Single cell transcriptomic analysis of murine lung development on hyperoxia-induced damage

Nature communications

2021 Mar 10

Hurskainen, M;Mižíková, I;Cook, DP;Andersson, N;Cyr-Depauw, C;Lesage, F;Helle, E;Renesme, L;Jankov, RP;Heikinheimo, M;Vanderhyden, BC;Thébaud, B;
PMID: 33692365 | DOI: 10.1038/s41467-021-21865-2

During late lung development, alveolar and microvascular development is finalized to enable sufficient gas exchange. Impaired late lung development manifests as bronchopulmonary dysplasia (BPD) in preterm infants. Single-cell RNA sequencing (scRNA-seq) allows for assessment of complex cellular dynamics during biological processes, such as development. Here, we use MULTI-seq to generate scRNA-seq profiles of over 66,000 cells from 36 mice during normal or impaired lung development secondary to hyperoxia with validation of some of the findings in lungs from BPD patients. We observe dynamic populations of cells, including several rare cell types and putative progenitors. Hyperoxia exposure, which mimics the BPD phenotype, alters the composition of all cellular compartments, particularly alveolar epithelium, stromal fibroblasts, capillary endothelium and macrophage populations. Pathway analysis and predicted dynamic cellular crosstalk suggest inflammatory signaling as the main driver of hyperoxia-induced changes. Our data provides a single-cell view of cellular changes associated with late lung development in health and disease.

Past, Present and Future of Cocaine- and Amphetamine-Regulated Transcript Peptide

Physiology & behavior

2021 Mar 08

Yosten, GLC;Haddock, CJ;Harada, CM;Almeida-Pereira, G;Kolar, GR;Stein, LM;Hayes, MR;Salvemini, D;Samson, WK;
PMID: 33705816 | DOI: 10.1016/j.physbeh.2021.113380

The existence of the peptide encoded by the cocaine- and amphetamine-regulated transcript (Cartpt) has been recognized since 1981, but it was not until 1995, that the gene encoding CART peptide (CART) was identified. With the availability of the predicted protein sequence of CART investigators were able to identify sites of peptide localization, which then led to numerous approaches attempting to clarify CART's multiple pharmacologic effects and even provide evidence of potential physiologic relevance. Although not without controversy, a picture emerged of the importance of CART in ingestive behaviors, reward behaviors and even pain sensation. Despite the wealth of data hinting at the significance of CART, in the absence of an identified receptor, the full potential for this peptide or its analogs to be developed into therapeutic agents remained unrealized. There was evidence favoring the action of CART via a G protein-coupled receptor (GPCR), but despite multiple attempts the identity of that receptor eluded investigators until recently. Now with the identification of the previously orphaned GPCR, GPR160, as a receptor for CART, focus on this pluripotent neuropeptide will in all likelihood experience a renaissance and the potential for the development of pharmcotherapies targeting GPR160 seems within reach.

Molecular correlates of muscle spindle and Golgi tendon organ afferents

Nature communications

2021 Mar 01

Oliver, KM;Florez-Paz, DM;Badea, TC;Mentis, GZ;Menon, V;de Nooij, JC;
PMID: 33649316 | DOI: 10.1038/s41467-021-21880-3

Proprioceptive feedback mainly derives from groups Ia and II muscle spindle (MS) afferents and group Ib Golgi tendon organ (GTO) afferents, but the molecular correlates of these three afferent subtypes remain unknown. We performed single cell RNA sequencing of genetically identified adult proprioceptors and uncovered five molecularly distinct neuronal clusters. Validation of cluster-specific transcripts in dorsal root ganglia and skeletal muscle demonstrates that two of these clusters correspond to group Ia MS afferents and group Ib GTO afferent proprioceptors, respectively, and suggest that the remaining clusters could represent group II MS afferents. Lineage analysis between proprioceptor transcriptomes at different developmental stages provides evidence that proprioceptor subtype identities emerge late in development. Together, our data provide comprehensive molecular signatures for groups Ia and II MS afferents and group Ib GTO afferents, enabling genetic interrogation of the role of individual proprioceptor subtypes in regulating motor output.

