Apobec-1重组腺病毒治疗肾性高脂血症的实验研究

发布时间：2018-06-19 23:40

本文选题：阿霉素肾病兔 + 蛋白尿　；参考：《安徽医科大学》2014年硕士论文

【摘要】：研究目的观察单侧肾切除阿霉素肾病兔ApoB mRNA编辑效率及其血、尿液生化和肾脏病理学改变。研究资料与方法 40只雄性6月龄新西兰兔，体重1000±100g，随机分为病例组（n=20只）、假手术组（n=20只）。①病例组：以3%的戊巴比妥30mg/Kg行腹腔注射麻醉，无菌条件下行左肾切除，1w后耳静脉注射阿霉素5mg/Kg，术后12周处死所有兔子留取肾脏及肝脏标本，滤纸吸干后称重，立即用10%中性福尔马林溶液固定，石蜡包埋；②假手术组：以3%的戊巴比妥30mg/Kg行腹腔注射麻醉，只探及肾包膜不切除左肾，1w后耳静脉注射生理盐水5mg/Kg，术后12周处死所有兔子留取肾脏标本，滤纸吸干后称重，立即用10%中性福尔马林溶液固定，石蜡包埋。另分别于试验前（0周）和试验结束（12周）检测血清肌酐(Scr)、尿素氮(BUN)、总蛋白(TP)、白蛋白(Alb)、24h尿蛋白(24h UPr)及载脂蛋白B-100、载脂蛋白B-48。酶法测定Scr、BUN、TP、Alb；放射免疫法检测血浆ApoB-100、ApoB-48；双缩脲比色法检测24小时尿蛋白浓度；苏木素一伊红(HE)染色观察肾脏病理改变；免疫组化观测肝脏ApoB-100及ApoB-48的组织表达。研究结果 ①病例组24h UPr和血清Scr、BUN、ApoB-100和ApoB-48显著高于假手术组，而血清TP，Alb则明显降低。 ②病例组肾脏肾脏肾上皮细胞肿胀，系膜细胞增生、系膜基质增多；肾间质增宽，，可见大量炎症细胞浸润。ApoB-100和ApoB-48以肾小球表达增高为主。研究结论单侧肾切除加1周后耳静脉注射阿霉素(5mg／kg)在术后12周可以成功建立肾病模型，具体表现为大量蛋白尿、低蛋白血症和明显的高脂血症，肾脏病理显示局灶性病变及肝脏组织ApoB-100和ApoB-48表达普遍增高。研究目的观察外源性Apobec-1基因导入新西兰兔体内后，肝脏apoB mRNA编辑效率及其对血脂代谢的影响。研究资料与方法 60只雄性6月龄新西兰兔，体重1000±100g，随机分为apobec-1腺病毒感染病例组（n=20）、假干预组（n=20)，假手术组（n=20)，置于代谢笼中饲养。①病例组：以3%的戊巴比妥30mg/Kg行腹腔注射麻醉，无菌条件下行左肾切除，1w后耳静脉注射阿霉素5mg/Kg，术后11周处死每组的半数兔子留取肝脏标本，其余兔子分别通过耳缘静脉注入Apobec-1重组腺病毒，饲养1周，摘除右侧肾脏和肝脏，滤纸吸干后称重，立即用10%中性福尔马林溶液固定，石蜡包埋；②假干预组：以3%的戊巴比妥30mg/Kg行腹腔注射麻醉，无菌条件下行左肾切除，1w后耳静脉注射阿霉素5mg/Kg，术后11周处死每组的半数兔子留取肝脏标本，其余兔子分别通过耳缘静脉注入空白腺病毒，饲养1周，摘除右侧肾脏和肝脏，滤纸吸干后称重，立即用10%中性福尔马林溶液固定，石蜡包埋；③假手术组：以3%的戊巴比妥30mg/Kg行腹腔注射麻醉，只探及肾包膜不切除左肾，1w后耳静脉注射生理盐水5mg/Kg，术后11周处死每组的半数兔子留取肝脏标本，其余兔子分别通过耳缘静脉注入Apobec-1重组腺病毒，饲养1周，摘除右侧肾脏和肝脏，滤纸吸干后称重，立即用10%中性福尔马林溶液固定，石蜡包埋。分别留取处死前后静脉血及24h尿液。研究结果 ①用药前病例组24h UPr、ApoB-100和ApoB-48、TG及TCH、VLDL、LDL-C显著高于假手术组，而与假干预组相差不大，而用药后24小时病例组尿蛋白较前有所缓解，ApoB-100、TG及TCH、VLDL、LDL-C显著下降，TP明显升高，ApoB-48上升不明显，假干预组及假手术组用药后改变与用药前差别不大。②导入重组腺病毒前后肝脏病理未发现有畸形改变；③Western Blot均显示病例组肝脏组织显著表达Apobec-1和ApoB-48。研究结论 Apobec-1重组腺病毒导入可在一定程度上起到降血脂和肾脏保护效应，尚未发现明显肝脏致畸改变。
[Abstract]:research objective
Objective To observe the editing efficiency of ApoB mRNA in unilateral nephrectomy rabbits with adriamycin nephropathy and their changes in blood, urine biochemistry and renal pathology.
Research materials and methods
40 male 6 month old New Zealand rabbits, weighing 1000 + 100g, were randomly divided into case group (n=20) and sham operation group (n=20 only). (1) case group: intraperitoneal injection of 3% pentobarbital 30mg/Kg, left nephrectomy under aseptic conditions, and adriamycin 5mg/Kg after 1W, all rabbits were sacrificed for kidney and liver specimens for 12 weeks after 12 weeks of operation and filter paper sucked. After dry weighing, immediately fixed with 10% neutral formalin solution, paraffin embedded; sham operation group: intraperitoneal injection of 3% pentobarbital 30mg/Kg, only the renal capsule did not remove the left kidney, 1W posterior auricular vein injection of saline 5mg/Kg, 12 weeks after the operation, all rabbits were left to take the kidney specimen, filter paper was weighed after sucking dry, immediately use 10% neutrality Faure Marin solution was fixed and paraffin was embedded. The serum creatinine (Scr), urea nitrogen (BUN), total protein (TP), albumin (Alb), 24h urine protein (24h UPr) and apolipoprotein B-100 were detected before the test (0 weeks) and the end of the experiment (12 weeks). The apolipoprotein B-48. enzyme method was used to determine Scr, BUN, TP, and arsenic. The urine protein concentration of 24 hours was measured by colorimetric method. The pathological changes of kidney were observed by hematoxylin and eosin (HE) staining, and the tissue expression of liver ApoB-100 and ApoB-48 was observed by immunohistochemistry.
Research results
(1) 24h UPr and serum Scr, BUN, ApoB-100 and ApoB-48 in case group were significantly higher than those in sham operation group, while serum TP and Alb decreased significantly.
