加味胃苓汤治疗肝硬化腹水的应用基础研究

发布时间：2018-06-02 21:40

本文选题：加味胃苓汤 + 中医药　；参考：《湖北中医药大学》2016年博士论文

【摘要】：目的:肝硬化腹水是临床上常见病及难治病之一,属于祖国医学“积聚”、“胁痛”、“臌胀”等范畴,临床上现代医学多以对症支持治疗为主,疗效一般。加味胃苓汤是门九章教授多年来根据肝病特点及临床实践自拟的治疗肝硬化腹水的有效经验方。本课题即主要通过临床观察和动物实验来进一步验证加味胃苓汤的疗效,为其治疗肝硬化腹水的有效性和科学性提供相关的实验数据,为加味胃苓汤的开发应用打下坚实的理论基础,为肝硬化腹水的临床治疗提供有益的参考。方法:研究分为实验研究和临床研究,包括三个部分,第一部分运用二甲基亚硝胺(DMN)建立肝硬化腹水的大鼠动物模型。选用雄性SD大鼠100只,重量在200~250g之间,随机选取10只作为正常对照组,其余90只作为模型组,用来制作肝硬化腹水模型。在熟悉环境一周后,每次给大鼠腹腔注射1%二甲基亚硝胺(DMN)1ml/kg,每周三天,每天一次,共12周。造模期间,密切关注大鼠体重,每次注射前称体重、测腹围,并与上一次注射前体重、腹围进行对比,体重明显下降,甚者下降率在30%以上,暂停腹腔注射。在第12周末进行腹腔穿刺观察有无腹水。为进一步判定造模是否成功,对穿刺有腹水的大鼠和正常对照组大鼠进行解剖以及病理、肝肾功能等方面的比较。第二部分在第一部分造模成功的基础上与其他药物比较,探讨加味胃苓汤对肝硬化腹水大鼠模型的治疗作用及机理研究。将第一部分获得的肝硬化腹水大鼠模型随机分成加味胃苓汤高剂量组(A组)、加味胃苓汤低剂量组(B组)、模型组(C组)、速尿+安体舒通组(D组)、扶正化瘀组(E组),每组各8只大鼠。在代谢笼内饲养大鼠24小时,留取24小时尿液,计算24小时净出水量后开始给药,除模型组给予等容量的生理盐水灌胃外,加味胃苓汤高剂量组、加味胃苓汤低剂量组给予加味胃苓汤,剂量分别为42.7g/kg体重、6.1g/kg体重,扶正化瘀组给予扶正化瘀胶囊0.075g/kg体重灌服,速尿+安体舒通组给予速尿和安体舒通0.66mg/kg体重灌服。以上各组灌胃液体量均为2ml,每日给药1次,其浓度及等效剂量按体重折算,共给药3周。观察大鼠的一般情况及体重变化,在灌胃的最后一天再次将实验大鼠饲养于代谢笼内,再次收集24小时尿液,计算24小时净出水量。测定门静脉压力并检测大鼠肝功能指标及NO、ET-1的含量和肝纤维化标志物四项。第三部分观察加味胃苓汤治疗肝硬化腹水的临床疗效,选择符合肝硬化腹水诊断标准的患者60例,随机将其分为治疗组和对照组。对照组30例,采用常规西医对症治疗;治疗组30例,在常规西医对症治疗的基础上配合加味胃苓汤,每日1剂,治疗一个月后。观察两组患者治疗前后的主要症状、体征(包括乏力、纳差、腹胀、尿少、黄疸、双下肢浮肿)、腹围、体重以及肝功能指标、腹部B超检查指标(包括脾厚、门静脉及脾静脉宽度)及其有无并发症等,同时监测24h尿量变化。结果:1 12周后,模型组大鼠有51只存活。经腹腔穿刺其中48只有腹水,相对成模率为94.1%。解剖可见造模组的肝脏肿大明显,表面不平;病理组织学检查提示肝硬化以及假小叶形成;门静脉压力(Ppv)较正常对照组明显升高(p0.01);总胆红素(TBIL)、丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转移酶(AST)较正常对照组明显升高(p0.01);白蛋白(ALB)较正常对照组明显降低(p0.05)。2大鼠的一般情况:正常大鼠在整个实验期间,体重逐渐增加,毛色光滑润泽,无异常行为表现。模型组动物体重增加缓慢,皮毛光泽度不高,精神倦怠不振,摄食减少,表现为易激惹。各组治疗开始后,大鼠进食、体重均有所增加,毛色较造模期间光润,能不同程度地减轻造模药物对大鼠的影响。其中,加味胃苓汤高剂量组、加味胃苓汤低剂量组较其它组最显著的变化就是治疗后大便由造模期间的稀溏便转为正常大便。2.1灌胃治疗后,经解剖,加味胃苓汤高剂量组、加味胃苓汤低剂量组大鼠腹水明显减少,色淡黄,模型组可见较多腹水,速尿+安体舒通组腹水微量,扶正化瘀组有少量腹水。加味胃苓汤高剂量组、加味胃苓汤低剂量组与模型组比较,差异具有统计学意义(p0.05),速尿+安体舒通组与模型组比较,差异具有显著统计学意义(p0.01),而扶正化瘀组与模型组比较,差异不具有统计学意义(p0.05);加味胃苓汤高剂量组、加味胃苓汤低剂量组与速尿+安体舒通组比较,差异不具有统计学意义(p0.05);与扶正化瘀组比较,差异具有统计学意义(p0.05)。2.2加味胃苓汤高剂量组、加味胃苓汤低剂量组与速尿+安体舒通组治疗后较治疗前24小时净出水量明显增加,差异具有显著统计学意义(p0.01);扶正化瘀组治疗后较治疗前24小时净出水量有所增加,前后比较差异不具有统计学意义(p0.05)。2.3各治疗组门静脉压力均有不同程度的降低,与模型组比较差异具有统计学意义(p0.05),其中加味胃苓汤高剂量组、加味胃苓汤低剂量组与速尿+安体舒通组比较,差异具有统计学意义(p0.05);加味胃苓汤高剂量组与扶正化瘀组比较,差异具有统计学意义(p0.05);加味胃苓汤低剂量组与扶正化瘀组比较,差异不具有统计学意义(p0.05)。表明加味胃苓汤可以降低大鼠肝硬化腹水门静脉压力,其中以加味胃苓汤大剂量组效果最好。2.4加味胃苓汤能明显减轻大鼠肝组织炎症与纤维增生的程度,其中尤以加味胃苓汤高剂量组改善最为明显,其作用与扶正化瘀胶囊组相当(p0.05)。2.5治疗后,加味胃苓汤高剂量组、加味胃苓汤低剂量组与扶正化瘀组均能不同程度的降低血清丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转移酶(AST)、球蛋白(GLB)和升高白蛋白(ALB)的水平,与模型组比较差异具有显著统计学意义(p0.01);与速尿+安体舒通组比较差异具有统计学意义(p0.05);加味胃苓汤高剂量组、加味胃苓汤低剂量组各指标分别与扶正化瘀组比较,差异不具有统计学意义(p0.05)。2.6治疗后,各治疗组大鼠NO及ET-1含量均有不同程度的降低,其中加味胃苓汤高剂量组、加味胃苓汤低剂量组与模型组比较差异有统计学意义(p0.05),速尿+安体舒通组、扶正化瘀组与模型组比较差异无统计学意义(p0.05);加味胃苓汤高剂量组、加味胃苓汤低剂量组与速尿+安体舒通组比较,差异具有统计学意义(p0.05);加味胃苓汤高剂量组、加味胃苓汤低剂量组与扶正化瘀组比较,差异具有统计学意义(p0.05)。2.7治疗后,加味胃苓汤高剂量组、加味胃苓汤低剂量组、扶正化瘀组透明质酸(HA)、层粘连蛋白(LN)、Ⅲ型前胶原(PCⅢ)及Ⅳ型胶原(CIV)均显著降低,与模型组比较差异具有统计学意义(P0.05或P0.01);速尿+安体舒通组与模型组比较无显著性差异(p0.05);与扶正化瘀组比较,加味胃苓汤高剂量组、加味胃苓汤低剂量组无显著性差异(P0.05)。与速尿+安体舒通组比较,加味胃苓汤高剂量组、加味胃苓汤低剂量组具有显著性差异(p0.01)。3临床综合疗效及中医证候疗效:治疗1个月临床综合疗效比较,治疗组显效率与总有效率分别为23.33%和93.33%,对照组分别为13.33%和70.0%,两组比较差异有统计学意义(P0.05)。3.1两组患者治疗前后主要症状、体征改善情况比较,治疗组在治疗后主要症状、及体征改善明显,与治疗前比较差异有显著统计学意义(P0.05);对照组患者治疗后主要症状及体征也有改善,与治疗前比较差异无统计学意义(P0.05)。