α-Linolenic acid promotes testosterone synthesis by improving mitochondrial function in primary rooster Leydig cells

IF 2.4 2区农林科学 Q3 REPRODUCTIVE BIOLOGY

Theriogenology Pub Date : 2024-11-01 DOI:10.1016/j.theriogenology.2024.10.026

Xuerui Chang , Danyang Li , Yong Guo , Xihui Sheng , Xiangguo Wang , Kai Xing , Longfei Xiao , Xueze Lv , Cheng Long , Xiaolong Qi

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Abstract

The present study aimed to investigate the direct effects of α-Linolenic acid (ALA) on the in vitro production of testosterone and the expression of key enzymes and proteins related to steroidogenesis in Leydig cells of roosters.

Methods

Purified primary Leydig cells isolated from 65-week-old roosters were purified and treated with different concentrations of ALA treatments: (0 μm/L [control], solvent control group (DMSO), 20 μM/L, 40 μM/L, and 80 μM/L) and cell counting-8 (CCK-8) for cell viability assay, Enzyme-linked immunosorbent assay (ELISA) kit for the determination of testosterone in cell supernatants, quantitative (real-time) PCR, and analysis of activities of antioxidants catalase (CAT), superoxide dismutase (SOD) and malondialdehyde (MDA), evaluation of mitochondrial membrane potential, pro- and anti-apoptotic proteins/genes Bcl-2, Bcl-2-associated X protein (Bax), apoptosis-inducing factor (AIF) were done respectively.

Results

Our results showed that ALA significantly increased testosterone secretion in primary rooster Leydig cells (P < 0.05), and 40 μM/L is the optimal dose. Leydig cells supplemented with ALA (20, 40, 80 μM) increased the expression of key enzymes and proteins 3β-hydroxysteroid dehydrogenase (3β-HSD), steroidogenic acute regulatory protein (StAR), cholesterol side-chain cleavage enzyme (P450scc) concerning steroidogenesis, enhanced antioxidant capability, improved mitochondrial biogenesis, and markedly improved the mitochondrial membrane potential (P < 0.05). Furthermore, the expression of the apoptosis-suppressive gene Bcl-2 was significantly increased, but Bax and AIF expression was decreased in the ALA group compared to that in the control group (P < 0.05).

Conclusion

ALA promoted testosterone production, enhanced steroidogenic enzyme expression, improved mitochondrial function, and antioxidant capacity, and reduced apoptosis in primary rooster Leydig cells, with 40 μM/L identified as the optimal concentration.

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α-亚麻酸通过改善原代公鸡莱迪格细胞线粒体功能促进睾酮合成

本研究旨在探讨α-亚麻酸（ALA）对公鸡Leydig细胞体外睾酮生成及与类固醇生成相关的关键酶和蛋白表达的直接影响。方法纯化从65周龄公鸡分离的原代Leydig细胞，并用不同浓度的ALA处理：(0μm/L[对照组]、溶剂对照组（DMSO）、20μM/L、40μM/L和80μM/L）和细胞计数-8（CCK-8）检测细胞活力，酶联免疫吸附试验（ELISA）试剂盒检测细胞上清液中的睾酮，定量（实时）PCR、分别对抗氧化剂过氧化氢酶（CAT）、超氧化物歧化酶（SOD）和丙二醛（MDA）的活性、线粒体膜电位、促凋亡和抗凋亡蛋白/基因 Bcl-2、Bcl-2 相关 X 蛋白（Bax）、凋亡诱导因子（AIF）进行了分析。结果表明，ALA能显著增加原代公鸡Leydig细胞的睾酮分泌量（P< 0.05），40 μM/L是最佳剂量。补充了 ALA（20、40、80 μM）的 Leydig 细胞增加了与类固醇生成有关的关键酶和蛋白 3β-羟基类固醇脱氢酶（3β-HSD）、类固醇生成急性调节蛋白（StAR）、胆固醇侧链裂解酶（P450scc）的表达，增强了抗氧化能力，改善了线粒体生物生成，并明显提高了线粒体膜电位（P <0.05）。此外，与对照组相比，ALA 组抑制细胞凋亡基因 Bcl-2 的表达明显增加，但 Bax 和 AIF 的表达减少（P < 0.05）。

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来源期刊

Theriogenology 农林科学-生殖生物学

CiteScore

5.50

自引率

14.30%

发文量

387

审稿时长

72 days

期刊介绍： Theriogenology provides an international forum for researchers, clinicians, and industry professionals in animal reproductive biology. This acclaimed journal publishes articles on a wide range of topics in reproductive and developmental biology, of domestic mammal, avian, and aquatic species as well as wild species which are the object of veterinary care in research or conservation programs.