微囊蛋白在小鼠胆囊胆固醇结石症形成中的作用
发布时间：2018-07-24 22:02
【摘要】：目的：研究微囊蛋白在致石饲料诱导的小鼠胆囊胆固醇结石症形成中的作用,为临床胆囊胆固醇结石症的诊疗提供新的思路。 方法：以结石易感小鼠C57BL/6小鼠为研究对象,分别给予普通饲料和含高脂高胆固醇致石饲料饲养4周。4%水合氯醛腹腔注射,待小鼠麻醉后,进行小鼠外科实验。依次剪开皮肤、皮下筋膜、腹膜,充分暴露肝胆组织,肉眼观察小鼠肝脏、胆囊外观及胆囊结石形成情况。结扎胆囊管及胆总管下端,在胆总管插入PE10号管1h留取胆汁-20℃保存。下腔静脉采血于含EDTA的生化管中,3000rpm4℃离心15min收集上层血清4℃保存。最后切取肝脏和胆囊组织,肝脏称重,肝胆组织保存于-80℃冰箱。全自动生化分析仪检测血脂及胆汁成分。RT-PCR及Westren blot分别检测小鼠肝脏及胆囊CAV1、CAV3、SR-B1、CCK-AR的基因和蛋白水平。 结果：致石饲料饲养4周后,实验组小鼠胆囊增大,胆囊内可见大量泥沙状胆固醇结石沉积,成石率为100%(6/6),而普通饲料饲养的对照组小鼠胆囊略小,均未发现有结石。实验组小鼠肝体重比显著增加(0.08±0.01vs0.05+0.01,P0.01),肝脏外观较灰暗、肿大,而普通饲料饮食的对照组小鼠肝脏色泽红润,体积较小实验组小鼠胆汁浑浊,胆固醇和磷脂含量较对照组明显升高(1.33±0.33vs0.21±0.11,P0.01；3.55±1.40vs1.55±0.63,P0.05),而总胆汁酸含量却显著下降(726.48±51.83vs839.83+23.75,P0.01)。实验组小鼠血清总胆固醇、高密度脂蛋白和低密度脂蛋白含量明显高于对照组(TC：4.22±0.46vs2.21±0.11,P0.01；HDL：1.86±0.10vs1.35±0.11,P0.01,LDL：2.18±0.44vs0.58±0.12,P0.01),甘油三酯、极低密度脂蛋白(水平未有明显改变(TG：0.75+0.04vs0.83+0.15,VLDL：0.18±0.12vs0.28±0.09)。与对照组小鼠相比,实验组小鼠肝脏和胆囊CAV1mRNA和蛋白表达均显著下降(肝脏CAV1基因和蛋白分别为0.53±±0.13vs1±±0.32,P0.01；0.39±0.07vs0.9±0.06,P0.01；胆囊CAV1基因和蛋白分别为0.44-0.22vs1±0.22,P0.01；1.04±0.07vs1.34±0.04,P0.01),胆囊CCKAR基因和蛋白水平亦明显降低(0.32±0.20vs1±0.25,P0.01；0.07±0.02vs0.35±0.04,P0.01),两组小鼠肝脏均未能检测到CCK-AR的表达。肝脏CAV3mRNA和蛋白表达增加(肝脏CAV3基因和蛋白分别为3.38±1.47vs1±0.17,P0.01；0.93±0.04vs0.5±±0.06,P0.01),胆囊CAV3mRNA水平改变没有统计学意义(0.87±0.27vs1±0.40),蛋白水平显著升高(0.25±0.03vs0.17±0.02,P0.05)；SR-B1基因和蛋白均未有明显改变(肝脏基因和蛋白分别为0.25±0.03vs0.17±0.02,0.9±0.04vs0.92±0.04；胆囊基因和蛋白分别为0.86±0.27vs1±0.35,0.89±0.03vs1.044±0.07)。 结论：含高脂高胆固醇高胆酸的致石餐作为一外界干预因素,能诱导小鼠血脂含量升高,胆道胆固醇高分泌,总胆汁酸合成减少,促使小鼠胆囊胆固醇结石形成。小鼠胆囊胆固醇结石产生伴随CAV1和CAV3在肝胆组织基因和蛋白表达水平发生变化,提示CAV1及CAV3可能与胆囊胆固醇结石的形成有关。SR-B1可能对C57BL/6小鼠胆囊胆固醇结石的形成无显著影响。CAV1在肝胆组织表达下降,可能造成胆固醇转运功能受损,脂质代谢紊乱,胆固醇在肝细胞内大量堆积。CAV3可能通过抑制胆囊CCKAR的表达,减弱胆囊平滑肌的收缩,胆囊动力低下,胆汁淤滞,促进胆固醇结晶析出,聚集以及结石形成。CAV1、CAV3有望成为临床胆囊胆固醇结石症治疗的药物靶标分子或/和诊断标志物。
[Abstract]:Objective: To study the role of microcystin in the formation of gallbladder cholesterol gallstone induced by stone feed in mice, and to provide a new idea for the diagnosis and treatment of cholelithiasis in clinical gallbladder.
