Abordagem terapêutica com um lisado bacteriano (om-85) frente a infecção pelo virus sars-cov-2 : um estudo experimental ex vivo
Autor(a) principal: | |
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Data de Publicação: | 2022 |
Tipo de documento: | Dissertação |
Idioma: | por |
Título da fonte: | Biblioteca Digital de Teses e Dissertações da PUC_RS |
Texto Completo: | https://tede2.pucrs.br/tede2/handle/tede/10412 |
Resumo: | Introduction: The COVID-19 pandemic, caused by the SARS-CoV-2 virus, affects many people around the world. Even with the development of vaccines, there is an urgent need to develop effective therapeutic approaches to combat and prevent this viral infection. OM-85 is an immunostimulating bacterial lysate capable of modulating the antiviral response, already used to prevent recurrent respiratory infections acute otitis media, pneumonia, rhinitis, pharyngotonsillitis, asthma, chronic obstructive pulmonary disease). Objective: To understand whether bacterial lysate (OM-85) is able to activate molecular antiviral mechanisms in SARS-CoV-2 infected cells. Methods: This is an ex vivo experimental study. Epithelial cells from nasal lavage of patients with COVID-19, diagnosed by RT-PCR, with up to 48 hours of hospital admission and within three weeks of the onset of symptoms, were cultured for 24 hours in the presence of OM-85 (10 μg /ml) and 2.5 ng/ml of IFNβ. RNA from cells was extracted and performed by RT-PCR, analysis of the relative expression of the SARS-CoV-2 N1 gene, the ACE2 virus receptor gene and the genes related to the antiviral response, ISG15, MAVS, MDA5, OASL, IRF3, RIG-I, IFNλ and with inflammatory response, IL-6, IL-8, mTOR, TRL2, TLSP, IL1B. Cell supernatant was collected for cytokine analysis by ELISA. In addition, A549WT and Calu-3 cell lines were cultured, infected with SARS-CoV-2 for one hour and treated with OM-85 for 24 hours. Results: Treatment with OM-85 reduced the viral gene expression of SARS-Cov-2 and increased the expression of the RIG-I molecule. Analyzes on the expression of IFNB, ISG15, IL-6, MDA5, OASL, IL1B had no statistical significance in their expression when comparing the group of untreated cells with the treated group. The gene expression of IL-8, MAVS, IRF3, mTOR, IFNλ, TRL2, TLSP and ACE2 was not detected. When analyzing the proteins IFNB, IL-6, IL-8, IL-10 and TNFα by ELISA test, there was no relevant difference between the control group and the treated group. This was repeated when we analyzed the correlations of these same proteins, as well as with the other cytokines, with the expression of SARS-CoV-2. As for the in vitro experiment, the analysis of the gene expression of the virus showed a tendency to reduce this expression in A549WT cells when treated with OM-85; which was not observed in the Calu-3 lineage. Conclusion: Our data complement previous studies that show a protective effect of OM-85 against SARS-CoV-2 infection in mice. Clinical trials are needed to confirm a possible therapeutic or prophylactic use against coronavirus infection. |
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Stein, Renato Tetelbomhttp://lattes.cnpq.br/8128743330371501Souza, Ana Paula Duarte dehttp://lattes.cnpq.br/7065225247536877http://lattes.cnpq.br/4073795741208559Cassão, Gisele2022-08-31T12:01:37Z2022-04-18https://tede2.pucrs.br/tede2/handle/tede/10412Introduction: The COVID-19 pandemic, caused by the SARS-CoV-2 virus, affects many people around the world. Even with the development of vaccines, there is an urgent need to develop effective therapeutic approaches to combat and prevent