Probes for INS

Postnatal liver development is characterized by hepatocyte growth, proliferation and functional maturation. Notably, canonical Wnt signaling in hepatocytes has been identified as an important regulator of final adult liver size and metabolic liver zonation. The cellular origin of Wnt ligands responsible for homeostatic liver/body weight ratio remained unclear, which was also attributable to a lack of suitable endothelial Cre driver mice. To comprehensively analyze the effects of hepatic angiocrine Wnt signaling on liver development and metabolic functions, we used endothelial subtype-specific Stab2-Cre driver mice to delete Wls from hepatic endothelial cells (HEC). The resultant Stab2-Cretg/wt;Wlsfl/fl (Wls-HECKO) mice were viable but showed a significantly reduced liver/body weight ratio. Specifically, ablation of angiocrine Wnt signaling impaired metabolic zonation in the liver, as shown by loss of pericentral, β-catenin-dependent target genes such as Glutamine Synthase (Glul), RhBg, Axin2 and CYP2E1 as well as by extended expression of periportal genes such as Arginase 1 (Arg1). Furthermore, endothelial subtype-specific expression of a c-terminally YFP-tagged Wls fusion protein in Wls-HECKO mice (Stab2-Cretg/wt ;Wlsfl/fl;Rosa26:Wls-YFPfl/wt [Wls-rescue]) restored metabolic liver zonation. Interestingly, lipid metabolism was altered in Wls-HECKO miceexhibiting significantly reduced plasma cholesterol levels, while maintaining normal plasma triglyceride and blood glucose concentrations. On the contrary, zonal expression of Endomucin, LYVE1 and other markers of HEC heterogeneity were not altered in Wls-HECKO livers.

Background Aging is accompanied by altered thinking (cognition) and feeling (mood), functions that depend on information processing by brain cortical cell microcircuits. We hypothesized that age-associated long-term functional and biological changes are mediated by gene transcriptomic changes within neuronal cell-types forming cortical microcircuits, namely excitatory pyramidal cells (PYC) and inhibitory GABAergic neurons expressing vasoactive intestinal peptide (Vip), somatostatin (Sst) and parvalbumin (Pvalb). Methods To test this hypothesis, we assessed locomotor, anxiety-like and cognitive behavioral changes between young (2 months, n=9) and old (22 months, n=12) male C57BL/6 mice, and performed frontal cortex cell-type specific molecular profiling, using laser-capture microscopy and RNA sequencing. Results were analyzed by neuroinformatics and validated by fluorescent in situ hybridization. Results Old-mice displayed increased anxiety and reduced working memory. The four cell-types displayed distinct age-related transcriptomes and biological pathway profiles, affecting metabolic and cell signaling pathways, and selective markers of neuronal vulnerability (Ryr3), resilience (Oxr1), and mitochondrial dynamics (Opa1), suggesting high age-related vulnerability of PYCs, and variable degree of adaptation in GABAergic neurons. Correlations between gene expression and behaviors suggest that changes in cognition and anxiety associated with age are partly mediated by normal age-related cell changes, and that additional age-independent decreases in synaptic and signaling pathways, notably in PYC and SST-neurons further contribute to behavioral changes. Conclusions Our study demonstrates cell-dependent differential vulnerability and coordinated cell-specific cortical microcircuit molecular changes with age. Collectively, the results suggest intrinsic molecular links between aging, cognition and mood-related behaviors with SST-neurons contributing evenly to both behavioral conditions.

The heterogeneity of cellular states in cancer has been linked to drug resistance, cancer progression and the presence of cancer cells with properties of normal tissue stem cells. Secreted Wnt signals maintain stem cells in various epithelial tissues, including in lung development and regeneration. Here we show that mouse and human lung adenocarcinomas display hierarchical features with two distinct subpopulations, one with high Wnt signalling activity and another forming a niche that provides the Wnt ligand. The Wnt responder cells showed increased tumour propagation ability, suggesting that these cells have features of normal tissue stem cells. Genetic perturbation of Wnt production or signalling suppressed tumour progression. Small-molecule inhibitors targeting essential posttranslational modification of Wnt reduced tumour growth and markedly decreased the proliferative potential of lung cancer cells, leading to improved survival of tumour-bearing mice. These results indicate that strategies for disrupting pathways that maintain stem-like and niche cell phenotypes can translate into effective anti-cancer therapies.

Detailed characterization of the cell types in the human brain requires scalable experimental approaches to examine multiple aspects of the molecular state of individual cells, as well as computational integration of the data to produce unified cell-state annotations. Here we report improved high-throughput methods for single-nucleus droplet-based sequencing (snDrop-seq) and single-cell transposome hypersensitive site sequencing (scTHS-seq). We used each method to acquire nuclear transcriptomic and DNA accessibility maps for >60,000 single cells from human adult visual cortex, frontal cortex, and cerebellum. Integration of these data revealed regulatory elements and transcription factors that underlie cell-type distinctions, providing a basis for the study of complex processes in the brain, such as genetic programs that coordinate adult remyelination. We also mapped disease-associated risk variants to specific cellular populations, which provided insights into normal and pathogenic cellular processes in the human brain. This integrative multi-omics approach permits more detailed single-cell interrogation of complex organs and tissues.

Hair growth disorders often carry a major psychological burden. Therefore, more effective human hair growth-modulatory agents urgently need to be developed. Here, we used the hypertrichosis-inducing immunosuppressant, Cyclosporine A (CsA), as a lead compound to identify new hair growth-promoting molecular targets. Through microarray analysis we identified the Wnt inhibitor, secreted frizzled related protein 1 (SFRP1), as being down-regulated in the dermal papilla (DP) of CsA-treated human scalp hair follicles (HFs) ex vivo. Therefore, we further investigated the function of SFRP1 using a pharmacological approach and found that SFRP1 regulates intrafollicular canonical Wnt/β-catenin activity through inhibition of Wnt ligands in the human hair bulb. Conversely, inhibiting SFRP1 activity through the SFRP1 antagonist, WAY-316606, enhanced hair shaft production, hair shaft keratin expression, and inhibited spontaneous HF regression (catagen) ex vivo. Collectively, these data (a) identify Wnt signalling as a novel, non-immune-inhibitory CsA target; (b) introduce SFRP1 as a physiologically important regulator of canonical β-catenin activity in a human (mini-)organ; and (c) demonstrate WAY-316606 to be a promising new promoter of human hair growth. Since inhibiting SFRP1 only facilitates Wnt signalling through ligands that are already present, this 'ligand-limited' therapeutic strategy for promoting human hair growth may circumvent potential oncological risks associated with chronic Wnt over-activation.

Gut mesenchyme provides key stem cell niche signals such as Wnt ligands, but how these signals are regulated is unclear. Because Hedgehog (Hh) signaling is critical for gut mesenchymal development and tumorigenesis, we investigated Hh-mediated mechanisms by analyzing mice deleted for key negative regulators of Hh signaling, Sufu and/or Spop, in the gut mesenchyme, and demonstrated their dosage-dependent roles. Although these mutants exhibit abnormal mesenchymal cell growth and functionally defective muscle layers, villification is completed with proper mesenchymal clustering, implying a permissive role for Hh signaling. These mesenchymal defects are partially rescued by Gli2 reduction. Consistent with increased epithelial proliferation caused by abnormal Hh activation in development, Sufu reduction promotes intestinal tumorigenesis, whereas Gli2 heterozygosity suppresses it. Our analyses of chromatin and GLI2 binding genomic regions reveal its transcriptional regulation of stem cell niche signals through enhancers, providing mechanistic insight into the intestinal stem cell niche in development and tumorigenesis