in the OCJ region. Clinical PD‐ and T₂‐weighted FSE sequences were also performed for comparison. The T₁‐weighted 3D IR‐UTE‐Cones sequence showed high resolution and contrast bright band of the normal OCJ region in the cadaveric joints. Normal OCJ appearances were also seen in healthy volunteers. Abnormal OCJ regions, manifested as ill‐defined, focal loss or non‐visualization of the high intensity band adjacent to the subchondral bone plate, were observed in the knee joints of both ex vivo and in vivo OA patients. The 3D IR‐UTE‐Cones sequence can image OCJ regions ex vivo and in vivo, with abnormalities depicted with high resolution and contrast. The technique may be useful for demonstrating involvement of OCJ regions in early OA.     

Single breath‐hold 3D cardiac T1 mapping using through‐time spiral GRAPPA

The quantification of cardiac T₁ relaxation time holds great potential for the detection of various cardiac diseases. However, as a result of both cardiac and respiratory motion, only one two‐dimensional T₁ map can be acquired in one breath‐hold with most current techniques, which limits its application for whole heart evaluation in routine clinical practice. In this study, an electrocardiogram (ECG)‐triggered three‐dimensional Look–Locker method was developed for cardiac T₁ measurement. Fast three‐dimensional data acquisition was achieved with a spoiled gradient‐echo sequence in combination with a stack‐of‐spirals trajectory and through‐time non‐Cartesian generalized autocalibrating partially parallel acquisition (GRAPPA) acceleration. The effects of different magnetic resonance parameters on T₁ quantification with the proposed technique were first examined by simulating data acquisition and T₁ map reconstruction using Bloch equation simulations. Accuracy was evaluated in studies with both phantoms and healthy subjects. These results showed that there was close agreement between the proposed technique and the reference method for a large range of T₁ values in phantom experiments. In vivo studies further demonstrated that rapid cardiac T₁ mapping for 12 three‐dimensional partitions (spatial resolution, 2 × 2 × 8 mm³) could be achieved in a single breath‐hold of ~12 s. The mean T₁ values of myocardial tissue and blood obtained from normal volunteers at 3 T were 1311 ± 66 and 1890 ± 159 ms, respectively. In conclusion, a three‐dimensional T₁ mapping technique was developed using a non‐Cartesian parallel imaging method, which enables fast and accurate T₁ mapping of cardiac tissues in a single short breath‐hold.     

Ribavirin modulates the conversion of human CD4+ CD25− T cell to CD4+ CD25+ FOXP3+ T cell via suppressing interleukin‐10‐producing regulatory T cell

Because regulatory T (Treg) cells play an important role in modulating the immune system response against both endogenous and exogenous antigens, their control is critical to establish immunotherapy against autoimmune disorders, chronic viral infections and tumours. Ribavirin (RBV), an antiviral reagent used with interferon, is known to polarize the T helper (Th) 1/2 cell balance toward Th1 cells. Although the immunoregulatory mechanisms of RBV are not fully understood, it has been expected that RBV would affect T reg cells to modulate the Th1/2 cell balance. To confirm this hypothesis, we investigated whether RBV modulates the inhibitory activity of human peripheral CD4⁺ CD25⁺ CD127⁻ T cells in vitro. CD4⁺ CD25⁺ CD127⁻ T cells pre-incubated with RBV lose their ability to inhibit the proliferation of CD4⁺ CD25⁻ T cells. Expression of Forkhead box P3 (FOXP3) in CD4⁺ CD25⁻ T cells was down-modulated when they were incubated with CD4⁺ CD25⁺ CD127⁻ T cells pre-incubated with RBV without down-modulating CD45RO on their surface. In addition, transwell assays and cytokine-neutralizing assays revealed that this effect depended mainly on the inhibition of interleukin-10 (IL-10) produced from CD4⁺ CD25⁺ CD127⁻ T cells. These results indicated that RBV might inhibit the conversion of CD4⁺ CD25⁻ FOXP3⁻ naive T cells into CD4⁺ CD25⁺ FOXP3⁺ adaptive Treg cells by down-modulating the IL-10-producing Treg 1 cells to prevent these effector T cells from entering anergy and to maintain Th1 cell activity. Taken together, our findings suggest that RBV would be useful for both elimination of long-term viral infections such as hepatitis C virus infection and for up-regulation of tumour-specific cellular immune responses to prevent carcinogenesis, especially hepatocellular carcinoma.     

