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1.
Sci Rep ; 11(1): 13533, 2021 06 29.
Article in English | MEDLINE | ID: covidwho-1387483

ABSTRACT

The host receptor for SARS-CoV-2, angiotensin-converting enzyme 2 (ACE2), is highly expressed in small intestine. Our aim was to study colonic ACE2 expression in Crohn's disease (CD) and non-inflammatory bowel disease (non-IBD) controls. We hypothesized that the colonic expression levels of ACE2 impacts CD course. We examined the expression of colonic ACE2 in 67 adult CD and 14 NIBD control patients using RNA-seq and quantitative (q) RT-PCR. We validated ACE2 protein expression and localization in formalin-fixed, paraffin-embedded matched colon and ileal tissues using immunohistochemistry. The impact of increased ACE2 expression in CD for the risk of surgery was evaluated by a multivariate regression analysis and a Kaplan-Meier estimator. To provide critical support for the generality of our findings, we analyzed previously published RNA-seq data from two large independent cohorts of CD patients. Colonic ACE2 expression was significantly higher in a subset of adult CD patients which was defined as the ACE2-high CD subset. IHC in a sampling of ACE2-high CD patients confirmed high ACE2 protein expression in the colon and ileum compared to ACE2-low CD and NIBD patients. Notably, we found that ACE2-high CD patients are significantly more likely to undergo surgery within 5 years of CD diagnosis, and a Cox regression analysis found that high ACE2 levels is an independent risk factor for surgery (OR 2.17; 95% CI, 1.10-4.26; p = 0.025). Increased intestinal expression of ACE2 is associated with deteriorated clinical outcomes in CD patients. These data point to the need for molecular stratification that can impact CD disease-related outcomes.


Subject(s)
Angiotensin-Converting Enzyme 2/metabolism , Crohn Disease/pathology , Adolescent , Adult , Angiotensin-Converting Enzyme 2/genetics , Crohn Disease/metabolism , Crohn Disease/surgery , Female , Humans , Ileum/metabolism , Ileum/pathology , Immunohistochemistry , Inflammatory Bowel Diseases/metabolism , Inflammatory Bowel Diseases/pathology , Male , Prognosis , Proportional Hazards Models , RNA, Messenger/chemistry , RNA, Messenger/metabolism , Risk Factors , Sequence Analysis, RNA , Young Adult
2.
Eur Rev Med Pharmacol Sci ; 25(15): 5063-5069, 2021 08.
Article in English | MEDLINE | ID: covidwho-1346861

ABSTRACT

OBJECTIVE: Vaccine-induced immune thrombocytopenia (VITT) is a new syndrome occurring primarily in healthy young adults, with a female predominance, after receiving the first dose of ChAdOx1 nCoV-19 vaccine. We describe VITT syndrome characterized by severe thrombosis and thrombocytopenia found in our patient, with fatal outcome. CASE REPORT: A 58-year-old man, after 13 days from the first administration of ChAdOx1 nCoV-19 vaccine (AstraZeneca), presented with abdominal pain, diarrhea and vomitus. Laboratory tests revealed a severe thrombocytopenia, low fibrinogen serum levels and marked increase of D-dimer serum levels. The patient quickly developed a multiple organ failure, till death, three days after the hospital admission. RESULTS: At histology, in the lungs, interalveolar septa appeared thickened with microthrombi in the capillaries and veins. Interalveolar septa appeared thickened and showed vascular proliferation. Thrombi were detected in the capillaries of glomerular tufts. In the hearth, thrombi were observed in veins and capillaries. In the liver, voluminous fibrin thrombi were diffusely observed in the branches of the portal vein. Microthrombi were also found in the vasa vasorum of the wall of abdominal aorta. In the brain, microthrombi were observed in the capillaries of the choroid plexuses. Diffuse hemorrhagic necrosis was observed in the intestinal wall with marked congestion of the venous vessels. CONCLUSIONS: In our patient, the majority of data necessary for a VITT final diagnosis were present: thrombocytopenia and thrombosis in pulmonary, portal, hepatic, renal and mesenteric veins, associated with a marked increase of D-dimer serum levels. The finding of cerebral thrombosis in choroid plexuses, is a new finding in VITT. These features are suggestive for a very aggressive form of VITT.


