Mitochondrial Superoxide Production Decreases on Glucose-Stimulated Insulin Secretion in Pancreatic β Cells Due to Decreasing Mitochondrial Matrix NADH/NAD + Ratio
Glucose-stimulated insulin secretion (GSIS) in pancreatic β cells was expected to enhance mitochondrial superoxide formation. Hence, we elucidated relevant redox equilibria. Unexpectedly, INS-1E cells at transitions from 3 (11 m ; pancreatic islets from 5 m ) to 25 m glucose decreased matrix superox...
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| Veröffentlicht in: | Antioxidants & redox signaling 2020-10, Vol.33 (12), p.789-815 |
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| creator | Plecitá-Hlavatá, Lydie Engstová, Hana Holendová, Blanka Tauber, Jan Špaček, Tomáš Petrásková, Lucie Křen, Vladimír Špačková, Jitka Gotvaldová, Klára Ježek, Jan Dlasková, Andrea Smolková, Katarína Ježek, Petr |
| description | Glucose-stimulated insulin secretion (GSIS) in pancreatic β cells was expected to enhance mitochondrial superoxide formation. Hence, we elucidated relevant redox equilibria.
Unexpectedly, INS-1E cells at transitions from 3 (11 m
; pancreatic islets from 5 m
) to 25 m
glucose decreased matrix superoxide release rates (MitoSOX Red monitoring validated by MitoB) and H
O
(mitoHyPer, subtracting mitoSypHer emission). Novel double-channel fluorescence lifetime imaging, approximating free mitochondrial matrix NADH
indicated its ∼20% decrease. Matrix NAD
increased on GSIS, indicated by the FAD-emission lifetime decrease, reflecting higher quenching of FAD by NAD
. The participation of pyruvate/malate and pyruvate/citrate redox shuttles, elevating cytosolic NADPH
(iNAP1 fluorescence monitoring) at the expense of matrix NADH
, was indicated, using citrate (2-oxoglutarate) carrier inhibitors and cytosolic malic enzyme silencing: All changes vanished on these manipulations.
C-incorporation from
C-L-glutamine into
C-citrate reflected the pyruvate/isocitrate shuttle. Matrix NADPH
(iNAP3 monitored) decreased. With decreasing glucose, the suppressor of Complex III site Q electron leak (S3QEL) suppressor caused a higher Complex I I
site contribution, but a lower superoxide fraction ascribed to the Complex III site III
. Thus, the diminished matrix NADH
/NAD
decreased Complex I flavin site I
superoxide formation on GSIS.
Mutually validated methods showed decreasing superoxide release into the mitochondrial matrix in pancreatic β cells on GSIS, due to the decreasing matrix NADH
/NAD
(NADPH
/NADP
) at increasing cytosolic NADPH
levels. The developed innovative methods enable real-time NADH/NAD
and NADPH/NADP
monitoring in any distinct cell compartment.
The export of reducing equivalents from mitochondria adjusts lower mitochondrial superoxide production on GSIS, but it does not prevent oxidative stress in pancreatic β cells. |
| doi_str_mv | 10.1089/ars.2019.7800 |
| format | Article |
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Unexpectedly, INS-1E cells at transitions from 3 (11 m
; pancreatic islets from 5 m
) to 25 m
glucose decreased matrix superoxide release rates (MitoSOX Red monitoring validated by MitoB) and H
O
(mitoHyPer, subtracting mitoSypHer emission). Novel double-channel fluorescence lifetime imaging, approximating free mitochondrial matrix NADH
indicated its ∼20% decrease. Matrix NAD
increased on GSIS, indicated by the FAD-emission lifetime decrease, reflecting higher quenching of FAD by NAD
. The participation of pyruvate/malate and pyruvate/citrate redox shuttles, elevating cytosolic NADPH
(iNAP1 fluorescence monitoring) at the expense of matrix NADH
, was indicated, using citrate (2-oxoglutarate) carrier inhibitors and cytosolic malic enzyme silencing: All changes vanished on these manipulations.
C-incorporation from
C-L-glutamine into
C-citrate reflected the pyruvate/isocitrate shuttle. Matrix NADPH
(iNAP3 monitored) decreased. With decreasing glucose, the suppressor of Complex III site Q electron leak (S3QEL) suppressor caused a higher Complex I I
site contribution, but a lower superoxide fraction ascribed to the Complex III site III
. Thus, the diminished matrix NADH
/NAD
decreased Complex I flavin site I
superoxide formation on GSIS.
