The Journal of clinical investigation
Yadav, VK;Berger, JM;Singh, P;Nagarajan, P;Karsenty, G;
PMID: 34905510 | DOI: 10.1172/JCI153752
Through their ability to regulate gene expression in most organs, glucocorticoid hormones influence numerous physiological processes and therefore are key regulators of organismal homeostasis. In bone, glucocorticoid hormones inhibit the expression of the hormone Osteocalcin for poorly understood reasons. Here we show that in a classical endocrine feedback loop, osteocalcin in return enhances the biosynthesis of glucocorticoid but also mineralocorticoid hormones (adrenal steroidogenesis) in rodents and primates. Conversely, inactivating osteocalcin signalling in adrenal glands significantly impairs adrenal growth and steroidogenesis in mice. Embryo-made osteocalcin is necessary for normal Sf1 expression in foetal adrenal cells and adrenal cell steroidogenic differentiation, it therefore determines the number of steroidogenic cells present in adrenal glands of adult animals. Embryonic not postnatal osteocalcin also governs adrenal growth, adrenal steroidogenesis, blood pressure, electrolyte equilibrium and the rise of circulating corticosterone during the acute stress response in adult offspring. This osteocalcin-dependent regulation of adrenal development and steroidogenesis occurs even in the absence of a functional of hypothalamus-pituitary-adrenal axis; this explains why osteocalcin administration during pregnancy promotes adrenal growth and steroidogenesis and improves survival of adrenocorticotropic hormone signalling-deficient animals. This study reveals that a bone-derived, embryonic hormone influences lifelong adrenal functions and organismal homeostasis in the mouse.
Jarmas, AE;Brunskill, EW;Chaturvedi, P;Salomonis, N;Kopan, R;
PMID: 34732708 | DOI: 10.1038/s41467-021-26626-9
Mammalian nephron endowment is determined by the coordinated cessation of nephrogenesis in independent niches. Here we report that translatome analysis in Tsc1+/- nephron progenitor cells from mice with elevated nephron numbers reveals how differential translation of Wnt antagonists over agonists tips the balance between self-renewal and differentiation. Wnt agonists are poorly translated in young niches, resulting in an environment with low R-spondin and high Fgf20 promoting self-renewal. In older niches we find increased translation of Wnt agonists, including R-spondin and the signalosome-promoting Tmem59, and low Fgf20, promoting differentiation. This suggests that the tipping point for nephron progenitor exit from the niche is controlled by the gradual increase in stability and possibly clustering of Wnt/Fzd complexes in individual cells, enhancing the response to ureteric bud-derived Wnt9b inputs and driving synchronized differentiation. As predicted by these findings, removing one Rspo3 allele in nephron progenitors delays cessation and increases nephron numbers in vivo.
Bertozzi, A;Wu, CC;Hans, S;Brand, M;Weidinger, G;
PMID: 34748730 | DOI: 10.1016/j.ydbio.2021.11.001
Zebrafish can achieve scar-free healing of heart injuries, and robustly replace all cardiomyocytes lost to injury via dedifferentiation and proliferation of mature cardiomyocytes. Previous studies suggested that Wnt/β-catenin signaling is active in the injured zebrafish heart, where it induces fibrosis and prevents cardiomyocyte cell cycling. Here, via targeting the destruction complex of the Wnt/β-catenin pathway with pharmacological and genetic tools, we demonstrate that Wnt/β-catenin activity is required for cardiomyocyte proliferation and dedifferentiation, as well as for maturation of the scar during regeneration. Using cardiomyocyte-specific conditional inhibition of the pathway, we show that Wnt/β-catenin signaling acts cell-autonomously to promote cardiomyocyte proliferation. Our results stand in contrast to previous reports and rather support a model in which Wnt/β-catenin signaling plays a positive role during heart regeneration in zebrafish.