Methods: the C57BL/6 mice were treated with common fodder and high cholesterol gallstone feed for 4 weeks. The mice were treated with chloral hydrate and chloral hydrate for 4 weeks. After the mice were anesthetized, the mice were treated with the skin, the subcutaneous fascia, the peritoneum, the liver and the hepatobiliary body, the liver and the outside of the gallbladder. The formation of cholecystolithiasis. Ligation of the cystic duct and the lower end of the common bile duct, the bile duct was inserted into the PE10 tube 1h to retain the bile -20 centigrade. The inferior vena cava was collected in a biochemical tube containing EDTA, 3000rpm4 centigrade centrifugation 15min to collect the upper serum at 4 C. Finally, the liver and gallbladder tissues were removed, the liver was weighed, and the hepatobiliary tissue was preserved at -80 centigrade refrigerator. .RT-PCR and Westren blot were detected by automatic biochemical analyzer to detect the gene and protein levels of CAV1, CAV3, SR-B1 and CCK-AR in liver and gallbladder of mice.
Results: after 4 weeks of feeding, the gallbladder in the experimental group was enlarged and a large number of silt like cholesterol stones were deposited in the gallbladder. The rate of stone formation was 100% (6/6), while the gallbladder in the control group of the control group of the normal feed was slightly smaller and no stones were found. The liver weight of the mice in the experimental group was increased (0.08 + 0.01vs0.05+0.01, P0.01), and the appearance of the liver was more gray. The liver color of the control group of the control group of the normal diet diet was red, and the bile was cloudy in the small experimental group and the content of cholesterol and phospholipid was significantly higher than that of the control group (1.33 + 0.33vs0.21 + 0.11, P0.01; 3.55 + 1.40vs1.55 + 0.63, P0.05), but the total bile acid content decreased significantly (726.48 + 51.83vs839.83+23.75, P0.01). The content of serum total cholesterol, high density lipoprotein and low density lipoprotein in the group of mice was significantly higher than that in the control group (TC:4.22 + 0.46vs2.21 + 0.11, P0.01; HDL:1.86 + 0.10vs1.35 + 0.11, P0.01, LDL:2.18 + 0.44vs0.58 + 0.12, P0.01), triglyceride and extremely low density lipoprotein (TG:0.75+0.04vs0.83+0.15, VLDL:0.18) 0.12vs0.28 + 0.09). Compared with the control group, the expression of CAV1mRNA and protein in the liver and gallbladder of the experimental group decreased significantly (the liver CAV1 gene and protein were 0.53 + + + 0.13vs1 + 0.32, P0.01, 0.39 + 0.07vs0.9 + 0.06, P0.01, and CAV1 gene and protein of the gallbladder were 0.44-0.22vs1 + 0.22, P0.01; 1.04 + 0.07vs1.34 + 0.04, P0.01). The CCKAR gene and protein level of gallbladder also decreased significantly (0.32 + 0.20vs1 + 0.25, P0.01, 0.07 + 0.02vs0.35 + 0.04, P0.01). The expression of CCK-AR was not detected in the two groups of mice, and the expression of CAV3mRNA and protein in the liver was increased (the liver CAV3 gene and protein were 3.38 + 1.47vs1 + 0.17, P0.01; 0.93 + 0.04vs0.5 + + 0.06, P0.01). The level of mRNA was not statistically significant (0.87 + 0.27vs1 + 0.40), the protein level increased significantly (0.25 + 0.03vs0.17 + 0.02, P0.05), and there was no significant change in the SR-B1 gene and protein (the liver gene and protein were 0.25 + 0.03vs0.17 + 0.02,0.9 + 0.04vs0.92 + 0.04, respectively, and the gene and protein of the gallbladder were 0.86 + 0.27vs1 + 0.35,0.89 + 0.03vs1., respectively. 044 + 0.07).
