Single-cell analysis of cardiogenesis reveals basis for organ-level developmental defects.

Organogenesis involves integration of diverse cell types; dysregulation of cell-type-specific gene networks results in birth defects, which affect 5% of live births. Congenital heart defects are the most common malformations, and result from disruption of discrete subsets of cardiac progenitor cells1, but the transcriptional changes in individual progenitors that lead to organ-level defects remain unknown. Here we used single-cell RNA sequencing to interrogate early cardiac progenitor cells as they become specified during normal and abnormal cardiogenesis, revealing how dysregulation of specific cellular subpopulations has catastrophic consequences. A network-based computational method for single-cell RNA-sequencing analysis that predicts lineage-specifying transcription factors2,3 identified Hand2 as a specifier of outflow tract cells but not right ventricular cells, despite the failure of right ventricular formation in Hand2-null mice4. Temporal single-cell-transcriptome analysis of Hand2-null embryos revealed failure of outflow tract myocardium specification, whereas right ventricular myocardium was specified but failed to properly differentiate and migrate. Loss of Hand2 also led to dysregulation of retinoic acid signalling and disruption of anterior-posterior patterning of cardiac progenitors. This work reveals transcriptional determinants that specify fate and differentiation in individual cardiac progenitor cells, and exposes mechanisms of disrupted cardiac development at single-cell resolution, providing a framework for investigating congenital heart defects

Context-Specific Transcription Factor Functions Regulate Epigenomic and Transcriptional Dynamics during Cardiac Reprogramming.

Ectopic expression of combinations of transcription factors (TFs) can drive direct lineage conversion, thereby reprogramming a somatic cell's identity. To determine the molecular mechanisms by which Gata4, Mef2c, and Tbx5 (GMT) induce conversion from a cardiac fibroblast toward an induced cardiomyocyte, we performed comprehensive transcriptomic, DNA-occupancy, and epigenomic interrogation throughout the reprogramming process. Integration of these datasets identified new TFs involved in cardiac reprogramming and revealed context-specific roles for GMT, including the ability of Mef2c and Tbx5 to independently promote chromatin remodeling at previously inaccessible sites. We also find evidence for cooperative facilitation and refinement of each TF's binding profile in a combinatorial setting. A reporter assay employing newly defined regulatory elements confirmed that binding of a single TF can be sufficient for gene activation, suggesting that co-binding events do not necessarily reflect synergy. These results shed light on fundamental mechanisms by which combinations of TFs direct lineage conversion

Context-Specific Transcription Factor Functions Regulate Epigenomic and Transcriptional Dynamics during Cardiac Reprogramming

Ectopic expression of combinations of transcription factors (TFs) can drive direct lineage conversion, thereby reprogramming a somatic cell's identity. To determine the molecular mechanisms by which Gata4, Mef2c, and Tbx5 (GMT) induce conversion from a cardiac fibroblast toward an induced cardiomyocyte, we performed comprehensive transcriptomic, DNA-occupancy, and epigenomic interrogation throughout the reprogramming process. Integration of these datasets identified new TFs involved in cardiac reprogramming and revealed context-specific roles for GMT, including the ability of Mef2c and Tbx5 to independently promote chromatin remodeling at previously inaccessible sites. We also find evidence for cooperative facilitation and refinement of each TF's binding profile in a combinatorial setting. A reporter assay employing newly defined regulatory elements confirmed that binding of a single TF can be sufficient for gene activation, suggesting that co-binding events do not necessarily reflect synergy. These results shed light on fundamental mechanisms by which combinations of TFs direct lineage conversion

Characterisation of emergent toxigenic M1UKStreptococcus pyogenes and associated sublineages

Supplementary Data File Supplementary Figure S1 Real-time PCR measurement of glpF2 transcription Supplementary Figure S2 GldA operon in different species  Supplementary Figure S3 SpeA production by non-invasive emm1 S. pyogenes  Supplementary Figure S4 String analysis of proteomic data: M1global vs. M1UK cellular fractions Supplementary Figure S5 String analysis of proteomic data: four M1 sublineages compared Supplementary Table S1  Strains used in experimental work Supplementary Table S2 Strains and WGS used for phylogenetic tree and SpeA quantification Supplementary Table S3 Chemically-defined medium composition  Supplementary Table S4 Primers used for RT-qPCR Numbers files: Proteomic comparisons for 5 strains per lineage of emm1 Streptococcus pyogenes cultured in chemically defined medium.  Experiment 1: M1global versus M1UK. 5 strains per group cultured in CDM. Bacterial preparations compared were: cell wall; cytosol; supernatant   Experiment 2; M1global versus M1_13snps versus M1_23snps versus M1UK. 5 strains per group cultured in CDM. Only cytosol compared. Excel spreadsheet Pairwise comparisons undertaken to creat volcano plots in Figures 4 and 5. Comparisons in Experiment 2 are M1global, M1_13snps, M1_23snps, and M1UK Excel sheet Pairwise comparisons used for volcano plots in Figures 4 and 5.</p

Empagliflozin in Patients with Chronic Kidney Disease

Background The effects of empagliflozin in patients with chronic kidney disease who are at risk for disease progression are not well understood. The EMPA-KIDNEY trial was designed to assess the effects of treatment with empagliflozin in a broad range of such patients. Methods We enrolled patients with chronic kidney disease who had an estimated glomerular filtration rate (eGFR) of at least 20 but less than 45 ml per minute per 1.73 m(2) of body-surface area, or who had an eGFR of at least 45 but less than 90 ml per minute per 1.73 m(2) with a urinary albumin-to-creatinine ratio (with albumin measured in milligrams and creatinine measured in grams) of at least 200. Patients were randomly assigned to receive empagliflozin (10 mg once daily) or matching placebo. The primary outcome was a composite of progression of kidney disease (defined as end-stage kidney disease, a sustained decrease in eGFR to &lt; 10 ml per minute per 1.73 m(2), a sustained decrease in eGFR of &amp; GE;40% from baseline, or death from renal causes) or death from cardiovascular causes. Results A total of 6609 patients underwent randomization. During a median of 2.0 years of follow-up, progression of kidney disease or death from cardiovascular causes occurred in 432 of 3304 patients (13.1%) in the empagliflozin group and in 558 of 3305 patients (16.9%) in the placebo group (hazard ratio, 0.72; 95% confidence interval [CI], 0.64 to 0.82; P &lt; 0.001). Results were consistent among patients with or without diabetes and across subgroups defined according to eGFR ranges. The rate of hospitalization from any cause was lower in the empagliflozin group than in the placebo group (hazard ratio, 0.86; 95% CI, 0.78 to 0.95; P=0.003), but there were no significant between-group differences with respect to the composite outcome of hospitalization for heart failure or death from cardiovascular causes (which occurred in 4.0% in the empagliflozin group and 4.6% in the placebo group) or death from any cause (in 4.5% and 5.1%, respectively). The rates of serious adverse events were similar in the two groups. Conclusions Among a wide range of patients with chronic kidney disease who were at risk for disease progression, empagliflozin therapy led to a lower risk of progression of kidney disease or death from cardiovascular causes than placebo