Loss of β‐arrestin 2 exacerbates experimental autoimmune encephalomyelitis with reduced number of Foxp3+ CD4+ regulatory T cells

β-Arrestins are well-known regulators and mediators of G protein-coupled receptor signalling, and accumulating evidence reveals that they are functionally involved in inflammation and autoimmune diseases. Of the two β-arrestins, β-arrestin 1 is documented to play regulatory roles in an animal model of multiple sclerosis (MS), whereas the role of β-arrestin 2 is less clear. Here, we show that β-arrestin 2-deficient mice displayed the exacerbated and sustained symptoms of experimental autoimmune encephalomyelitis (EAE), an animal model of MS. At the cellular level, deficiency of β-arrestin 2 led to a decreased number of Foxp3⁺ CD4⁺ regulatory T (Treg) cells in peripheral lymphoid organs of EAE mice. Consistently, our in vitro observations also revealed that loss of β-arrestin 2 impaired the conversion of Foxp3⁻ CD4⁺ T cells into Foxp3⁺ CD4⁺ inducible Treg cells. Taken together, our data suggest that β-arrestin 2 plays a regulatory role in MS, that is opposite to that of β-arrestin 1, in autoimmune diseases such as MS, which is at least partially through regulation of iTreg cell differentiation.     

Critical stoichiometric ratio of CD4+ CD25+ FoxP3+ regulatory T cells and CD4+ CD25− responder T cells influence immunosuppression in patients with B‐cell acute lymphoblastic leukaemia

Regulatory T (Treg) cells act to suppress activation of the immune system and thereby maintain immunological homeostasis and tolerance to self-antigens. The frequency and suppressing activity of Treg cells in general are high in different malignancies. We wanted to identify the role and regulation of CD4⁺ CD25⁺ FoxP3⁺ Treg cells in B-cell acute lymphoblastic leukaemia (B-ALL). We have included patients at diagnosis (n = 54), patients in clinical remission (n = 32) and normal healthy individuals (n = 35). These diagnosed patients demonstrated a lower number of CD4⁺ CD25⁺ cells co-expressing a higher level of FoxP3, interleukin-10, transforming growth factor-β and CD152/CTLA-4 than the normal population. Treg cells from patients showed a higher suppressive capability on CD4⁺ CD25⁻ responder T (Tresp) cells than normal. The frequency and immunosuppressive potential of CD4⁺ CD25⁺ FoxP3⁺ Treg cells became high with the progression of malignancy in B-ALL. Relative distribution of Tresp and Treg cells was only ˜5 : 1 in B-ALL but ˜35 : 1 in normal healthy individuals, further confirming the elevated immunosuppression in patients. A co-culture study at these definite ex vivo ratios, indicated that Treg cells from B-ALL patients exhibited higher immunosuppression than Treg cells from normal healthy individuals. After chemotherapy using the MCP841 protocol, the frequency of CD4⁺ CD25⁺ cells was gradually enhanced with the reduction of FoxP3, interleukin-10 positivity corresponded with disease presentation, indicating reduced immunosuppression. Taken together, our study indicated that the CD4⁺ CD25⁺ FoxP3⁺ Treg cells played an important role in immunosuppression, resulting in a positive disease-correlation in these patients. To the best of our knowledge, this is the first detailed report on the frequency, regulation and functionality of Treg cells in B-ALL.     

1α,25‐dihydroxyvitamin D3 acts via transforming growth factor‐β to up‐regulate expression of immunosuppressive CD73 on human CD4+ Foxp3– T cells

Vitamin D deficiency is associated with increased incidence and severity of various immune‐mediated diseases. Active vitamin D (1α,25‐dihydroxyvitamin D3; 1,25(OH)₂D3) up‐regulates CD4⁺ T‐cell expression of the purine ectonucleotidase CD39, a molecule that is associated with the generation of anti‐inflammatory adenosine. Here we aimed to investigate the direct impact of 1,25(OH)₂D3 on expression of the downstream ecto‐5′‐nucleotidase CD73 by human CD4 T cells, and components of the transforming growth factor‐β (TGF‐β) pathway, which have been implicated in the modulation of CD73 by murine T cells. At 10^?8 to 10^?7 m , 1,25(OH)₂D3 significantly increased expression of CD73 on peripheral human CD4⁺ T cells. Although 1,25(OH)₂D3 did not affect the mRNA expression of latent TGF‐β₁, 1,25(OH)₂D3 did up‐regulate expression of TGF‐β‐associated molecules [latency‐associated peptide (LAP), glycophorin A repetitions predominant (GARP), GP96, neuropilin‐1, thrombospondin‐1 and α_v integrin] which is likely to have contributed to the observed enhancement in TGF‐β bioactivity. CD73 was highly co‐expressed with LAP and GARP following 1,25(OH)₂D3 treatment, but unexpectedly, each of these cell surface molecules was expressed primarily on CD4⁺ Foxp3^– T cells, rather than CD4⁺ Foxp3⁺ T cells. Notably, neutralization of TGF‐β significantly impaired 1,25(OH)₂D3‐mediated induction of CD73. Collectively, we show that 1,25(OH)₂D3 enhances expression of CD73 on CD4⁺ Foxp3^– T cells in a process that is at least partially TGF‐β‐dependent. These data reveal an additional contributing mechanism by which vitamin D may be protective in immune‐mediated disease.     