In case group, renal renal epithelial cells were swollen, mesangial cells proliferated, mesangial matrix increased, renal interstitium widened, and a large number of inflammatory cells infiltrated.ApoB-100 and ApoB-48 with increased glomerular expression.
research conclusion
A nephrotic model could be successfully established at 12 weeks after unilateral nephrectomy plus adriamycin injection (5mg / kg) after 1 weeks. The specific manifestations were a large number of proteinuria, hypoproteinemia and obvious hyperlipidemia. The renal pathological manifestations of focal lesions and liver tissue ApoB-100 and ApoB-48 were generally increased.
research objective
To observe the apoB mRNA editing efficiency and the effect on lipid metabolism in the liver of New Zealand rabbits after the introduction of exogenous Apobec-1 gene.
Research materials and methods
60 male 6 month old New Zealand rabbits, weighing 1000 + 100g, were randomly divided into apobec-1 adenovirus infection case group (n=20), false intervention group (n=20), sham operation group (n=20) and kept in metabolic cage. (1) case group: intraperitoneal injection of 3% pentobarbital 30mg/Kg, left nephrectomy under no bacteria condition, and adriamycin 5mg/Kg after 1W, after 1W. Half of the rabbits in each group were sacrificed for 11 weeks to leave the liver specimens. The rest of the rabbits were injected with Apobec-1 recombinant adenovirus through the auricular vein. The right kidney and liver were removed for 1 weeks. The filter paper was removed and weighed after sucking the filter paper. 10% neutral Faure Marin solution was fixed and paraffin embedded. 2. The false intervention group was injected with 3% pentobarbital 30mg/Kg. Left nephrectomy under sterile conditions and adriamycin 5mg/Kg were injected into the auricular vein after 1W. Half of the rabbits in each group were sacrificed 11 weeks after the operation to leave the liver specimens. The rest of the rabbits were injected with the blank adenovirus through the ear vein. The right kidney and liver were removed for 1 weeks. The filter paper was weighed after sucking the filter paper. 10% neutral formalin solution was immediately fixed and paraffin paraffin was used. Buried; (3) sham operation group: intraperitoneal injection of 3% pentobarbital 30mg/Kg, only the renal capsule did not remove the left kidney, 1W posterior auricular vein injection of saline 5mg/Kg, 11 weeks after the operation, half of the rabbits in each group were sacrificed to leave the liver specimens, and the rest of the rabbits were injected with the Apobec-1 recombinant adenovirus through the ear vein, and the right kidney was removed for 1 weeks. After weighing with the liver, the filter paper was dried and weighed immediately. 10% neutral formalin solution was fixed and paraffin embedded. The venous blood and 24h urine were collected before and after the death.
Research results
(1) 24h UPr, ApoB-100 and ApoB-48, TG and TCH, VLDL, and LDL-C were significantly higher than those in the sham operation group, but the urine protein in the case group was significantly different from that in the sham intervention group, while the 24 hours after the drug use was relieved, ApoB-100, TG and TCH, VLDL, increased significantly. There was not much difference between the changes before and before the drug use. (2) there was no abnormal change in the liver pathology before and after the introduction of the recombinant adenovirus; (3) Western Blot showed that the liver tissues of the case group showed significant expression of Apobec-1 and ApoB-48.
research conclusion
Apobec-1 recombinant adenovirus can play a role in lowering blood lipid and renal protection to some extent, and has not found obvious liver teratogenic changes.
【学位授予单位】：安徽医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R699.2

【参考文献】