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1.
Methods Mol Biol ; 2813: 117-123, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38888774

RESUMO

The emergence of zoonotic viruses like severe acute respiratory syndrome coronavirus (SARS-CoV), Middle East respiratory syndrome coronavirus (MERS-CoV), and SARS-CoV-2 have significantly impacted global health and economy. The discovery of other viruses in wildlife reservoir species present a threat for future emergence in humans and animals. Therefore, assays that are less reliant on virus-specific information, such as neutralization assays, are crucial to rapidly develop diagnostics, understand virus replication and pathogenicity, and assess the efficacy of therapeutics against newly emerging viruses. Here, we describe the discontinuous median tissue culture infectious dose 50 (TCID50) assay to quantitatively determine the titer of any virus that can produce a visible cytopathic effect in infected cells.


Assuntos
Efeito Citopatogênico Viral , Animais , Humanos , SARS-CoV-2/patogenicidade , SARS-CoV-2/fisiologia , Chlorocebus aethiops , COVID-19/virologia , Células Vero , Replicação Viral , Técnicas de Cultura de Tecidos/métodos
2.
PLoS One ; 19(6): e0304504, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38870232

RESUMO

To determine why SARS-CoV-2 appears to thrive specifically well in meat packaging plants, we used SARS-CoV-2 Delta variant and meat packaging plant drain samples to develop mixed-species biofilms on materials commonly found within meat packaging plants (stainless steel (SS), PVC, and ceramic tile). Our data provides evidence that SARS-CoV-2 Delta variant remained viable on all the surfaces tested with and without an environmental biofilm after the virus was inoculated with the biofilm for 5 days at 7°C. We observed that SARS-CoV-2 Delta variant was able to remain infectious with each of the environmental biofilms by conducting plaque assay and qPCR experiments, however, we detected a significant reduction in viability post-exposure to Plant B biofilm on SS, PVC, and on ceramic tile chips, and to Plant C biofilm on SS and PVC chips. The numbers of viable SARS-CoV-2 Delta viral particles was 1.81-4.57-fold high than the viral inoculum incubated with the Plant B and Plant C environmental biofilm on SS, and PVC chips. We did not detect a significant difference in viability when SARS-CoV-2 Delta variant was incubated with the biofilm obtained from Plant A on any of the materials tested and SARS-CoV-2 Delta variant had higher plaque numbers when inoculated with Plant C biofilm on tile chips, with a 2.75-fold difference compared to SARS-CoV-2 Delta variant on tile chips by itself. In addition, we detected an increase in the biofilm biovolume in response to SARS-CoV-2 Delta variant which is also a concern for food safety due to the potential for foodborne pathogens to respond likewise when they come into contact with the virus. These results indicate a complex virus-environmental biofilm interaction which correlates to the different bacteria found in each biofilm. Our results also indicate that there is the potential for biofilms to protect SARS-CoV-2 from disinfecting agents and remaining prevalent in meat packaging plants.


Assuntos
Biofilmes , Embalagem de Alimentos , SARS-CoV-2 , Biofilmes/crescimento & desenvolvimento , SARS-CoV-2/fisiologia , SARS-CoV-2/isolamento & purificação , SARS-CoV-2/genética , Embalagem de Alimentos/métodos , Humanos , COVID-19/microbiologia , COVID-19/virologia , COVID-19/transmissão , Aço Inoxidável , Carne/microbiologia , Carne/virologia
3.
Open Biol ; 14(6): 230349, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38862017

RESUMO

Coronavirus disease 2019 (COVID-19) was initially considered a primarily respiratory disease but is now known to affect other organs including the heart and brain. A major route by which COVID-19 impacts different organs is via the vascular system. We studied the impact of apolipoprotein E (APOE) genotype and inflammation on vascular infectivity by pseudo-typed severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) viruses in mouse and human cultured endothelial cells and pericytes. Possessing the APOE4 allele or having existing systemic inflammation is known to enhance the severity of COVID-19. Using targeted replacement human APOE3 and APOE4 mice and inflammation induced by bacterial lipopolysaccharide (LPS), we investigated infection by SARS-CoV-2. Here, we show that infectivity was higher in murine cerebrovascular pericytes compared to endothelial cells and higher in cultures expressing APOE4. Furthermore, increasing the inflammatory state of the cells by prior incubation with LPS increased infectivity into human and mouse pericytes and human endothelial cells. Our findings provide insights into the mechanisms underlying severe COVID-19 infection, highlighting how risk factors such as APOE4 genotype and prior inflammation may exacerbate disease severity by augmenting the virus's ability to infect vascular cells.


Assuntos
COVID-19 , Células Endoteliais , Pericitos , SARS-CoV-2 , Pericitos/virologia , Pericitos/metabolismo , Pericitos/patologia , Humanos , Animais , SARS-CoV-2/fisiologia , SARS-CoV-2/patogenicidade , COVID-19/virologia , COVID-19/patologia , Camundongos , Células Endoteliais/virologia , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Fatores de Risco , Lipopolissacarídeos/farmacologia , Apolipoproteína E4/genética , Apolipoproteína E4/metabolismo , Apolipoproteína E3/genética , Apolipoproteína E3/metabolismo , Inflamação/virologia , Inflamação/patologia
4.
Nat Commun ; 15(1): 4996, 2024 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-38862527

RESUMO

Assessing the impact of SARS-CoV-2 on organelle dynamics allows a better understanding of the mechanisms of viral replication. We combine label-free holotomographic microscopy with Artificial Intelligence to visualize and quantify the subcellular changes triggered by SARS-CoV-2 infection. We study the dynamics of shape, position and dry mass of nucleoli, nuclei, lipid droplets and mitochondria within hundreds of single cells from early infection to syncytia formation and death. SARS-CoV-2 infection enlarges nucleoli, perturbs lipid droplets, changes mitochondrial shape and dry mass, and separates lipid droplets from mitochondria. We then used Bayesian network modeling on organelle dry mass states to define organelle cross-regulation networks and report modifications of organelle cross-regulation that are triggered by infection and syncytia formation. Our work highlights the subcellular remodeling induced by SARS-CoV-2 infection and provides an Artificial Intelligence-enhanced, label-free methodology to study in real-time the dynamics of cell populations and their content.