Mutually validated methods showed decreasing superoxide release into the mitochondrial matrix in pancreatic β cells on GSIS, due to the decreasing matrix NADH
/NAD
(NADPH
/NADP
) at increasing cytosolic NADPH
levels. The developed innovative methods enable real-time NADH/NAD
and NADPH/NADP
monitoring in any distinct cell compartment.
The export of reducing equivalents from mitochondria adjusts lower mitochondrial superoxide production on GSIS, but it does not prevent oxidative stress in pancreatic β cells.</description><identifier>ISSN: 1523-0864</identifier><identifier>EISSN: 1557-7716</identifier><identifier>DOI: 10.1089/ars.2019.7800</identifier><identifier>PMID: 32517485</identifier><language>eng</language><publisher>United States: Mary Ann Liebert, Inc</publisher><subject>Adenosine Triphosphate - metabolism ; Animals ; Beta cells ; Cell Respiration ; Chromatography, Liquid ; Citric acid ; Citric Acid - metabolism ; Emission ; Energy Metabolism ; Flavin ; Flavin-adenine dinucleotide ; Flavin-Adenine Dinucleotide - metabolism ; Fluorescence ; Forum Original Research Communication ; Glucose ; Glucose - metabolism ; Glutamine ; Hydrogen peroxide ; Hydrogen Peroxide - metabolism ; Insulin ; Insulin Secretion ; Insulin-Secreting Cells - metabolism ; Ketoglutaric acid ; Malate ; Malic enzyme ; Mass Spectrometry ; Membrane Potential, Mitochondrial ; Metabolic Networks and Pathways ; Metabolomics - methods ; Mitochondria ; Mitochondria - metabolism ; Monitoring ; NAD ; NAD - metabolism ; NADH ; NADP ; Nicotinamide adenine dinucleotide ; Oxidative stress ; Pancreas ; Pyruvic acid ; Rats ; Secretion ; Signal Transduction ; Superoxide ; Superoxides - metabolism</subject><ispartof>Antioxidants & redox signaling, 2020-10, Vol.33 (12), p.789-815</ispartof><rights>Copyright Mary Ann Liebert, Inc. Oct 20, 2020</rights><rights>Lydie Plecitá-Hlavatá et al. 2020; Published by Mary Ann Liebert, Inc. 2020 Lydie Plecitá-Hlavatá et al.</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c345t-4a5d26aabe3bf627e480b57b5a6a102df06c47de3fc72d5294558378370393a73</citedby><cites>FETCH-LOGICAL-c345t-4a5d26aabe3bf627e480b57b5a6a102df06c47de3fc72d5294558378370393a73</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>230,314,776,780,881,27903,27904</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/32517485$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Plecitá-Hlavatá, Lydie</creatorcontrib><creatorcontrib>Engstová, Hana</creatorcontrib><creatorcontrib>Holendová, Blanka</creatorcontrib><creatorcontrib>Tauber, Jan</creatorcontrib><creatorcontrib>Špaček, Tomáš</creatorcontrib><creatorcontrib>Petrásková, Lucie</creatorcontrib><creatorcontrib>Křen, Vladimír</creatorcontrib><creatorcontrib>Špačková, Jitka</creatorcontrib><creatorcontrib>Gotvaldová, Klára</creatorcontrib><creatorcontrib>Ježek, Jan</creatorcontrib><creatorcontrib>Dlasková, Andrea</creatorcontrib><creatorcontrib>Smolková, Katarína</creatorcontrib><creatorcontrib>Ježek, Petr</creatorcontrib><title>Mitochondrial Superoxide Production Decreases on Glucose-Stimulated Insulin Secretion in Pancreatic β Cells Due to Decreasing Mitochondrial Matrix NADH/NAD + Ratio</title><title>Antioxidants & redox signaling</title><addtitle>Antioxid Redox Signal</addtitle><description>Glucose-stimulated insulin secretion (GSIS) in pancreatic β cells was expected to enhance mitochondrial superoxide formation. Hence, we elucidated relevant redox equilibria.
Unexpectedly, INS-1E cells at transitions from 3 (11 m
; pancreatic islets from 5 m
) to 25 m
glucose decreased matrix superoxide release rates (MitoSOX Red monitoring validated by MitoB) and H
O
(mitoHyPer, subtracting mitoSypHer emission). Novel double-channel fluorescence lifetime imaging, approximating free mitochondrial matrix NADH
indicated its ∼20% decrease. Matrix NAD
increased on GSIS, indicated by the FAD-emission lifetime decrease, reflecting higher quenching of FAD by NAD
. The participation of pyruvate/malate and pyruvate/citrate redox shuttles, elevating cytosolic NADPH
(iNAP1 fluorescence monitoring) at the expense of matrix NADH
, was indicated, using citrate (2-oxoglutarate) carrier inhibitors and cytosolic malic enzyme silencing: All changes vanished on these manipulations.
