Transcript levels for selected ATP synthase membrane FO-subunits-including DAPIT-in INS-1E cells were found to be sensitive to lowering glucose down from 11 mM, in which these cells are routinely cultured. Depending on conditions, the diminished mRNA levels recovered when glucose was restored to 11 mM; or were elevated during further 120 min incubations with 20-mM glucose. Asking whether DAPIT expression may be elevated by hyperglycemia in vivo, we studied mice with hyaluronic acid implants delivering glucose for up to 14 days. Such continuous two-week glucose stimulations in mice increased DAPIT mRNA by >5-fold in isolated pancreatic islets (ATP synthase F1α mRNA by 1.5-fold). In INS-1E cells, the glucose-induced ATP increment vanished with DAPIT silencing (6% of ATP rise), likewise a portion of the mtDNA-copy number increment. With 20 and 11-mM glucose the phosphorylating/non-phosphorylating respiration rate ratio diminished to ~70% and 96%, respectively, upon DAPIT silencing, whereas net GSIS rates accounted for 80% and 90% in USMG5/DAPIT-deficient cells. Consequently, the sufficient DAPIT expression and complete ATP synthase assembly is required for maximum ATP synthesis and mitochondrial biogenesis, but not for insulin secretion as such. Elevated DAPIT expression at high glucose further increases the ATP synthesis efficiency.

• MeSH
• adenosintrifosfát metabolismus   MeSH
• beta-buňky cytologie   účinky léků   metabolismus   MeSH
• buněčné kultury MeSH
• buněčné linie MeSH
• glukosa aplikace a dávkování   farmakologie   MeSH
• konformace proteinů MeSH
• krysa rodu rattus MeSH
• kyselina hyaluronová chemie   MeSH
• membránové proteiny chemie   genetika   metabolismus   MeSH
• mitochondriální DNA účinky léků   genetika   MeSH
• mitochondrie účinky léků   genetika   metabolismus   MeSH
• molekulární modely MeSH
• myši MeSH
• upregulace * MeSH
• variabilita počtu kopií segmentů DNA účinky léků   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Methylene blue (MB) is a promising compound with a broad range of neuroprotective activity. One of therapeutic effects is the activation of mitochondrial biogenesis via Nrf2/ARE signaling cascade. Probably, mild oxidative stress caused by MB-depended H2O2 production is a trigger for activation of this signaling cascade. So mechanistically, MB can be regarded as prooxidant. We investigated the dose-dependent H2O2 production in intact brain mitochondria and showed the increase in the H2O2 production after adding as little as 50 nM MB. We have not found genotoxic effect of therapeutic concentration of MB to mitochondrial genome. 100 μM MB selectively damaged fragments of mitochondrial DNA, which correlated with the number of purine-T-G-purine (RTGR)-sequences in studied fragments. Furthermore, 20 μM MB combined with the red light caused the formation of singlet oxygen, which strongly damaged mitochondrial DNA in all studied fragments. We did not observe mitochondrial DNA lesions in brain after single intraperitoneal injection of MB in the concentration of 50 mg/kg. Furthermore, we showed the neuroprotective properties of MB pretreatments after rotenone injection. Therefore, we suggest that MB-induced mild oxidative stress does not have genotoxic effect on mitochondrial DNA.

• MeSH
• insekticidy škodlivé účinky   MeSH
• laboratorní zvířata MeSH
• methylenová modř farmakologie   terapeutické užití   MeSH
• mitochondriální DNA účinky léků   MeSH
• mitochondrie účinky léků   MeSH
• modely nemocí na zvířatech MeSH
• mozek metabolismus   účinky léků   ultrastruktura   MeSH
• myši MeSH
• neuroprotektivní látky MeSH
• peroxid vodíku farmakologie   metabolismus   MeSH
• poškození DNA genetika   účinky léků   MeSH
• rotenon * škodlivé účinky   MeSH
• techniky in vitro MeSH
• Check Tag
• myši MeSH
• Publikační typ
• práce podpořená grantem MeSH

• MeSH
• léčivé přípravky metabolismus   MeSH
• lidé MeSH
• metabolické vedlejší účinky léčivých látek MeSH
• mitochondriální DNA * účinky léků   MeSH
• mitochondrie * genetika   metabolismus   účinky léků   MeSH
• oxidativní fosforylace účinky léků   MeSH
• transport elektronů účinky léků   MeSH
• výměna plynů v plicích účinky léků   MeSH
• Check Tag
• lidé MeSH

