Autologous extracellular traps (ETs) influence T cell profiles and are not activating factors of human M1 macrophages
The formation of extracellular traps (ETs) is an important microbicidal functional mechanism of immune cells due to its implication in the pathogenesis of various diseases, including COVID-19. We set out to study the influence of ETs on the differentiation profiles of human lymphocytes and macrophag...
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| Formato: | Artículo revista |
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Universidad Nacional Córdoba. Facultad de Ciencias Médicas. Secretaria de Ciencia y Tecnología
2022
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| Acceso en línea: | https://revistas.unc.edu.ar/index.php/med/article/view/39024 |
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I10-R327-article-39024 |
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ojs |
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Universidad Nacional de Córdoba |
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I-10 |
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R-327 |
| container_title_str |
Revista de la Facultad de Ciencias Médicas de Córdoba |
| format |
Artículo revista |
| topic |
extracellular traps human leukocytes autologous culture trampas extracelulares leucocitos humanos cultivo autólogo |
| spellingShingle |
extracellular traps human leukocytes autologous culture trampas extracelulares leucocitos humanos cultivo autólogo Bailo , G Rodríguez, FM Carabajal Miotti , CL Ruiz de Frattari, S Vargas, AH González Silva, N Novak, ITC Autologous extracellular traps (ETs) influence T cell profiles and are not activating factors of human M1 macrophages |
| topic_facet |
extracellular traps human leukocytes autologous culture trampas extracelulares leucocitos humanos cultivo autólogo |
| author |
Bailo , G Rodríguez, FM Carabajal Miotti , CL Ruiz de Frattari, S Vargas, AH González Silva, N Novak, ITC |
| author_facet |
Bailo , G Rodríguez, FM Carabajal Miotti , CL Ruiz de Frattari, S Vargas, AH González Silva, N Novak, ITC |
| author_sort |
Bailo , G |
| title |
Autologous extracellular traps (ETs) influence T cell profiles and are not activating factors of human M1 macrophages |
| title_short |
Autologous extracellular traps (ETs) influence T cell profiles and are not activating factors of human M1 macrophages |
| title_full |
Autologous extracellular traps (ETs) influence T cell profiles and are not activating factors of human M1 macrophages |
| title_fullStr |
Autologous extracellular traps (ETs) influence T cell profiles and are not activating factors of human M1 macrophages |
| title_full_unstemmed |
Autologous extracellular traps (ETs) influence T cell profiles and are not activating factors of human M1 macrophages |
| title_sort |
autologous extracellular traps (ets) influence t cell profiles and are not activating factors of human m1 macrophages |
| description |
The formation of extracellular traps (ETs) is an important microbicidal functional mechanism of immune cells due to its implication in the pathogenesis of various diseases, including COVID-19. We set out to study the influence of ETs on the differentiation profiles of human lymphocytes and macrophages in autologous culture. The objectives were a) to cause the generation of ETs in vitro with lipopolysaccharide (LPS) or formylated peptides fMLFN-formyl-met-leu-phe (fMLP) and their isolation; b) to cause processing and presentation of the heterologous ovalbumin antigen (OVA) and c) mark by immunofluorescence (IF) molecules of the CD4 T profile, Th17 helper T cells and innate lymphoid cells 3 ILC3 (transcription factor RORɣ, orphan nuclear receptor); as well as the common leukocyte antigen CD45RO+ marker for T-cell activation and, the macrophage classic activation profile molecule M1, inducible nitric oxide synthetase (iNOS).
Autologous total leukocyte cell cultures were performed (n=10) from healthy human peripheral blood (donated by IHH, UNC, Ethically approved, HNC, FCM, UNC) that were challenged with OVA, and subjected to autologous ET stock. Controls: paired samples without challenge. Samples were taken at 24 and 72 h. IF with anti-RORɣ, anti-CD4, anti-CD45RO, anti-NOS2, and DNA staining with DAPI. Quantification of labeled cells was performed with FIJI program. Statistical treatment: t-test for paired samples.
At 24 hours, significant differences were observed in the percentages of positive cells for CD4 and CD45RO between paired control and OVA-challenged samples (*p<0.05) and between paired control and OVA- and ET-added samples (*p <0.05). In an independent experiment, significant differences were observed between those stimulated with OVA vs those stimulated with OVA and addition of ETs (*p<0.05). At 72 hours of culture, no significant differences were found between the paired samples in any case. There were no significant differences either at 24 or 72 hours of culture in the percentages of cells positive for RORɣ or in the percentages of cells positive for iNOS in different challenges.
