提高肝移植邊緣供肝質(zhì)量的研究進(jìn)展

關(guān)　正(綜述)，呂　毅(審校)

(西安交通大學(xué)醫(yī)學(xué)院第一附屬醫(yī)院 a.麻醉科,b.肝膽外科，西安 710061)

提高肝移植邊緣供肝質(zhì)量的研究進(jìn)展

關(guān)正a(綜述)，呂毅b※(審校)

(西安交通大學(xué)醫(yī)學(xué)院第一附屬醫(yī)院 a.麻醉科,b.肝膽外科，西安 710061)

摘要：由于等待肝移植的患者越來越多，而可用于移植的肝臟嚴(yán)重短缺，故提高邊緣供肝的質(zhì)量對(duì)于增加供肝來源具有重要意義。提高邊緣供肝質(zhì)量的最新研究包括對(duì)供體的藥物干預(yù)及循環(huán)支持、肝臟保存液的改進(jìn)、機(jī)械灌注的嘗試和更加合適的保存溫度以及氣態(tài)氧灌注的進(jìn)一步改進(jìn)。但其中大部分都局限在動(dòng)物研究階段，其與臨床應(yīng)用還存在一定的差距。該文就提高邊緣供肝質(zhì)量的研究新進(jìn)展予以綜述。

關(guān)鍵詞：邊緣供肝；肝移植；心臟死亡后器官捐獻(xiàn)；機(jī)械灌注

由于近年來供肝的短缺，嚴(yán)重影響了肝移植的發(fā)展，故人們開始使用老年、脂肪變性以及心臟死亡后器官捐獻(xiàn)(donation after cardiac death，DCD)肝臟來進(jìn)行肝移植；使用這類肝臟進(jìn)行肝移植后原發(fā)移植物無功能，出現(xiàn)膽管系統(tǒng)并發(fā)癥且再次肝移植發(fā)生率較高，故這類肝臟被稱為邊緣供肝[1-3]。Fernandez-Merino等[1]研究發(fā)現(xiàn)，進(jìn)行肝移植患者的生存率與供體年齡呈負(fù)相關(guān)，移植肝功能也有著同樣的規(guī)律。脂肪變性肝臟由于嚴(yán)重的線粒體損傷、能量代謝損害以及氧化應(yīng)激反應(yīng)的增強(qiáng)導(dǎo)致其對(duì)缺血/再灌注損傷的耐受性較差[4]。DCD供肝由于熱缺血時(shí)間較長，導(dǎo)致了膽管系統(tǒng)并發(fā)癥發(fā)生率明顯升高[5]。盡管邊緣供肝存在以上的缺陷，但由于肝移植供肝的短缺及等待移植患者的增加，邊緣供肝占肝移植供肝的比例仍在逐年升高[6]。因此，如何提高邊緣供肝的質(zhì)量，增加供肝的來源，成為目前研究的熱點(diǎn)。

1對(duì)肝臟供體的干預(yù)

對(duì)肝臟供體的干預(yù)包括藥物干預(yù)和循環(huán)輔助。前列腺素E1和蛋白酶抑制劑是近年來研究較為廣泛的干預(yù)藥物；前者通過下調(diào)黏附分子表達(dá)，抑制中性粒細(xì)胞向內(nèi)皮細(xì)胞的黏附從而對(duì)抗肝臟缺血再灌注損傷[7]。研究表明，給予大鼠10 μg/L的前列腺素E1，心搏停止30 min后切取肝臟，發(fā)現(xiàn)肝臟質(zhì)量明顯提高，損傷明顯減輕[8-9]。蛋白酶抑制劑通過減輕脂質(zhì)過氧化反應(yīng)從而明顯降低肝臟再灌注后的轉(zhuǎn)氨酶水平，縮小肝臟壞死面積[10]。利用體外膜肺氧合對(duì)循環(huán)進(jìn)行輔助，可恢復(fù)由于心搏停止而導(dǎo)致的肝臟細(xì)胞內(nèi)能量的損失；2005年，體外膜肺氧合第一次在DCD供體上作為切取肝臟前的循環(huán)輔助應(yīng)用并獲得了成功[11]。Noormohamed等[12]在豬心搏停止后30 min開始給予2 h的體外膜肺氧合，之后切取肝臟，發(fā)現(xiàn)亦可提高供肝的質(zhì)量。