3.2两组患者肝功能、凝血酶原活动度改善比较,治疗组治疗后ALB、PTA、A/G水平明显升高,与治疗前比较差异有统计学意义(P0.05),对照组ALB、PTA、A/G水平也有升高,但与治疗前比较,差异无统计学意义(P0.05);两组患者AST、ALT、TBIL水平治疗后比较,差异无统计学意义(P0.05),ALB、PTA、A/G水平治疗后比较,差异有统计学意义(P0.05)。3.3两组患者体重、腹围、平均24h尿量比较,两组患者治疗后体重、腹围与治疗前比较降低;与治疗前比较,平均24h尿量明显增多,差异有统计学意义(P0.05);对照组治疗前后体重、腹围差异无统计学意义(P0.05),平均24h尿量治疗前后差异有统计学意义。两组在治疗后体重、腹围、平均24h尿量比较差异有统计学意义(P0.05)。3.4两组患者腹部B超比较,两组患者在治疗后腹部B超指标(包括脾脏厚度、门静脉及脾静脉宽度)较治疗前有改善,差异有统计学意义(P0.05);对照组脾脏厚度、脾静脉宽度改善与治疗前比不如治疗组明显。两组患者治疗后B超指标(包括脾脏厚度、门静脉及脾静脉宽度)比较差异无统计学意义(P0.05)。3.5两组患者治疗后B超腹水量均有所减少,治疗组与治疗前比较差异具有统计学意义(P0.05);对照组治疗后与治疗前比较差异不具有统计学意义;两组治疗后比较,差异具有统计学意义(P0.05),说明治疗组消腹水的作用优于对照组。3.6两组患者治疗中并发症的发生率分别是13.33%和23.33%,并发症发生率比较差异无统计学意义(P0.05)。3.7两组患者在治疗后血、尿、便常规、一般体格检查、肾功能、心电图等指标与治疗前比较无明显异常变化;在整个治疗过程及治疗结束后未发生不良反应,提示加味胃苓汤治疗肝硬化腹水安全有效,并且无明显毒副作用。结论:1运用二甲基亚硝胺成功建立肝硬化腹水的大鼠模型,为进一步研究作准备。2加味胃苓汤可明显改变模型大鼠的一般状况,提高模型大鼠的活力。并具有明显的消腹水的作用。3加味胃苓汤能明显增加模型大鼠24小时净出水量,有较强的利水作用。4加味胃苓汤能够降低模型大鼠门静脉压力。5加味胃苓汤能够抑制胶原纤维增生、减轻肝组织炎症,提示加味胃苓汤有抗纤维化、保肝作用。6加味胃苓汤能够降低模型大鼠NO及ET-1含量,从而能够调整对肝脏血流的影响,改善门静脉血流和阻力的影响,这可能是加味胃苓汤能够降低模型大鼠门静脉压力、治疗腹水的作用机制之一。7加味胃苓汤治疗肝硬化腹水疗效肯定,能够明显改善患者的症状、体征、以及肝功能等。优于单纯的西医对照组,并且能够减少各种并发症的发生,减轻了患者的痛苦,提高了患者的生活质量,同时无不良反应发生,为我们治疗肝硬化腹水提供了很好的思路。8加味胃苓汤配合西医对症治疗的中西医结合组在提高患者疗效,缓解病人的临床症状,改善其肝功能方面优于单纯的西医对症治疗组;它可以改善患者临床症状,还可以改善患者肝功能的水平:ALT、AST、ALB、TB、PTA,能够稳定血浆白蛋白水平,改善凝血功能,减轻患者经济压力,减少出血等并发症,可以给患者获得更好的远期临床疗效。
[Abstract]:Objective: cirrhotic ascites is one of the common and refractory diseases in the clinic. It belongs to the category of "accumulation", "hypochondriac pain", "distension" and so on. In clinical modern medicine, most of the clinical modern medicine is mainly with symptomatic support treatment, and the curative effect is general. The nine chapters of Jiawei Ling Ling decoction are the treatment of liver cirrhosis ascites for many years according to the characteristics of liver disease and clinical practice. This topic is to further verify the curative effect of Jiawei Ling Ling soup through clinical observation and animal experiment, and provide the relevant experimental data for the effectiveness and science of the treatment of liver cirrhosis ascites, and lay a solid theoretical foundation for the development and application of the Jiawei Luling soup, and provide beneficial treatment for the clinical treatment of liver cirrhosis ascites. Methods: the study was divided into experimental and clinical studies, including three parts. The first part used two methylnitrosamine (DMN) to establish the rat model of cirrhotic ascites. 100 male SD rats were selected and 10 were randomly selected as the normal control group, and the other 90 were used as model groups to make liver cirrhosis. After one week of familiar environment, 1% two methylnitrosamine (DMN) 1ml/kg was injected into the abdominal cavity of rats, each Wednesday, once a day, for a total of 12 weeks. During the modeling period, the weight of rats was closely concerned, the weight was measured before each injection, the abdominal circumference was measured, and the weight of the previous injection was compared with the abdominal circumference, the weight decreased significantly and the decline rate was 30%. In order to further determine whether the model was successful, the second part of the first part of the model was compared with other drugs on the basis of the success of the first part of the model and the liver and kidney