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2.
Cell Discov ; 7(1): 13, 2021 Mar 09.
Artigo em Inglês | MEDLINE | ID: mdl-33750767

RESUMO

The current dogma in ophthalmology and vision research presumes the intraocular environment to be sterile. However, recent evidence of intestinal bacterial translocation into the bloodstream and many other internal organs including the eyes, found in healthy and diseased animal models, suggests that the intraocular cavity may also be inhabited by a microbial community. Here, we tested intraocular samples from over 1000 human eyes. Using quantitative PCR, negative staining transmission electron microscopy, direct culture, and high-throughput sequencing technologies, we demonstrated the presence of intraocular bacteria. The possibility that the microbiome from these low-biomass communities could be a contamination from other tissues and reagents was carefully evaluated and excluded. We also provide preliminary evidence that a disease-specific microbial signature characterized the intraocular environment of patients with age-related macular degeneration and glaucoma, suggesting that either spontaneous or pathogenic bacterial translocation may be associated with these common sight-threatening conditions. Furthermore, we revealed the presence of an intraocular microbiome in normal eyes from non-human mammals and demonstrated that this varied across species (rat, rabbit, pig, and macaque) and was established after birth. These findings represent the first-ever evidence of intraocular microbiota in humans.

3.
Cell ; 184(7): 1895-1913.e19, 2021 04 01.
Artigo em Inglês | MEDLINE | ID: mdl-33657410

RESUMO

A dysfunctional immune response in coronavirus disease 2019 (COVID-19) patients is a recurrent theme impacting symptoms and mortality, yet a detailed understanding of pertinent immune cells is not complete. We applied single-cell RNA sequencing to 284 samples from 196 COVID-19 patients and controls and created a comprehensive immune landscape with 1.46 million cells. The large dataset enabled us to identify that different peripheral immune subtype changes are associated with distinct clinical features, including age, sex, severity, and disease stages of COVID-19. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA was found in diverse epithelial and immune cell types, accompanied by dramatic transcriptomic changes within virus-positive cells. Systemic upregulation of S100A8/A9, mainly by megakaryocytes and monocytes in the peripheral blood, may contribute to the cytokine storms frequently observed in severe patients. Our data provide a rich resource for understanding the pathogenesis of and developing effective therapeutic strategies for COVID-19.


Assuntos
COVID-19/imunologia , Megacariócitos/imunologia , Monócitos/imunologia , RNA Viral , SARS-CoV-2/genética , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Criança , China , Estudos de Coortes , Citocinas/metabolismo , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , RNA Viral/sangue , RNA Viral/isolamento & purificação , Análise de Célula Única , Transcriptoma/imunologia , Adulto Jovem
5.
Invest Ophthalmol Vis Sci ; 61(2): 47, 2020 02 07.
Artigo em Inglês | MEDLINE | ID: mdl-32106294

RESUMO

Purpose: Microbial ecosystems interact with the human body and affect human health. The microbial community on the ocular surface remains an underexplored territory despite its importance as the first line of defense barrier that protects the eye and ultimately sight. We investigated how age and sex affected human ocular surface microbiome, and in the present study wanted to understand how geographic difference shaped the microbiome in the ocular surface. Methods: We collected conjunctival specimens of 172 eyes from 86 healthy volunteers living in three Chinese cities, namely, Guangzhou, Wenzhou, and Beijing. Using the direct metagenomic shotgun sequencing approach, we characterized how geographic difference affected the human ocular microbiome. Results: We surveyed the taxonomic composition and metabolic function of the microbiota on human ocular surface. We showed that the ocular surface microbiota was composed of bacteria, viruses, and fungi. A geographical difference in both composition and function of the conjunctival microbiome suggests that the environment people lived in shapes their conjunctival microbiome, especially the dominate species. Conclusions: Our study provides a reference catalog of the healthy conjunctival metagenome and raises a concern for environmental influences on the ocular surface microbiome.


