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1.
Front Immunol ; 13: 819058, 2022.
Article in English | MEDLINE | ID: covidwho-1834399

ABSTRACT

Vaccines for COVID-19 are now a crucial public health need, but the degree of protection provided by conventional vaccinations for individuals with compromised immune systems is unclear. The use of viral vectors to express neutralizing monoclonal antibodies (mAbs) in the lung is an alternative approach that does not wholly depend on individuals having intact immune systems and responses. Here, we identified an anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) monoclonal antibody, NC0321, which can efficiently neutralize a range of SARS-CoV-2 variants, including alpha, beta, delta, and eta. Both prophylactic and therapeutic NC0321 treatments effectively protected mice from SARS-CoV-2 infection. Notably, we adopted viral vector-mediated delivery of NC0321 IgG1 as an attractive approach to prevent SARS-CoV-2 infection. The NC0321 IgG1 expression in the proximal airway, expressed by a single direct in-vivo intranasal (I.N.) administration of a self-inactivating and recombinant lentiviral vector (rSIV.F/HN-NC0321), can protect young, elderly, and immunocompromised mice against mouse-adapted SARS-CoV-2 surrogate challenge. Long-term monitoring indicated that rSIV.F/HN-NC0321 mediated robust IgG expression throughout the airway of young and SCID mice, importantly, no statistical difference in the NC0321 expression between young and SCID mice was observed. A single I.N. dose of rSIV.F/HN-NC0321 30 or 180 days prior to SARS-CoV-2 challenge significantly reduced lung SARS-CoV-2 titers in an Ad5-hACE2-transduced mouse model, reconfirming that this vectored immunoprophylaxis strategy could be useful, especially for those individuals who cannot gain effective immunity from existing vaccines, and could potentially prevent clinical sequelae.


Subject(s)
COVID-19 , SARS-CoV-2 , Aged , Animals , Antibodies, Monoclonal/therapeutic use , Antibodies, Neutralizing , Antibodies, Viral , COVID-19/prevention & control , COVID-19 Vaccines , Humans , Immunoglobulin G , Mice , Mice, SCID , SARS-CoV-2/genetics , Spike Glycoprotein, Coronavirus
2.
J Am Soc Nephrol ; 32(9): 2242-2254, 2021 09.
Article in English | MEDLINE | ID: covidwho-1702796

ABSTRACT

BACKGROUND: Although coronavirus disease 2019 (COVID-19) causes significan t morbidity, mainly from pulmonary involvement, extrapulmonary symptoms are also major componen ts of the disease. Kidney disease, usually presenting as AKI, is particularly severe among patients with COVID-19. It is unknown, however, whether such injury results from direct kidney infection with COVID-19's causative virus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), or from indirect mechanisms. METHODS: Using ex vivo cell models, we sought to analyze SARS-CoV-2 interactions with kidney tubular cells and assess direct tubular injury. These models comprised primary human kidney epithelial cells (derived from nephrectomies) and grown as either proliferating monolayers or quiescent three-dimensional kidney spheroids. RESULTS: We demonstrated that viral entry molecules and high baseline levels of type 1 IFN-related molecules were present in monolayers and kidney spheroids. Although both models support viral infection and replication, they did not exhibit a cytopathic effect and cell death, outcomes that were strongly present in SARS-CoV-2-infected controls (African green monkey kidney clone E6 [Vero E6] cultures). A comparison of monolayer and spheroid cultures demonstrated higher infectivity and replication of SARS-CoV-2 in actively proliferating monolayers, although the spheroid cultures exhibited high er levels of ACE2. Monolayers exhibited elevation of some tubular injury molecules-including molecules related to fibrosis (COL1A1 and STAT6) and dedifferentiation (SNAI2)-and a loss of cell identity, evident by reduction in megalin (LRP2). The three-dimensional spheroids were less prone to such injury. CONCLUSIONS: SARS-CoV-2 can infect kidney cells without a cytopathic effect. AKI-induced cellular proliferation may potentially intensify infectivity and tubular damage by SARS-CoV-2, suggesting that early intervention in AKI is warranted to help minimize kidney infection.


