Macrophages Facilitate Electrical Conduction in the Heart.

Organ-specific functions of tissue-resident macrophages in the steady-state heart are unknown. Here, we show that cardiac macrophages facilitate electrical conduction through the distal atrioventricular node, where conducting cells densely intersperse with elongated macrophages expressing connexin 43. When coupled to spontaneously beating cardiomyocytes via connexin-43-containing gap junctions, cardiac macrophages have a negative resting membrane potential and depolarize in synchrony with cardiomyocytes. Conversely, macrophages render the resting membrane potential of cardiomyocytes more positive and, according to computational modeling, accelerate their repolarization. Photostimulation of channelrhodopsin-2-expressing macrophages improves atrioventricular conduction, whereas conditional deletion of connexin 43 in macrophages and congenital lack of macrophages delay atrioventricular conduction. In the Cd11bDTR mouse, macrophage ablation induces progressive atrioventricular block. These observations implicate macrophages in normal and aberrant cardiac conduction.

Perivascular Fibrosis Is Mediated by a KLF10-IL-9 Signaling Axis in CD4+ T Cells.

BACKGROUND: Perivascular fibrosis, characterized by increased amount of connective tissue around vessels, is a hallmark for vascular disease. Ang II (angiotensin II) contributes to vascular disease and end-organ damage via promoting T-cell activation. Despite recent data suggesting the role of T cells in the progression of perivascular fibrosis, the underlying mechanisms are poorly understood. METHODS: TF (transcription factor) profiling was performed in peripheral blood mononuclear cells of hypertensive patients. CD4-targeted KLF10 (Kruppel like factor 10)-deficient (Klf10fl/flCD4Cre+; [TKO]) and CD4-Cre (Klf10+/+CD4Cre+; [Cre]) control mice were subjected to Ang II infusion. End point characterization included cardiac echocardiography, aortic imaging, multiorgan histology, flow cytometry, cytokine analysis, aorta and fibroblast transcriptomic analysis, and aortic single-cell RNA-sequencing. RESULTS: TF profiling identified increased KLF10 expression in hypertensive human subjects and in CD4+ T cells in Ang II-treated mice. TKO mice showed enhanced perivascular fibrosis, but not interstitial fibrosis, in aorta, heart, and kidney in response to Ang II, accompanied by alterations in global longitudinal strain, arterial stiffness, and kidney function compared with Cre control mice. However, blood pressure was unchanged between the 2 groups. Mechanistically, KLF10 bound to the IL (interleukin)-9 promoter and interacted with HDAC1 (histone deacetylase 1) inhibit IL-9 transcription. Increased IL-9 in TKO mice induced fibroblast intracellular calcium mobilization, fibroblast activation, and differentiation and increased production of collagen and extracellular matrix, thereby promoting the progression of perivascular fibrosis and impairing target organ function. Remarkably, injection of anti-IL9 antibodies reversed perivascular fibrosis in Ang II-infused TKO mice and C57BL/6 mice. Single-cell RNA-sequencing revealed fibroblast heterogeneity with activated signatures associated with robust ECM (extracellular matrix) and perivascular fibrosis in Ang II-treated TKO mice. CONCLUSIONS: CD4+ T cell deficiency of Klf10 exacerbated perivascular fibrosis and multi-organ dysfunction in response to Ang II via upregulation of IL-9. Klf10 or IL-9 in T cells might represent novel therapeutic targets for treatment of vascular or fibrotic diseases.

Quantitative analysis of Y-Chromosome gene expression across 36 human tissues.

Little is known about how human Y-Chromosome gene expression directly contributes to differences between XX (female) and XY (male) individuals in nonreproductive tissues. Here, we analyzed quantitative profiles of Y-Chromosome gene expression across 36 human tissues from hundreds of individuals. Although it is often said that Y-Chromosome genes are lowly expressed outside the testis, we report many instances of elevated Y-Chromosome gene expression in a nonreproductive tissue. A notable example is EIF1AY, which encodes eukaryotic translation initiation factor 1A Y-linked, together with its X-linked homolog EIF1AX Evolutionary loss of a Y-linked microRNA target site enabled up-regulation of EIF1AY, but not of EIF1AX, in the heart. Consequently, this essential translation initiation factor is nearly twice as abundant in male as in female heart tissue at the protein level. Divergence between the X and Y Chromosomes in regulatory sequence can therefore lead to tissue-specific Y-Chromosome-driven sex biases in expression of critical, dosage-sensitive regulatory genes.