C-incorporation from
C-L-glutamine into
C-citrate reflected the pyruvate/isocitrate shuttle. Matrix NADPH
(iNAP3 monitored) decreased. With decreasing glucose, the suppressor of Complex III site Q electron leak (S3QEL) suppressor caused a higher Complex I I
site contribution, but a lower superoxide fraction ascribed to the Complex III site III
. Thus, the diminished matrix NADH
/NAD
decreased Complex I flavin site I
superoxide formation on GSIS.
Mutually validated methods showed decreasing superoxide release into the mitochondrial matrix in pancreatic β cells on GSIS, due to the decreasing matrix NADH
/NAD
(NADPH
/NADP
) at increasing cytosolic NADPH
levels. The developed innovative methods enable real-time NADH/NAD
and NADPH/NADP
monitoring in any distinct cell compartment.
The export of reducing equivalents from mitochondria adjusts lower mitochondrial superoxide production on GSIS, but it does not prevent oxidative stress in pancreatic β cells.</description><subject>Adenosine Triphosphate - metabolism</subject><subject>Animals</subject><subject>Beta cells</subject><subject>Cell Respiration</subject><subject>Chromatography, Liquid</subject><subject>Citric acid</subject><subject>Citric Acid - metabolism</subject><subject>Emission</subject><subject>Energy Metabolism</subject><subject>Flavin</subject><subject>Flavin-adenine dinucleotide</subject><subject>Flavin-Adenine Dinucleotide - metabolism</subject><subject>Fluorescence</subject><subject>Forum Original Research Communication</subject><subject>Glucose</subject><subject>Glucose - metabolism</subject><subject>Glutamine</subject><subject>Hydrogen peroxide</subject><subject>Hydrogen Peroxide - metabolism</subject><subject>Insulin</subject><subject>Insulin Secretion</subject><subject>Insulin-Secreting Cells - metabolism</subject><subject>Ketoglutaric acid</subject><subject>Malate</subject><subject>Malic enzyme</subject><subject>Mass Spectrometry</subject><subject>Membrane Potential, Mitochondrial</subject><subject>Metabolic Networks and Pathways</subject><subject>Metabolomics - methods</subject><subject>Mitochondria</subject><subject>Mitochondria - metabolism</subject><subject>Monitoring</subject><subject>NAD</subject><subject>NAD - metabolism</subject><subject>NADH</subject><subject>NADP</subject><subject>Nicotinamide adenine dinucleotide</subject><subject>Oxidative stress</subject><subject>Pancreas</subject><subject>Pyruvic acid</subject><subject>Rats</subject><subject>Secretion</subject><subject>Signal Transduction</subject><subject>Superoxide</subject><subject>Superoxides - metabolism</subject><issn>1523-0864</issn><issn>1557-7716</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2020</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpdkdtqFTEUhoMo9qCX3krAG6HMbo6T2TdC2bu2hVZLd70OmSTTpsxOtjmU-j4-gQ_iM5mxB6wQkrXIt_5krR-AdxjNMOrm-yqmGUF4PhMdQi_ANuZcNELg9uUUE9qgrmVbYCelG4QQwRi9BluUcCxYx7fBzzOXg74O3kSnRrgqGxvDnTMWnsdgis4ueLi0OlqVbII1ORqLDsk2q-zWZVTZGnjiUxmdh6uJ-1tRk3Plp6rsNPz9Cy7sOCa4LBbm8Kjn_BV8_vyZytHdwS8Hy-P9usE9eFEFwhvwalBjsm8fzl3w7fPh5eK4Of16dLI4OG00ZTw3THFDWqV6S_uhJcKyDvVc9Fy1CiNiBtRqJoylgxbEcDJnnHdU1IXonCpBd8Gne91N6dfWaOtzVKPcRLdW8YcMysnnN95dy6twK-ssSR15Ffj4IBDD92JTlmuXdG1deRtKkoTh6g8XCFf0w3_oTSjR1_YqxQghiPNJsLmndAwpRTs8fQYjOfkvq_9y8l9O_lf-_b8dPNGPhtM_PhiujA</recordid><startdate>20201020</startdate><enddate>20201020</enddate><creator>Plecitá-Hlavatá, Lydie</creator><creator>Engstová, Hana</creator><creator>Holendová, Blanka</creator><creator>Tauber, Jan</creator><creator>Špaček, Tomáš</creator><creator>Petrásková, Lucie</creator><creator>Křen, Vladimír</creator><creator>Špačková, Jitka</creator><creator>Gotvaldová, Klára</creator><creator>Ježek, Jan</creator><creator>Dlasková, Andrea</creator><creator>Smolková, Katarína</creator><creator>Ježek, Petr</creator><general>Mary Ann Liebert, Inc</general><general>Mary Ann Liebert, Inc., publishers</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QL</scope><scope>7QP</scope><scope>7T5</scope><scope>7TK</scope><scope>7TM</scope><scope>7TO</scope><scope>7U9</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>H94</scope><scope>P64</scope><scope>RC3</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>20201020</creationdate><title>Mitochondrial Superoxide Production Decreases