2單純低溫保存的改進(jìn)

肝臟保存液中加入蛋白酶抑制劑、表皮生長因子和白蛋白可明顯提高肝臟保存的效果[13-16]。蛋白酶抑制劑硼替佐米通過提高肝臟抗氧化酶水平來保護(hù)慢性酒精性肝損害[13]。在UW液(the university of wisconsin solution，UW)中加入小劑量硼替佐米(100 nmol/L)，對(duì)有脂肪變性的肝臟在4 ℃下冷保存24 h后，常溫再灌注2 h，發(fā)現(xiàn)肝臟質(zhì)量明顯提高，損傷明顯減輕[14]。Zaouali等[15]給IGL-1液(institute george lopez solution，IGL)中加入10 μg/L的表皮生長因子1，對(duì)脂肪變性的肝臟進(jìn)行離體24 h單純低溫保存；結(jié)果發(fā)現(xiàn)，與不加表皮生長因子1的肝臟相比，再灌注2 h后，其轉(zhuǎn)氨酶水平下降，膽汁產(chǎn)生增加，且線粒體損傷減輕，氧化應(yīng)激反應(yīng)減輕。在UW液中加入白蛋白發(fā)現(xiàn)，其具有更好的肝臟保護(hù)作用[16]；原位末端轉(zhuǎn)移酶標(biāo)記技術(shù)染色發(fā)現(xiàn)其完全抑制了細(xì)胞凋亡[17]。

3機(jī)械灌注下肝臟保存的嘗試

機(jī)械灌注保存通過下調(diào)促炎因子(如腫瘤壞死因子α、白細(xì)胞介素8、細(xì)胞間黏附分子1)的活性從而提高保存器官的功能[18]。Shigeta等[19]對(duì)豬DCD肝臟分別通過門靜脈和肝動(dòng)脈進(jìn)行持續(xù)機(jī)械灌注；與單純低溫保存相比，灌注液中天冬氨酸轉(zhuǎn)氨酶及乳酸脫氫酶水平明顯降低，且肝臟再灌注后存活率明顯提高。Fondevila等[20]對(duì)DCD肝臟采用低溫機(jī)械灌注與低溫保存的效果進(jìn)行了比較，發(fā)現(xiàn)接受低溫機(jī)械灌注移植肝的實(shí)驗(yàn)豬5 d生存率提高，生化檢查凝血酶原時(shí)間明顯改善。Vairetti等[21]發(fā)現(xiàn)，脂肪肝機(jī)械灌注保存效果明顯好于低溫保存。Guarrera等[22]第一次將低溫機(jī)械灌注肝臟保存應(yīng)用于臨床，該小組對(duì)比了20例分別進(jìn)行低溫機(jī)械灌注和單純低溫保存的肝臟質(zhì)量，發(fā)現(xiàn)前者早期移植物無功能比例下降，膽管系統(tǒng)的并發(fā)癥亦明顯降低。機(jī)械灌注的缺點(diǎn)是灌注過程中對(duì)肝竇內(nèi)皮細(xì)胞的損傷，在灌注液中加入羥乙基淀粉[23]或多巴胺[24]可減輕其對(duì)肝竇內(nèi)皮細(xì)胞的損傷，尤其是對(duì)DCD供肝。

