Effects of Antioxidant N-acetylcysteine Against Paraquat-Induced Oxidative Stress in Vital Tissues of Mice

Abstract

Paraquat (PQ) is a commonly used herbicide that induces oxidative stress via reactive oxygen species (ROS) generation. This study aimed to investigate the effects of the antioxidant N-acetylcysteine (NAC) against PQ-induced oxidative stress in mice. Male Balb/C mice (24) were randomly divided into 4 groups and treated for 3 weeks: 1) control (saline), 2) NAC (0.5% in diet), 3) PQ (20 mg/kg, IP) and 4) combination (PQ + NAC). Afterwards mice were sacrificed and oxidative stress markers were analyzed. Our data showed no significant change in serum antioxidant capacity. PQ enhanced lipid peroxidation (MDA) levels in liver tissue compared to control whereas NAC decreased MDA levels (p<0.05). NAC significantly increased MDA in brain tissue (p<0.05). PQ significantly depleted glutathione (GSH) levels in liver (p=0.001) and brain tissue (p<0.05) but non-significant GSH depletion in lung tissue. NAC counteracted PQ, showing a moderate increase GSH levels in liver and brain tissues. PQ significantly increased 8-oxodeoxyguanosine (8-OH-dG) levels (p<0.05) in liver tissue compared to control without a significant change in brain tissue. NAC treatment ameliorated PQ-induced oxidative DNA damage in the liver tissue. PQ significantly decreased the relative mtDNA amplification and increased the frequency of lesions in liver and brain tissue (p<0.0001), while NAC restored the DNA polymerase activity in liver tissue but not in brain tissue. In conclusion, PQ induced lipid peroxidation, oxidative nuclear DNA and mtDNA damage in liver tissues and depleted liver and brain GSH levels. NAC supplementation ameliorated the PQ-induced oxidative stress response in liver tissue of mice.

Keywords: Antioxidant Capacity; DNA Integrity; Glutathione levels; MDA levels; Mice; Oxidative Stress; Reactive Oxygen Species.

Figure 1

Antioxidant capacity in term of Trolox (mM) equivalent units in mice serum. Bars represent mean ±1.96 (SD) with distribution of samples per group in scatter blot column. Interval of confidence at 95% were as follow: Control: 2.54+/−1.96 (0.40), NAC: 1.74 +/− 1.96 (0.31), PQ: 1.71 +/− 1.96 (0.30), and PQ+NAC: 2.32 +/− 1.96 (0.34). The sample size of the experimental groups was as follow: Control (N=5), NAC (N=6), PQ (N=7), and PQ+NAC (N=7).

Figure 2

Glutathione (GSH) levels (μM) of (A) liver, (B) brain and (C) lung tissue of mice. Bars represent mean ±1.96 (SD) with distribution of samples per group in scatter blot column. (A) Interval of confidence at 95% were as follow: Control: 5.40+/−1.96 (0.42), NAC: 4.66 +/− 1.96 (0.43), PQ: 1.82 +/− 1.96 (0.40), and PQ+NAC: 3.34 +/− 1.96 (0.55). The sample size of the experimental groups was as follow: Control (N=6), NAC (N=7), PQ (N=5), and PQ+NAC (N=6). (B) Interval of confidence at 95% were as follow: Control: 22.50 +/−1.96 (0.80), NAC: 35.03 +/− 1.96 (1.35), PQ: 10.86 +/− 1.96 (1.24), and PQ+NAC: 23.01 +/− 1.96 (1.52). The sample size of the experimental groups was as follow: Control (N=4), NAC (N=5), PQ (N=6), and PQ+NAC (N=5). (C) Interval of confidence at 95% were as follow: Control: 0.09 +/−1.96 (0.19), NAC: 1.60 +/− 1.96 (0.30), PQ: 0 +/− 1.96 (0) and PQ+NAC: 2.92 +/− 1.96 (0.56). The sample size of the experimental groups was as follow: Control (N=6), NAC (N=7), PQ (N=7), and PQ+NAC (N=7).

