肝炎肝硬化“肝藏血”功能异常表现与微观指标的相关性研究

【作者】 赵丽红；

【导师】 王天芳；

【作者基本信息】 北京中医药大学 ， 中医诊断学， 2014， 博士

【摘要】 肝炎肝硬化是由于肝炎病毒长期作用于肝脏引起的一种慢性、进行性、弥漫性肝病的终末阶段,肝脏纤维组织增生,破坏正常肝小叶的结构,代之以再生结节和假小叶,使肝脏正常血液供应被破坏,临床表现为门静脉高压症及肝功能障碍等。根据中医理论,肝炎肝硬化的发生与发展过程存在肝主疏泄和藏血功能的异常,其中“肝藏血”功能异常主要表现为血瘀、阴虚等基本病机。本研究拟从分析肝炎肝硬化“肝藏血”功能异常表现与微观指标的相关性入手并由病理反推生理,为阐释“肝藏血”的现代科学内涵提供一定依据。目的1梳理“肝藏血”的理论与肝炎肝硬化的病理生理,分析中医对肝炎肝硬化病因病机的认识,构建探讨肝炎肝硬化“肝藏血”功能异常表现与微观指标相关性的研究思路。2基于临床横断面调查数据,分析肝炎肝硬化常见证候要素的分布特点,在此基础上,了解其“肝藏血”功能异常相关的证候要素血瘀、阴虚等的表现特点。3基于临床横断面调查数据,分析肝炎肝硬化“肝藏血”功能异常时微观指标的表现特点,为进一步探讨两者间的相关性提供参考依据。4基于临床横断面调查数据,分析肝炎肝硬化“肝藏血”功能异常表现与微观指标的相关性,并由病理反推生理,为阐释“肝藏血”的现代科学内涵提供一定依据。方法1构建研究思路通过梳理中医对“肝藏血”功能及功能异常表现和肝炎肝硬化病因病机的认识,以及西医对肝脏生理和肝炎肝硬化病理生理的认识,归纳肝炎肝硬化“肝藏血”功能异常表现的证候要素以及微观指标,构建探讨两者间相关性的研究思路。2临床数据采集采用前瞻性多中心的横断面流行病学调查方法,运用课题组统一制订的《肝炎肝硬化临床信息采集表》采集患者的中医症状、体征等临床表现以及实验室、影像学检查等微观指标。3制订《肝炎肝硬化常见证候要素辨识标准》(供本课题研究使用)参照现行的多个诊疗共识,结合前期文献回顾与临床调查结果,并经过二轮专家论证制订。4制订《肝炎肝硬化常见症状与证候要素量化表》根据信息采集表中对中医症状和体征的量化分级形成,证候要素量化得分为相关症状、体征得分的加和。5数据处理包括数据录入,数据清洗,数据类型转换等。6统计学方法应用SPSS13.0统计软件包进行。6.1分析分布特点计量资料采用独立样本t检验或Mann-Whitney秩和检验比较两组间的差异,采用单因素方差分析或Kruskal-Wallis H检验比较三组间的差异；计数资料采用卡方检验比较两组间的差异。统计推断以0.05作为检验水准。6.2分析相关性(1)线性趋势图：观察血瘀、阴虚的证候要素量化得分与微观指标水平均值的线性趋势。(2)简单相关分析：连续、分类变量分别采用Pearson、Spearman相关分析方法。(3)多元线性回归：以血瘀、阴虚的证候要素量化得分为因变量,微观指标水平为自变量,自变量筛选采用逐步回归法。(4)二分类Logistic回归分析：以是、否判定为血瘀或阴虚合并血证为因变量,以微观指标是、否异常为自变量,采用进入法分别进行单自变量的模型拟合。(5)逐步贝叶斯判别分析：以是、否判定为血瘀或(和)阴虚以及其是、否合并各血证作为因变量,以微观指标为自变量,自变量筛选采用Wilks’ Lambda法。结果1肝炎肝硬化“肝藏血”功能异常表现与微观指标相关性的研究思路肝炎肝硬化的发生发展过程与“肝藏血”功能异常的血瘀、阴虚等基本病机密切相关。以统计学方法为工具,探讨患者的血瘀、阴虚(或合并血证)与微观指标(血常规、凝血功能、肝功能、门静脉血流动力学、脾脏体积、胃镜检查、肝脏储备功能、血管活性物质等指标)的相关性。2肝炎肝硬化常见证候要素的分布特点2.1常见证候要素的分布801例肝炎肝硬化患者中,血瘀503例,阴虚448例,湿热257例,气滞438例,气虚517例,阳虚428例,水停462例。2.2血瘀、阴虚的分布在代偿期与失代偿期患者中,血瘀分别占52.50%、69.60%,量化得分分别为4.97±2.41、5.96±2.61(P<0.05)；阴虚分别占55.40%、56.30%,量化得分分别为5.33±2.05、5.55±2.15(P>0.05)。2.3血证的分布吐血、便血、紫斑、齿衄、鼻衄患者在代偿期/失代偿期分别有5例/38例,5例/33例,7例/16例,72例/78例,43例/70例。齿衄多合并鼻衄,吐血多合并便血,前者两期均有分布,后者主要分布在失代偿期。2.4血瘀、阴虚与血证组合的分布67.40%的吐血、71.00%的便血、91.30%的紫斑、92.00%的齿衄及96.40%的鼻衄患者均合并血瘀或(和)阴虚。血瘀、阴虚得分在血瘀、阴虚与(血瘀合阴虚)分别合并各血证的三组间存在差异,在合并紫斑、吐血、便血时均较高。3肝炎肝硬化“肝藏血”功能异常时微观指标的表现特点3.1肝炎肝硬化患者血瘀出现或程度加重时微观指标的表现特点分别比较患者血瘀组与非血瘀组、血瘀代偿期与失代偿期微观指标水平的差异以及血瘀在各微观指标是、否异常两组的分布和得分差异,在血瘀出现或程度加重时,微观指标的变化有：(1)红细胞、血红蛋白、红细胞压积、血小板计数、血小板平均体积降低；(2)凝血酶原时间延长、凝血酶原活动度降低、凝血酶原时间比值升高；(3)白蛋白、白球比、前白蛋白降低,球蛋白升高；(4)MELD、Child-Pugh评分升高；(5)脾脏长径增长、厚径增厚；(6)门静脉内径增宽、门静脉平均血流速度减低,等。比较患者血瘀分别合并吐血、便血、紫斑、齿衄、鼻衄时在各微观指标是否异常两组的分布差异,在血瘀合并血证出现时,微观指标的变化有：(1)红细胞、血红蛋白、红细胞压积、血小板计数降低；(2)凝血酶原活动度降低、凝血酶原时间比值升高；(3)总蛋白、白蛋白、白球比降低；(4)肝固有动脉峰值流速升高,等。以上差异均有统计学意义(P<0.05)。3.2肝炎肝硬化患者阴虚出现或程度加重时微观指标的表现特点分别比较患者阴虚组与非阴虚组、阴虚代偿期与失代偿期微观指标水平的差异以及阴虚在各微观指标是、否异常两组的分布和得分差异,在阴虚出现或程度加重时,微观指标的变化有：(1)凝血酶原活动度降低、凝血酶原时间比值升高、纤维蛋白原升高；(2)球蛋白升高、白球比降低；(3)红细胞、血红蛋白、红细胞压积、血小板平均体积降低；(4)脾脏厚径增厚；(5)肝固有动脉峰值流速升高、门静脉平均血流速度减低；(6)NO水平降低,等。比较患者阴虚分别合并吐血、便血、紫斑、齿衄、鼻衄时在各微观指标是、否异常两组的分布差异,在阴虚合并血证出现时,微观指标的变化有：(1)中性粒细胞百分比、红细胞、血红蛋白、红细胞压积、血小板计数降低；(2)凝血酶原活动度降低、凝血酶原时间比值升高；(3)MELD评分升高；(4)肝固有动脉内径扩张,等。以上差异均有统计学意义(P<0.05)。3.3肝炎肝硬化患者血瘀、阴虚与(血瘀合阴虚)三组间微观指标的表现特点比较血瘀组、阴虚组与(血瘀合阴虚)组及其分别合并吐血、便血、紫斑、齿衄、鼻衄时三组间指标水平的差异,微观指标的变化有：(1)血瘀或(血瘀合阴虚)与阴虚相比,白蛋白、白球比、总蛋白降低,MELD、Child-Pugh评分升高,门、脾静脉流速降低；(2)(血瘀合阴虚)与阴虚相比,红细胞降低、凝血酶原时间延长、凝血酶原活动度降低、凝血酶原时间比值升高；(3)阴虚或(血瘀合阴虚)与血瘀相比,5-HT、NO水平降低；(4)阴虚与血瘀相比,门静脉内径变窄、中性粒细胞百分比降低,等。