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GLP-1-, but not GDF-15-, receptor activation increases the number of IL-6-expressing cells in the external lateral parabrachial nucleus

Neuroendocrinology

2019 Mar 20

Anesten F, Mishra D, Dalmau Gasull A, Engstrom-Ruud L, Bellman J, Palsdottir V, Zhang FP, Trapp S, Skibicka KP, Poutanen M and Jansson JO
PMID: 30889580 | DOI: 10.1159/000499693

Background/Aims IL-6 in the hypothalamus and hindbrain is an important downstream mediator of suppression of body weight and food intake by glucagon-like peptide-1 (GLP-1) receptor stimulation. CNS GLP-1 is produced almost exclusively in prepro-glucagon neurons in the nucleus of the solitary tract. These neurons innervate energy balance-regulating areas, such as the external lateral parabrachial nucleus (PBNel); essential for induction of anorexia. Methods Using a validated novel IL-6-reporter mouse strain, we investigated the interactions in PBNel between GLP-1, IL-6 and calcitonin gene related peptide (CGRP, a well-known mediator of anorexia). We show that PBNel GLP-1R-containing cells highly (to about 80%) overlap with IL-6-containing cells on both protein and mRNA level. Results Intraperitoneal administration of a GLP-1 analogue exendin-4 to mice increased the proportion of IL-6 containing cells in PBNel 3-fold, while there was no effect in the rest of the lateral PBN. In contrast, injections of an anorexigenic peptide growth and differentiation factor 15 (GDF15) markedly increased the proportion of CGRP-containing cells, while IL-6-containing cells were not affected. Conclusion In summary, GLP-1R are found on IL-6 producing cells in PBNel, and GLP-1R stimulation leads to an increase in the proportion of cells with IL-6 reporter fluorescence, supporting IL-6 mediation of GLP-1 effects on energy balance.
Exploring the immunogenic properties of SARS-CoV-2 structural proteins: PAMP:TLR signaling in the mediation of the neuroinflammatory and neurologic sequelae of COVID-19

Brain, behavior, and immunity

2023 Apr 26

Frank, MG;Fleshner, M;Maier, SF;
PMID: 37116592 | DOI: 10.1016/j.bbi.2023.04.009

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) produces an array of neurologic and neuropsychiatric symptoms in the acute and post-acute phase of infection (PASC; post-acute sequelae of SARS-CoV-2 infection). Neuroinflammatory processes are considered key factors in the etiology of these symptoms. Several mechanisms underpinning the development of inflammatory events in the brain have been proposed including SARS-CoV-2 neurotropism and peripheral inflammatory responses (i.e., cytokine storm) to infection, which might produce neuroinflammation via immune-to-brain signaling pathways. In this review, we explore evidence in support of an alternate mechanism whereby structural proteins (e.g., spike and spike S1 subunit) derived from SARS-CoV-2 virions function as pathogen-associated molecular patterns (PAMPs) to elicit proinflammatory immune responses in the periphery and/or brain via classical Toll-Like Receptor (TLR) inflammatory pathways. We propose that SARS-CoV-2 structural proteins might directly produce inflammatory processes in brain independent of and/or in addition to peripheral proinflammatory effects, which might converge to play a causal role in the development of neurologic/neuropsychiatric symptoms in COVID-19.
Technical note on the exploration of COVID-19 in autopsy material

Journal of clinical pathology

2023 Jan 30

Humphries, MP;Bingham, V;Abdullah Sidi, F;Craig, S;Lara, B;El-Daly, H;O'Doherty, N;Maxwell, P;Lewis, C;McQuaid, S;Lyness, J;James, J;Snead, DRJ;Salto-Tellez, M;
PMID: 36717223 | DOI: 10.1136/jcp-2022-208525

