本文選題：肝疾病/酒精性　切入點(diǎn)：肝纖維化　出處：《第一軍醫(yī)大學(xué)》2007年碩士論文

【摘要】： 酒精性肝病(Alcoholic liver disease，ALD)是由于長(zhǎng)期大量飲酒導(dǎo)致的肝臟疾病。根據(jù)中華醫(yī)學(xué)會(huì)肝臟病學(xué)分會(huì)提出的ALD病理診斷標(biāo)準(zhǔn)，分輕型酒精性肝病(mild alcoholic injury，MAI)、酒精性脂肪肝(alcoholic fatty liver，AFL)、酒精性肝炎(alcoholic hepatic，AH)、酒精性肝纖維化(alcoholic hepatic fibrosis，AHF)、酒精性肝硬化(alcoholic cirrhosis，AC)5種類型。ALD是西方國(guó)家導(dǎo)致肝硬化的最主要的病因，也是十大常見死因之一。近年來(lái)隨著人們生活方式的改變，我國(guó)由酒精所至肝損害的發(fā)病率亦呈逐年上升趨勢(shì)，成為繼病毒性肝炎后導(dǎo)致肝損害的第二大病因。肝纖維化是肝細(xì)胞發(fā)生壞死或炎癥刺激時(shí)，肝內(nèi)纖維結(jié)締組織異常增生的病理過(guò)程，進(jìn)一步發(fā)展可引起肝小葉結(jié)構(gòu)改建、假小葉及結(jié)節(jié)形成，即肝硬化。國(guó)內(nèi)外學(xué)者對(duì)肝纖維化的可逆性已無(wú)異議，故酒精性肝纖維化對(duì)酒精性肝硬化的出現(xiàn)與否及疾病的預(yù)后起著決定性的作用，其演變和逆轉(zhuǎn)近年來(lái)倍受關(guān)注。為深入研究酒精性肝病的發(fā)病機(jī)制，篩選有效預(yù)防、治療本病的藥物及方法，建立與人類酒精性肝損傷病變過(guò)程相似的動(dòng)物模型具有積極的現(xiàn)實(shí)意義。 目的 本文第一部分旨在探索建立較為理想的小鼠酒精性肝纖維化模型，為研究骨髓衍生肝干細(xì)胞對(duì)酒精性肝損傷的修復(fù)提供動(dòng)物模型。本文第二部分對(duì)我院近5年來(lái)收治的105例酒精性肝病患者進(jìn)行臨床資料分析，探討酒精性肝病的發(fā)病特點(diǎn)、臨床表現(xiàn)及預(yù)后，以提高對(duì)酒精性肝病的認(rèn)識(shí)和重視，為早期發(fā)現(xiàn)、早期診斷、早期治療酒精性肝病提供依據(jù)。 方法和結(jié)果 1、小鼠酒精性肝纖維化模型的建立 Balb／c雌性小鼠100只，SPF級(jí)，5～6周齡，體重15～18g，適應(yīng)環(huán)境一周后，隨機(jī)分為造模組92只，對(duì)照組8只。造模組給予54度白酒每天灌胃2次共16周，同時(shí)以10％(v／v)白酒為唯一飲料。第1周～第3周末灌胃量為每次8mL／kg，第4周～第7周末灌胃量為每次10 mL／kg，第8周～第16周末灌胃量為每次12 mL／kg。對(duì)照組給予等量蒸餾水灌胃，飲用自來(lái)水。實(shí)驗(yàn)期間均予全價(jià)營(yíng)養(yǎng)顆粒飼料喂養(yǎng)。造模組于白酒灌胃的第4，8，12，16周末分別隨機(jī)抽樣選取6只小鼠，眼球采血后，頸椎脫臼處死，處死前禁食12h。處死后立即取出肝臟，稱重后以10％的甲醛固定，脫水后石臘包埋切片，蘇木精—伊紅染色(HE)和苦味酸—酸性品紅染色(VG)，光鏡觀察病理變化。血標(biāo)本用于測(cè)定生化指標(biāo)。對(duì)照組小鼠于第16周末處死，處理同造模組。實(shí)驗(yàn)過(guò)程中觀察小鼠的精神狀態(tài)、活動(dòng)情況、皮毛光澤度、食欲等。肝臟標(biāo)本觀察肝臟的大小、外形、色澤、質(zhì)地、切面情況。取血清進(jìn)行生化指標(biāo)的檢測(cè)，包括丙氨酸氨基轉(zhuǎn)移酶(ALT)，天冬氨酸氨基轉(zhuǎn)移酶(AST)，甘油三脂(TG)，總蛋白(TP)，白蛋白(ALB)。病理學(xué)觀察：肝組織常規(guī)石臘包埋切片，HE染色和VG染色，用于觀察肝臟病理學(xué)改變(肝細(xì)胞變性、壞死、炎性細(xì)胞浸潤(rùn)、膠原纖維增生等)，結(jié)果如下。 1．1實(shí)驗(yàn)動(dòng)物數(shù)量分析：實(shí)驗(yàn)過(guò)程中造模組共23只小鼠死亡，多為急性和亞急性死亡。原因主要有：窒息、消化道穿孔、急性胃擴(kuò)張和酒精中毒等。對(duì)照組全部成活。 1．2精神變化：灌胃初期，造模組小鼠出現(xiàn)精神萎靡、嗜睡、反應(yīng)遲鈍、活動(dòng)減少、食欲減退等現(xiàn)象，實(shí)驗(yàn)開始4周后上述癥狀減輕，但皮毛光澤度差，體重增長(zhǎng)緩慢。對(duì)照組小鼠則皮毛光澤、體態(tài)活潑、食欲正常，無(wú)嗜睡現(xiàn)象。 1．3肝臟標(biāo)本的外觀：對(duì)照組小鼠肝臟表面光滑細(xì)潤(rùn)、色紅、邊緣銳利、質(zhì)地中等。相比之下，造模組小鼠肝臟色澤較正常肝臟暗淡，表面充血。 