Lineage-tracing hematopoietic stem cell origins in vivo to efficiently make human HLF+ HOXA+ hematopoietic progenitors from pluripotent stem cells.

The developmental origin of blood-forming hematopoietic stem cells (HSCs) is a longstanding question. Here, our non-invasive genetic lineage tracing in mouse embryos pinpoints that artery endothelial cells generate HSCs. Arteries are transiently competent to generate HSCs for 2.5 days (∼E8.5-E11) but subsequently cease, delimiting a narrow time frame for HSC formation in vivo. Guided by the arterial origins of blood, we efficiently and rapidly differentiate human pluripotent stem cells (hPSCs) into posterior primitive streak, lateral mesoderm, artery endothelium, hemogenic endothelium, and >90% pure hematopoietic progenitors within 10 days. hPSC-derived hematopoietic progenitors generate T, B, NK, erythroid, and myeloid cells in vitro and, critically, express hallmark HSC transcription factors HLF and HOXA5-HOXA10, which were previously challenging to upregulate. We differentiated hPSCs into highly enriched HLF+ HOXA+ hematopoietic progenitors with near-stoichiometric efficiency by blocking formation of unwanted lineages at each differentiation step. hPSC-derived HLF+ HOXA+ hematopoietic progenitors could avail both basic research and cellular therapies.

Microstructure and Mechanical Properties of Gradient Interfaces in Wire Arc Additive Remanufacturing of Hot Forging Die Steel.

Hot forging dies are subjected to periodic thermal stress and often fail in the forms of thermal fatigue, wear, plastic deformation, and fracture. A gradient multi-material wire arc additive remanufacturing method for hot forging dies was proposed to extend the service life of hot forging dies and reduce total production costs. The properties of multi-material gradient interfaces play a critical role in determining the overall performance of the final products. In this study, the remanufacturing zone of a hot forging die was divided into three deposition layers: the transition layer, the intermediate layer, and the strengthening layer. Experiments of wire arc additive manufacturing with gradient material were conducted on a 5CrNiMo hot forging die steel. The microstructure, microhardness, bonding strength, and impact property of gradient interfaces were characterized and analyzed. The results revealed that the gradient additive layers and their interfaces were defect-free and that the gradient interfaces had obtained a high-strength metallurgical bonding. The microstructure of the gradient additive layers presented a gradient transformation process of bainite-to-martensite from the bottom to the top layer. The microhardness gradually increased from the substrate layer to the surface-strengthening layer, forming a three-level gradient in the range of 100 HV. The impact toughness values of the three interfaces were 46.15 J/cm2, 54.96 J/cm2, and 22.53 J/cm2, and the impact fracture morphology ranged from ductile fracture to quasi-cleavage fracture. The mechanical properties of the gradient interfaces showed a gradient increase in hardness and strength, and a gradient decrease in toughness. The practical application of hot forging die remanufactured by the proposed method had an increase of 37.5% in average lifespan, which provided scientific support for the engineering application of the gradient multi-material wire arc additive remanufacturing of hot forging dies.

Generating human artery and vein cells from pluripotent stem cells highlights the arterial tropism of Nipah and Hendra viruses.

Stem cell research endeavors to generate specific subtypes of classically defined "cell types." Here, we generate >90% pure human artery or vein endothelial cells from pluripotent stem cells within 3-4 days. We specified artery cells by inhibiting vein-specifying signals and vice versa. These cells modeled viral infection of human vasculature by Nipah and Hendra viruses, which are extraordinarily deadly (∼57%-59% fatality rate) and require biosafety-level-4 containment. Generating pure populations of artery and vein cells highlighted that Nipah and Hendra viruses preferentially infected arteries; arteries expressed higher levels of their viral-entry receptor. Virally infected artery cells fused into syncytia containing up to 23 nuclei, which rapidly died. Despite infecting arteries and occupying ∼6%-17% of their transcriptome, Nipah and Hendra largely eluded innate immune detection, minimally eliciting interferon signaling. We thus efficiently generate artery and vein cells, introduce stem-cell-based toolkits for biosafety-level-4 virology, and explore the arterial tropism and cellular effects of Nipah and Hendra viruses.

Increased ACTL6A occupancy within mSWI/SNF chromatin remodelers drives human squamous cell carcinoma.

Mammalian SWI/SNF (BAF) chromatin remodelers play dosage-sensitive roles in many human malignancies and neurologic disorders. The gene encoding the BAF subunit actin-like 6a (ACTL6A) is amplified early in the development of many squamous cell carcinomas (SCCs), but its oncogenic role remains unclear. Here we demonstrate that ACTL6A overexpression leads to its stoichiometric assembly into BAF complexes and drives their interaction and engagement with specific regulatory regions in the genome. In normal epithelial cells, ACTL6A was substoichiometric to other BAF subunits. However, increased ACTL6A levels by ectopic expression or in SCC cells led to near saturation of ACTL6A within BAF complexes. Increased ACTL6A occupancy enhanced polycomb opposition genome-wide to activate SCC genes and facilitated the co-dependent loading of BAF and TEAD-YAP complexes on chromatin. Both mechanisms appeared to be critical and function as a molecular AND gate for SCC initiation and maintenance, thereby explaining the specificity of the role of ACTL6A amplification in SCCs.

Dedifferentiation: the return road to repair the intestinal epithelium.

In the March 5 issue of Cell Stem Cell, (Murata K et al. Cell Stem Cell. 26(377-390):e376 2020) reported that intestinal stem cell recovery after injury is principally through Ascl2-dependent dedifferentiation of absorptive and secretory precursors in mice. This study provides evidence for robust regenerative capability of the intestinal epithelium via dedifferentiation of absorptive and secretory progenitors in the crypt.

Single-cell transcriptome analysis reveals differential nutrient absorption functions in human intestine.

The intestine plays an important role in nutrient digestion and absorption, microbe defense, and hormone secretion. Although major cell types have been identified in the mouse intestinal epithelium, cell type-specific markers and functional assignments are largely unavailable for human intestine. Here, our single-cell RNA-seq analyses of 14,537 epithelial cells from human ileum, colon, and rectum reveal different nutrient absorption preferences in the small and large intestine, suggest the existence of Paneth-like cells in the large intestine, and identify potential new marker genes for human transient-amplifying cells and goblet cells. We have validated some of these insights by quantitative PCR, immunofluorescence, and functional analyses. Furthermore, we show both common and differential features of the cellular landscapes between the human and mouse ilea. Therefore, our data provide the basis for detailed characterization of human intestine cell constitution and functions, which would be helpful for a better understanding of human intestine disorders, such as inflammatory bowel disease and intestinal tumorigenesis.