Three‐dimensional ultrashort echo time cones T1ρ (3D UTE‐cones‐T1ρ) imaging

We report a novel three‐dimensional (3D) ultrashort echo time (UTE) sequence employing Cones trajectory and T_1ρ preparation (UTE‐Cones‐T_1ρ) for quantitative T_1ρ assessment of short T₂ tissues in the musculoskeletal system. A basic 3D UTE‐Cones sequence was combined with a spin‐locking preparation pulse for T_1ρ contrast. A relatively short TR was used to decrease the scan time, which required T₁ measurement and compensation using 3D UTE‐Cones data acquisitions with variable TRs. Another strategy to reduce the total scan time was to acquire multiple Cones spokes (N_sp) after each T_1ρ preparation and fat saturation. Four spin‐locking times (TSL = 0–20 ms) were acquired over 12 min, plus another 7 min for T₁ measurement. The 3D UTE‐Cones‐T_1ρ sequence was compared with a two‐dimensional (2D) spiral‐T_1ρ sequence for the imaging of a spherical CuSO₄ phantom and ex vivo meniscus and tendon specimens, as well as the knee and ankle joints of healthy volunteers, using a clinical 3‐T scanner. The CuSO₄ phantom showed a T_1ρ value of 76.5 ± 1.6 ms with the 2D spiral‐T_1ρ sequence, as well as 85.7 ± 3.6 and 89.2 ± 1.4 ms for the 3D UTE‐Cones‐T_1ρ sequences with N_sp of 1 and 5, respectively. The 3D UTE‐Cones‐T_1ρ sequence provided shorter T_1ρ values for the bovine meniscus sample relative to the 2D spiral‐T_1ρ sequence (10–12 ms versus 16 ms, respectively). The cadaveric human Achilles tendon sample could only be imaged with the 3D UTE‐Cones‐T_1ρ sequence (T_1ρ = 4.0 ± 0.9 ms), with the 2D spiral‐T_1ρ sequence demonstrating near‐zero signal intensity. Human studies yielded T_1ρ values of 36.1 ± 2.9, 18.3 ± 3.9 and 3.1 ± 0.4 ms for articular cartilage, meniscus and the Achilles tendon, respectively. The 3D UTE‐Cones‐T_1ρ sequence allows volumetric T_1ρ measurement of short T₂ tissues in vivo.     

The role of 1α,25‐dihydroxyvitamin D3 and cytokines in the promotion of distinct Foxp3+and IL‐10+ CD4+ T cells

1α,25‐Dihydroxyvitamin D3 (1α25VitD3) has potent immunomodulatory properties. We have previously demonstrated that 1α25VitD3 promotes human and murine IL‐10‐secreting CD4⁺ T cells. Because of the clinical relevance of this observation, we characterized these cells further and investigated their relationship with Foxp3⁺ regulatory T (Treg) cells. 1α25VitD3 increased the frequency of both Foxp3⁺ and IL‐10⁺ CD4⁺T cells in vitro. However, Foxp3 was increased at high concentrations of 1α25VitD3 and IL‐10 at more moderate levels, with little coexpression of these molecules. The Foxp3⁺ and IL‐10⁺ T‐cell populations showed comparable suppressive activity. We demonstrate that the enhancement of Foxp3 expression by 1α25VitD3 is impaired by IL‐10. 1α25VitD3 enables the selective expansion of Foxp3⁺ Treg cells over their Foxp3^? T‐cell counterparts. Equally, 1α25VitD3 maintains Foxp3⁺ expression by sorted populations of human and murine Treg cells upon in vitro culture. A positive in vivo correlation between vitamin D status and CD4⁺Foxp3⁺ T cells in the airways was observed in a severe pediatric asthma cohort, supporting the in vitro observations. In summary, we provide evidence that 1α25VitD3 enhances the frequency of both IL‐10⁺ and Foxp3⁺ Treg cells. In a translational setting, these data suggest that 1α25VitD3, over a broad concentration range, will be effective in enhancing the frequency of Treg cells.     

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.     