Subject(s)
COVID-19 Vaccines/adverse effects , COVID-19/prevention & control , Purpura, Thrombocytopenic, Idiopathic/etiology , Thrombosis/etiology , Aorta/pathology , COVID-19/blood , COVID-19 Vaccines/administration & dosage , Choroid Plexus/pathology , Fibrin Fibrinogen Degradation Products/metabolism , Humans , Ileum/pathology , Kidney/pathology , Liver/pathology , Lung/pathology , Male , Middle Aged , Myocardium/pathology , Purpura, Thrombocytopenic, Idiopathic/blood , Thrombosis/blood
3.
Brief Bioinform ; 22(2): 914-923, 2021 03 22.
Article in English | MEDLINE | ID: covidwho-1343627

ABSTRACT

The novel coronavirus or COVID-19 has first been found in Wuhan, China, and became pandemic. Angiotensin-converting enzyme 2 (ACE2) plays a key role in the host cells as a receptor of Spike-I Glycoprotein of COVID-19 which causes final infection. ACE2 is highly expressed in the bladder, ileum, kidney and liver, comparing with ACE2 expression in the lung-specific pulmonary alveolar type II cells. In this study, the single-cell RNAseq data of the five tissues from different humans are curated and cell types with high expressions of ACE2 are identified. Subsequently, the protein-protein interaction networks have been established. From the network, potential biomarkers which can form functional hubs, are selected based on k-means network clustering. It is observed that angiotensin PPAR family proteins show important roles in the functional hubs. To understand the functions of the potential markers, corresponding pathways have been researched thoroughly through the pathway semantic networks. Subsequently, the pathways have been ranked according to their influence and dependency in the network using PageRank algorithm. The outcomes show some important facts in terms of infection. Firstly, renin-angiotensin system and PPAR signaling pathway can play a vital role for enhancing the infection after its intrusion through ACE2. Next, pathway networks consist of few basic metabolic and influential pathways, e.g. insulin resistance. This information corroborate the fact that diabetic patients are more vulnerable to COVID-19 infection. Interestingly, the key regulators of the aforementioned pathways are angiontensin and PPAR family proteins. Hence, angiotensin and PPAR family proteins can be considered as possible therapeutic targets. Contact: sagnik.sen2008@gmail.com, umaulik@cse.jdvu.ac.in Supplementary information: Supplementary data are available online.


Subject(s)
COVID-19/metabolism , SARS-CoV-2/pathogenicity , Algorithms , Angiotensin-Converting Enzyme 2/metabolism , COVID-19/virology , Humans , Ileum/metabolism , Ileum/pathology , Kidney/metabolism , Kidney/pathology , Liver/metabolism , Liver/pathology , Peroxisome Proliferator-Activated Receptors/metabolism , Protein Interaction Maps , Renin-Angiotensin System/physiology , Signal Transduction , Spike Glycoprotein, Coronavirus/metabolism , Urinary Bladder/metabolism , Urinary Bladder/pathology
5.
Virology ; 552: 43-51, 2021 01 02.
Article in English | MEDLINE | ID: covidwho-843443

ABSTRACT

This study focused on intestinal restitution including phenotype switching of absorptive enterocytes and the abundance of different enterocyte subtypes in weaned pigs after porcine epidemic diarrhea virus (PEDV) infection. At 10 days post-PEDV-inoculation, the ratio of villus height to crypt depth in both jejunum and ileum had restored, and the PEDV antigen was not detectable. However, enterocytes at the villus tips revealed epithelial-mesenchymal transition (EMT) in the jejunum in which E-cadherin expression decreased while expression of N-cadherin, vimentin, and Snail increased. Additionally, there was reduced expression of actin in microvilli and Zonula occludens-1 (ZO-1) in tight junctions. Moreover, the protein concentration of transforming growth factor ß1 (TGFß1), which mediates EMT and cytoskeleton alteration, was increased. We also found a decreased number of Peyer's patch M cells in the ileum. These results reveal incomplete restitution of enterocytes in the jejunum and potentially impaired immune surveillance in the ileum after PEDV infection.


