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Modification of DNA structure by reactive nitrogen species as a result of 2-methoxyestradiol–induced neuronal nitric oxide synthase uncoupling in metastatic osteosarcoma cells
2-methoxyestradiol (2-ME) is a physiological anticancer compound, metabolite of 17β-estradiol. Previously, our group evidenced that from mechanistic point of view one of anticancer mechanisms of action of 2-ME is specific induction and nuclear hijacking of neuronal nitric oxide synthase (nNOS), resu...
Autores principales: | , , , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Elsevier
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7162974/ https://www.ncbi.nlm.nih.gov/pubmed/32305006 http://dx.doi.org/10.1016/j.redox.2020.101522 |
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author | Gorska-Ponikowska, Magdalena Ploska, Agata Jacewicz, Dagmara Szkatula, Michal Barone, Giampaolo Lo Bosco, Giosuè Lo Celso, Fabrizio Dabrowska, Aleksandra M Kuban-Jankowska, Alicja Gorzynik-Debicka, Monika Knap, Narcyz Chmurzynski, Lech Dobrucki, Lawrence Wawrzyniec Kalinowski, Leszek Wozniak, Michal |
author_facet | Gorska-Ponikowska, Magdalena Ploska, Agata Jacewicz, Dagmara Szkatula, Michal Barone, Giampaolo Lo Bosco, Giosuè Lo Celso, Fabrizio Dabrowska, Aleksandra M Kuban-Jankowska, Alicja Gorzynik-Debicka, Monika Knap, Narcyz Chmurzynski, Lech Dobrucki, Lawrence Wawrzyniec Kalinowski, Leszek Wozniak, Michal |
author_sort | Gorska-Ponikowska, Magdalena |
collection | PubMed |
description | 2-methoxyestradiol (2-ME) is a physiological anticancer compound, metabolite of 17β-estradiol. Previously, our group evidenced that from mechanistic point of view one of anticancer mechanisms of action of 2-ME is specific induction and nuclear hijacking of neuronal nitric oxide synthase (nNOS), resulting in local generation of nitro-oxidative stress and finally, cancer cell death. The current study aims to establish the substantial mechanism of generation of reactive nitrogen species by 2-ME. We further achieved to identify the specific reactive nitrogen species involved in DNA-damaging mechanism of 2-ME. The study was performed using metastatic osteosarcoma 143B cells. We detected the release of biologically active (free) nitric oxide ((•)NO) with concurrent measurements of peroxynitrite (ONOO(−)) in real time in a single cell of 143B cell line by using (•)NO/ONOO(−) sensitive microsensors after stimulation with calcium ionophore. Detection of nitrogen dioxide ((•)NO(2)) and determination of chemical rate constants were carried out by a stopped-flow technique. The affinity of reactive nitrogen species toward the guanine base of DNA was evaluated by density functional theory calculations. Expression and localization of nuclear factor NF-kB was determined using imaging cytometry, while cell viability assay was evaluated by MTT assay. Herein, we presented that 2-ME triggers pro-apoptotic signalling cascade by increasing cellular reactive nitrogen species overproduction – a result of enzymatic uncoupling of increased nNOS protein levels. In particular, we proved that ONOO(−) and (•)NO(2) directly formed from peroxynitrous acid (ONOOH) and/or by auto-oxidation of (•)NO, are inducers of DNA damage in anticancer mechanism of 2-ME. Specifically, the affinity of reactive nitrogen species toward the guanine base of DNA, evaluated by density functional theory calculations, decreased in the order: ONOOH > ONOO(−) > (•)NO(2) > (•)NO. Therefore, we propose to consider the specific inducers of nNOS as an effective tool in the field of chemotherapy. |
format | Online Article Text |
id | pubmed-7162974 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | Elsevier |
record_format | MEDLINE/PubMed |
