Hematopoietic stem cells reversibly switch from dormancy to self-renewal during homeostasis and repair.

Bone marrow hematopoietic stem cells (HSCs) are crucial to maintain lifelong production of all blood cells. Although HSCs divide infrequently, it is thought that the entire HSC pool turns over every few weeks, suggesting that HSCs regularly enter and exit cell cycle. Here, we combine flow cytometry with label-retaining assays (BrdU and histone H2B-GFP) to identify a population of dormant mouse HSCs (d-HSCs) within the lin(-)Sca1+cKit+CD150+CD48(-)CD34(-) population. Computational modeling suggests that d-HSCs divide about every 145 days, or five times per lifetime. d-HSCs harbor the vast majority of multilineage long-term self-renewal activity. While they form a silent reservoir of the most potent HSCs during homeostasis, they are efficiently activated to self-renew in response to bone marrow injury or G-CSF stimulation. After re-establishment of homeostasis, activated HSCs return to dormancy, suggesting that HSCs are not stochastically entering the cell cycle but reversibly switch from dormancy to self-renewal under conditions of hematopoietic stress.

Deletion of the RNA-binding proteins ZFP36L1 and ZFP36L2 leads to perturbed thymic development and T lymphoblastic leukemia.

ZFP36L1 and ZFP36L2 are RNA-binding proteins (RBPs) that interact with AU-rich elements in the 3' untranslated region of mRNA, which leads to mRNA degradation and translational repression. Here we show that mice that lacked ZFP36L1 and ZFP36L2 during thymopoiesis developed a T cell acute lymphoblastic leukemia (T-ALL) dependent on the oncogenic transcription factor Notch1. Before the onset of T-ALL, thymic development was perturbed, with accumulation of cells that had passed through the beta-selection checkpoint without first expressing the T cell antigen receptor beta-chain (TCRbeta). Notch1 expression was higher in untransformed thymocytes in the absence of ZFP36L1 and ZFP36L2. Both RBPs interacted with evolutionarily conserved AU-rich elements in the 3' untranslated region of Notch1 and suppressed its expression. Our data establish a role for ZFP36L1 and ZFP36L2 during thymocyte development and in the prevention of malignant transformation.

Single cell tuning of Myc expression by antigen receptor signal strength and interleukin-2 in T lymphocytes.

Myc controls the metabolic reprogramming that supports effector T cell differentiation. The expression of Myc is regulated by the T cell antigen receptor (TCR) and pro-inflammatory cytokines such as interleukin-2 (IL-2). We now show that the TCR is a digital switch for Myc mRNA and protein expression that allows the strength of the antigen stimulus to determine the frequency of T cells that express Myc. IL-2 signalling strength also directs Myc expression but in an analogue process that fine-tunes Myc quantity in individual cells via post-transcriptional control of Myc protein. Fine-tuning Myc matters and is possible as Myc protein has a very short half-life in T cells due to its constant phosphorylation by glycogen synthase kinase 3 (GSK3) and subsequent proteasomal degradation. We show that Myc only accumulates in T cells exhibiting high levels of amino acid uptake allowing T cells to match Myc expression to biosynthetic demands. The combination of digital and analogue processes allows tight control of Myc expression at the population and single cell level during immune responses.

Notch signaling in the immune system.

The Notch signaling pathway regulates many aspects of embryonic development, as well as differentiation processes and tissue homeostasis in multiple adult organ systems. Disregulation of Notch signaling is associated with several human disorders, including cancer. In the last decade, it became evident that Notch signaling plays important roles within the hematopoietic and immune systems. Notch plays an essential role in the development of embryonic hematopoietic stem cells and influences multiple lineage decisions of developing lymphoid and myeloid cells. Moreover, recent evidence suggests that Notch is an important modulator of T cell-mediated immune responses. In this review, we discuss Notch signaling in hematopoiesis, lymphocyte development, and function as well as in T cell acute lymphoblastic leukemia.

Hes1 is a critical but context-dependent mediator of canonical Notch signaling in lymphocyte development and transformation.