Assuntos
Teorema de Bayes , COVID-19 , Gotículas Lipídicas , Mitocôndrias , SARS-CoV-2 , SARS-CoV-2/fisiologia , Humanos , COVID-19/virologia , COVID-19/metabolismo , Mitocôndrias/metabolismo , Gotículas Lipídicas/metabolismo , Gotículas Lipídicas/virologia , Inteligência Artificial , Nucléolo Celular/metabolismo , Nucléolo Celular/virologia , Replicação Viral , Núcleo Celular/metabolismo , Núcleo Celular/virologia , Animais , Chlorocebus aethiops , Células Vero
5.
Mol Med ; 30(1): 81, 2024 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-38862942

RESUMO

BACKGROUND: Studies have highlighted a possible crosstalk between the pathogeneses of COVID-19 and systemic lupus erythematosus (SLE); however, the interactive mechanisms remain unclear. We aimed to elucidate the impact of COVID-19 on SLE using clinical information and the underlying mechanisms of both diseases. METHODS: RNA-seq datasets were used to identify shared hub gene signatures between COVID-19 and SLE, while genome-wide association study datasets were used to delineate the interaction mechanisms of the key signaling pathways. Finally, single-cell RNA-seq datasets were used to determine the primary target cells expressing the shared hub genes and key signaling pathways. RESULTS: COVID-19 may affect patients with SLE through hematologic involvement and exacerbated inflammatory responses. We identified 14 shared hub genes between COVID-19 and SLE that were significantly associated with interferon (IFN)-I/II. We also screened and obtained four core transcription factors related to these hub genes, confirming the regulatory role of the IFN-I/II-mediated Janus kinase/signal transducers and activators of transcription (JAK-STAT) signaling pathway on these hub genes. Further, SLE and COVID-19 can interact via IFN-I/II and IFN-I/II receptors, promoting the levels of monokines, including interleukin (IL)-6/10, tumor necrosis factor-α, and IFN-γ, and elevating the incidence rate and risk of cytokine release syndrome. Therefore, in SLE and COVID-19, both hub genes and core TFs are enriched within monocytes/macrophages. CONCLUSIONS: The interaction between SLE and COVID-19 promotes the activation of the IFN-I/II-triggered JAK-STAT signaling pathway in monocytes/macrophages. These findings provide a new direction and rationale for diagnosing and treating patients with SLE-COVID-19 comorbidity.


Assuntos
COVID-19 , Estudo de Associação Genômica Ampla , Lúpus Eritematoso Sistêmico , SARS-CoV-2 , Transdução de Sinais , Humanos , COVID-19/genética , Lúpus Eritematoso Sistêmico/genética , SARS-CoV-2/fisiologia , Feminino , Janus Quinases/metabolismo , Fatores de Transcrição STAT/metabolismo , Fatores de Transcrição STAT/genética , Masculino , Transcriptoma , Perfilação da Expressão Gênica , Multiômica
6.
Sci Rep ; 14(1): 13134, 2024 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-38849411

RESUMO

The pandemic of coronavirus disease 19 (COVID-19), caused by severe respiratory syndrome coronavirus 2 (SARS-CoV-2), had severe repercussions for breast cancer patients. Increasing evidence indicates that SARS-CoV-2 infection may directly impact breast cancer biology, but the effects of SARS-CoV-2 on breast tumor cells are still unknown. Here, we analyzed the molecular events occurring in the MCF7, MDA-MB-231 and HCC1937 breast cancer cell lines, representative of the luminal A, basal B/claudin-low and basal A subtypes, respectively, upon SARS-CoV-2 infection. Viral replication was monitored over time, and gene expression profiling was conducted. We found that MCF7 cells were the most permissive to viral replication. Treatment of MCF7 cells with Tamoxifen reduced the SARS-CoV-2 replication rate, suggesting an involvement of the estrogen receptor in sustaining virus replication in malignant cells. Interestingly, a metagene signature based on genes upregulated by SARS-CoV-2 infection in all three cell lines distinguished a subgroup of premenopausal luminal A breast cancer patients with a poor prognosis. As SARS-CoV-2 still spreads among the population, it is essential to understand the impact of SARS-CoV-2 infection on breast cancer, particularly in premenopausal patients diagnosed with the luminal A subtype, and to assess the long-term impact of COVID-19 on breast cancer outcomes.


