Identifying Pathways and Networks Associated With the SARS-CoV-2 Cell Receptor ACE2 Based on Gene Expression Profiles in Normal and SARS-CoV-2-Infected Human Tissues

Abstract The angiotensin-converting enzyme 2 (ACE2) is a host cell receptor of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) that has infected more than six million people worldwide and has caused more than 370,000 deaths as of May 31, 2020. An investigation of ACE2 expression in human tissues may provide insights into the mechanism of SARS-CoV-2 infection. We identified pathways associated with ACE2 expression and gene co-expression networks of ACE2 in pan-tissue based on the gene expression profiles in human tissues. We found that the pathways significantly associated with ACE2 upregulation were mainly involved in immune, stromal signature, metabolism, cell growth and proliferation, and cancer and other diseases. The number of genes having a significant positive expression correlation with ACE2 in females far exceeded that in males. The estrogen receptors (ESR1 and ESR2) and androgen receptor (AR) genes had a significant positive expression correlation with ACE2 in pan-tissue. Meanwhile, the enrichment levels of immune cells were positively associated with the expression levels of ESR1 and ESR2, while they were inversely associated with the expression levels of AR in pan-tissue and in multiple individual tissues. It suggests that females are likely to have a more robust immune defense system against SARS-CoV-2 than males, partially explaining why females have better clinical outcomes of SARS-CoV-2 infections than males. Our data warrant further investigation for understanding the mechanism of SARS-CoV-2 infection.

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Single-cell immune repertoire and transcriptome sequencing reveals that clonally expanded and transcriptionally distinct lymphocytes populate the aged central nervous system in mice

Proceedings of The Royal Society B Biological Sciences ◽

10.1098/rspb.2020.2793 ◽

2021 ◽

Vol 288 (1945) ◽

pp. 20202793

Author(s):

Alexander Yermanos ◽

Daniel Neumeier ◽

Ioana Sandu ◽

Mariana Borsa ◽

Ann Cathrin Waindok ◽

...

Keyword(s):

Gene Expression ◽

Central Nervous System ◽

Nervous System ◽

B Cells ◽

Single Cell ◽

Somatic Hypermutation ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Cell Receptor ◽

The Central Nervous System

Neuroinflammation plays a crucial role during ageing and various neurological conditions, including Alzheimer's disease, multiple sclerosis and infection. Technical limitations, however, have prevented an integrative analysis of how lymphocyte immune receptor repertoires and their accompanying transcriptional states change with age in the central nervous system. Here, we leveraged single-cell sequencing to simultaneously profile B cell receptor and T cell receptor repertoires and accompanying gene expression profiles in young and old mouse brains. We observed the presence of clonally expanded B and T cells in the central nervous system of aged male mice. Furthermore, many of these B cells were of the IgM and IgD isotypes, and had low levels of somatic hypermutation. Integrating gene expression information additionally revealed distinct transcriptional profiles of these clonally expanded lymphocytes. Our findings implicate that clonally related T and B cells in the CNS of elderly mice may contribute to neuroinflammation accompanying homeostatic ageing.

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In Silico Discovery of Candidate Drugs against Covid-19

Viruses ◽

10.3390/v12040404 ◽

2020 ◽

Vol 12 (4) ◽

pp. 404 ◽

Cited By ~ 42

Author(s):

Claudia Cava ◽

Gloria Bertoli ◽

Isabella Castiglioni

Keyword(s):

Gene Expression ◽

In Silico ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Interaction Network ◽

