College of Animal Science, Xinjiang Agricultural University, Urumqi 830000, China.
Institute of Animal Husbandry and Veterinary Medicine, Zhejiang Academy of Agricultural Sciences, Hangzhou, Hangzhou 310021, China.
Poult Sci. 2024 Jun;103(6):103724. doi: 10.1016/j.psj.2024.103724. Epub 2024 Apr 4.
Sertoli cells (SC) are a type of important cells in the testes, which can provide transport proteins, regulatory proteins, growth factors, and other cytokines for the spermatogenic process. They participate in the regulation of the maturation and differentiation of spermatogenic cells and play an important supporting role in the migration, proliferation, and differentiation of germ cells at all levels in the testes. Previous studies found differential expression of LINC9137, miR-140-3p, and Sodium/Potassium Transporting ATPase Interacting 3 (NKAIN3) genesin high and low sperm motility goose testicular tissues. This study investigated the effects of the LINC9137-miR-140-3p-NKAIN3 signal axis on the proliferation and apoptosis of goose testicular sertoli cells at the cellular level, respectively. The results showed that through acridine orange staining, oil red O staining, Alkaline phosphatase (AKP) staining, and RT qPCR assay, it was comprehensively identified that the cultured testicular sertoli cells were purified in vitro. Through the dual luciferase activity detection test, it was found that LINC9137 has a targeted binding site with miR-140-3p and NKAIN3. In addition, this study found that overexpression of miR-140-3p significantly inhibited the expression of LINC9137 and NKAIN3 in sertoli cells, and their expression was significantly increased when miR-140-3p was interfered with. By measuring cell proliferation activity and apoptosis related gene expression, it was found that overexpression of LINC9137 decreased cell proliferation activity (P > 0.05), while the expression level of apoptosis factor Bcl2 Associated X Protein (Bax)/B-cell lymphoma-2 (Bcl2) increased (P > 0.05). On the contrary, when interfering with LINC9137, the cell proliferation activity of sertoli cells was significantly increased (P < 0.01), and the expression level of apoptosis factor Bax/Bcl2 was significantly reduced (P < 0.05); The effect of miR-140-3p on the proliferation and apoptosis of sertoli cells is opposite to that of LINC9137. Meanwhile, this study co transfected overexpressed LINC9137 and miR-140-3p plasmids into sertoli cells, and found that the effect of LINC9137 overexpression on supporting cell proliferation was weakened by miR-140-3p. This study elucidates the role and function of the LINC9137 miR-140-3p-NKAIN3 signaling axis in the development of goose testes and spermatogenesis, establishes a regulatory network related to spermatogenesis, and provides a theoretical basis for studying the genetic regulation of goose spermatogenesis.
支持细胞(SCs)是睾丸中一种重要的细胞类型,可为精子发生过程提供转运蛋白、调节蛋白、生长因子和其他细胞因子。它们参与调节精子细胞的成熟和分化,并在睾丸各级生精细胞的迁移、增殖和分化中发挥重要的支持作用。先前的研究发现,在高精子活力和低精子活力鹅睾丸组织中,LINC9137、miR-140-3p 和 Sodium/Potassium Transporting ATPase Interacting 3(NKAIN3)基因的表达存在差异。本研究分别在细胞水平上研究了 LINC9137-miR-140-3p-NKAIN3 信号轴对鹅睾丸支持细胞增殖和凋亡的影响。结果表明,通过吖啶橙染色、油红 O 染色、碱性磷酸酶(AKP)染色和 RT-qPCR 检测,综合鉴定体外培养的睾丸支持细胞得到了纯化。通过双荧光素酶活性检测试验,发现 LINC9137 与 miR-140-3p 和 NKAIN3 具有靶向结合位点。此外,本研究发现 miR-140-3p 的过表达显著抑制了支持细胞中 LINC9137 和 NKAIN3 的表达,而当 miR-140-3p 受到干扰时,它们的表达显著增加。通过测量细胞增殖活性和凋亡相关基因的表达,发现 LINC9137 的过表达降低了细胞增殖活性(P>0.05),而凋亡因子 Bcl2 相关 X 蛋白(Bax)/B 细胞淋巴瘤-2(Bcl2)的表达水平升高(P>0.05)。相反,当干扰 LINC9137 时,支持细胞的细胞增殖活性明显增加(P<0.01),凋亡因子 Bax/Bcl2 的表达水平明显降低(P<0.05);miR-140-3p 对支持细胞增殖和凋亡的作用与 LINC9137 相反。同时,本研究将过表达的 LINC9137 和 miR-140-3p 质粒共转染入支持细胞,发现 miR-140-3p 减弱了 LINC9137 对支持细胞增殖的支持作用。本研究阐明了 LINC9137-miR-140-3p-NKAIN3 信号轴在鹅睾丸和精子发生发育中的作用和功能,建立了与精子发生相关的调控网络,为研究鹅精子发生的遗传调控提供了理论依据。