Epac信號分子與哮喘關(guān)系研究進(jìn)展

黃革，高亞東

(武漢大學(xué)中南醫(yī)院呼吸內(nèi)科，湖北武漢430071)

Epac信號分子與哮喘關(guān)系研究進(jìn)展

黃革，高亞東

(武漢大學(xué)中南醫(yī)院呼吸內(nèi)科，湖北武漢430071)

支氣管哮喘是臨床常見的氣道慢性炎癥性疾病，以氣道高反應(yīng)性、慢性氣道炎癥和氣道重塑為病理生理特征，其發(fā)病機(jī)制目前尚未完全闡明。環(huán)磷酸腺苷(cAMP)是體內(nèi)重要的第二信使，參與調(diào)控機(jī)體新陳代謝、細(xì)胞鈣信號傳導(dǎo)、細(xì)胞生長與分化、凋亡等多種病理生理過程。長期以來，蛋白激酶A(PKA)被認(rèn)為是介導(dǎo)cAMP生物學(xué)效應(yīng)的唯一下游信號分子。但新近發(fā)現(xiàn)的新型cAMP靶分子——cAMP直接激活的交換蛋白(Epac)的發(fā)現(xiàn)打破了這一說法。大量研究證實，Epac可單獨(dú)或協(xié)同PKA介導(dǎo)cAMP的多種生物學(xué)效應(yīng)。本文就Epac在支氣管哮喘發(fā)病中的作用及其可能機(jī)制作一綜述，為尋找哮喘治療新靶點奠定基礎(chǔ)。

支氣管哮喘；環(huán)磷酸腺苷；cAMP直接激活的交換蛋白；蛋白激酶A

支氣管哮喘(簡稱哮喘)是臨床常見的氣道慢性炎癥性疾病，以氣道高反應(yīng)性、炎癥及重塑為重要特征[1]，且發(fā)病率呈逐年上升趨勢[2]。目前，哮喘的治療以緩解癥狀和控制急性發(fā)作為主。β2受體激動劑、磷酸二酯酶抑制劑等作為哮喘治療的一線藥物，其療效已被廣泛認(rèn)同，作用機(jī)制主要是通過升高細(xì)胞內(nèi)cAMP水平。PKA一直被認(rèn)為是cAMP下游唯一的效應(yīng)分子，介導(dǎo)cAMP的多種生物學(xué)效應(yīng)，直到1998年分別發(fā)表在Nature和Science的兩項研究發(fā)現(xiàn)cAMP直接激活的交換蛋白(Exchange protein directly activated by cAMP，Epac)分子才改變了這一觀念[3-4]。Epac作為新型cAMP靶分子，可單獨(dú)或協(xié)同PKA介導(dǎo)cAMP的多種生物學(xué)效應(yīng)。本文將主要介紹Epac在哮喘發(fā)病中的作用及其可能的機(jī)制。

1 新型cAMP靶分子——Epac

Epac主要功能是作為小G蛋白Rap1的特異性鳥嘌呤核苷酸交換因子(Guanine nucleotide exchange factor，GEF)，將Rap1結(jié)合的GDP(無活性)置換為GTP(有活性)，活化后的Rap1進(jìn)一步對各項生理活動起調(diào)控作用。Epac被證實存在兩種亞型：Epac1和Epac2，兩者在包括肺在內(nèi)的許多組織器官均有表達(dá)[3-4]，且在不同器官及器官的不同發(fā)育階段的表達(dá)量存在明顯差異[5]。Epac在結(jié)構(gòu)上包括調(diào)節(jié)區(qū)和催化區(qū)，前者含有高親和力的cAMP結(jié)合位點，后者含有一特異性的CDC25同源結(jié)構(gòu)域(CDC25 homology domain，CDC25-HD)，CDC25-HD具有GEF活性，可活化一系列信號分子，如Rap1、Rap2、PKB/Akt、Rho、R-Ras等，進(jìn)一步調(diào)控細(xì)胞生長、分化、粘附及凋亡等多種細(xì)胞生命活動[6]。Epac作為一個多功能信號分子，對免疫、呼吸、神經(jīng)、心血管及內(nèi)分泌等多個系統(tǒng)功能有其獨(dú)特的調(diào)控作用[7-8]。

2 Epac與哮喘

哮喘是多種細(xì)胞、細(xì)胞組分及炎癥介質(zhì)引起的氣道慢性炎癥性疾病，伴隨氣道阻塞和氣道結(jié)構(gòu)改變(即氣道重塑)。大量研究表明，Epac參與哮喘氣道高反應(yīng)性、炎癥及重塑進(jìn)程，在哮喘的發(fā)病中發(fā)揮重要作用。