以上差异均有统计学意义(P<0.05)。4肝炎肝硬化“肝藏血”功能异常表现与微观指标的相关性4.1肝炎肝硬化血瘀与微观指标的相关性总结血瘀与微观指标的相关性在线性趋势、简单相关分析、多元线性回归分析的表现,与血瘀呈正相关的微观指标有：(1)血小板计数、血小板平均体积降低；(2)凝血酶原时间延长、凝血酶原活动度降低、凝血酶原时间比值升高；(3)球蛋白升高,白蛋白、白球比降低；(4)脾脏厚径增厚；(5)门静脉平均血流速度减低、门静脉内径增宽；(6)AT-Ⅱ水平升高；(7)MELD、Child-Pugh评分升高；(8)红细胞、血红蛋白、红细胞压积降低,等。总结血瘀分别合并吐血、便血、紫斑、齿衄、鼻衄时与微观指标的相关性在简单相关分析和二分类Logistic回归分析的表现,与血瘀合并血证呈正相关的微观指标有：(1)凝血酶原活动度降低、凝血酶原时间比值升高；(2)红细胞、血红蛋白、红细胞压积降低；(3)肝固有动脉峰值流速升高,等。以上结果均有统计学意义(P<0.05)。4.2肝炎肝硬化阴虚与微观指标的相关性总结阴虚与微观指标的相关性在线性趋势、简单相关分析、多元线性回归分析的表现,与阴虚呈正相关的微观指标有：(1)凝血酶原活动度降低、凝血酶原时间比值升高、纤维蛋白原升高；(2)白球比、前白蛋白降低,球蛋白升高；(3)门静脉内径变窄、肝固有动脉峰值流速升高、肝固有动脉内径扩张；(4)中性粒细胞百分比降低；(5)5-HT、TNF-α、NO水平降低,等。总结阴虚分别合并吐血、便血、紫斑、齿衄、鼻衄时与微观指标的相关性在简单相关分析和二分类Logistic回归分析的表现,与阴虚合并血证呈正相关的微观指标有：(1)红细胞、血红蛋白、红细胞压积降低；(2)凝血酶原活动度降低、凝血酶原时间比值升高,(3)肝固有动脉内径扩张,等。以上结果均有统计学意义(P<0.05)。4.3肝炎肝硬化血瘀、阴虚与(血瘀合阴虚)三组间的判别分析在判别血瘀、阴虚与(血瘀合阴虚)时有贡献的微观指标为红细胞、白蛋白、门静脉内径、5-HT和TNF-α。在判别血瘀、阴虚与(血瘀合阴虚)分别合并吐血、便血、紫斑、齿衄、鼻衄时有贡献的微观指标为红细胞、白细胞、凝血酶原活动度、纤维蛋白原、Child-Pugh评分和NO。结论1随着肝炎肝硬化血瘀程度的加重,门静脉高压加重,脾肿大/脾功能亢进加重,肝脏储备功能降低,肝脏凝血功能障碍,等。(?)2随着肝炎肝硬化阴虚程度的加重,肝脏合成凝血酶原功能障碍,肝脏合成血浆蛋白功能障碍,门静脉高压加重,等。3从肝炎肝硬化的病理反推生理,初步表明,“肝藏血”与门静脉血流动力学、脾脏功能、肝脏合成血浆蛋白功能、肝脏合成凝血酶原功能、肝脏储备功能等有一定相关性。更多还原

【Abstract】 Hepatitis cirrhosis is the end-stage of chronic, progressive and diffuse liver disease due to the long-term effects of hepatitis virus on the liver. Liver fibrous tissue proliferated, normal lobular structures was disrupted and replaced with regenerative nodules and false lobules, resulting in normal blood supply to the liver was damaged, and its clinical menifestation included liver dysfunction and portal hypertension,etc. According to traditioanal Chinese medicine(TCM), the occurrence and development of hepatitis cirrhosis was affected by the dysfunction of liver controlling dispersion and housing blood. The specific contents of dysfunction of liver housing blood included blood stasis, yin deficiency, etc. The study focused on analyzing the correlation between dysfunction of liver housing blood and indicators from lab findings and imaging of hepatitis cirrhosis, supplying evidence for interpreting the modern scientific connotation of liver housing blood by inferring physiology from pathology.Objective1To build the working hypothesis by combing the theory of hepatitis cirrhosis and liver housing blood.2To understand the distribution features of common syndrome elements of hepatitis cirrhosis, and then understand the manifestation featres of blood stasis, yin deficiency, etc. based on the clinical cross-sectional survey data.3To understand the manifestation features of indicators when liver housing blood was dysfunction based on the clinical cross-sectional survey data and supply evidence for further analysis.4To supply evidence for interpreting the modern scientific connotation of dysfunction of liver housing blood by analyzing the correlation between the dysfunction manifestation of liver housing blood and indicators from lab findings and imaging based on the clinical cross-sectional survey data.Methods1Building the working hypothesisThe traditional Chinese medicine(TCM) theory