GRP78在糖尿病大鼠逼尿肌细胞内质网应激中的相关性研究

发布时间：2018-06-13 17:17

本文选题：糖尿病大鼠 + 逼尿肌　；参考：《山西医科大学》2015年硕士论文

【摘要】：目的:通过探讨GRP78在糖尿病大鼠逼尿肌细胞内质网应激病变发生发展,特别是逼尿肌细胞凋亡过程中的作用和机制,揭示GRP78在糖尿病大鼠逼尿肌细胞内质网应激病变发生发展中的动态演变规律变化,为DCP的病因学研究提供重要的实验依据。方法:在本研究中,我们选取GRP78作为ERS的标志性因子,取健康清洁级成年雌性Sprague-Dawley大鼠50只随机平均分为4组糖尿病组和1组正常对照组。糖尿病组采用一次注射法经腹腔注射链脲佐菌素(STZ)50mg/kg,注射后72小时行鼠尾静脉血糖浓度16.7mmol/L为模型成功。分别在4周、8周、12周、16周处死大鼠,切除膀胱组织,并去除周围脂肪和结缔组织,分别采用tunel法测定逼尿肌细胞凋亡率和电镜下观察逼尿肌细胞内质网形态学变化。用RT-PCR技术检测及Western-Blot技术动态研究糖尿病大鼠逼尿肌细胞凋亡水平与GRP78表达情况的相互关系,并进行所有数据的统计学分析。结果:糖尿病大鼠造模成功即出现多饮、多食、排尿增多,体型明显消瘦,毛发变黄变硬,光泽度下降,出现嗜睡对外界条件反应迟钝,个别糖尿病大鼠四肢溃烂,对疼痛不敏感,对照组大鼠饮食状况正常;糖尿病各组凋亡率随着时间的递增而升高,12w时升高趋势明显,凋亡趋势呈时间依赖性;在电镜下糖尿病组逼尿肌细胞的内质网细胞核周围大量内质网无序排列,体积增大,为白色不规则空泡样结构,浆膜附着核糖小体;RT-PCR检测结果显示,4w组和8w组明显高于对照组,12w组表达水平最高,是对照组的8.87倍,而16w组表达水平较12w有所下降。Western blot检测结果显示,GRP78蛋白表达水平4w组较对照组出现升高,8w组表达水平最高,是对照组的1.45倍,而12w组和16w组较8w组明显下降。结论:DCP逼尿肌细胞凋亡与内质网应激密切相关,其中GRP78表达水平的改变是至逼尿肌细胞凋亡的重要病变机制,随着糖尿病病程的进展,GRP78与DCP有明显的相关性。通过研究DCP的病变与GRP78的关系,会使我们对于糖尿病膀胱病因学机制的理解更加深入并且为其治疗提供新的潜在治疗靶点。
[Abstract]:Objective: to investigate the role and mechanism of GRP78 in the development of endoplasmic reticulum stress in detrusor cells in diabetic rats, especially in the process of detrusor cell apoptosis. To reveal the dynamic evolution of GRP78 in the development of endoplasmic reticulum stress in detrusor cells of diabetic rats, and to provide an important experimental basis for the etiological study of DCP. Methods: in this study, we selected GRP78 as the symbol of ERS, and 50 healthy and clean female Sprague-Dawley rats were randomly divided into 4 diabetic groups and 1 normal control group. In the diabetic group, 50 mg / kg streptozotocin (STZ) was injected intraperitoneally, and the blood glucose concentration of rat tail vein was taken at 72 hours after injection. The rats were killed at 4 weeks, 8 weeks and 12 weeks and 16 weeks, respectively. The bladder tissues were excised, and peripheral fat and connective tissues were removed. The apoptosis rate of detrusor cells and the morphological changes of endoplasmic reticulum of detrusor cells were measured by tunel method and electron microscope respectively. The relationship between the expression of GRP78 and apoptosis of detrusor cells in diabetic rats was studied by RT-PCR and Western-Blot, and all the data were analyzed statistically. Results: the success of model making in diabetic rats resulted in more drinking, more food, more urination, obvious weight loss, yellowing and hardening of hair, decreased gloss, dull response to external conditions, and limb ulceration in individual diabetic rats. The rats in the control group were not sensitive to pain, the diet of the control group was normal, the apoptotic rate of the diabetic groups increased with the increase of time, and the trend of apoptosis was time-dependent. Under electron microscope, a large number of endoplasmic reticulum (ER) cells around the endoplasmic reticulum (ER) nuclei in diabetic group were arranged disorderly, and the volume of endoplasmic reticulum (ER) was increased, which was a white irregular vacuolar structure. The results of RT-PCR analysis showed that the expression of ribosome in serous membrane was significantly higher in 4w group and 8w group than in control group (12w), 8.87 times higher than that in control group (8.87 times). The results of Western blot analysis showed that the expression level of GRP78 protein was the highest in the 4w group compared with the control group, which was 1.45 times higher than that in the control group, while the expression level in the 12w and 16w groups was significantly lower than that in the 8w group. Conclusion the apoptosis of detrusor cells is closely related to endoplasmic reticulum stress. The change of GRP78 expression level is an important pathological mechanism of detrusor cell apoptosis. There is a significant correlation between GRP78 and DCP with the progression of diabetes mellitus. By studying the relationship between DCP lesions and GRP78, we can further understand the etiological mechanism of diabetic bladder and provide a new potential therapeutic target for its treatment.
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R587.2

【相似文献】