Although canonical Notch signaling regulates multiple hematopoietic lineage decisions including T cell and marginal zone B cell fate specification, the downstream molecular mediators of Notch function are largely unknown. We showed here that conditional inactivation of Hes1, a well-characterized Notch target gene, in adult murine bone marrow (BM) cells severely impaired T cell development without affecting other Notch-dependent hematopoietic lineages such as marginal zone B cells. Competitive mixed BM chimeras, intrathymic transfer experiments, and in vitro culture of BM progenitors on Delta-like-expressing stromal cells further demonstrated that Hes1 is required for T cell lineage commitment, but dispensable for Notch-dependent thymocyte maturation through and beyond the beta selection checkpoint. Furthermore, our data strongly suggest that Hes1 is essential for the development and maintenance of Notch-induced T cell acute lymphoblastic leukemia. Collectively, our studies identify Hes1 as a critical but context-dependent mediator of canonical Notch signaling in the hematopoietic system.

Developing HSCs become Notch independent by the end of maturation in the AGM region.

The first definitive hematopoietic stem cells (dHSCs) in the mouse emerge in the dorsal aorta of the embryonic day (E) 10.5 to 11 aorta-gonad-mesonephros (AGM) region. Notch signaling is essential for early HSC development but is dispensable for the maintenance of adult bone marrow HSCs. How Notch signaling regulates HSC formation in the embryo is poorly understood. We demonstrate here that Notch signaling is active in E10.5 HSC precursors and involves both Notch1 and Notch2 receptors, but is gradually downregulated while they progress toward dHSCs at E11.5. This downregulation is accompanied by gradual functional loss of Notch dependency. Thus, as early as at final steps in the AGM region, HSCs begin acquiring the Notch independency characteristic of adult bone marrow HSCs as part of the maturation program. Our data indicate that fine stage-dependent tuning of Notch signaling may be required for the generation of definitive HSCs from pluripotent cells.

Notch Signaling Regulates the Homeostasis of Tissue-Restricted Innate-like T Cells.

Although Notch signaling plays important roles in lineage commitment and differentiation of multiple cell types including conventional T cells, nothing is currently known concerning Notch function in innate-like T cells. We have found that the homeostasis of several well-characterized populations of innate-like T cells including invariant NKT cells (iNKT), CD8ααTCRαß small intestinal intraepithelial lymphocytes, and innate memory phenotype CD8 T cells is controlled by Notch. Notch selectively regulates hepatic iNKT cell survival via tissue-restricted control of B cell lymphoma 2 and IL-7Rα expression. More generally, Notch regulation of innate-like T cell homeostasis involves both cell-intrinsic and -extrinsic mechanisms and relies upon context-dependent interactions with Notch ligand-expressing fibroblastic stromal cells. Collectively, using conditional ablation of Notch receptors on peripheral T cells or Notch ligands on putative fibroblastic stromal cells, we show that Notch signaling is indispensable for the homeostasis of three tissue-restricted populations of innate-like T cells: hepatic iNKT, CD8ααTCRαß small intestinal intraepithelial lymphocytes, and innate memory phenotype CD8 T cells, thus supporting a generalized role for Notch in innate T cell homeostasis.

NLRC5 exclusively transactivates MHC class I and related genes through a distinctive SXY module.

