Avenanthramide-C ameliorate doxorubicin-induced hepatotoxicity via modulating Akt/GSK-3β and Wnt-4/β-Catenin pathways in male rats

Abstract

Background: Doxorubicin (DOX) drugs used in cancer treatment can cause various adverse effects, including hepatotoxicity. Natural-derived constituents have shown promising effects in alleviating chemotherapy-induced toxicities. This study addressed the effect of Avenanthramides-C (AVN-C) treatment in rats with DOX-indued hepatotoxicity.

Methods: AutoDock Vina was used for the molecular docking investigations. In silico toxicity prediction for AVN-C and DOX was performed using the Pro Tox-III server. Four groups of ten male Sprague-Dawley rats were created: Group 1 (Gp1) served as a negative control, Gp2 received an intraperitoneal (i.p.) injection of AVN-C (10 mg/kg), Gp3 received an i.p. dose of DOX (4 mg/kg) weekly for a month, and Gp4 received the same dose of DOX as G3 and AVN-C as G2. Histopathological, molecular, and biochemical analyses were conducted 1 month later.

Results: The study showed that treatment with AVN-C significantly ameliorated DOX-induced hepatotoxicity in rats by restoring biochemical alterations, boosting antioxidant activity, reducing inflammation, and modulating the Akt/GSK-3β and Wnt-4/β-Catenin signaling pathways in male rats.

Conclusion: This study is the first to demonstrate the therapeutic effects of AVN-C therapy on DOX-induced liver damage in male rats. Therefore, AVN-C could have a pronounced palliative effect on the hepatotoxicity caused by DOX treatment. These findings suggest that AVN-C could potentially alleviate the hepatotoxicity associated with DOX-based chemotherapy.

Keywords: anti-inflammatory; antioxidants; avenanthramides; doxorubicin; hepatotoxicity; signaling pathway.

FIGURE 1

Schematic illustration of experimental design showing the different groups under the study after different treatment settings.

FIGURE 2

(A) Molecular docking analysis shows the interaction of DOX with Akt-1, the conventional H-bond (LYS385 and ASP387); the electrostatic interactions via Pi-cation (LYS39); the hydrophobic interactions (LYS39, LEU52, and LYS386). (B) DOX interaction with GSK-3β, conventional H-bond (ASN95, THR8, SER9, and GLY202); carbon H- Bond (GLY202); the hydrophobic interactions (LEU88). (C) DOX interaction with Wnt-4; conventional H-bond (ARG83, ILE303, GLN81, and ASP239); Electrostatic Pi-cation interactions (ARG85 and LYS235); Hydrophobic Pi-Alkyl interaction (ALA302, ALA302, and ILE303). (D) DOX interaction with β-Catenin, conventional H-bond (ARG515, LEU781, GLU568, and ASN432); carbon H- Bond (CYC429); Pi-cation interaction (HIS470); Pi-Alkyl interaction (ALA775 and PHE777).

FIGURE 3

(A) Molecular docking analysis shows the interaction of AVN-C with Akt-1, the conventional H-bond (SER50 and THR389); the hydrophobic Pi-Alkyl interactions (LEU52 and PRO388). (B) AVN-C interaction with GSK-3β, conventional H-bond (ARG92, ASP90, and GLY202); the hydrophobic Pi-Pi Stacked interactions (PHE93); the hydrophobic Pi-Alkyl interactions (VAL87 and ALA11). (C) AVN-C interaction with Wnt-4; conventional H-bond (ARG86, TRP87, and PRO277); Hydrophobic Pi-Sigma interaction (LEU308); Hydrophobic Pi-Alkyl interaction (ARG85). (D) AVN-C interaction with β-Catenin, conventional H-bond (GLU571, ASP778, and ARG469); Unfavorable donor-donor bond (ASN516).

FIGURE 4

Changes of the body weight (g) (A) and the relative liver weight (%) (B) in the different groups. The values were represented as means ± S.D. (n = 10). IBW: initial body weight; FBW: final body weight; AVN-C: avenanthramide-C; DOX: doxorubicin. Means that do not share letters indicated significant differences (*p < 0.05).

FIGURE 5

Hepatic levels of malondialdehyde (MDA) (A), reduced glutathione (GSH) (B), superoxide dismutase (SOD) (C), and catalase (CAT) (D) in the different groups. AVN-C: avenanthramide-C; DOX: doxorubicin. The values were represented as means ± S.D. (n = 10). Means that do not share letters indicated significant differences (* p < 0.05).

FIGURE 6

Hepatic levels of tumor necrosis factor alpha (TNF-α) (A), nuclear factor kappa-B (NF-κB) (B), interleukin-6 (IL-6) (C), interleukin-1β (IL-1β) (D), and cyclooxygenase-2 (COX-2) (E) in the different groups. AVN-C: avenanthramide-C; DOX: doxorubicin. The values were represented as means ± S.D. (n = 10). Means that do not share letters indicated significant differences (* p < 0.01).

FIGURE 7

Hepatic levels of RAC-alpha serine/threonine-protein kinase (Akt) (A), Glycogen synthase kinase (GSK-3β) (B), Wnt family member 4 (Wnt-4) (C), and β-Catenin (D) in the different groups. AVN-C: avenanthramide-C; DOX: doxorubicin. The values were represented as means ± S.D. (n = 10). Means that do not share letters indicated significant differences (* p < 0.05).

FIGURE 8

Relative mRNA expression levels of the hepatic RAC-alpha serine/threonine-protein kinase (Akt) (A), Glycogen synthase kinase (GSK-3β) (B), Wnt family member 4 (Wnt-4) (C), and β-Catenin (D) in the different groups. AVN-C: avenanthramide-C; DOX: doxorubicin. The values were represented as means ± S.D. (n = 10). Means that do not share letters indicated significant differences (*p < 0.01).

FIGURE 9

(A) A photomicrograph of the liver section from the negative control group shows organized hepatic architecture, centered nucleus (arrows), central veins (CV), normal hepatocytes (h), normal blood sinusoids (Bs) and Kupffer cells (k). (B) Liver section of AVN-C control group shows mostly normal hepatocytes, nucleus with normal Bs and (k) (C) Liver section of DOX-injected group demonstrated disorganization hepatic structures, congested CV (cCV), cellular infiltrations (*), vacuolated cytoplasm (V), pyknotic nuclei (dotted arrows). (D) Liver section of the DOX + AVN-C-treated group demonstrated the improvement in the hepatic organization, fewer congestion, binucleated hepatocytes, and cellular infiltrations (H&E × 400, scale bar = 50 μm). (E) Liver sections histological scores in the different groups.

FIGURE 10

Diagram shows the effect of AVN-C against DOX-induced hepatotoxicity through inhibiting oxidative stress, inflammation, and modulating the Akt/GSK-3β and Wnt-4/β-Catenin pathways.