function. The effect and mechanism of modified Wei Ling Decoction on the rat model of cirrhotic ascites. The first part of the liver cirrhosis ascites rat model was randomly divided into the high dose group (group A) of Jiawei Ling Ling Decoction (group B), the model group (group C), furosemide + ansasong group (group D), the group of Fuzheng Huayu (group E), 8 rats in each group. The rats were kept in the metabolic cage for 24 hours, leaving the urine for 24 hours and calculating the net water content for 24 hours. In addition to the model group, the high dose group of Wei Ling soup was added to the model group and the low dose group of Wei Ling decoction was given with the dosage of 42.7g/kg weight, 6.1g/kg weight, and the group of Fuzheng Huayu group were given the centralizability. Blood stasis capsule 0.075g/kg body weight perfusion, furosemide + anspironolactone group to give furosemide and anacontan 0.66mg/kg weight perfusion. The volume of gastric perfusion in all groups is 2ml, 1 times a day, the concentration and equivalent dose of weight conversion, a total of 3 weeks. Observe the general situation and body weight changes in rats, the last day in the gavage of rats will be experimental rats again. In the metabolic cage, the urine was collected for 24 hours, and the net water content of 24 hours was calculated. The portal pressure was measured and the liver function index of the rats and the content of NO, ET-1 and four markers of liver fibrosis were detected. The third part observed the clinical efficacy of Jiawei lling Decoction in the treatment of cirrhotic ascites, and chose 60 patients with the diagnostic criteria of cirrhosis ascites. The control group was randomly divided into the treatment group and the control group. 30 cases in the control group were treated with conventional western medicine, and 30 cases in the treatment group were combined with Jiawei Wei Ling Decoction on the basis of conventional western medicine treatment, 1 doses a day, and one month after treatment. The main symptoms and signs of the two groups before and after treatment were observed (including fatigue, difference, abdominal distension, urine, jaundice, jaundice, and lower limbs). Edema), abdominal circumference, weight and liver function index, abdominal B-ultrasound examination index (including splenic thickness, portal vein and splenic vein width) and its complications, and monitoring 24h urine volume. Results: after 112 weeks, 51 rats in the model group were alive. 48 of them had ascites through abdominal puncture, and the relative model rate was 94.1%., the liver was dissected. Histopathological examination suggested liver cirrhosis and false lobule formation; portal vein pressure (Ppv) was significantly higher than normal control group (P0.01); total bilirubin (TBIL), alanine aminotransferase (ALT), aspartate aminotransferase (AST) was significantly higher than normal control group (P0.01); albumin (ALB) was more obvious than normal control group. The general condition of reducing (P0.05).2 rats: the body weight increased gradually during the whole experiment, the hair color was smooth and moist, and there was no abnormal behavior. The weight of the model group increased slowly, the skin glossiness was not high, the mental