1．4組織的病理學(xué)改變：對(duì)照組HE染色標(biāo)本見肝細(xì)胞以中央靜脈為中心呈放射狀排列，肝小葉輪廓清晰，肝索排列整齊。造模組4周時(shí)可見輕度脂肪變性，肝細(xì)胞濁腫，胞漿中出現(xiàn)大小不等的脂滴；8周時(shí)，細(xì)胞索紊亂，，胞漿腫大疏松化，其內(nèi)細(xì)胞核固縮，可見肝細(xì)胞廣泛性的空泡樣變性，脂肪變性較4周模型組小鼠增多；12周時(shí)，肝細(xì)胞點(diǎn)狀及灶狀壞死較多見，匯管區(qū)有炎性細(xì)胞浸潤(rùn)，一些壞死區(qū)可見纖維細(xì)胞增生；16周時(shí)，炎癥壞死和纖維化更加明顯，匯管區(qū)有明顯紅色膠原纖維增生，并向周圍肝小葉內(nèi)延伸。 1．5血清生化指標(biāo)：各周造模組小鼠血清中白蛋白含量與正常對(duì)照組比較差異顯著(F=6.490，P=0.001)；各周模型組相比，血清白蛋白含量沒(méi)有顯著差異(F=0.620，P=0.610)。造模組血清總蛋白的含量與對(duì)照組相比，在造模的第4，8，12，16周時(shí)沒(méi)有明顯改變，差異不顯著(F=1.217，P=0.327)；造模各時(shí)間點(diǎn)間兩兩比較，總蛋白含量也沒(méi)有顯著性差異(F=0.965，P=0.429)。各周造模組小鼠血清中AST、ALT、TG含量和AST／ALT與對(duì)照組相比，除4周模型組AST／ALT、TG與正常對(duì)照組相比無(wú)顯著性差異外(P=0.083；P=0.318)，其余均有顯著性差異(F=10.281，P＜0.001；F=8.610，P=＜0.001；F=3.605，P=0.018；F=4.490，P=0.007)。 2、酒精性肝病105例臨床分析 依據(jù)患者的飲酒史、臨床表現(xiàn)、肝功能指標(biāo)及影像學(xué)、病理學(xué)檢查的結(jié)果，按照中華醫(yī)學(xué)會(huì)肝病學(xué)分會(huì)脂肪肝和酒精性肝病學(xué)組2006年2月修訂的酒精性肝病診療指南上的酒精性肝病臨床診斷標(biāo)準(zhǔn)，將我院近5年來(lái)收治的105例酒精性肝病患者分為酒精性脂肪肝組(AFL)、酒精性肝炎組(AH)、酒精性肝硬化組(AC)，對(duì)其數(shù)據(jù)進(jìn)行統(tǒng)計(jì)學(xué)分析，并做出臨床分析和討論，其結(jié)果如下。 2．1 ALD臨床表現(xiàn)與其他原因所致肝病類似，無(wú)特異性，常見癥狀為乏力、納差、發(fā)熱、黃疸、腹脹、嘔血、黑便。 2．2 ALD患者男性為主，日飲酒量45～600g，平均年限21年。本資料中ALD患者年齡在30～60之間的病例數(shù)最多(89例)。酒精性肝硬化67例，占63.8％。AC組日飲酒量同AFL組、AH組相比，差異顯著(P=0.001，P=0.013)；AC組飲酒時(shí)間(24.13±7.84)年同AFL組(16.21±12.43)年、AH組(16.00±10.09)年相比也均有顯著性差異(P=0.001，P=0.001)。 2．3 ALD患者的血清酶學(xué)水平有不同程度的異常，血清中ALT、AST、GGT的水平以AH組為最高，同AFL組相比，差異顯著(P＜0.001)；AH組患者血清中AST、ALT、GGT的水平同AC組患者相比，差異顯著(P＜0.001；P＜0.001；P=0.047)。各組間AST／ALT相比無(wú)顯著性差異(F=0.801，P=0.451)。血清ALB以AC組為最低，分別與AH組、AFL組相比均有顯著性差異(P=0.003；P＜0.001)。TBIL以AC組最高，3組間差異有顯著性意義(P＜0.001)。ALD患者死亡的主要原因是肝硬化的晚期并發(fā)癥。 結(jié)論 1、應(yīng)用白酒灌胃法成功的復(fù)制了小鼠酒精性肝纖維化模型，并在16周內(nèi)觀察到酒精性肝病的一些病理表現(xiàn)，如脂肪變性、炎癥改變、膠原纖維的增生。隨著酒精刺激時(shí)間的延長(zhǎng)，肝臟的損傷程度加重。 2、此種建模方法接近國(guó)人的飲酒習(xí)慣、方法簡(jiǎn)便易行、費(fèi)用低，可為研究骨髓衍生肝干細(xì)胞對(duì)酒精性肝損傷的修復(fù)提供較理想的動(dòng)物模型。 3、本研究中ALD患者高峰年齡在30～60之間，飲酒量、飲酒時(shí)間與酒精性肝病的發(fā)病關(guān)系密切。 4、因酒精性肝病臨床表現(xiàn)無(wú)特異性，對(duì)于有飲酒嗜好的病人，應(yīng)定期對(duì)AST、AST／ALT，GGT，TBIL等生化指標(biāo)監(jiān)測(cè)并結(jié)合影像學(xué)檢查，從而爭(zhēng)取早期診斷、早期治療、改善預(yù)后。
[Abstract]:Alcohol liver disease ( ALD ) is a major cause of liver injury due to chronic alcoholic liver disease ( ALD ) .






