Pinyopummintr T, Bavister B D
Department of Animal Health and Biomedical Sciences, University of Wisconsin, Madison, WI 53706, U.S.A.
Theriogenology. 1994;41(6):1241-9. doi: 10.1016/0093-691x(94)90481-w.
Bovine embryos, derived from in vitro matured (IVM)/in vitro fertilized (IVF) ova, were used to investigate the effects of timing of serum inclusion in the culture medium and different types of blood sera and heat inactivation of the serum on embryo development. In Experiment 1, oocytes at 18 h post insemination were allocated to 1 of the following 4 treatments: 1) TCM-199 + 0.1 mg/ml polyvinylalcohol (PVA), 2) TCM-199 supplemented with 10% bovine calf serum (BCS), 3) PVA medium followed by BCS medium at 47 h, or 4) PVA medium followed by BCS medium at 82 h. Supplementation with BCS at 18 h post insemination suppressed (P<0.05) development of morulae/blastocysts (17.6%) when compared with PVA (30.5%) or with serum supplementation at 47 or 82 h post insemination (32.4 and 27.6%, respectively). However, inclusion of BCS at 18, 47 or 82 h post insemination produced more blastocysts (16.8, 29.3 and 22.1%, respectively; P<0.05) than medium +PVA (8.8%). In Experiment 2, ova were cultured from 18 h to 42 h post insemination in PVA-medium, then >/=2-cell embryos were transferred into serum-supplemented medium for another 168 h. Fetal bovine serum (FBS) +/- heat-inactivation (56 degrees C for 30 min, = heated FBS) suppressed morula/blastocyst development compared with medium + PVA, medium + BCS or medium + heated BCS (P<0.05). Bovine calf serum was superior to FBS in supporting blastocyst development (35.1 and 15.2%, respectively), but there was no difference between BCS and heated BCS. However, heated FBS increased the proportion of blastocysts/>/=8-cell embryos compared with that of FBS (51.0 and 31.4%, respectively; P<0.05). These results indicate that the type of serum supplementation and the timing of its inclusion in the culture medium markedly affect bovine embryo development in vitro, and that heat inactivation of serum with high embryotrophic properties is not necessary.
使用源自体外成熟（IVM）/体外受精（IVF）卵子的牛胚胎，来研究培养基中血清添加时间、不同类型血清以及血清热灭活对胚胎发育的影响。在实验1中，将授精后18小时的卵母细胞分配到以下4种处理之一：1）TCM - 199 + 0.1 mg/ml聚乙烯醇（PVA）；2）补充10%胎牛血清（BCS）的TCM - 199；3）PVA培养基，然后在47小时时更换为BCS培养基；4）PVA培养基，然后在82小时时更换为BCS培养基。与PVA（30.5%）或授精后47小时或82小时添加血清（分别为32.4%和27.6%）相比，授精后18小时添加BCS会抑制（P<0.05）桑葚胚/囊胚的发育（17.6%）。然而，授精后18、47或82小时添加BCS产生的囊胚更多（分别为16.8%、29.3%和22.1%；P<0.05），高于培养基+PVA组（8.8%）。在实验2中，卵母细胞在授精后18小时至42小时在PVA培养基中培养，然后将≥2细胞胚胎转移到补充血清的培养基中再培养168小时。与培养基+PVA、培养基+BCS或培养基+热灭活BCS相比，胎牛血清（FBS）±热灭活（56℃30分钟，即热灭活FBS）会抑制桑葚胚/囊胚的发育（P<0.05）。在支持囊胚发育方面，胎牛血清优于FBS（分别为35.1%和15.2%），但BCS和热灭活BCS之间没有差异。然而，与FBS相比，热灭活FBS增加了囊胚/≥8细胞胚胎的比例（分别为51.0%和31.4%；P<0.05）。这些结果表明，血清补充的类型及其在培养基中的添加时间显著影响牛胚胎的体外发育，并且对于具有高胚胎营养特性的血清，热灭活并非必要。
