本文關鍵詞： 兔 肝臟解剖 SFSS 動物模型 門靜脈壓力 門-腔分流 肝再生　出處：《昆明醫(yī)科大學》2014年碩士論文　論文類型：學位論文

【摘要】：背景 小肝綜合征(small-for-size syndrome, SFSS)是發(fā)生在擴大肝切除或部分原位肝移植后由于肝臟體積過小導致功能上不能滿足受體的代謝需求而出現的一種臨床綜合征。SFSS最常見于成人—成人部分原位活體肝移植,供肝體積較小時可能導致受體門靜脈血流高灌注和持續(xù)性門靜脈高壓,由此引起一系列嚴重的病理生理紊亂。其臨床表現為：術后持續(xù)性黃疸、凝血功能障礙、門靜脈高壓、頑固性腹水,嚴重者進一步誘發(fā)膿毒血癥、胃腸道出血等并發(fā)癥。 SFSS的病因及發(fā)病機制至今尚未完全清楚,目前普遍認為主要是由于功能性肝臟體積過小,從而引起持續(xù)性門靜脈高壓,持續(xù)性的門靜脈高灌注進一步導致肝竇內皮細胞損傷繼而造成肝實質細胞的損傷,影響肝細胞再生,最終導致肝功能不全。除此之外,還可能與其他多種因素有關,如移植物的大小、潛在病變、肝再生能力、流入道和流出道情況,受者病情輕重等等。正因為如此,SFSS的防治措施也一直在不斷的積極探索之中。 目前預防SFSS的策略主要集中在以下3個方面[6]:(1)增加肝臟移植物的體積；(2)控制門靜脈的壓力；(3)改進肝臟靜脈回流的方式。大部分學者普遍認同門靜脈高灌注和門靜脈高壓對小體積肝臟移植物有破壞作用,因此降低門靜脈灌流和壓力也成為預防SFSS的主要手段。而目前常用的手術方式包括脾動脈結扎術和門-體分流術。 脾動脈結扎、栓塞操作簡單,創(chuàng)傷小,對門靜脈灌注壓力的調節(jié)作用有限,可并發(fā)術后門靜脈血栓和腹腔感染；門靜脈分流能有效調節(jié)門靜脈的灌注壓,近年來,一些研究者采用門靜脈-腔靜脈分流來防治SFSS的發(fā)生。但是,有學者發(fā)現門-腔靜脈分流在活體肝移植術后數周內雖能有效減少SFSS的發(fā)生,并能提高移植肝的存活率,后期卻出現移植物逐漸萎縮、肝再生受到抑制的現象。 由此看來,在擴大肝切除或部分肝移植術后維持適當的門靜脈灌注壓在余肝再生過程中發(fā)揮了重要作用。本實驗擬建立SFSS實驗動物模型,并實施門-腔分流調控門靜脈的灌注血流,觀察門靜脈灌注壓對肝再生的影響。 目的 1.對兔肝臟分葉及附屬管道進行應用解剖學研究,并在此基礎上建立兔SFSS動物模型。 2.在SFSS動物模型的基礎上實施門-腔分流,觀察門靜脈灌注壓力的變化對肝功能和肝再生的影響。方法 1.兔肝臟及其附屬管道應用解剖學研究：選擇由昆明醫(yī)科大學實驗動物中心提供的健康成年日本大耳兔20只,體重范圍1.5kg-2.5kg。經耳緣靜脈按照1ml/kg注射3%戊巴比妥鈉麻醉。固定其四肢,腹部褪毛。(1)肝臟在體與離體解剖學觀察：取正中切口進入腹腔,離斷肝周韌帶,充分暴露肝臟并對其進行在體解剖學觀察。處死動物后,經門靜脈灌注生理鹽水沖洗血管。完整取下肝臟對其進行離體解剖學觀察。(2)全肝及各肝葉稱重,并計算各肝葉占全肝臟及兔體重百分比。(3)制作門靜脈和肝靜脈管道灌注標本。經腸系膜上靜脈插管結扎其尾側端,注入管道鑄型劑(自凝造牙粉和自凝牙托水按10g：10ml混合)制作門靜脈管道鑄型標本。結扎肝后腔靜脈頭側、尾側端,同上注射管道鑄型劑制作肝靜脈管道鑄型標本。待管道鑄型標本硬固后,取出肝臟。將其放置于20%氫氧化鈉溶液中浸泡腐蝕顯露管道標本。SPSS13.0軟件對數據進行描述性分析,計量數據結果以“均數±標準差(x±s)”表示。 2. SFSS動物模型的建立：將42只兔隨機分為5組,分別為假手術組(n=2),A組(n=10)為切除“左中葉+右中葉”組,B組(n=10)為切除“左外葉+左中葉+右中葉”組,C組(n=10)為切除“尾狀葉+左中葉+右中葉+右外葉”組,D組(n=10)為切除“左外葉+左中葉+右中葉+右外葉”組。