Cuscutae Semen in depression-induced ovarian dysfunction: metabolomics with UPLC-QToF-MS in female mice

Abstract

The increasing prevalence of depression profoundly affects female ovarian health. Although Cuscutae Semen (CS) is acknowledged for treating reproductive disorders, its pharmacological mechanisms in depression-induced ovarian dysfunction remain insufficiently explored. This study investigated CS's effects in a chronic unpredictable mild stress (CUMS) mouse model of depression. Mice were divided into control, CUMS model, CS treatment and estradiol treatment group. Behavioral and biochemical analyses assessed depressive-like behaviors and hormone levels. Untargeted metabolomics utilizing ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry was applied to identify differential metabolites of CS in the treatment of depression-induced ovarian dysfunction. These findings were confirmed through real-time quantitative polymerase chain reaction assays. Based on the outcomes from behavioral and biochemical assays, CS effectively ameliorated the chronic unpredictable mild stress-induced reproductive ailment in mice. Ten differential metabolites were identified, highlighting the impact of CUMS and CS's ameliorative effects. Pathways linked to arachidonic acid metabolism, glycerophospholipid metabolism, linoleic acid metabolism, and steroid hormone biosynthesis were involved. Seven target genes further validated the metabolomic analysis. This study provides strong evidence of CS's therapeutic potential in alleviating depression-induced ovarian dysfunction, shedding light on its pharmacological mechanisms and supporting its use as a functional medical food.

Keywords: Cuscutae Semen; depression; metabolomics; ovarian dysfunction; ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry.

FIGURE 1

Evaluation of sucrose preference test (SPT), tail suspension test (TST) and focused swimming test (FST) for measuring depressive-like behavior. “CON”, “CUMS”, “CUMS+CS” and “CUMS+E” represented control group, chronic unpredictable mild stress group, Cuscutae Semen treatment group and estradiol treatment group, respectively. Mann-Whitney U test was used to calculate significant difference. ^## P < 0.01, compared with CON group. ^** P < 0.01, compared with CUMS group. Each group contained six samples, maintaining consistency across all groups.

FIGURE 2

Ovarian pathological examinations (A) and enumeration of follicles at diverse development stages (B) in female mice.“CON”, “CUMS”, “CUMS+CS” and “CUMS+E” represented control group, chronic unpredictable mild stress group, Cuscutae Semen treatment group and estradiol treatment group, respectively. Mann-Whitney U test was used to calculate significant difference. ^# P < 0.01, compared with CON group. Each group contained six samples, maintaining consistency across all groups.

FIGURE 3

Assessment of follicle-stimulating hormone (FSH), luteinizing hormone (LH), estradiol (E2) and anti-mullerian hormone (AMH) indicators in female mice. “CON”, “CUMS”, “CUMS+CS” and “CUMS+E” represented control group, chronic unpredictable mild stress group, Cuscutae Semen treatment group and estradiol treatment group, respectively. Mann-Whitney U test was used to calculate significant difference. ^## P < 0.01, compared with CON group. ^** P < 0.01, compared with CUMS group. Each group contained six samples, maintaining consistency across all groups.

FIGURE 4

(A) Principal component analysis (PCA) of peak areas detected in quality control (QC), control (CON), chronic unpredictable mild stress (CUMS) and Cuscutae Semen treatment (CUMS+CS) group. (B) Orthogonal partial least squares discrimination analysis (OPLS-DA) score scatter plot in comprising of CON-CUMS. (C) Results of 999 permutations for the OPLS-DA model.

FIGURE 5

The relative contents of 10 discriminant metabolites in the control (CON), chronic unpredictable mild stress (CUMS), and Cuscutae Semen treatment (CUMS+CS) group. The data on the relative contents of differential metabolites were subjected to logarithmic transformation with a base of 2. Mann-Whitney U test was used to calculate significant difference. ^# P < 0.05, ^## P < 0.01, compared with CON group. ^* P < 0.05, compared with CUMS group.

FIGURE 6

Metabolic pathway of mainly differential metabolites under effect of chronic unpredictable mild stress (CUMS) and Cuscutae Semen (CS). “CON” and “CUMS+CS” represented control group and CS treatment group, respectively. The color bar from blue to red represented low to high intensity after logarithmic transformation with a base of 10. Each group contained six samples, maintaining consistency across all groups.

FIGURE 7

Effect of Cuscutae Semen (CS) on the expression of targets in the treatment of chronic unpredictable mild stress (CUMS) detected by real-time quantitative polymerase chain reaction (n = 6). “CON” and “CUMS+CS” represented control group and CS treatment group, respectively. Mann-Whitney U test was used to calculate significant difference. ^# P < 0.05, ^## P < 0.01, compared with CON group. ^* P < 0.05, ^** P < 0.01, compared with CUMS group. Each group contained six samples, maintaining consistency across all groups.