4邊緣供肝保存的合適溫度研究

對(duì)供肝進(jìn)行-4 ℃的超低溫保存，與傳統(tǒng)的4 ℃低溫保存相比，肝臟再灌注后天冬氨酸轉(zhuǎn)氨酶、丙氨酸轉(zhuǎn)氨酶及乳酸脫氫酶水平明顯下降，組織學(xué)檢查未發(fā)現(xiàn)肝細(xì)胞、膽管及血管的損傷[25]。Charrueau等[26]分別于4 ℃、1 ℃及-0.5 ℃對(duì)遭遇熱缺血15 min的肝臟進(jìn)行24 h低溫保存，發(fā)現(xiàn)1 ℃下DCD肝臟保存效果最好，這與1 ℃下肝竇內(nèi)皮細(xì)胞的損傷最輕有關(guān)。對(duì)DCD肝臟的機(jī)械灌注保存研究發(fā)現(xiàn)，保持灌注液溫度在20 ℃左右時(shí)(亞低溫)，再灌注后丙氨酸轉(zhuǎn)氨酶釋放量最低、門靜脈阻力最小、膽汁產(chǎn)生量最大[27]，肝臟損傷明顯減輕[28]、腺苷三磷酸(adenosine triphosphate，ATP)水平明顯升高[29]。在對(duì)脂肪變性肝臟的保存中亦發(fā)現(xiàn)，亞低溫機(jī)械灌注有減輕肝臟損傷，提高肝臟功能的作用[30]。

5氣態(tài)氧灌注保存的研究進(jìn)展

氣態(tài)氧灌注肝臟保存的效果在動(dòng)物供肝保存中已得到了驗(yàn)證[31-32]，但由于其存在移植后移植物氣栓的可能而未得到應(yīng)用。Treckmann等[33]利用氣態(tài)氧逆行灌注聯(lián)合單純低溫保存，使5例由于熱缺血損傷而準(zhǔn)備舍棄的肝臟作為供肝成功地進(jìn)行了移植；在2年的隨訪中，接受這些肝臟患者的肝功能良好，未遭受再次肝移植。在Minor等[34]的研究中，對(duì)脂肪變性肝臟在低溫保存前給予90 min氣態(tài)氧灌注，結(jié)果肝酶釋放、脂質(zhì)過氧化、細(xì)胞凋亡及細(xì)胞自我吞噬減少，對(duì)氨的代謝、肝臟顯微形態(tài)及整體代謝狀況提高。Koetting等[35]在DCD肝臟低溫保存期間給予氣態(tài)氧灌注，發(fā)現(xiàn)肝細(xì)胞損傷明顯減少。以上研究均提示氣態(tài)氧灌注作為肝臟保存的輔助手段，具有一定的應(yīng)用價(jià)值。

6預(yù)防器官缺血/再灌注損傷的研究進(jìn)展

DCD供肝由于反復(fù)的缺血/再灌注損傷而導(dǎo)致移植物存活率下降，缺血性膽管病的發(fā)病率增加；而脂肪變性供肝及老年供肝易受缺血/再灌注損傷的影響，因此，將預(yù)防器官缺血/再灌注的方法應(yīng)用于對(duì)邊緣供肝獲取前供體的干預(yù)，有可能提高邊緣供肝的質(zhì)量。

6.1遠(yuǎn)端缺血預(yù)處理在器官缺血/再灌注損傷中的應(yīng)用遠(yuǎn)端缺血預(yù)處理是指對(duì)一個(gè)臟器的缺血預(yù)處理會(huì)引起遠(yuǎn)隔臟器的缺血耐受[36]。臨床上采用對(duì)一側(cè)肢體進(jìn)行5 min缺血5 min再灌注的缺血/再灌注處理，使機(jī)體產(chǎn)生啟動(dòng)因子(如腺苷、一氧化氮、緩激肽、阿片類物質(zhì)等)，其通過體液因子介導(dǎo)或神經(jīng)通路(如蛋白激酶C和ATP敏感性鉀離子通道)達(dá)到對(duì)遠(yuǎn)隔器官的保護(hù)作用。Kanoria等[37]研究發(fā)現(xiàn)，對(duì)兔后肢行3個(gè)循環(huán)的5 min缺血5 min再灌注，能降低全肝缺血/再灌注損傷。但是，如何實(shí)現(xiàn)遠(yuǎn)端缺血預(yù)處理對(duì)靶器官的最佳保護(hù)強(qiáng)度(包括最佳循環(huán)次數(shù)，缺血時(shí)間等)還需要進(jìn)一步研究；遠(yuǎn)端缺血預(yù)處理是否會(huì)造成局部組織損傷也還需要大量臨床試驗(yàn)來驗(yàn)證。