burnout was not vibrant, the feeding reduced, and the irritability. After the treatment of each group, the rats fed, the weight of the rat increased, Mao increased, Mao increased. It can reduce the effect of the light in the model during the process of modeling. Among them, the high dose group of Jiawei Ling Ling decoction, the most significant change of the low dose group of Jiawei Ling Ling Decoction and the other groups is the treatment after the treatment of the dilute stool during the period of the treatment to the normal stool.2.1 after the treatment. The abdominal water of the rats in the low dose group of Wei Ling Ling decoction was significantly reduced and the color was pale yellow. The model group could see more ascites, the ascitic water in the furosemide + analastong group, and a small amount of ascites in the group of Fuzheng Huayu Decoction. The difference of the high dosage group of Jiawei Ling Ling Decoction and the model group was compared with the model group (P0.05), and the ratio of the furosemide + analastong group to the model group was compared. The difference had significant statistical significance (P0.01), but the difference was not statistically significant (P0.05) in the group of Fuzheng Huayu and the model group (P0.05); the difference was not statistically significant (P0.05) in the high dose group of Jiawei Ling Ling Decoction and the low dose group of Jiawei Ling Ling Decoction and the group of furosemide + anolactone (P), and the difference was statistically significant (P 0.05) the high dose group of.2.2 added Wei Ling soup, the low dose group of Jiawei Ling Ling Decoction and the 24 hours after treatment, the net water content of the group was significantly increased, the difference has significant statistical significance (P0.01), and the net amount of water in the group of Fuzheng Huayu group was increased after 24 hours after treatment, and the difference was not statistically significant (p0. 05) the pressure of portal vein in each group of.2.3 was reduced in varying degrees, and the difference was statistically significant (P0.05). The difference was statistically significant (P0.05) in the high dose group of Jiawei Ling Ling decoction, the low dose group of Jiawei Ling Ling Decoction and the group of furosemide + anolactone (P0.05), and the difference between the high dose group and the group of Fuzheng Huayu group was poor. The difference was statistically significant (P0.05); the difference between the low dose group of Jiawei Ling Ling Decoction and the group of Fuzheng Huayu was not statistically significant (P0.05). It showed that Jiawei Ling decoction could reduce the pressure of portal vein in the ascites of the rat liver cirrhosis, and the best effect of the large dosage group of Jiawei Ling Ling Decoction (.2.4 plus Wei Ling Decoction) could obviously reduce the inflammation and inflammation of liver tissue in rats. The degree of fibrous hyperplasia, especially the high dose group of Jiawei Luling soup, was the most obvious. After the treatment of the group of Fuzheng Huayu Capsule (P0.05).2.5, the high dose group of Jiawei Ling Ling decoction, the low dose group of Jiawei Ling Decoction and the group of Fuzheng Huayu could reduce the serum alanine aminotransferase (ALT) and