Holly (Ilex latifolia Thunb.) Polyphenols Extracts Alleviate Hepatic Damage by Regulating Ferroptosis Following Diquat Challenge in a Piglet Model

Abstract

Background: Holly (Ilex latifolia Thunb.) polyphenols extracts (HPE) contain high amounts of polyphenols, including phenolic acids, triterpenoids, tannic acids, and so on, which have strong antioxidant function. This experiment was aimed to explore the protective effect and mechanism of HPE against hepatic injury induced by diquat. Methods: Thirty-two weaned piglets were allotted by a 2 × 2 factorial experiment design with diet type (basal diet vs. HPE diet) and diquat challenge (saline vs. diquat). On the 21st day, piglets were injected with diquat or saline. One week later, blood samples were collected. Then all piglets were slaughtered and hepatic samples were collected. Results: Dietary HPE supplementation improves hepatic morphology, the activities of plasma aspartate aminotransferase, alanine aminotransferase, and glutamyl transpeptidase, and enhances hepatic anti-oxidative capacity, while it regulates the expression of ferroptosis mediators (transferrin receptor protein 1, heat shock protein beta 1, solute carrier family 7 member 11, and glutathione peroxidase 4) in diquat-challenged piglets. Conclusion: These results indicate that dietary HPE supplementation enhances hepatic morphology and function, which is involved in modulating antioxidant capacity and ferroptosis.

Keywords: diquat; ferroptosis; holly polyphenols extracts; liver; oxidative stress; weaned piglets.

Figure 1

Effect of holly polyphenols extracts (HPE) supplementation on hepatic morphology after 7 d diquat challenge in weanling[-2mm]Q7 piglets. Representative liver photomicrographs are shown. (A) Pigs fed a basal diet and treated with 0.9% NaCl solution. (B) Pigs fed a HPE diet and treated with 0.9% NaCl solution. No obvious pathological changes were observed. (C) Pigs fed the same basal diet and challenged with diquat. Significant pathological changes of liver damage such as hepatic spindle cells disappear (a), hepatocyte caryolysis (b), hepatocyte karyopycnosis (c), hepatic cell cords arrangement in disorder were found. (D) Pigs fed a HPE diet and challenged with diquat. Liver damage was significantly alleviated. n = 8 (1 pig/pen). Original magnifications 400×. Scale bars = 22.4 μm.

Figure 2

The effect of dietary holly polyphenols extracts (HPE) supplementation on hepatocyte ultrastructure after 7 d diquat challenge in weanling pigs. Representative hepatocyte ultrastructures are shown. (A) Pigs fed a basal diet and treated with 0.9% NaCl solution. (B) Pigs fed a HPE diet and treated with 0.9% NaCl solution. (A) and (B) were no obvious ferroptosis characteristics. Presented as complete mitochondria (a), mitochondria with distinct cristae (b), normal rough endoplasmic reticulum (c), karyotheca integrity (d). (C) Pigs fed the same basal diet and challenged with diquat. Significant ferroptosis characteristics were observed, such as mitochondrial pyknosis (e), mitochondrial outer membrane rupture (f), mitochondrial cristae reduction (g), and slightly dilatations of rough endoplasmic reticulum (h) were found. (D) Pigs fed a HPE diet and challenged with diquat.

Figure 3

The effect of dietary holly polyphenols extracts (HPE) on protein expression of TFR1 (B), SLC7A11 (C), and GPX4 (D) of weaned pigs after 7 d diquat challenge. The bands were the representative Western blot images (A). Values are mean and pooled SEM, n = 8 (1 pig/pen). CON_S, piglets fed the basal diet and injected with saline; HP_S, piglets fed HPE diet and injected with saline; CON_DIQ, piglets fed the basal diet and challenged with diquat; HP_DIQ, piglets fed HPE diet and challenged with diquat.