Fractionated radiation suppresses Kruppel-like factor 2 pathway to a greater extent than by single exposure to the same total dose

Abstract

Kruppel-like factor 2 (KLF2) is a positive transcriptional regulator of several endothelial protective molecules, including thrombomodulin (TM), a surface receptor, and endothelial nitric oxide synthase (eNOS), an enzyme that generates nitric oxide (NO). Loss of TM and eNOS causes endothelial dysfunction, which results in suppressed generation of activated protein C (APC) by TM-thrombin complex and in upregulation of intercellular adhesion molecule 1 (ICAM-1). Mechanistic studies revealed that activation of extracellular signal-regulated kinase 5 (ERK5) via upregulation of myocyte enhancer factor 2 (MEF2) induces KLF2 expression. Radiation causes endothelial dysfunction, but no study has investigated radiation's effects on the KLF2 pathway. Because fractionated radiation is routinely used during cancer radiotherapy, we decided to delineate the effects of radiation dose fractionation on the KLF2 signaling cascade at early time points (up to 24 h). We exposed human primary endothelial cells to radiation as a series of fractionated or as a single exposure, with the same total dose delivered to each group. We measured the expression and activity of critical members of the KLF2 pathway at subsequent time points, and determined whether pharmacological upregulation of KLF2 can reverse the radiation effects. Compared to single exposure, fractionated radiation profoundly suppressed KLF2, TM, and eNOS levels, subdued APC generation, declined KLF2 binding ability to TM and eNOS promoters, enhanced ICAM-1 expression, and decreased expression of upstream regulators of KLF2 (ERK5 and MEF2). Pharmacological inhibitors of the mevalonate pathway prevented fractionatedradiation-induced suppression of KLF2, TM, and eNOS expression. Finally, fractionated irradiation to thoracic region more profoundly suppressed KLF2 and enhanced ICAM-1 expression than single exposure in the lung at 24 h. These data clearly indicate that radiation dose fractionation plays a critical role in modulating levels of KLF2, its upstream regulators, and its downstream target molecules in endothelial cells. Our findings will provide important insights for selecting fractionated regimens during radiotherapy and for developing strategies to alleviate radiotherapy-induced toxicity to healthy tissues. Endothelium-the inner lining of blood and lymphatic vessels-is composed of endothelial cells and forms a critical component of all the body's organs. Under normal physiological conditions, endothelium acts as a barrier between blood and surrounding tissues, maintains vascular tone, regulates blood homeostasis, provides an anticoagulant surface, and restricts proliferation of vascular smooth muscle cells 1. Mechanistic studies revealed that all these endothelial-mediated beneficial functions are primarily exerted by thrombomodulin (TM), an endothelial surface receptor, and endothelial nitric oxide synthase (eNOS), an enzyme that generates nitric oxide (NO) 2-6. TM has intrinsic antioxidant and anti-inflammatory properties 2,4 , and TM-thrombin complex enhances genera...

Krüppel-like Factor 10 as a Prognostic and Predictive Biomarker of Radiotherapy in Pancreatic Adenocarcinoma

“…KLF2 and KLF4 are positive regulators of endothelial-protective molecules such as nitric oxide and thrombomodulin. Compared with single-dose radiation, fractionated radiation markedly reduced the ERK5/KLF2 pathway and enhanced ICAM-1 expression, leading to endothelial dysfunction [77]. KLF4 and KLF5 may prevent radiation-induced intestinal injury by inhibiting apoptosis and modulating DNA repair pathways [78,79].…”

Challenges with Methods for Detecting and Studying the Transcription Factor Nuclear Factor Kappa B (NF-κB) in the Central Nervous System

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