MHC class II (MHCII) genes are transactivated by the NOD-like receptor (NLR) family member CIITA, which is recruited to SXY enhancers of MHCII promoters via a DNA-binding "enhanceosome" complex. NLRC5, another NLR protein, was recently found to control transcription of MHC class I (MHCI) genes. However, detailed understanding of NLRC5's target gene specificity and mechanism of action remained lacking. We performed ChIP-sequencing experiments to gain comprehensive information on NLRC5-regulated genes. In addition to classical MHCI genes, we exclusively identified novel targets encoding non-classical MHCI molecules having important functions in immunity and tolerance. ChIP-sequencing performed with Rfx5(-/-) cells, which lack the pivotal enhanceosome factor RFX5, demonstrated its strict requirement for NLRC5 recruitment. Accordingly, Rfx5-knockout mice phenocopy Nlrc5 deficiency with respect to defective MHCI expression. Analysis of B cell lines lacking RFX5, RFXAP, or RFXANK further corroborated the importance of the enhanceosome for MHCI expression. Although recruited by common DNA-binding factors, CIITA and NLRC5 exhibit non-redundant functions, shown here using double-deficient Nlrc5(-/-)CIIta(-/-) mice. These paradoxical findings were resolved by using a "de novo" motif-discovery approach showing that the SXY consensus sequence occupied by NLRC5 in vivo diverges significantly from that occupied by CIITA. These sequence differences were sufficient to determine preferential occupation and transactivation by NLRC5 or CIITA, respectively, and the S box was found to be the essential feature conferring NLRC5 specificity. These results broaden our knowledge on the transcriptional activities of NLRC5 and CIITA, revealing their dependence on shared enhanceosome factors but their recruitment to distinct enhancer motifs in vivo. Furthermore, we demonstrated selectivity of NLRC5 for genes encoding MHCI or related proteins, rendering it an attractive target for therapeutic intervention. NLRC5 and CIITA thus emerge as paradigms for a novel class of transcriptional regulators dedicated for transactivating extremely few, phylogenetically related genes.

The relationship between vitamin D status and muscle strength in young healthy adults from sunny climate countries currently living in the northeast of Scotland.

The current study examined the relationship between vitamin D status and muscle strength in young healthy adults: residents (>6 months) and newcomers (0-3 months), originally from sunny climate countries but currently living in the northeast of Scotland. Our longitudinal data found a positive, albeit small, relationship between vitamin D status and knee extensor isometric strength. INTRODUCTION: Vitamin D has been suggested to play a role in muscle health and function, but studies so far have been primarily in older populations for falls prevention and subsequent risk of fractures. METHODS: Vitamin D status was assessed in a healthy young adults from sunny climate countries (n = 71, aged 19-42 years) with 56% seen within 3 months of arriving in Aberdeen [newcomers; median (range) time living in the UK = 2 months (9-105 days)] and the remainder resident for >6 months [residents; 23 months (6-121 months)]. Participants attended visits every 3 months for 15 months. At each visit, fasted blood samples were collected for analysis of serum 25-hydroxyvitamin D [25(OH)D], parathyroid hormone (PTH), carboxy-terminal collagen crosslinks (CTX) and N-terminal propeptide of type I collagen (P1NP). Maximal voluntary contractions (MVC) were performed for grip strength (both arms) and for maximal isometric strength of the knee extensors (right knee). RESULTS: There were small seasonal variations in 25(OH)D concentrations within the newcomers and residents, but no seasonal variation in bone turnover markers. There was a positive, albeit small, association between 25(OH)D and knee extensor maximal isometric strength. Mixed modelling predicted that for each 1 nmol/L increase in 25(OH)D, peak torque would increase by 1 Nm (p = 0.04). CONCLUSIONS: This study suggests that vitamin D may be important for muscle health in young adults migrating from sunnier climates to high latitudes, yet the potential effect is small.

Longevity of daily oral vitamin D3 supplementation: differences in 25OHD and 24,25(OH)2D observed 2 years after cessation of a 1-year randomised controlled trial (VICtORy RECALL).