Coronavirus Disease 2019 (COVID-19) Coronary Vascular Thrombosis: Correlation with Neutrophil but Not Endothelial Activation.

Severe coronavirus disease 2019 (COVID-19) increases the risk of myocardial injury that contributes to mortality. This study used multiparameter immunofluorescence to extensively examine heart autopsy tissue of 7 patients who died of COVID-19 compared to 12 control specimens, with or without cardiovascular disease. Consistent with prior reports, no evidence of viral infection or lymphocytic infiltration indicative of myocarditis was found. However, frequent and extensive thrombosis was observed in large and small vessels in the hearts of the COVID-19 cohort, findings that were infrequent in controls. The endothelial lining of thrombosed vessels typically lacked evidence of cytokine-mediated endothelial activation, assessed as nuclear expression of transcription factors p65 (RelA), pSTAT1, or pSTAT3, or evidence of inflammatory activation assessed by expression of intracellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), tissue factor, or von Willebrand factor (VWF). Intimal EC lining was also generally preserved with little evidence of cell death or desquamation. In contrast, there were frequent markers of neutrophil activation within myocardial thrombi in patients with COVID-19, including neutrophil-platelet aggregates, neutrophil-rich clusters within macrothrombi, and evidence of neutrophil extracellular trap (NET) formation. These findings point to alterations in circulating neutrophils rather than in the endothelium as contributors to the increased thrombotic diathesis in the hearts of COVID-19 patients.

Triarylphosphine-Coordinated Bipyridyl Ru(II) Complexes Induce Mitochondrial Dysfunction.

While cancer cells rely heavily upon glycolysis to meet their energetic needs, reducing the importance of mitochondrial oxidative respiration processes, more recent studies have shown that their mitochondria still play an active role in the bioenergetics of metastases. This feature, in combination with the regulatory role of mitochondria in cell death, has made this organelle an attractive anticancer target. Here, we report the synthesis and biological characterization of triarylphosphine-containing bipyridyl ruthenium (Ru(II)) compounds and found distinct differences as a function of the substituents on the bipyridine and phosphine ligands. 4,4'-Dimethylbipyridyl-substituted compound 3 exhibited especially high depolarizing capabilities, and this depolarization was selective for the mitochondrial membrane and occurred within minutes of treatment in cancer cells. The Ru(II) complex 3 exhibited an 8-fold increase in depolarized mitochondrial membranes, as determined by flow cytometry, which compares favorably to the 2-fold increase observed by carbonyl cyanide chlorophenylhydrazone (CCCP), a proton ionophore that shuttles protons across membranes, depositing them into the mitochondrial matrix. Fluorination of the triphenylphosphine ligand provided a scaffold that maintained potency against a range of cancer cells but avoided inducing toxicity in zebrafish embryos at higher concentrations, displaying the potential of these Ru(II) compounds for anticancer applications. This study provides essential information regarding the role of ancillary ligands for the anticancer activity of Ru(II) coordination compounds that induce mitochondrial dysfunction.

Effects of Mineralocorticoid Receptor Blockade and Statins on Kidney Injury Marker 1 (KIM-1) in Female Rats Receiving L-NAME and Angiotensin II.