6.2嘌呤能信號(hào)轉(zhuǎn)導(dǎo)在器官缺血/再灌注損傷中的應(yīng)用經(jīng)由ATP參與的嘌呤能信號(hào)轉(zhuǎn)導(dǎo)及腺苷受體是目前對(duì)抗器官缺血/再灌注損傷的最新治療靶點(diǎn)[38]。在器官缺血情況下，細(xì)胞釋放ATP到細(xì)胞外，激活細(xì)胞外的ATP受體，后者激活炎性反應(yīng)，導(dǎo)致器官在缺血狀態(tài)下的損傷；同時(shí)，釋放到細(xì)胞外的ATP又會(huì)很快地轉(zhuǎn)化為腺苷，其作用與ATP受體相反，具有器官保護(hù)作用[39]。因此，作用于此代謝通路上的藥物被認(rèn)為具有對(duì)抗器官缺血/再灌注損傷的作用(如ATP受體拮抗劑等)；加速ATP向腺苷轉(zhuǎn)化的藥物有ATP雙磷酸酶、核苷酸酶、缺氧誘導(dǎo)因子等[40]。但目前還需要大量臨床隨機(jī)對(duì)照試驗(yàn)的驗(yàn)證，才能使其進(jìn)入臨床應(yīng)用。

6.3血紅素氧合酶1/一氧化碳體系在器官缺血/再灌注損傷中的應(yīng)用血紅素氧合酶1/一氧化碳體系分布于所有的組織和器官，其具有抗氧化應(yīng)激、抗細(xì)胞凋亡和抗炎癥反應(yīng)的作用[41]。血紅素氧合酶1/一氧化碳體系對(duì)冷保存肝臟具有保護(hù)作用，可顯著延長其保存時(shí)間,并通過減少巨噬細(xì)胞浸潤、抗細(xì)胞凋亡作用對(duì)移植肝臟出現(xiàn)的缺血/再灌注產(chǎn)生保護(hù)作用，提高器官移植后的存活率[42]。

7小結(jié)

提高邊緣供肝的質(zhì)量，可明顯增加肝移植供肝的來源，緩解供肝短缺的問題。近年來人們嘗試了多種新的保存藥物、保存技術(shù)，甚至各種保存溫度對(duì)邊緣供肝進(jìn)行了保存，取得了一定的進(jìn)展。另外，在對(duì)抗器官的缺血再灌注損傷方面，近年來亦有了一些新的進(jìn)展，但距進(jìn)入臨床應(yīng)用還存在一定的距離，且目前仍未見到對(duì)于老年供肝保存的研究，還需進(jìn)一步深入研究。

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The Research Progress of Improving the Quality of the Marginal Donor Liver in Liver Transplantation

GUANZhenga，LVYib.

(a.DepartmentofAnesthesiology,b.DepartmentofHepatobiliarySurgery,theFistHospitalAffiliatedtoXi′anJiaotongUniversity,Xi′an710061，China)

Abstract:There is a contradiction between the increasing patients waiting for liver transplantation and the shortage of donor liver,it is important to increasing the number of donor livers by improving the quality of the marginal donor liver.The new researches of improving the quality of the marginal donor liver include:the medication and the circulation support for the donor,the modified preserving solution,the use of machine perfusion preservation,the research of the best temperature for liver graft storage and the improving of gaseous oxygen perfusion.However,most of the researches are still in animal experiment stage,where a certain gap still exist to clinical application.Here is to make a review of the recent progress in improving the quality of marginal donor liver.

Key words：Marginal donor liver； Liver transplantation； Donation after cardiac death； Machine perfusion

收稿日期：2014-04-10修回日期：2014-08-07編輯：鄭雪

doi：10.3969/j.issn.1006-2084.2015.04.037

中圖分類號(hào)：R657.3

文獻(xiàn)標(biāo)識(shí)碼：A

文章編號(hào)：1006-2084(2015)04-0671-04