spelling | pubmed-71629742020-04-22 Modification of DNA structure by reactive nitrogen species as a result of 2-methoxyestradiol–induced neuronal nitric oxide synthase uncoupling in metastatic osteosarcoma cells Gorska-Ponikowska, Magdalena Ploska, Agata Jacewicz, Dagmara Szkatula, Michal Barone, Giampaolo Lo Bosco, Giosuè Lo Celso, Fabrizio Dabrowska, Aleksandra M Kuban-Jankowska, Alicja Gorzynik-Debicka, Monika Knap, Narcyz Chmurzynski, Lech Dobrucki, Lawrence Wawrzyniec Kalinowski, Leszek Wozniak, Michal Redox Biol Research Paper 2-methoxyestradiol (2-ME) is a physiological anticancer compound, metabolite of 17β-estradiol. Previously, our group evidenced that from mechanistic point of view one of anticancer mechanisms of action of 2-ME is specific induction and nuclear hijacking of neuronal nitric oxide synthase (nNOS), resulting in local generation of nitro-oxidative stress and finally, cancer cell death. The current study aims to establish the substantial mechanism of generation of reactive nitrogen species by 2-ME. We further achieved to identify the specific reactive nitrogen species involved in DNA-damaging mechanism of 2-ME. The study was performed using metastatic osteosarcoma 143B cells. We detected the release of biologically active (free) nitric oxide ((•)NO) with concurrent measurements of peroxynitrite (ONOO(−)) in real time in a single cell of 143B cell line by using (•)NO/ONOO(−) sensitive microsensors after stimulation with calcium ionophore. Detection of nitrogen dioxide ((•)NO(2)) and determination of chemical rate constants were carried out by a stopped-flow technique. The affinity of reactive nitrogen species toward the guanine base of DNA was evaluated by density functional theory calculations. Expression and localization of nuclear factor NF-kB was determined using imaging cytometry, while cell viability assay was evaluated by MTT assay. Herein, we presented that 2-ME triggers pro-apoptotic signalling cascade by increasing cellular reactive nitrogen species overproduction – a result of enzymatic uncoupling of increased nNOS protein levels. In particular, we proved that ONOO(−) and (•)NO(2) directly formed from peroxynitrous acid (ONOOH) and/or by auto-oxidation of (•)NO, are inducers of DNA damage in anticancer mechanism of 2-ME. Specifically, the affinity of reactive nitrogen species toward the guanine base of DNA, evaluated by density functional theory calculations, decreased in the order: ONOOH > ONOO(−) > (•)NO(2) > (•)NO. Therefore, we propose to consider the specific inducers of nNOS as an effective tool in the field of chemotherapy. Elsevier 2020-03-28 /pmc/articles/PMC7162974/ /pubmed/32305006 http://dx.doi.org/10.1016/j.redox.2020.101522 Text en © 2020 The Authors http://creativecommons.org/licenses/by/4.0/ This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Research Paper Gorska-Ponikowska, Magdalena Ploska, Agata Jacewicz, Dagmara Szkatula, Michal Barone, Giampaolo Lo Bosco, Giosuè Lo Celso, Fabrizio Dabrowska, Aleksandra M Kuban-Jankowska, Alicja Gorzynik-Debicka, Monika Knap, Narcyz Chmurzynski, Lech Dobrucki, Lawrence Wawrzyniec Kalinowski, Leszek Wozniak, Michal Modification of DNA structure by reactive nitrogen species as a result of 2-methoxyestradiol–induced neuronal nitric oxide synthase uncoupling in metastatic osteosarcoma cells |
title | Modification of DNA structure by reactive nitrogen species as a result of 2-methoxyestradiol–induced neuronal nitric oxide synthase uncoupling in metastatic osteosarcoma cells |
title_full | Modification of DNA structure by reactive nitrogen species as a result of 2-methoxyestradiol–induced neuronal nitric oxide synthase uncoupling in metastatic osteosarcoma cells |
title_fullStr | Modification of DNA structure by reactive nitrogen species as a result of 2-methoxyestradiol–induced neuronal nitric oxide synthase uncoupling in metastatic osteosarcoma cells |
title_full_unstemmed | Modification of DNA structure by reactive nitrogen species as a result of 2-methoxyestradiol–induced neuronal nitric oxide synthase uncoupling in metastatic osteosarcoma cells |
title_short | Modification of DNA structure by reactive nitrogen species as a result of 2-methoxyestradiol–induced neuronal nitric oxide synthase uncoupling in metastatic osteosarcoma cells |
title_sort | modification of dna structure by reactive nitrogen species as a result of 2-methoxyestradiol–induced neuronal nitric oxide synthase uncoupling in metastatic osteosarcoma cells |
topic | Research Paper |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7162974/ https://www.ncbi.nlm.nih.gov/pubmed/32305006 http://dx.doi.org/10.1016/j.redox.2020.101522 |
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