Kidney injury molecule-1 (KIM-1) is a biomarker of renal injury and a predictor of cardiovascular disease. Aldosterone, via activation of the mineralocorticoid receptor, is linked to cardiac and renal injury. However, the impact of mineralocorticoid receptor activation and blockade on KIM-1 is uncertain. We investigated whether renal KIM-1 is increased in a cardiorenal injury model induced by L-NAME/ANG II, and whether mineralocorticoid receptor blockade prevents the increase in KIM-1. Since statin use is associated with lower aldosterone, we also investigated whether administering eiSther a lipophilic statin (simvastatin) or a hydrophilic statin (pravastatin) prevents the increase in renal KIM-1. Female Wistar rats (8-10 week old), consuming a high salt diet (1.6% Na+), were randomized to the following conditions for 14 days: control; L-NAME (0.2 mg/mL in drinking water)/ANG II (225 ug/kg/day on days 12-14); L-NAME/ANG II + eplerenone (100 mg/kg/day p.o.); L-NAME/ANG II + pravastatin (20 mg/kg/day p.o.); L-NAME/ANG II + simvastatin (20 mg/kg/day p.o.). Groups treated with L-NAME/ANG II had significantly higher blood pressure, plasma and urine aldosterone, cardiac injury/stroke composite score, and renal KIM-1 than the control group. Both eplerenone and simvastatin reduced 24-h urinary KIM-1 (p = 0.0046, p = 0.031, respectively) and renal KIM-1 immunostaining (p = 0.004, p = 0.037, respectively). Eplerenone also reduced renal KIM-1 mRNA expression (p = 0.012) and cardiac injury/stroke composite score (p = 0.04). Pravastatin did not affect these damage markers. The 24-h urinary KIM-1, renal KIM-1 immunostaining, and renal KIM-1 mRNA expression correlated with cardiac injury/stroke composite score (p < 0.0001, Spearman ranked correlation = 0.69, 0.66, 0.59, respectively). In conclusion, L-NAME/ANG II increases renal KIM-1 and both eplerenone and simvastatin blunt this increase in renal KIM-1.

Natural and anthropogenic controls on lake water-level decline and evaporation-to-inflow ratio in the conterminous United States.

Lake water levels are integral to lake function, but hydrologic changes from land and water management may alter lake fluctuations beyond natural ranges. We constructed a conceptual model of multifaceted drivers of lake water-levels and evaporation-to-inflow ratio (Evap:Inflow). Using a structural equation modeling framework, we tested our model on 1) a national subset of lakes in the conterminous United States with minimal water management to describe natural drivers of lake hydrology and 2) five ecoregional subsets of lakes to explore regional variation in water management effects. Our model fit the national and ecoregional datasets and explained up to 47% of variation in Evap:Inflow, 38% of vertical water-level decline, and 79% of horizontal water-level decline (littoral exposure). For lakes with minimal water management, Evap:Inflow was related to lake depth (ß = -0.31) and surface inflow (ß = -0.44); vertical decline was related to annual climate (e.g., precipitation ß = -0.18) and water management (ß = -0.21); and horizontal decline was largely related to vertical decline (ß = 0.73) and lake morphometry (e.g., depth ß = -0.18). Anthropogenic effects varied by ecoregion and likely reflect differences in regional water management and climate. In the West, water management indicators were related to greater vertical decline (ß = 0.38), whereas in the Midwest, these indicators were related to more stable and full lake levels (ß = -0.22) even during drought conditions. National analyses show how human water use interacts with regional climate resulting in contrasting impacts to lake hydrologic variation in the US.

Trends and inequalities in distance to and use of nearest natural space in the context of the 20-min neighbourhood: A 4-wave national repeat cross-sectional study, 2013 to 2019.

The 20-min neighbourhood is a policy priority for governments worldwide; a key feature of this policy is providing access to natural space (NS) within 800 m of home. The study aims were to (1) examine the association between distance to nearest NS and frequent use over time and (2) examine whether frequent use and changes in use were patterned by income and housing tenure over time. Bi-annual Scottish Household Survey data were obtained for 2013 to 2019 (n:42128 aged 16+). Adults were asked the walking distance to their nearest NS, the frequency of visits to this space and their housing tenure, as well as age, sex and income. We examined the association between distance from home of nearest NS, housing tenure, and the likelihood of frequent NS use (visited once a week or more). Two-way interaction terms were further applied to explore variation in the association between tenure and frequent NS use over time. We found that 87% of respondents lived within 10 min walk of a NS, meeting the policy specification for a 20-min neighbourhood. Greater proximity to NS was associated with increased use; individuals living a 6-10 min walk and over 10 min walk were respectively 53% and 78% less likely to report frequent NS use than those living within a 5 min walk. Housing tenure was an important predictor of frequent NS use; private renters and homeowners were more likely to report frequent NS use than social renters. Our findings provide evidence that proximity to NS is a strong predictor of frequent use. Our study provides important evidence that time-based access measures alone do not consider deep-rooted socioeconomic variation in use of NS. Policy makers should ensure a nuanced lens is applied to operationalising and monitoring the 20-min neighbourhood to safeguard against exacerbating existing inequalities.