Assuntos
Neoplasias da Mama , COVID-19 , SARS-CoV-2 , Tamoxifeno , Replicação Viral , Humanos , Neoplasias da Mama/virologia , Neoplasias da Mama/patologia , COVID-19/virologia , Feminino , SARS-CoV-2/fisiologia , Linhagem Celular Tumoral , Tamoxifeno/farmacologia , Células MCF-7 , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica
7.
Vet Res ; 55(1): 71, 2024 May 31.
Artigo em Inglês | MEDLINE | ID: mdl-38822398

RESUMO

In the wake of the COVID-19 pandemic caused by SARS-CoV-2, questions emerged about the potential effects of Bacillus Calmette-Guérin (BCG) vaccine on the immune response to SARS-CoV-2 infection, including the neurodegenerative diseases it may contribute to. To explore this, an experimental study was carried out in BCG-stimulated and non-stimulated k18-hACE2 mice challenged with SARS-CoV-2. Viral loads in tissues determined by RT-qPCR, histopathology in brain and lungs, immunohistochemical study in brain (IHC) as well as mortality rates, clinical signs and plasma inflammatory and coagulation biomarkers were assessed. Our results showed BCG-SARS-CoV-2 challenged mice presented higher viral loads in the brain and an increased frequency of neuroinvasion, with the greatest differences observed between groups at 3-4 days post-infection (dpi). Histopathological examination showed a higher severity of brain lesions in BCG-SARS-CoV-2 challenged mice, mainly consisting of neuroinflammation, increased glial cell population and neuronal degeneration, from 5 dpi onwards. This group also presented higher interstitial pneumonia and vascular thrombosis in lungs (3-4 dpi), BCG-SARS-CoV-2 mice showed higher values for TNF-α and D-dimer values, while iNOS values were higher in SARS-CoV-2 mice at 3-4 dpi. Results presented in this study indicate that BCG stimulation could have intensified the inflammatory and neurodegenerative lesions promoting virus neuroinvasion and dissemination in this experimental model. Although k18-hACE2 mice show higher hACE2 expression and neurodissemination, this study suggests that, although the benefits of BCG on enhancing heterologous protection against pathogens and tumour cells have been broadly demonstrated, potential adverse outcomes due to the non-specific effects of BCG should be considered.


Assuntos
Vacina BCG , Encéfalo , COVID-19 , SARS-CoV-2 , Animais , Camundongos , Vacina BCG/administração & dosagem , COVID-19/imunologia , COVID-19/virologia , SARS-CoV-2/fisiologia , Encéfalo/patologia , Encéfalo/virologia , Carga Viral , Pulmão/patologia , Pulmão/virologia , Pulmão/imunologia , Enzima de Conversão de Angiotensina 2/metabolismo , Camundongos Transgênicos , Feminino
8.
Int J Mol Sci ; 25(11)2024 Jun 04.
Artigo em Inglês | MEDLINE | ID: mdl-38892373

RESUMO

SARS-CoV-2 infection has been recently shown to induce cellular senescence in vivo. A senescence-like phenotype has been reported in cystic fibrosis (CF) cellular models. Since the previously published data highlighted a low impact of SARS-CoV-2 on CFTR-defective cells, here we aimed to investigate the senescence hallmarks in SARS-CoV-2 infection in the context of a loss of CFTR expression/function. We infected WT and CFTR KO 16HBE14o-cells with SARS-CoV-2 and analyzed both the p21 and Ki67 expression using immunohistochemistry and viral and p21 gene expression using real-time PCR. Prior to SARS-CoV-2 infection, CFTR KO cells displayed a higher p21 and lower Ki67 expression than WT cells. We detected lipid accumulation in CFTR KO cells, identified as lipolysosomes and residual bodies at the subcellular/ultrastructure level. After SARS-CoV-2 infection, the situation reversed, with low p21 and high Ki67 expression, as well as reduced viral gene expression in CFTR KO cells. Thus, the activation of cellular senescence pathways in CFTR-defective cells was reversed by SARS-CoV-2 infection while they were activated in CFTR WT cells. These data uncover a different response of CF and non-CF bronchial epithelial cell models to SARS-CoV-2 infection and contribute to uncovering the molecular mechanisms behind the reduced clinical impact of COVID-19 in CF patients.


Assuntos
Brônquios , COVID-19 , Senescência Celular , Inibidor de Quinase Dependente de Ciclina p21 , Regulador de Condutância Transmembrana em Fibrose Cística , Células Epiteliais , Antígeno Ki-67 , SARS-CoV-2 , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Regulador de Condutância Transmembrana em Fibrose Cística/metabolismo , Humanos , Senescência Celular/genética , SARS-CoV-2/fisiologia , COVID-19/virologia , COVID-19/metabolismo , COVID-19/patologia , Células Epiteliais/metabolismo , Células Epiteliais/virologia , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Inibidor de Quinase Dependente de Ciclina p21/genética , Antígeno Ki-67/metabolismo , Brônquios/virologia , Brônquios/metabolismo , Brônquios/patologia , Brônquios/citologia , Fibrose Cística/metabolismo , Fibrose Cística/genética , Fibrose Cística/virologia , Fibrose Cística/patologia , Linhagem Celular
9.
Immunity ; 57(6): 1195-1214, 2024 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-38865966

RESUMO

Long COVID (LC) is a condition in which patients do not fully recover from the initial SARS-CoV-2 infection but rather have persistent or new symptoms for months to years following the infection. Ongoing research efforts are investigating the pathophysiologic mechanisms of LC and exploring preventative and therapeutic treatment approaches for patients. As a burgeoning area of investigation, LC research can be structured to be more inclusive, innovative, and effective. In this perspective, we highlight opportunities for patient engagement and diverse research expertise, as well as the challenges of developing definitions and reproducible studies. Our intention is to provide a foundation for collaboration and progress in understanding the biomarkers and mechanisms driving LC.