Conclusion: the stony meal containing high cholesterol and hypercholesterolic hypercholic acid can induce the increase of blood lipid content, the hypersecretion of cholesterol in the bile duct and the decrease of total bile acid synthesis in mice. The cholesterol gallstones of gallbladder in mice are associated with the gene and protein expression levels of CAV1 and CAV3 in the liver and gallbladder tissues. The results suggest that CAV1 and CAV3 may be related to the formation of gallbladder cholesterol stones..SR-B1 may have no significant effect on the formation of cholesterol gallstones in the gallbladder of C57BL/6 mice..CAV1 may decrease in the expression of.CAV1 in the liver and gallbladder tissue, which may cause impaired cholesterol transport function, lipid metabolism disorder, and a large accumulation of.CAV3 in the liver cells. The expression of CCKAR can weaken the contraction of the smooth muscle of the gallbladder, low gallbladder motility, cholestasis, promote the crystallization of cholesterol, accumulate and form.CAV1. CAV3 is expected to be a drug target molecule or / and diagnostic marker for the treatment of cholelithiasis in the gallbladder.
【学位授予单位】：浙江大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R575.6
本文编号：2142788
[Abstract]:Objective: To study the role of microcystin in the formation of gallbladder cholesterol gallstone induced by stone feed in mice, and to provide a new idea for the diagnosis and treatment of cholelithiasis in clinical gallbladder.
Methods: the C57BL/6 mice were treated with common fodder and high cholesterol gallstone feed for 4 weeks. The mice were treated with chloral hydrate and chloral hydrate for 4 weeks. After the mice were anesthetized, the mice were treated with the skin, the subcutaneous fascia, the peritoneum, the liver and the hepatobiliary body, the liver and the outside of the gallbladder. The formation of cholecystolithiasis. Ligation of the cystic duct and the lower end of the common bile duct, the bile duct was inserted into the PE10 tube 1h to retain the bile -20 centigrade. The inferior vena cava was collected in a biochemical tube containing EDTA, 3000rpm4 centigrade centrifugation 15min to collect the upper serum at 4 C. Finally, the liver and gallbladder tissues were removed, the liver was weighed, and the hepatobiliary tissue was preserved at -80 centigrade refrigerator. .RT-PCR and Westren blot were detected by automatic biochemical analyzer to detect the gene and protein levels of CAV1, CAV3, SR-B1 and CCK-AR in liver and gallbladder of mice.
Results: after 4 weeks of feeding, the gallbladder in the experimental group was enlarged and a large number of silt like cholesterol stones were deposited in the gallbladder. The rate of stone formation was 100% (6/6), while the gallbladder in the control group of the control group of the normal feed was slightly smaller and no stones were found. The liver weight of the mice in the experimental group was increased (0.08 + 0.01vs0.05+0.01, P0.01), and the appearance of the liver was more gray. The liver color of the control group of the control group of the normal diet diet was red, and the bile was cloudy in the small experimental group and the content of cholesterol and phospholipid was significantly higher than that of the control group (1.33 + 0.33vs0.21 + 0.11, P0.01; 3.55 + 1.40vs1.55 + 0.63, P0.05), but the total bile acid content decreased significantly (726.48 + 51.83vs839.83+23.75, P0.01). The content of serum total cholesterol, high density lipoprotein and low density lipoprotein in the group of mice was significantly higher than that in the control group (TC:4.22 + 0.46vs2.21 + 0.11, P0.01; HDL:1.86 + 0.10vs1.35 + 0.11, P0.01, LDL:2.18 + 0.44vs0.58 + 0.12, P0.01), triglyceride and extremely low density lipoprotein (TG:0.75+0.04vs0.83+0.15, VLDL:0.18) 0.12vs0.28 + 0.09). Compared with the control group, the expression of CAV1mRNA and protein in the liver and gallbladder of the experimental group decreased significantly (the liver CAV1 gene and protein were 0.53 + + + 0.13vs1 + 0.32, P0.01, 0.39 + 0.07vs0.9 + 0.06, P0.01, and CAV1 gene and protein of the gallbladder were 0.44-0.22vs1 + 0.22, P0.01; 1.04 + 0.07vs1.34 + 0.04, P0.01). The CCKAR gene and protein level of gallbladder also decreased significantly (0.32 + 0.20vs1 + 0.25, P0.01, 0.07 + 0.02vs0.35 + 0.04, P0.01). The expression of CCK-AR was not detected in the two groups of mice, and the expression of CAV3mRNA and protein in the liver was increased (the liver CAV3 gene and protein were 3.38 + 1.47vs1 + 0.17, P0.01; 0.93 + 0.04vs0.5 + + 0.06, P0.01). The level of mRNA was not statistically significant (0.87 + 0.27vs1 + 0.40), the protein level increased significantly (0.25 + 0.03vs0.17 + 0.02, P0.05), and there was no significant change in the SR-B1 gene and protein (the liver gene and protein were 0.25 + 0.03vs0.17 + 0.02,0.9 + 0.04vs0.92 + 0.04, respectively, and the gene and protein of the gallbladder were 0.86 + 0.27vs1 + 0.35,0.89 + 0.03vs1., respectively. 044 + 0.07).
