The Protective Impact of Salsola imbricata Leaf Extract From Taif Against Acrylamide-Induced Hepatic Inflammation and Oxidative Damage: The Role of Antioxidants, Cytokines, and Apoptosis-Associated Genes

Abstract

Salsola imbricata is a herbal plant native to Saudi Arabia, known for its antioxidative and anti-inflammatory properties. This study explored the protective effects of an ethanolic leaf extract of Salsola imbricata against the oxidative stress and hepatic injury caused by acrylamide. Rats received intragastric administrations of 20 mg/kg of body weight of acrylamide to induce hepatic injury, or 300 mg/kg of body weight of Salsola ethanolic extract orally for 7 days before acrylamide administration. The treatments were continued for 3 weeks. Blood and liver samples were collected from all the groups, and the following biochemical parameters were tested: serum ALT (alanine aminotransferase), AST (aspartate aminotransferase), GGT (gamma glutaryl transferase), urea, albumin, total proteins, catalase, SOD (superoxide dismutase), reduced glutathione (GSH), nitric oxide (NO), and MDA (malondialdehyde). Quantitative real-time PCR (qRT-PCR) was used to examine the expression of Nrf2 (Nuclear factor-erythroid factor 2-related factor 2), HO-1 (Hemoxygenase-1), COX-2 (Cyclooxgenase-2), TGF-β1 (transforming growth factor-beta1), Bax, and Bcl2 (B-cell lymphoma 2), which are associated with oxidative stress, fibrosis, apoptosis, and anti-apoptotic effects. The annexin and survivin immunoreactivity were examined at the immunohistochemical level. Pretreatment with the Salsola ethanolic extract reduced the negative impact of acrylamide on ALT, AST, GGT, urea, albumin, and total proteins. The Salsola ethanolic extract reversed acrylamide's effects on serum and tissue antioxidants. Nrf2/HO-1 expression was downregulated, while COX-2 and TGF-β1 were upregulated in the acrylamide-administered group and normalized by the pre-administration of Salsola ethanolic extract to the acrylamide experimental group. The immunoreactivity of annexin and survivin was restored in the experimental group administered Salsola ethanolic extract plus acrylamide. In conclusion, Salsola ethanolic extract inhibits and regulates the side effects induced in the liver by acrylamide. Salsola induced its impacts by regulating inflammation, oxidative stress, and apoptosis-/anti-apoptosis-associated genes at the biochemical, molecular, and cellular levels. Salsola is recommended as oxidative stress relievers against environmental toixicity at high altitude areas.

Keywords: Salsola imbricata; acrylamide; antioxidants; gene expression; hepatic toxicity; oxidative stress.

Figure 1

A schematic diagram for the experimental design used in the current study.

Figure 2

Total phenols and flavonoids of an ethanolic extract of Salsola imbricata.

Figure 3

The ameliorative impact of Salsola imbricata extract on the mRNA expression of catalase and SOD in acrylamide-administered rats. Graphic presentation of hepatic expression according to qRT-PCR analysis of catalase and SOD in different groups of rats after normalization with the housekeeping gene (actin). *p < 0.05 vs. the control group; ^#p < 0.05 vs. the other groups; ^$p < 0.05 vs. the acrylamide-administered group.

Figure 4

The ameliorative impact of Salsola imbricata extract on the mRNA expression of HO-1 and Nrf2 in acrylamide-administered rats (n = 10). Graphic presentation of hepatic expression by qRT-PCR analysis of HO-1 and Nrf2 in different groups of rats after normalization with the housekeeping gene (actin). *p < 0.05 vs. the control group; ^$p < 0.05 vs. the acrylamide-administered group.

Figure 5

The impact of Salsola imbricata extract on the mRNA expression of COX-2 and TGF-β1 in acrylamide-administered rats (n = 10). Graphic presentation of hepatic expression according to qRT-PCR analysis of COX-2 and TGF-β1 in different groups of rats after normalization with the housekeeping gene (actin). *p < 0.05 vs. the control group; ^$p < 0.05 vs. the acrylamide-administered group.

Figure 6

The impact of Salsola imbricata extract on the mRNA expression of Bax and Bcl2 in acrylamide-administered rats (n = 10). Graphic presentation of hepatic expression according to qRT-PCR analysis of Bax and Bcl2 in different groups of rats after normalization with the housekeeping gene (actin). *p < 0.05 vs. the control group; ^#p < 0.05 vs. the other groups; ^$p < 0.05 vs. the acrylamide-administered group.

Figure 7

Photomicrographs of hepatic sections stained with hematoxylin and eosin in the control group (A), the Salsola-treated group (C), the acrylamide-treated group (B), and the group co-treated with acrylamide and Salsola (D). Sections from control and Salsola extract groups (A,C) showed the liver consisted of central vein (CV) surrounded by hepatic cords (h). The cords consisted of large hepatocytes with centrally located nuclei and acidophilic cytoplasm. Sections from the acrylamide-treated rats (B) the liver tissue showed vacular degeneration (v), hydrobic degeneration (hy). The liver tissues showed proliferation of the vonkupher cells (k). The blood sinusoids showed lymphocytic infiltrates (s). Some heaptocytes cells were characterized by necrosis (n) table (6). Co-treatment of rats with acrylamide and Salsola (D) both of the vacuolar and the hydrobic degeneration were localized to few hepatocyte cells. Some cells still in necrotic pattern (n). The vonkupher cells still proliferated but not numerous compared with acrylamide group. Scale bar = 20 μm (original magnification = 200 ×). The morphometric analysis and scoring of Salsola imbricata extract against acrylamide toxicity were shown on Table 6. Scale bar = 20 μm (original magnification = 200 ×).

Figure 8

Photomicrographs of hepatic sections immunostained with annexin antibody in the control group (A), the Salsola-treated group (C), the acrylamide-treated group (B), and the group co-treated with Salsola and acrylamide (D). The intensity of immunostaining was weak in the control and Salsola-treated groups (A,C), strong in the acrylamide-treated group (B), and reduced to weak or moderate in the co-treated rats (D). Reactive liver cells (arrows) are frequently seen next to central veins (CVs). Scale bar = 20 μm (original magnification = 200 ×). The degree of positive immunoreactivity for annexin is graphed in (E). Densitometric values are statistically significant at *p < 0.05 vs. the control and Salsola-treated groups; ^#p < 0.05 vs. the acrylamide-administered group.

Figure 9

Photomicrographs of hepatic sections immunostained with survivin antibody, in the control group (A), the Salsola-treated group (C), the acrylamide-treated group (B), and the group co-treated with Salsola and acrylamide (D). The intensity of immunostaining was high in the control and Salsola-treated groups (A,C), reduced in the acrylamide-treated group (B), and restored to moderate levels in the co-treated rats (D). Reactive liver cells (arrows) are usually next to central veins (CVs). Scale bar = 20 μm (original magnification = 200 ×). The degree of positive immunoreactivity for survivin is graphed in (E). Densitometric values are statistically significant at *p < 0.05 vs. the control and Salsola-treated groups; ^#p < 0.05 vs. the acrylamide-administered group.

Figure 10

Graphical abstract represents the hepatoprotective impact of Salsola imbricata extract on acrylamide-induced inflammation, oxidative stress, and liver dysfunction.