2.1 Epac調(diào)控氣道反應(yīng)性氣道高反應(yīng)性(Airway hyperresponsibility，AHR)是哮喘基本特征之一。盡管關(guān)于AHR發(fā)生機(jī)制的研究已取得較大進(jìn)展，但其確切機(jī)制目前尚未完全闡明，參與的機(jī)制包括：Th細(xì)胞作用失衡；平滑肌細(xì)胞增生與肥大；黏液過度分泌；嗜酸粒細(xì)胞等多種炎癥細(xì)胞、炎癥介質(zhì)和細(xì)胞因子的作用等。雖然AHR形成機(jī)制復(fù)雜多樣，但氣道平滑肌(Airway smooth muscle，ASM)在其中所起作用是毋庸置疑的。cAMP激動劑如β2受體激動劑緩解哮喘支氣管痙攣的作用已被廣泛認(rèn)同，大多數(shù)觀點認(rèn)為PKA介導(dǎo)了這一效應(yīng)[9]。但也有研究表明，cAMP激動劑產(chǎn)生的舒張支氣管作用非PKA依賴性[10]。不乏證據(jù)表明Epac參與其中，有研究證實Epac可將RhoA/Rac1平衡點向Rac1偏移，從而降低ASM肌球蛋白輕鏈(Myosin light chain，MLC)磷酸化水平，介導(dǎo)ASM舒張[11]。另有Zieba等[12]研究發(fā)現(xiàn)，Epac可通過下調(diào)RhoA活性誘導(dǎo)ASM松弛。RhoA、Rac1皆屬Rho GTPase家族，Rho GTPase可通過調(diào)控肌動-肌球蛋白細(xì)胞骨架影響細(xì)胞運(yùn)動，在ASM收縮與舒張活動中起重要作用。氣道黏液的過度分泌與AHR的形成密切相關(guān)。氣道上皮細(xì)胞電解質(zhì)的跨膜轉(zhuǎn)運(yùn)(如Cl-的分泌)在氣道黏液的清除中發(fā)揮積極作用。Lau等[13]研究表明，Epac可增強(qiáng)二磷酸尿苷(Uridine diphosphate，UDP)誘導(dǎo)的人上皮細(xì)胞Cl-的分泌活動，從而促進(jìn)黏液的清除，減輕氣道高反應(yīng)性。

2.2 Epac調(diào)控氣道炎癥慢性氣道炎癥在哮喘的發(fā)生中占重要地位，越來越多的研究表明Epac在哮喘炎癥過程中發(fā)揮重要作用，且同時表現(xiàn)出抗炎和促炎作用。細(xì)胞屏障功能在抑制炎癥的擴(kuò)散中發(fā)揮重要作用。有研究顯示，Epac可通過活化小G蛋白Rap1/Rac維持肺血管內(nèi)皮細(xì)胞功能穩(wěn)定[14]，從而阻止炎癥的擴(kuò)散。Vang等[15]研究發(fā)現(xiàn)，Epac參與了cAMP對效應(yīng)T細(xì)胞的抑制作用。另有Zaslona等[16]研究表明，Epac抑制了Th2型細(xì)胞因子IL-13的生成。Epac經(jīng)PKB/Akt和GSK-3信號途徑，抑制細(xì)菌脂多糖(Lipopolysaccaride，LPS)誘導(dǎo)的巨噬細(xì)胞IFN-β的分泌[17]。Jing等[18]研究發(fā)現(xiàn)，Epac通過活化PKB/GSK信號途徑抑制了樹突狀細(xì)胞(Dendritic cell，DC)分泌趨化因子CCL3和CCL4，從而發(fā)揮抗炎效應(yīng)。腺苷(Adenosine)可通過與各種炎癥細(xì)胞表面的腺苷受體A2a(A2aR)結(jié)合發(fā)揮抗炎效應(yīng)，Scheibner等[19]研究發(fā)現(xiàn)，A2aR介導(dǎo)的抗炎作用與Epac抑制核因子-κB (Nuclear factor-κB，NF-κB)和一些炎性基因的表達(dá)有關(guān)。除上述抗炎作用外，Epac同時表現(xiàn)出一定的促炎作用。哮喘時血液循環(huán)中白細(xì)胞向氣道募集是炎癥的主要來源之一。有研究表明，Epac-Rap1通過活化整合素β1(Integrins β1)促進(jìn)了白細(xì)胞的粘附及游出[20]。Tan等[21]研究表明，Epac通過活化細(xì)胞外信號調(diào)節(jié)激酶1/2(Extracellular signal-regulated kinase 1/2，ERK1/2)和p38信號途徑，促進(jìn)巨噬細(xì)胞分泌IL-6和IL-1β等炎癥因子。有研究顯示，Epac協(xié)同PKA通過激活Rap1-ERK1/2信號途徑，促進(jìn)了緩激肽(Bradykinin)誘導(dǎo)的氣道平滑肌細(xì)胞IL-8的分泌[22]。另有Nagano等[23]研究證明，Epac可通過活化Rap-PLCε信號途徑，上調(diào)哮喘小鼠支氣管上皮細(xì)胞炎癥因子的分泌。IL-10是重要的免疫調(diào)節(jié)性因子，主要功能為抑制炎癥反應(yīng)和調(diào)控免疫細(xì)胞的分化和增殖。有研究表明，Epac可抑制外周血單個核細(xì)胞(Peripheral blood mononuclear cell，PBMC)分泌IL-10[24]，間接的促進(jìn)了炎癥反應(yīng)。因此，Epac作為一個多功能信號分子，可通過不同的信號途徑，在哮喘中同時發(fā)揮著抗炎及促炎作用。