Influence of ETs on T cell activation was observed and components of autologous ETs did not elicit classical activation of M1 macrophages. |
| publisher |
Universidad Nacional Córdoba. Facultad de Ciencias Médicas. Secretaria de Ciencia y Tecnología |
| publishDate |
2022 |
| url |
https://revistas.unc.edu.ar/index.php/med/article/view/39024 |
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I10-R327-article-390242024-04-15T16:14:45Z Autologous extracellular traps (ETs) influence T cell profiles and are not activating factors of human M1 macrophages Trampas extracelulares (ETs) autólogas influyen en perfiles de linfocitos T y no son factores de activación de macrófagos M1 humanos Bailo , G Rodríguez, FM Carabajal Miotti , CL Ruiz de Frattari, S Vargas, AH González Silva, N Novak, ITC extracellular traps human leukocytes autologous culture trampas extracelulares leucocitos humanos cultivo autólogo The formation of extracellular traps (ETs) is an important microbicidal functional mechanism of immune cells due to its implication in the pathogenesis of various diseases, including COVID-19. We set out to study the influence of ETs on the differentiation profiles of human lymphocytes and macrophages in autologous culture. The objectives were a) to cause the generation of ETs in vitro with lipopolysaccharide (LPS) or formylated peptides fMLFN-formyl-met-leu-phe (fMLP) and their isolation; b) to cause processing and presentation of the heterologous ovalbumin antigen (OVA) and c) mark by immunofluorescence (IF) molecules of the CD4 T profile, Th17 helper T cells and innate lymphoid cells 3 ILC3 (transcription factor RORɣ, orphan nuclear receptor); as well as the common leukocyte antigen CD45RO+ marker for T-cell activation and, the macrophage classic activation profile molecule M1, inducible nitric oxide synthetase (iNOS). Autologous total leukocyte cell cultures were performed (n=10) from healthy human peripheral blood (donated by IHH, UNC, Ethically approved, HNC, FCM, UNC) that were challenged with OVA, and subjected to autologous ET stock. Controls: paired samples without challenge. Samples were taken at 24 and 72 h. IF with anti-RORɣ, anti-CD4, anti-CD45RO, anti-NOS2, and DNA staining with DAPI. Quantification of labeled cells was performed with FIJI program. Statistical treatment: t-test for paired samples. At 24 hours, significant differences were observed in the percentages of positive cells for CD4 and CD45RO between paired control and OVA-challenged samples (*p<0.05) and between paired control and OVA- and ET-added samples (*p <0.05). In an independent experiment, significant differences were observed between those stimulated with OVA vs those stimulated with OVA and addition of ETs (*p<0.05). At 72 hours of culture, no significant differences were found between the paired samples in any case. There were no significant differences either at 24 or 72 hours of culture in the percentages of cells positive for RORɣ or in the percentages of cells positive for iNOS in different challenges. Influence of ETs on T cell activation was observed and components of autologous ETs did not elicit classical activation of M1 macrophages. La formación de trampas extracelulares (ETs) es un mecanismo funcional microbicida de células inmunes importante por su implicancia en la patogenia de diversas enfermedades, incluida COVID-19. Nos propusimos estudiar la influencia de ETs sobre la diferenciación de perfiles de linfocitos y macrófagos humanos en cultivo autólogo. Los objetivos fueron a) provocar la generación de ETs in vitro con lipopolisacarido (LPS) o péptidos formilados fMLFN-formyl-met-leu-phe (fMLP) y su aislamiento; b) provocar procesamiento y presentación del antígeno heterólogo ovoalbúmina (OVA) y c) marcar por inmunofluorescencia (IF) moléculas del perfil T CD4, células T helper Th17 y células linfoides innatas 3 ILC3 (factor de transcripción RORɣ, receptor nuclear huérfano); así como el antígeno común leucocitario CD45RO+ marcador de activación de células T y, la molécula del perfil de activación clásica de macrófagos M1, óxido nítrico sintetasa inducible (iNOS). Se realizaron (n=10) cultivos celulares autólogos de leucocitos totales de sangre periférica humana sana (donada por IHH, UNC, con aprobación Ética, HNC, FCM, UNC) que fueron desafiados con OVA, y sometidos al stock de ETs autólogas. Controles: muestras apareadas sin desafío. Se tomaron muestras a 24 y 72 h. IF con anti-RORɣ, anti-CD4, anti-CD45RO, anti-NOS2 y tinción de ADN con DAPI. Cuantificación de células marcadas con programa FIJI. Tratamiento estadístico: test t para muestras apareadas. A 24 hs se observaron diferencias significativas en los porcentajes de células positivas para CD4 y CD45RO entre muestras apareadas control y con desafío de OVA, (*p<0,05) y entre muestras apareadas control y con agregado de OVA y ETs (*p<0,05). En un experimento independiente se observaron diferencias significativas entre las estimuladas con OVA vs aquellas estimuladas con OVA y agregado de ETs (*p<0,05). A 72 hs de cultivo no se hallaron diferencias significativas entre las muestras apareadas en ningún caso. No hubo diferencias significativas tanto a 24 como a 72 hs de cultivo en los porcentajes de células positivas para RORɣ ni en los porcentajes de células positivas para iNOS en los diferentes desafíos. Se observó influencia de las ETs en la activación celular T y los componentes de las ETs autólogas no provocaron activación clásica de macrófagos M1. Universidad Nacional Córdoba. Facultad de Ciencias Médicas. Secretaria de Ciencia y Tecnología 2022-10-26 info:eu-repo/semantics/article info:eu-repo/semantics/publishedVersion . https://revistas.unc.edu.ar/index.php/med/article/view/39024 Revista de la Facultad de Ciencias Médicas de Córdoba.; Vol. 79 No. Suplemento JIC XXIII (2022): Suplemento JIC XXIII Revista de la Facultad de Ciencias Médicas de Córdoba; Vol. 79 Núm. Suplemento JIC XXIII (2022): Suplemento JIC XXIII Revista da Faculdade de Ciências Médicas de Córdoba; v. 79 n. Suplemento JIC XXIII (2022): Suplemento JIC XXIII 1853-0605 0014-6722 http://creativecommons.org/licenses/by-nc/4.0 |