Localized EMT reprograms glial progenitors to promote spinal cord repair

Developmental cell

2021 Feb 16

Klatt Shaw, D;Saraswathy, VM;Zhou, L;McAdow, AR;Burris, B;Butka, E;Morris, SA;Dietmann, S;Mokalled, MH;
PMID: 33609461 | DOI: 10.1016/j.devcel.2021.01.017

Anti-regenerative scarring obstructs spinal cord repair in mammals and presents a major hurdle for regenerative medicine. In contrast, adult zebrafish possess specialized glial cells that spontaneously repair spinal cord injuries by forming a pro-regenerative bridge across the severed tissue. To identify the mechanisms that regulate differential regenerative capacity between mammals and zebrafish, we first defined the molecular identity of zebrafish bridging glia and then performed cross-species comparisons with mammalian glia. Our transcriptomics show that pro-regenerative zebrafish glia activate an epithelial-to-mesenchymal transition (EMT) gene program and that EMT gene expression is a major factor distinguishing mammalian and zebrafish glia. Functionally, we found that localized niches of glial progenitors undergo EMT after spinal cord injury in zebrafish and, using large-scale CRISPR-Cas9 mutagenesis, we identified the gene regulatory network that activates EMT and drives functional regeneration. Thus, non-regenerative mammalian glia lack an essential EMT-driving gene regulatory network that reprograms pro-regenerative zebrafish glia after injury.

Evolving Up‐regulation of Biliary Fibrosis–Related Extracellular Matrix Molecules After Successful Portoenterostomy

Hepatology Communications

2021 Feb 09

Kyrönlahti, A;Godbole, N;Akinrinade, O;Soini, T;Nyholm, I;Andersson, N;Hukkinen, M;Lohi, J;Wilson, D;Pihlajoki, M;Pakarinen, M;Heikinheimo, M;
| DOI: 10.1002/hep4.1684

Successful portoenterostomy (SPE) improves the short‐term outcome of patients with biliary atresia (BA) by relieving cholestasis and extending survival with native liver. Despite SPE, hepatic fibrosis progresses in most patients, leading to cirrhosis and a deterioration of liver function. The goal of this study was to characterize the effects of SPE on the BA liver transcriptome. We used messenger RNA sequencing to analyze global gene‐expression patterns in liver biopsies obtained at the time of portoenterostomy (n = 13) and 1 year after SPE (n = 8). Biopsies from pediatric (n = 2) and adult (n = 2) organ donors and other neonatal cholestatic conditions (n = 5) served as controls. SPE was accompanied by attenuation of inflammation and concomitant up‐regulation of key extracellular matrix (ECM) genes. Highly overexpressed genes promoting biliary fibrosis and bile duct integrity, such as integrin subunit beta 6 and previously unreported laminin subunit alpha 3, emerged as candidates to control liver fibrosis after SPE. At a cellular level, the relative abundance of activated hepatic stellate cells and liver macrophages decreased following SPE, whereas portal fibroblasts (PFs) and cholangiocytes persisted. Conclusion: The attenuation of inflammation following SPE coincides with emergence of an ECM molecular fingerprint, a set of profibrotic molecules mechanistically connected to biliary fibrosis. The persistence of activated PFs and cholangiocytes after SPE suggests a central role for these cell types in the progression of biliary fibrosis.