Interrogation of immune response in autopsy material from patients with SARS-CoV-2 is potentially significant. We aim to describe a validated protocol for the exploration of the molecular physiopathology of SARS-CoV-2 pulmonary disease using multiplex immunofluorescence (mIF).The application of validated assays for the detection of SARS-CoV-2 in tissues, originally developed in our laboratory in the context of oncology, was used to map the topography and complexity of the adaptive immune response at protein and mRNA levels.SARS-CoV-2 is detectable in situ by protein or mRNA, with a sensitivity that could be in part related to disease stage. In formalin-fixed, paraffin-embedded pneumonia material, multiplex immunofluorescent panels are robust, reliable and quantifiable and can detect topographic variations in inflammation related to pathological processes.Clinical autopsies have relevance in understanding diseases of unknown/complex pathophysiology. In particular, autopsy materials are suitable for the detection of SARS-CoV-2 and for the topographic description of the complex tissue-based immune response using mIF.
IL-6 expression helps distinguish Castleman\'s disease from IgG4-related disease in the lung

BMC pulmonary medicine

2021 Jul 10

Kinugawa, Y;Uehara, T;Iwaya, M;Asaka, S;Kobayashi, S;Nakajima, T;Komatsu, M;Yasuo, M;Yamamoto, H;Ota, H;
PMID: 34246246 | DOI: 10.1186/s12890-021-01603-6

It is difficult to distinguish between multicentric Castleman's disease (MCD) and IgG4-related lung disease (IgG4-LD), an IgG4-related disease (IgG4-RD) in the lung.We focused on IL-6, which is elevated in MCD, to distinguish between MCD and IgG4-LD by RNAscope, a highly sensitive RNA in situ method. Six cases of MCD and four cases of IgG4-LD were selected.In all cases of MCD and IgG4-LD, 10 or more IgG4-positive cells were found in one high-power field. All MCD cases were inconsistent with the pathological IgG4-related comprehensive diagnostic criteria, but 2 of 6 cases had an IgG4/IgG ratio greater than 40%. In all IgG4-LD cases, histological features were consistent with the pathological IgG4-RD comprehensive diagnostic criteria. IL-6 expression was observed in all MCD and IgG4-LD cases except for one IgG4-LD biopsy. IL-6-expressing cells were mainly identified in the stroma. Sites of IL-6 expression were not characteristic and were sparse. IL-6 expression tended to be higher in MCD compared with IgG4-LD. A positive correlation was found between the IL-6 H-score and serum IL-6 level.Differences in IL-6 expression may help distinguish between MCD and IgG4-LD. In addition, the presence of high IL-6 levels may help elucidate the pathological mechanisms of IgG4-LD.
Fatal COVID-19 and Non-COVID-19 Acute Respiratory Distress Syndrome is associated with Incomplete Alveolar Type 1 Epithelial Cell Differentiation from the Transitional State Without Fibrosis

The American journal of pathology

2021 Dec 30

Ting, C;Aspal, M;Vaishampayan, N;Huang, SK;Riemondy, KA;Wang, F;Farver, C;Zemans, RL;
PMID: 34973949 | DOI: 10.1016/j.ajpath.2021.11.014

ARDS due to COVID-19 and other etiologies results from injury to the alveolar epithelial cell (AEC) barrier resulting in noncardiogenic pulmonary edema, which causes acute respiratory failure; clinical recovery requires epithelial regeneration. During physiologic regeneration in mice, AEC2s proliferate, exit the cell cycle, and transiently assume a transitional state before differentiating into AEC1s; persistence of the transitional state is associated with pulmonary fibrosis in humans. It is unknown whether transitional cells emerge and differentiate into AEC1s without fibrosis in human ARDS and why transitional cells differentiate into AEC1s during physiologic regeneration but persist in fibrosis. We hypothesized that incomplete but ongoing AEC1 differentiation from transitional cells without fibrosis may underlie persistent barrier permeability and fatal acute respiratory failure in ARDS. Immunostaining of postmortem ARDS lungs revealed abundant transitional cells in organized monolayers on alveolar septa without fibrosis. They were typically cuboidal or partially spread, sometimes flat, and occasionally expressed AEC1 markers. Immunostaining and/or interrogation of scRNAseq datasets revealed that transitional cells in mouse models of physiologic regeneration, ARDS, and fibrosis express markers of cell cycle exit but only in fibrosis express a specific senescence marker. Thus, in severe, fatal early ARDS, AEC1 differentiation from transitional cells is incomplete, underlying persistent barrier permeability and respiratory failure, but ongoing without fibrosis; senescence of transitional cells may be associated with pulmonary fibrosis.
Idiopathic multicentric Castleman’s disease: a clinicopathologic study in comparison with IgG4-related disease

