Lipidomics study of the therapeutic mechanism of Plantaginis Semen in potassium oxonate-induced hyperuricemia rat

Abstract

Background: Plantaginis Semen has been widely used as folk medicine and health care food against hyperuricemia (HUA) and gout, but its pharmacological mechanism remains unclear. This study investigated the therapeutic mechanism of Plantaginis Semen extract on potassium oxonate -induced HUA rats based on a lipidomics approach.

Methods: A model of HUA was established by potassium oxonate intragastric administration. 42 Sprague-Dawley (SD) male rats were randomly divided into the control group, model group, benzbromarone group (10 mg/kg) and three Plantaginis Semen groups (n = 7). The Plantaginis Semen groups were treated orally with Plantaginis Semen, 0.9375, 1.875 or 3.75 g/kg for 28 days. The levels of serum uric acid (UA), creatinine (Cr), triacylglycerol (TG) and tumor necrosis factor-α (TNF-α) were measured using enzyme-linked immunosorbent assay kits. Ultra performance liquid chromatography quadrupole time of flight mass spectrometry (UPLC-Q-TOF/MS) was used for the serum lipidomics analysis, multivariate statistical analysis and independent samples t-test were carried out for the pattern recognition and characteristic metabolites identification. The relative levels of critical regulatory factors were determined by quantitative real-time polymerase chain reaction (RT-qPCR).

Results: Compared with the model group, the levels of serum UA, Cr, TG and TNF-α were significantly (p < 0.05) decreased in benzbromarone and three Plantaginis Semen groups. With lipidomics analysis, significant lipid metabolic perturbations were observed in HUA rats, 13 metabolites were identified as potential biomarkers and glycerophospholipid metabolism pathway was most affected. These perturbations were partially restored via treatment of benzbromarone and Plantaginis Semen. Additionally, the mRNA expression levels of urate anion transporter 1 (URAT1) and phosphatidylinositol 3-kinase/protein kinases B (PI3K/Akt) were significantly decreased (p < 0.01) after treatment with benzbromarone and high dose of Plantaginis Semen.

Conclusions: Plantaginis Semen had significant effects on anti-HUA, anti-inflammatory and renal protection. It attenuated potassium oxonate-induced HUA through regulation of lipid metabolism disorder.

Keywords: Hyperuricemia; Lipid metabolism disorder; Lipidomics; Lowering uric acid; Plantaginis Semen.

Fig. 1

The results of serum biochemistry analysis. a UA level, b Cr level, c TG level, d TNF-α level. C, control group; M, model group; Y, benzbromarone group; CL, low dosage group; CM, medium dosage group; CH, high dosage group; values are given as the mean ± SD (n = 7), ANOVA, Tukey post hoc and nonparametric test were used for statistical analysis, **, P < 0.01 vs. control group. *, P < 0.05 vs. control group; ##, P < 0.01 vs. model group; #, P < 0.05 vs. model group

Fig. 2

The PCA score plot derived from UPLC-Q-TOF/MS profiles of serum sample from control group and model group

Fig. 3

OPLS-DA score plots (a) and the corresponding validation plots (b) with 200 times permutation tests obtained

Fig. 4

Metabolic profiles of rat serum in the control, model, benzbromarone, and different dose Plantaginis semen groups. a Metabolic profiles of rat serum in C, M and Y group, b Metabolic profiles of rat serum in C, M and CL group, c Metabolic profiles of rat serum in C, M and CM group, d Metabolic profiles of rat serum in C, M and CH group. C, control group; M, model group; Y, benzbromarone group; CL, low dosage group; CM, medium dosage group; CH, high dosage group

Fig. 5

Heat map analysis of relative contents of potential metabolites. (green through dark red corresponding to a progressive increase in concentration). C, control group; M, model group; Y, benzbromarone group; CL, low dosage group; CM, medium dosage group; CH, high dosage group

Fig. 6

Comparison of 13 biomarkers peak relative signal intensities in 6 groups. C, control group; M, model group; Y, benzbromarone group; CL, low dosage group; CM, medium dosage group; CH, high dosage group; boxplots show the 25, 50 and 75% percentiles. ANOVA, Tukey post hoc and nonparametric test were used for statistical analysis, **, P < 0.01 vs. control group. *, P < 0.05 vs. control group; ##, P < 0.01 vs. model group; #, P < 0.05 vs. model group

Fig. 7

six pathway related to changed biomarkers. a: Glycerophospholipid metabolism, b: Linoleic acid metabolism, c: Glycosylphosphatidylinositol (GPI)-anchor biosynthesis, d: alpha-Linolenic acid metabolism, e: Arachidonic acid metabolism, f: Steroid biosynthesis

Fig. 8

KEGG global metabolic network related to changed biomarkers. The purple textboxes represented the pathways, the yellow and green textboxes represented the significant and no detection metabolites. The arrows in red represented the up regulated metabolites. The arrows in blue represented direct or indirect connections between two metabolites

Fig. 9

Effects of Plantaginis semen on PI3k/Akt and UTAR1 mRNA expression. a mRNA expression levels of URAT1, b mRNA expression levels of PI3k, c mRNA expression levels of Akt. C, control group; M, model group; Y, benzbromarone group; CL, low dosage group; CM, medium dosage group; CH, high dosage group; values are given as the mean ± SD(n = 7), ANOVA, Tukey post hoc and nonparametric test were used for statistical analysis, **, P < 0.01 vs. control group. *, P < 0.05 vs. control group; ##, P < 0.01 vs. model group; #, P < 0.05 vs. model group