Reactivation of latent HIV-1 in vitro using an ethanolic extract from Euphorbia umbellata (Euphorbiaceae) latex

Euphorbia umbellata (E. umbellata) belongs to Euphorbiaceae family, popularly known as Janauba, and its latex contains a combination of phorbol esters with biological activities described to different cellular protein kinase C (PKC) isoforms. Here, we identified deoxi-phorbol esters present in E. umbellata latex alcoholic extract that are able to increase HIV transcription and reactivate virus from latency models. This activity is probably mediated by NF-kB activation followed by nuclear translocation and binding to the HIV LTR promoter. In addition, E. umbellata latex extract induced the production of pro inflammatory cytokines in vitro in human PBMC cultures. This latex extract also activates latent virus in human PBMCs isolated from HIV positive patients as well as latent SIV in non-human primate primary CD4+ T lymphocytes. Together, these results indicate that the phorbol esters present in E. umbellata latex are promising candidate compounds for future clinical trials for shock and kill therapies to promote HIV cure and eradication.Research and experimental expenses were funded by the grant E26/2015064289 from FAPERJinfo:eu-repo/semantics/publishedVersio

TITLE: Zika virus impairs growth in human neurospheres and brain organoids

SUMMARY: We provide evidence that Zika virus infects human iPS-derived neural stem cells, causing cell death and reduced growth in neurospheres and cerebral organoids

Use of the heteroduplex mobility assay and cell sorting to select genome sequences of the CCR5 gene in HEK 293T cells edited by transcription activator-like effector nucleases

Engineered nucleases such as zinc finger nucleases (ZFN) and transcription activator-like effector nucleases (TALEN) are one of the most promising tools for modifying genomes. These site-specific enzymes cause double- strand breaks that allow gene disruption or gene insertion, thereby facilitating genetic manipulation. The major problem associated with this approach is the labor-intensive procedures required to screen and confirm the cellular modification by nucleases. In this work, we produced a TALEN that targets the human CCR5 gene and developed a heteroduplex mobility assay for HEK 293T cells to select positive colonies for sequencing. This approach provides a useful tool for the quick detection and easy assessment of nuclease activity

Development of a rapid phenotypic test for HCV protease inhibitors with potential use in clinical decisions

Abstract Approximately 185 million people worldwide are chronically infected with hepatitis C virus (HCV). The first-wave of approved NS3 protease inhibitors (PIs) were Telaprevir and Boceprevir, which are currently discontinued. Simeprevir is a second-wave PI incorporated into the Brazilian hepatitis C treatment protocol. Drug resistance plays a key role in patients' treatment regimen. Here, we developed a simple phenotypic assay to evaluate the impact of resistance mutations in HCV NS3 protease to PIs, using a protein expression vector containing wild type NS3 protease domain and NS4A co-factor. We analyzed the impact of five resistance mutations (T54A, V36M, V158I, V170I and T54S+V170I) against Telaprevir, Boceprevir and Simeprevir. Protein purifications were performed with low cost methodology, and enzymatic inhibition assays were measured by FRET. We obtained recombinant proteases with detectable activity, and IC50 and fold change values for the evaluated PIs were determined. The variant T54A showed the highest reduction of susceptibility for the PIs, while the other four variants exhibited lower levels of reduced susceptibility. Interestingly, V170I showed 3.2-fold change for Simeprevir, a new evidence about this variant. These results emphasize the importance of enzymatic assays in phenotypic tests to determine which therapeutic regimen should be implemented