Oncotarget.

2018 Jan 09

Otani K, Inoue D, Fujikura K, Komori T, Abe-Suzuki S, Tajiri T, Itoh T, Zen Y.
PMID: - | DOI: 10.18632/oncotarget.24068

The present study aimed to compare clinicopathologic features between idiopathic multicentric Castleman’s disease (n=22) and IgG4-related disease (n=26). Histology was analyzed using lymph node and lung biopsies. The expression of IL-6 mRNA in tissue was also examined by in situ hybridization and real-time PCR. Patients with idiopathic multicentric Castleman’s disease were significantly younger than those with IgG4-related disease (p<0.001). Splenomegaly was observed in only idiopathic multicentric Castleman’s disease (p=0.002), while pancreatitis and sialo-dacryoadenitis were restricted to IgG4-related disease (both p<0.001). Serum IgG4 concentrations were commonly elevated at >135 mg/dL in both groups (p=0.270). However, the IgG4/IgG ratio in IgG4-related disease was significantly higher than that in Castleman’s disease (p<0.001). Histologically, sheet-like plasmacytosis was highly characteristic of idiopathic multicentric Castleman’s disease (p<0.001), while plasmacytic infiltration in IgG4-related disease was always associated with intervening lymphocytes. Similar to laboratory findings, the IgG4/IgG-positive plasma cell ratio, but not the IgG4-positive cell count, was significantly higher in IgG4-related disease (p=0.002). Amyloid-like hyalinized fibrosis was found in 6/8 lung biopsies (75%) of Castleman’s disease. The over-expression of IL-6 mRNA was not confirmed in tissue samples of Castleman’s disease by either in situhybridization or quantitative real-time PCR. In conclusion, useful data for a differential diagnosis appear to be age, affected organs, the serum IgG4/IgG ratio, sheet-like plasmacytosis in biopsies, and the IgG4/IgG-positive cell ratio on immunostaining. Since IL-6 was not over-expressed in tissue of idiopathic multicentric Castleman’s disease, IL-6 may be produced outside the affected organs, and circulating IL-6 may lead to lymphoplasmacytosis at nodal and extranodal sites.

InfectionCMA: A Cell MicroArray Approach for Efficient Biomarker Screening in In Vitro Infection Assays

Pathogens (Basel, Switzerland)

2022 Mar 03

Magalhães, AC;Ricardo, S;Moreira, AC;Nunes, M;Tavares, M;Pinto, RJ;Gomes, MS;Pereira, L;
PMID: 35335638 | DOI: 10.3390/pathogens11030313

The recently emerged severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has forced the scientific community to acquire knowledge in real-time, when total lockdowns and the interruption of flights severely limited access to reagents as the global pandemic became established. This unique reality made researchers aware of the importance of designing efficient in vitro set-ups to evaluate infectious kinetics. Here, we propose a histology-based method to evaluate infection kinetics grounded in cell microarray (CMA) construction, immunocytochemistry and in situ hybridization techniques. We demonstrate that the chip-like organization of the InfectionCMA has several advantages, allowing side-by-side comparisons between diverse cell lines, infection time points, and biomarker expression and cytolocalization evaluation in the same slide. In addition, this methodology has the potential to be easily adapted for drug screening.
Positive Retrospective SARS-CoV-2 Testing in a Case of Acute Respiratory Distress Syndrome of Unknown Etiology