To determine how long vitamin D lasts after supplementation ceases, the marker of status was measured 2 and 3 years after a 1-year trial. Compared to placebo, the proportion of vitamin D-deficient women was still lower, if they had taken daily vitamin D3, after 2 years, indicating its longevity. INTRODUCTION: The purpose of this study was to determine longevity of vitamin D status following cessation of vitamin D3 supplementation, 2 and 3 years after a 1-year randomised, double-blind placebo controlled trial and to investigate possible predictive factors. METHODS: Caucasian non-smoking postmenopausal women randomised to ViCtORY (2009-2010), who had not taken vitamin D supplements since the trial ended, were invited to attend follow-up visits. Total 25-hydroxyvitamin D (25OHD) and 24,25-dihydroxyvitamin D (24,25OH2D) were measured by dual tandem mass spectrometry of serum samples following removal of protein and de-lipidation; the original randomised controlled trial (RCT) samples were re-analysed simultaneously. Vitamin D-binding protein (VDBP) was measured by monoclonal immunoassay. RESULTS: In March 2012 and March 2013, 159 women (mean (SD) age 67.6 (2.1) years) re-attended, equally distributed between the original treatment groups: daily vitamin D3 (400 IU, 1000 IU) and placebo. One month after the RCT ended (March 2010), the proportion of women in placebo, 400 IU and 1000 IU vitamin D3 groups, respectively, with 25OHD < 25 nmol/L was 15, 0 and 0 (chi-square p < 0.001, n = 46, 44, 54). After 2 years (March 2012), it was 22, 4 and 4% (p = 0.002, n = 50, 48, 57); after 3 years, it was 23, 13 and 15% (p = 0.429, n = 48, 45, 52). The respective proportions of women with 24,25OH2D < 2.2 nmol/L were 50, 2 and 2% (1 month, p < 0.001, n = 46, 44, 54); 42, 33 and 12% (2 years, p = 0.002, n = 50, 48, 57); and 45, 27 and 29% (3 years, p = 0.138, n = 47, 45, 51). VDBP was a predictor of circulating 25OHD longevity (beta for VDBP in µg/mL 0.736; 95% CI 0.216-1.255, p = 0.006) but not 24,25OH2D. CONCLUSION: Four hundred international units or 1000 IU of daily vitamin D3 showed benefits over placebo 2 years after supplementation ceased in keeping 25OHD > 25 nmol/L.

Vitamin E homologues α- and γ-tocopherol are not associated with bone turnover markers or bone mineral density in peri-menopausal and post-menopausal women.

UNLABELLED: In a large cohort of older women, we investigated the relationships that different forms of vitamin E may have with bone turnover markers and bone mineral density (BMD). We found a suggestive positive association between serum alpha-tocopherol and BMD at the femoral neck, but no other clinically relevant observations. INTRODUCTION: Vitamin E has anti-oxidant and anti-inflammatory properties hypothesized to benefit bone, but limited studies exist regarding its homologues. We examined circulating and dietary α- and γ-tocopherols with bone turnover markers (BTMs) and bone mineral density (BMD), and the role of inflammation in this relationship. METHODS: We performed two cross-sectional analyses from two visits (V2, 1997-1999, n = 3883; V3, 2007-2011, n = 2130) of the Aberdeen Prospective Osteoporosis Screening Study. Dietary and supplement intakes by food frequency questionnaire were assessed at both visits. V2 BTMs (urinary free pyridinoline and deoxypyridinoline, serum N-terminal propeptide of type 1 collagen) and V3 serum α- and γ-tocopherols, inflammatory markers (interleukin-6 [IL-6], serum amyloid A [SAA], high-sensitivity C-reactive protein [hs-CRP], E-selectin) and dual X-ray absorptiometry BMD at the femoral neck and lumbar spine were collected. Food sources of tocopherol homologues and diet-serum correlations were determined. The relationships between dietary tocopherols and BTMs (V2), and dietary and serum tocopherols with BMD (V3) were examined by multivariable regression (adjusting for age, cholesterol, inflammatory markers, carotenoids, body mass index, physical activity level, alcohol intake, smoking status and national deprivation category). RESULTS: Serum γ-tocopherol was associated with increasing concentrations of hs-CRP, SAA and E-selectin (P-trend all <0.0001), while α-tocopherol was associated with decreasing concentrations of IL-6 and hs-CRP (P-trend all <0.001). Controlling for covariates, serum α-tocopherol was positively associated with BMD at the femoral neck (ß = 0.002, P = 0.04) among those not reporting vitamin E supplementation. CONCLUSION: We did not find biologically meaningful results between dietary and tocopherol homologues with BTMs or BMD.

Reference data for jumping mechanography in Canadian children, adolescents and young adults.