Dada, LA;Welch, LC;Magnani, ND;Ren, Z;Han, H;Brazee, PL;Celli, D;Flozak, AS;Weng, A;Herrerias, MM;Kryvenko, V;Vadász, I;Runyan, CE;Abdala-Valencia, H;Shigemura, M;Casalino-Matsuda, SM;Misharin, AV;Budinger, GRS;Gottardi, CJ;Sznajder, JI;
PMID: 36626234 | DOI: 10.1172/jci.insight.159331
Persistent symptoms and radiographic abnormalities suggestive of failed lung repair are among the most common symptoms in patients with COVID-19 after hospital discharge. In mechanically ventilated patients with ARDS secondary to SARS-CoV-2 pneumonia, low tidal volumes to reduce ventilator-induced lung injury necessarily elevate blood CO2 levels, often leading to hypercapnia. The role of hypercapnia on lung repair after injury is not completely understood. Here, using a mouse model of hypercapnia exposure, cell lineage-tracing, spatial transcriptomics and 3D-cultures, we show that hypercapnia limits β-catenin signaling in AT2 cells, leading to their reduced proliferative capacity. Hypercapnia alters expression of major Wnts in PDGFRα+-fibroblasts from those maintaining AT2 progenitor activity towards those that antagonize β-catenin signaling thereby limiting progenitor function. Constitutive activation of β-catenin signaling in AT2 cells or treatment of organoid cultures with recombinant WNT3A protein bypasses the inhibitory effects of hypercapnia. Inhibition of AT2 proliferation in hypercapnic patients may contribute to impaired lung repair after injury, preventing sealing of the epithelial barrier, increasing lung flooding, ventilator dependency and mortality. .
Lav, R;Krivanek, J;Anthwal, N;Tucker, AS;
PMID: 36931279 | DOI: 10.1016/j.stemcr.2023.02.004
Stem cell regulation plays a crucial role during development and homeostasis. Here, an essential source of Wnts from Gli1+ stem/progenitor cells was identified in the murine molar. Loss of Wnt production in Gli1+ apical stem/progenitor cells led to loss of Axin2 at the root apex, mis-regulation of SOX9, loss of BMP and Hh signaling, and truncation of root development. In the absence of Wnt signals, the root epithelium lost its integrity and epithelial identity. This phenotype could be partially mimicked by loss of Sox9 in the Gli1 population. Stabilization of Wnt signaling in the apical papilla led to rapid unordered differentiation of hard tissues and fragmentation of the epithelial root sheath. Wnt signaling from Gli1+ stem/progenitor cells, therefore, orchestrates root development, coordinating mesenchymal and epithelial interactions via SOX9 to regulate stem/progenitor cell expansion and differentiation. Our results demonstrate that disparate stem/progenitor cell populations are unified in their fundamental signaling interactions.
A cellular and spatial map of the choroid plexus across brain ventricles and ages
Dani, N;Herbst, RH;McCabe, C;Green, GS;Kaiser, K;Head, JP;Cui, J;Shipley, FB;Jang, A;Dionne, D;Nguyen, L;Rodman, C;Riesenfeld, SJ;Prochazka, J;Prochazkova, M;Sedlacek, R;Zhang, F;Bryja, V;Rozenblatt-Rosen, O;Habib, N;Regev, A;Lehtinen, MK;
PMID: 33932339 | DOI: 10.1016/j.cell.2021.04.003
The choroid plexus (ChP) in each brain ventricle produces cerebrospinal fluid (CSF) and forms the blood-CSF barrier. Here, we construct a single-cell and spatial atlas of each ChP in the developing, adult, and aged mouse brain. We delineate diverse cell types, subtypes, cell states, and expression programs in epithelial and mesenchymal cells across ages and ventricles. In the developing ChP, we predict a common progenitor pool for epithelial and neuronal cells, validated by lineage tracing. Epithelial and fibroblast cells show regionalized expression by ventricle, starting at embryonic stages and persisting with age, with a dramatic transcriptional shift with maturation, and a smaller shift in each aged cell type. With aging, epithelial cells upregulate host-defense programs, and resident macrophages upregulate interleukin-1β (IL-1β) signaling genes. Our atlas reveals cellular diversity, architecture and signaling across ventricles during development, maturation, and aging of the ChP-brain barrier.
Am J Respir Crit Care Med. 2018 Dec 15.