Dichloroacetate (DCA) is beneficial in cancer therapy because it induces apoptosis and decreases cancer growth in vitro and in vivo without affecting non-cancer cells. DCA stimulates the activity of the enzyme pyruvate dehydrogenase by inhibiting pyruvate dehydrogenase kinase. Consequently, DCA promotes oxidative phosphorylation after glycolysis. Therefore, DCA produces changes in energy metabolism that could affect the mitochondrial network and mitophagy. This investigation determined the effects of DCA treatment on mitophagy in human neuroblastoma SH-SY5Y cells. SH-SY5Y cells were cultured and distributed into 3 groups: control, NH4Cl and chloroquine. Each group was treated with DCA at 0, 5, 30 and 60 mM for 16 h. Samples were analyzed for cell viability, mtDNA copy number, mitochondrial network morphology and expression of key proteins involved in mitochondrial dynamics, such as LC3b, FIS1, OPA1, PARKIN and PINK1. In all groups, DCA caused a decrease in cell viability, an induction of autophagy in a dose-dependent manner and a decrease in Tim23, FIS1 and PARKIN protein expression, leading to profound morphological changes in the mitochondrial network resulting in shorter and more fragmented filaments. However, TFAM protein levels remained unchanged. Similarly, the mitochondrial copy number was not significantly different among the treatment groups. In conclusion, DCA induces mitophagy and remodeling of the mitochondrial network. In this remodeling, DCA induces a decrease in the expression of key proteins involved in protein degradation and mitochondrial dynamics but does not significantly affect the mtDNA density. Blocking late phase autophagy increases the effects of DCA, suggesting that autophagy protects the cell, at least partially, against DCA.

• MeSH
• autofagie účinky léků   MeSH
• chlorochin farmakologie   MeSH
• kyselina dichloroctová farmakologie   MeSH
• lidé MeSH
• mitochondriální DNA účinky léků   MeSH
• mitochondriální proteiny metabolismus   MeSH
• mitofagie účinky léků   MeSH
• nádorové buněčné linie MeSH
• neuroblastom genetika   metabolismus   patologie   MeSH
• regulace genové exprese u nádorů účinky léků   MeSH
• variabilita počtu kopií segmentů DNA MeSH
• viabilita buněk účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

BACKGROUND/AIMS: Deleterious effects of saturated fatty acids in skeletal muscle cells are well known but their impact on mitochondrial respiration has not been well studied. Mitochondrial oxidative damage has been implicated to play a role in their effect. The purpose of this study was to evaluate viability, mtDNA integrity and mitochondrial respiration in C2C12 myoblasts and myotubes exposed to palmitate and to test the effect of mitochondria-targeted antioxidants MitoQ and MitoTEMPOL in preventing palmitate-induced damage. METHODS: Cells were treated with tested compounds, mtDNA damage was detected by quantitative PCR and mitochondrial respiration was measured using an extracellular flux analyzer XF24. RESULTS: Palmitate caused mtDNA damage, which was associated with reduced mitochondrial respiration and cell death in myoblasts but not in myotubes. MitoTEMPOL was able to prevent palmitate-induced mtDNA damage in myoblasts but failed to prevent cell death. MitoQ did not show any protective effect and both compounds markedly inhibited mitochondrial respiration. CONCLUSION: Our results indicate that skeletal muscle progenitor cells could be the first target of the deleterious action of palmitate, as myoblasts appeared to be more sensitive to its effects than myotubes possibly in part due to a lower spare respiratory capacity in the former. Only MitoTEMPOL prevented palmitate-induced mtDNA damage but neither antioxidant was able to prevent cell death and both antioxidants had a marked negative effect on respiration.

• MeSH
• antioxidancia metabolismus   MeSH
• buněčná smrt účinky léků   MeSH
• kultivované buňky MeSH
• mitochondriální DNA účinky léků   metabolismus   MeSH
• mitochondrie účinky léků   metabolismus   patologie   MeSH
• myoblasty účinky léků   patologie   MeSH
• myši MeSH
• palmitany farmakologie   MeSH
• poškození DNA MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

• MeSH
• amidy chemie   imunologie   MeSH
• antifungální látky chemie   imunologie   MeSH
• chloramfenikol chemie   imunologie   MeSH
• cyklické peptidy chemie   imunologie   MeSH
• finanční podpora výzkumu jako téma MeSH
• kofein chemie   imunologie   MeSH
• kvasinky účinky léků   MeSH
• mitochondriální DNA účinky léků   MeSH
• mitochondrie fyziologie   účinky léků   MeSH
• Saccharomyces cerevisiae účinky léků   MeSH