OBJECTIVES: To provide age- and sex-specific reference data for mechanography-derived parameters of muscle function in Canadian children and youth using the single two-legged jump (S2LJ) with hands-on-waist. METHODS: Our sample included 2017 observations from 715 participants (9-21 years; 338 girls). Participants performed three S2LJ with hands-on-waist on a force platform (Leonardo Mechanograph, Novotec). Outcomes were maximum peak power (Pmax), Pmax/mass, peak force/body weight (Fmax/BW), force efficiency, maximum jump height (Hmax), and velocity (Vmax). We used the LMS method to construct age- and sex-specific percentile curves and mixed effects models to examine sex and ethnic differences. RESULTS: With the exception of Efficiency, mechanography outcomes were greater in girls (4-40%, p<0.05) than boys at age 9. Boys' advantage in mechanography parameters emerged in adolescence (age 11-13 years; 3-65%, p<0.05) and persisted into young adulthood, except for Fmax/BW which was not greater in boys until age 17 (4-10%, p<0.05). Mechanography outcomes were 3-9% (p<0.05) greater in Asian compared with white participants. CONCLUSIONS: We provide the first reference data for the S2LJ using the hands-on-waist protocol in children, youth and young adults. These data support previous findings using freely moving arms and can be used when evaluating muscle function in pediatric studies.

[Better Reporting of Interventions: Template for Intervention Description and Replication (TIDieR) Checklist and Guide]. / Die TIDieR Checkliste und Anleitung - ein Instrument für eine verbesserte Interventionsbeschreibung und Replikation.

Without a complete published description of interventions, clinicians and patients cannot reliably implement interventions that are shown to be useful, and other researchers cannot replicate or build on research findings. The quality of description of interventions in publications, however, is remarkably poor. To improve the completeness of reporting, and ultimately the replicability, of interventions, an international group of experts and stakeholders developed the Template for Intervention Description and Replication (TIDieR) checklist and guide. The process involved a literature review for relevant checklists and research, a Delphi survey of an international panel of experts to guide item selection, and a face-to-face panel meeting. The resultant 12-item TIDieR checklist (brief name, why, what (materials), what (procedure), who intervened, how, where, when and how much, tailoring, modifications, how well (planned), how well (actually carried out)) is an extension of the CONSORT 2010 statement (item 5) and the SPIRIT 2013 statement (item 11). While the emphasis of the checklist is on trials, the guidance is intended to apply across all evaluative study designs. This paper presents the TIDieR checklist and guide, with a detailed explanation of each item, and examples of good reporting. The TIDieR checklist and guide should improve the reporting of interventions and make it easier for authors to structure the accounts of their interventions, reviewers and editors to assess the descriptions, and readers to use the information.

LRF-mediated Dll4 repression in erythroblasts is necessary for hematopoietic stem cell maintenance.

Hematopoietic stem cells (HSCs) are the most primitive cells in the hematopoietic system and are under tight regulation for self-renewal and differentiation. Notch signals are essential for the emergence of definitive hematopoiesis in mouse embryos and are critical regulators of lymphoid lineage fate determination. However, it remains unclear how Notch regulates the balance between HSC self-renewal and differentiation in the adult bone marrow (BM). Here we report a novel mechanism that prevents HSCs from undergoing premature lymphoid differentiation in BM. Using a series of in vivo mouse models and functional HSC assays, we show that leukemia/lymphoma related factor (LRF) is necessary for HSC maintenance by functioning as an erythroid-specific repressor of Delta-like 4 (Dll4) expression. Lrf deletion in erythroblasts promoted up-regulation of Dll4 in erythroblasts, sensitizing HSCs to T-cell instructive signals in the BM. Our study reveals novel cross-talk between HSCs and erythroblasts, and sheds a new light on the regulatory mechanisms regulating the balance between HSC self-renewal and differentiation.

Deficits in distal radius bone strength, density and microstructure are associated with forearm fractures in girls: an HR-pQCT study.