NEDD9 Is a Novel and Modifiable Mediator of Platelet-Endothelial Adhesion in the Pulmonary Circulation.

Rationale: Data on the molecular mechanisms that regulate platelet-pulmonary endothelial adhesion under conditions of hypoxia are lacking, but may have important therapeutic implications. Objectives: To identify a hypoxia-sensitive, modifiable mediator of platelet-pulmonary artery endothelial cell adhesion and thrombotic remodeling. Methods: Network medicine was used to profile protein-protein interactions in hypoxia-treated human pulmonary artery endothelial cells. Data from liquid chromatography-mass spectrometry and microscale thermophoresis informed the development of a novel antibody (Ab) to inhibit platelet-endothelial adhesion, which was tested in cells from patients with chronic thromboembolic pulmonary hypertension (CTEPH) and three animal models in vivo. Measurements and Main Results: The protein NEDD9 was identified in the hypoxia thrombosome network in silico. Compared with normoxia, hypoxia (0.2% O2) for 24 hours increased HIF-1α (hypoxia-inducible factor-1α)-dependent NEDD9 upregulation in vitro. Increased NEDD9 was localized to the plasma-membrane surface of cells from control donors and patients with CTEPH. In endarterectomy specimens, NEDD9 colocalized with the platelet surface adhesion molecule P-selectin. Our custom-made anti-NEDD9 Ab targeted the NEDD9-P-selectin interaction and inhibited the adhesion of activated platelets to pulmonary artery endothelial cells from control donors in vitro and from patients with CTEPH ex vivo. Compared with control mice, platelet-pulmonary endothelial aggregates and pulmonary hypertension induced by ADP were decreased in NEDD9-/- mice or wild-type mice treated with the anti-NEDD9 Ab, which also decreased chronic pulmonary thromboembolic remodeling in vivo. Conclusions: The NEDD9-P-selectin protein-protein interaction is a modifiable target with which to inhibit platelet-pulmonary endothelial adhesion and thromboembolic vascular remodeling, with potential therapeutic implications for patients with disorders of increased hypoxia signaling pathways, including CTEPH.

Severe triple vessel disease secondary to IgG4-related coronary periarteritis.

BACKGROUND: Immunoglobulin G4-related disease is a rare systemic inflammatory disease that can lead to vascular manifestations such as periarteritis. CASE PRESENTATION: A 41-year-old man with stress angina was referred for coronary bypass surgery due to triple vessel coronary disease. CONCLUSIONS: Operative findings revealed significant adhesions and dense peri-coronary and periaortic thickening, also involving the left internal mammary artery. The IgG4-associated disease was confirmed by aortic pathology. The stress angina subsequently improved with the initiation of treatment with prednisone and rituximab.

A web-based tool for assessing the condition of benthic diatom assemblages in streams and rivers of the conterminous United States.

Benthic diatom assemblages are known to be indicative of water quality but have yet to be widely adopted in biological assessments in the United States due to several limitations. Our goal was to address some of these limitations by developing regional multi-metric indices (MMIs) that are robust to inter-laboratory taxonomic inconsistency, adjusted for natural covariates, and sensitive to a wide range of anthropogenic stressors. We aggregated bioassessment data from two national-scale federal programs and used a data-driven analysis in which all-possible combinations of 2-7 metrics were compared for three measures of performance. After ranking the best-performing MMIs, we selected the final MMIs by evaluating stress-response relations in independent regional datasets of diatom samples paired with measures of several water-quality stressors, including herbicides and streamflow flashiness. Each regional MMI performed well at calibration sites and represented diverse aspects of the structure and function of diatom communities. Most metrics included in the best MMIs were modeled to account for natural variation including climate, topography, soil characteristics, lithology, and groundwater influence on streamflow. MMI performance improved with higher numbers of component metrics, but this effect diminished beyond six metrics. Component metrics of MMIs were associated with a broad suite of measured stressors in every region, including salinity, nutrients, herbicides, and streamflow flashiness. We provide a web-based software application that allows users in the conterminous United States to apply our MMIs to their own datasets and compare MMI scores from their sites to a broader regional context.

