醫(yī)學(xué)免費(fèi)論文:NF�拨蔅及其相關(guān)炎性因子致大鼠移植肝冷保存損傷的機(jī)制

醫(yī)學(xué)免費(fèi)論文:NF�拨蔅及其相關(guān)炎性因子致大鼠移植肝冷保存損傷的機(jī)制

【摘要】 目的 通過大鼠移植肝在不同冷保存時間再灌注后核因子�拨蔅(nuclear transcription factor�拨蔅, NF�拨蔅)及其相關(guān)炎性因子的激活情況，判斷冷保存時間、NF�拨蔅激活以及移植肝功能損傷三者之間的聯(lián)系。方法 用Wistar大鼠建立原位肝移植模型，假手術(shù)組(A組)作為對照組，觀察大鼠移植肝在4℃ UW液中保存45min(B組)、120min(C組)、180min(D組)以及再灌注3h后肝組織、血清中NF�拨蔅、腫瘤壞死因子�拨�(TNF�拨�)、白細(xì)胞介素��1β(IL��1β)和肝損傷指標(biāo)丙氨酸轉(zhuǎn)氨酶(ALT)、天冬氨酸轉(zhuǎn)氨酶(AST)和肝細(xì)胞凋亡的變化情況。結(jié)果 隨著移植肝冷保存時間的延長NF�拨蔅逐漸激活(P<0.05)，并使肝組織內(nèi)TNF�拨�、IL��1β水平上調(diào)(P<0.05);再灌注后，NF�拨蔅近一步激活(P<0.05)，肝組織中TNF�拨�、IL��1β進(jìn)一步上調(diào)(P<0.05)，ALT、AST和肝細(xì)胞凋亡指數(shù)明顯升高(P<0.05)，肝功能損害加重。結(jié)論 NF�拨蔅在供肝的冷保存階段隨時間的延長呈誘導(dǎo)性激活，再灌注后呈爆發(fā)式激活，并通過上調(diào)其靶基因TNF�拨�、IL��1β的轉(zhuǎn)錄產(chǎn)生肝臟損傷，這可能是肝臟冷保存再灌注損傷發(fā)生的重要機(jī)制。

【關(guān)鍵詞】 肝移植;器官保存;冷保存再灌注損傷;核因子�拨蔅

The effect of NF�拨蔅 and correlated inflammatory factors

in rat donor liver after cold preservation

JIANG An, LIU Feng, LIU Chang, SONG Yu�瞝ong, MENG Ke�瞱ei, L�a Yi

1. Department of Hepatobiliary Surgery, the First Affiliated Hospital, Medical School of Xi�餫n Jiaotong University, Xi�餫n 710061; 2. Department of General Surgery,

the Second Affiliated Hospital, Medical School of Xi�餫n Jiaotong University, Xi�餫n 710004;3. Department of Hepatobiliary Surgery, Qianfoshan Hospital, Ji�餹an 250014;

4. Department of Anesthesiology, Shaanxi Provincial People�餾 Hospital, Xi�餫n 710068;5. Department of Hepatobiliary Surgery, Yuhuangding Hospital, Yantai 264000, China

