In Vivo Endothelial NO Activation by Losartan Is Independent of Blood Pressure Lowering and EXP3179/EXP3174 Metabolites Yet Require First Passage Metabolism

Abstract

Background and Purpose: Losartan is a well-established anti-hypertensive angiotensin II (AngII) receptor type 1 (ATR1) blocker (ARB) with unexpected therapeutic properties in a range of diseases not linked to hypertension, such as chronic lung diseases, Alzheimer's disease, muscular dystrophy and viral infections. This suggests that there is more to losartan than simple AngII-ATR1 inhibition. We have recently shown that losartan can unexpectedly activate endothelial function in vivo via the endothelial release of protective nitric oxide (NO), which is typically associated with aerobic exercise and vascular homeostasis. Hence, losartan activation of endothelial NO release may be behind its protective non-blood pressure-related effects, although how this occurs is unknown. Losartan is a prodrug with many active metabolites that might be behind some of its unexpected therapeutic properties. Losartan first-passage metabolism in the liver converts it to EXP3179 (losartan-carboxaldehyde) believed to be devoid of ATR1 blocking effects followed by further metabolism into EXP3174 (losartan-carboxylic acid), the main ATR1 blocker. Our goal is to determine whether first passage metabolism is required for losartan to activate endothelial NO release, or if bypassing liver metabolism via intraperitoneal (IP) injections of losartan metabolites EXP3179/3174 will result is similar biological activites, an indication that metabolites of losartan are not involved in this biological activity. Experimental Approach: C57B6/J mice were treated with 1) losartan potassium-salt (LOS-PS) for 2 weeks via drinking water (liver first passage-dependent; 0.6 g/L) or 2) daily IP injections (first passage-independent) of LOS-PS, EXP3179 or EXP3174 (10 mg/kg). Systemic BP was noninvasively measured by a tail cuff system in mice anesthetized with isoflurane. NO-dependent vasodilatory properties of losartan, EXP3179 and EXP3174 were evaluated in mouse aortic rings using dual-wire myograph systems. Key Results: When losartan was delivered in drinking water, NO-dependent vasorelaxation was increased by 57.6%. In stark contrast, IP injection of losartan resulted in unexpected increases in contractility by 124.9% compared to vehicle control, whereas IP EXP3179 and EXP3174 also increased vasoconstriction by 69.8% and 72.2% respectively. Vasodilatory effects of losartan were undetected in the presence of L-NAME (NO synthase inhibitor), confirming that losartan exerts its effects in a NO-dependent manner. Moreover, BP was decreased in all drug treated group of mice, confirming that all drugs were biologically active and blocking ATR1. Conclusion and implications: Our data unexpectedly show that losartan increases endothelial function in mice only when given in drinking water, which suggests that non-EXP3179/74 metabolites may play an important role in endothelial NO activation. This also suggests that losartan-induced BP lowering is separate from its endothelial NO activating properties.