the aminotransferase of aspartic acid in different degrees. The levels of AST, globulin (GLB) and elevated albumin (ALB) were significantly different from those in the model group (P0.01), and the difference was statistically significant (P0.05) compared with the group of furosemide + analosontin (P0.05); the different indexes of the low dose group of Jiawei Ling Ling Decoction and the low dose group of Jiawei Ling Ling Decoction were compared with that of the Fuzheng Huayu group. After P0.05.2.6 treatment, the contents of NO and ET-1 in each group of rats were reduced in varying degrees. There was a significant difference between the high dose group of Jiawei Ling Ling Decoction and the low dose group of Jiawei Ling Ling soup with the model group (P0.05). There was no significant difference between the group of furosemide + analacontan and the group of Fuzheng Huayu and model group (P0.05). The high dose group of Wei Ling decoction, the low dose group of Jiawei Ling Ling Decoction and the group of furosemide + anolactone, the difference has statistical significance (P0.05); the high dose group of Jiawei Ling Ling decoction, the low dose group of Jiawei Ling Ling Decoction and the group of Fuzheng Huayu group, the difference has statistical significance (P0.05) after.2.7 treatment, the high dose group of Jiawei Ling Ling Decoction and the low dose group of Jiawei Luling soup Hyaluronic acid (HA), laminin (LN), type III procollagen (PC III) and type IV collagen (CIV) in the group of Fuzheng Huayu group were significantly lower than those in the model group (P0.05 or P0.01), and there was no significant difference between the model group and the model group (P0.05). There was no significant difference in the low dose group of Wei Ling Decoction (P0.05). Compared with the group of furosemide + anacontan, the high dose group of Jiawei Ling Ling Decoction and the low dose group of Jiawei Ling Ling decoction had significant difference (P0.01) the clinical comprehensive curative effect of.3 and the curative effect of TCM syndrome: the effective and total effective rate of the treatment group was 23.33% and 93.33%, respectively, for the 1 months of clinical treatment. The control group was 13.33% and 70% respectively. The difference between the two groups was statistically significant (P0.05), the main symptoms before and after treatment in the.3.1 two groups, the improvement of physical signs, the major symptoms after treatment, and the improvement of the physical signs were significant (P0.05), and the main symptoms and body after treatment in the control group. The difference was not statistically significant (P0.05) before treatment (P0.05), there was no significant difference in the liver function and prothrombin activity in the.3.2 two groups. The level of ALB, PTA, and A/G in the treatment group was significantly higher than that before treatment (P0.05), and the level of ALB, PTA, and A/G in the control group was also elevated, but there was no statistical difference compared with that before treatment. Learning significance (P0.05); two groups of patients with AST, ALT, TBIL level after treatment, the difference was not statistically significant (P0.05), ALB, PTA, A/G level after treatment compared, the difference was