this viral infection. OM-85 is an immunostimulating bacterial lysate capable of modulating the antiviral response, already used to prevent recurrent respiratory infections acute otitis media, pneumonia, rhinitis, pharyngotonsillitis, asthma, chronic obstructive pulmonary disease). Objective: To understand whether bacterial lysate (OM-85) is able to activate molecular antiviral mechanisms in SARS-CoV-2 infected cells. Methods: This is an ex vivo experimental study. Epithelial cells from nasal lavage of patients with COVID-19, diagnosed by RT-PCR, with up to 48 hours of hospital admission and within three weeks of the onset of symptoms, were cultured for 24 hours in the presence of OM-85 (10 μg /ml) and 2.5 ng/ml of IFNβ. RNA from cells was extracted and performed by RT-PCR, analysis of the relative expression of the SARS-CoV-2 N1 gene, the ACE2 virus receptor gene and the genes related to the antiviral response, ISG15, MAVS, MDA5, OASL, IRF3, RIG-I, IFNλ and with inflammatory response, IL-6, IL-8, mTOR, TRL2, TLSP, IL1B. Cell supernatant was collected for cytokine analysis by ELISA. In addition, A549WT and Calu-3 cell lines were cultured, infected with SARS-CoV-2 for one hour and treated with OM-85 for 24 hours. Results: Treatment with OM-85 reduced the viral gene expression of SARS-Cov-2 and increased the expression of the RIG-I molecule. Analyzes on the expression of IFNB, ISG15, IL-6, MDA5, OASL, IL1B had no statistical significance in their expression when comparing the group of untreated cells with the treated group. The gene expression of IL-8, MAVS, IRF3, mTOR, IFNλ, TRL2, TLSP and ACE2 was not detected. When analyzing the proteins IFNB, IL-6, IL-8, IL-10 and TNFα by ELISA test, there was no relevant difference between the control group and the treated group. This was repeated when we analyzed the correlations of these same proteins, as well as with the other cytokines, with the expression of SARS-CoV-2. As for the in vitro experiment, the analysis of the gene expression of the virus showed a tendency to reduce this expression in A549WT cells when treated with OM-85; which was not observed in the Calu-3 lineage. Conclusion: Our data complement previous studies that show a protective effect of OM-85 against SARS-CoV-2 infection in mice. Clinical trials are needed to confirm a possible therapeutic or prophylactic use against coronavirus infection.Introdução: A pandemia de COVID-19, causada pelo vírus SARS-CoV-2, acomete muitas pessoas ao redor do mundo. Mesmo com o desenvolvimento de vacinas, há necessidade urgente em se desenvolver abordagens terapêuticas eficazes no combate e prevenção dessa infecção viral. O OM-85 é um lisado bacteriano, imunoestimulador, capaz de modular a resposta antiviral, já utilizado para prevenção de infecções respiratórias recorrentes (otite média aguda, pneumonia, rinite, faringoamigdalite, asma, doença pulmonar obstrutiva crônica). Objetivo: Entender se o lisado bacteriano (OM-85) é capaz de ativar mecanismos moleculares antivirais em células infectadas por SARS-CoV-2. Métodos: Trata-se de um estudo experimental ex vivo. As células epiteliais provenientes do lavado nasal de pacientes com COVID-19, diagnosticados por RT-PCR, com até 48h de internação hospitalar e em até três semanas do início de sintomas, foram cultivadas por 24 horas na presença de OM-85 (10 μg/ml) e 2,5 ng/ml de IFNβ. O RNA das células foi extraído e foi realizada por RT-PCR, a análise da expressão relativa do gene SARS-CoV-2 N1, do gene do receptor do vírus ACE2 e dos genes relacionados com a resposta antiviral, ISG15, MAVS, MDA5, OASL, IRF3, RIG-I, IFNλ e com resposta inflamatória, IL-6, IL-8, mTOR, TRL2, TLSP, IL1B. O sobrenadante das células foi coletado para análise das citocinas por ELISA. Além disso, linhagens celulares A549WT e Calu-3 foram cultivadas, infectadas com SARS-CoV-2 por uma hora e tratadas com OM-85 por 24 horas. Resultados: Tratamento com OM-85 reduziu a expressão gênica viral do SARS-Cov-2 e aumentou a expressão da molécula RIG-I. Análises na expressão de IFNB, ISG15, IL-6, MDA5, OASL, IL1B não tiveram significância estatística na sua expressão quando comparados o grupo de células não tratadas com o grupo tratado. Já a expressão gênica de IL-8, MAVS, IRF3, mTOR, IFNλ, TRL2, TLSP e ACE2 não foi detectada. Ao analisarmos as proteínas IFNB, IL-6, IL-8, IL-10 e TNFα por teste de ELISA, não houve diferença relevante entre o grupo controle e grupo tratado. Isso se repetiu ao analisarmos as correlações dessas mesmas proteínas, bem como com as demais citocinas com a expressão de SARS-CoV-2. Quanto ao experimento in vitro a análise da expressão gênica do vírus demonstrou uma tendência na redução dessa expressão em células A549WT quando tratadas com OM-85; o que não foi observado na linhagem Calu-3. Conclusão: Nossos dados complementam estudos prévios que mostram efeito protetor do OM-85 frente à infecção por SARS-CoV-2 em camundongos. Ensaios clínicos são necessários para confirmar uma possível utilização terapêutica ou profilática frente a infecção por coronavírus.Submitted by PPG Pediatria e Saúde da Criança (pediatria-pg@pucrs.br) on 2022-08-29T16:05:30Z No. of bitstreams: 1 DISSERTAÇÃO corrigida 2 - Homologação Gisele Cassao.pdf: 1985648 bytes, checksum: 7e75881b72c163ccfcf2c1bd6920db97 (MD5)Approved for entry into archive by Sheila Dias (sheila.dias@pucrs.br) on 2022-08-31T11:56:33Z (GMT) No. of bitstreams: 1 DISSERTAÇÃO corrigida 2 - Homologação Gisele Cassao.pdf: 1985648 bytes, checksum: 7e75881b72c163ccfcf2c1bd6920db97 (MD5)Made available in DSpace on 2022-08-31T12:01:37Z (GMT). No. of bitstreams: 1 DISSERTAÇÃO corrigida 2 - Homologação Gisele Cassao.pdf: 1985648 bytes, checksum: 7e75881b72c163ccfcf2c1bd6920db97 (MD5) Previous issue date: 2022-04-18Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPESapplication/pdfhttps://tede2.pucrs.br/tede2/retrieve/185175/DIS_GISELE_CASSAO_CONFIDENCIAL.pdf.jpgporPontifícia Universidade Católica do Rio Grande do SulPrograma de Pós-Graduação em Medicina/Pediatria e Saúde da CriançaPUCRSBrasilEscola de MedicinaPandemia COVID-19;Células EpiteliaisPacientes; IFNB; OM-85COVID 19 Pandemic;Epithelial CellsPatients; IFN-I; OM-85CIENCIAS DA SAUDE::MEDICINAMEDICINA::SAUDE MATERNO-INFANTILCLINICA MEDICA::PEDIATRIAAbordagem terapêutica com um lisado bacteriano (om-85) frente a infecção pelo virus sars-cov-2 : um estudo experimental ex vivoinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/masterThesisTrabalho será publicado como artigo ou livro60 meses31/08/2027557290555552975733500500500600600600-224747486637135387-969369452308786627-8067417953925345752-70271217037790659553590462550136975366info:eu-repo/semantics/openAccessreponame:Biblioteca Digital de Teses e Dissertações da PUC_RSinstname:Pontifícia Universidade Católica do Rio Grande do Sul (PUCRS)instacron:PUC_RSTHUMBNAILDIS_GISELE_CASSAO_CONFIDENCIAL.pdf.jpgDIS_GISELE_CASSAO_CONFIDENCIAL.pdf.jpgimage/jpeg4132https://tede2.pucrs.br/tede2/bitstream/tede/10412/5/DIS_GISELE_CASSAO_CONFIDENCIAL.pdf.jpg909d200ca241f9948586a84e0c7be17dMD55TEXTDIS_GISELE_CASSAO_CONFIDENCIAL.pdf.txtDIS_GISELE_CASSAO_CONFIDENCIAL.pdf.txttext/plain1612https://tede2.pucrs.br/tede2/bitstream/tede/10412/4/DIS_GISELE_CASSAO_CONFIDENCIAL.pdf.txt5c3af87d17adf03debd8fd327e9db7c5MD54ORIGINALDIS_GISELE_CASSAO_CONFIDENCIAL.pdfDIS_GISELE_CASSAO_CONFIDENCIAL.pdfapplication/pdf557565https://tede2.pucrs.br/tede2/bitstream/tede/10412/3/DIS_GISELE_CASSAO_CONFIDENCIAL.pdfedac3c864e1e945329ed2eebd6fe1209MD53LICENSElicense.txtlicense.txttext/plain; charset=utf-8590https://tede2.pucrs.br/tede2/bitstream/tede/10412/1/license.txt220e11f2d3ba5354f917c7035aadef24MD51tede/104122022-08-31 12:00:15.451oai:tede2.pucrs.br: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Biblioteca Digital de Teses e Dissertaçõeshttp://tede2.pucrs.br/tede2/PRIhttps://tede2.pucrs.br/oai/requestbiblioteca.central@pucrs.br||opendoar:2022-08-31T15:00:15Biblioteca Digital de Teses e Dissertações da PUC_RS - Pontifícia Universidade Católica do Rio Grande do Sul (PUCRS)false |
dc.title.por.fl_str_mv |
Abordagem terapêutica com um lisado bacteriano (om-85) frente a infecção pelo virus sars-cov-2 : um estudo experimental ex vivo |
title |
Abordagem terapêutica com um lisado bacteriano (om-85) frente a infecção pelo virus sars-cov-2 : um estudo experimental ex vivo |
spellingShingle |
Abordagem terapêutica com um lisado bacteriano (om-85) frente a infecção pelo virus sars-cov-2 : um estudo experimental ex vivo Cassão, Gisele Pandemia COVID-19; Células Epiteliais Pacientes; IFNB; OM-85 COVID 19 Pandemic; Epithelial Cells Patients; IFN-I; OM-85 CIENCIAS DA SAUDE::MEDICINA MEDICINA::SAUDE MATERNO-INFANTIL CLINICA MEDICA::PEDIATRIA |
title_short |
Abordagem terapêutica com um lisado bacteriano (om-85) frente a infecção pelo virus sars-cov-2 : um estudo experimental ex vivo |
title_full |
Abordagem terapêutica com um lisado bacteriano (om-85) frente a infecção pelo virus sars-cov-2 : um estudo experimental ex vivo |
title_fullStr |
Abordagem terapêutica com um lisado bacteriano (om-85) frente a infecção pelo virus sars-cov-2 : um estudo experimental ex vivo |
title_full_unstemmed |
Abordagem terapêutica com um lisado bacteriano (om-85) frente a infecção pelo virus sars-cov-2 : um estudo experimental ex vivo |
title_sort |
Abordagem terapêutica com um lisado bacteriano (om-85) frente a infecção pelo virus sars-cov-2 : um estudo experimental ex vivo |
author |
Cassão, Gisele |
author_facet |
Cassão, Gisele |
author_role |
author |
dc.contributor.advisor1.fl_str_mv |
Stein, Renato Tetelbom |
dc.contributor.advisor1Lattes.fl_str_mv |
http://lattes.cnpq.br/8128743330371501 |
dc.contributor.advisor-co1.fl_str_mv |
Souza, Ana Paula Duarte de |
dc.contributor.advisor-co1Lattes.fl_str_mv |
http://lattes.cnpq.br/7065225247536877 |
dc.contributor.authorLattes.fl_str_mv |
http://lattes.cnpq.br/4073795741208559 |
dc.contributor.author.fl_str_mv |
Cassão, Gisele |
contributor_str_mv |
Stein, Renato Tetelbom Souza, Ana Paula Duarte de |
dc.subject.por.fl_str_mv |
Pandemia COVID-19; Células Epiteliais Pacientes; IFNB; OM-85 |
topic |
Pandemia COVID-19; Células Epiteliais Pacientes; IFNB; OM-85 COVID 19 Pandemic; Epithelial Cells Patients; IFN-I; OM-85 CIENCIAS DA SAUDE::MEDICINA MEDICINA::SAUDE MATERNO-INFANTIL CLINICA MEDICA::PEDIATRIA |
dc.subject.eng.fl_str_mv |
COVID 19 Pandemic; Epithelial Cells Patients; IFN-I; OM-85 |
dc.subject.cnpq.fl_str_mv |
CIENCIAS DA SAUDE::MEDICINA MEDICINA::SAUDE MATERNO-INFANTIL CLINICA MEDICA::PEDIATRIA |
description |
