维生素 E 通过抑制氧化应激缓解鹅卵泡颗粒细胞凋亡

doi:10.3969/j.issn.1674-7968.2023.08.010

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摘要氧化应激介导的鹅(Anser cygnoides)卵泡颗粒细胞(granulosa cells, GCs)凋亡影响鹅产蛋性能。本研究旨在探讨抗氧化剂维生素 E (vitamin E, VE)对鹅卵泡 GCs 凋亡的缓解作用。通过添加 H2O2 (100 μmol/L)建立鹅卵泡 GCs 氧化应激模型,并在体外检测氧化应激对鹅卵泡 GCs 凋亡的影响。在该模型中分别添加 4 种不同浓度的原花青素、白藜芦醇、没食子酸和VE,筛选出最佳缓解剂及最适浓度。进一步采用ELISA、qPCR 和免疫荧光检测等技术探讨最佳缓解剂 VE 对氧化应激介导的鹅卵泡 GCs 凋亡的作用。结果表明,100 μmol/L H2O2 处理 12 h 极显著促进鹅卵泡 GCs 凋亡(P<0.01) ;4 种不同抗氧化剂中,40 μmol/L VE 对氧化应激模型的缓解作用最佳 (P<0.01),添加后可减少细胞乳酸脱氢酶 (lactate dehydrogenase, LDH)的释放量(P<0.05),降低氧化应激相关基因超氧化物歧化酶 1 (superoxide dismutase 1, SOD-1)、SOD-2、过氧化氢酶 (catalase, CAT) 表达量、环氧化酶 2 (cyclooxygenase, COX-2)和活性氧 (reactive oxygen species, ROS)含量(P<0.05);细胞增殖活力显著上升(P<0.05)且呈剂量依赖性,抗凋亡基因B 细胞淋巴瘤/白血病-2 (B-cell lymphoma/leukemia-2, Bcl-2)的表达量显著上升(P<0.05),促凋亡基因半胱氨酸的天冬氨酸蛋白水解酶 3 (cysteinyl aspartate specific proteinase 3, Caspase3)、Caspase9 和肿瘤抑制基因肿瘤蛋白 p53 (tumor protein p53, p53)的表达量显著下降(P<0.05),细胞凋亡数显著降低(P<0.05)。本研究表明,在氧化应激诱导条件下,鹅卵泡 GCs 存在明显的凋亡,添加 VE 可抑制氧化应激缓解鹅卵泡GCs 凋亡,本研究可为提高鹅的产蛋性能提供参考。

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	皇甫奕雯
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	陈国宏
	徐琪
	张钰

关键词 ：维生素 E, 氧化应激, 凋亡, 鹅卵泡颗粒细胞

Abstract：Oxidative stress mediates apoptosis of goose (Anser cygnoides) follicular granulosa cells (GCs), thus affecting laying performance. To investigate the effects of vitamin E (VE) on goose follicular GCs, an oxidative stress model was established by adding H2O2 (100 μmol/L), and the effects of oxidative stress on apoptosis in goose follicular granulosa cells were examined in vitro. Four different concentrations of proanthocyanidins, resveratrol, gallic acid and VE were added to the model, and the best reliever and optimum concentration were screened out. Furthermore, ELISA, qPCR and immunofluorescence detection techniques were used to explore the effect of the best alleviator VE on the apoptosis of goose follicle GCs mediated by oxidative stress. The results showed that 100 μmol/L H2O2 treatment for 12 h extremely significantly increased the level of apoptosis in follicular GCs (P<0.01). Among the 4 different antioxidants, 40 μmol/L VE had the best alleviation effect on the oxidative stress model (P<0.01). The addition of VE significantly inhibited the release of lactate dehydrogenase (LDH) (P<0.05), and reduced the expression of oxidative stress-related genes superoxide dismutase 1 (SOD-1), SOD-2, catalase (CAT), cyclooxygenase 2 (COX-2) and reactive oxygen species (ROS) content (P<0.05). Besides, GCs proliferation viability increased significantly (P<0.05) and there was a phenomenon of dose dependence. The expression levels of anti-apoptotic genes B-cell lymphoma/leukemia-2 (Bcl2) were significantly increased (P<0.05), but pro-apoptotic genes cysteinyl aspartate specific proteinase 3 (Caspase3), Caspase9 and tumor protein p53 (p53) were significantly decreased (P<0.05), as well as the number of apoptotic cells were significantly reduced after VE treatment (P<0.05). In conclusion, the goose follicle GCs had obvious apoptosis under oxidative stress. Adding VE could inhibit oxidative stress and relieve goose follicle GCs apoptosis. The study can provide reference for improving goose laying performance.

Key words： Vitamin E Oxidative stress Apoptosis Goose follicular granulosa cells

收稿日期: 2022-05-03

ZTFLH:

S831.2

基金资助:江苏现代农业产业技术体系建设专项资金(JATS[2020]435); 江苏省高等学校大学生创新创业训练计划项目(202111117115Y)

通讯作者: ** yuzhang@yzu.edu.cn

作者简介: *同等贡献作者

引用本文:

皇甫奕雯, 宋莉娜, 侯丽娥, 张扬, 陈国宏, 徐琪, 张钰. 维生素 E 通过抑制氧化应激缓解鹅卵泡颗粒细胞凋亡[J]. 农业生物技术学报, 2023, 31(8): 1659-1670.
HUANGFU Yi-Wen, SONG Li-Na, HOU Li-E, ZHNAG Yang, CHEN Guo-Hong, XU Qi, ZHANG Yu. Vitamin E Relieves the Apoptosis of Goose (Anser cygnoides) Granulosa Cells by Inhibiting Oxidative Stress. 农业生物技术学报, 2023, 31(8): 1659-1670.

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http://journal05.magtech.org.cn/Jwk_ny/CN/10.3969/j.issn.1674-7968.2023.08.010 或 http://journal05.magtech.org.cn/Jwk_ny/CN/Y2023/V31/I8/1659

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