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1.
EuropePMC; 2020.
Preprint in English | EuropePMC | ID: ppcovidwho-317546

ABSTRACT

Preexisting diabetes increases the risk of a severe course of the pandemic coronavirus disease 2019 (COVID-19). Vice versa, exacerbations of a preexisting diabetes as well as new-onset diabetes have been reported upon SARS-CoV-2 infection. Thus, there is an imperative need to clarify whether human pancreatic endocrine cells organized within an islet of Langerhans are permissive for and affected by SARS-CoV-2 infection, and to elucidate the mechanisms underlying the development of diabetes upon COVID-19. Here, we (i) defined ACE2 and TMPRSS2 expression patterns in human pancreatic endocrine and exocrine cell types, (ii) employed human pancreatic islet cultures to demonstrate susceptibility to SARS-CoV-2 infection and to viral replication in β-cells, (iii) showed that SARS-CoV-2 attenuates glucose-stimulated insulin secretion, and (iv) tested remdesivir as eventually effective to prevent β-cell failure. In addition, we (v) visualized viral particles replicating in endocrine pancreatic cells and define their subcellular localization patterns via transmission electron microscopy, and finally (vi) present examples of cell type specific pancreatic infection patterns of COVID-19 deceased patients. Overall, our data demonstrate that SARS-CoV-2 can infect both the exocrine and endocrine compartments of the pancreas and can perturb β-cell integrity, which might lead to an increased risk for diabetes.

2.
Nat Commun ; 12(1): 6855, 2021 11 25.
Article in English | MEDLINE | ID: covidwho-1537312

ABSTRACT

The bat sarbecovirus RaTG13 is a close relative of SARS-CoV-2, the cause of the COVID-19 pandemic. However, this bat virus was most likely unable to directly infect humans since its Spike (S) protein does not interact efficiently with the human ACE2 receptor. Here, we show that a single T403R mutation increases binding of RaTG13 S to human ACE2 and allows VSV pseudoparticle infection of human lung cells and intestinal organoids. Conversely, mutation of R403T in the SARS-CoV-2 S reduces pseudoparticle infection and viral replication. The T403R RaTG13 S is neutralized by sera from individuals vaccinated against COVID-19 indicating that vaccination might protect against future zoonoses. Our data suggest that a positively charged amino acid at position 403 in the S protein is critical for efficient utilization of human ACE2 by S proteins of bat coronaviruses. This finding could help to better predict the zoonotic potential of animal coronaviruses.


Subject(s)
Angiotensin-Converting Enzyme 2/chemistry , Protein Binding , SARS-CoV-2/genetics , SARS-CoV-2/metabolism , Spike Glycoprotein, Coronavirus/chemistry , Spike Glycoprotein, Coronavirus/genetics , Animals , COVID-19/virology , COVID-19 Vaccines , Caco-2 Cells , Cloning, Molecular , HEK293 Cells , Humans , Molecular Dynamics Simulation , Mutation , Replicon , Species Specificity , Stem Cells , Zoonoses
3.
Nat Commun ; 12(1): 4584, 2021 07 28.
Article in English | MEDLINE | ID: covidwho-1387354

ABSTRACT

Interferon-induced transmembrane proteins (IFITMs 1, 2 and 3) can restrict viral pathogens, but pro- and anti-viral activities have been reported for coronaviruses. Here, we show that artificial overexpression of IFITMs blocks SARS-CoV-2 infection. However, endogenous IFITM expression supports efficient infection of SARS-CoV-2 in human lung cells. Our results indicate that the SARS-CoV-2 Spike protein interacts with IFITMs and hijacks them for efficient viral infection. IFITM proteins were expressed and further induced by interferons in human lung, gut, heart and brain cells. IFITM-derived peptides and targeting antibodies inhibit SARS-CoV-2 entry and replication in human lung cells, cardiomyocytes and gut organoids. Our results show that IFITM proteins are cofactors for efficient SARS-CoV-2 infection of human cell types representing in vivo targets for viral transmission, dissemination and pathogenesis and are potential targets for therapeutic approaches.


