Protective effect of the new prepared Atractylodes macrocephala Koidz polysaccharide on fatty liver hemorrhagic syndrome in laying hens

Abstract

Fatty liver hemorrhage syndrome (FLHS) is the most common noninfectious cause of death in backyard chickens worldwide, which can cause a sudden drop in egg production in the affected flocks and cause huge losses to the laying hens breeding industry. In this study, we prepared polysaccharide from Atractylodes macrocephala Koidz (PAMK) by one-step alcohol precipitation. The structural analysis found that PAMK with a molecular weight of 2.816 × 10³ Da was composed of glucose and mannose, in a molar ratio of 0.582 to 0.418. Furthermore, we investigated the hepatoprotective effects of PAMK on high-energy and low-protein (HELP) diet-induced FLHS in laying hens. The results showed that the hens' livers of the HELP diet showed yellowish-brown, greasy, and soft, whereas the supplement of PAMK (200 mg/kg or 400 mg/kg) could alleviate such pathological changes. The liver index, the abdominal fat percentage, and liver injury induced by the HELP diet were reduced in PAMK (200 mg/kg or 400 mg/kg). Supplementing 200 mg/kg or 400 mg/kg PAMK showed improvements of the antioxidant capacity in laying hens. Furthermore, we found that the HELP diet increased the expression of hepatic lipogenesis genes and decreased the expression of fatty acid β-oxidation genes, which could be reversed by 200 mg/kg or 400 mg/kg PAMK supplementation. Nevertheless, there is no difference between the addition of 40 mg/kg PAMK and the HELP group. Collectively, these results showed that PAMK supplements could ameliorate HELP diet-induced liver injury through regulating activities of antioxidant enzymes and hepatic lipid metabolism. Therefore, PAMK could be a potential feedstuff additive to alleviate FLHS in laying hens.

Keywords: Atractylodes macrocephala Koidz; fatty liver hemorrhagic syndrome; lipid metabolism; polysaccharide.

Figure 1

Characterization of PAMK. (A) UV spectrum of PAMK. (B) Standard monosaccharides and PAMK analysis by GC-MS. 1, Rhamnose; 2, fucose; 3, arabinose; 4, xylose; 5, mannose; 6, glucose; 7, galactose. (C) FT-IR spectra of PAMK. (D) ¹H spectrum and (E) ¹³C spectrum of PAMK. Abbreviations: FT-IR, Fourier transform–infrared; GC-MS, gas chromatography–mass spectrometry; PAMK, polysaccharide from Atractylodes macrocephala Koidz; UV, ultraviolet.

Figure 2

Weekly egg laying rate of the hens. ∗∗P < 0.01 as compared with the control group; ^#P < 0.05 as compared with the HELP group; ^##P < 0.01 as compared with the HELP group (n = 10). Abbreviations: CON, control; HELP, high energy and low protein; HELP + P-40, high energy and low protein +40 mg/kg PAMK; HELP + P-200, high energy and low protein +200 mg/kg PAMK; HELP + P-400, high energy and low protein +400 mg/kg PAMK; PAMK, polysaccharide from Atractylodes macrocephala Koidz.

Figure 3

The effect of PAMK in the liver index and abdominal fat percentage. (A) Liver index. (B) Abdominal fat percentage. ∗∗P < 0.01 as compared with the CON group; ^#P < 0.05 as compared with the HELP group; ^##P < 0.01 as compared with the HELP group (n = 10). Abbreviations: CON, control; HELP, high energy and low protein; HELP + P-40, high energy and low protein +40 mg/kg PAMK; HELP + P-200, high energy and low protein +200 mg/kg PAMK; HELP + P-400, high energy and low protein +400 mg/kg PAMK; PAMK, polysaccharide from Atractylodes macrocephala Koidz.