Assuntos
COVID-19 , Síndrome de COVID-19 Pós-Aguda , SARS-CoV-2 , Humanos , Biomarcadores , Pesquisa Biomédica , COVID-19/imunologia , COVID-19/virologia , SARS-CoV-2/fisiologia , SARS-CoV-2/imunologia
10.
Nat Commun ; 15(1): 5112, 2024 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-38879641

RESUMO

Virus infectivity is traditionally determined by endpoint titration in cell cultures, and requires complex processing steps and human annotation. Here we developed an artificial intelligence (AI)-powered automated framework for ready detection of virus-induced cytopathic effect (DVICE). DVICE uses the convolutional neural network EfficientNet-B0 and transmitted light microscopy images of infected cell cultures, including coronavirus, influenza virus, rhinovirus, herpes simplex virus, vaccinia virus, and adenovirus. DVICE robustly measures virus-induced cytopathic effects (CPE), as shown by class activation mapping. Leave-one-out cross-validation in different cell types demonstrates high accuracy for different viruses, including SARS-CoV-2 in human saliva. Strikingly, DVICE exhibits virus class specificity, as shown with adenovirus, herpesvirus, rhinovirus, vaccinia virus, and SARS-CoV-2. In sum, DVICE provides unbiased infectivity scores of infectious agents causing CPE, and can be adapted to laboratory diagnostics, drug screening, serum neutralization or clinical samples.


Assuntos
Inteligência Artificial , Efeito Citopatogênico Viral , Microscopia , SARS-CoV-2 , Humanos , SARS-CoV-2/patogenicidade , SARS-CoV-2/fisiologia , Microscopia/métodos , COVID-19/virologia , Redes Neurais de Computação , Animais , Vaccinia virus/fisiologia , Vaccinia virus/patogenicidade , Saliva/virologia , Chlorocebus aethiops , Células Vero , Rhinovirus/patogenicidade , Rhinovirus/fisiologia , Linhagem Celular
11.
Front Immunol ; 15: 1368946, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38881905

RESUMO

Background: In early infected or severe coronavirus disease 2019 (COVID-19) patients, circulating NK cells are consistently reduced, despite being highly activated or exhausted. The aim of this paper was to establish whether severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike glycoprotein (SP) may directly trigger NK cells and through which receptor(s). Methods: SP-stimulated human NK cells have been evaluated for the expression of activation markers, cytokine release, and cytotoxic activity, as well as for gene expression profiles and NF-kB phosphorylation, and they have been silenced with specific small interfering RNAs. Results: SPs from the Wuhan strain and other variants of concern (VOCs) directly bind and stimulate purified NK cells by increasing activation marker expression, cytokine release, and cytolytic activity, prevalently in the CD56brightNK cell subset. VOC-SPs differ in their ability to activate NK cells, G614, and Delta-Plus strains providing the strongest activity in the majority of donors. While VOC-SPs do not trigger ACE2, which is not expressed on NK cells, or other activating receptors, they directly and variably bind to both Toll-like receptor 2 (TLR2) and TLR4. Moreover, SP-driven NK cell functions are inhibited upon masking such receptors or silencing the relative genes. Lastly, VOC-SPs upregulate CD56dimNK cell functions in COVID-19 recovered, but not in non-infected, individuals. Conclusions: TLR2 and TLR4 are novel activating receptors for SP in NK cells, suggesting a new role of these cells in orchestrating the pathophysiology of SARS-CoV-2 infection. The pathogenic relevance of this finding is highlighted by the fact that free SP providing NK cell activation is frequently detected in a SARS-CoV-2 inflamed environment and in plasma of infected and long-COVID-19 subjects.


Assuntos
COVID-19 , Células Matadoras Naturais , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Receptor 2 Toll-Like , Receptor 4 Toll-Like , Humanos , Células Matadoras Naturais/imunologia , Células Matadoras Naturais/metabolismo , Glicoproteína da Espícula de Coronavírus/imunologia , Glicoproteína da Espícula de Coronavírus/metabolismo , SARS-CoV-2/imunologia , SARS-CoV-2/fisiologia , COVID-19/imunologia , COVID-19/virologia , Receptor 4 Toll-Like/metabolismo , Receptor 4 Toll-Like/imunologia , Receptor 2 Toll-Like/metabolismo , Receptor 2 Toll-Like/imunologia , Ativação Linfocitária/imunologia , Citocinas/metabolismo , Citocinas/imunologia , Enzima de Conversão de Angiotensina 2/metabolismo , Enzima de Conversão de Angiotensina 2/genética , Enzima de Conversão de Angiotensina 2/imunologia
12.
EBioMedicine ; 104: 105181, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38838469

