Uridine Prevents Fenofibrate-Induced Fatty Liver

Citation: Le TT, Urasaki Y, Pizzorno G ( Uridine Prevents Fenofibrate-Induced Fatty Liver Thuc T. Le 0 1 Yasuyo Urasaki 0 1 Giuseppe Pizzorno 0 1 Vasu D. Appanna, Laurentian University, Canada 0 Funding: This work was partially supported by the Nevada INBRE Program of the National Center for Research Resources (P20RR-016464, TTL), the American Cancer Society (IRG-08-062-04, TTL), and the Vons Breast Cancer Research Award (TTL and GP). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript 1 1 Nevada Cancer Institute, Las Vegas, Nevada, United States of America, 2 Desert Research Institute , Las Vegas, Nevada , United States of America Uridine, a pyrimidine nucleoside, can modulate liver lipid metabolism although its specific acting targets have not been identified. Using mice with fenofibrate-induced fatty liver as a model system, the effects of uridine on liver lipid metabolism are examined. At a daily dosage of 400 mg/kg, fenofibrate treatment causes reduction of liver NAD+/NADH ratio, induces hyper-acetylation of peroxisomal bifunctional enzyme (ECHD) and acyl-CoA oxidase 1 (ACOX1), and induces excessive accumulation of long chain fatty acids (LCFA) and very long chain fatty acids (VLCFA). Uridine co-administration at a daily dosage of 400 mg/kg raises NAD+/NADH ratio, inhibits fenofibrate-induced hyper-acetylation of ECHD, ACOX1, and reduces accumulation of LCFA and VLCFA. Our data indicates a therapeutic potential for uridine co-administration to prevent fenofibrate-induced fatty liver. - Uridine has been widely tested in clinical trials for treatments of neurological disorders, liver dysfunction, and cancer [1]. Uridine is well-known to have positive neurological and systemic effects [1 4]. However, lack of understanding of its biological activity hinders effective usage of uridine to modulate human physiology in both healthy and diseased states. Previously, a linkage between pyrimidine biosynthesis pathway and liver lipid metabolism was reported through the use of transgenic uridine phosphorylase 1 (UPase1-TG) mice with overexpression of UPase1 and depleted endogenous uridine source [5]. UPase1 is an enzyme that catalyzes the reversible conversion of uridine into uracil and regulates uridine homeostasis [6]. UPase1-TG mice exhibited fatty liver phenotype, which could be reversed with dietary uridine supplementation. Uridine was found to modulate liver lipid metabolism although its specific acting targets have not been identified [5]. The liver is an important source of uridine, where circulating plasma uridine is degraded in a single pass and replaced with newly synthesized uridine [7]. Most tissues lack the ability to synthesize uridine and rely on plasma for uridine supply [8]. Thus, the liver serves as an effective regulator of whole-body uridine homeostasis. The concentration of circulating uridine is highly conserved across species of between 2 mM to 4 mM [8,9]. To maintain circulating uridine homeostasis, the liver has multiple robust means to manage surges in plasma uridine concentration due to dietary intakes. Uridine could be salvaged into pyrimidine nucleotide pool of UTP, CTP, and TTP, or catabolized into balanine and acetyl-CoA [1]. Acute surges of uridine or its metabolites in the liver have the ability affect other energy metabolism processes as evidence by the ability of dietary uridine supplementation to modulate liver lipid metabolism [5]. The liver is also a primary site for drug detoxification, which renders it highly susceptible to drug-induced fatty liver [10]. Druginduced fatty liver is a well-known side effect of many currently FDA-approved drugs [1113]. Most drugs cause fatty liver by inhibiting hepatic fatty acid oxidation [14,15]. Fatty liver due to chronic drug usage increases the risk for the development of nonalcoholic fatty liver disease such as steatohepatitis and cirrhosis [16,17]. Current clinical approach to the prevention of fatty liver is dependent on the management of obesity or obesity-associated metabolic diseases, often via pharmaceutical means [18]. However, this approach is problematic when the drugs themselves are contributors to the development of fatty liver condition. In this study, the ability of uridine to modulate liver lipid metabolism is evaluated in a C57bl/6 mouse model with druginduced fatty liver. Previously, our lab reported that fenofibrate, when administered at high dosage, induced severe hepatic microvesicular steatosis in mice [19]. Fenofibrate is a peroxisome proliferator-activated receptor-a (PPAR-a) agonist known for its blood lipid-lowering effects [20,21]. Fenofibrate is widely prescribed for the treatment of dyslipidemia, type 2 diabetes, and the metabolic syndrome [22]. Fenofibrate, via PPAR-a, stimulates the remodeling of hepatic lipid metabolism and promotes fatty acid oxidation [23,24]. However, inhibitory effects of fenofibrate on fatty acid oxidation have also been reported in rodents at high dosage [25,26]. Fenofibrate induces hepatocarcinoma and fatty liver in rodents but not in humans [19,27,28]. Uridine is coadministered with fenofibrate via dietary supplementation and the effects of uridine on liver lipid metabolism are evaluated in C57bl/ 6 mice. We aim to evaluate the therapeutic potential of uridine for the prevention of drug-induced fatty liver. First, the relationship between endogenous liver uridine concentration and fenofibrate-induced fatty liver was evaluated in C57bl/6 mice and mice with disrupted uridine homeostasis, UPase1-/- and UPase1-TG mice of C57bl/6 background. UPase1-/mice had genetic knock-out of UPase1 and elevated endogenous liver uridine concentration of 43 mM compared to 6 mM in C57bl/6 mice [6]. In contrast, UPase1-TG mice had genetic knock-in of UPase1 and reduced endogenous liver uridine concentration of 0.5 mM [5]. Using CARS microscopy as a sensitive means to visualize liver lipid [19,29,30], both C57bl/6 mice and UPase1-/- mice did not exhibit any visible fatty liver phenotype (Figure 1A). UPase1-TG mice with endogenous 0.5 mM of liver uridine concentration exhibited mild microvesicular steatosis [5,19]. Administration of fenofibrate at 400 mg/kg/ day induced a 5-fold and a 3-fold increase in liver lipid content in C57bl/6 and UPase1-/- mice, respectively (Figure 1B). On the other hand, administration of fenofibrate induced a 2-fold increase in liver lipid content in UPase1-TG mice compared to untreated UPase1-TG mice, or a 6-fold increase in liver lipid content compared to untreated C57bl/6 or UPase1-/- mice. Thus, depletion of endogenous liver uridine concentration aggravated, whereas elevation of endogenous liver uridine concentration alleviated, fenofibrate-induced fatty liver. Next, the impact of exogenous uridine supplementation on fatty liver phenotype was evaluated in C57bl/6, UPase1-/-, and UPase1TG mice. Consistent with our previous findings, dietary uridine suppleme (...truncated)