on Glucose-Stimulated Insulin Secretion in Pancreatic β Cells Due to Decreasing Mitochondrial Matrix NADH/NAD + Ratio</title><author>Plecitá-Hlavatá, Lydie ; Engstová, Hana ; Holendová, Blanka ; Tauber, Jan ; Špaček, Tomáš ; Petrásková, Lucie ; Křen, Vladimír ; Špačková, Jitka ; Gotvaldová, Klára ; Ježek, Jan ; Dlasková, Andrea ; Smolková, Katarína ; Ježek, Petr</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c345t-4a5d26aabe3bf627e480b57b5a6a102df06c47de3fc72d5294558378370393a73</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2020</creationdate><topic>Adenosine Triphosphate - metabolism</topic><topic>Animals</topic><topic>Beta cells</topic><topic>Cell Respiration</topic><topic>Chromatography, Liquid</topic><topic>Citric acid</topic><topic>Citric Acid - metabolism</topic><topic>Emission</topic><topic>Energy Metabolism</topic><topic>Flavin</topic><topic>Flavin-adenine dinucleotide</topic><topic>Flavin-Adenine Dinucleotide - metabolism</topic><topic>Fluorescence</topic><topic>Forum Original Research Communication</topic><topic>Glucose</topic><topic>Glucose - metabolism</topic><topic>Glutamine</topic><topic>Hydrogen peroxide</topic><topic>Hydrogen Peroxide - metabolism</topic><topic>Insulin</topic><topic>Insulin Secretion</topic><topic>Insulin-Secreting Cells - metabolism</topic><topic>Ketoglutaric acid</topic><topic>Malate</topic><topic>Malic enzyme</topic><topic>Mass Spectrometry</topic><topic>Membrane Potential, Mitochondrial</topic><topic>Metabolic Networks and Pathways</topic><topic>Metabolomics - methods</topic><topic>Mitochondria</topic><topic>Mitochondria - metabolism</topic><topic>Monitoring</topic><topic>NAD</topic><topic>NAD - metabolism</topic><topic>NADH</topic><topic>NADP</topic><topic>Nicotinamide adenine dinucleotide</topic><topic>Oxidative stress</topic><topic>Pancreas</topic><topic>Pyruvic acid</topic><topic>Rats</topic><topic>Secretion</topic><topic>Signal Transduction</topic><topic>Superoxide</topic><topic>Superoxides - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Plecitá-Hlavatá, Lydie</creatorcontrib><creatorcontrib>Engstová, Hana</creatorcontrib><creatorcontrib>Holendová, Blanka</creatorcontrib><creatorcontrib>Tauber, Jan</creatorcontrib><creatorcontrib>Špaček, Tomáš</creatorcontrib><creatorcontrib>Petrásková, Lucie</creatorcontrib><creatorcontrib>Křen, Vladimír</creatorcontrib><creatorcontrib>Špačková, Jitka</creatorcontrib><creatorcontrib>Gotvaldová, Klára</creatorcontrib><creatorcontrib>Ježek, Jan</creatorcontrib><creatorcontrib>Dlasková, Andrea</creatorcontrib><creatorcontrib>Smolková, Katarína</creatorcontrib><creatorcontrib>Ježek, Petr</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Calcium & Calcified Tissue Abstracts</collection><collection>Immunology Abstracts</collection><collection>Neurosciences Abstracts</collection><collection>Nucleic Acids Abstracts</collection><collection>Oncogenes and Growth Factors Abstracts</collection><collection>Virology and AIDS Abstracts</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>Engineering Research Database</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Genetics Abstracts</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Antioxidants & redox signaling</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Plecitá-Hlavatá, Lydie</au><au>Engstová, Hana</au><au>Holendová, Blanka</au><au>Tauber, Jan</au><au>Špaček, Tomáš</au><au>Petrásková, Lucie</au><au>Křen, Vladimír</au><au>Špačková, Jitka</au><au>Gotvaldová, Klára</au><au>Ježek, Jan</au><au>Dlasková, Andrea</au><au>Smolková, Katarína</au><au>Ježek, Petr</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Mitochondrial Superoxide Production Decreases on Glucose-Stimulated Insulin Secretion in Pancreatic β Cells Due to Decreasing Mitochondrial Matrix NADH/NAD + Ratio</atitle><jtitle>Antioxidants & redox signaling</jtitle><addtitle>Antioxid Redox Signal</addtitle><date>2020-10-20</date><risdate>2020</risdate><volume>33</volume><issue>12</issue><spage>789</spage><epage>815</epage><pages>789-815</pages><issn>1523-0864</issn><eissn>1557-7716</eissn><abstract>Glucose-stimulated insulin secretion (GSIS) in pancreatic β cells was expected to enhance mitochondrial superoxide formation. Hence, we elucidated relevant redox equilibria.