LPA signaling acts as a cell-extrinsic mechanism to initiate cilia disassembly and promote neurogenesis

Nature communications

2021 Jan 28

Hu, HB;Song, ZQ;Song, GP;Li, S;Tu, HQ;Wu, M;Zhang, YC;Yuan, JF;Li, TT;Li, PY;Xu, YL;Shen, XL;Han, QY;Li, AL;Zhou, T;Chun, J;Zhang, XM;Li, HY;
PMID: 33510165 | DOI: 10.1038/s41467-021-20986-y

Dynamic assembly and disassembly of primary cilia controls embryonic development and tissue homeostasis. Dysregulation of ciliogenesis causes human developmental diseases termed ciliopathies. Cell-intrinsic regulatory mechanisms of cilia disassembly have been well-studied. The extracellular cues controlling cilia disassembly remain elusive, however. Here, we show that lysophosphatidic acid (LPA), a multifunctional bioactive phospholipid, acts as a physiological extracellular factor to initiate cilia disassembly and promote neurogenesis. Through systematic analysis of serum components, we identify a small molecular-LPA as the major driver of cilia disassembly. Genetic inactivation and pharmacological inhibition of LPA receptor 1 (LPAR1) abrogate cilia disassembly triggered by serum. The LPA-LPAR-G-protein pathway promotes the transcription and phosphorylation of cilia disassembly factors-Aurora A, through activating the transcription coactivators YAP/TAZ and calcium/CaM pathway, respectively. Deletion of Lpar1 in mice causes abnormally elongated cilia and decreased proliferation in neural progenitor cells, thereby resulting in defective neurogenesis. Collectively, our findings establish LPA as a physiological initiator of cilia disassembly and suggest targeting the metabolism of LPA and the LPA pathway as potential therapies for diseases with dysfunctional ciliogenesis.

Pathology of COVID 19 associated acute kidney injury

Clinical Kidney Journal

2021 Jan 24

Sharma, P;Ng, J;Bijol, V;Jhaveri, K;Wanchoo, R;
| DOI: 10.1093/ckj/sfab003

Acute kidney injury (AKI) is common among hospitalized patients with Coronavirus Infectious Disease 2019 (COVID-19), with the occurrence of AKI ranging from 0.5% to 80%. An improved knowledge of the pathology of AKI in COVID-19 is crucial to mitigate and manage AKI and to improve the survival of patients who develop AKI during COVID-19. In this review, we summarize the published cases and case series of various kidney pathology seen with COVID-19. Both live kidney biopsies and autopsy series suggest acute tubular injury as the most commonly encountered pathology. Collapsing glomerulopathy and thrombotic microangiopathy are other encountered pathologies noted in both live and autopsy tissues. Other rare findings such as ANCA vasculitis, Anti GBM disease, and podocytopathies have been reported. Although direct viral infection of the kidney is possible, it is certainly not a common or even widespread finding reported at the time of this writing (November 2020).

A subset of spinal dorsal horn interneurons crucial for gating touch-evoked pain-like behavior

Proceedings of the National Academy of Sciences of the United States of America

2021 Jan 19

Tashima, R;Koga, K;Yoshikawa, Y;Sekine, M;Watanabe, M;Tozaki-Saitoh, H;Furue, H;Yasaka, T;Tsuda, M;
PMID: 33431693 | DOI: 10.1073/pnas.2021220118

A cardinal, intractable symptom of neuropathic pain is mechanical allodynia, pain caused by innocuous stimuli via low-threshold mechanoreceptors such as Aβ fibers. However, the mechanism by which Aβ fiber-derived signals are converted to pain remains incompletely understood. Here we identify a subset of inhibitory interneurons in the spinal dorsal horn (SDH) operated by adeno-associated viral vectors incorporating a neuropeptide Y promoter (AAV-NpyP+) and show that specific ablation or silencing of AAV-NpyP+ SDH interneurons converted touch-sensing Aβ fiber-derived signals to morphine-resistant pain-like behavioral responses. AAV-NpyP+ neurons received excitatory inputs from Aβ fibers and transmitted inhibitory GABA signals to lamina I neurons projecting to the brain. In a model of neuropathic pain developed by peripheral nerve injury, AAV-NpyP+ neurons exhibited deeper resting membrane potentials, and their excitation by Aβ fibers was impaired. Conversely, chemogenetic activation of AAV-NpyP+ neurons in nerve-injured rats reversed Aβ fiber-derived neuropathic pain-like behavior that was shown to be morphine-resistant and reduced pathological neuronal activation of superficial SDH including lamina I. These findings suggest that identified inhibitory SDH interneurons that act as a critical brake on conversion of touch-sensing Aβ fiber signals into pain-like behavioral responses. Thus, enhancing activity of these neurons may offer a novel strategy for treating neuropathic allodynia.