Subject(s)
Angiotensin-Converting Enzyme 2/genetics , Antigens, Differentiation/genetics , Membrane Proteins/genetics , RNA-Binding Proteins/genetics , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus/genetics , Amino Acid Sequence , Angiotensin-Converting Enzyme 2/antagonists & inhibitors , Angiotensin-Converting Enzyme 2/metabolism , Antibodies, Neutralizing/pharmacology , Antigens, Differentiation/metabolism , Binding Sites , COVID-19/virology , Gene Expression Regulation , Host-Pathogen Interactions/drug effects , Host-Pathogen Interactions/genetics , Humans , Interferon-beta/pharmacology , Membrane Proteins/antagonists & inhibitors , Membrane Proteins/metabolism , Protein Binding , Protein Interaction Domains and Motifs , RNA, Small Interfering/genetics , RNA, Small Interfering/metabolism , RNA-Binding Proteins/antagonists & inhibitors , RNA-Binding Proteins/metabolism , SARS-CoV-2/drug effects , SARS-CoV-2/metabolism , Sequence Alignment , Sequence Homology, Amino Acid , Spike Glycoprotein, Coronavirus/metabolism , Virus Attachment/drug effects
4.
Z Gastroenterol ; 59(11): 1205-1213, 2021 Nov.
Article in German | MEDLINE | ID: covidwho-1324455

ABSTRACT

SARS-CoV-2 is a novel human pathogenic coronavirus whose predilection for the respiratory tract has given rise to a rapid pandemic spread via airborne particles. Organ-specific susceptibility is substantially determined by the density of cell surface expression of ACE2, which is exploited by viral spike protein as a receptor molecule to mediate adhesion and, thus, to permit internalization of the viral genome into the host cell. Based on an ample data set derived from clinical studies and case reports, evidence suggests that distinct cell populations of the digestive and olfactory-gustatory system are equally equipped with membrane-bound ACE2, rendering them "vulnerable" to SARS-CoV-2. Numerous reports on concomitant gastrointestinal complaints and laboratory abnormalities are thought to reflect a relevant degree of organ dysfunction and underscore the tropism of SARS-CoV-2 for the digestive tract. Organoids are three-dimensional in vitro replicas of organ tissue which, owing to their organotypic complex cellular composition and functional resemblance to primary cells, are particularly appreciated for basic research in the field of infectious diseases. This review specifically addresses the involvement of digestive organs by SARS-CoV-2 and outlines the significant contribution of organoid- and primary-cell culture-based models to gaining a deeper understanding of the underlying pathophysiological processes.


Subject(s)
COVID-19 , Communicable Diseases , Gastrointestinal Tract , Humans , Organoids , SARS-CoV-2 , Virus Internalization
5.
Nat Commun ; 12(1): 1726, 2021 03 19.
Article in English | MEDLINE | ID: covidwho-1142436

ABSTRACT

SARS-CoV-2 is a respiratory pathogen and primarily infects the airway epithelium. As our knowledge about innate immune factors of the respiratory tract against SARS-CoV-2 is limited, we generated and screened a peptide/protein library derived from bronchoalveolar lavage for inhibitors of SARS-CoV-2 spike-driven entry. Analysis of antiviral fractions revealed the presence of α1-antitrypsin (α1AT), a highly abundant circulating serine protease inhibitor. Here, we report that α1AT inhibits SARS-CoV-2 entry at physiological concentrations and suppresses viral replication in cell lines and primary cells including human airway epithelial cultures. We further demonstrate that α1AT binds and inactivates the serine protease TMPRSS2, which enzymatically primes the SARS-CoV-2 spike protein for membrane fusion. Thus, the acute phase protein α1AT is an inhibitor of TMPRSS2 and SARS-CoV-2 entry, and may play an important role in the innate immune defense against the novel coronavirus. Our findings suggest that repurposing of α1AT-containing drugs has prospects for the therapy of COVID-19.