Zika virus disrupts molecular fingerprinting of human neurospheres

Zika virus (ZIKV) has been associated with microcephaly and other brain abnormalities; however, the molecular consequences of ZIKV to human brain development are still not fully understood. Here we describe alterations in human neurospheres derived from induced pluripotent stem (iPS) cells infected with the strain of Zika virus that is circulating in Brazil. Combining proteomics and mRNA transcriptional profiling, over 500 proteins and genes associated with the Brazilian ZIKV infection were found to be differentially expressed. These genes and proteins provide an interactome map, which indicates that ZIKV controls the expression of RNA processing bodies, miRNA biogenesis and splicing factors required for self-replication. It also suggests that impairments in the molecular pathways underpinning cell cycle and neuronal differentiation are caused by ZIKV. These results point to biological mechanisms implicated in brain malformations, which are important to further the understanding of ZIKV infection and can be exploited as therapeutic potential targets to mitigate it7CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQCOORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPESFINANCIADORA DE ESTUDOS E PROJETOS - FINEPFUNDAÇÃO CARLOS CHAGAS FILHO DE AMPARO À PESQUISA DO ESTADO DO RIO DE JANEIRO - FAPERJFUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESPsem informaçãosem informaçãosem informaçãosem informação14/21035-0; 14/14881-1; 13/08711-3; 14/10068-

Zika virus disrupts molecular fingerprinting of human neurospheres

Submitted by Sandra Infurna ([email protected]) on 2018-04-03T16:26:16Z No. of bitstreams: 1 anamariab_filippis_etal_IOC_2017.pdf: 1347352 bytes, checksum: b3a68b2d8ea28413682153a717faa4c4 (MD5)Approved for entry into archive by Sandra Infurna ([email protected]) on 2018-04-03T16:44:04Z (GMT) No. of bitstreams: 1 anamariab_filippis_etal_IOC_2017.pdf: 1347352 bytes, checksum: b3a68b2d8ea28413682153a717faa4c4 (MD5)Made available in DSpace on 2018-04-03T16:44:04Z (GMT). No. of bitstreams: 1 anamariab_filippis_etal_IOC_2017.pdf: 1347352 bytes, checksum: b3a68b2d8ea28413682153a717faa4c4 (MD5) Previous issue date: 2017Instituto D´Or de Pesquisa e Educação. Rio de Janeiro, Brasil / Universidade Federal do Rio de Janeiro. Instituto de Ciências Biológicas. Rio de Janeiro, RJ, Brasil.Instituto D´Or de Pesquisa e Educação. Rio de Janeiro, Brasil / Universidade de Campinas. Departamento de Bioquímica e Biologia Tecidual. Campinas, SP, Brasil.Instituto Evandro Chagas. Centro de Inovação Tecnológica. Belém, PA, BrasilInstituto D´Or de Pesquisa e Educação. Rio de Janeiro, Brasil.Universidade Federal do Rio de Janeiro. Instituto de Biologia. Rio de Janeiro, RJ, BrasilInstituto D´Or de Pesquisa e Educação. Rio de Janeiro, Brasil.Instituto D´Or de Pesquisa e Educação. Rio de Janeiro, Brasil.Universidade Federal do Rio de Janeiro. Instituto de Biologia. Rio de Janeiro, RJ, BrasilUniversidade de Campinas. Departamento de Bioquímica e Biologia Tecidual. Campinas, SP, Brasil.Instituto D´Or de Pesquisa e Educação. Rio de Janeiro, Brasil.Fundação Oswaldo Cruz Fiocruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, Brasil.Instituto D´Or de Pesquisa e Educação. Rio de Janeiro, Brasil.Universidade Federal do Rio de Janeiro. Instituto de Biologia. Rio de Janeiro, RJ, BrasilUniversidade Federal do Pará. Belém, PA, Brasil.Fundação Oswaldo Cruz Fiocruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, Brasil.Instituto Evandro Chagas. Centro de Inovação Tecnológica. Belém, PA, BrasilUniversidade Federal do Rio de Janeiro. Instituto de Biologia. Rio de Janeiro, RJ, BrasilUniversidade de Campinas. Departamento de Bioquímica e Biologia Tecidual. Campinas, SP, Brasil.Instituto D´Or de Pesquisa e Educação. Rio de Janeiro, Brasil / Universidade Federal do Rio de Janeiro. Instituto de Ciências Biológicas. Rio de Janeiro, RJ, Brasil.Zika virus (ZIKV) has been associated with microcephaly and other brain abnormalities; however, the molecular consequences of ZIKV to human brain development are still not fully understood. Here we describe alterations in human neurospheres derived from induced pluripotent stem (iPS) cells infected with the strain of Zika virus that is circulating in Brazil. Combining proteomics and mRNA transcriptional profiling, over 500 proteins and genes associated with the Brazilian ZIKV infection were found to be differentially expressed. These genes and proteins provide an interactome map, which indicates that ZIKV controls the expression of RNA processing bodies, miRNA biogenesis and splicing factors required for self-replication. It also suggests that impairments in the molecular pathways underpinning cell cycle and neuronal differentiation are caused by ZIKV. These results point to biological mechanisms implicated in brain malformations, which are important to further the understanding of ZIKV infection and can be exploited as therapeutic potential targets to mitigate it

