A comprehensive transcriptomic analysis of the bisphenol A affected kidney in mice

Abstract

Introduction: Bisphenol A (BPA) is a substance belonging to the endocrine-disrupting chemicals, globally used in the production of polycarbonate plastics. It has been found that BPA enhances carcinogenesis, triggers obesity and exerts a pathogenic effect in several disorders, such as type 2 diabetes, asthma, or increased blood pressure. Recent studies have revealed, that BPA has a harmful impact on the kidneys function, therefore, the current research aimed to explore the specific molecular changes triggered in these organs after oral BPA exposure in mice. Materials and Methods: The experiment was carried out on 12 (3-month-old) female mice. Six mice served as controls. The other 6 mice were treated with BPA in the drinking water at a dose of 50 mg/kg b. w. for 3 months. Then animals were euthanized, the kidneys were collected, and extracted RNA was used to perform RNA-seq. Results: Applied multistep bioinformatics revealed 433 differentially expressed genes (DEGs) in the BPA-treated kidneys (232 upregulated and 201 downregulated). Additionally, 95 differentially expressed long-noncoding RNAs (DELs) were revealed in BPA samples. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) annotations indicated that BPA exposure resulted in profound changes in several essential processes, such as oxidative phosphorylation, mitochondrial and ribosome function, or chemical carcinogenesis. Conclusion: The obtained novel results suggest that BPA has a harmful impact on the fundamental processes of the kidney and significantly impairs its function by inducing mitochondrial dysfunction leading to oxidative stress and reactive oxygen species production.

Keywords: RNA-Seq; bisphenol A; kidney; mitochondrial dysfunction; oxidative stress.

FIGURE 1

Expression profiles overview of kidney differentially expressed transcriptionally active regions (TARs) under BPA influence. Volcano plot with binary logarithmic values of fold change (log2FC; X-axis) plotted against negative logarithmic adjusted p-values (-log10 (p-value); Y-axis). The dotted horizontal line indicates a negative logarithmic adjusted p-value cut-off (0.05).

FIGURE 2

Circular heatmap visualization of differentially expressed genes (DEGs) and long non-coding RNAs (DELs) in BPA-affected and CTR-control libraries. The 12 upper tracks visualize the normalized (Z-score) expressions for DEGs in each biological replicate. The large part of the circle (green-red) depicts DEGs and the smaller part (blue-yellow) describes DELs. The inner track shows the correlation links between the co-expressed DEGs and DELs, whereas blue links depict positive (>0.9) and yellow negative (<-0.9) Pearson’s correlation.

FIGURE 3

Volcano plot showing the percentage of splicing inclusions difference (ΔPSI) against the statistical significance (-log10FDR) of DASs identified within genes of murine BPA-affected kidneys vs. control samples. The dashed lines indicate the applied cut-off thresholds, described in the text.

FIGURE 4

Circular visualization of differentially alternative splicing events (DASs) occurring after BPA treatment. The five-scale color heatmaps (outer track) represent percentage inclusion values (PSI) in experimental (BPA) and control (CTR) samples. The middle track shows dPSI values (red–higher inclusion level in BPA, blue–higher inclusion level in CTR). Color links join common genes with more than one DAS classified in different types of alternative splicing events.

FIGURE 5

Sashimi plot visualizing the detected coverage of RNA-Seq reads on the reference genome and the average values of reads combining distant genome fragments (black blocks underneath the graphs) in CTR and BPA groups. Displayed fragments of (A) HSF1, (B) MCFD2 and (C) SON genes were classified as statistically significant differentiated alternative splicing events.

FIGURE 6

Gene Ontology (GO) enrichment dot-plot (A) of abundances (size of dots) and significance (color of dots) of the top GO terms. (B) Circos-plot relationship of differentially expressed genes (DEGs) engaged in kidney function under BPA influence significantly associated with four selected GO enriched terms. Gene symbols with logarithmic values (blue-red scale) of fold change (log2FC) are located on the left side of the circos. Color links merge genes with the GO terms (cytoplasmic translation, oxidative phosphorylation, electron transport chain and cellular respiration) on the right side.

FIGURE 7

Enrichment Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of differentially expressed genes (DEGs) engaged in “Oxidative phosphorylation” signaling pathway induced in the kidney after BPA exposure. Red rectangles represent upregulated genes. Logarithmic fold change (log2FC) red-green scale describes gene expression values.