RESUMO

BACKGROUND: Although several SARS-CoV-2-related coronaviruses (SC2r-CoVs) were discovered in bats and pangolins, the differences in virological characteristics between SARS-CoV-2 and SC2r-CoVs remain poorly understood. Recently, BANAL-20-236 (B236) was isolated from a rectal swab of Malayan horseshoe bat and was found to lack a furin cleavage site (FCS) in the spike (S) protein. The comparison of its virological characteristics with FCS-deleted SARS-CoV-2 (SC2ΔFCS) has not been conducted yet. METHODS: We prepared human induced pluripotent stem cell (iPSC)-derived airway and lung epithelial cells and colon organoids as human organ-relevant models. B236, SARS-CoV-2, and artificially generated SC2ΔFCS were used for viral experiments. To investigate the pathogenicity of B236 in vivo, we conducted intranasal infection experiments in hamsters. FINDINGS: In human iPSC-derived airway epithelial cells, the growth of B236 was significantly lower than that of the SC2ΔFCS. A fusion assay showed that the B236 and SC2ΔFCS S proteins were less fusogenic than the SARS-CoV-2 S protein. The infection experiment in hamsters showed that B236 was less pathogenic than SARS-CoV-2 and even SC2ΔFCS. Interestingly, in human colon organoids, the growth of B236 was significantly greater than that of SARS-CoV-2. INTERPRETATION: Compared to SARS-CoV-2, we demonstrated that B236 exhibited a tropism toward intestinal cells rather than respiratory cells. Our results are consistent with a previous report showing that B236 is enterotropic in macaques. Altogether, our report strengthens the assumption that SC2r-CoVs in horseshoe bats replicate primarily in the intestinal tissues rather than respiratory tissues. FUNDING: This study was supported in part by AMED ASPIRE (JP23jf0126002, to Keita Matsuno, Kazuo Takayama, and Kei Sato); AMED SCARDA Japan Initiative for World-leading Vaccine Research and Development Centers "UTOPIA" (JP223fa627001, to Kei Sato), AMED SCARDA Program on R&D of new generation vaccine including new modality application (JP223fa727002, to Kei Sato); AMED SCARDA Hokkaido University Institute for Vaccine Research and Development (HU-IVReD) (JP223fa627005h0001, to Takasuke Fukuhara, and Keita Matsuno); AMED Research Program on Emerging and Re-emerging Infectious Diseases (JP21fk0108574, to Hesham Nasser; JP21fk0108493, to Takasuke Fukuhara; JP22fk0108617 to Takasuke Fukuhara; JP22fk0108146, to Kei Sato; JP21fk0108494 to G2P-Japan Consortium, Keita Matsuno, Shinya Tanaka, Terumasa Ikeda, Takasuke Fukuhara, and Kei Sato; JP21fk0108425, to Kazuo Takayama and Kei Sato; JP21fk0108432, to Kazuo Takayama, Takasuke Fukuhara and Kei Sato; JP22fk0108534, Terumasa Ikeda, and Kei Sato; JP22fk0108511, to Yuki Yamamoto, Terumasa Ikeda, Keita Matsuno, Shinya Tanaka, Kazuo Takayama, Takasuke Fukuhara, and Kei Sato; JP22fk0108506, to Kazuo Takayama and Kei Sato); AMED Research Program on HIV/AIDS (JP22fk0410055, to Terumasa Ikeda; and JP22fk0410039, to Kei Sato); AMED Japan Program for Infectious Diseases Research and Infrastructure (JP22wm0125008 to Keita Matsuno); AMED CREST (JP21gm1610005, to Kazuo Takayama; JP22gm1610008, to Takasuke Fukuhara; JST PRESTO (JPMJPR22R1, to Jumpei Ito); JST CREST (JPMJCR20H4, to Kei Sato); JSPS KAKENHI Fund for the Promotion of Joint International Research (International Leading Research) (JP23K20041, to G2P-Japan Consortium, Keita Matsuno, Takasuke Fukuhara and Kei Sato); JST SPRING (JPMJSP2108 to Shigeru Fujita); JSPS KAKENHI Grant-in-Aid for Scientific Research C (22K07103, to Terumasa Ikeda); JSPS KAKENHI Grant-in-Aid for Scientific Research B (21H02736, to Takasuke Fukuhara); JSPS KAKENHI Grant-in-Aid for Early-Career Scientists (22K16375, to Hesham Nasser; 20K15767, to Jumpei Ito); JSPS Core-to-Core Program (A. Advanced Research Networks) (JPJSCCA20190008, to Kei Sato); JSPS Research Fellow DC2 (22J11578, to Keiya Uriu); JSPS Research Fellow DC1 (23KJ0710, to Yusuke Kosugi); JSPS Leading Initiative for Excellent Young Researchers (LEADER) (to Terumasa Ikeda); World-leading Innovative and Smart Education (WISE) Program 1801 from the Ministry of Education, Culture, Sports, Science and Technology (MEXT) (to Naganori Nao); Ministry of Health, Labour and Welfare (MHLW) under grant 23HA2010 (to Naganori Nao and Keita Matsuno); The Cooperative Research Program (Joint Usage/Research Center program) of Institute for Life and Medical Sciences, Kyoto University (to Kei Sato); International Joint Research Project of the Institute of Medical Science, the University of Tokyo (to Terumasa Ikeda and Takasuke Fukuhara); The Tokyo Biochemical Research Foundation (to Kei Sato); Takeda Science Foundation (to Terumasa Ikeda and Takasuke Fukuhara); Mochida Memorial Foundation for Medical and Pharmaceutical Research (to Terumasa Ikeda); The Naito Foundation (to Terumasa Ikeda); Hokuto Foundation for Bioscience (to Tomokazu Tamura); Hirose Foundation (to Tomokazu Tamura); and Mitsubishi Foundation (to Kei Sato).


Assuntos
COVID-19 , Quirópteros , SARS-CoV-2 , Animais , SARS-CoV-2/genética , SARS-CoV-2/fisiologia , Humanos , COVID-19/virologia , Quirópteros/virologia , Glicoproteína da Espícula de Coronavírus/metabolismo , Glicoproteína da Espícula de Coronavírus/genética , Organoides/virologia , Organoides/metabolismo , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/virologia , Cricetinae , Furina/metabolismo , Células Epiteliais/virologia , Células Vero , Chlorocebus aethiops
13.
Front Immunol ; 15: 1398369, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38835759