Enrichment Analysis ◽

Basic Mechanism ◽

Cell Receptor ◽

The Cancer Genome Atlas ◽

Pathway Enrichment Analysis

Previous studies reported that Angiotensin converting enzyme 2 (ACE2) is the main cell receptor of SARS-CoV and SARS-CoV-2. It plays a key role in the access of the virus into the cell to produce the final infection. In the present study we investigated in silico the basic mechanism of ACE2 in the lung and provided evidences for new potentially effective drugs for Covid-19. Specifically, we used the gene expression profiles from public datasets including The Cancer Genome Atlas, Gene Expression Omnibus and Genotype-Tissue Expression, Gene Ontology and pathway enrichment analysis to investigate the main functions of ACE2-correlated genes. We constructed a protein-protein interaction network containing the genes co-expressed with ACE2. Finally, we focused on the genes in the network that are already associated with known drugs and evaluated their role for a potential treatment of Covid-19. Our results demonstrate that the genes correlated with ACE2 are mainly enriched in the sterol biosynthetic process, Aryldialkylphosphatase activity, adenosylhomocysteinase activity, trialkylsulfonium hydrolase activity, acetate-CoA and CoA ligase activity. We identified a network of 193 genes, 222 interactions and 36 potential drugs that could have a crucial role. Among possible interesting drugs for Covid-19 treatment, we found Nimesulide, Fluticasone Propionate, Thiabendazole, Photofrin, Didanosine and Flutamide.

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Molecular Determinants of Lymph Node Microenvironment Induced Host Immune Tolerance In CLL: Role for CAV1, PTPN6, and PKCβ In the Process

Blood ◽

10.1182/blood.v116.21.1367.1367 ◽

2010 ◽

Vol 116 (21) ◽

pp. 1367-1367

Author(s):

Christine Gilling ◽

Amit Mittal ◽

Vincent Nganga ◽

Vicky Palmer ◽

Dennis D. Weisenburger ◽

...

Keyword(s):

Gene Expression ◽

Mouse Model ◽

Immune Tolerance ◽

Conflicts Of Interest ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Cell Receptor ◽

Differentially Expressed ◽

Expression Levels ◽

Aggressive Disease

Abstract Abstract 1367 Previously, we have shown that gene expression profiles (GEP) of CLL cells from lymph nodes (LN), bone marrow (BM), and peripheral blood (PB) are significantly different from each other. Among the major pathways associated with differential gene expression, a “tolerogenic signature” involved in host immune tolerance is significant in regulating CLL progression. The genes associated with the tolerogenic signature are significantly differentially expressed in patient LN-CLL compared to BM-CLL and PB-CLL, suggesting that LN-CLL cells induce this immune tolerance. From 83 differentially expressed genes identified by GEP that are associated with immune dysregulation, we selected eleven genes (CAV1, PTPN6, PKCb, ZWINT, IL2Ra, CBLC, CDC42, ZNF175, ZNF264, IL10, and HLA-G) for validation studies to determine whether these genes are also dysregulated in the Emu-TCL1 mouse model of CLL. The results demonstrate a trend of upregulation of these genes as determined by qRT-PCR in the LN-tumor microenvironment. To further evaluate the kinetics of selected gene expression during tumor progression, we determined the expression levels of Cav1, Ptpn6, and Pkcb at 12, 24, and 36 weeks of CLL development in the Em-TCL1 mouse model. We found that the expression of all three genes increased as a function of age, indicating a correlation of gene expression with disease progression. In addition, as CLL progressed in these mice there was a marked decrease in CD4+ and CD8+ T cells. The murine data were further validated using CLL cells from the same patients with indolent versus aggressive disease indicating a similar trend in expression as CLL progressed (n=4). Furthermore, patient data analyzed by Kaplan Meier analyses of the expression levels of the selected genes indicated a significant association between down-regulation of PTPN6 (p=0.031) and up-regulation of ZWINT (p<0.001) with clinical outcome as determined by a shorter time to treatment (p<0.05). Functional analysis by knockdown of CAV1 and PKCb in primary patient CLL cells determined by MTT assay showed a decrease in proliferation following knockdown of these genes (p<0.005). Protein-interaction modeling revealed regulation of CAV1 and PTPN6 by one another. Additionally, the PTPN6 protein regulates B cell receptor (BCR) signaling and subsequently the BCR regulates PKCb. Therefore, these data from both mice and humans with CLL, argue that an aggressive disease phenotype is paralleled by expression of genes associated with immune suppression. In particular, evidence presented here suggests, dysregulation of CAV1, PTPN6, ZWINT, and PKCb expression promotes CLL progression. Disclosures: No relevant conflicts of interest to declare.

