Involvement of Nrf2, JAK and COX pathways in acetaminophen-induced nephropathy: Role of some antioxidants

Abstract

Objectives: Acetaminophen (APAP)-induced nephrotoxicity is detrimental consequence for which there has not been a standardized therapeutic regimen. Although, N-acetylcysteine (NAC) is a well-known antidote used in APAP-induced hepatotoxicity, its benefit in nephrotoxicity caused by APAP is almost lacking. This study aimed to compare the possible protective effect of thymoquinone (TQ), curcumin (CR), and α-lipoic acid (α-LA), either in solo or in combination regimens with that of NAC against APAP-induced renal injury.

Design and method: Rats were divided into nine groups; control group, APAP intoxicated group (1000 mg/kg; orally), and the remaining seven groups received, in addition to APAP, oral doses of NAC, TQ, CR, α-LA, CR plus TQ, TQ plus α-LA, or CR plus α-LA. The first dose of the aforementioned antioxidants was given 24 h before APAP, and then the second dose was given 2 h after APAP, whereas the last dose was given 10 h after administration of APAP.

Results: Treatment with APAP elevated kidney markers like serum uric acid, urea, and creatinine. In addition, it increased the serum level of tumor necrosis factor alpha (TNF-α), interleukin-1beta (IL-1β) and thiobarbituric acid reactive species (TBARS). Also, the protein expression of renal janus kinase (JAK) and cyclooxygenase (COX)-2 were all upregulated by APAP. In contrast, the expression of Nrf2 and the renal levels of superoxide dismutase and glutathione were downregulated. Treatment with the indicated natural antioxidants resulted in amelioration of the aberrated parameters through exhibiting anti-inflammatory, antioxidant and free radical-scavenging effects with a variable degree.

Conclusion: The combined administration of CR and TQ exerted the most potent protection against APAP-induced nephrotoxicity through its anti-inflammatory and free radical-scavenging effects (antioxidant) which were comparable to that of NAC-treatment.

Keywords: Acetaminophen; COX; Curcumin; JAK; N-acetylcysteine; Nrf2; Thymoquinone; α-Lipoic acid.

Fig. 1

NAC, CR, TQ, α-LA alone or in combination attenuate renal oxidative stress in a single dose of APAP-intoxicated rats. (a) Thiobarbituric Acid Reactive Species (TBARS) (b) superoxide dismutase (SOD), (c) glutathione (GSH) and (d) nitric oxide (NO) levels in the kidney tissue of rats were measured following a single dose of APAP followed by treatment with NAC, CR, TQ, α-LA or their selected combinations. Data are expressed as mean ± SEM (N = 6). ^***P ≤ 0.001 vs control, ^πππP ≤ 0.001 vs APAP group, ^$$$P ≤ 0.01, ^$P ≤ 0.05 vs NAC group.

Fig. 2

Protein expression of JAK and Nrf2 in renal tissue following administration of a single dose of APAP and all treated group. Western blot was used to determine the protein expression in treatment groups. After treatments, sections of kidney were homogenized and the total protein was extracted and subjected to separation on SDS–PAGE gel, then protein bands were transferred to PVDF membranes. After blocking, the membranes were incubated with the primary antibodies of JAK and Nrf2 using β-actin as an internal control. After incubation with the corresponding secondary antibodies, the membranes were visualized with ECL reagent exposed to X-ray. Data are expressed as mean ± SEM (N = 6). ^***P ≤ 0.001 vs control, ^πππP ≤ 0.001 vs APAP group, P ≤ 0.01, ^$$$P ≤ 0.001 vs NAC group.

Fig. 3

Protein expression of COX1 and COX2 in renal tissue following administration of a single dose of APAP and all treated group. Western blot was used to determine the protein expression in treatment groups. After treatments, sections of kidney were homogenized and the total protein was extracted and subjected to separation on SDS–PAGE gel, then protein bands were transferred to PVDF membranes. After blocking, the membranes were incubated with the primary antibodies of COX1 and COX2 using β-actin as an internal control. After incubation with the corresponding secondary antibodies, the membranes were visualized with ECL reagent exposed to X-ray. Data are mean ± SEM (N = 6). ^***P ≤ 0.001 vs control, ^πππP ≤ 0.001 vs APAP group, P ≤ 0.01, ^$$$P ≤ 0.001 vs NAC group.