Subject(s)
Coronavirus Infections/veterinary , Enterocytes/pathology , Epithelial-Mesenchymal Transition , Gastroenteritis, Transmissible, of Swine/pathology , Peyer's Patches/pathology , Porcine epidemic diarrhea virus/pathogenicity , Animals , Cadherins/metabolism , Coronavirus Infections/immunology , Coronavirus Infections/pathology , Coronavirus Infections/virology , Gastroenteritis, Transmissible, of Swine/immunology , Gastroenteritis, Transmissible, of Swine/virology , Ileum/immunology , Ileum/pathology , Intestinal Mucosa/pathology , Jejunum/immunology , Jejunum/pathology , Microvilli/pathology , Swine , Tight Junctions/pathology , Transforming Growth Factor beta1/metabolism , Weaning
6.
J Crohns Colitis ; 15(3): 485-498, 2021 Mar 05.
Article in English | MEDLINE | ID: covidwho-756892

ABSTRACT

BACKGROUND: Patients with inflammatory bowel disease [IBD] are considered immunosuppressed, but do not seem more vulnerable for COVID-19. Nevertheless, intestinal inflammation has shown to be an important risk factor for SARS-CoV-2 infection and prognosis. Therefore, we investigated the role of intestinal inflammation on the viral intestinal entry mechanisms, including ACE2, in IBD. METHODS: We collected inflamed and uninflamed mucosal biopsies from Crohn's disease [CD] [n = 193] and ulcerative colitis [UC] [n = 158] patients, and from 51 matched non-IBD controls for RNA sequencing, differential gene expression, and co-expression analysis. Organoids from UC patients were subjected to an inflammatory mix and processed for RNA sequencing. Transmural ileal biopsies were processed for single-cell [sc] sequencing. Publicly available colonic sc-RNA sequencing data, and microarrays from tissue pre/post anti-tumour necrosis factor [TNF] therapy, were analysed. RESULTS: In inflamed CD ileum, ACE2 was significantly decreased compared with control ileum [p = 4.6E-07], whereas colonic ACE2 was higher in inflamed colon of CD/UC compared with control [p = 8.3E-03; p = 1.9E-03]. Sc-RNA sequencing confirmed this ACE2 dysregulation and exclusive epithelial ACE2 expression. Network analyses highlighted HNF4A as key regulator of ileal ACE2, and pro-inflammatory cytokines and interferon regulating factors regulated colonic ACE2. Inflammatory stimuli upregulated ACE2 in UC organoids [p = 1.7E-02], but not in non-IBD controls [p = 9.1E-01]. Anti-TNF therapy restored colonic ACE2 regulation in responders. CONCLUSIONS: Intestinal inflammation alters SARS-CoV-2 coreceptors in the intestine, with opposing dysregulations in ileum and colon. HNF4A, an IBD susceptibility gene, seems an important upstream regulator of ACE2 in ileum, whereas interferon signalling might dominate in colon.


Subject(s)
Angiotensin-Converting Enzyme 2/immunology , COVID-19 , Colitis, Ulcerative , Colon , Crohn Disease , Hepatocyte Nuclear Factor 4 , Ileum , Interferons/immunology , SARS-CoV-2/physiology , Biopsy/methods , COVID-19/immunology , COVID-19/pathology , COVID-19/physiopathology , Colitis, Ulcerative/immunology , Colitis, Ulcerative/pathology , Colitis, Ulcerative/virology , Colon/immunology , Colon/pathology , Colon/virology , Crohn Disease/immunology , Crohn Disease/pathology , Crohn Disease/virology , Cytokines/immunology , Female , Gene Expression Regulation , Hepatocyte Nuclear Factor 4/genetics , Hepatocyte Nuclear Factor 4/immunology , Humans , Ileum/immunology , Ileum/pathology , Ileum/virology , Male , Middle Aged , Sequence Analysis, RNA , Signal Transduction , Single-Cell Analysis
7.
J Virol ; 94(17)2020 08 17.
Article in English | MEDLINE | ID: covidwho-601769