Assuntos
Túnica Conjuntiva/microbiologia , Microbiota , Fatores Etários , China , DNA Bacteriano/análise , DNA Fúngico/análise , DNA Viral/análise , Feminino , Humanos , Masculino , Metagenoma , Fatores Sexuais
6.
Bioinformatics ; 36(5): 1577-1583, 2020 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-31626280

RESUMO

MOTIVATION: Microbiome analyses of clinical samples with low microbial biomass are challenging because of the very small quantities of microbial DNA relative to the human host, ubiquitous contaminating DNA in sequencing experiments and the large and rapidly growing microbial reference databases. RESULTS: We present computational subtraction-based microbiome discovery (CSMD), a bioinformatics pipeline specifically developed to generate accurate species-level microbiome profiles for clinical samples with low microbial loads. CSMD applies strategies for the maximal elimination of host sequences with minimal loss of microbial signal and effectively detects microorganisms present in the sample with minimal false positives using a stepwise convergent solution. CSMD was benchmarked in a comparative evaluation with other classic tools on previously published well-characterized datasets. It showed higher sensitivity and specificity in host sequence removal and higher specificity in microbial identification, which led to more accurate abundance estimation. All these features are integrated into a free and easy-to-use tool. Additionally, CSMD applied to cell-free plasma DNA showed that microbial diversity within these samples is substantially broader than previously believed. AVAILABILITY AND IMPLEMENTATION: CSMD is freely available at https://github.com/liuyu8721/csmd. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.


Assuntos
Metagenoma , Microbiota , Biologia Computacional , Humanos , Metagenômica , Software
7.
Curr Mol Med ; 19(6): 434-442, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31288713

RESUMO

BACKGROUND: Age-related macular degeneration (AMD) is a progressive and irreversible eye disease. The anti-vascular endothelial growth factor (VEGF) therapy has revolutionized the treatment of neovascular AMD. However, the expense for such treatment is quite high. METHODS: We used a traditional Chinese medicine ZQMT as an alternative therapeutic regimen for AMD. We employed two in vivo animal models mimicking dry and wet AMD respectively to assess the therapeutic efficacy of ZQMT on treating AMD-related retinopathy. AMD-related retinopathy in Crb1rd8 mice was evaluated from week 1 to 8 by fundus photography. Laser-induced choroidal neovascularization (CNV) was evaluated by fluorescein angiography and histopathology. RESULTS: ZQMT increased CX3CR1 expression in murine CD4+ T cells either cultured in vitro or directly isolated from animals treated with ZQMT. We also performed both in vitro and in vivo studies to confirm that ZQMT has no apparent toxic effects. ZQMT alleviated AMD-related retinopathy in both Crb1rd8 and CNV models. Depletion of CCL2 and CX3CR1 in Crb1rd8 mice abolished the efficacy of ZQMT, suggesting that CCL2 and/or CX3CR1 may underlie the mechanisms of ZQMT in treating AMD-related retinopathy in mice. CONCLUSION: In summary, our study supports the protective roles of a traditional Chinese medicine ZQMT in AMD.


Assuntos
Degeneração Macular/tratamento farmacológico , Medicina Tradicional Chinesa , Animais , Apoptose/efeitos dos fármacos , Biomarcadores , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Modelos Animais de Doenças , Expressão Gênica , Imunofenotipagem , Degeneração Macular/diagnóstico , Degeneração Macular/etiologia , Degeneração Macular/metabolismo , Camundongos , Camundongos Transgênicos , Índice de Gravidade de Doença
8.
J Gene Med ; 20(2-3): e3007, 2018 02.
Artigo em Inglês | MEDLINE | ID: mdl-29323771

RESUMO

BACKGROUND: Polypoidal choroidal vasculopathy (PCV) and neovascular age-related macular degeneration (nAMD) share a similar phenotype but are different in their clinical manifestations, responses to treatment and prognosis. Whether PCV is a subtype of AMD or a distinct entity from nAMD remains unknown. Therefore, we performed a whole-exome sequencing based association analysis to compare the genetic architecture of PCV and nAMD in Han Chinese. METHODS: Whole-exome sequencing analysis was performed on 21 nAMD cases, 20 PCV cases and 20 healthy controls. As a follow-up validation, 145 nAMD cases, 160 PCV cases and 193 controls were genotyped using the Sequenom MassARRAY platform (Sequenom, San Diego, CA, USA). RESULTS: A novel variant, c.6196A>G in the IGFN1 gene, was significantly associated with only PCV (combined p = 7.1 × 10-11 , odds ratio = 9.44), but not with nAMD (combined p = 0.683, odds ratio = 1.30). The minor allele G conferred an increased risk of PCV. CONCLUSIONS: The findings of the present study indicate that, although some of the susceptibility loci are shared between PCV and nAMD, a unique genetic signature may decide the pathogenesis of PCV.