Subject(s)
COVID-19/drug therapy , SARS-CoV-2/drug effects , Serine Endopeptidases/metabolism , Serine Proteinase Inhibitors/pharmacology , alpha 1-Antitrypsin/pharmacology , Antibodies, Viral/blood , Antiviral Agents/pharmacology , COVID-19/blood , Caco-2 Cells , Humans , Immunoglobulin G/blood , Molecular Docking Simulation , Spike Glycoprotein, Coronavirus/metabolism , Virus Internalization/drug effects , Virus Replication/drug effects
6.
Cell ; 184(9): 2384-2393.e12, 2021 04 29.
Article in English | MEDLINE | ID: covidwho-1141659

ABSTRACT

The global spread of SARS-CoV-2/COVID-19 is devastating health systems and economies worldwide. Recombinant or vaccine-induced neutralizing antibodies are used to combat the COVID-19 pandemic. However, the recently emerged SARS-CoV-2 variants B.1.1.7 (UK), B.1.351 (South Africa), and P.1 (Brazil) harbor mutations in the viral spike (S) protein that may alter virus-host cell interactions and confer resistance to inhibitors and antibodies. Here, using pseudoparticles, we show that entry of all variants into human cells is susceptible to blockade by the entry inhibitors soluble ACE2, Camostat, EK-1, and EK-1-C4. In contrast, entry of the B.1.351 and P.1 variant was partially (Casirivimab) or fully (Bamlanivimab) resistant to antibodies used for COVID-19 treatment. Moreover, entry of these variants was less efficiently inhibited by plasma from convalescent COVID-19 patients and sera from BNT162b2-vaccinated individuals. These results suggest that SARS-CoV-2 may escape neutralizing antibody responses, which has important implications for efforts to contain the pandemic.


Subject(s)
Antibodies, Neutralizing/immunology , Antibodies, Viral/immunology , SARS-CoV-2/immunology , Animals , COVID-19/immunology , COVID-19/therapy , COVID-19/virology , Cell Line , Drug Resistance, Viral , Humans , Immunization, Passive , Kinetics , Membrane Fusion , Models, Molecular , Neutralization Tests , Serine Endopeptidases/metabolism , Solubility , Spike Glycoprotein, Coronavirus/immunology , Vaccination , Virus Internalization
7.
Nat Metab ; 3(2): 149-165, 2021 02.
Article in English | MEDLINE | ID: covidwho-1065968

ABSTRACT

Infection-related diabetes can arise as a result of virus-associated ß-cell destruction. Clinical data suggest that the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), causing the coronavirus disease 2019 (COVID-19), impairs glucose homoeostasis, but experimental evidence that SARS-CoV-2 can infect pancreatic tissue has been lacking. In the present study, we show that SARS-CoV-2 infects cells of the human exocrine and endocrine pancreas ex vivo and in vivo. We demonstrate that human ß-cells express viral entry proteins, and SARS-CoV-2 infects and replicates in cultured human islets. Infection is associated with morphological, transcriptional and functional changes, including reduced numbers of insulin-secretory granules in ß-cells and impaired glucose-stimulated insulin secretion. In COVID-19 full-body postmortem examinations, we detected SARS-CoV-2 nucleocapsid protein in pancreatic exocrine cells, and in cells that stain positive for the ß-cell marker NKX6.1 and are in close proximity to the islets of Langerhans in all four patients investigated. Our data identify the human pancreas as a target of SARS-CoV-2 infection and suggest that ß-cell infection could contribute to the metabolic dysregulation observed in patients with COVID-19.


Subject(s)
Islets of Langerhans/virology , SARS-CoV-2/growth & development , Aged , Aged, 80 and over , Angiotensin-Converting Enzyme 2/biosynthesis , Angiotensin-Converting Enzyme 2/genetics , COVID-19/physiopathology , Cells, Cultured , Diabetes Mellitus , Female , Humans , Islets of Langerhans/cytology , Islets of Langerhans/physiopathology , Male , Pancreas, Exocrine/cytology , Pancreas, Exocrine/physiopathology , Pancreas, Exocrine/virology , Pancreatic Diseases/etiology , Pancreatic Diseases/virology , Serine Endopeptidases/biosynthesis , Serine Endopeptidases/genetics , Virus Internalization , Virus Replication
8.
Cells Tissues Organs ; 209(4-6): 155-164, 2020.
Article in English | MEDLINE | ID: covidwho-1042717