經耳緣靜脈按照1ml/kg注射3%戊巴比妥鈉麻醉。固定四肢,腹部褪毛,碘伏消毒,鋪無菌單。取腹部正中切口進入腹腔。應用無血切肝技術,分別對A組、B組、C組及D組實施肝葉切除術。觀察術后兔活動及進食情況。術前和術后存活兔分別于第1、3、5、7天麻醉后經耳緣靜脈采血留取標本檢測ALT、TB、PT,然后開腹獲取肝組織標本行組織病理學檢查。術后死亡兔行肝臟剖檢。計量數據結果以“均數±標準差(x±s)”表示,應用SPSS13.0統(tǒng)計軟件對數據進行F檢驗和x2檢驗,檢驗水準a=0.05。 3.在SFSS的基礎上實施門-腔分流,調控門靜脈灌注壓力,觀察肝再生：將120只兔隨機分為3組,分別為E組(n=40)為“SFSS模型”組,F組(n=40)為‘'SFSS基礎上的門-腔分流”組,G組(n=40)為“門-腔分流調控”組。同樣經耳緣靜脈以1ml/kg注射3%戊巴比妥鈉麻醉,之后固定四肢,腹部褪毛,碘伏消毒,鋪無菌單。取腹部正中切口進入腹腔。應用無血切肝技術,分別對E組、F組及G組實施“左外葉+左中葉+右中葉”的肝葉切除術,同時對F組、G組進一步行改良后的門-腔靜脈分流術。觀察術后兔生存情況。術前和術后E組、F組存活兔分別于第1、3、5、7、9、11、13、15、17、19天再次開腹測量門靜脈壓力值并對余肝進行稱重觀察肝再生。而G組存活兔則統(tǒng)一于術后第7天開腹結扎封閉分流口后分別于第9、11、13、15、17、19天開腹測量門靜脈壓力值及余肝重量進一步觀察調控門-腔分流后的門靜脈壓力值變化對肝再生的影響。計量數據結果以“均數±標準差(x±s)”表示,應用SPSS13.0統(tǒng)計軟件對生存率組間比較行Log Rank檢驗,組間差異采用重復測量方差分析,以P0.05有統(tǒng)計學意義。 結果 1.兔肝肝裂明顯,依據肝葉形態(tài)、肝裂走形和門靜脈主干分支形式將兔肝臟分為五葉：尾狀葉、左外葉、左中葉、右中葉和右外葉。全肝質量為(60.13±16.11)g,占其質量百分比為(2.88±0.06)%,各肝葉質量分別為(3.93±1.13)g、(15.93±3.50)g、(14.83±3.31) g、(15.08±4.34) g、(12.08±3.55) g占全肝質量百分比分別為(6.22±1.02)%、(25.44±2.55)%、(23.72±2.71)%、(24.15±5.21)%、(19.32±4.23)%。左中葉和右中葉根部肝組織融合,其余各肝葉相對獨立,尾狀葉包括相對獨立的乳頭突和尾狀突兩部分。各肝葉有相對獨立的Glisson系統(tǒng)且肝靜脈走行于肝蒂內 2.根據上述測得的各肝葉質量及不同肝葉組合占全肝質量百分比數據,A、B、C、D四組保留肝葉占全肝質量百分比分別約為50%、28%、25%和6%,術后一周存活率分別為100%、50%、20%和0%。術后A組與假手術組比較,ALT、TB、PT數值變化不明顯(P0.05)；B、C兩組ALT、TB、PT在術后第1天明顯升高,第3天達到高峰,隨后逐漸降低,至第7天仍較對照組和A組增高(P0.05)；而B、C兩組數值術后第1、3、5、7天組間比較,C組雖較B組有所升高,但兩者差值較小(P0.05)。剖檢術后7d內自然死亡的兔肝,可見不同程度的淤血、淤膽、出血,余肝腫脹、壞死,胃腸道淤血,渾濁腹水；相比之下,存活7d者則可見肝體積明顯增大,與周圍組織致密粘連,邊緣圓鈍,似圓盤狀,腹腔內少量清亮腹水。肝組織病檢可見：A組匯管區(qū)無明顯有絲分裂象和多倍體細胞；B、C兩組肝組織淤血水腫明顯,可見中央小靜脈及肝竇擴張,匯管區(qū)中性粒細胞浸潤明顯,毛細血管內膽栓形成,肝實質可見點狀壞死,肝細胞氣球樣變和脂肪變性,隨著時間延長,肝實質細胞增生逐漸活躍,并逐步出現多倍體細胞,但肝實質細胞增生比A組延遲。 3.