Case reports in pulmonology

2021 Aug 28

Burkett, A;McElwee, S;Margaroli, C;Bajpai, P;Elkholy, A;Manne, U;Wille, K;Benson, P;
PMID: 34513107 | DOI: 10.1155/2021/5484239

In order to elucidate the cause of acute respiratory distress syndrome of unknown etiology in a pre-pandemic patient, molecular techniques were used for detection of SARS-CoV-2. We used a SARS-CoV-2 nucleocapsid protein immunofluorescence stain to retrospectively identify an individual with diffuse alveolar damage on autopsy histology who had negative respiratory virus panel results in February, 2020, in Birmingham, Alabama. In situ hybridization for SARS-CoV-2 RNA revealed evidence of widespread multiorgan SARS-CoV-2 infection. This death antecedes the first reported death of a State of Alabama resident diagnosed with SARS-CoV-2 by 26 days.
SARS-CoV-2 airway infection results in the development of somatosensory abnormalities in a hamster model

Science signaling

2023 May 09

Serafini, RA;Frere, JJ;Zimering, J;Giosan, IM;Pryce, KD;Golynker, I;Panis, M;Ruiz, A;tenOever, BR;Zachariou, V;
PMID: 37159520 | DOI: 10.1126/scisignal.ade4984

Although largely confined to the airways, SARS-CoV-2 infection has been associated with sensory abnormalities that manifest in both acute and chronic phenotypes. To gain insight on the molecular basis of these sensory abnormalities, we used the golden hamster model to characterize and compare the effects of infection with SARS-CoV-2 and influenza A virus (IAV) on the sensory nervous system. We detected SARS-CoV-2 transcripts but no infectious material in the cervical and thoracic spinal cord and dorsal root ganglia (DRGs) within the first 24 hours of intranasal virus infection. SARS-CoV-2-infected hamsters exhibited mechanical hypersensitivity that was milder but prolonged compared with that observed in IAV-infected hamsters. RNA sequencing analysis of thoracic DRGs 1 to 4 days after infection suggested perturbations in predominantly neuronal signaling in SARS-CoV-2-infected animals as opposed to type I interferon signaling in IAV-infected animals. Later, 31 days after infection, a neuropathic transcriptome emerged in thoracic DRGs from SARS-CoV-2-infected animals, which coincided with SARS-CoV-2-specific mechanical hypersensitivity. These data revealed potential targets for pain management, including the RNA binding protein ILF3, which was validated in murine pain models. This work elucidates transcriptomic signatures in the DRGs triggered by SARS-CoV-2 that may underlie both short- and long-term sensory abnormalities.
Primate neural retina upregulates IL-6 and IL-10 in response to a herpes simplex vector suggesting the presence of a pro-/anti-inflammatory axis.

Exp Eye Res.

2016 May 08

Sauter MM, Brandt CR.
PMID: 27170050 | DOI: 10.1016/j.exer.2016.05.003

Injection of herpes simplex virus vectors into the vitreous of primate eyes induces an acute, transient uveitis. The purpose of this study was to characterize innate immune responses of macaque neural retina tissue to the herpes simplex virus type 1-based gene delivery vector hrR3. PCR array analysis demonstrated the induction of the pro-inflammatory cytokine IL-6, as well as the anti-inflammatory cytokine IL-10, following hrR3 exposure. Secretion of IL-6 was detected by ELISA and cone photoreceptors and Muller cells were the predominant IL-6 positive cell types. RNA in situ hybridization confirmed that IL-6 was expressed in photoreceptor and Muller cells. The IL-10 positive cells in the inner nuclear layer were identified as amacrine cells by immunofluorescence staining with calretinin antibody. hrR3 challenge resulted in activation of NFκB (p65) in Muller glial cells, but not in cone photoreceptors, suggesting a novel regulatory mechanism for IL-6 expression in cone cells. hrR3 replication was not required for IL-6 induction or NFκB (p65) activation. These data suggest a pro-inflammatory (IL-6)/anti-inflammatory (IL-10) axis exists in neural retina and the severity of acute posterior uveitis may be determined by this interaction. Further studies are needed to identify the trigger for IL-6 and IL-10 induction and the mechanism of IL-6 induction in cone cells.

