Chlorogenic acid in the oxidative stress injury triggered by Shuang-Huang-Lian injection

Abstract

Injections of Chinese herbs are a novel approach to prepare traditional Chinese medicines. However, as injections of Chinese herbs have been extensively used, adverse drug reactions (ADRs) have been on the increase. Additionally, the mechanism for injections of Chinese herbs remains unclear. This study explored the potential role played by chlorogenic acid (CGA) in initiating oxidative stress injury triggered by the utilization of injections of Chinese herbs and the underlying mechanism. A total of 90 male Wistar rats were raised for varying periods by using Shuang-Huang-Lian (SHL) injection or CGA in diverse dosages. Western blot analysis examined the expression of nicotinamide adenine dinucleotide phosphate oxidase subunits, spectrophotometry was used to examine the activity taken by catalase, ELISA was used to examine the concentrations of inflammatory factors in serum, and intravital microscopy was employed to examine the microcirculation. The results showed that the excessive peroxide production induced by CGA in high-dose or SHL in the venule walls may well be through nicotinamide adenine dinucleotide phosphate oxidase along with a decline in the activity of catalase, and led to imbalance of basal levels of pro-(TNF-α) and anti-(IL-10) inflammatory cytokines. On the basis of the aforementioned results, the mechanism hidden behind the adverse effects of CGA induced by irrational use of Chinese herbal injection can be identified from a deeper perspective.

Keywords: Chinese herbal injection; NADPH oxidase; chlorogenic acid; drug safety; reactive oxygen species.

Figure 1.

The experimental flow diagram. IL-10, interleukin-10; TNF-α, tumor necrosis factor-α.

Figure 2.

Concentrations of IL-10 and TNF-α in rat serum. (A) IL-10 content (pg/ml); (B) TNF-α content (pg/ml); (C) the ratio of IL-10 to TNF-α. Data are expressed as mean ± SEM of six animals. *P<0.05 vs. control group (day 7), ^#p<0.05 vs. control group (day 14). IL-10, interleukin-10; TNF-α, tumor necrosis factor-α; L-CGA, low-dose CGA; L-SHL, low-dose SHL; H-CGA, high-dose CGA; H-SHL, high-dose SHL.

Figure 3.

The concentrations of CAT in rat serum. CAT activity analysis (U/ml). Data are expressed as mean ± SEM of six animals. *P<0.05 vs. control group (day 7), ^#p<0.05 vs. control group (day 14). L-CGA, low-dose CGA; L-SHL, low-dose SHL; H-CGA, high-dose CGA; H-SHL, high-dose SHL.

Figure 4.

The protein expression of Nox4, p22^phox and p47^phox in the ileum tissues of rats. (A and E) Representative western blots of Nox4, p22^phox and p47^phox at day 7 and 14. The protein expression of (B and F) Nox4, (C and G) p22^phox, and (D and H) p47^phox at day 7 and 14. Lower columns: quantification of the western blotting results shown in (A and E). Data are expressed as means ± SEM of three animals. *P<0.05 vs. control group (day 7), ^#p<0.05 vs. control group (day 14). L-CGA, low-dose CGA; L-SHL, low-dose SHL; H-CGA, high-dose CGA; H-SHL, high-dose SHL.

Figure 5.

The protein expression of Nox4, p22^phox and p47^phox in the lung tissues of rats. (A and E) Representative western blots of Nox4, p22^phox and p47^phox at day 7 and 14. The protein expression of (B and F) Nox4, (C and G) p22^phox, and (D and H) p47^phox at day 7 and 14. Lower columns: quantification of the western blotting results shown in (A and E). Data are expressed as means ± SEM of three animals. *P<0.05 vs. control group (day 7). L-CGA, low-dose CGA; L-SHL, low-dose SHL; H-CGA, high-dose CGA; H-SHL, high-dose SHL.

Figure 6.

The protein expression of Nox4, p22^phox and p47^phox in the brain tissues of rats. (A and E) Representative western blots of Nox4, p22^phox and p47^phox at day 7 and 14. The protein expression of (B and F) Nox4, (C and G) p22^phox, and (D and H) p47^phox at day 7 and 14. Lower columns: quantification of the western blotting results shown in (A and E). Data are expressed as means ± SEM of three animals. *P<0.05 vs. control group (day 7). L-CGA, low-dose CGA; L-SHL, low-dose SHL; H-CGA, high-dose CGA; H-SHL, high-dose SHL.