Polymorphonuclear Cell Chemotaxis and Suicidal NETosis: Simultaneous Observation Using fMLP, PMA, H7, and Live Cell Imaging
Chemotaxis and the formation of suicidal neutrophil extracellular traps (suicidal NETosis) are key functions of polymorphonuclear cells (PMNs). Neutrophil extracellular traps in particular are known to be significantly involved in the severity of inflammatory and immunological disorders such as rheu...
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| Format: | Article |
| Language: | English |
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Wiley
2020-01-01
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| Series: | Journal of Immunology Research |
| Online Access: | http://dx.doi.org/10.1155/2020/1415947 |
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| author | Delou Pai Michael Gruber Sophie-Marie Pfaehler Andre Bredthauer Karla Lehle Benedikt Trabold |
| author_facet | Delou Pai Michael Gruber Sophie-Marie Pfaehler Andre Bredthauer Karla Lehle Benedikt Trabold |
| author_sort | Delou Pai |
| collection | DOAJ |
| description | Chemotaxis and the formation of suicidal neutrophil extracellular traps (suicidal NETosis) are key functions of polymorphonuclear cells (PMNs). Neutrophil extracellular traps in particular are known to be significantly involved in the severity of inflammatory and immunological disorders such as rheumatoid arthritis and Crohn’s disease. Therefore, detailed knowledge of PMNs is essential for analyzing the mechanisms involved in, and developing new therapies for, such diseases. To date, no standard method to analyze these cell activities has been established. This study used in vitro live cell imaging to simultaneously observe and analyze PMN functions. To demonstrate this, the effects of phorbol-12-myristat-13-acetat (PMA, 0.1-10 nM), N-formylmethionine-leucyl-phenylalanine (fMLP, 10 nM), and protein kinase C inhibitor 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H7) on PMN chemotaxis and suicidal NETosis were studied. PMA (1 nM-10 nM) resulted in significant concentration-dependent behavior in chemotaxis and an earlier onset of maximum oxidative burst and NET formation of up to 44%. When adding H7, PMA-triggered PMN functions were reduced, demonstrating that all three functions rely mostly on protein kinase C (PKC) activity, while PKC is not essential for fMLP-induced PMN activity. Thus, the method here described can be used to objectively quantify PMN functions and, especially through the regulation of the PKC pathway, could be useful in further clinical studies of immunological disorders. |
| format | Article |
| id | doaj-art-7a4a940ea61e419db2b19afc8dd40681 |
| institution | OA Journals |
| issn | 2314-8861 2314-7156 |
| language | English |
| publishDate | 2020-01-01 |
| publisher | Wiley |
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| series | Journal of Immunology Research |
| spelling | doaj-art-7a4a940ea61e419db2b19afc8dd406812025-08-20T02:07:35ZengWileyJournal of Immunology Research2314-88612314-71562020-01-01202010.1155/2020/14159471415947Polymorphonuclear Cell Chemotaxis and Suicidal NETosis: Simultaneous Observation Using fMLP, PMA, H7, and Live Cell ImagingDelou Pai0Michael Gruber1Sophie-Marie Pfaehler2Andre Bredthauer3Karla Lehle4Benedikt Trabold5Department of Anesthesiology, University Medical Center Regensburg, Franz-Josef-Strauß-Allee 11, 93053 Regensburg, GermanyDepartment of Anesthesiology, University Medical Center Regensburg, Franz-Josef-Strauß-Allee 11, 93053 Regensburg, GermanyDepartment of Anesthesiology, University Medical Center Regensburg, Franz-Josef-Strauß-Allee 11, 93053 Regensburg, GermanyDepartment of Anesthesiology, University Medical Center Regensburg, Franz-Josef-Strauß-Allee 11, 93053 Regensburg, GermanyDepartment of Cardiothoracic Surgery, University Medical Center Regensburg, Franz-Josef-Strauß-Allee 11, 93053 Regensburg, GermanyDepartment of Anesthesiology, University Medical Center Regensburg, Franz-Josef-Strauß-Allee 11, 93053 Regensburg, GermanyChemotaxis and the formation of suicidal neutrophil extracellular traps (suicidal NETosis) are key functions of polymorphonuclear cells (PMNs). Neutrophil extracellular traps in particular are known to be significantly involved in the severity of inflammatory and immunological disorders such as rheumatoid arthritis and Crohn’s disease. Therefore, detailed knowledge of PMNs is essential for analyzing the mechanisms involved in, and developing new therapies for, such diseases. To date, no standard method to analyze these cell activities has been established. This study used in vitro live cell imaging to simultaneously observe and analyze PMN functions. To demonstrate this, the effects of phorbol-12-myristat-13-acetat (PMA, 0.1-10 nM), N-formylmethionine-leucyl-phenylalanine (fMLP, 10 nM), and protein kinase C inhibitor 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H7) on PMN chemotaxis and suicidal NETosis were studied. PMA (1 nM-10 nM) resulted in significant concentration-dependent behavior in chemotaxis and an earlier onset of maximum oxidative burst and NET formation of up to 44%. When adding H7, PMA-triggered PMN functions were reduced, demonstrating that all three functions rely mostly on protein kinase C (PKC) activity, while PKC is not essential for fMLP-induced PMN activity. Thus, the method here described can be used to objectively quantify PMN functions and, especially through the regulation of the PKC pathway, could be useful in further clinical studies of immunological disorders.http://dx.doi.org/10.1155/2020/1415947 |
| spellingShingle | Delou Pai Michael Gruber Sophie-Marie Pfaehler Andre Bredthauer Karla Lehle Benedikt Trabold Polymorphonuclear Cell Chemotaxis and Suicidal NETosis: Simultaneous Observation Using fMLP, PMA, H7, and Live Cell Imaging Journal of Immunology Research |
| title | Polymorphonuclear Cell Chemotaxis and Suicidal NETosis: Simultaneous Observation Using fMLP, PMA, H7, and Live Cell Imaging |
| title_full | Polymorphonuclear Cell Chemotaxis and Suicidal NETosis: Simultaneous Observation Using fMLP, PMA, H7, and Live Cell Imaging |
| title_fullStr | Polymorphonuclear Cell Chemotaxis and Suicidal NETosis: Simultaneous Observation Using fMLP, PMA, H7, and Live Cell Imaging |
| title_full_unstemmed | Polymorphonuclear Cell Chemotaxis and Suicidal NETosis: Simultaneous Observation Using fMLP, PMA, H7, and Live Cell Imaging |
| title_short | Polymorphonuclear Cell Chemotaxis and Suicidal NETosis: Simultaneous Observation Using fMLP, PMA, H7, and Live Cell Imaging |
| title_sort | polymorphonuclear cell chemotaxis and suicidal netosis simultaneous observation using fmlp pma h7 and live cell imaging |
| url | http://dx.doi.org/10.1155/2020/1415947 |
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