ABSTRACT

Usually, pandemic COVID-19 disease, caused by SARS-CoV2, presents with mild respiratory symptoms such as fever, cough, but frequently also with anosmia and neurological symptoms. Virus-cell fusion is mediated by angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2) with their organ expression pattern determining viral tropism. Clinical presentation suggests rapid viral dissemination to the central nervous system leading frequently to severe symptoms including viral meningitis. Here, we provide a comprehensive expression landscape of ACE2 and TMPRSS2 proteins across human postmortem nasal and olfactory tissue. Sagittal sections through the human nose complemented with immunolabelling of respective cell types represent different anatomically defined regions including olfactory epithelium, respiratory epithelium of the nasal conchae and the paranasal sinuses along with the hardly accessible human olfactory bulb. ACE2 can be detected in the olfactory epithelium as well as in the respiratory epithelium of the nasal septum, the nasal conchae, and the paranasal sinuses. ACE2 is located in the sustentacular cells and in the glandular cells in the olfactory epithelium as well as in the basal cells, glandular cells, and epithelial cells of the respiratory epithelium. Intriguingly, ACE2 is not expressed in mature or immature olfactory receptor neurons and basal cells in the olfactory epithelium. Similarly, ACE2 is not localized in the olfactory receptor neurons albeit the olfactory bulb is positive. Vice versa, TMPRSS2 can also be detected in the sustentacular cells and the glandular cells of the olfactory epithelium. Our findings provide the basic anatomical evidence for the expression of ACE2 and TMPRSS2 in the human nose, olfactory epithelium, and olfactory bulb. Thus, they are substantial for future studies that aim to elucidate the symptom of SARS-CoV2 induced anosmia via the olfactory pathway.


Subject(s)
Angiotensin-Converting Enzyme 2/analysis , COVID-19/pathology , Nasal Mucosa/pathology , Olfactory Bulb/pathology , SARS-CoV-2/isolation & purification , Serine Endopeptidases/analysis , COVID-19/diagnosis , Humans , Nasal Mucosa/virology , Nose/pathology , Nose/virology , Olfactory Bulb/virology , Olfactory Mucosa/pathology , Olfactory Mucosa/virology
9.
Cell Mol Gastroenterol Hepatol ; 11(4): 935-948, 2021.
Article in English | MEDLINE | ID: covidwho-917333

ABSTRACT

BACKGROUND AND AIMS: The COVID-19 pandemic has spread worldwide and poses a severe health risk. While most patients present mild symptoms, descending pneumonia can lead to severe respiratory insufficiency. Up to 50% of patients show gastrointestinal symptoms like diarrhea or nausea, intriguingly associating with prolonged symptoms and increased severity. Thus, models to understand and validate drug efficiency in the gut of COVID-19 patients are of urgent need. METHODS: Human intestinal organoids derived from pluripotent stem cells (PSC-HIOs) have led, due to their complexity in mimicking human intestinal architecture, to an unprecedented number of successful disease models including gastrointestinal infections. Here, we employed PSC-HIOs to dissect SARS-CoV-2 pathogenesis and its inhibition by remdesivir, one of the leading drugs investigated for treatment of COVID-19. RESULTS: Immunostaining for viral entry receptor ACE2 and SARS-CoV-2 spike protein priming protease TMPRSS2 showed broad expression in the gastrointestinal tract with highest levels in the intestine, the latter faithfully recapitulated by PSC-HIOs. Organoids could be readily infected with SARS-CoV-2 followed by viral spread across entire PSC-HIOs, subsequently leading to organoid deterioration. However, SARS-CoV-2 spared goblet cells lacking ACE2 expression. Importantly, we challenged PSC-HIOs for drug testing capacity. Specifically, remdesivir effectively inhibited SARS-CoV-2 infection dose-dependently at low micromolar concentration and rescued PSC-HIO morphology. CONCLUSIONS: Thus, PSC-HIOs are a valuable tool to study SARS-CoV-2 infection and to identify and validate drugs especially with potential action in the gut.


