Wnt信號(hào)轉(zhuǎn)導(dǎo)通路與結(jié)直腸癌的研究進(jìn)展

李伶俐　陳小燕　朱　蓉　趙　逵*

遵義醫(yī)學(xué)院(563003)1　遵義醫(yī)學(xué)院附屬醫(yī)院消化內(nèi)科2

Wnt信號(hào)轉(zhuǎn)導(dǎo)通路與結(jié)直腸癌的研究進(jìn)展

李伶俐1陳小燕2朱蓉2趙逵2*

遵義醫(yī)學(xué)院(563003)1遵義醫(yī)學(xué)院附屬醫(yī)院消化內(nèi)科2

結(jié)直腸癌是胃腸道最常見(jiàn)的惡性腫瘤之一，發(fā)病機(jī)制尚未完全明確，目前認(rèn)為是一個(gè)多步驟、多階段參與的疾病。Wnt信號(hào)轉(zhuǎn)導(dǎo)通路是進(jìn)化上高度保守的信號(hào)系統(tǒng)，可調(diào)控細(xì)胞生長(zhǎng)、運(yùn)動(dòng)和分化，且在胚胎發(fā)育、腫瘤發(fā)生等過(guò)程中起重要作用。人類多種惡性腫瘤中均存在Wnt通路異常，90%以上的結(jié)直腸癌存在Wnt經(jīng)典信號(hào)轉(zhuǎn)導(dǎo)通路激活[1]。本文就Wnt信號(hào)轉(zhuǎn)導(dǎo)通路與結(jié)直腸癌的研究進(jìn)展作一綜述。

一、Wnt信號(hào)轉(zhuǎn)導(dǎo)通路

1. Wnt經(jīng)典信號(hào)轉(zhuǎn)導(dǎo)通路：Wnt經(jīng)典信號(hào)轉(zhuǎn)導(dǎo)通路即Wnt/β-catenin通路。Wnt通路的模式是：無(wú)Wnt信號(hào)時(shí)，降解復(fù)合體與β-catenin結(jié)合使其磷酸化，β-catenin隨即脫離復(fù)合體，通過(guò)β-轉(zhuǎn)導(dǎo)重復(fù)相容蛋白(β-TrCP)泛素化，從而被蛋白酶體降解；Wnt信號(hào)刺激時(shí)，Wnt信號(hào)誘導(dǎo)軸蛋白(Axin)與磷酸化的低密度脂蛋白受體相關(guān)蛋白(LRP)結(jié)合，降解復(fù)合體分離，β-catenin在胞質(zhì)中穩(wěn)定存在，并轉(zhuǎn)位入胞核，與T細(xì)胞因子(TCF)結(jié)合，形成轉(zhuǎn)錄復(fù)合體，激活Lgr5等靶基因。近年來(lái)，基于內(nèi)源性降解復(fù)合體的研究提出一種新的Wnt信號(hào)轉(zhuǎn)導(dǎo)通路模式：無(wú)Wnt信號(hào)時(shí)，降解復(fù)合體通過(guò)β-TrCP與β-catenin結(jié)合，使其磷酸化和泛素化后被降解；Wnt信號(hào)刺激時(shí)，降解復(fù)合物與磷酸化LRP結(jié)合，然后結(jié)合β-catenin使其磷酸化，但此過(guò)程可被β-TrCP泛素化阻止，導(dǎo)致β-catenin在胞質(zhì)內(nèi)積聚[2]。

2. Wnt非經(jīng)典信號(hào)轉(zhuǎn)導(dǎo)通路：目前有多種Wnt非經(jīng)典通路，如Wnt/PCP、Wnt-RAP1、Wnt-Ror2、Wnt-PKA、Wnt-GSK3MT、Wnt-aPKC、Wnt-RYK、Wnt-mTOR以及Wnt/Ca2+、Wnt/Wg等，多種通路可重疊出現(xiàn)。其中Wnt/Ca2+通路尤為重要，其由Wnt5a或Wnt4激活后可抑制包括結(jié)直腸癌在內(nèi)的多種腫瘤[3]。

