Astaxanthin from Haematococcus pluvialis prevents high-fat diet-induced hepatic steatosis and oxidative stress in mice by gut-liver axis modulating properties

Scope: Evidence is mounting that astaxanthin (ATX), a xanthophyll carotenoid, used as a nutritional supplement to prevent chronic metabolic diseases. The present study aims to identify the potential function of ATX supplementation in preventing steatohepatitis and hepatic oxidative stress in diet-induced obese mice. Methods and Results: In this study, ATX as dose of 0.25%, 0.5% and 0.75% have orally administered to mice along with a high-fat diet (HFD) to investigate the role of ATX in regulating liver lipid metabolism and gut microbiota. The study showed that ATX dose-dependently reduces body weight, lipid droplet formation, hepatic triglycerides and ameliorated hepatic steatosis and oxidative stress. 0.75% ATX altered the levels of 34 lipid metabolites related to hepatic cholesterol and fatty acid metabolism which might be associated with downregulation of lipogenesis-related genes and upregulation of bile acid biosynthesis-related genes. The result also revealed that ATX alleviates HFD-induced gut microbiota dysbiosis by significantly inhibiting the growth of obesity-related Parabacteroides and Desulfovibrio while promoting the growth of Allobaculum and Akkermansia. Conclusion: The study results suggested that dietary ATX may prevent the development of hepatic steatosis and oxidative stress with the risk of metabolic disease by gut-liver axis modulating properties.Please refer to ReadMe file.Funding provided by: National Natural Science Foundation of ChinaCrossref Funder Registry ID: http://dx.doi.org/10.13039/501100001809Award Number: 31902394The body weight and food intake were recorded daily for 63 days. To avoid error values, the measurement of weight was repeated three times for each mouse. Total RNA was extracted from liver tissue using TRIzol reagent (Shenggong BBI Life, Shanghai, China) according to the manufacturer's instructions. Then, cDNA was synthesized from total RNA using the PrimeScript Reverse Transcription reagent kit (Takara, Dalian, China). Quantitative PCR was conducted in triplicate for each group to detect gene expression. The quantitative analysis of AMPK, SREBP1c, ACC, CPT-1, PPARα, PPARγ, LXRα, SCD-1, PGC-1, FAS, CYP27A1, and CYP7A1 mRNA expression in the liver was measured in triplicate for each group by quantitative PCR. According to the SYBR Premix Ex Taq II (Takara, Dalian, China), the thermal cycle of qPCR was reacted on the CFX 96 Real-Time PCR Detection system (BIO-RAD, Hercules, CA, USA) under the following conditions: 95 °C for 10 min, then 40 cycles of 95 °C for 15 s, 60 °C for 30 s, and 72 °C for 30 s. Table S2 (supplement files) shows the PCR primer sequences of each gene, and the target genes were normalized to the reference gene GAPDH. The 2−ΔΔCt method was used to calculate relative gene expression. The caecal contents were sent to Shanghai Personal Biotechnology Co., Ltd. to investigate microbial diversity through 16S rRNA analysis on the Illumina MiSeq platform. When the microbial DNA was isolated, polymerase chain reaction (PCR) of the V3-V4 region of the bacterial 16S rRNA gene was performed using the forwards primer 5′-ACTCCTACGGGAGGCAGCA-3′ and the reverse primer 5′-GGACTACHVGGGTWTCTAAT-3′ according to the manufacturer's protocol (24-25). A previous study illustrated the analytical conditions and detailed parameters (18). All experiments were biologically repeated three times, and the data were analysed with Social Sciences (SPSS 16.0) statistical software and are presented as the mean ± SD. Multiple comparisons among treatments were statistically analysed using Duncan's multiple range test in one-way analysis of variance (ANOVA) (P < 0.05). Origin 9.1 was used to draw charts