FKBP5基因对绵羊卵泡颗粒细胞功能的影响

doi:10.11843/j.issn.0366-6964.2024.09.019

畜牧兽医学报 ›› 2024, Vol. 55 ›› Issue (9): 3947-3956.doi: 10.11843/j.issn.0366-6964.2024.09.019

FKBP5基因对绵羊卵泡颗粒细胞功能的影响

古丽米热·阿布都热依木¹^,²^,³(), 张欣如¹^,²^,³, 吴阳升¹^,²^,³, 陈莹¹^,²^,³, 汪立芹¹^,²^,³, 徐新明¹^,²^,³, 黄俊成¹^,²^,³^,*(), 林嘉鹏¹^,²^,³^,*()

1. 农业农村部草食家畜遗传育种与繁殖重点实验室，乌鲁木齐 830011
2. 新疆动物生物技术重点实验室，乌鲁木齐 830011
3. 新疆畜牧科学院生物技术研究所，乌鲁木齐 830011

收稿日期:2024-02-27 出版日期:2024-09-23 发布日期:2024-09-27
通讯作者: 黄俊成,林嘉鹏 E-mail:gulimire127@163.com;h_jc@sina.com;linjiapeng5188@163.com
作者简介:古丽米热·阿布都热依木(1985-), 女，新疆乌鲁木齐人，硕士，主要从事动物遗传育种与繁殖研究，E-mail：gulimire127@163.com
基金资助:
新疆维吾尔自治区重点实验室开放课题(2020D04019);自治区重大科技专项(2023A02011);国家自然科学基金(31860646);自治区科技支疆项目(2022E02017);自治区“三农”骨干人才培养项目(2022SNGGNT091)

Effects of FKBP5 on Function of Sheep Follicular Granulosa Cells

古丽米热·阿布都热依木¹^,²^,³(), Xinru ZHANG¹^,²^,³, Yangsheng WU¹^,²^,³, Ying CHEN¹^,²^,³, Liqin WANG¹^,²^,³, Xinming XU¹^,²^,³, Juncheng HUANG¹^,²^,³^,*(), Jiapeng LIN¹^,²^,³^,*()

1. Key Laboratory of Genetics Breeding and Reproduction of Grass Feeding Livestock of Ministry of Agriculture and Rural Affairs, Urumqi 830011, China
2. Key Laboratory of Animal Biotechnology of Xinjiang, Urumqi 830011, China
3. Institute of Biotechnology, Xinjiang Academy of Animal Sciences, Urumqi 830011, China

Received:2024-02-27 Online:2024-09-23 Published:2024-09-27
Contact: Juncheng HUANG, Jiapeng LIN E-mail:gulimire127@163.com;h_jc@sina.com;linjiapeng5188@163.com

摘要/Abstract

摘要：

旨在探讨FKBP5在绵羊卵巢颗粒细胞(GCs)中的潜在功能。本研究从乌鲁木齐市华凌屠宰场的20个2岁左右处于性成熟阶段的阿勒泰羊卵巢中分离GCs。将试验分为4组：GC(空白对照)、GC+NC(转染siRNA-NC)、GC+ siFKBP5(转染siRNA-FKBP5)、GC+ siFKBP5+LH(促黄体生成素处理转染siRNA-FKBP5)。利用细胞免疫荧光检测FKBP5蛋白在GCs中的定位。通过EdU、Tunel和Western blot检测其对细胞增殖、凋亡的影响；通过Western blot检测其对AKT和ERK信号通路的影响；通过ELISA检测其对E2、P4水平的影响。结果表明，FKBP5表达于GCs细胞质中。FKBP5表达量随着LH浓度不断升高；5 IU·mL^-1 LH作用4 h，FKBP5表达较高(P < 0.001)。干扰FKBP5后显著抑制了细胞增殖，并显著降低PCNA蛋白表达量(P < 0.001)；显著促进了细胞凋亡，增加了BAX的表达并减少了Bcl-2的表达(P < 0.001)；显著抑制了GCs中AKT和ERK信号通路的激活以及E2和P4的分泌(P < 0.001)。LH处理部分改善了干扰FKBP5对GCs的影响(P < 0.05)。本研究结果表明，FKBP5影响绵羊GCs的增殖、凋亡、AKT和ERK信号通路以及E2和P4的分泌，其功能受LH的影响。这些结果为进一步研究FKBP5在绵羊卵泡发育中的作用提供了理论依据。

