mPTP opening differently affects electron transport chain and oxidative phosphorylation at succinate and NAD-dependent substrates oxidation in permeabilized rat hepatocytes
Mitochondrial Ca2+ overload may trigger the opening of mitochondrial permeability transition pore (mPTP) and its prolonged activation leads to cell death. ATP synthase is considered as a possible molecular component of the pore. The aim of this study was to investigate the state of oxidative phospho...
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National Academy of Sciences of Ukraine, Palladin Institute of Biochemistry
2020-08-01
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| Series: | The Ukrainian Biochemical Journal |
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| Online Access: | http://ukrbiochemjournal.org/wp-content/uploads/2020/09/Mazur_4_20.pdf |
| _version_ | 1827775529836609536 |
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| author | H. M. Mazur V. M. Merlavsky B. O. Manko V. V. Manko |
| author_facet | H. M. Mazur V. M. Merlavsky B. O. Manko V. V. Manko |
| author_sort | H. M. Mazur |
| collection | DOAJ |
| description | Mitochondrial Ca2+ overload may trigger the opening of mitochondrial permeability transition pore (mPTP) and its prolonged activation leads to cell death. ATP synthase is considered as a possible molecular component of the pore. The aim of this study was to investigate the state of oxidative phosphorylation at Ca2+-induced activation of mPTP in permeabilized hepatocytes. Hepatocytes were isolated by two-stage Seglen method. Permeabilization was performed using digitonin. Oxygen consumption rate was measured with Clark electrode. Oxidative phosphorylation was determined as the ratio of the ADP-stimulated respiration and substrate-stimulated respiration rates (ADP/S). It was established that increasing of Ca2+ concentration in the medium inhibited oligomycin effects and suppressed ADP- and FCCP-stimulated respiration upon succinate or glutamate, pyruvate and malate mixture oxidation. The mPTP inhibitor cyclosporin A did not directly affect respiration and oxidative phosphorylation after elevation of Ca2+ concentration and mPTP activation. When cyclosporine A was added before increasing Ca2+ concentration, the electron transport chain function (FCCP-stimulated respiration) was not impaired while the partial disruption of oxidative phosphorylation (ADP-stimulated respiration) was observed only upon succinate oxidation. The results obtained showed that inhibition of oxidative phosphorylation was the primary event in mPTP activation, possibly due to the involvement of ATP synthase in pore opening. In the case of NAD-dependent substrates oxidation that effect was stronger and faster than at succinate oxidation, due to the lower mitochondria energization. |
| first_indexed | 2024-03-11T13:50:46Z |
| format | Article |
| id | doaj.art-f6badd0acf924136b1c9037588ca7f32 |
| institution | Directory Open Access Journal |
| issn | 2409-4943 2413-5003 |
| language | English |
| last_indexed | 2024-03-11T13:50:46Z |
| publishDate | 2020-08-01 |
| publisher | National Academy of Sciences of Ukraine, Palladin Institute of Biochemistry |
| record_format | Article |
| series | The Ukrainian Biochemical Journal |
| spelling | doaj.art-f6badd0acf924136b1c9037588ca7f322023-11-02T09:07:53ZengNational Academy of Sciences of Ukraine, Palladin Institute of BiochemistryThe Ukrainian Biochemical Journal2409-49432413-50032020-08-01924142310.15407/ubj92.04.014mPTP opening differently affects electron transport chain and oxidative phosphorylation at succinate and NAD-dependent substrates oxidation in permeabilized rat hepatocytesH. M. Mazur 0V. M. Merlavsky 1B. O. Manko2https://orcid.org/0000-0002-6905-8580V. V. Manko3https://orcid.org/0000-0002-4406-1732Ivan Franko National University of Lviv, UkraineIvan Franko National University of Lviv, UkraineIvan Franko National University of Lviv, UkraineIvan Franko National University of Lviv, UkraineMitochondrial Ca2+ overload may trigger the opening of mitochondrial permeability transition pore (mPTP) and its prolonged activation leads to cell death. ATP synthase is considered as a possible molecular component of the pore. The aim of this study was to investigate the state of oxidative phosphorylation at Ca2+-induced activation of mPTP in permeabilized hepatocytes. Hepatocytes were isolated by two-stage Seglen method. Permeabilization was performed using digitonin. Oxygen consumption rate was measured with Clark electrode. Oxidative phosphorylation was determined as the ratio of the ADP-stimulated respiration and substrate-stimulated respiration rates (ADP/S). It was established that increasing of Ca2+ concentration in the medium inhibited oligomycin effects and suppressed ADP- and FCCP-stimulated respiration upon succinate or glutamate, pyruvate and malate mixture oxidation. The mPTP inhibitor cyclosporin A did not directly affect respiration and oxidative phosphorylation after elevation of Ca2+ concentration and mPTP activation. When cyclosporine A was added before increasing Ca2+ concentration, the electron transport chain function (FCCP-stimulated respiration) was not impaired while the partial disruption of oxidative phosphorylation (ADP-stimulated respiration) was observed only upon succinate oxidation. The results obtained showed that inhibition of oxidative phosphorylation was the primary event in mPTP activation, possibly due to the involvement of ATP synthase in pore opening. In the case of NAD-dependent substrates oxidation that effect was stronger and faster than at succinate oxidation, due to the lower mitochondria energization.http://ukrbiochemjournal.org/wp-content/uploads/2020/09/Mazur_4_20.pdffccphepatocytesmitochondrial permeability transition poreoxidative substratesсyclosporin a |
| spellingShingle | H. M. Mazur V. M. Merlavsky B. O. Manko V. V. Manko mPTP opening differently affects electron transport chain and oxidative phosphorylation at succinate and NAD-dependent substrates oxidation in permeabilized rat hepatocytes The Ukrainian Biochemical Journal fccp hepatocytes mitochondrial permeability transition pore oxidative substrates сyclosporin a |
| title | mPTP opening differently affects electron transport chain and oxidative phosphorylation at succinate and NAD-dependent substrates oxidation in permeabilized rat hepatocytes |
| title_full | mPTP opening differently affects electron transport chain and oxidative phosphorylation at succinate and NAD-dependent substrates oxidation in permeabilized rat hepatocytes |
| title_fullStr | mPTP opening differently affects electron transport chain and oxidative phosphorylation at succinate and NAD-dependent substrates oxidation in permeabilized rat hepatocytes |
| title_full_unstemmed | mPTP opening differently affects electron transport chain and oxidative phosphorylation at succinate and NAD-dependent substrates oxidation in permeabilized rat hepatocytes |
| title_short | mPTP opening differently affects electron transport chain and oxidative phosphorylation at succinate and NAD-dependent substrates oxidation in permeabilized rat hepatocytes |
| title_sort | mptp opening differently affects electron transport chain and oxidative phosphorylation at succinate and nad dependent substrates oxidation in permeabilized rat hepatocytes |
| topic | fccp hepatocytes mitochondrial permeability transition pore oxidative substrates сyclosporin a |
| url | http://ukrbiochemjournal.org/wp-content/uploads/2020/09/Mazur_4_20.pdf |
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