Hydrogen peroxide modulates energy metabolism and oxidative stress in cultures of permanent human Müller cells MIO‐M1

In this study the influence of hydrogen peroxide (H2 O2 ) on the redox state, NADH protein binding, and mitochondrial membrane potential in Müller cells is investigated. Cultures of permanent human Müller cells MIO-M1 were exposed to H2 O2 in 75 µM and 150 µM concentration for two hours. Fluorescence emission spectra and lifetimes were measured by two-photon microscopy (excitation wavelength: 740 nm) at the mitochondria which were identified in the microscopic images by their fluorescence properties (spectra and intensity). Two hours of H2 O2 exposure did not impair viability of MIO-M1 cells in culture. Whereas the ratio of flavine- to NADH fluorescence intensity did not change under either H2 O2 concentration, the mean lifetime was significantly different between controls, not exposed to H2 O2 , and the 150 µM H2 O2 exposure (972 ± 63 ps vs. 1152 ± 64 ps, p = 0.014). One hour after cessation of the H2 O2 exposure, the value retuned to that of the control (983 ± 36 ps). A hyperpolarization of the mitochondrial membrane under 150 µM H2 O2 was found. These findings suggest a shift form free to protein-bound NADH in mitochondria as well as a hyperpolarization of their inner membrane which could be related to an impairment of Müller cell function despite their preserved viability. Exposure of human Müller cells to hydrogen peroxide for two hours results in a reversible change of protein binding of mitochondrial NADH upon unchanged redox ratio. The mitochondrial membrane potential is increased during exposure.