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Single-cell immune repertoire and transcriptome sequencing reveals that clonally expanded and transcriptionally distinct lymphocytes populate the aged central nervous system in mice

10.1101/2020.05.04.077081 ◽

2020 ◽

Author(s):

Alexander Yermanos ◽

Daniel Neumeier ◽

Ioana Sandu ◽

Mariana Borsa ◽

Ann Cathrin Waindok ◽

...

Keyword(s):

Gene Expression ◽

Central Nervous System ◽

Nervous System ◽

B Cells ◽

Single Cell ◽

Somatic Hypermutation ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Cell Receptor ◽

The Central Nervous System

AbstractNeuroinflammation plays a crucial role during ageing and various neurological conditions, including Alzheimer’s disease, multiple sclerosis and infection. Technical limitations, however, have prevented an integrative analysis of how lymphocyte immune receptor repertoires and their accompanying transcriptional states change with age in the central nervous system (CNS). Here, we leveraged single-cell sequencing to simultaneously profile B cell receptor (BCR) and T cell receptor (TCR) repertoires and accompanying gene expression profiles in young and old mouse brains. We observed the presence of clonally expanded B and T cells in the central nervous system (CNS) of aged mice. Furthermore, many of these B cells were of the IgM and IgD isotype and had low levels of somatic hypermutation. Integrating gene expression information additionally revealed distinct transcriptional profiles of these clonally expanded lymphocytes. Our findings implicate that clonally related T and B cells in the CNS of elderly mice may contribute to neuroinflammation accompanying homeostatic ageing.

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Gene Expression Profile of the Hypothalamus in DNP-KLH Immunized Mice Following Electroacupuncture Stimulation

Evidence-based Complementary and Alternative Medicine ◽

10.1093/ecam/nep222 ◽

2011 ◽

Vol 2011 ◽

pp. 1-8 ◽

Cited By ~ 2

Author(s):

Sun Kwang Kim ◽

Jeungshin Kim ◽

Eunjung Ko ◽

Hyunseong Kim ◽

Deok-Sang Hwang ◽

...

Keyword(s):

Gene Expression ◽

Immune System ◽

Microarray Analysis ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Variable Region ◽

Cell Receptor ◽

Control Group ◽

Oligoadenylate Synthetase ◽

Expression Levels

Clinical evidence indicates that electroacupuncture (EA) is effective for allergic disorder. Recent animal studies have shown that EA treatment reduces levels of IgE and Th2 cytokines in BALB/c mice immunized with 2,4-dinitrophenylated keyhole limpet protein (DNP-KLH). The hypothalamus, a brain center of the neural-immune system, is known to be activated by EA stimulation. This study was performed to identify and characterize the differentially expressed genes in the hypothalamus of DNP-KLH immunized mice that were stimulated with EA or only restrained. To this aim, we conducted a microarray analysis to evaluate the global gene expression profiles, using the hypothalamic RNA samples taken from three groups of mice: (i) normal control group (no treatments); (ii) IMH group (DNP-KLH immunization + restraint); and (iii) IMEA group (immunization + EA stimulation). The microarray analysis revealed that total 39 genes were altered in their expression levels by EA treatment. Ten genes, including T-cell receptor alpha variable region family 13 subfamily 1 (Tcra-V13.1), heat shock protein 1B (Hspa1b) and 2′–5′oligoadenylate synthetase 1F (Oas1f), were up-regulated in the IMEA group when compared with the IMH group. In contrast, 29 genes, including decay accelerating factor 2 (Daf2), NAD(P)H dehydrogenase, quinone 1 (Nqo1) and programmed cell death 1 ligand 2 (Pdcd1lg2) were down-regulated in the IMEA group as compared with the IMH group. These results suggest that EA treatment can modulate immune response in DNP-KLH immunized mice by regulating expression levels of genes that are associated with innate immune, cellular defense and/or other kinds of immune system in the hypothalamus.

