Transcriptome analysis of flathead grey mullet (Mugil cephalus) ovarian development induced by recombinant gonadotropin hormones

Single chain recombinant gonadotropin hormones (rGths), recombinant follicle-stimulating hormone (rFsh) and recombinant luteinizing hormone (rLh), produced in Chinese Hamster Ovary cells induced the entire process of vitellogenesis in the flathead grey mullet (Mugil cephalus) and produced viable eggs and larvae. The aim of the present study was to characterize the ovarian molecular pathways and temporal gene expression patterns throughout vitellogenesis induced by rFsh and rLh. For this, five flathead grey mullet females that were arrested at early stages of gonad development were selected from an experimental group and were sampled to follow ovarian development induced with rGths. Briefly, the fish were weekly treated with increasing doses of rFsh —from 6, 9 to 12 µg kg-1 — and after four weeks, rFsh was combined with rLh —increasing doses of 2.5, 4 to 6 µg kg-1—. Then, when oocyte diameter was = 550 µm, only rLh —9 and 12 µg kg-1— was administered on a three-day-basis to complete oocyte growth to ~ 600 µm. Ovarian samples were collected by cannulation from the same five females at four sampling points during the induced gonadal development; (1) before rGth treatment from initial arrested gonad (Stage I), (2) after four weeks of rFsh administration (Stage II), (3) after the combined treatment with rFsh and rLh (Stage III), and (4) after rLh treatment (Stage IV).

在中国仓鼠卵巢细胞（Chinese Hamster Ovary cells）中重组表达的单链重组促性腺激素（single chain recombinant gonadotropin hormones, rGths）、重组促卵泡激素（recombinant follicle-stimulating hormone, rFsh）与重组促黄体激素（recombinant luteinizing hormone, rLh），可诱导鲻鱼（Mugil cephalus）完成完整的卵黄发生过程，并产生可存活的受精卵及幼体。本研究旨在解析由rFsh与rLh诱导的卵黄发生全程中，卵巢的分子通路及时序基因表达模式。为此，本研究从实验组中选取5尾性腺发育停滞于早期阶段的雌性鲻鱼，通过连续采样追踪rGths诱导的卵巢发育进程。简言之，实验鱼每周接受递增剂量的rFsh注射，剂量梯度为6、9至12 µg·kg⁻¹；给药四周后，改为rFsh联合rLh给药，rLh的剂量梯度为2.5、4至6 µg·kg⁻¹。当卵母细胞直径达到550 µm时，调整给药方案为每三天单次给予rLh（剂量为9与12 µg·kg⁻¹），直至卵母细胞生长至约600 µm。在诱导性腺发育的四个时间节点，研究人员通过插管法从同一5尾雌性鲻鱼体内采集卵巢样本：(1) rGths处理前，取自初始停滞的性腺（阶段I）；(2) 连续四周给予rFsh后（阶段II）；(3) 完成rFsh与rLh联合给药后（阶段III）；(4) 单独给予rLh后（阶段IV）。