Functional characterization of the dural sinuses as a neuroimmune interface

Cell

2021 Jan 18

Rustenhoven, J;Drieu, A;Mamuladze, T;de Lima, KA;Dykstra, T;Wall, M;Papadopoulos, Z;Kanamori, M;Salvador, AF;Baker, W;Lemieux, M;Da Mesquita, S;Cugurra, A;Fitzpatrick, J;Sviben, S;Kossina, R;Bayguinov, P;Townsend, RR;Zhang, Q;Erdmann-Gilmore, P;Smirnov, I;Lopes, MB;Herz, J;Kipnis, J;
PMID: 33508229 | DOI: 10.1016/j.cell.2020.12.040

Despite the established dogma of central nervous system (CNS) immune privilege, neuroimmune interactions play an active role in diverse neurological disorders. However, the precise mechanisms underlying CNS immune surveillance remain elusive; particularly, the anatomical sites where peripheral adaptive immunity can sample CNS-derived antigens and the cellular and molecular mediators orchestrating this surveillance. Here, we demonstrate that CNS-derived antigens in the cerebrospinal fluid (CSF) accumulate around the dural sinuses, are captured by local antigen-presenting cells, and are presented to patrolling T cells. This surveillance is enabled by endothelial and mural cells forming the sinus stromal niche. T cell recognition of CSF-derived antigens at this site promoted tissue resident phenotypes and effector functions within the dural meninges. These findings highlight the critical role of dural sinuses as a neuroimmune interface, where brain antigens are surveyed under steady-state conditions, and shed light on age-related dysfunction and neuroinflammatory attack in animal models of multiple sclerosis.

The versican-hyaluronan complex provides an essential extracellular matrix niche for Flk1+ hematoendothelial progenitors

Matrix biology : journal of the International Society for Matrix Biology

2021 Jan 14

Nandadasa, S;O'Donnell, A;Murao, A;Yamaguchi, Y;Midura, RJ;Olson, L;Apte, SS;
PMID: 33454424 | DOI: 10.1016/j.matbio.2021.01.002

Little is known about extracellular matrix (ECM) contributions to formation of the earliest cell lineages in the embryo. Here, we show that the proteoglycan versican and glycosaminoglycan hyaluronan are associated with emerging Flk1+ hematoendothelial progenitors at gastrulation. The mouse versican mutant Vcanhdf lacks yolk sac vasculature, with attenuated yolk sac hematopoiesis. CRISPR/Cas9-mediated Vcan inactivation in mouse embryonic stem cells reduced vascular endothelial and hematopoietic differentiation within embryoid bodies, which generated fewer blood colonies, and had an impaired angiogenic response to VEGF165. Hyaluronan was severely depleted in Vcanhdf embryos, with corresponding upregulation of the hyaluronan-depolymerase TMEM2. Conversely, hyaluronan-deficient mouse embryos also had vasculogenic suppression but with increased versican proteolysis. VEGF165 and Indian hedgehog, crucial vasculogenic factors, utilized the versican-hyaluronan matrix, specifically versican chondroitin sulfate chains, for binding. Versican-hyaluronan ECM is thus an obligate requirement for vasculogenesis and primitive hematopoiesis, providing a vasculogenic factor-enriching microniche for Flk1+ progenitors from their origin at gastrulation.