Triple‐echo steady‐state T2 relaxometry of the human brain at high to ultra‐high fields

Quantitative MRI techniques, such as T₂ relaxometry, have demonstrated the potential to detect changes in the tissue microstructure of the human brain with higher specificity to the underlying pathology than in conventional morphological imaging. At high to ultra‐high field strengths, quantitative MR‐based tissue characterization benefits from the higher signal‐to‐noise ratio traded for either improved resolution or reduced scan time, but is impaired by severe static (B₀) and transmit (B₁) field heterogeneities. The objective of this study was to derive a robust relaxometry technique for fast T₂ mapping of the human brain at high to ultra‐high fields, which is highly insensitive to B₀ and B₁ field variations. The proposed method relies on a recently presented three‐dimensional (3D) triple‐echo steady‐state (TESS) imaging approach that has proven to be suitable for fast intrinsically B₁‐insensitive T₂ relaxometry of rigid targets. In this work, 3D TESS imaging is adapted for rapid high‐ to ultra‐high‐field two‐dimensional (2D) acquisitions. The achieved short scan times of 2D TESS measurements reduce motion sensitivity and make TESS‐based T₂ quantification feasible in the brain. After validation in vitro and in vivo at 3 T, T₂ maps of the human brain were obtained at 7 and 9.4 T. Excellent agreement between TESS‐based T₂ measurements and reference single‐echo spin‐echo data was found in vitro and in vivo at 3 T, and T₂ relaxometry based on TESS imaging was proven to be feasible and reliable in the human brain at 7 and 9.4 T. Although prominent B₀ and B₁ field variations occur at ultra‐high fields, the T₂ maps obtained show no B₀‐ or B₁‐related degradations. In conclusion, as a result of the observed robustness, TESS T₂ may emerge as a valuable measure for the early diagnosis and progression monitoring of brain diseases in high‐resolution 2D acquisitions at high to ultra‐high fields. Copyright © 2014 John Wiley & Sons, Ltd.     

Rapid simultaneous acquisition of T1 and T2 mapping images using multishot double spin‐echo EPI and automated variations of TR and TE (ms‐DSEPI‐T12)

A rapid method of simultaneous T₁ and T₂ measurement is presented which uses a segmented echo‐planar readout with varying repetition times (TR) and echo times (TE). This method is useful in T₁ mapping for analysis of dynamic contrast enhanced MRI (DCE‐MRI), where T₁ can be used to estimate contrast agent concentration. In the application of this method to dynamic imaging, the equilibrium magnetization is measured on pre‐contrast images and incorporated into post‐contrast T₁ calculations for improved accuracy. Simultaneous T₂ measurement allows correction of T₂ effects in the T₁ map which may occur at high contrast agent concentrations, and is performed without significant imaging time penalty. Phantom and in vivo results show the usefulness of this technique for analysis of contrast enhancement kinetics. Accurate rapid contrast agent concentration measurement may be useful for analyzing the distribution and kinetics of contrast agents or labeled pharmaceuticals. Copyright © 2009 John Wiley & Sons, Ltd.     

NKG2D ligation relieves 2B4‐mediated NK‐cell self‐tolerance in mice

Along with MHC class I (MHCI), 2B4 provides nonredundant NK‐cell inhibition in mice. The immunoregulatory role of 2B4 has been increasingly appreciated in models of tumor and viral infection, however, the interactions among 2B4, MHCI, and other activating NK‐cell receptors remain uncertain. Here, we dissect the influence of two distinct inhibitory pathways in modulating NK‐cell‐mediated control of tumors expressing strong activating ligands, including RAE‐1γ. In vitro cytotoxicity and in vivo peritoneal clearance assays using MHCI⁺CD48⁺ (RMA‐neo), MHCI⁺CD48⁺RAE‐1γ (RMA‐RAE‐1γ), MHCI^?CD48⁺ (RMA‐S‐neo), and MHCI^?CD48⁺RAE‐1γ (RMA‐S‐RAE‐1γ) tumor lines demonstrated that NKG2D activation supersedes the inhibitory effect of both 2B4‐ and MHCI‐mediated immune‐tolerance systems. Furthermore, 2B4KO mice subcutaneously challenged with RMA‐neo and RMA‐S‐neo exhibited reduced tumor growth and significantly prolonged survival compared with WT mice, implying that 2B4 is constitutively engaged in the NK‐cell tolerance mechanism in vivo. Nevertheless, the inhibitory effect of 2B4 is significantly attenuated when NK cells encountered highly stressed tumor cells expressing RAE‐1γ, resulting in an immune response shift toward NK‐cell activation and tumor regression. Therefore, our data highlight the importance of the 2B4‐mediated inhibitory system as an alternate self‐tolerance mechanism, whose role can be modulated by the strength of activating receptor signaling within the tumor microenvironment.     

