Beclin1 mRNA和Atg5 mRNA在多囊卵巢综合征患者冠丘颗粒细胞中的表达及其意义

发布时间：2018-03-31 16:17

本文选题：多囊卵巢综合征　切入点：自噬　出处：《河北医科大学》2017年硕士论文

【摘要】：目的:探究多囊卵巢综合征与非多囊卵巢综合征患者放射冠及卵丘颗粒细胞(冠丘颗粒细胞)中自噬相关基因Beclin1 m RNA和Atg5 m RNA的表达是否存在差异及其意义,为研究多囊卵巢综合征(Polycystic Ovarian Syndrome,PCOS)发生、发展中潜在的分子机制提供新线索。方法:对照组入组标准:女方年龄35岁;月经规律;主要因男方因素要求行卵胞浆内单精子显微注射(Intracytoplasmic sperm injection,ICSI)助孕;无盆腔手术史;无子宫内膜异位症;且术前检查各项指标均无异常;无烟酒嗜好吸毒史。多囊卵巢综合征组入组标准(根据2003年鹿特丹标准):女方年龄35岁;主要因男方因素要求行ICSI助孕;无盆腔手术史;无子宫内膜异位症;且术前检查各项指标均无异常;无烟酒嗜好吸毒史;稀发排卵或无排卵;超声表现为双侧卵巢有12个以上直径为2-9mm卵泡;有或无高雄激素的临床表现和(或)高雄激素血症(除外其他高雄激素疾病)。按以上入组标准收取2016年3月1日至2016年6月30日于河北医科大学第二医院生殖医学科首次接受助孕且新鲜周期可移植的患者,多囊卵巢综合征患者组17例和对照组25例废弃的冠丘颗粒细胞及相应患者的临床资料:年龄,身体质量指数(Body Mass Index,BMI),促排方案,扳机日血清促黄体生成素(Luteotropic hormone,LH)、雌性激素(Estrogen,E2)、孕激素(Progesterone,P),获卵数,受精率,是否妊娠(移植后28天经阴道超声提示宫内孕,并探及胚芽及心管搏动视为妊娠)。用Real-Time PCR(Polymerase Chain Reaction)测定冠丘颗粒细胞中Beclin1 m RNA和Atg5 m RNA的表达,根据荧光定量原始检测结果,按照2-△△ct相对定量计算公式计算出各样品的目的基因相对定量结果。用SPSS 17.0统计学分析软件进行数据处理。符合正态分布的计量资料用均数±标准差表示,不符合正态分布的计量资料用中位数(四分位数间距)表示;符合正态分布且方差齐的两样本比较用t检验,不符合正态分布的两样本比较用秩和检验;计数资料的比较用X2检验;两资料均为正态分布的相关采用Pearson相关,不满足条件者采用Spearman秩相关分析。以上均以P0.05为差异有统计学意义。结果:1 Beclin1 m RNA和Atg5 m RNA在两组中均有表达。Beclin1 m RNA和Atg5 m RNA在多囊卵巢综合征组的相对表达量分别是1.612±1.068和0.990(0.96),其中Beclin1 m RNA在多囊卵巢综合征组的表达量高于对照组,差异有统计学意义(P0.05)。2将两组患者的临床资料(BMI,扳机日血清LH、E2、P,获卵数,受精率,妊娠情况)进行分析。与对照组相比,多囊卵巢综合征组BMI值25.048±3.557高于对照组22.660(3.580),且有统计学差异(P0.05);扳机日血清LH在对照组和多囊卵巢综合征组的统计结果分别为1.910(1.140)、1.190(0.620),且在对照组表现出相对高水平,此差异有统计学意义(P0.05);两组妊娠率(妊娠例数/该组总例数)的比较,对照组为15/25,多囊卵巢综合征组为4/17,对照组妊娠率高于多囊卵巢综合征组,差异有统计学意义(P0.05)。扳机日血清E2、P,获卵数、受精率两组资料无统计学差异(P0.05)。用Pearson相关分别分析Beclin1 m RNA的表达与多囊卵巢综合征组中BMI、血清P值、受精率之间的相关性;用Spearman秩相关分别分析Beclin1 m RNA的表达与多囊卵巢综合征组中血清LH值、血清E2值、获卵数之间的相关性。研究显示Beclin1 m RNA的表达与多囊卵巢综合征组中BMI、获卵数的相关系数分别为ρ=0.332、r=0.310,均有正相关关系,但是均无统计学意义(P0.05);Beclin1 m RNA的表达与多囊卵巢综合征组中其他临床资料无相关性。比较19例妊娠者与23例非妊娠者Beclin1 m RNA和Atg5 m RNA的表达情况,在非妊娠组中两基因表达活性较高,且Beclin1 m RNA在两组中表达差异有统计学意义(P0.05)。结论:1 Beclin1 m RNA和Atg5 m RNA在两组中均有表达,提示细胞自噬普遍存在于冠丘颗粒细胞中。2 Beclin1 m RNA在两组之间的表达差异有统计学意义(P0.05),且在多囊卵巢综合征组的表达量高于对照组,可从一个方面提示多囊卵巢综合征组的自噬活性高于对照组,推测颗粒细胞可通过增强自噬活性,对抗和修复卵巢微环境变化所带来的损伤。3 Beclin1和Atg5分别在自噬体启动形成、延伸中起重要作用。Beclin1 m RNA在两组之间的表达差异有统计学意义(P0.05),Atg5m RNA在两组之间的表达无统计学差异(P0.05),表明PCOS患者冠丘颗粒细胞自噬活性改变及过程,仍需要大样本和后续研究的进一步支持。4 多囊卵巢综合征组冠丘颗粒细胞自噬活性增加与BMI,扳机日血清LH、E2、P关系不密切,即以上临床资料并不是影响冠丘颗粒细胞自噬活性改变的主要因素,也不是通过影响冠丘颗粒细胞自噬活性导致该组妊娠率降低。5 妊娠与非妊娠之间的比较,自噬活性存在差异,且非妊娠组自噬活性较高,仍需进行更多临床资料分析,寻找自噬活性存在差异的原因。6 PCOS患者高雄激素、血糖、血脂、空腹胰岛素、是否存在烟雾暴露等是否会影响冠丘颗粒细胞自噬过程,是否会通过改变冠丘颗粒细胞自噬而影响卵子质量进而影响妊娠率仍需进一步讨论及验证。7 PCOS与PCO患者中冠丘颗粒细胞自噬情况是否有差异,亦需要进一步探索。8 本文讨论冠丘颗粒细胞自噬与PCOS的关系,对PCOS发生、发展中潜在的分子机制研究提供了新思路,但是仍有较多亟待研究讨论的问题需要去探索。
[Abstract]:Objective: To explore the polycystic ovary syndrome in patients with polycystic ovary syndrome and non coronal radiation and cumulus cells (Corona cumulus granulosa cells) whether the expression of autophagy related gene Beclin1 m RNA and Atg5 m RNA the difference and significance of polycystic ovary syndrome (Polycystic Ovarian Syndrome, PCOS), provide new clues to the molecular in the development of potential mechanism. Methods: control group group: the woman 35 years of age; menstrual regularity; mainly due to male factors required for intracytoplasmic sperm injection (Intracytoplasmic sperm, injection, ICSI) of pregnancy; no history of pelvic surgery; without endometriosis; and preoperative indicators examination no abnormal; no smoking and drinking history of drug abuse. Patients enrolled PCOS (according to 2003 Rotterdam standard): the age of 35 years; the