Abstract 12822: Regulation of Endothelial Metabolism by Krüppel-like Factor 2 and Shear Stress
Cellular metabolism is a crucial regulator of cell phenotype. Flow hemodynamics control endothelial cell (EC) phenotype and may play a role in metabolic reprogramming of ECs. The flow responsive transcription factor KLF2 is a pivotal mediator of the quiescent state of ECs, but the mechanism is unclear. We assessed the hypothesis that KLF2 regulates endothelial quiescence by controlling endothelial metabolism.
Laminar flow exposure and KLF2 over expression in HUVECs reduced glucose uptake in-vitro (-1.46 fold ± 0.09 and -1.23 fold ± 0.05, both p≤0.05). Consistently, infusion of 2-NBD labeled glucose in Langendorff perfused hearts of inducible EC specific KLF2 knockout and wild type mice and subsequent staining with CD31 and flow cytometry analyses showed an increased ex vivo uptake of glucose (1.37 fold ± 0.17, p≤0.05) in ECs lacking KLF2. KLF2 also reduced mitochondrial activity (-1.41 fold ± 0.11, p≤0.05) without altering NAD+ levels. Furthermore, laminar flow and KLF2 reduced mitochondrial content (-1.48 fold ± 0.19, p≤0.05 and -1.65 fold ± 0.07, p≤0.05), confirming that flow and KLF2 reduces metabolic activity of ECs. Bioenergetic measurements (Seahorse XF) revealed that KLF2 reduces mitochondrial respiration (-1.5 fold ± 0.01, p≤0.001) and glycolytic acidification (3.07 fold ± 0.08, p≤0.01).
Consistent with the reduction in glycolysis, KLF2 reduces the expression of key enzymes in the glycolytic pathway such as hexokinase 2, which catalyzes the first essential step of glucose metabolism (-1.71 fold ± 0.04, p≤0.05), the rate-limiting enzymes PFKFB3 (-1.61 fold ± 0.06, p≤0.05) and PFK-1 (-1.53 fold, ± 0.06, p≤0.01) and the key glycolytic genes PGM5 and PDK4. Deep sequencing analysis revealed a similar change in gene expression patterns in laminar flow-stimulated ECs versus static controls. Phospho-kinase array analysis showed that KLF2 decreased levels of HSP60 (-1.81 fold ± 0.06, p≤0.001), an essential mitochondrial protein and the key metabolic energy sensor AMPKa1 (-2.35 fold ± 0.02, p≤0.05), which may explain the inhibition of mitochondrial respiration by KLF2.
In conclusion, our data demonstrate that KLF2 attenuates both glycolysis and mitochondrial respiration in ECs and thus induces a quiescent “phalanx” phenotype of the endothelium.
- © 2013 by American Heart Association, Inc.