Moderate Exercise Inhibits Age-Related Inflammation, Liver Steatosis, Senescence, and Tumorigenesis

Journal of immunology (Baltimore, Md. : 1950)

2021 Jan 13

Bianchi, A;Marchetti, L;Hall, Z;Lemos, H;Vacca, M;Paish, H;Green, K;Elliott, B;Tiniakos, D;Passos, JF;Jurk, D;Mann, DA;Wilson, CL;
PMID: 33441438 | DOI: 10.4049/jimmunol.2001022

Age-related chronic inflammation promotes cellular senescence, chronic disease, cancer, and reduced lifespan. In this study, we wanted to explore the effects of a moderate exercise regimen on inflammatory liver disease and tumorigenesis. We used an established model of spontaneous inflammaging, steatosis, and cancer (nfkb1-/- mouse) to demonstrate whether 3 mo of moderate aerobic exercise was sufficient to suppress liver disease and cancer development. Interventional exercise when applied at a relatively late disease stage was effective at reducing tissue inflammation (liver, lung, and stomach), oxidative damage, and cellular senescence, and it reversed hepatic steatosis and prevented tumor development. Underlying these benefits were transcriptional changes in enzymes driving the conversion of tryptophan to NAD+, this leading to increased hepatic NAD+ and elevated activity of the NAD+-dependent deacetylase sirtuin. Increased SIRT activity was correlated with enhanced deacetylation of key transcriptional regulators of inflammation and metabolism, NF-κB (p65), and PGC-1α. We propose that moderate exercise can effectively reprogram pre-established inflammatory and metabolic pathologies in aging with the benefit of prevention of disease.

Embryonic Microglia Interact with Hypothalamic Radial Glia during Development and Upregulate the TAM Receptors MERTK and AXL following an Insult

Cell reports

2021 Jan 05

Rosin, JM;Marsters, CM;Malik, F;Far, R;Adnani, L;Schuurmans, C;Pittman, QJ;Kurrasch, DM;
PMID: 33406432 | DOI: 10.1016/j.celrep.2020.108587

Despite a growing appreciation for microglial influences on the developing brain, the responsiveness of microglia to insults during gestation remains less well characterized, especially in the embryo when microglia themselves are still maturing. Here, we asked if fetal microglia could coordinate an innate immune response to an exogenous insult. Using time-lapse imaging, we showed that hypothalamic microglia actively surveyed their environment by near-constant "touching" of radial glia projections. However, following an insult (i.e., IUE or AAV transduction), this seemingly passive touching became more intimate and long lasting, ultimately resulting in the retraction of radial glial projections and degeneration into small pieces. Mechanistically, the TAM receptors MERTK and AXL were upregulated in microglia following the insult, and Annexin V treatment inhibited radial glia breakage and engulfment by microglia. These data demonstrate a remarkable responsiveness of embryonic microglia to insults during gestation, a critical window for neurodevelopment. Crown

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	Description
sense Example: Hs-LAG3-sense	Standard probes for RNA detection are in antisense. Sense probe is reverse complent to the corresponding antisense probe.
Intron# Example: Mm-Htt-intron2	Probe targets the indicated intron in the target gene, commonly used for pre-mRNA detection
Pool/Pan Example: Hs-CD3-pool (Hs-CD3D, Hs-CD3E, Hs-CD3G)	A mixture of multiple probe sets targeting multiple genes or transcripts
No-XSp Example: Hs-PDGFB-No-XMm	Does not cross detect with the species (Sp)
XSp Example: Rn-Pde9a-XMm	designed to cross detect with the species (Sp)
O# Example: Mm-Islr-O1	Alternative design targeting different regions of the same transcript or isoforms
CDS Example: Hs-SLC31A-CDS	Probe targets the protein-coding sequence only
EnEm	Probe targets exons n and m
En-Em	Probe targets region from exon n to exon m
Retired Nomenclature
tvn Example: Hs-LEPR-tv1	Designed to target transcript variant n
ORF Example: Hs-ACVRL1-ORF	Probe targets open reading frame
UTR Example: Hs-HTT-UTR-C3	Probe targets the untranslated region (non-protein-coding region) only
5UTR Example: Hs-GNRHR-5UTR	Probe targets the 5' untranslated region only
3UTR Example: Rn-Npy1r-3UTR	Probe targets the 3' untranslated region only
Pan Example: Pool	A mixture of multiple probe sets targeting multiple genes or transcripts

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