Purpose






The first part of this paper is to explore the establishment of an ideal model of alcoholic liver fibrosis in mice , and to provide an animal model for the study of the repair of alcoholic liver injury by bone marrow derived liver stem cells . The second part of this paper deals with the clinical data of 105 patients with alcoholic liver disease treated in our hospital in recent 5 years , and discusses the pathogenesis , clinical manifestation and prognosis of alcoholic liver disease , so as to improve the awareness and attention of alcoholic liver disease . It provides the basis for early detection , early diagnosis and early treatment of alcoholic liver disease .






Methods and Results






1 . Establishment of model of alcoholic liver fibrosis in mice






Blood samples were used to measure biochemical indexes . The rats were randomly divided into two groups at 8 mL / kg every 8 mL / kg and 8 weeks to 16 weeks .






1.1 The number of experimental animals showed that 23 mice died during the experiment , mostly acute and subacute death . The main reasons were asphyxia , perforation of digestive tract , acute gastric dilatation and alcoholism . All the control groups were alive .






P < 0.001 ;






1.3 Appearance of the liver specimen : the liver surface of the control group was smooth and smooth , the color was red , the edge was sharp , and the texture was middle . In contrast , the liver of the model mice had a normal liver color , and the surface was filled with blood .






1.4 The pathological changes of the liver tissues were as follows : the control group HE staining samples showed that the liver cells were radially arranged in the central vein , the hepatic lobule was clear and the liver was arranged orderly .
At 8 weeks , the cell line was disturbed , the cytoplasm was loose , the cells in its inner cells were pyknosis , the vacuolar degeneration of hepatocytes was seen , and the number of mice with fatty degeneration was more than 4 weeks .
At 12 weeks , the hepatocyte punctate and focal necrosis were more common , inflammatory cell infiltration in the manifold area , and the proliferation of some necrotic areas .
At 16 weeks , inflammatory necrosis and fibrosis were more evident , and the manifold area had a marked red collagen fiber hyperplasia and extended to the surrounding liver leaflets .






1.5 Serum biochemical indexes : The serum albumin content of every peripheral group was significantly different from that of the normal control group ( F = 6.490 , P = 0.001 ) .
There was no significant difference in serum albumin content ( F = 0.620 , P = 0.610 ) . Compared with the control group , there was no significant difference between the serum total protein content and the control group ( F = 1.217 , P = 0.327 ) .
There was no significant difference ( F = 0.965 , P = 0.429 ) . Compared with control group , AST , ALT , TG and AST / ALT were not significantly different from normal control group ( P = 0.083 ) .
There was significant difference ( F = 10.281 , P < 0.001 ) .
F=8.610,P=錛

本文編號(hào)：1698756