RESUMO

Introduction: Although many studies have underscored the importance of T cells, phenotypically and functionally, fewer have studied the functions of myeloid cells in COVID disease. In particular, the potential role of myeloid cells such as monocytes and low-density neutrophils (LDNs) in innate responses and particular in the defense against secondary bacterial infections has been much less documented. Methods: Here, we compared, in a longitudinal study, healthy subjects, idiopathic fibrosis patients, COVID patients who were either hospitalized/moderate (M-) or admitted to ICU (COV-ICU) and patients in ICU hospitalized for other reasons (non-COV-ICU). Results: We show that COVID patients have an increased proportion of low-density neutrophils (LDNs), which produce high levels of proteases (particularly, NE, MMP-8 and MMP-9) (unlike non-COV-ICU patients), which are partly responsible for causing type II alveolar cell damage in co-culture experiments. In addition, we showed that M- and ICU-COVID monocytes had reduced responsiveness towards further live Pseudomonas aeruginosa (PAO1 strain) infection, an important pathogen colonizing COVID patients in ICU, as assessed by an impaired secretion of myeloid cytokines (IL-1, TNF, IL-8,…). By contrast, lymphoid cytokines (in particular type 2/type 3) levels remained high, both basally and post PAO1 infection, as reflected by the unimpaired capacity of T cells to proliferate, when stimulated with anti-CD3/CD28 beads. Discussion: Overall, our results demonstrate that COVID circulatory T cells have a biased type 2/3 phenotype, unconducive to proper anti-viral responses and that myeloid cells have a dual deleterious phenotype, through their LDN-mediated damaging effect on alveolar cells and their impaired responsiveness (monocyte-mediated) towards bacterial pathogens such as P. aeruginosa.


Assuntos
COVID-19 , Monócitos , Neutrófilos , Infecções por Pseudomonas , Pseudomonas aeruginosa , SARS-CoV-2 , Humanos , COVID-19/imunologia , Pseudomonas aeruginosa/imunologia , Monócitos/imunologia , Masculino , Feminino , Pessoa de Meia-Idade , SARS-CoV-2/imunologia , SARS-CoV-2/fisiologia , Infecções por Pseudomonas/imunologia , Neutrófilos/imunologia , Idoso , Citocinas/metabolismo , Citocinas/imunologia , Adulto , Estudos Longitudinais , Leucócitos Mononucleares/imunologia , Pulmão/imunologia , Pulmão/patologia , Pulmão/microbiologia
14.
Sci Adv ; 10(23): eadj4735, 2024 Jun 07.
Artigo em Inglês | MEDLINE | ID: mdl-38838150

RESUMO

Why individuals with Down syndrome (DS) are more susceptible to SARS-CoV-2-induced neuropathology remains elusive. Choroid plexus (ChP) plays critical roles in barrier function and immune response modulation and expresses the ACE2 receptor and the chromosome 21-encoded TMPRSS2 protease, suggesting its substantial role in establishing SARS-CoV-2 infection in the brain. To explore this, we established brain organoids from DS and isogenic euploid iPSC that consist of a core of functional cortical neurons surrounded by a functional ChP-like epithelium (ChPCOs). DS-ChPCOs recapitulated abnormal DS cortical development and revealed defects in ciliogenesis and epithelial cell polarity in ChP-like epithelium. We then demonstrated that the ChP-like epithelium facilitates infection and replication of SARS-CoV-2 in cortical neurons and that this is increased in DS. Inhibiting TMPRSS2 and furin activity reduced viral replication in DS-ChPCOs to euploid levels. This model enables dissection of the role of ChP in neurotropic virus infection and euploid forebrain development and permits screening of therapeutics for SARS-CoV-2-induced neuropathogenesis.


Assuntos
Encéfalo , COVID-19 , Plexo Corióideo , Síndrome de Down , Organoides , SARS-CoV-2 , Serina Endopeptidases , Plexo Corióideo/virologia , Plexo Corióideo/metabolismo , Plexo Corióideo/patologia , Organoides/virologia , Organoides/metabolismo , Organoides/patologia , Humanos , SARS-CoV-2/fisiologia , COVID-19/virologia , COVID-19/patologia , COVID-19/metabolismo , Serina Endopeptidases/metabolismo , Serina Endopeptidases/genética , Síndrome de Down/metabolismo , Síndrome de Down/patologia , Síndrome de Down/genética , Encéfalo/virologia , Encéfalo/patologia , Encéfalo/metabolismo , Neurônios/metabolismo , Neurônios/virologia , Neurônios/patologia , Replicação Viral , Células-Tronco Pluripotentes Induzidas/metabolismo , Células-Tronco Pluripotentes Induzidas/virologia , Furina/metabolismo , Furina/genética , Enzima de Conversão de Angiotensina 2/metabolismo , Enzima de Conversão de Angiotensina 2/genética , Tropismo Viral
15.
Front Immunol ; 15: 1381059, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38855100

RESUMO

Background: Understanding COVID-19 outcomes remains a challenge. While numerous biomarkers have been proposed for severity at admission, limited exploration exists for markers during the infection course, especially for the requirement of oxygen therapy. This study investigates the potential of eosinophil count normalization as a predictor for oxygen weaning during the initial wave of the pandemic. Methods: A retrospective study was conducted between March and April 2020 (first wave) among adults admitted directly to a medicine ward. Biological abnormalities, including lymphocyte count, eosinophil count, and C-reactive protein (CRP), were gathered daily during the first week of admission according to oxygen level. In case of worsening, oxygen level was censored at 15 L/min. The primary aim was to assess whether eosinophil count normalization predicts a subsequent decrease in oxygen requirements. Results: Overall, 132 patients were admitted, with a mean age of 59.0 ± 16.3 years. Of the patients, 72% required oxygen, and 20.5% were admitted to the intensive care unit after a median delay of 48 hours. The median CRP at admission was 79 (26-130) mg/L, whereas the eosinophil count was 10 (0-60)/mm3. Eosinophil count normalization (≥100/mm3) by day 2 correlated significantly with decreased oxygen needs (<2 L) with hazard ratio (HR) = 3.7 [1.1-12.9] (p = 0.04). Likewise, CRP < 80 mg/L was associated with reduced oxygen requirements (p < 0.001). Predictors, including underlying chronic respiratory disease, exhibited a trend toward a negative association (p = 0.06). Conclusion: The study highlights the relationship between eosinophil count and CRP, with implications for predicting oxygen weaning during COVID-19. Further research is warranted to explore the relevance of these biomarkers in other respiratory infections.