Activation of Immune System May Cause Pathophysiological Changes in the Myocardium of SARS-CoV-2 Infected Monkey Model

Cells

2022 Feb 10

Rabbani, MY;Rappaport, J;Gupta, MK;
PMID: 35203260 | DOI: 10.3390/cells11040611

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is an extremely contagious disease whereby the virus damages the host's respiratory tract via entering through the ACE2 receptor. Cardiovascular disorder is being recognized in the majority of COVID-19 patients; yet, the relationship between SARS-CoV-2 and heart failure has not been established. In the present study, SARS-CoV-2 infection was induced in the monkey model. Thereafter, heart tissue samples were collected, and pathological changes were analyzed in the left ventricular tissue by hematoxylin and eosin, trichrome, and immunohistochemical staining specific to T lymphocytes and macrophages. The findings revealed that SARS-CoV-2 infection induces several pathological changes in the heart, which cause cardiomyocyte disarray, mononuclear infiltrates of inflammatory cells, and hypertrophy. Furthermore, collagen-specific staining showed the development of cardiac fibrosis in the interstitial and perivascular regions in the hearts of infected primates. Moreover, the myocardial tissue samples displayed multiple foci of inflammatory cells positive for T lymphocytes and macrophages within the myocardium. These findings suggest the progression of the disease, which can lead to the development of severe complications, including heart failure. Additionally, SARS-CoV-2 antigen staining detected the presence of virus particles in the myocardium. Thus, we found that SARS-CoV-2 infection is characterized by an exaggerated inflammatory immune response in the heart, which possibly contributes to myocardial remodeling and subsequent fibrosis.
Reduced Pathogenicity of the SARS-CoV-2 Omicron Variant in Hamsters

Med (New York, N.Y.)

2022 Mar 17

McMahan, K;Giffin, V;Tostanoski, LH;Chung, B;Siamatu, M;Suthar, MS;Halfmann, P;Kawaoka, Y;Piedra-Mora, C;Jain, N;Ducat, S;Kar, S;Andersen, H;Lewis, MG;Martinot, AJ;Barouch, DH;
PMID: 35313451 | DOI: 10.1016/j.medj.2022.03.004

The SARS-CoV-2 Omicron (B.1.1.529) variant has proven highly transmissible and has outcompeted the Delta variant in many regions of the world. Early reports have also suggested that Omicron may result in less severe clinical disease in humans. Here we show that Omicron is less pathogenic than prior SARS-CoV-2 variants in Syrian golden hamsters.Hamsters were inoculated with either SARS-CoV-2 Omicron or other SARS-CoV-2 variants. Animals were followed for weight loss, and upper and lower respiratory tract tissues were assessed for viral loads and histopathology.Infection of hamsters with the SARS-CoV-2 WA1/2020, Alpha, Beta, or Delta strains led to 4-10% weight loss by day 4 and 10-17% weight loss by day 6. In contrast, infection of hamsters with two different Omicron challenge stocks did not result in any detectable weight loss, even at high challenge doses. Omicron infection led to substantial viral replication in both the upper and lower respiratory tracts but demonstrated lower viral loads in lung parenchyma and reduced pulmonary pathology compared with WA1/2020 infection.These data suggest that the SARS-CoV-2 Omicron variant may result in robust upper respiratory tract infection but less severe lower respiratory tract clinical disease compared with prior SARS-CoV-2 variants.Funding for this study was provided by NIH grant CA260476, the Massachusetts Consortium for Pathogen Readiness, the Ragon Institute, and the Musk Foundation.

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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
EnEmProbe targets exons n and m
En-EmProbe 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

Enabling research, drug development (CDx) and diagnostics

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