of the dysfunction manifestation of liver housing blood and hepatitis cirrhosis pathogenesis and the western medicine theory of liver physiology and hepatitis cirrhosis pathophysiology was combed. The syndrome elements of the dysfunction manifestation of liver housing blood was established. The working hypothesis was built.2Data collectionNationwide multi-center cross-sectional epidemiological survey was conducted. TCM symptoms, signs and indicators from lab findings and imaging were collected by using the unified "hepatitis cirrhosis case survey form" formulated by research group.3"Standards of hepatitis cirrhosis syndrome elements differentiation"The standards were formulated based on current multiple clinic consensus, literature review, clinical survey data and experts evaluation.4"Quantization form of hepatitis cirrhosis symptoms and syndrome elements"The form was formulated based on the quantitative classification of symptoms and signs in the "hepatitis cirrhosis case survey form".5Data processingIncluding data entry, data cleaning and data type conversion.6Statistical methodsSPSS13.0statistical package was adopted.6.1Analyzing the distribution features(1) Independent sample t test or Mann-Whitney rank sum test was used to compare the difference between two groups and one-way ANOVA or Kruskal-Wallis H test was used to compare the differences among three groups when the data was measurement.(2)The chi-square test was used to compare the difference between two groups when the data was count.Statistical inference took0.05as the significance level.6.2Analyzing the correlation(1)Linear trend:a linear trend was used to observe the liner trend between syndrome elements quantized score of blood stasis or yin deficiency and indicators’average level.(2) Simple correlation analysis:pearson and spearman correlation analysis were respectively used to analyze continuous and categorical variables.(3)Multiple linear regression:syndrome elements quantized score of blood stasis or yin deficiency as the dependent variable, the indicators level as independent variable, the variable selection using stepwise regression method. (4) Binary Logistic Regression Analysis:whether blood stasis or yin deficiency combined with bleeding syndrome being diagnosed as the dependent variable, whether indicators being abnormal as independent variables, single independent variable in fitting the model were respectively conducted using the enter method.(5) Stepwise Bayes discriminant analysis:whether blood stasis or yin deficiency being diagnosed, and whether blood stasis or yin deficiency combined with bleeding syndrome being diagnosed as the dependent variables, indicators as the independent variables, variables selection using Wilks’Lambda method.Results1Building the working hypothesis of the studyThe occurrence and development of hepatitis cirrhosis was affected by blood stasis and yin deficiency, which were the dysfunction manifestation of liver housing blood. The correlation between the syndrome elements of blood stasis, yin deficiency(or combined with bleeding syndrome) and