GC-TOF/MS-based metabolomic profiling of estrogen deficiency-induced obesity in ovariectomized rats

Abstract

Aim: To explore the alteration of endogenous metabolites and identify potential biomarkers using metabolomic profiling with gas chromatography coupled a time-of-flight mass analyzer (GC/TOF-MS) in a rat model of estrogen-deficiency-induced obesity.

Methods: Twelve female Sprague-Dawley rats six month of age were either sham-operated or ovariectomized (OVX). Rat blood was collected, and serum was analyzed for biomarkers using standard colorimetric methods with commercial assay kits and a metabolomic approach with GC/TOF-MS. The data were analyzed using multivariate statistical techniques.

Results: A high body weight and body mass index inversely correlated with serum estradiol (E2) in the OVX rats compared to the sham rats. Estrogen deficiency also significantly increased serum total cholesterol, triglycerides, and low-density lipoprotein cholesterol. Utilizing GC/TOF-MS-based metabolomic analysis and the partial least-squares discriminant analysis, the OVX samples were discriminated from the shams. Elevated levels of cholesterol, glycerol, glucose, arachidonic acid, glutamic acid, glycine, and cystine and reduced alanine levels were observed. Serum glucose metabolism, energy metabolism, lipid metabolism, and amino acid metabolism were involved in estrogen-deficiency-induced obesity in OVX rats.

Conclusion: The series of potential biomarkers identified in the present study provided fingerprints of rat metabolomic changes during obesity and an overview of multiple metabolic pathways during the progression of obesity involving glucose metabolism, lipid metabolism, and amino acid metabolism.

Figure 1

Body weight, body mass index, and uterus index from OVX and the sham rats. (A) Body weight of the rats was recorded weekly during the experiment. (B) Body mass index observed in the pre-surgery and weeks 3 and 6 post-surgery groups. (C) Uterus index. Values with a superscript are significantly different from the sham group (^bP<0.05, ^cP<0.01).

Figure 2

GC-TOF/MS chromatograph of a serum sample obtained from a sham-operated rat. 1, Alanine; 2, Lactic acid; 3, β-Hydroxybutyric acid; 4, Valine; 5, Leucine; 6, Glycerol; 7, Phosphoric acid; 8, Glycine; 9, Isoleucine; 10, Cysteine; 11, Glyceraldehyde 3-phosphate; 12, Creatinine; 13, Glutamic acid; 14, Glutamine; 15, Glucose; 16, Hexadecanoic acid; 17, Uric acid; 18, Oleic acid; 19, Octadecadienoic acid; 20, Octadecanoic acid; 21, Arachidonic acid; 22, Cholesterol.

Figure 3

PLS-DA score plots of serum samples. (A) PLS-DA score plots of OVX rats over the whole experiment (pre-surgery, weeks 3 and 6 post-surgery). (B) PLS-DA score plots of OVX rats and sham rats at 6 weeks post-surgery. The x-axis and y-axis are labeled as PC1 (the first principal component) and PC2 (the second principal component), respectively. One data point stands for one subject: ▪, OVX rats prior to surgery; □, OVX rats at week 6 post-surgery; •, sham rats at week 6 post-surgery; ▴, OVX rats at week 3 post-surgery.

Figure 4

Regulation of glycometabolism, lipid metabolism, and amino acid metabolism by estrogen deficiency (significant difference from the endogenous metabolite is indicated in bold).