辛世杰

(中國(guó)醫(yī)科大學(xué)第一附屬醫(yī)院 血管/甲狀腺外科，遼寧 沈陽 110001)

自異體血管植入受者開始，受者免疫系統(tǒng)便開始持續(xù)不斷地攻擊移植血管，在反復(fù)打擊與抗打擊中移植血管逐漸發(fā)生變化——血管內(nèi)膜向心性增厚、中膜平滑肌細(xì)胞凋亡、外膜纖維化，最終導(dǎo)致移植器官缺血、功能衰竭，此被稱為移植物血管病，由于其可出現(xiàn)纖維脂質(zhì)斑塊、內(nèi)膜出現(xiàn)細(xì)胞外基質(zhì)沉積、炎細(xì)胞浸潤(rùn)等類似于動(dòng)脈粥樣硬化的改變，故亦稱作移植物動(dòng)脈硬化。據(jù)國(guó)際心肺移植協(xié)會(huì)(International Society for Heart and Lung Transplantation,ISHLT) 報(bào)道，心臟移植術(shù)后1、5、10年AV發(fā)病率逐年增高，分別為8%、30%、50%，已成為慢性排斥反應(yīng)期患者死亡的主要病因[1]。AV形成過程經(jīng)歷4個(gè)階段，第1階段：移植后數(shù)天到數(shù)周血管壁滲入炎癥細(xì)胞，受體針對(duì)移植物發(fā)生免疫反應(yīng)；第2階段：異體免疫反應(yīng)導(dǎo)致內(nèi)皮細(xì)胞損傷、剝脫，中膜平滑肌細(xì)胞凋亡，甚至可以觀察到整個(gè)中膜完全沒有平滑肌細(xì)胞，殘存彈力膜支架；第3階段：失去正常功能的損傷血管的重建；第4階段：新生內(nèi)膜中內(nèi)皮細(xì)胞、平滑肌細(xì)胞的失去控制的增殖，導(dǎo)致管腔狹窄、閉塞，最終導(dǎo)致移植物失去功能。在上述過程中免疫因素持續(xù)存在并始終參與，故其在AV中的作用由此可見一斑。

1 細(xì)胞免疫介導(dǎo)的損傷

當(dāng)血液復(fù)流后受者T細(xì)胞活化為效應(yīng)T細(xì)胞，后者發(fā)揮細(xì)胞毒作用直接殺傷移植血管細(xì)胞。T細(xì)胞需經(jīng)過識(shí)別供者主要組織相容性復(fù)合體 (major histocompatibility complex, MHC)及共刺激分子作用才能被完全活化，活化信號(hào)均由抗原提呈細(xì)胞(antigen presenting cell, APC)傳遞，樹突狀細(xì)胞、巨噬細(xì)胞等專職APC在這一過程中起主要作用，血管內(nèi)皮細(xì)胞等非專職APC亦參與其中。T細(xì)胞識(shí)別有直接識(shí)別和間接識(shí)別兩種方式，區(qū)別在于前者是識(shí)別移植物中供者APC表面的同種異型抗原，而后者識(shí)別自身APC提呈的同種異型抗原。間接識(shí)別是導(dǎo)致AV的主要途徑，最直接的證據(jù)是將供者抗原提呈細(xì)胞去除后仍然存在AV；但在缺乏識(shí)別能力的動(dòng)物模型中也觀察到AV，提示這兩種識(shí)別方式同時(shí)存在[2-3]。

抗原識(shí)別使T細(xì)胞初步活化并賦予其適應(yīng)性免疫應(yīng)答的特性，在共刺激分子的作用下才可完全活化，只有完全活化的T細(xì)胞才能進(jìn)一步分泌細(xì)胞因子和表達(dá)細(xì)胞因子受體，反之T細(xì)胞克隆失能。CD40、CD80、CD86表達(dá)于抗原提呈細(xì)胞，其內(nèi)源性配體CD28、CD154表達(dá)于T細(xì)胞，將該共刺激通路阻斷雖然促進(jìn)了免疫耐受，但是并不能限制AV的發(fā)展[4]；而阻斷可誘導(dǎo)共刺激分子(inducible costimulator, ICOS)與其配體結(jié)合，則減輕內(nèi)膜中平滑肌樣細(xì)胞的增殖程度[5]。并不是所有的共刺激分子都發(fā)揮促進(jìn)T細(xì)胞增殖的作用，如程序性死亡因子-1(programmed death, PD-1)，其有PDL-1、PD-L-2兩個(gè)配體，阻斷PD-1/PDL-1加劇了排斥反應(yīng)，但阻斷PD-1/PDL-2卻未觀察到上述表現(xiàn)[6]；T細(xì)胞免疫球蛋白粘蛋白分子-1(T-cell immunoglobulin and mucin domain containing molecule, TIM-1)具有激活和抑制T細(xì)胞功能的雙重作用，將其阻斷后可抑制Th17而減輕AV[7-8]?？梢姴煌泊碳し肿訉?duì)T細(xì)胞作用不同，即便是同一因子亦可導(dǎo)致不同效應(yīng)，提示共刺激分子對(duì)T細(xì)胞功能調(diào)控存在更加精密的機(jī)制。T細(xì)胞完全活化便分化為效應(yīng)T細(xì)胞，后者分泌細(xì)胞毒性分子，經(jīng)穿孔素/顆粒酶途徑及Fas/FasL途徑激活Caspase級(jí)聯(lián)反應(yīng)，促使靶細(xì)胞凋亡[9]；分泌細(xì)胞因子調(diào)節(jié)免疫和促進(jìn)自身增殖。