Conclusion: the stony meal containing high cholesterol and hypercholesterolic hypercholic acid can induce the increase of blood lipid content, the hypersecretion of cholesterol in the bile duct and the decrease of total bile acid synthesis in mice. The cholesterol gallstones of gallbladder in mice are associated with the gene and protein expression levels of CAV1 and CAV3 in the liver and gallbladder tissues. The results suggest that CAV1 and CAV3 may be related to the formation of gallbladder cholesterol stones..SR-B1 may have no significant effect on the formation of cholesterol gallstones in the gallbladder of C57BL/6 mice..CAV1 may decrease in the expression of.CAV1 in the liver and gallbladder tissue, which may cause impaired cholesterol transport function, lipid metabolism disorder, and a large accumulation of.CAV3 in the liver cells. The expression of CCKAR can weaken the contraction of the smooth muscle of the gallbladder, low gallbladder motility, cholestasis, promote the crystallization of cholesterol, accumulate and form.CAV1. CAV3 is expected to be a drug target molecule or / and diagnostic marker for the treatment of cholelithiasis in the gallbladder.
【学位授予单位】：浙江大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R575.6
【相似文献】
相关期刊论文 前10条
1 梁旭方,黄芬;微囊蛋白基因及其与疾病关系研究进展[J];生物化学与生物物理进展;2003年03期
2 邸辉;焦保华;;微囊蛋白1的生物学功能及在恶性肿瘤中的作用[J];临床荟萃;2010年06期
3 陈文勤;席晓薇;;细胞质膜微囊蛋白-1在恶性肿瘤中的研究进展[J];现代妇产科进展;2011年10期
4 徐炜;顾栋桦;平金良;姚根有;;细胞质膜微囊蛋白-1在皮肤恶性黑色素瘤中的表达及临床意义[J];浙江实用医学;2010年04期
5 崔海宏;杜静平;韩英;;微囊蛋白-1的功能及其在结肠癌中的作用[J];国际消化病杂志;2009年01期
6 姚康,徐标;细胞质膜微囊-微囊蛋白及其相关信号分子[J];中国病理生理杂志;2001年04期
7 邸辉;焦保华;;Caveolae、Caveolin-1的功能及在肿瘤中的作用研究进展[J];中国医疗前沿;2009年22期
8 周立君;刘晶;;同型半胱氨酸对脐静脉内皮细胞微囊蛋白-1的蛋白及其mRNA表达的影响[J];中国应用生理学杂志;2009年03期
9 刘柏炎;俞悦;易健;陈雪梅;蔡光先;;细胞质膜微囊蛋白1敲除小鼠神经干细胞培养与生物学特性[J];中国组织工程研究;2014年23期
10 曾潍贤;戴元荣;;微囊蛋白-1蛋白参与哮喘气道平滑肌细胞增殖中ERK1/2调控以及罗红霉素的干预[J];中国临床药理学与治疗学;2014年06期
相关硕士学位论文 前1条
1 刘姗;微囊蛋白在小鼠胆囊胆固醇结石症形成中的作用[D];浙江大学;2014年
，本文编号：2142788
本文链接：https://www.wllwen.com/yixuelunwen/mazuiyixuelunwen/2142788.html
最近更新
教材专著