Physical habitat in conterminous US streams and rivers, Part 1: Geoclimatic controls and anthropogenic alteration.

Anthropogenic alteration of physical habitat structure in streams and rivers is increasingly recognized as a major cause of impairment worldwide. As part of their assessment of the status and trends in the condition of rivers and streams in the U.S., the U.S. Environmental Protection Agency's (USEPA) National Aquatic Resource Surveys (NARS) quantify and monitor channel size and slope, substrate size and stability, instream habitat complexity and cover, riparian vegetation cover and structure, anthropogenic disturbance activities, and channel-riparian interaction. Like biological assemblages and water chemistry, physical habitat is strongly controlled by natural geoclimatic factors that can obscure or amplify the influence of human activities. We developed a systematic approach to estimate the deviation of observed river and stream physical habitat from that expected in least-disturbed reference conditions. We applied this approach to calculate indices of anthropogenic alteration of three aspects of physical habitat condition in the conterminous U.S. (CONUS): streambed sediment size and stability, riparian vegetation cover, and instream habitat complexity. The precision and responsiveness of these indices led the USEPA to use them to evaluate physical habitat condition in CONUS rivers and streams. The scores of these indices systematically decreased with greater anthropogenic disturbance at river and stream sites in the CONUS and within ecoregions, which we interpret as a response of these physical habitat indices to anthropogenic influences. Although anthropogenic activities negatively influenced all three physical habitat indices in the least-disturbed sites within most ecoregions, natural geoclimatic and geomorphic factors were the dominant influences. For sites over the full range of anthropogenic disturbance, analyses of observed/expected sediment characteristics showed augmented flood flows and basin and riparian agriculture to be the leading predictors of streambed instability and excess fine sediments. Similarly, basin and riparian agriculture and non-agricultural riparian land uses were the leading predictors of reduced riparian vegetation cover complexity in the CONUS and within ecoregions. In turn, these reductions in riparian vegetation cover and complexity, combined with reduced summer low flows, were the leading predictors of instream habitat simplification. We conclude that quantitative measures of physical habitat structure are useful and important indicators of the impacts of human activities on stream and river condition.

Physical habitat in conterminous US streams and Rivers, part 2: A quantitative assessment of habitat condition.

Rigorous assessments of the ecological condition of water resources and the effect of human activities on those waters require quantitative physical, chemical, and biological data. The U.S. Environmental Protection Agency's river and stream surveys quantify river and stream bed particle size and stability, instream habitat complexity and cover, riparian vegetation cover and structure, and anthropogenic disturbance activities. Physical habitat is strongly controlled by natural geoclimatic factors that co-vary with human activities. We expressed the anthropogenic alteration of physical habitat as O/E ratios of observed habitat metric values divided by values expected under least-disturbed reference conditions, where site-specific expected values vary given their geoclimatic and geomorphic context. We set criteria for good, fair, and poor condition based on the distribution of O/E values in regional least-disturbed reference sites. Poor conditions existed in 22-24% of the 1.2 million km of streams and rivers in the conterminous U.S. for riparian human disturbance, streambed sediment and riparian vegetation cover, versus 14% for instream habitat complexity. Based on the same four indicators, the percentage of stream length in poor condition within 9 separate U.S. ecoregions ranged from 4% to 42%. Associations of our physical habitat indices with anthropogenic pressures demonstrate the scope of anthropogenic habitat alteration; habitat condition was negatively related to the level of anthropogenic disturbance nationally and in nearly all ecoregions. Relative risk estimates showed that streams and rivers with poor sediment, riparian cover complexity, or instream habitat cover conditions were 1.4 to 2.6 times as likely to also have fish or macroinvertebrate assemblages in poor condition. Our physical habitat condition indicators help explain deviations in biological conditions from those observed among least-disturbed sites and inform management actions for rehabilitating impaired waters and mitigating further ecological degradation.

