兰州大学机构库 >学院待认领
尿激酶在肾结石形成过程中作用机制的研究
Alternative TitleMechanism of urokinase in the formation of kidney stone
明星
Thesis Advisor岳中瑾
2014-05-28
Degree Grantor兰州大学
Place of Conferral兰州
Degree Name硕士
Keyword肾结石 尿激酶 草酸 肾小管上皮细胞
Abstract目的:观察在尿激酶作用下体外草酸钙晶体的形成和聚集情况,为尿激酶对草酸钙结石的抑制作用提供直接证据;检测机体血清尿激酶含量,探讨尿激酶在机体内环境中对肾结石的抑制作用;通过对尿激酶在草酸对MDCK细胞损伤中作用的研究,探讨尿激酶对尿路结石抑制作用的机制。证明尿激酶对肾结石形成的抑制作用,为肾结石的防治提供新的治疗方向。 方法:首先,利用草酸钙晶体在体外不同条件下生长和聚集情况,证明尿激酶对肾结石的抑制作用,分别配制蒸馏水(空白组)、含有不同浓度枸橼酸(对照组)及尿激酶(实验组)的成石溶液,25℃过夜,用倒置显微镜观察各组成石溶液中结晶的大小形态和聚集情况;其次,随机选取肾结石患者27例(结石组)和体检健康者24例(对照组),应用双抗体夹心酶联免疫吸附法(ELISA)法检测两组血清尿激酶水平,自动生化分析仪检测血清钙离子水平和肌酐水平,分析尿激酶与钙离子和肌酐之间的相关性;同时,培养正常MDCK细胞,在原代培养第5天时,随机分为3组(空白组、草酸组、尿激酶组),分别加入正常培养液、含有草酸(5mM)的培养液及含有草酸(5mM)和尿激酶(10万U)的培养液,于2 h后分别测定各组细胞培养液中丙二醛、乳酸脱氢酶的水平;于0.5h,1h,1.5h,2h时采用CCK-8试剂盒检测细胞活力,倒置显微镜下观察细胞表面晶体粘附情况。 结果:体外成石溶液中,空白组晶体明显比对照组和实验组大,并且更易于相互聚集,对照组和实验组草酸钙晶体的成长和聚集很大程度上被抑制;血清试验中,结石组血清尿激酶水平低于对照组(ug/L: 0.49±0.15vs1.44±0.55)(P<0.05),钙离子水平(mmol/L: 2.30±0.18vs2.18±0.13)、肌酐浓度(umol/L: (54.38,126.26)vs(46.53,83.24))高于对照组(P<0.05),结石组尿激酶水平与钙、肌酐均呈负相关(r分别是-0.741、-0.459,P<0.05);细胞实验中,①草酸组和尿激酶组细胞培养基中丙二醛与乳酸脱氢酶水平均明显高于对照组 (P<0.05);与尿激酶组相比,草酸组细胞细胞培养基中丙二醛及乳酸脱氢酶含量更高(P<0.05)。②各组细胞活力均有所下降,尿激酶组相对于草酸组下降幅度显著降低。③倒置显微镜下尿激酶组草酸钙晶体于细胞表面粘附程度明显轻于草酸组。 结论:尿激酶能够减轻肾小管上皮细胞的损伤,抑制草酸钙晶体的生长和聚集,从而抑制结石的形成。
Other AbstractObjective: To observe calcium oxalate crystal growth and aggregation in vitro with the urokinase, to provide experimental evidence of inhibition of urokinase to calcium oxalate stones; to test the serum level of urokinase of organism and discuss the inhibition of the urokinase to kidney stone; to study the effect of urokinase in MDCK cells damage induced by oxalate, to discuss the inhibitive mechanism of urokinase in renal calculus. To prove the inhibition of urokinase to the formation of stone and provide a new therapeutic direction for kidney stone. Methods: Firstly, Based on calcium oxalate crystal growth and aggregation, distilled water (Blank group), different concentrations of citrate (control group) and urokinase (experimental group) were prepared and added into stone solution, size and aggregation of oxalate crystal of the solutions under inverted microscope were observed to prove inhibition of urokinase in vitro. 27 cases of patients with calculus (stone group) and 24 healthy people (control group) were divided into two groups, the serum level of urokinase of all subjects was measured by enzyme-linked immuno-sorbent assay(ELISA), biochemical detection method test serum calcium level and renal function (serum creatinine), analysis correlation of urokinase and calcium ion or creatinine. MDCK cells were cultured respectively. On the fifth day of primary culture, cultured cells were exposed to oxalate (5 mM)for 2 hours with or without 100.000 units urokinase in the medium, Malonaldehyde(MDA) and lactic dehydrogenase(LDH)of the medium was determined; The activity of MDCK was also determined by CCK-8 at 0.5h, 1h, 1.5h, 2h; The cell surface adhesion of crystals was observed under inverted microscope. Results: crystal volume in blank group is significantly bigger and easier to mutual aggregation than control group and experimental group, calcium oxalate crystal growth and aggregation in control group and experimental group are heavyly suppressed; Serum level of urokinase with stone group is lower than normal group (μg/L:0.49±0.15vs1.44±0.55,P<0.05), calcium level higher than those in control group (mmol/L:2.30±0.18vs2.18±0.13,P<0.05), serum creatinine concentration (umol/L:(54.38,126.26)vs(46.53,83.24),P<0.05),Stone group of urokinase level change with calcium and creatinine level were negatively correlated (r=-0.741/-0.459 respectively,P<0.05). In cell experiments, MDA and LDH of oxalate and urokinase group were significantly higher than contro...
URL查看原文
Language中文
Document Type学位论文
Identifierhttps://ir.lzu.edu.cn/handle/262010/222914
Collection学院待认领
Affiliation临床医学院
Recommended Citation
GB/T 7714
明星. 尿激酶在肾结石形成过程中作用机制的研究[D]. 兰州. 兰州大学,2014.
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