Assuntos
COVID-19 , Eosinófilos , Oxigenoterapia , SARS-CoV-2 , Humanos , COVID-19/sangue , COVID-19/imunologia , COVID-19/terapia , COVID-19/epidemiologia , Masculino , Pessoa de Meia-Idade , Feminino , Estudos Retrospectivos , Eosinófilos/imunologia , Idoso , Contagem de Leucócitos , SARS-CoV-2/fisiologia , Adulto , Hospitalização , Biomarcadores/sangue , Proteína C-Reativa/análise , Proteína C-Reativa/metabolismo , Oxigênio/metabolismo , Oxigênio/sangue
16.
Front Immunol ; 15: 1348229, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38855114

RESUMO

Introduction: The COVID-19 pandemic represented one of the most significant challenges to researchers and healthcare providers. Several factors determine the disease severity, whereas none alone can explain the tremendous variability. The Single nucleotide variants (SNVs) in angiotensin-converting enzyme-2 (ACE2) and transmembrane serine protease type-2 (TMPRSS2) genes affect the virus entry and are considered possible risk factors for COVID-19. Methods: We compiled a panel of gene variants from both genes and used in-silico analysis to predict their significance. We performed biological validation to assess their capacity to alter the ACE2 interaction with the virus spike protein. Subsequently, we conducted a retrospective comparative genome analysis on those variants in the Emirati patients with different disease severity (total of 96) along with 69 healthy control subjects. Results: Our results showed that the Emirati population lacks the variants that were previously reported as associated with disease severity, whereas a new variant in ACE2 "Chr X:g.15584534" was associated with disease severity specifically among female patients. In-silico analysis revealed that the new variant can determine the ACE2 gene transcription. Several cytokines (GM-CSF and IL-6) and chemokines (MCP-1/CCL2, IL-8/CXCL8, and IP-10/CXCL10) were markedly increased in COVID-19 patients with a significant correlation with disease severity. The newly reported genetic variant of ACE2 showed a positive correlation with CD40L, IL-1ß, IL-2, IL-15, and IL-17A in COVID-19 patients. Conclusion: Whereas COVID-19 represents now a past pandemic, our study underscores the importance of genetic factors specific to a population, which can influence both the susceptibility to viral infections and the level of severity; subsequently expected required preparedness in different areas of the world.


Assuntos
Enzima de Conversão de Angiotensina 2 , COVID-19 , Citocinas , Polimorfismo de Nucleotídeo Único , SARS-CoV-2 , Serina Endopeptidases , Humanos , COVID-19/genética , Enzima de Conversão de Angiotensina 2/genética , Feminino , Masculino , SARS-CoV-2/fisiologia , Citocinas/sangue , Citocinas/genética , Serina Endopeptidases/genética , Emirados Árabes Unidos/epidemiologia , Pessoa de Meia-Idade , Adulto , Estudos Retrospectivos , Índice de Gravidade de Doença , Idoso
17.
Sci Rep ; 14(1): 12406, 2024 05 30.
Artigo em Inglês | MEDLINE | ID: mdl-38811809

RESUMO

Lung adenocarcinoma (LUAD) is the most common and aggressive subtype of lung cancer, and coronavirus disease 2019 (COVID-19) has become a serious public health threat worldwide. Patients with LUAD and COVID-19 have a poor prognosis. Therefore, finding medications that can be used to treat COVID-19/LUAD patients is essential. Bioinformatics analysis was used to identify 20 possible metformin target genes for the treatment of COVID-19/LUAD. PTEN and mTOR may serve as hub target genes of metformin. Metformin may be able to cure COVID-19/LUAD comorbidity through energy metabolism, oxidoreductase NADH activity, FoxO signalling pathway, AMPK signalling system, and mTOR signalling pathway, among other pathways, according to the results of bioinformatic research. Metformin has ability to inhibit the proliferation of A549 cells, according to the results of colony formation and proliferation assays. In A549 cells, metformin increased glucose uptake and lactate generation, while decreasing ATP synthesis and the NAD+/NADH ratio. In summary, PTEN and mTOR may be potential targets of metformin for the treatment of COVID-19/LUAD. The mechanism by which metformin inhibits lung adenocarcinoma cell proliferation may be related to glucose metabolism regulated by PI3K/AKT signalling and mTOR signalling pathways. Our study provides a new theoretical basis for the treatment of COVID-19/LUAD.


Assuntos
Adenocarcinoma de Pulmão , Tratamento Farmacológico da COVID-19 , COVID-19 , Proliferação de Células , Glucose , Neoplasias Pulmonares , Metformina , PTEN Fosfo-Hidrolase , Transdução de Sinais , Serina-Treonina Quinases TOR , Metformina/farmacologia , Metformina/uso terapêutico , Humanos , Células A549 , Glucose/metabolismo , Serina-Treonina Quinases TOR/metabolismo , COVID-19/metabolismo , COVID-19/virologia , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/metabolismo , Proliferação de Células/efeitos dos fármacos , PTEN Fosfo-Hidrolase/metabolismo , Adenocarcinoma de Pulmão/tratamento farmacológico , Adenocarcinoma de Pulmão/metabolismo , Adenocarcinoma de Pulmão/patologia , Transdução de Sinais/efeitos dos fármacos , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/fisiologia , Metabolismo Energético/efeitos dos fármacos
18.
Commun Biol ; 7(1): 526, 2024 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-38702425

