Comparative transcriptome analysis of the protective effects of Korean Red Ginseng against the influence of bisphenol A in the liver and uterus of ovariectomized mice

Abstract

Background: Bisphenol A (BPA), known as an endocrine disruptor, is widely used in the world. BPA is reported to cause inflammation-related diseases. Korean Red Ginseng (KRG) has been used safely in human for a long time for the treatment of diverse diseases. KRG has been reported of its mitigating effect on menopausal symptoms and suppress adipose inflammation. Here, we investigate the protective effect of orally administered KRG on the impacts of BPA in the liver and uterus of menopausal mice model.

Methods: The transcriptome analysis for the effects of BPA on mice liver was evaluated by Gene Expression Omnibus (GEO) database-based data (GSE26728). In vivo assay to evaluate the protective effect of KRG on BPA impact in ovariectomized (OVX) mice were designed and analyzed by RNA sequencing.

Results: We first demonstrated that BPA induced 12 kinds of gene set in the liver of normal mice. The administration of BPA and KRG did not change body, liver, and uterine weight in OVX mice. KRG downregulated BPA-induced inflammatory response and chemotaxis-related gene expression. Several gene set enrichment analysis (GSEA)-derived inflammatory response genes increased by BPA were inhibited by KRG in OVX mice.

Conclusion: Our data suggest that BPA has commonly influenced inflammatory response effects on both normal and OVX mice. KRG protects against BPA impact of inflammatory response and chemotaxis in OVX mouse models. Our comparative analysis will provide new insight into the efficacy of KRG on endocrine disrupting chemicals and OVX mouse.

Keywords: Bisphenol A; Korean Red Ginseng; Transcriptome analysis.

Fig. 1

Gene set enrichment plot of genes regulated by BPA. GSE26728 includes microarray data of CD-1 mice at 9 weeks of age treated with low and high doses of BPA (50 μg/kg/day and 5,000 μg/kg/day, respectively) for 28 days in their feed. The results of GSEA of DEGs between negative control and BPA treatment groups were visualized using the JavaGSEA package. The most significantly enriched signaling pathways were selected based on the normalized enrichment score (NES), with a nominal P-value < 0.05, and false discovery rate q-value < 0.25. The y axis represents the enrichment score, and the x axis lists genes that showed high levels of expression induced by BPA treatment among the 22,514 total genes. The black bar indicates the locations of the genes in each gene set. BPA, bisphenol A; DEGs, differentially expressed genes.

Fig. 2

KRG with or without BPA did not alter the liver or uterus weights. Treatment with BPA and KRG was started 1 week after ovariectomy. Ovariectomized CD-1 mice were treated with BPA (200 mg/kg/day) diluted in corn oil and KRG (1.2 g/kg/day) diluted in triple distilled water every day for 7 days via oral administration. Body weight (A), uterine weight (B), and liver weight (C) were recorded. BPA, bisphenol A; KRG, Korean Red Ginseng.

Fig. 3

Differential gene expression induced by BPA treatment with or without KRG in mouse liver and uterus. The levels of mRNA transcripts in mouse liver and uterus were determined by the library preparation and sequencing method. Venn diagrams show the classification of DEGs in the BPA group and BPA plus KRG group in the mouse liver (A) and uterus (B). Numbers represent the numbers of genes showing a more than twofold change in expression compared with negative controls. Upper numbers, upregulated genes; lower numbers, downregulated genes; middle numbers, contraregulated genes. Under the same conditions, genes with different expression levels between the BPA plus KRG group and the BPA-only group in the mouse liver (C) and uterus (D) were selected and charted by the percentage of genes in each gene ontology out of the total number of genes. The numbers of genes upregulated or downregulated in the liver (E) and uterus (F) in the BPA plus KRG group are shown in red and green bar graphs, respectively. BPA, bisphenol A; KRG, Korean Red Ginseng; DEGs, differentially expressed genes.

Fig. 4

Induction of inflammatory response and chemotaxis-related genes by BPA was reduced by KRG. Functional annotation of genes regulated by BPA plus KRG compared with those regulated by BPA alone. Gene Ontology analysis of regulated DEGs in the liver and uterus (A). Genes showing equal expression are marked with red boxes. Heat map plots of DEGs related to the inflammatory response and chemotaxis. The genes upregulated by BPA were identified and evaluated in the BPA plus KRG group and visualized as heat maps (B). Total RNA of the liver and uterus was extracted, and the expression level of the VCAM-1 and CCR7 mRNA was analyzed by real-time PCR (C). TNF-α protein levels in the liver and uterus were analyzed by ELISA (D). ∗P < 0.05, ∗∗P < 0.01. BPA vs OVX-control; #P < 0.05, ##P < 0.01 BPA vs BPA with KRG. BPA, bisphenol A; KRG, Korean Red Ginseng; OVX, ovariectomized; PCR, polymerase chain reaction; ELISA, enzyme-linked immunosorbent assay; DEGs, differentially expressed genes.

Fig. 5

Induction of inflammatory response–related genes by BPA was reduced by KRG. Heat map of enriched hallmark inflammatory response gene set from GSE26728 (A). The effects of KRG and BPA on the expression of inflammatory response–related genes. The gene set enriched in GSE26728 was collected and applied to the liver and uterus of the BPA group and BPA plus KRG group, and the results are visualized as heat maps (B). BPA, bisphenol A; KRG, Korean Red Ginseng.