A central aspect of aging research concerns the question as to when individuality in lifespan arises 1. We have now discovered that a transient increase in reactive oxygen species (ROS), which occurs naturally during early development in a subpopulation of synchronized Caenorhabditis elegans, sets processes into motion that increase stress resistance, improve redox homeostasis and ultimately prolong lifespan in those animals. We find that these effects are linked to the global ROS-mediated decrease in developmental histone H3K4me3 levels. Studies in HeLa cells confirmed that global H3K4me3 levels are ROS-sensitive, and that depletion of H3K4me3 levels increases stress resistance in mammalian cell cultures. In vitro studies identified the Set1/MLL histone methyltransferase as the redox sensitive unit of the H3K4-trimethylating COMPASS complex. Our findings imply a novel link between early-life events, ROS-sensitive epigenetic marks, stress resistance and lifespan.
c Nitric oxide, produced in pancreatic  cells in response to proinflammatory cytokines, plays a dual role in the regulation of -cell fate. While nitric oxide induces cellular damage and impairs -cell function, it also promotes -cell survival through activation of protective pathways that promote -cell recovery. In this study, we identify a novel mechanism in which nitric oxide prevents -cell apoptosis by attenuating the DNA damage response (DDR). Nitric oxide suppresses activation of the DDR (as measured by ␥H2AX formation and the phosphorylation of KAP1 and p53) in response to multiple genotoxic agents, including camptothecin, H 2 O 2 , and nitric oxide itself, despite the presence of DNA damage. While camptothecin and H 2 O 2 both induce DDR activation, nitric oxide suppresses only camptothecin-induced apoptosis and not H 2 O 2 -induced necrosis. The ability of nitric oxide to suppress the DDR appears to be selective for pancreatic  cells, as nitric oxide fails to inhibit DDR signaling in macrophages, hepatocytes, and fibroblasts, three additional cell types examined. While originally described as the damaging agent responsible for cytokine-induced -cell death, these studies identify a novel role for nitric oxide as a protective molecule that promotes -cell survival by suppressing DDR signaling and attenuating DNA damage-induced apoptosis.
Edited by Jeffrey E. Pessin Oxidative stress is thought to promote pancreatic -cell dysfunction and contribute to both type 1 and type 2 diabetes. Reactive oxygen species (ROS), such as superoxide and hydrogen peroxide, are mediators of oxidative stress that arise largely from electron leakage during oxidative phosphorylation. Reports that -cells express low levels of antioxidant enzymes, including catalase and GSH peroxidases, have supported a model in which -cells are ill-equipped to detoxify ROS. This hypothesis seems at odds with the essential role of -cells in the control of metabolic homeostasis and organismal survival through exquisite coupling of oxidative phosphorylation, a prominent ROS-producing pathway, to insulin secretion. Using glucose oxidase to deliver H 2 O 2 continuously over time and Amplex Red to measure extracellular H 2 O 2 concentration, we found here that -cells can remove micromolar levels of this oxidant. This detoxification pathway utilizes the peroxiredoxin/ thioredoxin antioxidant system, as selective chemical inhibition or siRNA-mediated depletion of thioredoxin reductase sensitized -cells to continuously generated H 2 O 2. In contrast, when delivered as a bolus, H 2 O 2 induced the DNA damage response, depleted cellular energy stores, and decreased -cell viability independently of thioredoxin reductase inhibition. These findings show that -cells have the capacity to detoxify micromolar levels of H 2 O 2 through a thioredoxin reductase-dependent mechanism and are not as sensitive to oxidative damage as previously thought. Although oxidative stress in pancreatic -cells has been widely implicated in the pathogenesis of both type 1 and type 2 diabetes, reactive oxygen species (ROS), 6 under physiological conditions, are important signaling molecules necessary for
Background:The mechanisms that control  cell fate following cytokine-and nitric oxide-induced damage remain unknown. Results: Cytokine-induced nitric oxide activates ATM and ATM-dependent caspase activation in  cells. Conclusion: ATM regulates the induction of apoptosis in cytokine-treated  cells. Significance: These studies define a role for DNA damage and ATM activation in nitric oxide-induced  cell apoptosis.