statistically significant (P0.05).3.3 two group of body weight, abdominal circumference, average 24h urine volume, two groups of patients after treatment weight, abdominal circumference and before treatment before treatment, compared with before treatment, average 24h The urine volume was significantly increased, the difference was statistically significant (P0.05). There was no significant difference between the weight and abdominal circumference of the control group before and after treatment (P0.05), and the difference of the average 24h urine volume before and after treatment was statistically significant. The difference of the weight, abdominal circumference and mean 24h urine volume after treatment in the two groups was significant (P0.05) in the two groups of patients with abdominal B ultrasound comparison, and the two groups of patients were compared. After treatment, the index of abdominal B-ultrasound (including the thickness of the spleen, the width of the portal vein and the splenic vein) was better than before the treatment (P0.05); the spleen thickness of the control group and the width of the splenic vein were better than those in the treatment group. The two groups of patients after treatment (including the thickness of the spleen, the width of the portal vein and the splenic vein) were poor after treatment. No statistical significance (P0.05).3.5 two groups of patients with B ultrasonic water volume decreased, the treatment group compared with before treatment was statistically significant (P0.05); the control group after treatment and before treatment, the difference was not statistically significant; the two groups after treatment, the difference was statistically significant (P0.05), indicating the treatment group to eliminate ascites in the treatment group. The incidence of complications in the treatment group of.3.6 two groups was 13.33% and 23.33%, respectively, and there was no significant difference in the incidence of complications (P0.05). There were no significant changes in blood, urine, routine, general physical examination, renal function and electrocardiogram in group.3.7 two. And there is no adverse reaction after the treatment, suggesting that Jiawei Ling soup is safe and effective in the treatment of cirrhotic ascites, and there is no obvious side effect. Conclusion: 1 the rat model of cirrhotic ascites by using two methylnitrosamine can be successfully established, and the general condition of the model rats can be obviously changed by the preparation of.2 added Wei Ling Soup for further study, and the model can be improved. The activity of the rat and the effect of obvious ascites elimination.3 can obviously increase the net water content of the model rats for 24 hours, and the strong water effect of.4 can reduce the pressure of the rat's portal vein, which can reduce the portal pressure of the model rats,.5 and Wei Ling soup can inhibit the proliferation of collagen fiber and reduce the inflammation of the liver tissue. Vitamin.6 and Wei Ling decoction can reduce the content of NO and ET-1 in model rats, thereby adjusting the effect of liver blood flow and improving the portal.
【学位授予单位】：湖北中医药大学
【学位级别】：博士
【学位授予年份】：2016
【分类号】：R259

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