indicators(routine blood test, clotting function, liver function, portal hemodynamics, liver ultrasound, endoscopy, liver reserve function, vasoactive substances, etc.) were explored by using statistical methods as the tools.2The distribution features of syndrome elements of hepatitis cirrhosis(1) The distribution of syndrome elements:503cases of patients were diagnosed blood stasis,448case of yin deficiency,257cases of damp heat,438cases of qi stagnation,517cases of qi deficiency,428cases of yang deficiency,462cases of water retention during all the801cases of hepatitis cirrhosis patients.(2)The distribution of blood stasis and yin deficiency:In the compensated and decompensated stages, patients with blood stasis accounted for52.50%and69.60%and quantified score were4.97±2.41and5.96±2.61respectively (P<0.05); patients with yin deficiency accounted for55.40%and56.30%and quantified score were5.33±2.05and5.55±2.15respectively (P>0.05)(3) The distribution of bleeding syndrome:In the compensated&decompensated stage, patients with hematemesis, melena, ecchymosis, gum bleeding and epistaxis accounted for5&38,5&33,7&16,72&78and43&70cases respectively. In addition, gum bleeding usually combined with epistaxis, which was most distributed in the two stages. Hematemesis combined with melena, which was distributed in the decompensated stage.(4) The distribution of combination of blood stasis or yin deficiency with bleeding syndrome:67.4%of hematemesis patients,71%of melena patients,91.3%of ecchymosis patients,92%of gum bleeding patients and96.4%of epistaxis patients combined with blood stasis or(and) yin deficiency. The quantified score of blood stasis and yin deficiency exists difference among the three groups of blood stasis, yin deficiency and their combination and both the score was much higher when combined with ecchymosis, hematemesis or melena than other bleeding syndrome.3The manifestation features of indicators when liver housing blood dysfunction3.1The manifestation features of indicators when blood stasis existed or aggravatedWhen blood stasis existed or aggravated, the changes of indicators contained:(1) reduced erythrocytes, hemoglobin, hematocrit, platelets, mean platelet volume, etc.;(2)prolonged prothrombin time, reduced prothrombin time activity percentage, increased international normalized ratio;(3) reduced albumin, albumin/globulin and prealbumin, elevated globulin;(4) increased MELD, Child-Pugh score;(5)growed spleen diameter and thickness;(6)widened portal vein, reduced portal vein average flow velocity, etc. through comparing the difference of indicator level between blood stasis and not blood stasis groups, blood stasis in the compensated and decompensated stages, and the difference of blood stasis’s distribution and quantified score between the normal and abnormal indicators groups.When blood stasis combined with bleeding syndrome existed, the changes of indicators contained:(1) reduced erythrocytes, hemoglobin, hematocrit and platelets;(2) reduced prothrombin time activity percentage, increased international normalized ratio;(3) reduced