Wang Jie, Xu Guijiang, Cai Yu, Wang Lihong, Zhang Wei
Jiangsu Agri-Animal Husbandry Vocational College, Taizhou, 225300, Jiangsu, People's Republic of China.
Yangzhou Maternal and Child Health Care Service Center, The Affiliated Hospital of Yangzhou University Medical College, Yangzhou, 225009, Jiangsu, People's Republic of China.
In Vitro Cell Dev Biol Anim. 2022 Jan;58(1):21-28. doi: 10.1007/s11626-021-00618-x. Epub 2022 Jan 6.
The apoptosis of granulosa cells can result in follicular atresia, a key process in follicle selection and development. The related molecular mechanisms were explored to study the effects of liver receptor homolog-1 (Lrh-1) on apoptosis, sodium-titanium system, and steroid synthesis of granulosa cells from Hu sheep in vitro. After constructing, designing, and synthesizing the Lrh-1 overexpression vector, liposomes were used to transfect Hu sheep granulosa cells. Thereafter, qRT-PCR and Western blot were used to detect the mechanism of Lrh-1 on the titanium system and steroid synthesis of Hu sheep granulosa cells. The overexpression efficiency was determined by fluorescence 48 h after liposome transfection into the Hu sheep granulosa cells in vitro culture. The transfection efficiency was higher, hence providing a basis for subsequent experiments. However, the protein level and transcriptional level of Lrh-1 significantly increased after transfection with Lrh-1 overexpression vector in Hu sheep granulosa cells. We further revealed that after Lrh-1 overexpression in Hu sheep granulosa cells, the expression of the pro-apoptotic gene Bax decreased significantly, while that of the anti-apoptotic gene Bcl-2 increased significantly, as well as the sodium peptide system. Moreover, the expression levels of natriuretic peptide precursor A, B, and C (NPPA, NPPB, NPPC) all showed an upward trend, while the expression levels of steroid synthesis-related genes (P450arom and P450scc) decreased. The above results showed that Lrh-1 overexpression influenced the apoptosis, sodium peptide system, and steroid synthesis.
颗粒细胞凋亡可导致卵泡闭锁，这是卵泡选择和发育的关键过程。本研究旨在探讨肝脏受体同源物-1（Lrh-1）对湖羊颗粒细胞凋亡、钠钛系统及类固醇合成的影响及其相关分子机制。构建、设计并合成Lrh-1过表达载体后，采用脂质体转染湖羊颗粒细胞。随后，运用qRT-PCR和蛋白质免疫印迹法检测Lrh-1对湖羊颗粒细胞钛系统和类固醇合成的作用机制。通过对体外培养的湖羊颗粒细胞进行脂质体转染48 h后的荧光检测来确定过表达效率。转染效率较高，为后续实验提供了依据。然而，在湖羊颗粒细胞中转染Lrh-1过表达载体后，Lrh-1的蛋白水平和转录水平显著升高。我们进一步发现，湖羊颗粒细胞中Lrh-1过表达后，促凋亡基因Bax的表达显著降低，而抗凋亡基因Bcl-2的表达显著增加，钠肽系统也发生了变化。此外，利钠肽前体A、B和C（NPPA、NPPB、NPPC）的表达水平均呈上升趋势，而类固醇合成相关基因（P450arom和P450scc）的表达水平下降。上述结果表明，Lrh-1过表达影响了细胞凋亡、钠肽系统和类固醇合成。
