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. 2020 Dec 17:8:616771.
doi: 10.3389/fcell.2020.616771. eCollection 2020.

Bisphenol B Exposure Disrupts Mouse Oocyte Meiotic Maturation in vitro Through Affecting Spindle Assembly and Chromosome Alignment

Affiliations

Bisphenol B Exposure Disrupts Mouse Oocyte Meiotic Maturation in vitro Through Affecting Spindle Assembly and Chromosome Alignment

Shou-Xin Zhang et al. Front Cell Dev Biol. .

Abstract

Bisphenol B (BPB), a substitute of bisphenol A (BPA), is widely used in the polycarbonate plastic and resins production. However, BPB proved to be not a safe alternative to BPA, and as an endocrine disruptor, it can harm the health of humans and animals. In the present study, we explored the effects of BPB on mouse oocyte meiotic maturation in vitro. We found that 150 μM of BPB significantly compromised the first polar body extrusion (PBE) and disrupted the cell cycle progression with meiotic arrest. The spindle assembly and chromosome alignment were disordered after BPB exposure, which was further demonstrated by the aberrant localization of p-MAPK. Also, BPB exposure increased the acetylation levels of α-tubulin. As a result, the spindle assemble checkpoint (SAC) was continuously provoked, contributing to meiotic arrest. We further demonstrated that BPB severely induced DNA damage, but the ROS and ATP production were not altered. Furthermore, the epigenetic modifications were changed after BPB exposure, as indicated by increased K3K9me3 and H3K27me3 levels. Besides, the pattern of estrogen receptor α (ERα) dynamics was disrupted with a mass gathering on the spindle in BPB-exposed oocytes. Our collective results indicated that exposure to BPB compromised meiotic maturation and damaged oocyte quality by affecting spindle assembly and chromosome alignment, acetylation of α-tubulin, DNA damage, epigenetic modifications, and ERα dynamics in mouse oocytes.

Keywords: DNA damage; bisphenol B; chromosome alignment; epigenetic modifications; spindle assembly.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

FIGURE 1
FIGURE 1
BPB exposure compromised the meiotic maturation of mouse oocytes. (A) Images depicting oocyte maturation with BPB (0, 50, 100, 150, 200 μM) exposure for 14 h. Bar,100 μm; Bar, 100 μm (small graph). (B) Rate of oocytes that extruded the first PB in control and BPB-treated groups. Control, n = 117; BPB, n = 88. Significantly different (P < 0.05); ∗∗∗Significantly different (P < 0.001) compared with the corresponding control.
FIGURE 2
FIGURE 2
BPB exposure disturbed cell cycle procession of meiotic maturation in mouse oocytes. (A) Spindle and chromosome depiction during oocyte developmental stages (GV, GVBD and Pro-MI, MI, A/TI, and MII). α-tubulin, green; DNA, blue. Bar, 20 μm. (B) The cell cycle distribution was quantified in the control and BPB groups. Control, n = 90; BPB, n = 90. Significantly different (P < 0.05); ∗∗Significantly different (P < 0.01); ∗∗∗Significantly different (P < 0.001).
FIGURE 3
FIGURE 3
BPB exposure disturbed the MI spindle assembly and chromosome alignment. Only the oocytes that reach to GVBD at 2 h in the control and BPB treated group were cultured for 8 h for subsequent analysis. (A) Spindle morphology and chromosome alignment depiction in control and BPB-treated oocytes. α-tubulin, green; DNA, blue. Bar, 20 μm. (B) Aberrant spindle morphology rate following BPB exposure. Control, n = 92; BPB, n = 91. ∗∗Significantly different (P < 0.01). (C) Aberrant chromosomal alignment rate following BPB exposure. Control, n = 92; BPB, n = 91. ∗∗Significantly different (P < 0.01). (D) Depiction of p-MAPK position in control and BPB-exposed oocytes. p-MAPK, red; α-tubulin, green; DNA, blue. Bar, 20 μm.
FIGURE 4
FIGURE 4
BPB exposure increased the acetylation level of α-tubulin. (A) Acetylated α-tubulin depiction in control and BPB-treated oocytes. Acetylated α-tubulin, red; DNA, blue. Bar, 20 μm. (B) Quantification of fluorescence intensity for acetylated α-tubulin in control, and BPB-exposed oocytes. Control, n = 34; BPB, n = 29. ∗∗∗Significantly different (P < 0.0001).
FIGURE 5
FIGURE 5
BPB exposure continuously activated the spindle assembly checkpoint. BubR1 depiction in control and BPB-exposed oocytes at ATI stage. BubR1, green. DNA, red. Bar, 20 μm.
FIGURE 6
FIGURE 6
BPB exposure increased DNA damage in mouse oocytes. (A) Images depicting DNA damage in control and BPB-treated oocytes. γ.H2A.X, red; DNA, blue. Bar, 20 μm. (B) Quantitative analysis of the fluorescence intensity of γ.H2A.X in control and BPB-treated oocytes. Control, n = 33; BPB, n = 32. ∗∗Significantly different (P < 0.01). (C) ROS depiction in control and BPB-exposed oocytes. ROS, red. Bar, 100 μm. (D) ROS fluorescence intensity in control and BPB-exposed oocytes. Control, n = 37; BPB, n = 36. No significant difference (P > 0.05). (E) The ATP content in control and BPB-treated oocytes. Control, n = 30; BPB, n = 30. No Significantly (P > 0.05).
FIGURE 7
FIGURE 7
BPB exposure altered epigenetic modification in mouse oocytes. (A) Images depicting H3K27me3 in control and BPB-treated oocytes. H3K27me3, red. Bar, 10 μm. (B) H3K27me3 fluorescence intensity in control and BPB-exposed oocytes. Control, n = 29; BPB, n = 31. Significantly different (P < 0.05). (C) Images depicting H3K9me3 in control and BPB-treated oocytes. H3K9me3, red. Bar, 10 μm. (D) H3Kme3 fluorescence intensity in control and BPB-exposed oocytes. Control, n = 30; BPB, n = 29. ∗∗∗Significantly different (P < 0.0001).
FIGURE 8
FIGURE 8
BPB exposure changed the distribution pattern of ERα. (A) ERα depiction in control and BPB-exposed. ERα, red; α-tubulin, green; DNA, blue. Bar, 20 μm. (B) ERα fluorescence intensity on the spindles in control and BPB-exposed oocytes. Control, n = 34; BPB, n = 33. ∗∗Significantly different (P < 0.01).

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