Reyfman PA, Walter JM, Joshi N, Anekalla KR, McQuattie-Pimentel AC, Chiu S, Fernandez R, Akbarpour M, Chen CI, Ren Z, Verma R, Abdala-Valencia H, Nam K, Chi M, Han S, Gonzalez-Gonzalez FJ, Soberanes S, Watanabe S, Williams KJN, Flozak AS, Nicholson TT, Morgan VK, Winter DR, Hinchcliff M, Hrusch CL, Guzy RD, Bonham CA, Sperling AI, Bag R, Hamanaka RB, Mutlu GM, Yeldandi AV, Marshall SA, Shilatifard A, Amaral LAN, Perlman H, Sznajder JI, Argento AC, Gillespie CT, Dematte J, Jain M, Singer BD, Ridge KM, Lam AP, Bharat A, Bhorade SM, Gottardi CJ, Budinger GRS, Misharin AV.
PMID: 30554520 | DOI: 10.1164/rccm.201712-2410OC
Abstract RATIONALE: The contributions of diverse cell populations in the human lung to pulmonary fibrosis pathogenesis are poorly understood. Single-cell RNA sequencing can reveal changes within individual cell populations during pulmonary fibrosis that are important for disease pathogenesis. OBJECTIVES: To determine whether single-cell RNA sequencing can reveal disease-related heterogeneity within alveolar macrophages, epithelial cells or other cell types in lung tissue from subjects with pulmonary fibrosis compared with controls. METHODS: We performed single-cell RNA sequencing on lung tissue obtained from eight transplant donors and eight recipients with pulmonary fibrosis and on one bronchoscopic cryobiospy sample from a patient with idiopathic pulmonary fibrosis. We validated these data in using in situ RNA hybridization, immunohistochemistry, and bulk RNA-sequencing on flow-sorted cells from 22 additional subjects. MEASUREMENTS AND MAIN RESULTS: We identified a distinct, novel population of profibrotic alveolar macrophages exclusively in patients with fibrosis. Within epithelial cells, the expression of genes involved in Wnt secretion and response was restricted to non-overlapping cells. We identified rare cell populations including airway stem cells and senescent cells emerging during pulmonary fibrosis. We developed a web-based tool to explore these data. CONCLUSIONS: We generated a single cell atlas of pulmonary fibrosis. Using this atlas we demonstrated heterogeneity within alveolar macrophages and epithelial cells from subjects with pulmonary fibrosis. These results support the feasibility of discovery-based approaches using next generation sequencing technologies to identify signaling pathways for targeting in the development of personalized therapies for patients with pulmonary fibrosis.
Pepe-Mooney BJ, Dill MT, Alemany A, Ordovas-Montanes J, Matsushita Y, Rao A, Sen A, Miyazaki M, Anakk S, Dawson PA, Ono N, Shalek AK, van Oudenaarden A, Camargo FD.
PMID: 31080134 | DOI: 10.1016/j.stem.2019.04.004
The liver can substantially regenerate after injury, with both main epithelial cell types, hepatocytes and biliary epithelial cells (BECs), playing important roles in parenchymal regeneration. Beyond metabolic functions, BECs exhibit substantial plasticity and in some contexts can drive hepatic repopulation. Here, we performed single-cell RNA sequencing to examine BEC and hepatocyte heterogeneity during homeostasisand after injury. Instead of evidence for a transcriptionally defined progenitor-like BEC cell, we found significant homeostatic BEC heterogeneity that reflects fluctuating activation of a YAP-dependent program. This transcriptional signature defines a dynamic cellular state during homeostasis and is highly responsive to injury. YAP signaling is induced by physiological bile acids (BAs), required for BEC survival in response to BA exposure, and is necessary for hepatocyte reprogramming into biliary progenitors upon injury. Together, these findings uncover molecular heterogeneity within the ductal epithelium and reveal YAP as a protective rheostat and regenerative regulator in the mammalian liver.