根據上述實驗結果,選擇實驗B組建立較穩(wěn)定的SFSS動物模型,并在其基礎上實施門-腔分流手術。術后19天E、F、G三組存活率分別為10%、30%和60%。術后E組表現為典型的SFSS特點,余肝組織充血水腫,肝實質破壞較多,肝再生表現不明顯,并且隨著時間的延長,余肝逐漸出現不能程度的壞死、胃腸道淤血等情況,實驗動物相繼死亡,最后僅4只存活時間達到19天。而F、G兩組因在擴大肝切除的基礎上實施門-腔分流手術,術后短時間內降低了門靜脈的灌注壓力,從而減少了過高的門靜脈灌注血流量對余肝組織的損壞作用,余肝組織充血水腫程度較輕,術后動物死亡率均較E組明顯下降,生存時間有所延長,并表現出不同程度的肝臟再生。至術后7天,因F組的門靜脈壓力持續(xù)降低,對余肝再生的機體代謝支持逐漸減少,從而導致肝臟再生減緩,肝功能逐漸惡化,則繼續(xù)出現動物死亡現象,最后有12只存活時間達到19天。因此,G組在術后第7天又再次開腹結扎門-腔分流口后,門靜脈灌注壓力再次逐漸升高,從而滿足了余肝持續(xù)再生的機體代謝需求,動物死亡率較F組進一步下降,肝臟再生明顯,最后有24只存活時間達到19天。 結論 1.兔肝臟解剖學既與犬、鼠、豬等哺乳類動物類似,又具有其自身的特點。本實驗通過對兔肝臟在體、離體解剖學和肝臟管道灌注標本的觀察,規(guī)范了文獻中對兔肝解剖學的混亂描述。并在此基礎上,通過測量全肝及各肝葉質量,計算出各肝葉所占全肝質量百分比,為在該方面的基礎研究提供數據支持。 2.根據兔肝解剖學特點行不同質量百分比肝葉切除術制作SFSS模型,當保留右外葉和手術切除相對困難的尾狀葉后,即余肝占全肝質量百分比約為28%時,術后存活率較高并可表現出典型的SFSS。而且兔體積適中,圍手術期管理和手術操作相對簡便,可作為研究SFSS發(fā)病機制及防治措施較為理想的實驗動物模型。 3.門-腔分流術后可明顯降低門靜脈灌注壓,促進肝再生,有效延長術后動物生存時間；過度分流,門靜脈灌注壓低于正常值則肝再生受到抑制。在擴大肝切除術后發(fā)生SFSS情況下,采用門-腔分流手術維持適度增高的門靜脈灌注壓可明顯促進肝再生。
[Abstract]:background
Small liver syndrome (small-for-size, syndrome, SFSS) occurs in hepatic resection or partial orthotopic liver transplantation after due to liver volume is too small to function can not meet the demand of the metabolic receptor a clinical syndrome.SFSS is most common in adult adult living donor liver transplantation for partial orthotopic liver volume is small, may lead to the receptor of portal vein blood flow perfusion and high persistence of portal hypertension, which caused a series of serious pathophysiological disorder. Its clinical manifestations include persistent postoperative jaundice, coagulopathy, portal hypertension, refractory ascites, which further induce sepsis, gastrointestinal bleeding and other complications.
