rhFSH干预对玻璃化冻融小鼠卵巢中葡萄糖神经酰胺转移酶表达及卵泡凋亡的影响

Effect of rhFSH Intervention on the Expression of UDP-glucose Ceramide Glucosyltransferase and Follicles Apoptosis in Mice Freezing-thawing Ovary with Vitrification

目的通过观察重组人卵泡刺激素(rh FSH)干预对玻璃化冻融小鼠卵巢中葡萄糖神经酰胺转移酶(UGCG)表达以及卵泡凋亡的影响,提供rh FSH降调神经酰胺的含量从而达到抗凋亡作用的实验室依据。方法将4周龄C57BL/6J小鼠卵巢随机分为三组:新鲜对照组(FCG)、玻璃化冻存对照组(VCG)、300 IU·L^(-1)rh FSH干预玻璃化冻存组(rh FSH-VG)。通过对冻融卵巢内各级卵泡计数、免疫组织化学技术及Western blot技术,观察并分析各组卵巢内葡萄糖神经酰胺转移酶(UGCG)、抗凋亡蛋白Bcl2和促凋亡蛋白Bax在卵巢不同细胞中的定位和蛋白表达量。结果与FCG组相比,两个冻融组卵巢结构较疏松,但rh FSH-VG组卵泡基质结构较VCG组整齐且较致密。原始卵泡、初级卵泡、正常卵泡和卵泡总数的计数及UGCG和Bcl2的表达显示,rh FSH-VG组高于VCG组(P<0.05),但这两个冻融组均低于FCG组(P<0.05)。闭锁卵泡计数和Bax的表达显示rh FSH-VG组低于VCG组(P<0.05),但这两个冻融组均高于FCG组(P<0.05)。结论玻璃化冻融过程中添加rh FSH可以上调UGCG和Bcl2的表达,下调Bax表达,提示其可能通过降调神经酰胺的含量从而达到抗凋亡目的。

Objective To investigating the effect of recombinant human follitropin（ rh FSH） intervention on the expression of UDP- glucose ceramide glucosyltransferase（ UGCG） and follicles apoptosis in mice freezing- thawing ovary with vitrification,then to provide laboratory basis that rh FSH can down regulate the content of ceramide so as to achieve the antiapoptotic effect. Methods Four- week- old C57 BL /6J mice ovarian were used and randomized into three groups: fresh control group（ FCG）,vitrification control group（ VCG）,rh FSH intervented vitrification group（ rh FSH- VG） treated with 300 IU·L-1rh FSH during freezing- thawing. By follicle counting at all levels,immunohistochemistry and western blot in the freezing- thawing ovary,the location and quantity of protein expression of intracellular UGCG,antiapoptotic proteins Bcl2 and promoting apoptosis proteins Bax in different cells of ovary were observed. Results The morphology of follicle showed good in FCG group while two freezing- thawing groups of follicle morphological structure was loose. Compared with VCG group,rh FSH- VG group of follicle matrix structure was in order and more dense.Primordial follicle,primary follicle,normal follicle,total follicle counting and the expression of UGCG and Bcl2 showed that rh FSH- VG group was significantly higher than VCG group（ P < 0. 05）,but these two freezing- thawing groups were all significantly lower than FCG group（ P < 0. 05）. Atresia follicle counting and the expression of Bax showed that rh FSH- VG group was significantly lower than VCG group（ P < 0. 05）,but these two freezing- thawing groups were all significantly higher than FCG group（ P < 0. 05）. Conclusion In process of freezing- thawing ovary with vitrification,the intervention of rh FSH can up regulate the expression of UGCG,which illustrated that rh FSH can down regulate the content of ceramide so as to achieve the aim of antiapoptotic.