Park Kyu-Mi, Kim Kyu-Jun, Jin Minghui, Han Yongquan, So Kyoung-Ha, Hyun Sang-Hwan
Institute for Stem Cell and Regenerative Medicine (ISCRM), Chungbuk National University, Cheongju 28644, Korea.
Laboratory of Veterinary Embryology and Biotechnology (VETEMBIO), Veterinary Medical Center and College of Veterinary Medicine, Chungbuk National University, Cheongju 28644, Korea.
Asian-Australas J Anim Sci. 2019 Dec;32(12):1844-1853. doi: 10.5713/ajas.19.0162. Epub 2019 Aug 3.
We investigated how pituitary adenylate cyclase-activating polypeptide (PACAP) affects embryonic development during pre-in vitro maturation (pre-IVM) using porcine oocytes isolated from small follicles.
We divided the follicles into the experimental groups by size (SF, small follicles; MF, medium follicles) and treated with and without PACAP and cultured for 18 hours (Pre-SF[-]PACAP; without PACAP, Pre-SF[+]PACAP; with PACAP) before undergoing IVM. The gene expression related to extracellular matrix formation (amphiregulin, epiregulin, and hyaluronan synthase 2 [HAS2]) and apoptosis (Bcl-2-associated X [BAX], B-cell lymphoma 2, and cysteine-aspartic acid protease 3) was investigated after maturation. The impact on developmental competence was assessed by the cleavage and blastocyst rate and total cell number of blastocysts in embryos generated from parthenogenesis (PA) and in vitro fertilization (IVF).
Cleavage rates in the Pre-SF(+)PACAP after PA were significantly higher than SF and Pre-SF(-)PACAP (p<0.05). The cleavage rates between MF and Pre- SF(+)PACAP groups yielded no notable differences after IVF. Pre-SF(+)PACAP displayed the higher rate of blastocyst formation and greater total cell number than SF and Pre-SF(-)PACAP (p<0.05). Cumulus cells showed significant upregulation of HAS2 mRNA in the Pre-SF(+)PACAP compared to the SF (p<0.05). In comparison to other groups, the Pre-SF(+)PACAP group displayed a downregulation in mRNA expression of BAX in matured oocytes (p<0.05).
The PACAP treatment during pre-IVM improved the developmental potential of porcine oocytes derived from SF by regulating cumulus expansion and apoptosis of oocytes.
我们研究了垂体腺苷酸环化酶激活多肽（PACAP）如何利用从小卵泡中分离出的猪卵母细胞在体外成熟前（pre-IVM）阶段影响胚胎发育。
我们根据卵泡大小将其分为实验组（SF，小卵泡；MF，中等卵泡），并在进行体外成熟（IVM）前，分别用或不用PACAP处理并培养18小时（Pre-SF[-]PACAP；无PACAP，Pre-SF[+]PACAP；有PACAP）。成熟后，研究与细胞外基质形成相关的基因表达（双调蛋白、表皮调节素和透明质酸合酶2 [HAS2]）以及凋亡相关基因表达（Bcl-2相关X蛋白[BAX]、B细胞淋巴瘤2和半胱天冬酶3）。通过孤雌生殖（PA）和体外受精（IVF）产生的胚胎的卵裂率、囊胚率和囊胚总细胞数来评估对发育能力的影响。
PA后Pre-SF(+)PACAP组的卵裂率显著高于SF组和Pre-SF(-)PACAP组（p<0.05）。IVF后，MF组和Pre-SF(+)PACAP组之间的卵裂率没有显著差异。Pre-SF(+)PACAP组的囊胚形成率更高且总细胞数比SF组和Pre-SF(-)PACAP组更多（p<0.05）。与SF组相比，Pre-SF(+)PACAP组的卵丘细胞中HAS2 mRNA显著上调（p<0.05）。与其他组相比，Pre-SF(+)PACAP组成熟卵母细胞中BAX的mRNA表达下调（p<0.05）。
体外成熟前阶段的PACAP处理通过调节卵丘扩展和卵母细胞凋亡，提高了源自小卵泡的猪卵母细胞的发育潜力。