2 抗體和補(bǔ)體介導(dǎo)的損傷

在AV患者中發(fā)現(xiàn)抗MHC抗體、供者特異性抗體，其可經(jīng)過多種直接或間接途徑影響血管內(nèi)皮細(xì)胞的功能[10-11]。IgG抗體與MHC-Ⅰ抗原結(jié)合后可使內(nèi)皮細(xì)胞迅速釋放粘附分子如假血友病因子、P-選擇素，趨化因子如MCP-1、IL-8 、RANTES及生長(zhǎng)因子如血小板源性生長(zhǎng)因子(platelet derived growth factor, PDGF)、堿性成纖維細(xì)胞生長(zhǎng)因子(basic fibroblast growth factor, bFGF)，這些細(xì)胞因子經(jīng)過各種途徑參與AV[12]。MHC分子還可改變細(xì)胞骨架結(jié)構(gòu)，以利于炎細(xì)胞黏附和增殖。內(nèi)皮細(xì)胞表面的抗MHC-Ⅰ抗體可增強(qiáng)Rho-GTP活性，使Rho激酶磷酸化，導(dǎo)致張力纖維重組從而改變細(xì)胞骨架結(jié)構(gòu)；同時(shí)RhoA通過磷脂酰激醇-3-激酶途徑促進(jìn)內(nèi)皮細(xì)胞增殖[13]。沉默黏著斑激酶(focal adhesion kinase, FAK) 后絲氨酸、樁蛋白和異位樁蛋白的磷酸化與MHC-Ⅰ分子的結(jié)合顯著減少，提示肌動(dòng)蛋白依賴的分子聚集致細(xì)胞骨架改變是MHC-Ⅰ分子發(fā)揮生物學(xué)作用的路徑[14]。在AV患者中發(fā)現(xiàn)毛細(xì)血管內(nèi)皮細(xì)胞表達(dá)S6核糖體蛋白(S6 ribosomal protein, S6RP)[15]，S6RP位于雷帕霉素復(fù)合體1(mammalian target of rapamycin complex 1, mTORC1)下游，mTORC1由Raptor和GβL共同組成，其可使真核起始4E結(jié)合蛋白和激活p70S6激酶、S6RP磷酸化而促進(jìn)細(xì)胞增殖，沉默Raptor、Rictor后可阻斷MHC-Ⅰ對(duì)血管內(nèi)皮細(xì)胞的增殖作用[16]；mTORC2可通過ERK通路促進(jìn)細(xì)胞增殖，敲除Raptor后發(fā)現(xiàn)MHC-Ⅰ促增殖作用取消，但敲除Rictor卻未能阻斷上述作用[17]；可見mTOR通路是MHC-Ⅰ參與AV的重要途徑，但調(diào)節(jié)機(jī)制有所差異。

補(bǔ)體系統(tǒng)激活能使抗體改變內(nèi)皮細(xì)胞、白細(xì)胞功能，抗體與C1結(jié)合后可被酶切為C2及C4，隨后二者形成復(fù)合體并再次被酶切為C3，C3和C4又可產(chǎn)生生物活性片段C3a、C3b及C4a、C4b，C3b與B因子結(jié)合使其大量沉著并促進(jìn)白細(xì)胞浸潤(rùn)、黏附；C3b還能激活C5產(chǎn)生活性片段C5a和C5b，前者對(duì)白細(xì)胞有強(qiáng)烈的趨化作用，而后者與C6、C9形成膜攻擊復(fù)合體溶解靶細(xì)胞[12]?？贵w和補(bǔ)體還可通過非經(jīng)典的NF- κB信號(hào)通路激活受者T細(xì)胞，從而誘發(fā)細(xì)胞介導(dǎo)的免疫損傷，激活補(bǔ)體系統(tǒng)意義在于放大了免疫效應(yīng)[18]。