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Systemic Transcriptional Responses to Hemarthrosis and FVIII Replacement in FVIII-Deficient Mice

Blood ◽

10.1182/blood-2018-99-120092 ◽

2018 ◽

Vol 132 (Supplement 1) ◽

pp. 381-381

Author(s):

Esther J Cooke ◽

Srila Gopal ◽

John Shimashita ◽

Chanond A Nasamran ◽

Kathleen M Fisch ◽

...

Keyword(s):

Gene Expression ◽

T Cell ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Cell Receptor ◽

Post Injury ◽

Transcriptional Responses ◽

T Cell Receptor Signaling ◽

Deficient Mice ◽

Cell Receptor Signaling

Abstract Introduction Hemarthrosis in patients with hemophilia (PWH) leads to local inflammation and vascular changes in the joint, but little is known about the extent and nature of systemic responses to joint bleeding. Since the spleen is a major systemic immune-modulatory organ, we quantified changes in splenic gene expression profiles in FVIII-deficient mice at baseline and after induced hemarthrosis, and in the presence and absence of FVIII replacement therapy. Methods Hemarthrosis was induced in FVIII-deficient mice by sub-patellar needle puncture +/- 100 IU/kg recombinant human FVIII (rhFVIII) intravenously 2 hours before and 6 hours after injury. Spleens were harvested on day 3 or 2 weeks post-injury (n=3-5). Spleens from uninjured mice +/- rhFVIII treatment served as controls. RNA libraries were prepared using the NEBNext Ultra II Directional RNA Library Prep Kit and sequenced on an Illumina NextSeq500 platform (single-end; 75bp reads). The limma-voom method (R Bioconductor) was used for differential expression analyses. The criteria for differential expression were: i) a log fold-change (logFC) >1 or <-1, and ii) an adjusted p value <0.05. Functional enrichment was performed using Signaling Pathway Impact Analysis and the STRING database of protein-protein interactions. Results Knee injury in FVIII-deficient mice caused gross hemarthrosis that was largely prevented with rhFVIII prophylaxis (day 2 hematocrit: 26.4% and 46.3%). Pronounced alterations in splenic gene expression profiles occurred in vehicle-treated mice on day 3 post-injury, with 4227 differentially expressed genes (DEG) and 41 perturbed pathways. This response was markedly improved with rhFVIII treatment (386 DEG; 5 pathways), and almost entirely corrected by 2 weeks. Multiple pathways relating to immune processes, inflammation, and cell survival were highly perturbed on day 3 post-injury, including cytokine-cytokine receptor interactions (pNDE=1.4x10-6) and cell cycle (pNDE=5.3x10-5). The cell cycle pathway remained significantly altered despite rhFVIII treatment (pNDE=3.5x10-6), while other pathways were comparable to uninjured mice. Analysis of the top 50 DEG that are mutual to both treatment groups revealed a striking difference in directionality, with up-regulation in the vehicle group, and down-regulation in the rhFVIII group. Together, these findings demonstrate a significant effect of rhFVIII treatment on systemic transcriptional responses to joint bleeding. Treatment with rhFVIII in the absence of hemarthrosis resulted in only 97 DEG on day 3 by the same criteria, and 233 DEG after lowering the logFC threshold from (-)1 to (-)0.5. Of these 233 genes, STRING analysis revealed perturbation of the platelet activation pathway (FDR: 7.7x10-10) for up-regulated genes (93 DEG) and the T cell receptor signaling pathway (FDR: 1.3x10-8) for down-regulated genes (140 DEG), which may corroborate a role of T cell responses to rhFVIII treatment in the development of inhibitors. These responses to rhFVIII were unique to uninjured mice and did not occur in mice with induced hemarthrosis and rhFVIII treatment. Conclusions Joint bleeding in hemophilic mice leads to acute, profound changes in multiple systemic pathways, including immune and inflammatory processes, as shown by gene expression profiling. Prophylactic treatment with rhFVIII largely corrects this response to hemarthrosis. Interestingly, in the absence of hemarthrosis, rhFVIII treatment affects platelet activation and T cell receptor signaling, whereby further analyses are required to determine if these effects stimulate or dampen immune responses. This approach can be used to explore the systemic mechanisms contributing to progressive hemophilic arthropathy in PWH, elucidate immune responses that may facilitate inhibitor formation, and lead to development of novel therapeutic strategies. Disclosures von Drygalski: UniQure BV, Bayer, Bioverativ/Sanofi, Pfizer, Novo Nordisk, Biomarin, Shire, CSL Behring: Consultancy.

