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목적 : Transcription factor 4 (TCF4) has been implicated in the pathogenesis of Fuchs endothelial corneal dystrophy (FECD). In this study, we investigated the role of TCF4 in the regeneration of human corneal endothelial cells (HCECs).
방법 : HCECs were cultured, and TCF4 was blocked by siRNA(si-TCF4) or activated using CRISPR/Cas9 systems for TCF4(pl-TCF4). Cell viability was assessed using a WST-8 assay kit, and mitochondrial membrane potential was measured using the JC-1 dye. Intracellular reactive oxygen species(ROS) formation was measured using a DCF-DA probe. 5'-bromo-2'-deoxy-uridine incorporation assay was used to determine the cell proliferation rate. The cell cycle was analyzed by quantitation of DNA content. Protein expression was evaluated by western blotting. Induction of autophagy and apoptosis were also evaluated.
결과 : TCF4 regulated the expression of GFAP, OCT3/4, and β-catenin/GSK3β in cultured HCECs. ΔΨm was reduced by blocking TCF4 and was elevated by its overexpression. TCF4 regulates mitochondrial functions including intracellular ROS levels, ATP concentration, and AMPK activation. Cell viability and cell proliferation, as well as CDK1 and cyclin D1 expression were all regulated by TCF4. Similarly, LC3-II expression was also regulated by TCF4, indicating an effect on autophagy. Furthermore, TCF4 interacted with transcriptional factors including KLF4 and NFAT.
결론 : In conclusion, TCF4 regulates mitochondrial function, energy production, cell proliferation, and autophagy in cultured HCECs. TCF4 may be a molecular target for regeneration of HCECs in HCEC diseases.
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