Outcomes for Australian children with relapsed/refractory acute lymphoblastic leukaemia treated with blinatumomab.

We report on the Australian experience of blinatumomab for treatment of 24 children with relapsed/refractory precursor B-cell acute lymphoblastic leukaemia (B-ALL) and high-risk genetics, resulting in a minimal residual disease (MRD) response rate of 58%, 2-year progression-free survival (PFS) of 39% and 2-year overall survival of 63%. In total, 83% (n = 20/24) proceeded to haematopoietic stem cell transplant, directly after blinatumomab (n = 12) or following additional salvage therapy (n = 8). Four patients successfully received CD19-directed chimeric antigen receptor T-cell therapy despite prior blinatumomab exposure. Inferior 2-year PFS was associated with MRD positivity (20%, n = 15) and in KMT2A-rearranged infants (15%, n = 9). Our findings highlight that not all children with relapsed/refractory B-ALL respond to blinatumomab and factors such as blast genotype may affect prognosis.

Simulating the density reduction and equity impact of potential tobacco retail control policies.

BACKGROUND: Reducing the provision of tobacco is important for decreasing inequalities in smoking and smoking-related harm. Various policies have been proposed to achieve this, but their impacts-particularly on equity-are often unknown. Here, using national-level data, we simulate the impacts of potential policies designed to reduce tobacco outlet density (TOD). METHODS: Tobacco retailer locations (n=9030) were geocoded from Scotland's national register, forming a baseline. Twelve policies were developed in three types: (1) regulating type of retailer selling tobacco, (2) regulating location of tobacco sales, and (3) area-based TOD caps. Density reduction was measured as mean percentage reduction in TOD across data zones and number of retailers nationally. Equity impact was measured using regression-based Relative Index of Inequality (RII) across income deprivation quintiles. RESULTS: Policies restricting tobacco sales to a single outlet type ('Supermarket'; 'Liquor store'; 'Pharmacy') caused >80% TOD reduction and >90% reduction in the number of tobacco outlets nationally. However, RIIs indicated that two of these policies ('Liquor store', 'Pharmacy') increased socioeconomic inequalities in TOD. Equity-promoting policies included 'Minimum spacing' and exclusion zones around 'Child spaces'. The only policy to remove statistically significant TOD inequalities was the one deliberately targeted to do so ('Reduce clusters'). CONCLUSIONS: Using spatial simulations, we show that all selected policies reduced provision of tobacco retailing to varying degrees. However, the most 'successful' at doing so also increased inequalities. Consequently, policy-makers should consider how the methods by which tobacco retail density is reduced, and success measured, align with policy aims.

Using point-of-sale data to examine tobacco pricing across neighbourhoods in Scotland.

OBJECTIVES: To assess the geographical variation in tobacco price (cigarettes and roll-your-own (RYO) tobacco) in convenience stores across Scotland and how this relates to neighbourhood income deprivation, tobacco retail outlet density and urban/rural status. METHODS: Tobacco price data from 124 566 shopping baskets purchased in 274 convenience stores during 1 week in April 2018 were obtained through an electronic point-of-sale system. These data were combined with neighbourhood-level measures of income deprivation, tobacco retail outlet density and urban/rural status. We examined brand price for 12 of the most popular cigarette brands and 3 RYO brands and variations in purchases by price segment; multivariable regression analysis assessed associations between area variables and tobacco price. RESULTS: Most stores sold tobacco in all price segments. The lowest priced subvalue brands were the most popular in all neighbourhoods but were most dominant in shops in more deprived neighbourhoods. When total sales were assessed, overall purchase price varied significantly by neighbourhood income deprivation; packets of 20 cigarettes were 50 pence (5.6%) lower and RYO 34 pence (2.7%) lower among shops in the two highest income deprivation quintiles relative to the lowest. Analysis of individual brands showed that for 3 of the 12 cigarette brands considered, average prices were 12-17 pence lower in more deprived neighbourhoods with the most popular RYO brand 15 pence lower. There was limited evidence of a relationship with tobacco retail outlet density. CONCLUSION: Across Scottish convenience stores, the purchase price of cigarettes and RYO was lower in more income-deprived neighbourhoods. The lower prices primarily reflect greater sales of cheap brands in these areas, rather than retailers reducing the prices of individual brands.

