Aldo-keto reductase-7A protects liver cells and tissues from acetaminophen-induced oxidative stress and hepatotoxicity

Abstract

Aldo-keto reductase-7A (AKR7A) is an enzyme important for bioactivation and biodetoxification. Previous studies suggested that Akr7a might be transcriptionally regulated by oxidative stress-responsive transcription factor nuclear factor erythroid 2 p45-related factor 2 (Nrf2), a protein highly responsive to acetaminophen (APAP) or its intermediate metabolite, N-acetyl-p-benzoquinoneimine (NAPQI). This study was, therefore, carried out to investigate whether Akr7a is involved in the protection against APAP-induced oxidative stress and hepatotoxicity. We found that in response to APAP or NAPQI exposure, Akr7a3 mRNA and protein were significantly up-regulated in vitro in human HepG2 and LO2 cells. Similarly, strong induction was observed for Akr7a5 in mouse AML12 hepatocytes exposed to APAP. In vivo in wild-type rats, significant up-regulation of hepatic AKR7A1 protein was observed after administration of APAP. On the other hand, depletion of Nrf2 reduced the expression of Akr7a3, suggesting that Nrf2, indeed, contributes significantly to the induction of Akr7a. Moreover, loss of cell viability in Nrf2-depleted cells was significantly rescued by coexpression of AKR7A3. Furthermore, increased AKR7A3 in HepG2 cells was associated with the up-regulation of oxidative stress-related enzymes to enhance cellular antioxidant defense, which appeared to contribute significantly to protection against APAP-induced toxicity. In a line of transgenic rats overexpressing AKR7A1, increased AKR7A1 stimulated the expression of Nrf2 and other Nrf2-regulated genes, but did not better protect rats from APAP insults. In contrast, depletion of Akr7a5 in vitro in cultured AML12 cells or depletion of Akr7a1 in vivo in rat liver greatly increased APAPinduced hepatotoxicity. Conclusion: AKR7A proteins are significantly up-regulated in response to APAP/NAPQI exposure to contribute significantly to protection against APAP-induced hepatotoxicity. AKR7A mediates this protection, in part, through enhancing hepatocellular antioxidant defense. (HEPATOLOGY 2011;54:1322-1332 A cetaminophen (also called paracetamol, N-acetyl-p-aminophenol, Tylenol V R , and APAP) is a clinically widely used drug with potent analgesic and antipyretic activities. At therapeutic doses, most ingested APAP is metabolized to phenolic glucuronide and sulfate conjugates in the liver by glucuronyltransferases and sulfotransferases and is subsequently excreted in the urine 1 ; however, a small portion is oxidized by cytochrome P450 (CYP450) 2E1 to generate a highly reactive, cytotoxic intermediate: N-acetyl-pAbbreviations: AFAR, aflatoxin aldehyde reductase; AKR, aldo-keto reductase; AKR7A, aldo-keto reductase-7A; ALT, alanine aminotransferase; APAP, acetaminophen; ARE, anti-oxidant response element; AST, aspartate transaminase; Cat, catalase; CMV, cytomegalovirus; CYP450, cytochrome P450; DMSO, dimethylsulfoxide; ERE, estrogen response element; G6PDH, glucose-6-phosphate dehydrogenase; Gpx-1/2, glutathione peroxidase-1/2; GSH, glutathione; GST, glutathione S-transferase; IDH-1,...

Overexpression of Aldose Reductase Render Mouse Hepatocytes More Sensitive to Acetaminophen Induced Oxidative Stress and Cell Death

“…We previously demonstrated in APAP-induced hepatotoxicity that overexpression of aldo-keto reductase-7a (AKR7A) in HepG2 caused significant hepatoprotection against APAP-induced oxidative stress and cell death [14]. In the present study, we have shown that APAP-induced hepatotoxicity and increased expression levels of aldose reductase in AML-12, and plasmid overexpressing aldose reductase augmented APAP induced oxidative stress and cell death.…”

“…Moreover, inhibition of aldose reductase renders J774A.1 macrophage cell line more susceptible to acrolein or hydrogen peroxide-induced cell death [17]. Also we previously demonstrated that increased AKR7A3 in HepG2 cells was associated with the up-regulation of oxidative stress-related enzymes to enhance cellular antioxidant defense, which appeared to contribute significantly to protection against APAP-induced toxicity [14]. Therefore, we suggest that aldose reductase may provide a mechanism for the cellular detoxification of APAP.…”

Aldo-Keto Reductase Regulation by the Nrf2 System: Implications for Stress Response, Chemotherapy Drug Resistance, and Carcinogenesis

“…39,46 The human AKR genes AKR1B1 , AKR1B10 , AKR1C1 , AKR1C2 , AKR1C3 , AKR7A2 , and AKR7A3 are all regulated by the Keap1/Nrf2 system. 7,9,10,47–52 Although many studies measure NQO1 induction as a read out of Nrf2 activation, often the level of induction of NQO1 is a quite modest 2–3 fold by comparison to the induction seen in the AKR genes, which can be 1–2 orders of magnitude greater. 52,53 In fact, the AKR gene response can be one of the most robust signatures of Nrf2 activation in humans, yet less attention is placed on the regulation of these genes and its consequences.…”

“…Knockdown of Nrf2 in HepG2 cells led to a reduction of AKR7A3 mRNA and AKR7A3 protein and increased sensitivity to acetaminophen-induced cytotoxicity. 47 Analysis of the human AKR gene promoters identified the existence of multiple AREs in 13/15 human genes, and many have subsequently been found to be functional, Table 2. 46 …”

Adenosine 5′-monophosphate blocks acetaminophen toxicity by increasing ubiquitination-mediated ASK1 degradation

“…Although much APAP is metabolized via conjugation with glucuronic acid and sulfate and then excreted, a portion of APAP is converted by cytochrome P-450 metabolism to a reactive quinone form, N-acetyl-p-benzoquinone imine (NAPQI), which is inactivated by conjugation with glutathione (GSH) [4]. Once the pool of GSH is exhausted, any remained N-acetyl-p-benzoquinone imine covalently binds cellular macromolecules induces a series of molecular events that include alkylation of proteins, membrane lipid peroxidation, mitochondrial dysfunction, imbalance of intracellular calcium, formation of reactive oxygen species and reactive nitrogen species [5–6]. APAP-induced hepatocellular damage and necrotic and apoptotic cell death can result in severe centrilobular hepatotoxicity and acute liver failure [7–8].…”