Lipidomics study of Liujunzi decoction in hyperlipidemia rats with spleen deficiency based on UPLC-Q-TOF/MS

Abstract

Hyperlipidemia refers to the abnormal levels of triglyceride (TG), total cholesterol (TC), low-density lipoprotein (LDL-C) and high-density lipoprotein (HDL-C) in peripheral blood circulation. It is a predominant risk factor underlying cardiovascular and cerebrovascular diseases, including coronary heart disease and atherosclerosis. Furthermore, it is also one of the most prevalent chronic diseases globally. Liujunzi Decoction is the basic prescription for the treatment of spleen and stomach diseases. It can tonify the spleen and qi, remove dampness, and reduce turbidity. Moreover, it is also clinically used for the treatment of spleen deficiency hyperlipidemia. However, its metabolites and therapeutic effect on spleen deficiency hyperlipidemia have not been comprehensively determined in vitro and in vivo. This study established a rat model of spleen deficiency hyperlipidemia by inducing starvation and satiety disorders, exhaustion swimming, and intragastric administration of the fat emulsion. To identify related metabolite changes and serum lipid composition, UPLC-Q-TOF-MS, PCA, and OPLS-DA lipidological methods were performed. The results demonstrated significant changes in rat's signs during the modeling process, which were consistent with the criteria for the syndrome differentiation of spleen deficiency in traditional Chinese medicine. Furthermore, this study identified 100 potential biomarkers in rat serum, of which 52 were associated with lipid synthesis, such as LPC, PC, PI, PE, PA, Cer, SM, etc. The pathways involved were glycerol phospholipid, sphingomyelin, and glycerol ester metabolisms. After the Liujunzi decoction intervention, 56 potential biomarkers were observed in the high-dose group, alleviating the metabolic spectrum imbalance by reducing metabolite levels. In addition, metabolic pathway disturbances were markedly improved. This study provides references for future studies on Liujunzi decoction and furnishes essential data for assessing the relationships between chemical constituents and pharmacological activities of Liujunzi decoction.

Keywords: Lipid metabolism; Liujunzi decoction; Spleen deficiency hyperlipidemia; UPLC-Q-TOF-MS.

Fig. 1

Weight changes during modeling for each group of rats.

Fig. 2

Comparison of the morphology of stomach, small intestine, and liver (100x HE staining). Note: A1: Gastric tissue of blank control group, complete gastric mucosa with clear layers. B1: Small intestinal tissue of the blank control group, the structure of each layer of the small intestinal wall was intact. C1: Liver tissue of the blank control group, hepatocytes had normal morphology. A2: Gastric tissue of the hyperlipidemia group, with intact gastric mucosa and clear layers. B2: Small intestinal tissue of the hyperlipidemia group, the structure of each layer of the small intestinal wall was intact. C2: Liver tissue of the hyperlipidemia group, mild steatosis and fat vacuoles. A3: Gastric tissue of the spleen deficiency hyperlipidemia group, the gastric mucosa inflammatory cells infiltrated. B3: Small intestinal tissue of spleen deficiency hyperlipidemia group, a large number of inflammatory cell infiltration, intestinal mucosa propria edema. C3: Liver tissue of spleen deficiency hyperlipidemia group, steatosis, fat vacuoles, and a small amount of inflammatory cell infiltration. Red arrows indicate inflammatory cell infiltration. Red squares indicate the presence of fat vacuoles. (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 3

The PCA score plot derived from UPLC-Q-TOF/MS profiles of serum sample between the two groups Note: A: model group (M, green color)-blank (K, yellow color), B: low-dose group (G1, red color)-model group (M, green color), C: medium-dose group (G2, blue color)-model group (M, green color), D: high-dose group (G3, purple color)-model group (M, green color). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 4

Score plot and permutation test results of OPLS-DA model Note: A1: Score plot of model group (M, green color)-blank (K, yellow color), A2: Permutation test of model group (M, green color)-blank (K, yellow color); B1: Score plot of low-dose group (G1, red color)-model group (M, green color), B2: Permutation test of low-dose group (G1, red color)-model group (M, green color); C1: Score plot of medium-dose group (G2, blue color)-model group (M, green color), C2: Permutation test of medium-dose group (G2, blue color)-model group (M, green color); D1: Score plot of high-dose group (G3, purple color)-model group (M, green color), D2: Permutation test of high-dose group (G3, purple color)-model group (M, green color). (For interpretation of the references to color in this figure legend, the reader is referred to the Web version of this article.)

Fig. 5

Cluster analysis of serum metabolites(A: ESI+, B: ESI-).

Fig. 6

The difference in Metabolite Screening for high-dose group versus M Group lipid bubble diagram, and cluster analysis heat map. Note: A1: ESI+, Bubble diagram of high-dose group (G3)-model group (M), A2: ESI+, Cluster analysis heat map of high-dose group (G3)-model group (M); ESI-, Bubble diagram of high-dose group (G3)-model group (M), A2: ESI-, Cluster analysis heat map of high-dose group (G3)-model group (M).