ABSTRACT: Objective To establish a model of rat orthotopic liver transplantation and investigate the relationship among cold preservation time, activation of nuclear transcription factor�拨蔅 (NF�拨蔅) and donor preservation injury after liver transplantation. Methods Orthotopic liver transplantation was performed in Wistar rats which were randomly divided into the following groups according to the different duration of liver cold storage in UW solution: group A (sham operation, n=10), group B (45min cold storage group, n=10), group C (120min cold storage group, n=10), and group D (180min cold storage group, n=10). The expression of NF�拨蔅 in liver before and after transplantation was measured by electrophoretic mobility shift assays; protein expression of tumor necrosis factor�瞐lpha (TNF�拨�) and interleukin��1 beta (IL��1β) in the liver was measured by immunohistochemistry; the serum TNFα and IL��1β, alanine aminotransferase (ALT), aspartate aminotransferase (AST) and hepatic cell apoptosis were examined. Results With extended cold storage duration, the activity of NF�拨蔅 in the donor liver increased (P<0.05, group D vs. groups A, B and C). TNF�拨� and IL��1β levels also increased (P<0.05, group D vs. groups A, B and C). Donor liver reperfusion injury was gradually aggravated with the prolonging of graft cold preservation. Both the serum TNF�拨� and IL��1β levels increased highly (P<0.05 group A vs. groups B, C and D),and the expression of NF�拨蔅 in the liver significantly increased (P<0.05, group A vs. groups D, B and C) with gradual prolonging of graft cold preservation time. The serum ALT and AST level and apoptosis index level elevated greatly (P<0.05, group A vs. groups D, B and C). Conclusion NF�拨蔅 of donor liver was activated inductively in cold preservation phase and activated explosively in reperfusion phase. The longer cold preservation time was, the higher NF�拨蔅 level in donor liver became. NF�拨蔅 led to the expression of TNF�拨� and IL��1β in donor liver. Inflammatory factors are one of the most important mechanisms for the donor liver injury after liver transplantation醫(yī).學(xué).全.在.線52667788.cn.

KEY WORDS: liver transplantation; organ preservation; cold preservation�瞱arm reperfusion injury; nuclear transcription factor�拨蔅

移植肝冷保存再灌注損傷(cold preservation�瞱arm reperfusion injury, P/RI)是移植肝功能不良(poor graft function, PGF)和原發(fā)性無功能(primary nonfunction, PNF)的直接原因[1]。PGF及其嚴(yán)重形式PNF的發(fā)生率分別占移植總數(shù)的5%～10%[2]和1.4%～8.5%[3]。PNF占術(shù)后1周內(nèi)再次肝移植病因的81%和再次肝移植總病因的21.7%[1]，危害嚴(yán)重。冷保存時間超過20h，PNF發(fā)生率是保存4h以內(nèi)的4倍[4]，所以冷保存時間是PNF的獨(dú)立預(yù)測因子[5]。而且，P/RI還可導(dǎo)致多器官功能不全和膽管非吻合口狹窄等并發(fā)癥[6]，嚴(yán)重影響患者預(yù)后。因此，如何最大限度地減輕供肝冷保存損傷、提高移植物成活率是當(dāng)今肝移植領(lǐng)域面臨的重要課題。本實(shí)驗(yàn)通過研究移植肝P/RI過程中核轉(zhuǎn)錄因子�拨蔅(nuclear transcription factor�拨蔅, NF�拨蔅)及其相關(guān)炎性因子的表達(dá)變化，探討NF�拨蔅在移植肝冷保存中的作用機(jī)制。

1 材料與方法

1.1 動物模型的制作

清潔級健康成年封閉群雄性Wistar大鼠70只(第四軍醫(yī)大學(xué)實(shí)驗(yàn)動物中心提供)，體重(272±31)g。術(shù)前禁食12h，不禁飲，清潔手術(shù)。氯胺酮100mg/kg體重腹腔注射麻醉。采用改良兩袖套法建立大鼠全血供原位肝移植模型[7]。

1.2 實(shí)驗(yàn)動物分組

依據(jù)供肝冷保存時間的不同，將Wistar大鼠隨機(jī)分為4組，每組10只：A組，假手術(shù)組;B組，冷保存45min再灌注3h組;C組，冷保存120min再灌注3h組;D組，冷保存180min再灌注3h組。供肝以4℃ UW液保存，冷保存結(jié)束前，留取乳頭葉腹側(cè)葉片。移植肝植入3h后，留取乳頭葉背側(cè)葉片，并取血留作標(biāo)本。

1.3 觀察指標(biāo)及檢測方法

1.3.1 肝組織NF�拨蔅水平的測定

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