TGF‐β downregulates KLRG1 expression in mouse and human CD8+ T cells

The inhibitory receptor killer cell lectin‐like receptor G1 (KLRG1) and the integrin α_E (CD103) are expressed by CD8⁺ T cells and both are specific for E‐cadherin. However, KLRG1 ligation by E‐cadherin inhibits effector T‐cell function, whereas binding of CD103 to E‐cadherin enhances cell–cell interaction and promotes target cell lysis. Here, we demonstrate that KLRG1 and CD103 expression in CD8⁺ T cells from untreated and virus‐infected mice are mutually exclusive. Inverse correlation of KLRG1 and CD103 expression was also found in human CD8⁺ T cells‐infiltrating hepatocellular carcinomas. As TGF‐β is known to induce CD103 expression in CD8⁺ T cells, we examined whether this cytokine also regulates KLRG1 expression. Indeed, our data further reveal that TGF‐β signaling in mouse as well as in human CD8⁺ T cells downregulates KLRG1 expression. This finding provides a rationale for the reciprocal expression of KLRG1 and CD103 in different CD8⁺ T‐cell subsets. In addition, it points to the limitation of KLRG1 as a marker for terminally differentiated CD8⁺ T cells if lymphocytes from tissues expressing high levels of TGF‐β are analyzed.     

TGF‐β interactions with IL‐1 family members trigger IL‐4‐independent IL‐9 production by mouse CD4+ T cells

TGF‐β and IL‐4 were recently shown to selectively upregulate IL‐9 production by naïve CD4⁺ T cells. We report here that TGF‐β interactions with IL‐1α, IL‐1β, IL‐18, and IL‐33 have equivalent IL‐9‐stimulating activities that function even in IL‐4‐deficient animals. This was observed after in vitro antigenic stimulation of immunized or unprimed mice and after polyclonal T‐cell activation. Based on intracellular IL‐9 staining, all IL‐9‐producing cells were CD4⁺ and 80–90% had proliferated, as indicated by reduced CFSE staining. In contrast to IL‐9, IL‐13 and IL‐17 were strongly stimulated by IL‐1 and either inhibited (IL‐13) or were unaffected (IL‐17) by addition of TGF‐β. IL‐9 and IL‐17 production also differed in their dependence on IL‐2 and regulation by IL‐1/IL‐23. As IL‐9 levels were much lower in Th2 and Th17 cultures, our results identify TGF‐β/IL‐1 and TGF‐β/IL‐4 as the main control points of IL‐9 synthesis.     

Inhibition of R5‐tropic HIV type‐1 replication in CD4+ natural killer T cells by γδ T lymphocytes

After the development of highly active anti‐retroviral therapy, it became clear that the majority of emergent HIV‐1 is macrophage‐tropic and infects CD4⁺, CCR5‐expressing cells (R5‐tropic). There are three distinct cell populations, R5‐tropic, HIV‐1‐susceptible CD4⁺ cells: (i) natural killer T (NKT) cells, (ii) dendritic cells and macrophages, and (iii) tissue‐associated T cells residing primarily at mucosal surfaces. We have confirmed that CD4⁺ NKT cells derived from peripheral blood mononuclear cells (PBMCs) predominantly express CCR5 rather than CXCR4, whereas the reverse is true for CD4⁺ T cells derived from circulating PBMCs, and that R5‐tropic HIV‐1 expands efficiently in the CD4⁺ NKT cells. Moreover, when PBMCs depleted of CD8α⁺ cells were stimulated in the presence of α‐galactosylceramide (α‐GalCer) and R5‐tropic HIV‐1 [NL(AD8)], the production of HIV‐1 virions was not suppressed, whereas, similar to the untreated PBMCs, depletion of CD8β⁺ cells from PBMCs significantly inhibited virion production. These findings suggest that CD8αα⁺ but not CD8αβ⁺ cells may have the ability to inhibit R5‐tropic HIV‐1 replication in CD4⁺ NKT cells. Here, we show that co‐culturing R5‐tropic HIV‐1‐infected CD4⁺ NKT cells with CD8αα⁺ γδ T cells, in particular Vγ1Vδ1 cells, but not with CD8αα⁺ NKT cells or CD8αα⁺ dendritic cells, inhibits HIV‐1 replication mainly by secreting chemokines, such as macrophage inflammatory proteins 1α and 1β and RANTES. Collectively, these results indicate the importance of CD8αα⁺ γδ T cells in the control of R5‐tropic HIV‐1 replication and persistence in CD4⁺ NKT cells.