PDCD4对奶山羊乳腺上皮细胞的凋亡及β-酪蛋白和TG合成的影响

doi:10.11843/j.issn.0366-6964.2023.04.008

Abstract

Abstract: The aim of this study was to investigate the regulatory effects of programmed cell death 4 (PDCD4) on the apoptosis of mammary epithelial cells (GMECs) and milk composition in goat. In this study, the dual luciferase reporter assay was used to verify the targeting relationship between miR-21-5p and PDCD4, and the regulatory effects of miR-21-5p on PDCD4 was detected by RT-qPCR and Western blot. The PDCD4 small interfering RNA (si-PDCD4) and PDCD4 overexpression vector (pc3.1-PDCD4) were transfected into GMECs to explore the effect of PDCD4. EdU and CCK-8 tests were used to detect the effect of PDCD4 on the proliferation of GMECs, flow cytometry was chosen to detect its effect on apoptosis of GMECs. After transfection, the ELISA kit was used to detect the secretion of β-casein and triglyceride (TG) in GMECs. Finally, the Western blot tests was used to detect the protein expression of PI3K, mTOR, S6K, ERK, Bcl-2, Bax in GMECs. The results of the dual luciferase reporter assay showed that PDCD4 was one of the target genes of miR-21-5p. The results of the EdU, CCK-8 and flow cytometry tests showed that the activity of GMECs was significantly decreased after overexpression of PDCD4 (P<0.01), indicated that PDCD4 inhibited the proliferation of GMECs and promoted the apoptosis of GMECs (P<0.01); the activity of GMECs was significantly increased after interfering PDCD4 (P<0.01), the pro-apoptotic level of GMECs was significantly reduced (P<0.01). The results of the ELISA kit test showed that the secretion of β-casein and TG in GMECs was significantly decreased after overexpression of PDCD4 (P<0.01), and they were significantly increased after interfering PDCD4 (P<0.01). The results of the Western blot test showed that, compared with the control group, the protein expressions of phosphorylated PI3K, mTOR, S6K and ERK were significantly decreased after overexpression PDCD4 (P<0.05), the protein expression of Bax was significantly up-regulated (P<0.01), and the protein expression of Bcl-2 was significantly down-regulated (P<0.01). After interfering PDCD4, the protein expression of phosphorylated PI3K, mTOR, S6K and ERK were significantly increased (P<0.05), the protein expression of Bax was significantly decreased (P<0.05), and the expression of Bcl-2 was significantly up-regulated (P<0.05). This study indicates that PDCD4 promotes the apoptosis of dairy goat mammary epithelial cells by activating the ERK/Bcl-2/Bax signaling pathway, and decreases the secretion of β-casein and TG in dairy goat mammary epithelial cells by inhibiting PI3K/mTOR/S6K signaling pathway. And miR-21-5p targets PDCD4 and can regulate its expression in GMECs.

Key words: PDCD4, miR-21-5p, milk goat, apoptosis, beta-casein, TG

CLC Number:

S827.2

ZHU Junru, WEI Wei, PEI Dangshuai, ZHANG Fenque, DUAN Yu, GUO Yongfeng, JIANG Yue, XIA Shuli, HAN Jing, HOU Jinxing, AN Xiaopeng. Effects of PDCD4 on the Apoptosis of Dairy Goat Mammary Epithelial Cells and the Synthesis of β-casein and TG[J]. Acta Veterinaria et Zootechnica Sinica, 2023, 54(4): 1429-1440.

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Effects of PDCD4 on the Apoptosis of Dairy Goat Mammary Epithelial Cells and the Synthesis of β-casein and TG

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