Aspirin may influence cellular energy status

Abstract

In our previous findings, we have demonstrated that aspirin/acetyl salicylic acid (ASA) might induce sirtuins via aryl hydrocarbon receptor (Ah receptor). Induction effects included an increase in cellular paraoxonase 1 (PON1) activity and apolipoprotein A1 (ApoA1) gene expression. As predicted, ASA and salicylic acid (SA) treatment resulted in generation of H₂O₂, which is known to be an inducer of mitochondrial gene Sirt4 and other downstream target genes of Sirt1.

Our current mass spectroscopic studies further confirm the metabolism of the drugs ASA and SA. Our studies show that HepG2 cells readily converted ASA to SA, which was then metabolized to 2,3-DHBA. HepG2 cells transfected with aryl hydrocarbon receptor siRNA upon treatment with SA showed the absence of a DHBA peak as measured by LC–MS/MS. MS studies for Sirt1 action also showed a peak at 180.9 m/z for the deacetylated and chlorinated product formed from N-acetyl lε-lysine. Thus an increase in Sirt4, Nrf2, Tfam, UCP1, eNOS, HO1 and STAT3 genes could profoundly affect mitochondrial function, cholesterol homeostasis, and fatty acid oxidation, suggesting that ASA could be beneficial beyond simply its ability to inhibit cyclooxygenase.

Introduction

Acetylsalicylic acid (ASA) has an analgesic property and serves as an important modulator of cyclooxygenase (COX), and thus the production of thromboxanes (Vane, 1971, Vane and Botting, 2003, Rowlinson et al., 2003). The beneficial actions of ASA have been mainly attributed to this property; ASA is believed to be readily hydrolyzed by enzymes, including paraoxonase 1 (PON1) (Santanam and Parthasarathy, 2006) to salicylic acid (SA). ASA has a short plasma half-life of less than 15 min and the product, SA, has profound biological actions of its own. The half-life of SA exceeds several hours and is concentration-dependent. In addition to its well-known analgesic actions (Williams and Hennekens, 2004, Hennekens et al., 1989, Claria and Serhan, 1995), SA also has been extensively studied as a hydroxyl radical trap (John et al., 1993). It is excreted as a glucuronide and other conjugates (Gutierrez et al., 2006), suggesting that it undergoes metabolic conversion in the liver. However, SA is known to be metabolized by liver microsomes enzymatically as well as non-enzymatically (Grootveld and Halliwell, 1986) to 2,3- and 2,5- dihydroxy benzoic acid (DHBA) by hydroxyl radicals. These dihydroxybenzene derivatives, such as 2,3- or 2,5-DHBA, are expected to autooxidize to form quinones and reactive oxygen species, particularly hydrogen peroxide (H₂O₂) (Kamble et al., 2013). Reactive oxygen species and H₂O₂ have been known to be attributed to an increase in the induction of sirtuins (Sirt) such as Sirt1, Sirt3 and Sirt4 (Alcendor et al., 2004, Kwon and Ott, 2008). Recent studies showed that there are few protein substrates for Sirt 2–7 which play an important role in multiple disease-relevant pathways largely encompassing mitochondrial energetics and inflammatory cardiomyopathy. Increased Sirt1 activity would decrease glucose levels, increase the number of mitochondria, regulate insulin sensitivity and lower body weight (Elliott and Jirousek, 2008). Among other sirtuins, Sirt4 is a mitochondrial protein that does not display nicotinamide adenine dinucleotide (NAD)-dependent deacetylase activity, but it utilizes NAD for carrying out adenosine diphosphate (ADP)–ribosylation of glutamate dehydrogenase (GDH) in the mitochondria. Nevertheless, ADP–ribosylation of GDH represses GDH activity by Sirt4, which limits metabolism of glutamate and glutamine to generate adenosine triphosphate (ATP) (Haigis et al., 2006). Thus, we tested the ability of ASA and SA to generate H₂O₂ to induce Sirt4, Nrf2, Tfam, UCP1, eNOS and STAT3 genes responsible for mitochondrial function and liver×receptorα, farnesoid×receptor and PPARα genes involved in lipogenesis and cholesterol homeostasis.