Assuntos
Proteínas de Transporte/genética , Neovascularização de Coroide/genética , Predisposição Genética para Doença , Degeneração Macular/genética , Proteínas Adaptadoras de Transdução de Sinal , Idoso , Neovascularização de Coroide/patologia , Feminino , Estudos de Associação Genética , Genótipo , Humanos , Degeneração Macular/patologia , Masculino , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único , Sequenciamento Completo do Exoma
9.
J Cell Mol Med ; 22(3): 1826-1839, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-29193684

RESUMO

Although antiviral drugs are available for the treatment of influenza infection, it is an urgent requirement to develop new antiviral drugs regarding the emergence of drug-resistant viruses. The nucleoprotein (NP) is conserved among all influenza A viruses (IAVs) and has no cellular equivalent. Therefore, NP is an ideal target for the development of new IAV inhibitors. In this study, we identified a novel anti-influenza compound, ZBMD-1, from a library of 20,000 compounds using cell-based influenza A infection assays. We found that ZBMD-1 inhibited the replication of H1N1 and H3N2 influenza A virus strains in vitro, with an IC50 ranging from 0.41-1.14 µM. Furthermore, ZBMD-1 inhibited the polymerase activity and specifically impaired the nuclear export of NP. Further investigation indicated that ZBMD-1 binds to the nuclear export signal 3 (NES3) domain and the dimer interface of the NP pocket. ZBMD-1 also protected mice that were challenged with lethal doses of A/PR/8/1934 (H1N1) virus, effectively relieving lung histopathology changes, as well as strongly inhibiting the expression of pro-inflammatory cytokines/chemokines, without inducing toxicity effects in mice. These results suggest that ZBMD-1 is a promising anti-influenza compound which can be further investigated as a useful strategy against IAVs in the future.


Assuntos
Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Antivirais/farmacologia , Vírus da Influenza A/efeitos dos fármacos , Proteínas de Ligação a RNA/antagonistas & inibidores , Proteínas do Core Viral/antagonistas & inibidores , Replicação Viral/efeitos dos fármacos , Animais , Humanos , Vírus da Influenza A Subtipo H1N1/efeitos dos fármacos , Vírus da Influenza A Subtipo H1N1/metabolismo , Vírus da Influenza A Subtipo H3N2/efeitos dos fármacos , Vírus da Influenza A Subtipo H3N2/metabolismo , Vírus da Influenza A/metabolismo , Concentração Inibidora 50 , Masculino , Camundongos Endogâmicos BALB C , Proteínas do Nucleocapsídeo , Infecções por Orthomyxoviridae/prevenção & controle , Infecções por Orthomyxoviridae/virologia , Proteínas de Ligação a RNA/metabolismo , Bibliotecas de Moléculas Pequenas/farmacologia , Proteínas do Core Viral/metabolismo
10.
Invest Ophthalmol Vis Sci ; 58(14): 6030-6037, 2017 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-29196767

RESUMO

Purpose: A growing body of evidence suggests that the microbiome of the ocular surface confers potent immunoregulatory functions and has a key role in the physiologic maintenance of healthy eyes and in the pathogenesis of ocular diseases. Although the microbiome is known to be affected by age and sex, the influence of these factors on ocular surface microbiota in healthy adults remains largely unknown. Methods: Ocular surface microbiome samples were obtained from the inferior bulbar conjunctiva of 48 young and 42 old adults at Zhongshan Ophthalmic Center. Using metagenomic shotgun sequencing, we characterized the sex- and age-differences in conjunctival microbiome profiles of healthy adults. Results: Male and female groups differed only in the ß diversity of bacterial communities, while there were significant differences in bacterial composition, metabolic functions, and the abundance of antibiotic resistance genes between young and old adult groups. Conclusions: Our findings suggest that age and sex collectively shape the conjunctival microbiome, and may change the immune homeostasis of the ocular surface through alterations of its commensal microbiome.


