The effects of AICAR and rapamycin on mitochondrial function in immortalized mitochondrial DNA mutator murine embryonic fibroblasts

• Published in: Biology open 2018, Vol.7 (11)
• Main Authors: Delic, Vedad, Noble, Kenyaria, Zivkovic, Sandra, Phan, Tam-Anh, Reynes, Christian, Zhang, Yumeng, Phillips, Oluwakemi, Claybaker, Charles, Ta, Yen, Dinh, Vinh B, Cruz, Josean, Prolla, Tomas A, Bradshaw, Patrick C
• Format: Article
• Language: English
• Subjects: Aging; AMP kinase; AMP-activated protein kinase; DNA polymerase; DNA-directed DNA polymerase; Electron transport; Embryo fibroblasts; Exonuclease; Fibroblasts; Kinases; Mitochondria; Mitochondrial DNA; mTOR; Mutation; Oxygen consumption; Phenotypes; Pyruvate addiction; Pyruvic acid; Rapamycin; Reactive oxygen species; Respiration; Signaling; Stem cells; TOR protein
• ISSN: 2046-6390; 2046-6390
• DOI: 10.1242/bio.033852

Mitochondrial DNA mutations accumulate with age and may play a role in stem cell aging as suggested by the premature aging phenotype of mitochondrial DNA polymerase gamma (POLG) exonuclease-deficient mice. Therefore, E1A immortalized murine embryonic fibroblasts (MEFs) from POLG exonuclease-deficient and wild-type (WT) mice were constructed. Surprisingly, when some E1A immortalized MEF lines were cultured in pyruvate-containing media they slowly became addicted to the pyruvate. The POLG exonuclease-deficient MEFs were more sensitive to several mitochondrial inhibitors and showed increased reactive oxygen species (ROS) production under standard conditions. When cultured in pyruvate-containing media, POLG exonuclease-deficient MEFs showed decreased oxygen consumption compared to controls. Increased AMP-activated protein kinase (AMPK) signaling and decreased mammalian target of rapamycin (mTOR) signaling delayed aging and influenced mitochondrial function. Therefore, the effects of 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR), an AMPK activator, or rapamycin, an mTOR inhibitor, on measures of mitochondrial function were determined. Rapamycin treatment transiently increased respiration only in WT MEFs and, under most conditions, increased ATP levels. Short term AICAR treatment transiently increased ROS production and, under most conditions, decreased ATP levels. Chronic AICAR treatment decreased respiration and ROS production in WT MEFs. These results demonstrate the context-dependent effects of AICAR and rapamycin on mitochondrial function.