Subject(s)
Acute Kidney Injury/etiology , Acute Kidney Injury/virology , COVID-19/complications , SARS-CoV-2/pathogenicity , Spheroids, Cellular/virology , Animals , Cells, Cultured , Chlorocebus aethiops , Cohort Studies , Cytopathogenic Effect, Viral , Epithelial Cells/pathology , Epithelial Cells/virology , Host Microbial Interactions , Humans , Interferon Type I/metabolism , Kidney/immunology , Kidney/pathology , Kidney/virology , Mice , Mice, Inbred NOD , Mice, SCID , Models, Biological , Pandemics , Receptors, Virus/metabolism , Retrospective Studies , SARS-CoV-2/physiology , Spheroids, Cellular/pathology , Vero Cells , Virus Replication
3.
Sci Rep ; 11(1): 17263, 2021 08 26.
Article in English | MEDLINE | ID: covidwho-1550348

ABSTRACT

Dexamethasone (Dex) is a highly insoluble front-line drug used in cancer therapy. Data from clinical trials indicates that the pharmacokinetics of Dex vary considerably between patients and prolonging drug exposure rather than increasing absolute dose may improve efficacy. Non-toxic, fully biodegradable Dex loaded nanovectors (NV) were formulated, via simple direct hydration within 10 min, as a vehicle to extend exposure and distribution in vivo. Dex-NV were just as effective as the free drug against primary human leukemia cells in vitro and in vivo. Importantly, high levels of DMSO solvent were not required in the NV formulations. Broad distribution of NV was seen rapidly following inoculation into mice. NV accumulated in major organs, including bone marrow and brain, known sanctuary sites for ALL. The study describes a non-toxic, more easily scalable system for improving Dex solubility for use in cancer and can be applied to other medical conditions associated with inflammation.


Subject(s)
Dexamethasone/administration & dosage , Drug Delivery Systems/methods , Nanostructures/chemistry , Polymers/chemistry , Precursor T-Cell Lymphoblastic Leukemia-Lymphoma/drug therapy , Xenograft Model Antitumor Assays/methods , Animals , Antineoplastic Agents, Hormonal/administration & dosage , Antineoplastic Agents, Hormonal/chemistry , Antineoplastic Agents, Hormonal/pharmacokinetics , Child , Dexamethasone/chemistry , Dexamethasone/pharmacokinetics , Drug Liberation , Humans , Kaplan-Meier Estimate , Mice, Inbred NOD , Mice, Knockout , Mice, SCID , Precursor T-Cell Lymphoblastic Leukemia-Lymphoma/metabolism , Treatment Outcome , Tumor Cells, Cultured , Young Adult
4.
J Clin Invest ; 131(21)2021 11 01.
Article in English | MEDLINE | ID: covidwho-1495789

ABSTRACT

To explore how the immune system controls clearance of SARS-CoV-2, we used a single-cell, mass cytometry-based proteomics platform to profile the immune systems of 21 patients who had recovered from SARS-CoV-2 infection without need for admission to an intensive care unit or for mechanical ventilation. We focused on receptors involved in interactions between immune cells and virus-infected cells. We found that the diversity of receptor repertoires on natural killer (NK) cells was negatively correlated with the viral clearance rate. In addition, NK subsets expressing the receptor DNAM1 were increased in patients who more rapidly recovered from infection. Ex vivo functional studies revealed that NK subpopulations with high DNAM1 expression had cytolytic activities in response to target cell stimulation. We also found that SARS-CoV-2 infection induced the expression of CD155 and nectin-4, ligands of DNAM1 and its paired coinhibitory receptor TIGIT, which counterbalanced the cytolytic activities of NK cells. Collectively, our results link the cytolytic immune responses of NK cells to the clearance of SARS-CoV-2 and show that the DNAM1 pathway modulates host-pathogen interactions during SARS-CoV-2 infection.


Subject(s)
COVID-19/immunology , COVID-19/virology , Killer Cells, Natural/immunology , Receptors, Natural Killer Cell/immunology , SARS-CoV-2/immunology , Adolescent , Adult , Aged , Animals , Antigens, Differentiation, T-Lymphocyte/immunology , Cell Adhesion Molecules/immunology , Cohort Studies , Cytotoxicity, Immunologic , Female , Heterografts , Host Microbial Interactions/immunology , Humans , Immunophenotyping , In Vitro Techniques , Ligands , Male , Mice , Mice, SCID , Middle Aged , NK Cell Lectin-Like Receptor Subfamily D/immunology , Pandemics , Receptors, Immunologic/immunology , Receptors, Virus/immunology , Viral Load , Young Adult
5.
Cells ; 10(10)2021 09 29.
Article in English | MEDLINE | ID: covidwho-1444117