Assuntos
Bactérias/genética , Túnica Conjuntiva/microbiologia , DNA Bacteriano/análise , Infecções Oculares Bacterianas/microbiologia , Imunidade Inata , Microbiota , Adulto , Fatores Etários , Idoso , Idoso de 80 Anos ou mais , Bactérias/isolamento & purificação , Infecções Oculares Bacterianas/imunologia , Feminino , Voluntários Saudáveis , Humanos , Masculino , Pessoa de Meia-Idade , Análise de Sequência de DNA , Fatores Sexuais , Adulto Jovem
11.
J Virol ; 90(8): 3966-3980, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26842467

RESUMO

UNLABELLED: The viral ribonucleoprotein (vRNP) complex of influenza A viruses (IAVs) contains an RNA-dependent RNA polymerase complex (RdRp) and nucleoprotein (NP) and is the functional unit for viral RNA transcription and replication. The vRNP complex is an important determinant of virus pathogenicity and host adaptation, implying that its function can be affected by host factors. In our study, we identified host protein Moloney leukemia virus 10 (MOV10) as an inhibitor of IAV replication, since depletion of MOV10 resulted in a significant increase in virus yield. MOV10 inhibited the polymerase activity in a minigenome system through RNA-mediated interaction with the NP subunit of vRNP complex. Importantly, we found that the interaction between MOV10 and NP prevented the binding of NP to importin-α, resulting in the retention of NP in the cytoplasm. Both the binding of MOV10 to NP and its inhibitory effect on polymerase activity were independent of its helicase activity. These results suggest that MOV10 acts as an anti-influenza virus factor through specifically inhibiting the nuclear transportation of NP and subsequently inhibiting the function of the vRNP complex. IMPORTANCE: The interaction between the influenza virus vRNP complex and host factors is a major determinant of viral tropism and pathogenicity. Our study identified MOV10 as a novel host restriction factor for the influenza virus life cycle since it inhibited the viral growth rate. Conversely, importin-α has been shown as a determinant for influenza tropism and a positive regulator for viral polymerase activity in mammalian cells but not in avian cells. MOV10 disrupted the interaction between NP and importin-α, suggesting that MOV10 could also be an important host factor for influenza virus transmission and pathogenicity. Importantly, as an interferon (IFN)-inducible protein, MOV10 exerted a novel mechanism for IFNs to inhibit the replication of influenza viruses. Furthermore, our study potentially provides a new drug design strategy, the use of molecules that mimic the antiviral mechanism of MOV10.


Assuntos
Transporte Ativo do Núcleo Celular , Vírus da Influenza A/fisiologia , RNA Helicases/metabolismo , Proteínas de Ligação a RNA/metabolismo , Proteínas do Core Viral/metabolismo , Animais , Linhagem Celular Tumoral , Cães , Inibidores Enzimáticos/metabolismo , Células HEK293 , Humanos , Células Madin Darby de Rim Canino , Proteínas do Nucleocapsídeo , Ligação Proteica , Proteínas de Ligação a RNA/antagonistas & inibidores , Proteínas de Ligação a RNA/isolamento & purificação , RNA Polimerase Dependente de RNA/antagonistas & inibidores , RNA Polimerase Dependente de RNA/metabolismo , Proteínas do Core Viral/antagonistas & inibidores , Proteínas do Core Viral/isolamento & purificação
12.
Virology ; 486: 15-26, 2015 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-26379090

RESUMO

Human immunodeficiency virus type 1 (HIV-1) exploits multiple host factors during its replication. The REV/RRE-dependent nuclear export of unspliced/partially spliced viral transcripts needs the assistance of host proteins. Recent studies have shown that MOV10 overexpression inhibited HIV-1 replication at various steps. However, the endogenous MOV10 was required in certain step(s) of HIV-1 replication. In this report, we found that MOV10 potently enhances the nuclear export of viral mRNAs and subsequently increases the expression of Gag protein and other late products through affecting the Rev/RRE axis. The co-immunoprecipitation analysis indicated that MOV10 interacts with Rev in an RNA-independent manner. The DEAG-box of MOV10 was required for the enhancement of Rev/RRE-dependent nuclear export and the DEAG-box mutant showed a dominant-negative activity. Our data propose that HIV-1 utilizes the anti-viral factor MOV10 to function as a co-factor of Rev and demonstrate the complicated effects of MOV10 on HIV-1 life cycle.


Assuntos
Infecções por HIV/enzimologia , HIV-1/metabolismo , RNA Helicases/metabolismo , RNA Viral/metabolismo , Elementos de Resposta , Produtos do Gene rev do Vírus da Imunodeficiência Humana/metabolismo , Transporte Biológico , Produtos do Gene gag/genética , Produtos do Gene gag/metabolismo , Infecções por HIV/genética , Infecções por HIV/virologia , HIV-1/genética , Interações Hospedeiro-Patógeno , Humanos , Ligação Proteica , Estrutura Terciária de Proteína , RNA Helicases/química , RNA Helicases/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Viral/genética , Produtos do Gene rev do Vírus da Imunodeficiência Humana/genética
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