total protein, albumin, and albumin/globulin;(4) elevated hepatic artery peak velocity, etc. through comparing the difference of distribution of blood stasis combined with hematemesis, melena, ecchymosis, gum bleeding and epistaxis respectively between the normal and abnormal indicators groups.The above differences were statistically significant (P<0.05)3.2The manifestation features of indicators when yin deficiency existed or aggravatedWhen yin deficiency existed or aggravated, the changes of indicators contained:(1) reduced prothrombin time activity percentage, increased international normalized ratio, elevated fibrinogen;(2) elevated globulin, reduced albumin/globulin;(3) reduced erythrocytes, hemoglobin, hematocrit, and mean platelet volume, etc.(4)growed spleen thickness;(5)elevated hepatic artery peak velocity, reduced portal vein average flow velocity;(6) reduced NO level, etc. through comparing the difference of indicator level between yin deficiency and not yin deficiency groups, yin deficiency in the compensated and decompensated stages, and the difference of yin deficiency’s distribution and quantified score between the normal and abnormal indicators groups.When yin deficiency combined with bleeding syndrome existed, the changes of indicators contained:(1) reduced percentage of neutrophils, erythrocytes, hemoglobin, hematocrit, platelets, mean platelet volume;(2) reduced prothrombin time activity percentage, increased international normalized ratio;(3)increased MELD score;(4)widened hepatic artery, etc. through comparing the difference of distribution of yin deficiency combined with hematemesis, melena, ecchymosis, gum bleeding and epistaxis respectively between the normal and abnormal indicators groups.The above differences were statistically significant (P<0.05)3.3The manifestation features of indicators among the three groups of blood stasis, yin deficiency and their combination(1) albumin, albumin/globulin and total protein reduced, MELD, Child-Pugh score increased, portal vein and spleen vein average flow velocity reduced when blood stasis or their combination compared with yin deficiency group;(2) erythrocytes reduced, prothrombin time prolonged, prothrombin time activity percentage reduced, international normalized ratio increased when their combination compared with yin deficiency group;(3)5-HT, NO level reduced when yin deficiency or their combination compared with blood stasis group;(4) portal vein narrowed, percentage of neutrophils reduced when yin deficiency compared with blood stasis group. The above differences were statistically significant (P<0.05)4The correlation between dysfunction manifestation of liver housing blood and indicators of hepatitis cirrhosis patients 4.1The correlation between blood stasis and indicators of hepatitis cirrhosis patientsThe indicators that positively related with blood stasis included:(l)reduced platelets, mean platelet volume;(2) prolonged prothrombin time, reduced prothrombin time activity percentage, increased international normalized ratio;(3) reduced albumin, albumin/globulin, elevated