Kim Hye-soo, Lee Gab-sang, Kim Ji-hye, Kang Sung-keun, Lee Byeong-chun, Hwang Woo-suk
Department of Theriogenology and Biotechnology, College of Veterinary Medicine, Seoul National University, Seoul 151-742, South Korea.
Theriogenology. 2006 Mar 1;65(4):831-44. doi: 10.1016/j.theriogenology.2005.06.016. Epub 2005 Dec 13.
The present study investigated the expression of ligand and receptor for leptin, and the effect of leptin supplementation on preimplantation development of porcine in vitro fertilized (IVF) and somatic cell nuclear transfer (SCNT) embryos. The IVF embryos were produced using frozen boar semen and SCNT embryos were obtained by nuclear transfer of fetal fibroblasts into enucleated oocytes. The protein expression of leptin ligand and receptor was investigated in in vitro matured oocytes, 2-, 4- and 8-cell embryos, morulae and blastocysts derived from IVF and SCNT using immunofluorescence. Both the ligand and receptor were detected in in vitro matured oocytes and all stage of IVF and SCNT embryos. The IVF and SCNT embryos were cultured in modified North Carolina State University (mNCSU)-23 medium supplemented with various concentrations (0, 1, 10, 100 or 1000 ng/mL) of leptin. The rates of cleavage at day 2 and blastocyst formation at day 7, and cell number of blastocysts were monitored as experimental parameters. In SCNT embryos, supplementing with 1000 ng/mL leptin significantly (P<0.05) increased the rate of blastocysts formation (20.2% versus 12.9%) and total cell number (54.6+/-17.4 versus 45.1+/-15.2) compared to the control group. In IVF embryos, leptin supplementation did not affect preimplantation embryo development and cell number in blastocysts. In conclusion, the present study demonstrated the expression of leptin ligand and receptor and the embryotropic effect of leptin in SCNT embryos.
本研究调查了瘦素配体和受体的表达,以及补充瘦素对猪体外受精(IVF)和体细胞核移植(SCNT)胚胎植入前发育的影响。IVF胚胎使用冷冻公猪精液产生,SCNT胚胎通过将胎儿成纤维细胞核移植到去核卵母细胞中获得。使用免疫荧光法研究了IVF和SCNT来源的体外成熟卵母细胞、2-细胞、4-细胞和8-细胞胚胎、桑葚胚和囊胚中瘦素配体和受体的蛋白表达。在体外成熟卵母细胞以及IVF和SCNT胚胎的所有阶段均检测到了配体和受体。IVF和SCNT胚胎在添加了不同浓度(0、1、10、100或1000 ng/mL)瘦素的改良北卡罗来纳州立大学(mNCSU)-23培养基中培养。监测第2天的卵裂率、第7天的囊胚形成率以及囊胚的细胞数量作为实验参数。在SCNT胚胎中,与对照组相比,添加1000 ng/mL瘦素显著(P<0.05)提高了囊胚形成率(20.2%对12.9%)和总细胞数(54.6±17.4对45.1±15.2)。在IVF胚胎中,补充瘦素不影响植入前胚胎发育和囊胚中的细胞数量。总之,本研究证明了瘦素配体和受体的表达以及瘦素对SCNT胚胎的促胚胎生长作用。