鞣花酸及其代谢物对山羊卵巢颗粒细胞促增殖、抗氧化、抑凋亡的效果对比研究

doi:10.11843/j.issn.0366-6964.2025.12.024

摘要/Abstract

摘要： 旨在研究鞣花酸(EA)及其代谢物尿石素A(UA)、尿石素B(UB)和尿石素C(UC)对贵州黑山羊卵巢颗粒细胞(OGCs)促增殖、抗氧化、抑凋亡的效果对比，以期寻找相较于EA更优的选择。本研究通过采集健康且性成熟的羊卵巢组织，分离培养OGCs，经鉴定后用于后续研究。同时，基于课题组前人的研究发现，1 μmol·L^-1浓度的EA对OGCs具有最优的生物学效应。因此，试验分为5组：对照组(CON组)、EA组、UA组、UB组、UC组，处理组分别添加1 μmol·L^-1对应药物。随后，采用CCK-8检测添加各组药物0、24、36及48 h对OGCs细胞增殖的影响，采用细胞划痕试验检测其对OGCs细胞迁移的影响，采用ROS试剂盒检测其对OGCs的 ROS水平影响，最后通过qRT-PCR检测其对抗凋亡基因Bcl-2，促凋亡基因Bax、Caspase-3、Caspase-9，炎症因子IL-1β、IL-6、TNF-α及繁殖相关基因GDF9、BMPR-1B、CYP19A1、FSHβ mRNA表达的影响。UA对OGCs的增殖效果最好，其次是EA与UB，最后是UC；细胞划痕试验发现UA显著促进了细胞迁移能力，UB显著抑制了细胞迁移，且EA与UA相比，两者之间并无显著性差异；ROS试验发现，UA与EA显著降低了细胞内ROS含量；qRT-PCR结果发现，与CON组相比，UA、UB、UC、EA组中促凋亡基因CASP3、CASP9、BAX的mRNA表达水平显著降低，而在抗凋亡方面，UA组中BCL-2的表达量显著高于其他组。对炎症因子mRNA表达水平的检测结果显示，UA、UC、EA组IL-1β 相对表达量显著低于CON组，且UA组mRNA表达量最低；各组IL-6 的表达量与CON组无显著差异，但TNF-α含量均显著降低。对繁殖相关基因的检测结果显示，UA、EA组GDF9、BMPR-1B、CYP19A1和FSHβ的表达量均显著高于CON组，其中UA组表达量最高。本研究发现，与EA及EA其他代谢物相比，UA在促进OGCs增殖、迁移、抗氧化、抗凋亡及调控生殖-炎症相关基因表达方面具有显著优势，可作为哺乳动物繁殖调控的重要添加剂进行深入研究。

关键词: 鞣花酸, 卵巢颗粒细胞, 代谢产物, 效果对比研究

Abstract: This study aimed to compare the effects of ellagic acid (EA) and its metabolites urolithin A (UA), urolithin B (UB), and urolithin C (UC) on promoting proliferation, enhancing antioxidant activity, inhibiting apoptosis in Guizhou black goat ovarian granulosa cells (OGCs), in order to identify a superior alternative to EA.Ovarian tissues from healthy, sexually mature goats were collected to isolate and culture OGCs, which were validated for subsequent experiments. At the same time, based on previous research findings of the research group, 1 μmol·L^-1concentration of EA had the optimal biological effect on OGCs. Therefore,cells were divided into 5 groups: control (CON), EA, UA, UB, and UC, with treatment groups supplemented with 1 μmol·L^-1 of the corresponding compounds. Proliferation was assessed via CCK-8 assay at 0, 24, 36, and 48 h. Cell migration was evaluated using a scratch assay. Intracellular ROS levels were measured using ROS detection kits. RT-qPCR was employed to analyze mRNA expression of anti-apoptotic (Bcl-2), pro-apoptotic (Bax, Caspase-3, Caspase-9), inflammatory factors (IL-1β, IL-6, TNF-α), and reproduction-related genes (GDF9, BMPR-1B, CYP19A1, FSHβ). UA exhibited the strongest pro-proliferative effect on OGCs, followed by EA and UB, with UC showing the weakest activity. The scratch assay revealed that UA significantly enhanced cell migration, whereas UB inhibited migration; no significant difference was observed between EA and UA. ROS levels were markedly reduced in UA and EA groups. qRT-PCR results demonstrated that UA, UB, UC, and EA significantly downregulated pro-apoptotic genes (CASP3, CASP9, BAX) compared to the CON group. UA upregulated BCL-2 expression most prominently. Among inflammatory factors, UA, UC, and EA significantly suppressed IL-1β expression (with UA being the lowest), there was no significant differences between CON group and other groups, while TNF-α levels decreased across all groups. UA and EA significantly enhanced the expression of reproduction-related genes (GDF9, BMPR-1B, CYP19A1, FSHβ), with UA showing the highest efficacy. In this study, compared to EA and other metabolites, UA demonstrated superior effects in promoting OGCs proliferation and migration, reducing oxidative stress and apoptosis, and modulating reproductive-inflammatory gene expression. These findings highlight UA as a promising candidate for further research as a reproductive regulatory additive in mammals.

Key words: ellagic acid, ovarian granulosa cells, metabolites, comparative study of effects

中图分类号:

S827.3

陈怡, 刘彬, 张习录, 陆情梅, 潘智仁, 施晓丽, 罗志军, 赵佳福. 鞣花酸及其代谢物对山羊卵巢颗粒细胞促增殖、抗氧化、抑凋亡的效果对比研究[J]. 畜牧兽医学报, 2025, 56(12): 6219-6231.

CHEN Yi, LIU Bin, ZHANG Xilu, LU Qingmei, PAN Zhiren, SHI Xiaoli, LUO Zhijun, ZHAO Jiafu. A Comparative Study on the Effects of Ellagic Acid and Its Metabolites on Proliferation Promotion, Antioxidation, Apoptosis Inhibition of Goat Ovarian Granulosa Cells[J]. Acta Veterinaria et Zootechnica Sinica, 2025, 56(12): 6219-6231.

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