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Gene Expression Profiling of Microdissected Hodgkin Reed Sternberg Cells: Molecular Subtypes and Treatment Outcome Correlations.

Blood ◽

10.1182/blood.v114.22.268.268 ◽

2009 ◽

Vol 114 (22) ◽

pp. 268-268

Author(s):

Christian Steidl ◽

Tang Lee ◽

Pedro Farinha ◽

Adele Telenius ◽

Merrill Boyle ◽

...

Keyword(s):

Gene Expression ◽

Treatment Failure ◽

B Cell ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Cell Receptor ◽

P Value ◽

Receptor Signaling ◽

Hematopoietic Progenitor ◽

Cell Receptor Signaling

Abstract Abstract 268 INTRODUCTION: Classical Hodgkin lymphoma (cHL) is unique among lymphomas due to the scarcity of the malignant Hodgkin Reed Sternberg (HRS) cells, which are derived from clonal germinal center B cells. Investigations using laser capture microdissection permit more detailed analysis of these cells. However, most recent studies were limited by low case numbers and lack of available clinical data. PATIENTS AND METHODS: We studied 29 cases of cHL and the 5 HL lines KMH2, HDLM2, L428, L540, and L1236 by gene expression profiling. All patients were treated at the BC Cancer Agency between 1984 and 2006 and received at least 4 cycles of polychemotherapy and stage-dependent radiotherapy. The cohort also included 5 biopsies taken at relapse. Treatment failure was defined as disease progression or relapse at any time (n=14) after initiation of treatment; treatment success as absence of progression (n=15). We used laser microdissection (Molecular Machines & Industries Cellcut with Nikon Eclipse TE2000-S microscope) to study the enriched HRS cell compartment separately from the microenvironment. RNA extraction was performed on pools of 1000 microdissected HRS cells in each case. Gene expression profiles were generated using Affymetrix HG UA133 2.0 Plus arrays using two-cycle labeling reactions. HRS cell profiles were compared to microdissected germinal centers (GC), and HL cell line profiles compared to enriched tonsillar CD77+ centroblasts (MACS cell separation, Miltenyi). Furthermore, we compared gene expression profiles of treatment failures to those of treatment successes. RESULTS: We identified 1342 differentially expressed probesets (fold change >5, False Discovery Rate (FDR) adjusted p value <0.001) between HRS and GC cells. Using overrepresentation analysis we found genes involved in NFκB, JAK/Stat, IL-6, IL-9 signaling, cytotoxic T lymphocyte-mediated apoptosis and IL-15 production to be significantly over-expressed in HRS cells and genes involved in B cell, T cell receptor signaling and many transcriptional regulators such as FOXO1, E2F5, IRF8, NFATC1, NFYB, POU2AF1 to be significantly under-expressed in HRS cell. Comparison of these data to differentially expressed genes in the HL cell lines (1004 genes, fold change >5, FDR-adjusted p value <0.001) showed significant overlap of genes involved in proliferation, apoptosis, IL15 signaling and B cell receptor signaling, while overrepresentation of metabolism genes was unique to the cell lines. Hierarchical clustering of all 29 primary HL cases identified 3 separate clusters characterized by 1) a cytotoxic signature, 2) TNF/TGFB receptor signaling or 3) a residual B cell signature. Dichotomizing the profiles into the two treatment outcome groups demonstrated that NFγB signaling, complement system genes and genes involved in the developmental process of hematopoietic progenitor cells, macrophages and blood vessels were overexpressed in treatment failure samples. DISCUSSION: Using microdissection of HRS cells in a large number of cases we were able to further characterize the unique expression program of HL and refine the data inventory about dysregulated cellular functions and pathways in this disease. Overexpression of genes associated with NFκB, complement and hematopoietic progenitor cells proliferation correlate strongly with treatment failure. Further study using immunohistochemistry is currently ongoing to validate these findings and to develop clinically useful biomarkers. Disclosures: Gascoyne: Roche Canada: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding.

