ABSTRACT
Replicative and chronological lifespan are two different modes of cellular aging. Chronological lifespan is defined as the duration during which quiescent normal cells retain their capacity to re-enter the proliferative cycle. This study investigated whether changes in metabolism occur during aging of quiescent normal human fibroblasts (NHFs) and the mechanisms that regulate these changes. Bioenergetics measurements were taken in quiescent NHFs from younger (newborn, 3-day, 5-month, and 1-year) and older (58-, 61-, 63-, 68-, and 70-year) healthy donors as well as NHFs from the same individual at different ages (29, 36, and 46 years). Results show significant changes in cellular metabolism during aging of quiescent NHFs: Old NHFs exhibit a significant decrease in glycolytic flux and lactate levels, and increase in oxygen consumption rate (OCR) and ATP levels compared to young NHFs. Results from the Seahorse XF Cell Mito Stress Test show that old NHFs with a lower Bioenergetic Health Index (BHI) are more prone to oxidative stress compared to young NHFs with a higher BHI. The increase in OCR in old NHFs is associated with a shift in mitochondrial dynamics more toward fusion. Genetic knockdown of mitofusin 1 (MFN1) and optic atrophy 1 (OPA1) in old NHFs decreased OCR and shifted metabolism more toward glycolysis. Downregulation of MFN1 and OPA1 also suppressed the radiation-induced increase in doubling time of NHFs. In summary, results show that a metabolic shift from glycolysis in young to mitochondrial respiration in old NHFs occurs during chronological lifespan, and MFN1 and OPA1 regulate this process.
Subject(s)
Aging/genetics , GTP Phosphohydrolases/genetics , Glycolysis/genetics , Mitochondria/genetics , Mitochondrial Membrane Transport Proteins/genetics , Oxidative Phosphorylation , Adult , Aged , Aging/metabolism , Cell Division , Cell Respiration/genetics , Cells, Cultured , Fibroblasts/cytology , Fibroblasts/metabolism , GTP Phosphohydrolases/antagonists & inhibitors , GTP Phosphohydrolases/metabolism , Gene Expression Regulation , Humans , Infant , Infant, Newborn , Middle Aged , Mitochondria/metabolism , Mitochondrial Dynamics , Mitochondrial Membrane Transport Proteins/antagonists & inhibitors , Mitochondrial Membrane Transport Proteins/metabolism , Oxygen Consumption , RNA, Small Interfering/genetics , RNA, Small Interfering/metabolism , Signal TransductionABSTRACT
The immutans (im) variegation mutant of Arabidopsis has green and white-sectored leaves due to the absence of fully functional plastid terminal oxidase (PTOX), a plastoquinol oxidase in thylakoid membranes. PTOX appears to be at the nexus of a growing number of biochemical pathways in the plastid, including carotenoid biosynthesis, PSI cyclic electron flow, and chlororespiration. During the early steps of chloroplast biogenesis, PTOX serves as an alternate electron sink and is a prime determinant of the redox poise of the developing photosynthetic apparatus. Whereas a lack of PTOX causes the formation of photooxidized plastids in the white sectors of im, compensating mechanisms allow the green sectors to escape the effects of the mutation. This manuscript provides an update on PTOX, the mechanism of im variegation, and findings about im compensatory mechanisms.