While insulinoma cells have been developed and proven to be extremely useful in studies focused on mechanisms controlling β-cell function and viability, translating findings to human β-cells has proven difficult because of the limited access to human islets and the absence of suitable insulinoma cell lines of human origin. Recently, a human β-cell line, EndoC-βH1, has been derived from human fetal pancreatic buds. The purpose of this study was to determine whether human EndoC-βH1 cells respond to cytokines in a fashion comparable to human islets. Unlike most rodent-derived insulinoma cell lines that respond to cytokines in a manner consistent with rodent islets, EndoC-βH1 cells fail to respond to a combination of cytokines (IL-1, IFN-γ, and TNF) in a manner consistent with human islets. Nitric oxide, produced following inducible nitric oxide synthase (iNOS) expression, is a major mediator of cytokine-induced human islet cell damage. We show that EndoC-βH1 cells fail to express iNOS or produce nitric oxide in response to this combination of cytokines. Inhibitors of iNOS prevent cytokine-induced loss of human islet cell viability; however, they do not prevent cytokine-induced EndoC-βH1 cell death. Stressed human islets or human islets expressing heat shock protein 70 (HSP70) are resistant to cytokines, and, much like stressed human islets, EndoC-βH1 cells express HSP70 under basal conditions. Elevated basal expression of HSP70 in EndoC-βH1 cells is consistent with the lack of iNOS expression in response to cytokine treatment. While expressing HSP70, EndoC-βH1 cells fail to respond to endoplasmic reticulum stress activators, such as thapsigargin. These findings indicate that EndoC-βH1 cells do not faithfully recapitulate the response of human islets to cytokines. Therefore, caution should be exercised when making conclusions regarding the actions of cytokines on human islets when using this human-derived insulinoma cell line.
In this report, we show that nitric oxide suppresses DNA damage response (DDR) signaling in the pancreatic β-cell line INS 832/13 and rat islets by inhibiting intermediary metabolism. Nitric oxide is known to inhibit complex IV of the electron transport chain and aconitase of the Krebs cycle. Non-β cells compensate by increasing glycolytic metabolism to maintain ATP levels; however, β cells lack this metabolic flexibility, resulting in a nitric oxide-dependent decrease in ATP and NAD+. Like nitric oxide, mitochondrial toxins inhibit DDR signaling in β cells by a mechanism that is associated with a decrease in ATP. Non-β cells compensate for the effects of mitochondrial toxins with an adaptive shift to glycolytic ATP generation that allows for DDR signaling. Forcing non-β cells to derive ATP via mitochondrial respiration (replacing glucose with galactose in the medium) and glucose deprivation sensitizes these cells to nitric oxide-mediated inhibition of DDR signaling. These findings indicate that metabolic flexibility is necessary to maintain DDR signaling under conditions in which mitochondrial oxidative metabolism is inhibited and support the inhibition of oxidative metabolism (decreased ATP) as one protective mechanism by which nitric oxide attenuates DDR-dependent β-cell apoptosis.
The tumorigenic potential of human pluripotent stem cells (hPSCs) is a major limitation to the widespread use of hPSC derivatives in the clinic. Here, we demonstrate that the small molecule STF-31 is effective at eliminating undifferentiated hPSCs across a broad range of cell culture conditions with important advantages over previously described methods that target metabolic processes. Although STF-31 was originally described as an inhibitor of glucose transporter 1, these data support the reclassification of STF-31 as a specific NAD + salvage pathway inhibitor through the inhibition of nicotinamide phosphoribosyltransferase (NAMPT). These findings demonstrate the importance of an NAD + salvage pathway in hPSC biology and describe how inhibition of NAMPT can effectively eliminate hPSCs from culture. These results will advance and accelerate the development of safe, clinically relevant hPSCderived cell-based therapies. STEM CELLS TRANSLATIONAL MEDICINE 2015;4:483-493 SIGNIFICANCEThe tumorigenic potential of human pluripotent stem cells (hPSCs) is a major limitation to the widespread use of hPSC derivatives in the clinic. This study provides detailed analyses of cellular metabolic flux to define an efficient strategy for selective hPSC elimination that is effective across many culture conditions and does not have cytotoxic effects on hPSC-derived progeny. Of broad significance to the stem cell and regenerative medicine fields, this study also highlights the importance of examining the effect of in vitro culturing parameters when evaluating the efficacy of hPSC-elimination strategies, especially those that target metabolic processes.
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