Unexpectedly, INS-1E cells at transitions from 3 (11 m
; pancreatic islets from 5 m
) to 25 m
glucose decreased matrix superoxide release rates (MitoSOX Red monitoring validated by MitoB) and H
O
(mitoHyPer, subtracting mitoSypHer emission). Novel double-channel fluorescence lifetime imaging, approximating free mitochondrial matrix NADH
indicated its ∼20% decrease. Matrix NAD
increased on GSIS, indicated by the FAD-emission lifetime decrease, reflecting higher quenching of FAD by NAD
. The participation of pyruvate/malate and pyruvate/citrate redox shuttles, elevating cytosolic NADPH
(iNAP1 fluorescence monitoring) at the expense of matrix NADH
, was indicated, using citrate (2-oxoglutarate) carrier inhibitors and cytosolic malic enzyme silencing: All changes vanished on these manipulations.
C-incorporation from
C-L-glutamine into
C-citrate reflected the pyruvate/isocitrate shuttle. Matrix NADPH
(iNAP3 monitored) decreased. With decreasing glucose, the suppressor of Complex III site Q electron leak (S3QEL) suppressor caused a higher Complex I I
site contribution, but a lower superoxide fraction ascribed to the Complex III site III
. Thus, the diminished matrix NADH
/NAD
decreased Complex I flavin site I
superoxide formation on GSIS.
Mutually validated methods showed decreasing superoxide release into the mitochondrial matrix in pancreatic β cells on GSIS, due to the decreasing matrix NADH
/NAD
(NADPH
/NADP
) at increasing cytosolic NADPH
levels. The developed innovative methods enable real-time NADH/NAD
and NADPH/NADP
monitoring in any distinct cell compartment.
The export of reducing equivalents from mitochondria adjusts lower mitochondrial superoxide production on GSIS, but it does not prevent oxidative stress in pancreatic β cells.</abstract><cop>United States</cop><pub>Mary Ann Liebert, Inc</pub><pmid>32517485</pmid><doi>10.1089/ars.2019.7800</doi><tpages>27</tpages><oa>free_for_read</oa></addata></record> |
| fulltext | fulltext |
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| ispartof | Antioxidants & redox signaling, 2020-10, Vol.33 (12), p.789-815 |
| issn | 1523-0864 1557-7716 |
| language | eng |
| recordid | cdi_pubmedcentral_primary_oai_pubmedcentral_nih_gov_7482716 |
| source | MEDLINE; Alma/SFX Local Collection |
| subjects | Adenosine Triphosphate - metabolism Animals Beta cells Cell Respiration Chromatography, Liquid Citric acid Citric Acid - metabolism Emission Energy Metabolism Flavin Flavin-adenine dinucleotide Flavin-Adenine Dinucleotide - metabolism Fluorescence Forum Original Research Communication Glucose Glucose - metabolism Glutamine Hydrogen peroxide Hydrogen Peroxide - metabolism Insulin Insulin Secretion Insulin-Secreting Cells - metabolism Ketoglutaric acid Malate Malic enzyme Mass Spectrometry Membrane Potential, Mitochondrial Metabolic Networks and Pathways Metabolomics - methods Mitochondria Mitochondria - metabolism Monitoring NAD NAD - metabolism NADH NADP Nicotinamide adenine dinucleotide Oxidative stress Pancreas Pyruvic acid Rats Secretion Signal Transduction Superoxide Superoxides - metabolism |
| title | Mitochondrial Superoxide Production Decreases on Glucose-Stimulated Insulin Secretion in Pancreatic β Cells Due to Decreasing Mitochondrial Matrix NADH/NAD + Ratio |
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