Mukhopadhyay, B;Holovac, K;Schuebel, K;Mukhopadhyay, P;Cinar, R;Iyer, S;Marietta, C;Goldman, D;Kunos, G;
PMID: 36966147 | DOI: 10.1038/s41420-023-01400-6
The proliferation and differentiation of hepatic progenitor cells (HPCs) drive the homeostatic renewal of the liver under diverse conditions. Liver regeneration is associated with an increase in Axin2+Cnr1+ HPCs, along with a marked increase in the levels of the endocannabinoid anandamide (AEA). But the molecular mechanism linking AEA signaling to HPC proliferation and/or differentiation has not been explored. Here, we show that in vitro exposure of HPCs to AEA triggers both cell cycling and differentiation along with increased expression of Cnr1, Krt19, and Axin2. Mechanistically, we found that AEA promotes the nuclear localization of the transcription factor β-catenin, with subsequent induction of its downstream targets. Systemic analyses of cells after CRISPR-mediated knockout of the β-catenin-regulated transcriptome revealed that AEA modulates β-catenin-dependent cell cycling and differentiation, as well as interleukin pathways. Further, we found that AEA promotes OXPHOS in HPCs when amino acids and glucose are readily available as substrates, but AEA inhibits it when the cells rely primarily on fatty acid oxidation. Thus, the endocannabinoid system promotes hepatocyte renewal and maturation by stimulating the proliferation of Axin2+Cnr1+ HPCs via the β-catenin pathways while modulating the metabolic activity of their precursor cells.
Gao, F;Li, C;Danopoulos, S;Al Alam, D;Peinado, N;Webster, S;Borok, Z;Kohbodi, GA;Bellusci, S;Minoo, P;
PMID: 35385750 | DOI: 10.1016/j.celrep.2022.110608
The lung alveolus is lined with alveolar type 1 (AT1) and type 2 (AT2) epithelial cells. During alveologenesis, increasing demand associated with expanding alveolar numbers is met by proliferating progenitor AT2s (pAT2). Little information exists regarding the identity of this population and their niche microenvironment. We show that during alveologenesis, Hedgehog-responsive PDGFRa(+) progenitors (also known as SCMFs) are a source of secreted trophic molecules that maintain a unique pAT2 population. SCMFs are in turn maintained by TGFβ signaling. Compound inactivation of Alk5 TβR2 in SCMFs reduced their numbers and depleted the pAT2 pool without impacting differentiation of daughter cells. In lungs of preterm infants who died with bronchopulmonary dysplasia, PDGFRa is reduced and the number of proliferative AT2s is diminished, indicating that an evolutionarily conserved mechanism governs pAT2 behavior during alveologenesis. SCMFs are a transient cell population, active only during alveologenesis, making them a unique stage-specific niche mesodermal cell type in mammalian organs.
Coquenlorge S, Yin WC, Yung T, Pan J, Zhang X, Mo R, Belik J, Hui CC, Kim TH.
PMID: 31167144 | DOI: 10.1016/j.celrep.2019.05.016
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
McCarthy, N;Tie, G;Madha, S;He, R;Kraiczy, J;Maglieri, A;Shivdasani, RA;
PMID: 36924771 | DOI: 10.1016/j.devcel.2023.02.012
Wnt and Rspondin (RSPO) signaling drives proliferation, and bone morphogenetic protein inhibitors (BMPi) impede differentiation, of intestinal stem cells (ISCs). Here, we identify the mouse ISC niche as a complex, multi-layered structure that encompasses distinct mesenchymal and smooth muscle populations. In young and adult mice, diverse sub-cryptal cells provide redundant ISC-supportive factors; few of these are restricted to single cell types. Niche functions refine during postnatal crypt morphogenesis, in part to oppose the dense aggregation of differentiation-promoting BMP+ sub-epithelial myofibroblasts at crypt-villus junctions. Muscularis mucosae, a specialized muscle layer, first appears during this period and supplements neighboring RSPO and BMPi sources. Components of this developing niche are conserved in human fetuses. The in vivo ablation of mouse postnatal smooth muscle increases BMP signaling activity, potently limiting a pre-weaning burst of crypt fission. Thus, distinct and progressively specialized mesenchymal cells together create the milieu that is required to propagate crypts during rapid organ growth and to sustain adult ISCs.