3 細(xì)胞因子介導(dǎo)的損傷

細(xì)胞因子主要來源于免疫細(xì)胞、內(nèi)皮細(xì)胞、平滑肌細(xì)胞和上皮細(xì)胞[19]。在AV中T細(xì)胞產(chǎn)生的IFN-γ最為重要，其激活巨噬細(xì)胞，促使內(nèi)皮細(xì)胞分泌粘附因子、趨化因子募集炎細(xì)胞，并通過增強(qiáng)抗原提呈細(xì)胞表達(dá)MHC-Ⅰ類和MHC-Ⅱ分子放大免疫效應(yīng)。敲除IFN-γ后未觀察到AV，使用抗IFN-γ抗體處理AV明顯減輕[20]。IFN-γ主要由Th1細(xì)胞產(chǎn)生，自然殺傷細(xì)胞細(xì)胞毒性T細(xì)胞、樹突狀細(xì)胞等分泌量相對(duì)少。IFN-γ是巨噬細(xì)胞的激活劑，后者活化后合成IL-12、IL-18，繼而進(jìn)一步激活T細(xì)胞，形成正反饋循環(huán)[21]。在IL-12、IL-18刺激下血管平滑肌細(xì)胞亦可分泌IFN-γ，而后者可直接促進(jìn)血管平滑肌細(xì)胞增殖[22]，這一作用還與IFN-γ所致的內(nèi)皮細(xì)胞對(duì)一氧化氮合成酶活性調(diào)節(jié)失能有關(guān)[23]。值得注意的是IFN-γ對(duì)移植血管中膜的平滑肌細(xì)胞和內(nèi)膜中的平滑肌樣細(xì)胞作用不同[24]。腫瘤壞死因子-α(tumour necrosis factor-α, TNF-α)是一種多效促炎因子，其通過促進(jìn)IL-12的產(chǎn)生進(jìn)而促使IFN-γ生成，敲除供者TNF受體明顯減輕AV[25]。

趨化因子是一類使細(xì)胞發(fā)生趨化運(yùn)動(dòng)的小分子蛋白(8-10kD)，可分為CXC、CC、XC及CX3C四個(gè)家族，其中以前二者最常見。趨化因子必須通過與其對(duì)應(yīng)的受體結(jié)合才能發(fā)揮生物學(xué)功能，MCP-1是典型的CC類趨化因子，血管內(nèi)皮細(xì)胞、平滑肌細(xì)胞、巨噬細(xì)胞均可合成，通過與CCR2受體配體結(jié)合，可以招募單核細(xì)胞、T細(xì)胞和自然殺傷細(xì)胞；而同屬CC類的細(xì)胞因子RANTES則在血小板和移植血管新生內(nèi)膜中的平滑肌樣細(xì)胞中表達(dá)。敲除MCP-1或CCR-2后均能減少單個(gè)核細(xì)胞的聚集以及遲發(fā)型超敏反應(yīng)對(duì)移植血管的損傷[26-27]。IFN-γ可誘導(dǎo)產(chǎn)生一些趨化因子，如IP10(IFN-inducible protein 10)， Mig(monokine induced by IFN-γ)、I-TAC(IFN-inducible T-cell α-chemoattractant)，與AV相關(guān)性更明顯[28-29]。內(nèi)皮祖細(xì)胞在趨化因子的作用下定向歸巢至移植物損傷血管內(nèi)膜，要完成黏附并定位于損傷的部位還需要血管內(nèi)膜表達(dá)各種黏附分子的幫助。研究顯示，移植物血管內(nèi)皮細(xì)胞表面可以表達(dá)多種黏附分子，包括細(xì)胞間黏附分子(inter cellular adhesion molecule-1, ICAM-l)、血管細(xì)胞黏附分子(vaseular cell adhesion molceule-1,VCAM-1)、P-選擇素以及E-選擇素等，對(duì)促進(jìn)內(nèi)皮細(xì)胞黏附發(fā)揮重要作用[30-32]。

4 展望

盡管缺血-再灌注損傷、感染等非免疫因素在一定程度參與了AV的發(fā)生、發(fā)展，但免疫因素仍是AV的核心機(jī)制。由于免疫細(xì)胞分泌的多種細(xì)胞因子和趨化因子，使得免疫因素和非免疫因素相互作用，錯(cuò)綜復(fù)雜的病因?qū)е翧V難防、難治；另外，動(dòng)脈粥樣硬化與AV在臨床表現(xiàn)、病理機(jī)制方面有相似之處，應(yīng)注意甄別?？傊?，AV的治療方案應(yīng)以免疫調(diào)節(jié)為基礎(chǔ)，并重細(xì)胞因子調(diào)控的原則制定，而進(jìn)一步發(fā)掘AV的發(fā)病機(jī)制是解決這一臨床議題的根本。

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