The etiology and pathogenesis of SFSS has not yet entirely clear, the prevailing view was mainly due to functional liver volume is too small, resulting in persistent portal hypertension, persistent portal hyperperfusion and further lead to injury of sinusoidal endothelial cells and caused liver parenchyma cell damage, liver cell regeneration, eventually lead to liver function. In addition, it may be related to other factors, such as graft size, potential lesions, liver regeneration, inflow and outflow tract, the severity and so on. Because of this, SFSS control measures have also been active in exploring.
At present, SFSS prevention strategies mainly focus on the following 3 aspects: (1) [6] increased liver graft volume; (2) control of portal vein pressure; (3) improvement of hepatic venous return way. Most scholars generally agree on portal vein perfusion and portal hypertension have a destructive effect on the small volume of liver graft plants, therefore reduce portal vein pressure and perfusion has become the main means of preventing SFSS. And the current commonly used surgical methods including splenic artery ligation and portosystemic shunt.
Ligation of splenic artery embolization, simple operation, small trauma, portal vein perfusion pressure regulation is limited, can be complicated with postoperative portal vein thrombosis and infection of abdominal cavity; portal vein shunt can effectively regulate the portal vein perfusion pressure, in recent years, some researchers use the portal vein to vena cava shunt occurred to the prevention and treatment of SFSS. However, there are researchers found that portacaval shunt in living donor liver transplantation after a few weeks can effectively reduce the incidence of SFSS, and can improve the survival rate of liver transplantation, but late graft gradually atrophic, inhibited the phenomenon of liver regeneration.
Therefore, to maintain the proper pressure in the portal vein perfusion during liver regeneration than plays an important role in hepatic resection or liver transplantation. This study was to establish the animal model of SFSS, and the implementation of perfusion portocaval shunt regulation of portal vein, to observe the effect of portal vein perfusion pressure on liver regeneration.
objective
1. the applied anatomy of the rabbit's liver lobules and accessory pipes was carried out, and a rabbit SFSS animal model was established on the basis of this.
2. the portal cavity shunt was carried out on the basis of the SFSS animal model, and the effects of the changes of portal vein perfusion pressure on liver function and liver regeneration were observed.
Study on applied anatomy of 1. rabbit liver and its affiliated pipeline: from the experimental animal center of Kunming Medical University health 20 adult Japanese white rabbits, weight 1.5kg-2.5kg. intravenously with 1ml/kg injection of 3% pentobarbital anesthesia. The fixation of limbs, abdominal faded hair. (1) the liver in vivo and in vitro anatomy observation: the median incision into the abdominal cavity, severed ligaments around the liver, liver and fully exposed were observed in vivo anatomy on it. Kill the animal, via the portal vein infusion of saline irrigation vessels. The complete pathological anatomy observation in vitro. (2) and the total hepatic lobe of the liver and liver and weighing, calculation the rabbit liver weight percentage. Ye Zhanquan (3) made of portal vein and hepatic vein perfusion specimens. After ligation of the superior mesenteric vein intubation caudal, injection pipe casting agents (self curing polymer and self curing denture water by 10g:10ml Mixed) making cast specimens of portal vein ligation of hepatic vena cava pipeline. Cephalic, caudal, ditto injection pipe casting agents making cast specimens of hepatic vein duct. To cast specimens of hardened pipe, remove the liver. Placed in 20% sodium hydroxide solution immersion exposure pipe specimens.SPSS13.0 for descriptive analysis of data the results of measurement data, the standard deviation (x + s) ".
2. SFSS鍔ㄧ墿妯″瀷鐨勫緩绔嬶細灝，

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