Figure 3

Lipid peroxidation (MDA) levels of (A) liver, (B) brain and (C) lung tissue of mice. Bars represent mean ±1.96 (SD) with distribution of samples per group in scatter blot column. (A) Interval of confidence at 95% were as follow: Control: 11.02+/−1.96 (0.47), NAC: 12.92 +/− 1.96 (0.66), PQ: 16.77 +/− 1.96 (0.64), and PQ+NAC: 14.69 +/− 1.96 (0.78). The sample size of the experimental groups was as follow: Control (N=6), NAC (N=7), PQ (N=7), and PQ+NAC (N=7). (B) Interval of confidence at 95% were as follow: Control: 6.33+/−1.96 (0.33), NAC: 13.28 +/− 1.96 (0.71), PQ: 11.58 +/− 1.96 (0.59), and PQ+NAC: 14.47 +/− 1.96 (0.84). The sample size of the experimental groups was as follow: Control (N=6), NAC (N=7), PQ (N=7), and PQ+NAC (N=6). (C) Interval of confidence at 95% were as follow: Control: 23.31+/−1.96 (1.18), NAC: 22.93 +/− 1.96 (1.18), PQ: 21.46 +/− 1.96 (1.01), and PQ+NAC: 22.42 +/− 1.96 (0.91). The sample size of the experimental groups was as follow: Control (N=6), NAC (N=7), PQ (N=7), and PQ+NAC (N=7).

Figure 4

Mitochondrial DNA damage (A) Relative amplification of mtDNA and (B) Poisson analysis for detection of lesions frequency in mtDNA both of liver and brain tissues of mice. Bars represent mean ±1.96 (SD) with distribution of samples per group in scatter blot column. (A) Relative amplification for liver tissue-Interval of confidence at 95% were as follow: Control: 1+/−1.96(0), NAC: 0.92 +/− 1.96 (0.10), PQ: 0.71 +/− 1.96 (0.07), and PQ+NAC: 0.90 +/− 1.96 (0.09). The sample size of the experimental groups was as follow: Control (N=5), NAC (N=5), PQ (N=4), and PQ+NAC (N=5). Relative amplification for brain tissue-Interval of confidence at 95% were as follow: Control: 1+/−1.96(0), NAC: 0.95 +/− 1.96 (0.18), PQ: 0.63 +/− 1.96 (0.16), and PQ+NAC: 0.69 +/− 1.96 (0.15). The sample size of the experimental groups was as follow: Control (N=5), NAC (N=4), PQ (N=5), and PQ+NAC (N=5). (B) Poisson analysis for liver tissue-Interval of confidence at 95% were as follow: NAC: 0.07 +/− 1.96 (0.10), PQ: 0.33 +/− 1.96 (0.08), and PQ+NAC: 0.07 +/− 1.96 (0.10). The sample size of the experimental groups was as follow: NAC (N=5), PQ (N=4), and PQ+NAC (N=5). Poisson analysis for brain tissue-Interval of confidence at 95% were as follow: NAC: 0.11 +/− 1.96 (0.14), PQ: 0.47 +/− 1.96 (0.20), and PQ+NAC: 0.30 +/− 1.96 (0.16). The sample size of the experimental groups was as follow: NAC (N=4), PQ (N=5), and PQ+NAC (N=4).

Figure 5

The levels of oxidized DNA base 8-oxodeoxyguanosine (8-OHdG) in (A) liver and (B) brain tissues of mice. Bars represent mean ±1.96 (SD) with distribution of samples per group in scatter blot column. (A) Interval of confidence at 95% was as follow: Control: 1.15+/−1.96 (0.30), NAC: 1.02 +/− 1.96 (0.19), PQ: 1.46 +/− 1.96 (0.19), and PQ+NAC: 0.85 +/− 1.96 (0.22). The sample size of the experimental groups was as follow: Control (N=5), NAC (N=5), PQ (N=5), and PQ+NAC (N=4). (B) Interval of confidence at 95% were as follow: Control: 1.50+/−1.96 (0.31), NAC: 1.33 +/− 1.96 (0.20), PQ: 1.82 +/− 1.96 (0.34), and PQ+NAC: 1.37 +/− 1.96 (0.25). The sample size of the experimental groups was as follow: Control (N=4), NAC (N=5), PQ (N=4), and PQ+NAC (N=4).