Meeting Abstract
Potential mechanisms for improving desiccation tolerance can be gleaned from animals that survive severe water stress in nature. Mechanisms often include accumulation of trehalose, expression of late embryogenesis abundant (LEA) proteins, and suppression of oxidative phosphorylation. We used hypoxia preconditioning to achieve metabolic adjustments in human hepatoma HepG2 cells. Cells were incubated under hypoxia (0.5% O2) for 24 h, which promoted elevation of HIF-1α and a 62% decrease in respiration (measured under normoxia directly after hypoxia) compared to controls. HepG2 cells were transfected to express AfrLEA2, as well as a trehalose transporter to enable sugar uptake. The S-nitrosating agent MitoSNO was administered to some cells prior to removal from hypoxia to attenuate the surge of superoxide generated by mitochondria upon reoxygenation. Addition of 10 µM MitoSNO eliminated the ROS burst, as measured by oxidation of dihydroethidium. Cells were rapidly dehydrated to a residual water content of 0.225 g H2O/g dry mass via spin-drying and then immediately rehydrated. Growth profiles across 7 days post-rehydration showed that cells receiving hypoxia preconditioning exhibited significantly greater cell proliferation than normoxic controls. Proliferation was further bolstered by the addition of MitoSNO under hypoxia prior to drying. When day 7 cell counts for normoxic controls were set to 100%, the value for hypoxia preconditioned cells was 132.6 ± 15.0% and that for hypoxia plus MitoSNO was 183.1 ± 18.3% (means ± SD, n = 6). These findings indicate that metabolic preconditioning improves biostability of mammalian cells after acute desiccation. [Supported by NSF grant IOS-1457061/IOS-1456809 and NIH grant 2-RO1-DK046270-14A1]