Figure 4

Pathological observation results of the liver. (A) Histopathological observation. (B) H&E staining × 200. (C) Oil Red O staining × 400. Abbreviations: CON, control; H&E, Hematoxylin-eosin staining; HELP, high energy and low protein; HELP + P-40, high energy and low protein +40 mg/kg PAMK; HELP + P-200, high energy and low protein +200 mg/kg PAMK; HELP + P-400, high energy and low protein +400 mg/kg PAMK; PAMK, polysaccharide from Atractylodes macrocephala Koidz.

Figure 5

(A) ALT and (B) AST levels in plasma. Values were expressed as mean ± SEM in each group. ∗∗P < 0.01 as compared with the CON group; ^#P < 0.05 as compared with the HELP group; ^##P < 0.01 as compared with the HELP group (n = 10). Abbreviations: ALT, alanine aminotransferase; AST, aspartate aminotransferase; CON, control; HELP, high energy and low protein; HELP + P-40, high energy and low protein +40 mg/kg PAMK; HELP + P-200, high energy and low protein +200 mg/kg PAMK; HELP + P-400, high energy and low protein +400 mg/kg PAMK; PAMK, polysaccharide from Atractylodes macrocephala Koidz.

Figure 6

Biochemical indexes change in plasma and liver tissue homogenate. (A) TG, (B) TC, (C) HDL-C, and (D) LDL-C levels in plasma. (E) TG, (F) TC, (G) HDL-C, and (H) LDL-C levels in hepatic. Values were expressed as mean ± SEM in each group. ∗∗P < 0.01 as compared with the CON group; ^#P < 0.05 as compared with the HELP group; ^##P < 0.01 as compared with the HELP group (n = 10). Abbreviations: CON, control; HDL-C, high-density lipoprotein cholesterol; HELP, high energy and low protein; HELP + P-40, high energy and low protein +40 mg/kg PAMK; HELP + P-200, high energy and low protein +200 mg/kg PAMK; HELP + P-400, high energy and low protein +400 mg/kg PAMK; LDL-C, low-density lipoprotein cholesterol; PAMK, polysaccharide from Atractylodes macrocephala Koidz; TC, total cholesterol; TG, triglyceride.

Figure 7

The effects of PAMK on antioxidant capacities. (A) Antioxidant enzyme activity in liver. (B) The content of MDA in liver. (C) Quantitative real-time PCR determination of hepatic mRNA expression of CAT, HO-1, GSH-PX, NQO1. Values were expressed as mean ± SEM in each group. ∗∗P < 0.01 as compared with the CON group; ^#P < 0.05 as compared with the HELP group; ^##P < 0.01 as compared with the HELP group (n = 10). Abbreviations: CAT, catalase; CON, control; GSH-Px, glutathione peroxidase; HELP, high energy and low protein; HELP + P-40, high energy and low protein +40 mg/kg PAMK; HELP + P-200, high energy and low protein +200 mg/kg PAMK; HELP + P-400, high energy and low protein +400 mg/kg PAMK; HO-1, heme oxygenase; MDA, malondialdehyde; NQO1, quinone oxidoreductase; PAMK, polysaccharide from Atractylodes macrocephala Koidz; T-SOD, total superoxide dismutase.

Figure 8

The effects of PAMK on hepatic lipid metabolism. Quantitative real-time PCR determination of hepatic mRNA expression of genes involved in fatty acid synthesis (A), fatty acid β-oxidation (B). Values were expressed as mean ± SEM in each group. ∗∗P < 0.01 as compared with the CON group; ^#P < 0.05 as compared with the HELP group; ^##P < 0.01 as compared with the HELP group (n = 10). Abbreviations: ATGL, adipose triglyceride lipase; CON, control; CPT-1, carnitine palmitoyl transferase-1; FAS, fatty acid synthase; HELP, high energy and low protein; HELP + P-40, high energy and low protein +40 mg/kg PAMK; HELP + P-200, high energy and low protein +200 mg/kg PAMK; HELP + P-400, high energy and low protein +400 mg/kg PAMK; ACC, acetyl-CoA carboxylase; PAMK, polysaccharide from Atractylodes macrocephala Koidz; PPARα, peroxisome proliferators–activated receptor-α; SCD-1, stearoyl coenzyme dehydrogenase-1.