ABSTRACT

Coronaviruses (CoVs) have repeatedly emerged from wildlife hosts and infected humans and livestock animals to cause epidemics with significant morbidity and mortality. CoVs infect various organs, including respiratory and enteric systems, as exemplified by newly emerged severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The constellation of viral factors that contribute to developing enteric disease remains elusive. Here, we investigated CoV interferon antagonists for their contribution to enteric pathogenesis. Using an infectious clone of an enteric CoV, porcine epidemic diarrhea virus (icPEDV), we generated viruses with inactive versions of interferon antagonist nonstructural protein 1 (nsp1), nsp15, and nsp16 individually or combined into one virus designated icPEDV-mut4. Interferon-responsive PK1 cells were infected with these viruses and produced higher levels of interferon responses than were seen with wild-type icPEDV infection. icPEDV-mut4 elicited robust interferon responses and was severely impaired for replication in PK1 cells. To evaluate viral pathogenesis, piglets were infected with either icPEDV or icPEDV-mut4. While the icPEDV-infected piglets exhibited clinical disease, the icPEDV-mut4-infected piglets showed no clinical symptoms and exhibited normal intestinal pathology at day 2 postinfection. icPEDV-mut4 replicated in the intestinal tract, as revealed by detection of viral RNA in fecal swabs, with sequence analysis documenting genetic stability of the input strain. Importantly, icPEDV-mut4 infection elicited IgG and neutralizing antibody responses to PEDV. These results identify nsp1, nsp15, and nsp16 as virulence factors that contribute to the development of PEDV-induced diarrhea in swine. Inactivation of these CoV interferon antagonists is a rational approach for generating candidate vaccines to prevent disease and spread of enteric CoVs, including SARS-CoV-2.IMPORTANCE Emerging coronaviruses, including SARS-CoV-2 and porcine CoVs, can infect enterocytes, cause diarrhea, and be shed in the feces. New approaches are needed to understand enteric pathogenesis and to develop vaccines and therapeutics to prevent the spread of these viruses. Here, we exploited a reverse genetic system for an enteric CoV, porcine epidemic diarrhea virus (PEDV), and outline an approach of genetically inactivating highly conserved viral factors known to limit the host innate immune response to infection. Our report reveals that generating PEDV with inactive versions of three viral interferon antagonists, nonstructural proteins 1, 15, and 16, results in a highly attenuated virus that does not cause diarrhea in animals and elicits a neutralizing antibody response in virus-infected animals. This strategy may be useful for generating live attenuated vaccine candidates that prevent disease and fecal spread of enteric CoVs, including SARS-CoV-2.


Subject(s)
Coronavirus Infections/immunology , Coronavirus/immunology , Interferons/immunology , Porcine epidemic diarrhea virus/immunology , Vaccines, Attenuated/immunology , Viral Nonstructural Proteins/antagonists & inhibitors , Animals , Betacoronavirus/immunology , COVID-19 , Chlorocebus aethiops , Coronavirus Infections/prevention & control , Diarrhea/pathology , Diarrhea/virology , Disease Models, Animal , Endoribonucleases/antagonists & inhibitors , Feces/virology , Ileum/pathology , Immunity, Innate , Jejunum/pathology , Pandemics , Pneumonia, Viral/immunology , Porcine epidemic diarrhea virus/genetics , RNA, Viral , RNA-Dependent RNA Polymerase , SARS-CoV-2 , Swine , Swine Diseases/virology , Vero Cells , Viral Nonstructural Proteins/genetics , Viral Nonstructural Proteins/immunology
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