Abstract 5867: The Beneficial Effect of Raloxifene in Aortic Valve Calcification is Associated with Up-Regulation of Gas6-Axl Pathway

Abstract

Aortic valve calcification (AVC) is an inflammatory regulated process. Growth arrest-specific 6 (Gas6) is involved in atherosclerosis and inflammation; in addition the Gla domain of Gas6 directly inhibits mineralization. We sought to assess the role of Gas6 in AVC using our unique animal model of reversible AVC based on a uremia-inducing diet. We also evaluated the possible role of raloxifene (a selective estrogen modulator), which has demonstrated anti inflammatory effects on blood vessels, in the pathogenesis of AVC. Aortic valves were obtained from four groups of rats (n=10 each): calcified valves- from rats fed with the uremia-inducing diet for 7 weeks (group A), valves after calcification resolution following diet cessation (group B), valves from rats fed with the same diet who also received subcutaneous raloxifen (1 mg/kg/day) (group C), and control valves. Valves were examined using multislice computed tomography (MSCT), histology, RT-PCR and western blot. Histological evaluation of calcified valves (group A) revealed positive staining for calcium deposits and osteoblast’s markers (osteopontin, Runx-2). Gene expression analyses of these valves revealed 2.5 times decrease in Gas6 level compared to control valves (p<0.01). Western blot confirmed these results, and further showed a significant decrease of 33 % in Axl (Gas6 receptor) level along with down-regulation of the AKT survival pathway. Resolution of AVC (group B) was demonstrated using MSCT and histology; it was associated with down-regulation of osteoblast features, and up-regulation of both Gas6-Axl and AKT pathways. MSCT of raloxifene treated rats (group C) showed only minimal valve calcification compared with group A (Agatston score 23 ± 7 vs. 105± 15; p< 0.05). The beneficial effect of raloxifene was also confirmed by histology and was associated with up-regulation of Gas6 and Axl (30% and 40% increase respectively compared with group A; p<0.05) and of the AKT pathway. Down-regulation of the Gas6-Axl pathway is a critical step in inducing AVC. The reversibility of AVC was associated with up-regulation of the Gas6-Axl and AKT pathways. These pathways were significantly restored by raloxifene, which may be a promising treatment for AVC.