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. 2022 Oct 1;323(4):R571-R580.
doi: 10.1152/ajpregu.00068.2022. Epub 2022 Aug 15.

Effect of muscone on anti-apoptotic ability of muskrat prostate primary cells

Yu Zhang  1 , Xuefei Zhao  1   2 , Xin Man  1 , Yue Ma  1   3 , Wei Zhang  1   3 , Yu Wang  1 , Xing Lei  1 , Suying Bai  1   3
Affiliations

Effect of muscone on anti-apoptotic ability of muskrat prostate primary cells

Yu Zhang et al. Am J Physiol Regul Integr Comp Physiol. .

Abstract

Muskrat is a small fur animal with a pair of scent glands that can secrete muskrat musk during breeding season. The consensus is muskrat musk functions as a pheromone, but we hypothesized it has a broader role. In previous research, we found the presence of muscone in muskrat musk. To study whether the muscone can affect the apoptosis of muskrat prostate, we carried out the following investigations. Primary muskrat prostate cells were cultured and treated with muscone. Then we drew cell proliferation curves by applying the CCK-8 and used TdT-mediated dUTP nick end labeling (TUNEL) to detect apoptosis. Levels of mRNA transcription and protein expression of Bcl-2 as well as Bax were detected by qRT-PCR and the Western blot. Meanwhile, we collected tissue samples of muskrat prostates and froze sections to analyze the fluorescence signal intensity of BCL-2 and BAX via immunofluorescence. Under the treatment of 30 μmol/L muscone, the proliferation rate of the experimental group exceeded that of the control group, and the proportion of cells undergoing apoptosis was lower in the experimental group. The qRT-PCR and Western blot result showed that, in the experimental group, the ratio of Bcl-2 to Bax mRNA transcription levels increased by 2.85 times and their corresponding protein expression ratio increased by 2.37 times (P < 0.05). Immunofluorescence results were consistent with the cell experiment's results. The fluorescence signal intensity of BCL-2 was higher in the breeding season than nonbreeding season but vice versa for BAX. Based on these results, we speculate that the muscone could regulates prostate development by inhibiting apoptosis.

Keywords: apoptosis; muscone; muskrat (Ondatra zibethicus); primary culture; prostate.

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Conflict of interest statement

No conflicts of interest, financial or otherwise, are declared by the authors.

Figures

Figure 1.
Figure 1.
Primary cultured muskrat prostate cells. Scale bars of A–C are 50, 20, and 10 μm, respectively.
Figure 2.
Figure 2.
A: cell proliferation curve of the EG group and CG group. B: cell survival rate of the EG group vs. CG group. Different uppercase letters indicate significant differences within EG group, and different lowercase letters indicate significant differences within CG group. *Significant difference between EG group and CG group. CG, control group; EG, experimental group.
Figure 3.
Figure 3.
A: TdT-mediated dUTP nick end labeling (TUNEL) result for the CG group. B: TUNEL result for the EG group. C: positive control. Arrow pattern, positive cells in the state of apoptosis (scale bars, 40 μm). CG, control group; EG, experimental group.
Figure 4.
Figure 4.
A: relative expression levels of Bcl-2 and Bax mRNA in the EG group vs. CG group. B: relative expression of BCL-2 and BAX proteins in the EG group vs. CG group. *Significant difference between EG group and CG group. CG, control group; EG, experimental group.
Figure 5.
Figure 5.
Western blotting results for BCL-2, BAX, and β-ACTIN proteins in the EG group and CG group. CG, control group; EG, experimental group.
Figure 6.
Figure 6.
Immunofluorescence staining of BAX protein in the muskrat prostate of breeding season (A–C) and nonbreeding season (D–F). Immunofluorescence staining of BCL-2 protein in the muskrat prostate of breeding season (G–I) and nonbreeding season (J–L; scale bars, 100 μm).
Figure 7.
Figure 7.
Sketch to show seasonal development of the prostate regulated by muscone.
See this image and copyright information in PMC

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