Addition of Thiotepa to Total Body Irradiation and Cyclophosphamide Conditioning for Allogeneic Hematopoietic Stem Cell Transplantation in Pediatric Acute Lymphoblastic Leukemia.

Total body irradiation (TBI)/cyclophosphamide (CY) is a standard-of-care conditioning regimen in allogeneic hematopoietic stem cell transplant (HSCT) for pediatric acute lymphoblastic leukemia (ALL). This study sought to identify whether the addition of thiotepa (TT) to TBI/CY improves HSCT outcomes for pediatric patients with ALL. A retrospective analysis was performed on 347 pediatric ALL patients who underwent HSCT between 1995 and 2015, with 242 receiving TBI/CY/TT and 105 patients receiving TBI/CY. There were no statistical differences in age, donor source, or complete remission status between the 2 groups. Comparison of the TBI/CY/TT versus TBI/CY groups demonstrated no difference in transplant-related mortality at 1 (11% versus 11%), 5 (13% versus 16%), or 10 years (16% versus 16%). There was lower relapse in the TBI/CY/TT group at 1 (14% versus 26%), 5 (24% versus 36%), 10 (26% versus 37%), and 15 years (26% versus 37%) (P= .02) but was not statistically significant on multivariate analysis. The TBI/CY/TT group showed a trend toward improved disease-free survival (DFS) at 5 (59% versus 47%), 10 (56% versus 46%), and 15 years (49% versus 40%) (P = .05) but was not statistically significant on multivariate analysis. Comparing overall survival at 5 (62% versus 53%), 10 (57% versus 50%), and 15 years (50% versus 44%) demonstrated no statistical difference between the 2 groups. The addition of thiotepa to TBI/CY demonstrated no increase in transplant-related mortality for pediatric ALL HSCT but was unable to demonstrate significant benefit in disease control. Minimal residual disease status remained the key risk factor impacting both relapse and DFS. More studies are warranted to better clarify the benefits of using thiotepa in conditioning for ALL HSCT.

Diversity of XMEN Disease: Description of 2 Novel Variants and Analysis of the Lymphocyte Phenotype.

Variants in MAGT1 have been identified as the cause of an immune deficiency termed X-linked immunodeficiency with magnesium defect, Epstein-Barr virus (EBV) infection and neoplasia (XMEN) disease. Here, we describe 2 cases of XMEN disease due to novel mutations in MAGT1, one of whom presented with classical features of XMEN disease and another who presented with a novel phenotype including probable CNS vasculitis, HHV-8 negative multicentric Castelman disease and severe molluscum contagiosum, thus highlighting the clinical diversity that may be seen in this condition. Peripheral blood immunophenotyping of these 2 patients, together with an additional 4 XMEN patients, revealed reduced NKG2D expression, impaired CD28 expression on CD8+ T cells, CD4+ T cell lymphopenia, an inverted CD4:CD8 ratio and decreased memory B cells. In addition, we showed for the first time alterations to the CD8+ T cell memory compartment, reduced CD56hi NK cells, MAIT and iNKT cells, as well as compromised differentiation of naïve CD4+ T cells into IL-21-producing Tfh-type cells in vitro. Both patients were treated with supplemental magnesium with limited benefit. However, one patient has undergone allogeneic haematopoietic stem cell transplant, with full donor chimerism and immune reconstitution. These results expand our understanding of the clinical and immunological phenotype in XMEN disease, adding to the current literature, which we further discuss here.