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Young Joo Shin, Tae-Young Chung, Soonil Kwon, Joon-Young Hyon; Transcription factor 4 Regulates Mitochondrial Function and Energy Metabolism in cultured Human Corneal Endothelial Cells. Invest. Ophthalmol. Vis. Sci. 2017;58(8):1440.
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© ARVO (1962-2015); The Authors (2016-present)
To investigate the role of TCF4 in the mitochondrial functions of human corneal endothelial cells (HCECs).
HCECs were cultured, and TCF4 was blocked by siRNA (si-TCF4) or activated using CRISPR/dCas9 systems for TCF4 (pl-TCF4). Cell viability was assessed using a WST-8 assay kit, and mitochondrial membrane potential (ΔΨm) was measured using the JC-1 dye. Intracellular reactive oxygen species (ROS) formation was measured using a DCF-DA probe. Protein expression was evaluated by western blotting. Induction of autophagy and apoptosis were also evaluated.
A mitochondrial viability stain demonstrated that mitochondrial viability was reduced by blocking TCF4 and elevated by its overexpression (p <0.05). ΔΨm was measured using JC-1, and was decreased by blocking TCF4 and elevated by TCF4 overexpression (p <0.05). Intracellular ROS production was evaluated by dichlorofluorescein (DCF) fluorescence. Intracellular ROS formation increased with TCF4 inhibition and decreased with its overexpression (p < 0.05). Relative adenosine triphosphate (ATP) production decreased upon TCF4 inhibition, but was elevated by its overexpression (p = 0.009 and 0.026). Activation of AMP-activated protein kinase (AMPK) decreased with inhibition of TCF4 expression, however TCF4 overexpression has the opposite effect (p < 0.05). In contrast, sirtuin 1 (SIRT1) expression was elevated by blocking TCF4 and reduced by its overexpression (p <0.05).
TCF4 regulates mitochondrial function and energy production in cultured HCECs.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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