UNLABELLED: Forearm fractures are common during growth. We studied bone strength in youth with a recent forearm fracture. In girls, suboptimal bone strength was associated with fractures. In boys, poor balance and physical inactivity may lead to fractures. Prospective studies will confirm these relationships and identify targets for prevention strategies. INTRODUCTION: The etiology of pediatric forearm fractures is unclear. Thus, we examined distal radius bone strength, microstructure, and density in children and adolescents with a recent low- or moderate-energy forearm fracture and those without forearm fractures. METHODS: We assessed the non-dominant (controls) and non-fractured (cases) distal radius (7% site) using high-resolution peripheral quantitative computed tomography (HR-pQCT) (Scanco Medical AG) in 270 participants (girls: cases n = 47, controls n = 61 and boys: cases n = 88, controls n = 74) aged 8-16 years. We assessed standard anthropometry, maturity, body composition (dual energy X-ray absorptiometry (DXA), Hologic QDR 4500 W) physical activity, and balance. We fit sex-specific logistic regression models for each bone outcome adjusting for maturity, ethnicity, height, and percent body fat. RESULTS: In girls, impaired bone strength (failure load, ultimate stress) and a high load-to-strength ratio were associated with low-energy fractures (odds ratios (OR) 2.8-4.3). Low total bone mineral density (Tt.BMD), bone volume ratio, trabecular thickness, and cortical BMD and thickness were also associated with low-energy fractures (ORs 2.0-7.0). In boys, low Tt.BMD, but not bone strength, was associated with low-energy fractures (OR = 1.8). Boys with low-energy fractures had poor balance and higher percent body fat compared with controls (p < 0.05). Boys with fractures (both types) were less active than controls (p < 0.05). CONCLUSIONS: Forearm fracture etiology appears to be sex-specific. In girls, deficits in bone strength are associated with fractures. In boys, a combination of poor balance, excess body fat, and low physical activity may lead to fractures. Prospective studies are needed to confirm these relationships and clarify targets for prevention strategies.

Bone micro-architecture of elite alpine skiers is not reflected by bone mineral density.

UNLABELLED: Bone quality is affected by muscle forces and external forces. We investigated how micro-architecture is influenced in elite alpine skiers who have received high loading levels throughout their adolescent bone development. Bone strength was higher in skiers, likely due to external forces, but muscle forces may also be a significant contributor. INTRODUCTION: Impact loading and muscle forces affect bone quality, but little is known about how they influence 3 dimensional aspects of bone structure. This study investigated bone quality in female and male elite alpine skiers using high-resolution peripheral quantitative computed tomography (HR-pQCT). METHODS: HR-pQCT at the distal radius and tibia, whole-body lean mass, and muscle strength were assessed in 10 female (22.7 ± 3.9 years) and 12 male (25.5 ± 3.3 years) Canadian national alpine team athletes and compared to recreationally active female (N = 10, 23.8 ± 3.2 years) and male (N = 12; 23.7 ± 3.6 years) control subjects. HR-pQCT standard parameters and customized cortical and finite element (FE) analyses were performed and analyzed using one-way ANOVA and Pearson's correlation. RESULTS: Male and female skiers had stronger bones than controls at radius (38-49 %, p < 0.001) and tibia (24-28 %, p < 0.001). This result was not consistently reflected by total bone mineral density (BMD) because higher trabecular BMD occurred in parallel with lower cortical BMD, which was due to a redistribution of mineral leading to a shift of the endocortical margin toward a thicker cortex. The endocortical regional adaptation was likely responsible for the greater strength of the athletes' bones. Lean mass and muscle strength was 29 to 90 % greater (p < 0.001) in athletes compared to controls. Good associations between muscle strength and FE-estimated bone strength were found (r = 0.63 to 0.80; p < 0.001), although micro-architecture was more strongly associated with muscle outcomes in females than males. CONCLUSIONS: Higher bone strength in elite alpine skiers is achieved through micro-architectural adaptation that is not apparent by BMD measurements alone. The improved micro-architecture at radius and tibia suggests that muscle forces may play an important role in bone adaptation.

Functional education of invariant NKT cells by dendritic cell tuning of SHP-1.

