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      Distinct and common expression of receptors for inflammatory mediators in vagal nodose versus jugular capsaicin-sensitive/TRPV1-positive neurons detected by low input RNA sequencing

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          Abstract

          Capsaicin-sensitive sensory C-fibers derived from vagal ganglia innervate the visceral organs, and respond to inflammatory mediators and noxious stimuli. These neurons play an important role in maintenance of visceral homeostasis, and contribute to the symptoms of visceral inflammatory diseases. Vagal sensory neurons are located in two ganglia, the jugular ganglia (derived from the neural crest), and the nodose ganglia (from the epibranchial placodes). The functional difference, especially in response to immune mediators, between jugular and nodose neurons is not fully understood. In this study, we microscopically isolated murine nodose and jugular capsaicin-sensitive / Trpv1-expressing C-fiber neurons and performed transcriptome profiling using ultra-low input RNA sequencing. RNAseq detected genes with significantly differential expression in jugular and nodose neurons, which were mostly involved in neural functions. Transcriptional regulators, including Cited1, Hoxb5 and Prdm12 showed distinct expression patterns in the two C-fiber neuronal populations. Common and specific expression of immune receptor proteins was characterized in each neuronal type. The expression of immune receptors that have received little or no attention from vagal sensory biologists is highlighted including receptors for certain chemokines (CXCLs), interleukins (IL-4) and interferons (IFNα, IFNγ). Stimulation of immune receptors with their cognate ligands led to activation of the C-fibers in isolated functional assays.

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          Silencing Nociceptor Neurons Reduces Allergic Airway Inflammation.

          Lung nociceptors initiate cough and bronchoconstriction. To elucidate if these fibers also contribute to allergic airway inflammation, we stimulated lung nociceptors with capsaicin and observed increased neuropeptide release and immune cell infiltration. In contrast, ablating Nav1.8(+) sensory neurons or silencing them with QX-314, a charged sodium channel inhibitor that enters via large-pore ion channels to specifically block nociceptors, substantially reduced ovalbumin- or house-dust-mite-induced airway inflammation and bronchial hyperresponsiveness. We also discovered that IL-5, a cytokine produced by activated immune cells, acts directly on nociceptors to induce the release of vasoactive intestinal peptide (VIP). VIP then stimulates CD4(+) and resident innate lymphoid type 2 cells, creating an inflammatory signaling loop that promotes allergic inflammation. Our results indicate that nociceptors amplify pathological adaptive immune responses and that silencing these neurons with QX-314 interrupts this neuro-immune interplay, revealing a potential new therapeutic strategy for asthma.
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            Central terminal sensitization of TRPV1 by descending serotonergic facilitation modulates chronic pain.

            The peripheral terminals of primary nociceptive neurons play an essential role in pain detection mediated by membrane receptors like TRPV1, a molecular sensor of heat and capsaicin. However, the contribution of central terminal TRPV1 in the dorsal horn to chronic pain has not been investigated directly. Combining primary sensory neuron-specific GCaMP3 imaging with a trigeminal neuropathic pain model, we detected robust neuronal hyperactivity in injured and uninjured nerves in the skin, soma in trigeminal ganglion, and central terminals in the spinal trigeminal nucleus. Extensive TRPV1 hyperactivity was observed in central terminals innervating all dorsal horn laminae. The central terminal TRPV1 sensitization was maintained by descending serotonergic (5-HT) input from the brainstem. Central blockade of TRPV1 or 5-HT/5-HT3A receptors attenuated central terminal sensitization, excitatory primary afferent inputs, and mechanical hyperalgesia in the territories of injured and uninjured nerves. Our results reveal central mechanisms facilitating central terminal sensitization underlying chronic pain. Copyright © 2014 Elsevier Inc. All rights reserved.
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              Hoxb5 marks long-term haematopoietic stem cells revealing a homogenous perivascular niche

              The hematopoietic stem cell (HSC) is arguably the most extensively characterized tissue stem cell. Since its identification by prospective isolation 1 , complex multi-parameter flow cytometric isolation of phenotypic subsets has facilitated studies on many aspects of HSC biology including, self-renewal 2–4 , differentiation, aging, niche 5 , and diversity 6–8 . Here we demonstrate by unbiased multi-step screening, identification of a single gene, Hoxb5 (homeobox B5 also known as Hox-2.1), whose expression in the bone marrow (BM) is limited to the long-term HSC (LT-HSC) in mice. Utilizing a single-color tri-mCherry reporter mouse driven by endogenous Hoxb5 regulation, only the Hoxb5-positive HSCs exhibit long-term reconstitution capacity after transplantation in primary transplant recipients, and critically, in secondary recipients. Only 7–35% of various previously defined immunophenotypic HSCs are LT-HSCs. Finally, by in situ imaging of mouse BM, we show that >94% of LT-HSC (Hoxb5+) are directly attached to VE-cadherin-positive cells, implicating a perivascular space as a near homogenous localization of the LT-HSC.
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                Author and article information

