Cornejo-Cortés M A, Sánchez-Torres C, Vázquez-Chagoyán J C, Suárez-Gómez H M, Garrido-Fariña G, Meraz-Ríos M A
Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional, Department of Molecular Biomedicine (CINVESTAV-IPN), Av. IPN 2508, CP 07360, Mexico City, Mexico.
Lab Anim. 2006 Jan;40(1):87-95. doi: 10.1258/002367706775404471.
The present study was performed to optimize a superovulation protocol in rats in order to produce a large number of good-quality embryos suitable to develop rat embryonic stem (rES) cells. We first evaluated the ovulation kinetics of three rat strains: Wistar, Fisher and ACI/N. Animals (n=30 per strain) were treated with 50 IU of pregnant mare serum gonadotrophin (PMSG), and ovulation was induced with 50 IU of human chorionic gonadotrophin (hCG) 50 h apart. Next, we evaluated the dose-response curves of PMSG and hCG in Wistar rats in order to obtain the highest number of embryos. The parameters evaluated for superovulation efficiency were: percentage of mated females, percentage of pregnant females and the average number of embryos collected per female. The results of these experiments suggested that the best dose combination was 50 IU for each hormone. Subsequent experiments, again with Wistar rats, were designed to test which of four hormonal combination treatments (30/30, 30/50, 50/30, and 50/50 IU of PMSG/hCG) will produce the largest numbers of good-quality embryos. Embryo quality was evaluated by embryo development uniformity, embryo morphology, embryo survival in an in vitro culture and embryo ability to generate rES-like cells. Results from these experiments showed that 30/50 IU of PMSG/hCG was the treatment that induced the best embryo quality. In conclusion, our results indicated that, in Wistar rats, the most appropriate hormonal combination dose for superovulation protocols with high number of good-quality embryos was 30 IU of PMSG and 50 IU of hCG given 50 h apart. We are performing further studies with rES-like cells produced with the present methodology to evaluate if they are able to participate in the production of germ-line chimeras.
本研究旨在优化大鼠超排卵方案，以产生大量适合用于培养大鼠胚胎干细胞（rES）的优质胚胎。我们首先评估了三种大鼠品系（Wistar、Fisher和ACI/N）的排卵动力学。对每组30只动物分别用50 IU孕马血清促性腺激素（PMSG）进行处理，并在50小时后用50 IU人绒毛膜促性腺激素（hCG）诱导排卵。接下来，我们评估了Wistar大鼠中PMSG和hCG的剂量反应曲线，以获得最多数量的胚胎。评估超排卵效率的参数包括：交配雌鼠的百分比、怀孕雌鼠的百分比以及每只雌鼠收集到的胚胎平均数量。这些实验结果表明，每种激素的最佳剂量组合为50 IU。随后再次以Wistar大鼠进行实验，旨在测试四种激素组合处理（30/30、30/50、50/30和50/50 IU的PMSG/hCG）中哪种能产生最多数量的优质胚胎。通过胚胎发育均匀性、胚胎形态、体外培养中的胚胎存活率以及胚胎生成rES样细胞的能力来评估胚胎质量。这些实验结果表明，30/50 IU的PMSG/hCG处理诱导出的胚胎质量最佳。总之，我们的结果表明，在Wistar大鼠中，产生大量优质胚胎的超排卵方案最合适的激素组合剂量是间隔50小时给予30 IU PMSG和50 IU hCG。我们正在用目前方法产生的rES样细胞进行进一步研究，以评估它们是否能够参与种系嵌合体的产生。