Invariant NKT (iNKT) cells play key roles in host defense by recognizing lipid Ags presented by CD1d. iNKT cells are activated by bacterial-derived lipids and are also strongly autoreactive toward self-lipids. iNKT cell responsiveness must be regulated to maintain effective host defense while preventing uncontrolled stimulation and potential autoimmunity. CD1d-expressing thymocytes support iNKT cell development, but thymocyte-restricted expression of CD1d gives rise to Ag hyperresponsive iNKT cells. We hypothesized that iNKT cells require functional education by CD1d(+) cells other than thymocytes to set their correct responsiveness. In mice that expressed CD1d only on thymocytes, hyperresponsive iNKT cells in the periphery expressed significantly reduced levels of tyrosine phosphatase SHP-1, a negative regulator of TCR signaling. Accordingly, heterozygous SHP-1 mutant mice displaying reduced SHP-1 expression developed a comparable population of Ag hyperresponsive iNKT cells. Restoring nonthymocyte CD1d expression in transgenic mice normalized SHP-1 expression and iNKT cell reactivity. Radiation chimeras revealed that CD1d(+) dendritic cells supported iNKT cell upregulation of SHP-1 and decreased responsiveness after thymic emigration. Hence, dendritic cells functionally educate iNKT cells by tuning SHP-1 expression to limit reactivity.

Notch signaling regulates follicular helper T cell differentiation.

Follicular helper T (TFH) cells are specialized in providing help for B cell differentiation and Ab secretion. Several positive and negative regulators of TFH cell differentiation have been described but their control is not fully understood. In this study, we show that Notch signaling in T cells is a major player in the development and function of TFH cells. T cell-specific gene ablation of Notch1 and Notch2 impaired differentiation of TFH cells in draining lymph nodes of mice immunized with T-dependent Ags or infected with parasites. Impaired TFH cell differentiation correlated with deficient germinal center development and the absence of high-affinity Abs. The impact of loss of Notch on TFH cell differentiation was largely independent of its effect on IL-4. These results show a previously unknown role for Notch in the regulation of TFH cell differentiation and function with implications for the control of this T cell population.

The fall and rise of corticomotor excitability with cancellation and reinitiation of prepared action.

The sudden cancellation of a motor action, known as response inhibition (RI), is fundamental to human motor behavior. The behavioral selectivity of RI can be studied by cueing cancellation of only a subset of a planned response, which markedly delays the remaining executed components. The present study examined neurophysiological mechanisms that may contribute to these delays. In two experiments, human participants received single- and paired-pulse transcranial magnetic stimulation while performing a bimanual anticipatory response task. Participants performed most trials bimanually (Go trials) and were sometimes cued to cancel the response with one hand while responding with the other (Partial trials). Motor evoked potentials were recorded from left first dorsal interosseous (FDI) as a measure of corticomotor excitability (CME) during Go and Partial trials. CME was temporally modulated during Partial trials in a manner that reflected anticipation, suppression, and subsequent initiation of a reprogrammed response. There was an initial increase in CME, followed by suppression 175 ms after the stop signal, even though the left hand was not cued to stop. A second increase in excitability occurred prior to the (delayed) response. We propose an activation threshold model to account for nonselective RI. To investigate the inhibitory component of our model, we investigated short-latency intracortical inhibition (sICI), but results indicated that sICI cannot fully explain the observed temporal modulation of CME. These neurophysiological and behavioural results indicate that the default mode for reactive partial cancellation is suppression of a unitary response, followed by response reinitiation with an inevitable time delay.

DL4-mediated Notch signaling is required for the development of fetal αß and γδ T cells.

T-cell development depends upon interactions between thymocytes and thymic epithelial cells (TECs). The engagement of delta-like 4 (DL4) on TECs by Notch1 expressed by blood-borne BM-derived precursors is essential for T-cell commitment in the adult thymus. In contrast to the adult, the earliest T-cell progenitors in the embryo originate in the fetal liver and migrate to the nonvascularized fetal thymus via chemokine signals. Within the fetal thymus, some T-cell precursors undergo programmed TCRγ and TCRδ rearrangement and selection, giving rise to unique γδ T cells. Despite these fundamental differences between fetal and adult T-cell lymphopoiesis, we show here that DL4-mediated Notch signaling is essential for the development of both αß and γδ T-cell lineages in the embryo. Deletion of the DL4 gene in fetal TECs results in an early block in αß T-cell development and a dramatic reduction of all γδ T-cell subsets in the fetal thymus. In contrast to the adult, no dramatic deviation of T-cell precursors to alternative fates was observed in the fetal thymus in the absence of Notch signaling. Taken together, our data reveal a common requirement for DL4-mediated Notch signaling in fetal and adult thymopoiesis.