【摘要】：目的： 單次劑量照射制備大鼠放射性肝纖維化實(shí)驗(yàn)動物模型，觀察造模后各個(gè)時(shí)期大鼠肝功能、肝臟標(biāo)本病理學(xué)形態(tài)、血清學(xué)細(xì)胞因子以及肝臟免疫組化指標(biāo)等改變。并初步探討大鼠造模后相關(guān)細(xì)胞因子及TGF-β_1/Smads信號傳導(dǎo)通路在放射性肝纖維化中的作用，揭示放射性肝纖維化的發(fā)病機(jī)制，為臨床預(yù)防和治療放射性肝纖維化提供理論基礎(chǔ)及實(shí)驗(yàn)依據(jù)。 方法： 將60只實(shí)驗(yàn)大鼠隨機(jī)分為正常對照組（A組）10只、照射組（B組）50只。除正常對照組外，，B組大鼠用10%水合氯醛0.3ml/100g全麻后，美國VARIN21-EX直線加速器照射大鼠右半肝，源皮距為100cm，劑量率600cGy/min，30Gy/單次，照射野2.5cm×2.5cm，每只動物右側(cè)胸腔及胃腸均用鉛塊遮擋，照射后等待大鼠自然蘇醒，A組大鼠只麻醉不進(jìn)行照射。 于照射后第2、4、8、12、26周末分批分別剖殺。10%水合氯醛腹腔麻醉（0.3ml/100g體重）后，稱重并記錄，自腹主動脈取血，室溫靜置30min，在3000r/min的條件下離心10min，收集上清液置入凍存管中，在-80℃條件下凍存?zhèn)溆茫缓髾z測大鼠血清中AST、ALT的活性，ELISA法測血清中TGF-β_1、TNF-α、IL-6、HA、PC-Ⅲ、LN含量，堿水解法檢測大鼠肝臟組織中Hyp含量；然后立即處死大鼠，迅速切開腹腔。A組10只大鼠于第4周末處死做對照。取右肝置于10%中性甲醛溶液中固定，常規(guī)石蠟包埋，行HE染色和Masson染色及免疫組化檢測肝臟組織TGF-β_1、Smad3/4/7、PDGF-BB、TIMP-1、MMP13蛋白表達(dá)情況。 結(jié)果： 與對照組（A組）比較，B組受照射肝組織表現(xiàn)出放射性肝炎及肝纖維化，照射組大鼠血清中AST，ALT酶活性和TGF-β_1水平自第4周升明顯升高（P＜0.05或P＜0.01），于第12周末達(dá)到高峰；照射組大鼠血清中HA、PC-Ⅲ、LN含量和肝臟組織中Hyp含量于照射后第4周顯著升高（P＜0.05或P＜0.01），一直呈上升趨勢，照射后26周值最高；病理結(jié)果顯示，大鼠肝臟病變呈慢性進(jìn)行性肝纖維化改變。免疫組化結(jié)果顯示，照射組大鼠肝臟組織中TGF-β_1、Smad3/4蛋白表達(dá)于照射后第2周開始升高（P＜0.05），照射后第12周達(dá)到高峰，26周開始下降；Smad7蛋白表達(dá)于照射后第2周明顯減少（P＜0.05），照射后第12周達(dá)最低水平。另外，照射組大鼠肝臟組織中PDGF-BB和TIMP-1蛋白表達(dá)于照射2周升高，照射后26周最高，MMP-13蛋白表達(dá)在照射后降低，26周時(shí)表達(dá)最少。 結(jié)論： 1.單次單劑量（30Gy）照射建立大鼠右肝放射性肝纖維化模型穩(wěn)定可靠，動物死亡率較低（15%左右），便于動態(tài)觀察放射性肝纖維化的變化規(guī)律，為防治放射性肝纖維化的實(shí)驗(yàn)研究提供一種科學(xué)、可行、實(shí)用的動物模型； 2.放射性肝纖維化與TGF-β_1/Smads信號傳導(dǎo)通路存在內(nèi)在聯(lián)系，TGF-β_1/Smads信號傳導(dǎo)通路改變可能是引起放射性肝纖維化的重要機(jī)制之一。
[Abstract]:Objective: to establish an experimental animal model of radiation hepatic fibrosis in rats by single dose irradiation, and observe the liver function and pathological morphology of liver samples in each period after the establishment of the model. Changes in serum cytokines and liver immunohistochemical markers. The role of related cytokines and TGF- 尾 1 / Smads signal transduction pathway in radiation-induced liver fibrosis in rats was also studied, and the pathogenesis of radiation hepatic fibrosis was revealed. To provide theoretical basis and experimental basis for clinical prevention and treatment of radioactive liver fibrosis. Methods: 60 experimental rats were randomly divided into normal control group (group A, n = 10) and irradiation group (group B, n = 50). In addition to the normal control group, rats in group B were treated with 10% chloral hydrate 0.3ml/100g general anesthesia. The right half liver was irradiated by VARIN21-EX linear accelerator in the United States. The skin distance was 100 cm, the dose rate was 600cGy 路min ~ (30) Gy / time, 2.5cm 脳 2.5 cm, and the right thorax and gastrointestinal tract of each animal were occluded by lead. Rats in group A were anesthetized and not irradiated after irradiation. At the end of the week, the abdominal anaesthesia (0.3ml/100g body weight) of 0.10% chloral hydrate was dissected in batches, weighing and recording. Blood was taken from the abdominal aorta, rest at room temperature for 30 min, centrifuged for 10 min under the condition of 3000r/min, and the supernatant was collected and placed in the frozen tube. After cryopreservation at -80 鈩

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