                Contributors
                Role: Data curationRole: Formal analysisRole: MethodologyRole: SoftwareRole: ValidationRole: VisualizationRole: Writing – original draftRole: Writing – review & editing
                Role: Data curationRole: Formal analysisRole: Funding acquisitionRole: InvestigationRole: MethodologyRole: ValidationRole: VisualizationRole: Writing – original draftRole: Writing – review & editing
                Role: Data curationRole: InvestigationRole: Validation
                Role: Data curationRole: InvestigationRole: Validation
                Role: Data curationRole: InvestigationRole: Validation
                Role: Data curationRole: InvestigationRole: Validation
                Role: ConceptualizationRole: Project administration
                Role: Data curationRole: InvestigationRole: MethodologyRole: Validation
                Role: ConceptualizationRole: Project administration
                Role: ConceptualizationRole: Funding acquisitionRole: Project administrationRole: ResourcesRole: SupervisionRole: Writing – original draftRole: Writing – review & editing
                Role: ConceptualizationRole: Funding acquisitionRole: Project administrationRole: ResourcesRole: SupervisionRole: Writing – original draftRole: Writing – review & editing
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                5 October 2017
                2017
                : 12
                : 10
                : e0185985
                Affiliations
                [1 ] Department of Respiratory, Inflammatory, and Autoimmunity, MedImmune, LLC, Gaithersburg, Maryland, United States of America
                [2 ] Division of Allergy & Clinical Immunology, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, United States of America
                [3 ] Department of Pathophysiology, Biomedical Center Martin, Jessenius Faculty of Medicine, Comenius University in Bratislava, Martin, Slovakia
                [4 ] Department of Neuroscience, Johns Hopkins School of Medicine, Baltimore, Maryland, United States of America
                [5 ] Department of Translational Medicine, MedImmune, LLC, Gaithersburg, Maryland, United States of America
                University of California Los Angeles, UNITED STATES
                Author notes

                Competing Interests: JW, SK, PB and RK are full time employees and shareholders of MedImmune LLC. GS has been a full time employee and shareholder of MedImmune until recently. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

                Author information
                http://orcid.org/0000-0003-4662-8584
                Article
                PONE-D-17-17019
                10.1371/journal.pone.0185985
                5628920
                28982197
                3b8e6481-032d-4ad3-a5de-feab411deebe
                © 2017 Wang et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 3 May 2017
                : 23 September 2017
                Page count
                Figures: 6, Tables: 2, Pages: 20
                Funding
                Funded by: Johns Hopkins University-Medimmune CollaborativeAgreement
                Award ID: 90057138
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/100000002, National Institutes of Health;
                Award ID: HL137807
                Award Recipient :
                Funded by: funder-id http://dx.doi.org/10.13039/100000002, National Institutes of Health;
                Award ID: HL122228
                Award Recipient :
                Funded by: Department of Education, Slovakia
                Award ID: APVV-15-0163
                Award Recipient :
                Funded by: Department of Education, Slovakia
                Award ID: VEGA 1/0226/15
                Award Recipient :
                Funded by: MedImmune, LLC
                Award Recipient :
                Funded by: MedImmune, LLC
                Award Recipient :
                Funded by: MedImmune, LLC
                Award Recipient :
                Funded by: MedImmune, LLC
                Award Recipient :
                Funded by: MedImmune, LLC
                Award Recipient :
                The work was supported by the following: 1. BU received funding from Johns Hopkins University-Medimmune CollaborativeAgreement, # 90057138, ( http://releases.jhu.edu/2013/12/11/medimmune/); 2. BU received funding from NIH HL137807 and HL122228, ( www.nih.gov); 3. MK was partially supported by APVV-15-0163 and VEGA 1/0226/15 (Department of Education, Slovakia) ( http://www.apvv.sk/?lang=en, https://www.minedu.sk/vedecka-grantova-agentura-msvvas-sr-asav-vega/); 4. JW, SK, PB and RK are full time employees and shareholders of MedImmune, LLC ( https://www.medimmune.com/). GS has been a full time employee and shareholder of MedImmune until recently. His current affiliation is Cellcion, LLC ( https://cellcion.com/). Funding bodies did not have a role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The specific roles of all authors are articulated in the "author contributions" section.
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                Custom metadata
                Analysis results of RNA sequencing are within the paper and its Supporting Information files. All the raw and processed RNA sequencing data has been uploaded to Gene Expression Omnibus with accession number GSE102123 ( https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE102123).

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