外源性内脂素对高龄雌鼠卵巢功能的影响

发布时间：2018-03-10 11:37

本文选题：外源性内脂素　切入点：诱导排卵　出处：《河北联合大学》2014年硕士论文　论文类型：学位论文

【摘要】：目的通过检测注射外源性内脂素的高龄雌鼠卵巢组织血管内皮生长因子（vascular endothelial growth factor，VEGF）和内分泌腺性血管内皮生长因子（endocrine gland vascular endothelial growth factor，EG-VEGF）的表达来探讨内脂素对高龄雌鼠卵巢功能和卵子质量的影响，为临床促排卵中卵巢低反应患者的用药提供实验依据。方法将两组不同周龄段（12～16周，32～36周）的SPF级雌性SD大鼠每组各60只，每组内再随机分为两组。在诱导排卵的基础上，应用外源性内脂素的作为治疗组，将仅诱导排卵的一组作为对照组。第一步，诱导排卵，于发情期给予四组雌性SD大鼠腹腔内注射15IU孕母马血清促性腺激素，同时，治疗组予100ng/ml重组鼠内脂素。第二步，注射48小时后，向治疗组与对照组大鼠腹腔内注射50IU人绒毛膜促性腺激素（HCG）。第三步，立即将这些雌鼠和雄鼠进行单独交配。第四步，交配18小时后，颈椎脱臼处死雌鼠，无菌条件下取出双侧输卵管中的受精卵进行体外培养；取出双侧卵巢，右侧卵巢用甲醛固定，免疫组化检测微血管密度（MVD），左侧卵巢的一部分立即置于Trizol中，用于Realtime-PCR检测卵巢VEGF、EG-VEGF的mRNA表达，另一部分则用于Western Blotting检测VEGF、EG-VEGF的蛋白表达。实验数据采用SPSS17.0软件进行统计学分析，计量资料以平均数±标准差（x s）表示，采用单因素方差分析判断组间差异，两组间数据比较采用SNK检验，P0.05认为有统计学意义。结果1年轻对照组平均合子数、卵裂数、卵裂率、平均胚胎数高于大龄对照组，，差异有统计学意义（P＜0.05），大龄治疗组高于大龄对照组，差异有统计学意义（P＜0.05）；而年轻治疗组、年轻对照组两组之间比较，差异无统计学意义（P＞0.05）。2年轻对照组卵巢MVD、VEGF及EG-VEGF的表达水平高于大龄对照组，差异有统计学意义（P＜0.05），大龄治疗组高于大龄对照组，差异有统计学意义（P＜0.05）；而年轻治疗组、年轻对照组两组之间比较，差异无统计学意义（P＞0.05）。结论1诱导排卵的高龄雌鼠的平均合子数、卵裂数、卵裂率、平均胚胎数随周龄的增加而下降，提示高龄雌鼠生殖能力下降。2给予外源性Qg脂素后治疗组高龄雌鼠的平均合子数、卵裂数、卵裂率、平均胚胎数明显增加，提示外源性Qg脂素可能改善高龄雌鼠卵巢功能，提高其生殖能力。3给予外源性Qg脂素后高龄雌鼠卵巢MVD、VEGF、EG-VEGF的表达水平明显提高，提示外源性Qg脂素可能是通过促进卵巢血管的生成从而改善高龄雌鼠的卵巢功能和生殖能力。
[Abstract]:Objective to investigate the expression of vascular endothelial growth factor (VEGF) and secreted glandular vascular endothelial growth factor (VEGF) in ovarian tissues of elderly female rats injected with exogenous endolipin, and to explore the effect of endocrine gland vascular endothelial growth factor-EG-VEGFF on the oocytes of elderly female rats. The effects of nest function and egg quality, To provide experimental evidence for the treatment of ovarian hyporeactive patients in clinical ovulation promotion. Methods two groups of female SD rats of SPF grade were randomly divided into two groups. On the basis of ovulation induction, exogenous endolipin was used as treatment group. In the first step, ovulation was induced by intraperitoneal injection of serum gonadotropin in female SD rats of 15 IU during estrus. Meanwhile, the treatment group was given 100ng / ml recombinant rat lipopolipin. After 48 hours of injection, 50 IU human chorionic gonadotropin (HCG) was injected intraperitoneally into the treatment group and the control group. In the third step, the female and male mice were mated separately. 4th steps, 18 hours after mating, the female rats were killed by cervical dislocations. Fertilized eggs from bilateral oviducts were removed for culture in vitro under aseptic conditions, bilateral ovaries were removed, the right ovaries were fixed with formaldehyde, microvessel density was detected by immunohistochemistry, and a portion of the left ovary was placed in Trizol immediately. The Realtime-PCR was used to detect the expression of EG-VEGF in ovary and the other part was used to detect the protein expression of EG-VEGF in VEGFU by Western Blotting. The experimental data were analyzed statistically by SPSS17.0 software, and the measured data were expressed as mean 卤standard deviation (x / s). Single factor analysis of variance was used to judge the difference between the two groups. The data of the two groups were compared by SNK test (P0.05). Results 1the average number of zygotes, cleavage, cleavage rate and the number of embryos in the young control group were higher than those in the older control group (P < 0.05), the difference was statistically significant (P < 0.05), and the difference was statistically significant in the older treatment group than in the older control group (P < 0.05), while in the young treatment group, the difference was significant (P < 0.05). There was no significant difference between the two groups (P > 0.05). 2 the expression of EG-VEGF and VEGF in the ovary of the young control group was higher than that of the older control group (P < 0.05), and the expression of EG-VEGF in the older group was higher than that in the older control group (P < 0.05). The difference was statistically significant (P < 0.05), but there was no significant difference between the young treatment group and the young control group (P > 0.05). Conclusion 1 the average number of zygotes, cleavage, cleavage rate and the average number of embryos decreased with the increase of week age. It was suggested that the average number of zygotes, cleavage, cleavage rate and the average number of embryos were significantly increased in the treatment group, which suggested that exogenous QG lipopolipin might improve the ovarian function of the aged female rats. After treated with exogenous QG lipopolipin, the expression of EG-VEGF in ovaries of aged female rats was significantly increased, suggesting that exogenous QG lipopolipin might improve the ovarian function and reproductive ability of elderly female rats by promoting the formation of ovarian blood vessels.
【学位授予单位】：河北联合大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R714.8

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