Subject(s)
Adenosine Monophosphate/analogs & derivatives , Alanine/analogs & derivatives , COVID-19/drug therapy , COVID-19/metabolism , Human Embryonic Stem Cells , Intestinal Mucosa , Organoids , SARS-CoV-2/physiology , Virus Replication/drug effects , Adenosine Monophosphate/pharmacology , Alanine/pharmacology , Caco-2 Cells , Human Embryonic Stem Cells/metabolism , Human Embryonic Stem Cells/pathology , Human Embryonic Stem Cells/virology , Humans , Intestinal Mucosa/metabolism , Intestinal Mucosa/pathology , Intestinal Mucosa/virology , Organoids/metabolism , Organoids/pathology , Organoids/virology
10.
J Virol Methods ; 286: 113965, 2020 12.
Article in English | MEDLINE | ID: covidwho-741388

ABSTRACT

Pandemic SARS-CoV-2 infection has rapidly developed into a socioeconomic and humanitarian catastrophe. Basic principles to prevent SARS-CoV-2 transmission are social distancing, face masks, contact tracing and early detection of SARS-CoV-2. To meet these requirements, virtually unlimited test capacities delivering results in a rapid and reliable manner are a prerequisite. Here, we provide and validate such a rapid, convenient and efficient kit-independent detection of SARS-CoV-2 RNA, termed COVID-quick-DET. This straightforward method operates with simple proteinase K treatment and repetitive heating steps with a sensitivity of 94.6% in head-to-head comparisons with kit-based isolation methods. This result is supported by data obtained from serially diluted SARS-CoV-2 virus stocks. Given its cost- and time-effective operation, COVID-quick-DET might be best suited for countries with general shortage or temporary acute scarcity of resources and equipment.


Subject(s)
Betacoronavirus/genetics , Betacoronavirus/isolation & purification , Clinical Laboratory Techniques/methods , Coronavirus Infections/diagnosis , Coronavirus Infections/virology , Pneumonia, Viral/diagnosis , Pneumonia, Viral/virology , RNA, Viral/isolation & purification , Reagent Kits, Diagnostic , COVID-19 , COVID-19 Testing , Diagnostic Tests, Routine , Endopeptidase K/chemistry , Heating , Humans , Pandemics , RNA, Viral/genetics , SARS-CoV-2 , Sensitivity and Specificity
11.
Clin Med (Lond) ; 20(6): e218-e221, 2020 11.
Article in English | MEDLINE | ID: covidwho-736876

ABSTRACT

Here, we advocate a highly favourable opportunity for the treatment of COVID-19 disease by repurposing a long-serving medical agent with an excellent history of clinical use, namely heparin. Heparin is best known as an anticoagulant, but it also exhibits direct antiviral activity against many enveloped viruses and has anti-inflammatory activity. The high incidence of thromboembolic events in COVID-19 patients suggests that coagulopathy plays an important role in the SARS-CoV-2 pathogenesis. This already makes heparin a unique, potentially curative agent that can be used immediately to help resolve the ongoing crisis associated with SARS-CoV-2 infection and COVID-19 disease. We demonstrate here in vitro that heparin does indeed inhibit SARS-CoV-2 infection. The three concurrent modes of activity of heparin (antiviral, anticoagulant and anti-inflammatory) against SARS-CoV-2/COVID-19 form a unique therapeutic combination. Thus, repurposing of heparin to fight SARS-CoV-2 and COVID-19 appears to be a powerful, readily available measure to address the current pandemic.


Subject(s)
Anti-Inflammatory Agents/therapeutic use , Anticoagulants/therapeutic use , Antiviral Agents/therapeutic use , Coronavirus Infections/drug therapy , Heparin/therapeutic use , Pneumonia, Viral/drug therapy , Administration, Inhalation , Betacoronavirus , COVID-19 , Drug Repositioning , Humans , Pandemics , SARS-CoV-2
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