RESUMO

COVID-19, caused by SARS-CoV-2, can lead to a severe inflammatory disease characterized by significant lymphopenia. However, the underlying cause for the depletion of T-cells in COVID-19 patients remains incompletely understood. In this study, we assessed the presence of different T-cell subsets in the progression of COVID-19 from mild to severe disease, with a focus on TCF1 expressing progenitor T-cells that are needed to replenish peripheral T-cells during infection. Our results showed a preferential decline in TCF1+ progenitor CD4 and CD8+ T-cells with disease severity. This decline was seen in various TCF1+ subsets including naive, memory and effector-memory cells, and surprisingly, was accompanied by a loss in cell division as seen by a marked decline in Ki67 expression. In addition, TCF1+ T-cells showed a reduction in pro-survival regulator, BcL2, and the appearance of a new population of TCF1 negative caspase-3 expressing cells in peripheral blood from patients with severe disease. The decline in TCF1+ T-cells was also seen in a subgroup of severe patients with vitamin D deficiency. Lastly, we found that sera from severe patients inhibited TCF1 transcription ex vivo which was attenuated by a blocking antibody against the cytokine, interleukin-12 (IL12). Collectively, our findings underscore the potential significance of TCF1+ progenitor T-cells in accounting for the loss of immunity in severe COVID-19 and outline an array of markers that could be used to identify disease progression.


Assuntos
COVID-19 , Fator 1-alfa Nuclear de Hepatócito , SARS-CoV-2 , Índice de Gravidade de Doença , Humanos , COVID-19/imunologia , COVID-19/patologia , Masculino , Feminino , SARS-CoV-2/imunologia , SARS-CoV-2/fisiologia , Fator 1-alfa Nuclear de Hepatócito/metabolismo , Fator 1-alfa Nuclear de Hepatócito/genética , Pessoa de Meia-Idade , Linfócitos T CD8-Positivos/imunologia , Adulto , Linfócitos T CD4-Positivos/imunologia , Idoso , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo
19.
Sci Total Environ ; 931: 172945, 2024 Jun 25.
Artigo em Inglês | MEDLINE | ID: mdl-38703849

RESUMO

The coagulation process has a high potential as a treatment method that can handle pathogenic viruses including emerging enveloped viruses in drinking water treatment process which can lower infection risk through drinking water consumption. In this study, a surrogate enveloped virus, bacteriophage Փ6, and surrogate non-enveloped viruses, including bacteriophage MS-2, T4, ՓX174, were used to evaluate removal efficiencies and mechanisms by the conventional coagulation process with alum, poly­aluminum chloride, and ferric chloride at pH 5, 7, and 9 in turbid water. Also, treatability of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a recent virus of global concern by coagulation was evaluated as SARS-CoV-2 can presence in drinking water sources. It was observed that an increase in the coagulant dose enhanced the removal efficiency of turbidity and viruses, and the condition that provided the highest removal efficiency of enveloped and non-enveloped viruses was 50 mg/L of coagulants at pH 5. In addition, the coagulation process was more effective for enveloped virus removal than for the non-enveloped viruses, and it demonstrated reduction of SARS-CoV-2 Omicron BA.2 over 0.83-log with alum. According to culture- and molecular-based assays (qPCR and CDDP-qPCR), the virus removal mechanisms were floc adsorption and coagulant inactivation. Through inactivation with coagulants, coagulants caused capsid destruction, followed by genome damage in non-enveloped viruses; however, damage to a lipid envelope is suggested to contribute to a great extend for enveloped virus inactivation. We demonstrated that conventional coagulation is a promising method for controlling emerging and re-emerging viruses in drinking water.


Assuntos
SARS-CoV-2 , Purificação da Água , Purificação da Água/métodos , SARS-CoV-2/fisiologia , COVID-19 , Água Potável/virologia , Água Potável/química , Compostos de Alúmen , Microbiologia da Água , Betacoronavirus/fisiologia , Floculação , Compostos de Alumínio , Compostos Férricos/química
20.
Front Immunol ; 15: 1380697, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38715608

RESUMO

The Corona Virus Disease (COVID-19), caused by the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2), has quickly spread worldwide and resulted in significant morbidity and mortality. Although most infections are mild, some patients can also develop severe and fatal myocarditis. In eukaryotic RNAs, 5-methylcytosine (m5C) is a common kind of post-transcriptional modification, which is involved in regulating various biological processes (such as RNA export, translation, and stability maintenance). With the rapid development of m5C modification detection technology, studies related to viral m5C modification are ever-increasing. These studies have revealed that m5C modification plays an important role in various stages of viral replication, including transcription and translation. According to recent studies, m5C methylation modification can regulate SARS-CoV-2 infection by modulating innate immune signaling pathways. However, the specific role of m5C modification in SARS-CoV-2-induced myocarditis remains unclear. Therefore, this review aims to provide insights into the molecular mechanisms of m5C methylation in SARS-CoV-2 infection. Moreover, the regulatory role of NSUN2 in viral infection and host innate immune response was also highlighted. This review may provide new directions for developing therapeutic strategies for SARS-CoV-2-associated myocarditis.


Assuntos
COVID-19 , Miocardite , SARS-CoV-2 , Miocardite/virologia , Miocardite/imunologia , Miocardite/terapia , Miocardite/genética , Humanos , COVID-19/imunologia , COVID-19/genética , COVID-19/terapia , SARS-CoV-2/fisiologia , Metilação , 5-Metilcitosina/metabolismo , Imunidade Inata , Tratamento Farmacológico da COVID-19 , Animais , RNA Viral/genética , RNA Viral/metabolismo , Processamento Pós-Transcricional do RNA
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