ABSTRACT

Mesenchymal stem cells (MSCs) are multipotent adult stem cells present in virtually all tissues; they have a potent self-renewal capacity and can differentiate into multiple cell types. They also affect the ambient tissue by the paracrine secretion of numerous factors in vivo, including the induction of other stem cells' differentiation. In vitro, the culture media supernatant is named secretome and contains soluble molecules and extracellular vesicles that retain potent biological function in tissue regeneration. MSCs are considered safe for human treatment; their use does not involve ethical issues, as embryonic stem cells do not require genetic manipulation as induced pluripotent stem cells, and after intravenous injection, they are mainly found in the lugs. Therefore, these cells are currently being tested in various preclinical and clinical trials for several diseases, including COVID-19. Several affected COVID-19 patients develop induced acute respiratory distress syndrome (ARDS) associated with an uncontrolled inflammatory response. This condition causes extensive damage to the lungs and may leave serious post-COVID-19 sequelae. As the disease may cause systemic alterations, such as thromboembolism and compromised renal and cardiac function, the intravenous injection of MSCs may be a therapeutic alternative against multiple pathological manifestations. In this work, we reviewed the literature about MSCs biology, focusing on their function in pulmonary regeneration and their use in COVID-19 treatment.


Subject(s)
COVID-19/blood , COVID-19/therapy , Lung/physiology , Mesenchymal Stem Cell Transplantation/methods , Mesenchymal Stem Cells/cytology , Regeneration/physiology , Animals , COVID-19/drug therapy , Cell Differentiation , Cell- and Tissue-Based Therapy , Culture Media , Extracellular Vesicles , Humans , Inflammation , Mice , Mice, SCID , Phenotype , Pneumonia/blood , Pneumonia/immunology , Pneumonia/therapy , Respiratory Distress Syndrome , SARS-CoV-2 , Thromboembolism/blood , Thromboembolism/immunology , Thromboembolism/therapy
6.
Retrovirology ; 18(1): 13, 2021 06 05.
Article in English | MEDLINE | ID: covidwho-1257950

ABSTRACT

Humanized mice model human disease and as such are used commonly for research studies of infectious, degenerative and cancer disorders. Recent models also reflect hematopoiesis, natural immunity, neurobiology, and molecular pathways that influence disease pathobiology. A spectrum of immunodeficient mouse strains permit long-lived human progenitor cell engraftments. The presence of both innate and adaptive immunity enables high levels of human hematolymphoid reconstitution with cell susceptibility to a broad range of microbial infections. These mice also facilitate investigations of human pathobiology, natural disease processes and therapeutic efficacy in a broad spectrum of human disorders. However, a bridge between humans and mice requires a complete understanding of pathogen dose, co-morbidities, disease progression, environment, and genetics which can be mirrored in these mice. These must be considered for understanding of microbial susceptibility, prevention, and disease progression. With known common limitations for access to human tissues, evaluation of metabolic and physiological changes and limitations in large animal numbers, studies in mice prove important in planning human clinical trials. To these ends, this review serves to outline how humanized mice can be used in viral and pharmacologic research emphasizing both current and future studies of viral and neurodegenerative diseases. In all, humanized mouse provides cost-effective, high throughput studies of infection or degeneration in natural pathogen host cells, and the ability to test transmission and eradication of disease.


Subject(s)
Disease Models, Animal , Immunity, Innate , Mice, SCID , Neurodegenerative Diseases/immunology , Animals , HIV-1/immunology , Mice
7.
J Am Soc Nephrol ; 32(9): 2242-2254, 2021 09.
Article in English | MEDLINE | ID: covidwho-1266593

