Abstract 15392: Radiation-Induced Coronary Microvascular Leakage is Mediated by Loss of Endothelial Tight Junction Protein Claudin-1

Abstract

Background: Ionizing radiation (IR) exposure to the heart during cancer therapy induces coronary microvascular leakage. Increased microvascular porosity allows inflammatory cell extravasation into the perivascular and myocardial interstitial space leading to fibrotic end organ damage. Hypothesis: Radiation-induced coronary microvascular leakage is mediated by a loss of a constitutively expressed tight-junction protein, claudin-1 (cldn1). Methods: We studied the function of cldn1 in human coronary vascular endothelial cells (HMVECs) and in mice using oligonucleotide-based cldn1 gene deletion. For the gain of function, we developed vascular-specific (VE-cadherin promotor-linked) claudin-1 overexpressing transgenic (TG) mice. We studied the effect of IR in cldn1 loss and gain of function models. Capillary permeability was assessed by FITC-dextran permeability and Evans blue dye injection, in vitro and in vivo, respectively . The extent of end-organ damage was examined by analyzing myocardial and perivascular fibrosis. Results: In HMVECs, cldn1 gene silencing (control, 1.0±0.34; siRNA, 0.026±0.03; p<0.001) led to increased permeability (control, 1.0±0.01, siRNA, 1.3±0.01; p<001). In mice, genetically engineered anti-cldn1 siRNA administration knocked down myocardial cldn1 expression (control, 1.0±0.18; siRNA, 0.51±0.11; p=0.04). Functionally, cldn1 knockdown led to increased microvascular extravasation (control, 1.0±0.23, cldn1-siRNA 0.38±0.27 p=0.002). Cardiac morphometry in these mice showed myocardial fibrosis (control, 1.0±0.19; siRNA, 1.29±0.35; p=0.048). Cardiac irradiation significantly decreased myocardial cldn1 (control 1.0±0.27, IR 0.45±0.31; p=0.001) and dye leakage (control 1.0±0.18, IR 1.28±0.22; p=0.007). Conversely, endothelium-targeted cldn1 overexpression in mice prevented abnormal microvasculature permeability (WT 1.0±0.23, TG 0.38±0.27 p=0.002) and conferred protection from IR-induced fibroinflammatory changes. Conclusions: Our study shows that cldn1 is crucial for maintaining coronary microvascular permeability. Prevention of cldn1 degradation or cldn1 reconstitution using novel pharmacotherapeutic approaches can prevent cardiac end-organ damage induced by cardiac IR.