关键词: FKBP5, 颗粒细胞, 细胞增殖凋亡, AKT和ERK信号通路, 类固醇

Abstract:

This study aimed to investigate the potential function of FKBP5 in ovine ovarian granulosa cells (GCs). GCs were isolated from the ovaries of twenty approximately 2-year-old sexually mature Altay sheep obtained from the Hualing slaughterhouse in Urumqi. The experiment was divided into 4 groups: GC (blank control), GC+NC (transfected with siRNA-NC), GC+siFKBP5 (transfected with siRNA-FKBP5), and GC+siFKBP5+LH (treated with luteinizing hormone and transfected with siRNA-FKBP5). Immunofluorescence was used to detect the localization of FKBP5 protein in GCs. Its effects on cell proliferation and apoptosis were examined using EdU, Tunel assays, and Western Blot analysis. Its impacts on the AKT and ERK signaling pathways were assessed by Western blot, while the levels of E2 and P4 were measured by ELISA. FKBP5 was found to be expressed in the cytoplasm of GCs. Its expression increased with the increasing concentration of LH, reaching a peak at 5 IU·mL^-1 LH after 4 h of treatment (P < 0.001). Knockdown of FKBP5 significantly inhibited cell proliferation, as evidenced by a reduction in PCNA protein expression (P < 0.001). It also significantly promoted cell apoptosis, with an increase in BAX expression and a decrease in Bcl-2 expression (P < 0.001). Additionally, FKBP5 knockdown markedly suppressed the activation of the AKT and ERK signaling pathways and the secretion of E2 and P4 (P < 0.001). LH treatment partially mitigated the effects of FKBP5 knockdown on GCs (P < 0.05). The results indicate that FKBP5 influences the proliferation and apoptosis of ovine GCs, as well as the activation of the AKT and ERK signaling pathways and the secretion of E2 and P4. Its function is modulated by LH. These findings provide a theoretical basis for further research on the role of FKBP5 in ovine follicular development.

Key words: FKBP5, granulosa cells, cell proliferation and apoptosis, AKT and ERK signaling pathways, steroids

中图分类号:

S826.2

古丽米热·阿布都热依木, 张欣如, 吴阳升, 陈莹, 汪立芹, 徐新明, 黄俊成, 林嘉鹏. FKBP5基因对绵羊卵泡颗粒细胞功能的影响[J]. 畜牧兽医学报, 2024, 55(9): 3947-3956.

古丽米热·阿布都热依木, Xinru ZHANG, Yangsheng WU, Ying CHEN, Liqin WANG, Xinming XU, Juncheng HUANG, Jiapeng LIN. Effects of FKBP5 on Function of Sheep Follicular Granulosa Cells[J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(9): 3947-3956.

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si-RNA名称 si-RNA name	引物序列(5′→3′) Primers sequence
si-NC	S：UUCUCCGAACGUGUCACGUTT AS：CGUGACACGUUCGGAGAATT
si-416	S：GUCCAUUACAGAGGGAAAUTT AS：AUUUCCCUCUGUAAUGGACTT
si-1023	S：GGGAGAUGGAUACCAAAGATT AS：UCUUUGGUAUCCAUCUCCCTT
si-1523	S：GAGGCCAGUAAAGCAAUGATT AS：UCAUUGCUUUACUGGCCUCTT

基因名称 Gene name	引物序列(5′→3′) Primer sequence	产物大小/bp Product size	退火温度/℃ Temperature
FKBP5	F: CAGTCTCCCCAAAATTCCCTC R: ATCAAACATCCTTCCACCACA	196	58
β-actin	F: TGACCCAGATCATGTTTGAGA R: CAAGGTCCAGACGCAGGAT	186	58

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FKBP5基因对绵羊卵泡颗粒细胞功能的影响

Effects of FKBP5 on Function of Sheep Follicular Granulosa Cells

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