Specific targeting of Egr-1 reduces neointimal lesions and improves RV remodeling in PAH

Abstract

Purpose: Pulmonary arterial hypertension (PAH) is a progressive pulmonary vascular disease with a high mortality and to date no curable treatment. Increased pulmonary blood flow, as in congenital heart disease, is an essential trigger for the development of neointimal lesions that are characteristic for PAH. We recently identified a flow-specific signal, Egr-1, which was activated in both experimental and human PAH. The aim of the present study was to identify the role of Egr-1 in the development of neointimal lesions in experimental PAH. Methods: In wistar rats (n=76), flow-associated PAH was created by combining monocrotaline with an aortocaval (av) shunt. Egr-1 was inhibited via chronic intravenous delivery (every 48h) of catalytic oligodeoxynucleotides (called DNAzymes) specifically targeting Egr-1 in vivo via RNA degradation. Starting at av-shunt, rats were randomized for either DNAzyme therapy (PAH\_DZ), scrambled DNAzymes (PAH\_SCR; scrambled oligonucleotides that do not recognize Egr-1 RNA) or vehicle (PAH_VEH). Sham operated rats served as control (Con). Increased flow was confirmed by echocardiography. At both 1 week and 3 weeks after increased flow animals underwent hemodynamic evaluation and were sacrificed for further histopathological analysis. Results: At 3 weeks PAH rats had increased vascular occlusion (33%±3, vs. 2±0%, p<0.001) and wall thickness (5.6μm±0.7 vs. 0.3±0.0 p<0.001), presence of neointimal lesions (36% vs. 0% of all arterioles), and subsequently increased right ventricular (RV) peak pressure (60mmHg±2 vs. 24±1, p<0.001) and hypertrophy (RV/BW ratio 1.5mg/g±0.1 vs. 0.6±0.1, p<0.001). Treatment with DNAzymes targeting Egr-1 attenuated vascular remodeling as early as 1 week after flow. At 3 weeks DNAzyme treatment significantly reduced vascular occlusion (-47%, p<0.001), wall thickness (-48%, p<0.001) and the development of neointimal lesions (-69%, p<0.001) compared to PAH\_VEH. PAH\_SCR showed no change compared to PAH\_VEH. Moreover, DNAzyme treatment significantly attenuated pulmonary vascular resistance (-27%, p<0.05), RV peak pressure (-12%, p<0.05) and RV hypertrophy (RV/body weight -27%, p<0.05) compared to PAH\_VEH and PAH_SCR. Conclusion: Here we show that RNA specific targeting of Egr-1 attenuates pulmonary vascular remodeling and disease development in a neointimal model of PAH. These data suggest that Egr-1 is an important regulator of PAH development and may possibly form a therapeutic target for future PAH treatment.