ABSTRACT

BACKGROUND: Although coronavirus disease 2019 (COVID-19) causes significan t morbidity, mainly from pulmonary involvement, extrapulmonary symptoms are also major componen ts of the disease. Kidney disease, usually presenting as AKI, is particularly severe among patients with COVID-19. It is unknown, however, whether such injury results from direct kidney infection with COVID-19's causative virus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), or from indirect mechanisms. METHODS: Using ex vivo cell models, we sought to analyze SARS-CoV-2 interactions with kidney tubular cells and assess direct tubular injury. These models comprised primary human kidney epithelial cells (derived from nephrectomies) and grown as either proliferating monolayers or quiescent three-dimensional kidney spheroids. RESULTS: We demonstrated that viral entry molecules and high baseline levels of type 1 IFN-related molecules were present in monolayers and kidney spheroids. Although both models support viral infection and replication, they did not exhibit a cytopathic effect and cell death, outcomes that were strongly present in SARS-CoV-2-infected controls (African green monkey kidney clone E6 [Vero E6] cultures). A comparison of monolayer and spheroid cultures demonstrated higher infectivity and replication of SARS-CoV-2 in actively proliferating monolayers, although the spheroid cultures exhibited high er levels of ACE2. Monolayers exhibited elevation of some tubular injury molecules-including molecules related to fibrosis (COL1A1 and STAT6) and dedifferentiation (SNAI2)-and a loss of cell identity, evident by reduction in megalin (LRP2). The three-dimensional spheroids were less prone to such injury. CONCLUSIONS: SARS-CoV-2 can infect kidney cells without a cytopathic effect. AKI-induced cellular proliferation may potentially intensify infectivity and tubular damage by SARS-CoV-2, suggesting that early intervention in AKI is warranted to help minimize kidney infection.


Subject(s)
Acute Kidney Injury/etiology , Acute Kidney Injury/virology , COVID-19/complications , SARS-CoV-2/pathogenicity , Spheroids, Cellular/virology , Animals , Cells, Cultured , Chlorocebus aethiops , Cohort Studies , Cytopathogenic Effect, Viral , Epithelial Cells/pathology , Epithelial Cells/virology , Host Microbial Interactions , Humans , Interferon Type I/metabolism , Kidney/immunology , Kidney/pathology , Kidney/virology , Mice , Mice, Inbred NOD , Mice, SCID , Models, Biological , Pandemics , Receptors, Virus/metabolism , Retrospective Studies , SARS-CoV-2/physiology , Spheroids, Cellular/pathology , Vero Cells , Virus Replication
8.
Theranostics ; 11(13): 6607-6615, 2021.
Article in English | MEDLINE | ID: covidwho-1231569

ABSTRACT

SARS-CoV-2 infection, which is responsible for the current COVID-19 pandemic, can cause life-threatening pneumonia, respiratory failure and even death. Characterizing SARS-CoV-2 pathogenesis in primary human target cells and tissues is crucial for developing vaccines and therapeutics. However, given the limited access to clinical samples from COVID-19 patients, there is a pressing need for in vitro/in vivo models to investigate authentic SARS-CoV-2 infection in primary human lung cells or tissues with mature structures. The present study was designed to evaluate a humanized mouse model carrying human lung xenografts for SARS-CoV-2 infection in vivo. Methods: Human fetal lung tissue surgically grafted under the dorsal skin of SCID mice were assessed for growth and development after 8 weeks. Following SARS-CoV-2 inoculation into the differentiated lung xenografts, viral replication, cell-type tropism and histopathology of SARS-CoV-2 infection, and local cytokine/chemokine expression were determined over a 6-day period. The effect of IFN-α treatment against SARS-CoV-2 infection was tested in the lung xenografts. Results: Human lung xenografts expanded and developed mature structures closely resembling normal human lung. SARS-CoV-2 replicated and spread efficiently in the lung xenografts with the epithelial cells as the main target, caused severe lung damage, and induced a robust pro-inflammatory response. IFN-α treatment effectively inhibited SARS-CoV-2 replication in the lung xenografts. Conclusions: These data support the human lung xenograft mouse model as a useful and biological relevant tool that should facilitate studies on the pathogenesis of SARS-CoV-2 lung infection and the evaluation of potential antiviral therapies.


Subject(s)
COVID-19/immunology , Disease Models, Animal , Lung/pathology , Respiratory Mucosa/cytology , SARS-CoV-2/immunology , Aborted Fetus , Animals , COVID-19/pathology , COVID-19/virology , Cells, Cultured , Epithelial Cells/virology , Heterografts , Humans , Lung/immunology , Lung/virology , Lung Transplantation , Male , Mice , Mice, SCID , Primary Cell Culture , SARS-CoV-2/pathogenicity , Virus Replication
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