main factors for the man asked for ICSI help pregnant; no history of pelvic surgery; no Endometriosis; and the preoperative indicators were no exception; no smoking and drinking history of drug abuse; dilute ovulation or anovulation; ultrasound showed bilateral ovaries with more than 12 2-9mm diameter follicles; clinical and performance with or without hormone (or Kaohsiung) Kaohsiung hormones (except other hormone Kaohsiung disease) according to the above standard charge into the group. From March 1, 2016 to June 30, 2016 in the second hospital of Hebei Medical University Department of reproductive medicine for the first time and the fresh cycle can help pregnant transplant patients, patients with polycystic ovary syndrome group and control group of 17 cases of clinical data of 25 cases of granular cell and Crown Hill abandoned the corresponding patients: age, body mass index (Body, Mass Index, BMI), cupai scheme, the trigger on serum luteinizing hormone (Luteotropic, hormone, LH), female sex hormones (Estrogen, E2), progesterone (Progesterone, P), the number of oocytes, fertilization rate, pregnancy (transplantation After 28 days of vaginal ultrasound tip intrauterine pregnancy, and to explore and germ and heart beat as pregnancy). Real-Time PCR (Polymerase Chain Reaction) expression in granulosa cells of Beclin1 m Crown Hill RNA Atg5 and m RNA assay, fluorescence quantitative detection based on the original results, according to the 2- delta CT relative quantitative calculation formula to calculate the target gene of each sample relative quantitative results. The data were analyzed using SPSS 17 statistical analysis software. With normal distribution measurement data is represented by the standard deviation, does not conform to the normal distribution of the measurement data with the median (four percentile interval); accord with normal distribution and homogeneity of two samples with t test does not meet the normal distribution of two samples compared with rank test; compared with count data X2 test; two normal distribution data are related to the use of Pearson, does not meet the conditions by using Spearman rank correlation The analysis above is P0.05. The difference was statistically significant. Results: there were two groups in the expression of 1 Beclin1 m RNA and Atg5 m RNA.Beclin1 m and Atg5 RNA relative expression of M RNA in polycystic ovary syndrome group were 1.612 + 1.068 and 0.990 (0.96), the Beclin1 m RNA in PCOS the expression of syndrome group than in the control group, the difference was statistically significant (P0.05.2) clinical data of the two groups (BMI, LH, serum E2, trigger, P, number of oocytes, fertilization rate, pregnancy) were analyzed. Compared with the control group, polycystic ovary syndrome group BMI = 25.048 + 3.557 22.660 more than that of control group (3.580), and there was statistical difference (P0.05); the trigger on serum LH in the control group and the polycystic ovary syndrome group comprehensive statistical results were 1.910 (1.140), 1.190 (0.620), and in the control group showed a relatively high level, the difference was statistically significant (P0.05); two the pregnancy rate of group ( The number of cases of pregnancy / the total number of cases) compared to control group 15/25, polycystic ovary syndrome group 4/17 control group, the pregnancy rate was higher in the PCOS group, the difference was statistically significant (P0.05). The trigger on serum E2, P, the number of