Chloroquine, an Endocytosis Blocking Agent, Inhibits Zika Virus Infection in Different Cell Models

Zika virus (ZIKV) infection in utero might lead to microcephaly and other congenital defects. Since no specific therapy is available thus far, there is an urgent need for the discovery of agents capable of inhibiting its viral replication and deleterious effects. Chloroquine is widely used as an antimalarial drug, anti-inflammatory agent, and it also shows antiviral activity against several viruses. Here we show that chloroquine exhibits antiviral activity against ZIKV in Vero cells, human brain microvascular endothelial cells, human neural stem cells, and mouse neurospheres. We demonstrate that chloroquine reduces the number of ZIKV-infected cells in vitro, and inhibits virus production and cell death promoted by ZIKV infection without cytotoxic effects. In addition, chloroquine treatment partially reveres morphological changes induced by ZIKV infection in mouse neurospheres

X-ray structure of O-methyl-acrocol and anti-cancer, anti-parasitic, anti-bacterial and anti-Zika virus evaluations of the Brazilian palm tree Acrocomia totai

Acrocomia totai Mart ("macauba") is a palm tree native from Brazil, whose potential for biodiesel production has been widely explored. In spite of the industrial interest in the oil from the nuts, little is known about the potential applications of other parts of the plant, especially in the pharmacological area. A phytochemical study of the plant thorns led to the identification of a new compound 3-(R)-methoxy-21-(R)-H-hop-22(29)-en-30-ol 1, two known triterpenes 2-3, four steroids 4-7 and a stilbene, piceatannol 8. The structures were elucidated by spectroscopic analyses, including 1D and 2D NMR and low and high resolution mass spectrometry. Compound 1 was purified as crystals, which allowed the determination of the absolute configuration of the asymmetric carbons by analysis of the X-ray diffraction spectrum. Biological tests were performed with crude extract (CE), fractions and isolated compound. The assays showed activity for CE against lung carcinoma (GI(50) 59.2 mu g mL(-1)). The ethyl acetate fraction (EAF) showed efficacy against many tumor cell lines, and the tests showed the most prominent activity for breast cancer (GI(50) 10.4 mu g mL(-1)), glioma (GI(50) 77.3 mu g mL(-1)), uterine cervix (SiHa) HPV 16 (IC50 39.8 mu g mL(-1)), (HeLa) HPV 18 (IC50 12.0 pg mL(-1)) and Caco-2 (IC50 40.0 mu g mL(-1)) and showed bacteriostatic action against Staphylococcus aureus (MIC 50 mu g mL(-1)). Piceatannol 8 isolated from EAF showed activity against the protozoan which causes leishmaniosis (IC50 58.4 mu g mL(-1)). For Ttypanosorna cruzi, the methanol fraction (EC50 15.5 mu g mL(-1)), CE (20.5 mu g(-1)), and HEF (43.8 mu g mL(-1)) were the most active, being highly selective for the protozoan and less toxic against Vero cells. The compound 8 was further tested against Zika virus MR 766 strain on MOI 2, however the assays showed no inhibition against virus infection109483492CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO - CNPQCOORDENAÇÃO DE APERFEIÇOAMENTO DE PESSOAL DE NÍVEL SUPERIOR - CAPESFUNDAÇÃO ARAUCÁRIA DE APOIO AO DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO DO ESTADO DO PARANÁ - FAFUNDAÇÃO DE AMPARO À PESQUISA E INOVAÇÃO DO ESTADO DE SANTA CATARINA - FAPESCsem informaçãosem informaçãosem informaçãosem informaçã