Introduction: The COVID-19 pandemic, caused by the SARS-CoV-2 virus, affects many people around the world. Even with the development of vaccines, there is an urgent need to develop effective therapeutic approaches to combat and prevent this viral infection. OM-85 is an immunostimulating bacterial lysate capable of modulating the antiviral response, already used to prevent recurrent respiratory infections acute otitis media, pneumonia, rhinitis, pharyngotonsillitis, asthma, chronic obstructive pulmonary disease). Objective: To understand whether bacterial lysate (OM-85) is able to activate molecular antiviral mechanisms in SARS-CoV-2 infected cells. Methods: This is an ex vivo experimental study. Epithelial cells from nasal lavage of patients with COVID-19, diagnosed by RT-PCR, with up to 48 hours of hospital admission and within three weeks of the onset of symptoms, were cultured for 24 hours in the presence of OM-85 (10 μg /ml) and 2.5 ng/ml of IFNβ. RNA from cells was extracted and performed by RT-PCR, analysis of the relative expression of the SARS-CoV-2 N1 gene, the ACE2 virus receptor gene and the genes related to the antiviral response, ISG15, MAVS, MDA5, OASL, IRF3, RIG-I, IFNλ and with inflammatory response, IL-6, IL-8, mTOR, TRL2, TLSP, IL1B. Cell supernatant was collected for cytokine analysis by ELISA. In addition, A549WT and Calu-3 cell lines were cultured, infected with SARS-CoV-2 for one hour and treated with OM-85 for 24 hours. Results: Treatment with OM-85 reduced the viral gene expression of SARS-Cov-2 and increased the expression of the RIG-I molecule. Analyzes on the expression of IFNB, ISG15, IL-6, MDA5, OASL, IL1B had no statistical significance in their expression when comparing the group of untreated cells with the treated group. The gene expression of IL-8, MAVS, IRF3, mTOR, IFNλ, TRL2, TLSP and ACE2 was not detected. When analyzing the proteins IFNB, IL-6, IL-8, IL-10 and TNFα by ELISA test, there was no relevant difference between the control group and the treated group. This was repeated when we analyzed the correlations of these same proteins, as well as with the other cytokines, with the expression of SARS-CoV-2. As for the in vitro experiment, the analysis of the gene expression of the virus showed a tendency to reduce this expression in A549WT cells when treated with OM-85; which was not observed in the Calu-3 lineage. Conclusion: Our data complement previous studies that show a protective effect of OM-85 against SARS-CoV-2 infection in mice. Clinical trials are needed to confirm a possible therapeutic or prophylactic use against coronavirus infection. |
publishDate |
2022 |
dc.date.accessioned.fl_str_mv |
2022-08-31T12:01:37Z |
dc.date.issued.fl_str_mv |
2022-04-18 |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
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info:eu-repo/semantics/masterThesis |
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masterThesis |
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https://tede2.pucrs.br/tede2/handle/tede/10412 |
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https://tede2.pucrs.br/tede2/handle/tede/10412 |
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por |
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por |
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Pontifícia Universidade Católica do Rio Grande do Sul |
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Programa de Pós-Graduação em Medicina/Pediatria e Saúde da Criança |
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PUCRS |
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Brasil |
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Escola de Medicina |
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Pontifícia Universidade Católica do Rio Grande do Sul |
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