2.3 Epac調(diào)控氣道重塑有些哮喘患者承受持續(xù)癥狀以及進(jìn)行性肺功能降低，對其氣道進(jìn)行組織學(xué)檢查，往往會發(fā)現(xiàn)氣道發(fā)生了明顯的結(jié)構(gòu)改變，包括氣道上皮和網(wǎng)狀層增厚、上皮下膠原沉積、平滑肌細(xì)胞增生肥大、黏液腺增生及新生血管形成等病理改變[25]，即所謂氣道重塑。氣道平滑肌細(xì)胞(Airway smooth muscle cells，ASMCs)增生與肥大是哮喘氣道重塑最顯著病理特征之一。ASMCs有潛在的增殖、遷移及合成分泌功能，這也正是其導(dǎo)致氣道重塑的主要機(jī)制。Kassel等[26]研究表明，Epac參與β2受體激動劑抑制ASMCs的增殖效應(yīng)。ASMCs在絲裂原刺激下會發(fā)現(xiàn)表型轉(zhuǎn)換，由收縮型轉(zhuǎn)換為增殖分泌型，促進(jìn)哮喘疾病的發(fā)展。有研究顯示，Epac通過抑制ERK1/2轉(zhuǎn)錄阻止ASMCs表型轉(zhuǎn)換[27]，起到抗重塑作用。肺成纖維細(xì)胞在哮喘氣道重塑中同樣發(fā)揮重要作用。當(dāng)成纖維細(xì)胞受到刺激時，可轉(zhuǎn)化成肌成纖維細(xì)胞，后者可分泌大量包括Ⅰ、Ⅲ、Ⅴ型膠原蛋白及纖維連接蛋白在內(nèi)的細(xì)胞外基質(zhì)和促炎因子，此外成纖維細(xì)胞可分泌基質(zhì)金屬蛋白酶(Matrix metalloprotinease，MMP-9)及其抑制物組織性基質(zhì)金屬蛋白酶抑制物(Tissue inhibitor of matrix metalloproteinase，TIMP-1)，MMP-9可分解細(xì)胞外基質(zhì)，若MMP-9與TIMP-1失衡，會導(dǎo)致細(xì)胞外基質(zhì)的沉積[28]。Huang等[29]研究證實，Epac1通過活化Rap1抑制了人肺成纖維細(xì)胞的增殖。同時，Haag等[30]研究證明，Epac1而非PKA抑制了人肺成纖維細(xì)胞增殖。Oldenburger等[31]研究表明，Epac1可抑制香煙煙霧誘導(dǎo)的氣道重塑過程。因此可知，Epac可通過多種途徑抑制氣道重塑。

3 展望

cAMP作為重要的第二信使分子，參與調(diào)控體內(nèi)多種生理病理過程。臨床上常用哮喘治療藥物，如β2受體激動劑、磷酸二酯酶抑制劑、前列腺素E2等均是通過增加細(xì)胞內(nèi)cAMP水平來發(fā)揮效應(yīng)的。PKA作為cAMP下游效應(yīng)分子參與介導(dǎo)cAMP的多種效應(yīng)，而新近發(fā)現(xiàn)的Epac同樣介導(dǎo)發(fā)揮cAMP的生物學(xué)功能。Epac有調(diào)控哮喘氣道高反應(yīng)性、炎癥及重塑過程的作用，但在炎癥方面，Epac似乎表現(xiàn)出相互對抗的作用，如抗炎和促炎作用，說明Epac調(diào)控疾病過程的信號途徑紛繁復(fù)雜，有待研究進(jìn)一步闡明。鑒于Epac在哮喘中發(fā)揮重要作用，或許在不久的將來會成為哮喘治療的新型靶分子。

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Research progress of the relationship between Epac and asthma.

HUANG Ge,GAO Ya-dong.Department of Respiratory Medicine,Zhongnan Hospital of Wuhan University,Wuhan 430071,Hubei,CHINA

Asthma is a chronic airway inflammatory disorder characterized by airway hyperresponsibility, chronic inflammation and airway remodeling,of which the mechanisms are not fully illustrated.Cyclic adenosine monophosphate(cAMP)is a very important second messenger which controls a range of diverse physiological or pathological processes,including metabolism,calcium signal,cell growth and differentiation,and apoptosis.Protein kinase A(PKA)had been considered as the only downstream target of cAMP.However,the recently reported target of cAMP, the exchange protein directly activated by cAMP(Epac),had changed the previous point.A large number of studies had shown that Epac regulates many biological processes either alone and(or)in concert with PKA.In this review,we will discuss the roles and probable mechanisms of Epac in asthma.

Asthma;Cyclic adenosine monophosphate(cAMP);Exchange protein directly activated by cAMP(Epac);Protein kinaseA(PKA)

R562.2+5

A

1003—6350（2015）12—1791—03

2015-01-12)

湖北省自然科學(xué)基金(編號：2014CFB202)

高亞東。E-mail：gaoyadong@gmail.com

doi∶10.3969/j.issn.1003-6350.2015.12.0641