枸杞多糖干预对提高小鼠冻存卵巢移植效果的相关机制研究

Frozen thawed ovarian autotransplantation is a strategy to preserve fertility of young patients with cancer, freeze-thaw hypoxic ischemic damage after transplantation has become the main bottleneck of prevention to improve the life and function of the grafts. The study to reduce ovarian damage after freeze-thaw and transplantation induced by oxidative stress has caused concern. A number of studies have demonstrated that Lycium barbarum polysaccharides (LBP) play a protective effect on cells by antioxidant. According to our research,the effect of LBP improve the freeze-thaw fetal ovarian on follicle survival rate after xenotransplantation is better than that of the antioxidant beta mercaptoethanol. However, to make the LBP with no toxic and side effects to play a protection role in ovarian freeze-thaw and transplantation, the related of the dose and its mechanism should be clearly. Therefore, this study intend to make LBP intervention on mouse ovarian in the process of vitrification, to explore the optimal dose of LBP protection and the related mechanism by detecting the cold shock proteins after thawn, key protein of oxidative stress , normal follicles, the cell gap junction protein between oocytes and granulosa and Nrf/ARE signal pathway detection . In order to provide scientific evidence of LBP which can play a role in freeze-thaw ovary transplanted through oxidative stress pathways, by making the model of anticancer treatment receptor under the chemotherapy drugs, blood vessels perfusion with fluorescence, use orthogonal experimental to observe if LBP intervention can improve rapid revascularization and fertility potential of grafts of chemotherapy drug receptor and provide scientific basis for the application of LBP in the preservation of fertility in patients with chemotherapy.

冻融卵巢的自体移植是保存青年癌症患者生育力的一个策略，冻融移植后的缺血缺氧损伤已成为阻碍提高移植卵巢功能寿命的主要瓶颈。降低氧化应激致冻融移植卵巢损伤的的研究已引起关注。大量研究证实枸杞子的有效成分枸杞多糖(LBP)通过抗氧化发挥细胞保护作用，本组前期工作显示，LBP提高冻融胎儿卵巢异种移植后卵泡存活率的作用优于抗氧化剂β-巯基乙醇。然而，要发挥无毒无副作用的LBP在卵巢冻融移植中的保护作用，须明确相关的剂量及机制。故本研究拟对小鼠卵巢进行玻璃化冻存过程中的LBP干预，通过检测解冻后冷应激蛋白、氧化应激关键蛋白、正常卵泡百分比、卵母细胞与颗粒细胞间缝隙连接蛋白及氧化应激Nrf/ARE信号通路检测，探索最适LBP保护剂量和相关机制。通过血管荧光灌注、正交试验，观察接受化疗药物受体经LBP干预后是否能起到使移植卵巢血流快速重建及提高生育潜能，为LBP在化疗患者生育力保存中的应用提供科学依据。

冻融卵巢的自体移植是保存青年癌症患者生育力的一个策略，冻融移植后的缺血缺氧损伤已成为阻碍提高移植卵巢功能寿命的主要瓶颈。降低氧化应激致冻融移植卵巢损伤的研究已引起关注。故本研究拟对小鼠卵巢进行玻璃化冻存过程中的 LBP 干预，通过检测解冻后冷应激蛋白、氧化应激关键蛋白、正常卵泡百分比、卵母细胞与颗粒细胞间缝隙连接蛋白及氧化应激 Nrf/ARE 信号通路检测，探索最适 LBP 保护剂量和相关机制。通过血管荧光灌注、正交试验，观察接受化疗药物受体经 LBP 干预后是否能起到使移植卵巢血流快速重建及提高生育潜能，得到400μg/ml 的枸杞多糖用于小鼠卵巢组织的玻璃化冻存，可有效的抑制冻存卵巢内卵泡的凋亡，从而提高冻存卵泡的活性，在冷冻保护剂以及培养液内加入枸杞多糖和Vit-C 后，对照组相比，可上调VEGF/VEGFR和缝隙连接蛋白Cx43和Cx37的表达，说明枸杞多糖有利于小鼠卵巢组织的玻璃化冻存，为 LBP 在化疗患者生育力保存中的应用提供科学依据。通过ELISA法检测了冷冻卵巢组织中CIRBP、 Rbm3、8-OHdG 、Sod1、MDA和GSH-Px，说明了枸杞多糖有利于卵巢组织的冷冻保存。通过对化疗受体经 LBP干预后对卵巢移植的血流重建及卵巢储备影响以及枸杞多糖全程干预对玻璃化冻融卵巢原位移植后的生育力保护作用的研究，也证实了LBP干预也有助于小鼠半卵巢冻存后原位移植的生育力恢复研究。