oocytes, fertilization rate of two groups had no significant difference (P0.05) by Pearson. Expression of Beclin1 m RNA were related with polycystic ovary syndrome group BMI, serum P value, the correlation between the rate of fertilization; using Spearman rank correlation analysis respectively the expression of Beclin1 m RNA and polycystic ovary syndrome group, serum LH level, serum E2 value, a correlation between the number of eggs the study showed that the expression of Beclin1. M RNA and polycystic ovary syndrome in the group of BMI, the correlation coefficient of the number of oocytes were p =0.332, r=0.310 had a significant positive correlation, but had no statistical significance (P0.05); the expression of Beclin1 m RNA in women with polycystic ovary syndrome and other clinical data of syndrome group No correlation. Compare the expression of 19 cases of pregnancy and 23 cases of non pregnancy Beclin1 m RNA and Atg5 m RNA, two gene expression was higher in non pregnant group, and Beclin1 m RNA in the two groups there was significant difference in the expression (P0.05). Conclusion: 1 Beclin1 m RNA and Atg5 m RNA in the two groups were expressed, suggesting that autophagy exists in the Crown Hill of.2 in granulosa cells of Beclin1 m RNA expression had statistical significance between the two groups (P0.05), and the expression of polycystic ovary syndrome group than in the control group, from a hint of polycystic ovary syndrome group of autophagy. Higher than the control group, that of granulosa cells by enhancing autophagy activity against ovarian microenvironment and repair the damage caused by.3 Beclin1 and Atg5 respectively in autophagosome formation started,.Beclin1 plays an important role in the expression of RNA m between the two groups in extension There was statistical significance (P0.05), no significant difference in the expression of Atg5m RNA between the two groups (P0.05), Crown Hill granulosa cells showed that PCOS patients and the change of autophagy process, further support.4 still need large sample of polycystic ovary syndrome group and follow-up study of corona cumulus granulosa cells autophagy activity increased with BMI, on the trigger serum LH, E2, P do not close relationship, namely the clinical data above is not the main factor of change of autophagy particle corona cumulus effect is not affected by the Crown Hill granulosa cell autophagic activity leads to the pregnancy rate of group.5 decreased comparison between pregnancy and non pregnancy, autophagic activity differences, and non pregnancy group of autophagy. High, still need more clinical data analysis, find the reasons for the difference of autophagic activity in patients with PCOS.6 Kaohsiung hormone, blood glucose, blood lipid, fasting insulin, the presence of smoke exposure will affect the Crown Hill etc. Autophagy in granulosa cells process, whether by changing the Crown Hill and the egg quality of granulosa cell autophagy thereby affecting the pregnancy rate still need further discussion and verification of.7 PCOS and PCO in patients with granulosa cell autophagy Crown Hill is whether there are differences, also need to further explore the.8 this paper discusses the relationship between autophagy and PCOS particles on the Crown Hill, PCOS. Provides a new way to study the molecular mechanisms of the development potential, but there are still many problems need to be discussed urgently to explore.

【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R711.75

【参考文献】