globulin;(4) growed spleen thickness;(5)widened portal vein, reduced portal vein average flow velocity;(6) elevated AT-II level;(7) increased MELD, Child-Pugh score;(8) reduced erythrocytes, hemoglobin, hematocrit, etc. based on linear trend, simple correlation analysis and multiple linear regression.The indicators that positively related with blood stasis combined with bleeding syndrome included:(1)reduced prothrombin time activity percentage, increased international normalized ratio;(2)reduced erythrocytes, hemoglobin, hematocrit;(3)elevated hepatic artery peak velocity, etc. based on the simple correlation analysis and binary logistic regression analysis between indicators and blood stasis combined with bleeding syndrome.The above results were statistically significant (P<0.05)4.2The correlation between yin deficiency and indicators of hepatitis cirrhosis patientsThe indicators that positively related with yin deficiency included:(1)reduced prothrombin time activity percentage, increased international normalized ratio, elevated fibrinogen;(2) reduced albumin/globulin and prealbumin, elevated globulin;(3) narrowed portal vein, elevated hepatic artery peak velocity, widened hepatic artery;(4)reduced percentage of neutrophils;(5)reduced5-HT, TNF-a, NO level, etc. based on linear trend, simple correlation analysis and multiple linear regression.The indicators that positively related with yin deficiency combined with bleeding syndrome included:(1)reduced erythrocytes, hemoglobin, hematocrit;(2) reduced prothrombin time activity percentage, increased international normalized ratio;(3)widened hepatic artery, etc. based on the simple correlation analysis and binary logistic regression analysis between indicators and yin deficiency combined with bleeding syndrome.The above results were statistically significant (P<0.05)4.3Discriminant analysis among the three groups of blood stasis, yin deficiency and their combination of hepatitis cirrhosis patients The indicators that contributed to discriminate blood stasis, yin deficiency and their combination were erythrocytes, albumin, portal vein,5-HT and TNF-a.The indicators that contributed to discriminate blood stasis, yin deficiency and their combination accompanied with5bleeding syndrome respectively were erythrocytes, leukocytes, prothrombin time activity percentage, fibrinogen, Child-Pugh score and NO level.Conclusion1With the severity of blood stasis in hepatitis cirrhosis patients, portal hypertension and splenomegaly/hypersplenism aggravated, liver reserve function decreased, liver coagulation dysfunctioned, etc.2With the severity of yin deficiency in hepatitis cirrhosis patients, liver’s synthesis of prothrombin dysfunctioned, liver’s synthesis of plasma protein dysfunctioned, portal hypertension aggravated, etc.3Inferring physiology from pathology of hepatitis corrihosis, the correlation preliminarily indicated that liver housing blood might associated with portal hemodynamics, spleen function, liver’s synthesis function of plasma protein, liver’s synthesis function of prothrombin, liver reserve function, etc.更多还原