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Platypus: an open-access software for integrating lymphocyte single-cell immune repertoires with transcriptomes

NAR Genomics and Bioinformatics ◽

10.1093/nargab/lqab023 ◽

2021 ◽

Vol 3 (2) ◽

Author(s):

Alexander Yermanos ◽

Andreas Agrafiotis ◽

Raphael Kuhn ◽

Damiano Robbiani ◽

Josephine Yates ◽

...

Keyword(s):

Gene Expression ◽

Single Cell ◽

Somatic Hypermutation ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Cell Receptor ◽

Antigen Specificity ◽

Immune Repertoire ◽

High Resolution Analysis ◽

Gene Usage

Abstract High-throughput single-cell sequencing (scSeq) technologies are revolutionizing the ability to molecularly profile B and T lymphocytes by offering the opportunity to simultaneously obtain information on adaptive immune receptor repertoires (VDJ repertoires) and transcriptomes. An integrated quantification of immune repertoire parameters, such as germline gene usage, clonal expansion, somatic hypermutation and transcriptional states opens up new possibilities for the high-resolution analysis of lymphocytes and the inference of antigen-specificity. While multiple tools now exist to investigate gene expression profiles from scSeq of transcriptomes, there is a lack of software dedicated to single-cell immune repertoires. Here, we present Platypus, an open-source software platform providing a user-friendly interface to investigate B-cell receptor and T-cell receptor repertoires from scSeq experiments. Platypus provides a framework to automate and ease the analysis of single-cell immune repertoires while also incorporating transcriptional information involving unsupervised clustering, gene expression and gene ontology. To showcase the capabilities of Platypus, we use it to analyze and visualize single-cell immune repertoires and transcriptomes from B and T cells from convalescent COVID-19 patients, revealing unique insight into the repertoire features and transcriptional profiles of clonally expanded lymphocytes. Platypus will expedite progress by facilitating the analysis of single-cell immune repertoire and transcriptome sequencing.

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Platypus: an open-access software for integrating lymphocyte single-cell immune repertoires with transcriptomes

10.1101/2020.11.09.374280 ◽

2020 ◽

Author(s):

Alexander Yermanos ◽

Andreas Agrafiotis ◽

Josephine Yates ◽

Chrysa Papadopoulou ◽

Damiano Robbiani ◽

...

Keyword(s):

Gene Expression ◽

Single Cell ◽

Somatic Hypermutation ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Cell Receptor ◽

Antigen Specificity ◽

Immune Repertoire ◽

Single Cell Sequencing ◽

Gene Usage

AbstractHigh-throughput single-cell sequencing (scSeq) technologies are revolutionizing the ability to molecularly profile B and T lymphocytes by offering the opportunity to simultaneously obtain information on adaptive immune receptor repertoires (VDJ repertoires) and transcriptomes. An integrated quantification of immune repertoire parameters such as germline gene usage, clonal expansion, somatic hypermutation and transcriptional states opens up new possibilities for the high-resolution analysis of lymphocytes and the inference of antigen-specificity. While multiple tools now exist to investigate gene expression profiles from scSeq of transcriptomes, there is a lack of software dedicated to single-cell immune repertoires. Here, we present Platypus, an open-source software platform providing a user-friendly interface to investigate B cell receptor (BCR) and T cell receptor (TCR) repertoires from single-cell sequencing experiments. Platypus provides a framework to automate and ease the analysis of single-cell immune repertoires while also incorporating transcriptional information involving unsupervised clustering, gene expression, and gene ontology. To showcase the capabilities of Platypus, we use it to analyze and visualize single-cell immune repertoires and transcriptomes from B and T cells from convalescent COVID-19 patients, revealing unique insight into the repertoire features and transcriptional profiles of clonally expanded lymphocytes. Platypus will expedite progress by increasing accessibility to the broader immunology community by facilitating the analysis of single-cell immune repertoire and transcriptome sequencing.

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