二、Wnt信號(hào)轉(zhuǎn)導(dǎo)通路主要組成成分

1. Wnt蛋白：①Wnt蛋白脂?；揎棧篧nt蛋白中高度保守的半胱氨酸與棕櫚酸酯通過(guò)硫脂鍵結(jié)合后被脂?；痆4]。研究[5-6]發(fā)現(xiàn)，Wnt蛋白去除半胱氨酸后，活性喪失，提示W(wǎng)nt蛋白活性受脂?；{(diào)控。②Wnt蛋白的轉(zhuǎn)運(yùn)：Wls是7次跨膜蛋白，存在于高爾基體網(wǎng)絡(luò)、核內(nèi)體和質(zhì)膜，可與Wnt蛋白結(jié)合，將Wnt蛋白從高爾基體運(yùn)至質(zhì)膜，且對(duì)Wnt蛋白穩(wěn)定分泌起重要作用。研究[7]顯示，胞內(nèi)轉(zhuǎn)運(yùn)復(fù)合體retromer亦對(duì)Wnt信號(hào)途徑十分重要，其參與Wnt蛋白的轉(zhuǎn)運(yùn)，通過(guò)逆向轉(zhuǎn)運(yùn)將核內(nèi)體中的Wls運(yùn)回至高爾基網(wǎng)絡(luò)，從而避免被溶酶體降解。

2. β-catenin：β-catenin 是Wnt/β-catenin通路的中樞成分。正常成熟細(xì)胞中，大部分β-catenin與細(xì)胞膜上的E-鈣黏蛋白結(jié)合，參與細(xì)胞間黏附、生長(zhǎng)、增殖等過(guò)程，少部分與胞質(zhì)降解復(fù)合體結(jié)合被降解。β-catenin的黏附作用和信號(hào)轉(zhuǎn) 導(dǎo)功能獨(dú)立存在[8]。Wnt信號(hào)活化可提高β-catenin水平[9]。

三、Wnt信號(hào)轉(zhuǎn)導(dǎo)通路的調(diào)節(jié)

1. β-catenin對(duì)Wnt通路的調(diào)節(jié)：降解復(fù)合體可調(diào)控胞質(zhì)內(nèi)β-catenin的穩(wěn)定性，后者對(duì)經(jīng)典Wnt信號(hào)轉(zhuǎn)導(dǎo)通路起關(guān)鍵作用[10]。β-catenin降解復(fù)合體可使β-catenin磷酸化，而后被泛素-蛋白酶體系統(tǒng)降解，使其保持低水平，從而關(guān)閉Wnt通路[11]。WTX是一種抑癌蛋白，參與β-catenin降解復(fù)合體功能，促進(jìn)β-catenin降解，發(fā)揮抑癌作用。目前關(guān)于WTX在Wnt通路中的作用存在爭(zhēng)議[12]。

2. Lgr對(duì)Wnt通路的調(diào)節(jié)：Glinka等[13]的研究顯示，Lgr4、Lgr5是R-脊柱蛋白受體，其可介導(dǎo)R-脊柱蛋白參與Wnt經(jīng)典信號(hào)通路。在生殖器、眼睛、膀胱、腎、毛囊、下顎、舌頭、胃、腸道等多種器官中均有Lgr4、Lgr5表達(dá)[14-16]。對(duì)胃腸道腫瘤的研究[15]發(fā)現(xiàn)，Lgr5是Wnt通路的靶基因，胃腸道中Lgr5陽(yáng)性表達(dá)的干細(xì)胞可發(fā)揮組織重建作用。Walker等[17]對(duì)結(jié)直腸癌細(xì)胞株的研究顯示，Lgr5可拮抗Wnt信號(hào)，調(diào)控細(xì)胞黏附，抑制腫瘤生長(zhǎng)。

3. 金屬結(jié)合蛋白(Dpr)對(duì)Wnt通路的調(diào)節(jié)：Dpr的功能域在進(jìn)化上高度保守，對(duì)于調(diào)控Wnt信號(hào)的機(jī)制尚未明確。Dpr可與散亂蛋白(DSH)結(jié)合，誘導(dǎo)DSH在溶酶體中降解，從而抑制Wnt經(jīng)典通路和非經(jīng)典通路激活[18]。Gao等[19]亦發(fā)現(xiàn)Dpr1能同時(shí)負(fù)性調(diào)控細(xì)胞質(zhì)和細(xì)胞核中的Wnt信號(hào)途徑。但是，Gloy等[20]對(duì)爪蟾的研究顯示，Dpr1能協(xié)同加強(qiáng)DSH的作用。因此，Dpr1對(duì)Wnt信號(hào)通路的調(diào)節(jié)作用尚存爭(zhēng)議。

四、Wnt通路與結(jié)直腸癌的關(guān)系

1. 基因異常：①β-catenin基因突變：β-catenin基因缺失或突變導(dǎo)致β-catenin不能降解，在胞質(zhì)內(nèi)積累，引起腫瘤發(fā)生。雖然β-catenin基因突變可導(dǎo)致Wnt信號(hào)活化，但大多數(shù)結(jié)腸癌患者顯示了β-catenin的異質(zhì)性，表明Wnt信號(hào)調(diào)控的復(fù)雜性[21]。②APC基因突變：多數(shù)結(jié)直腸癌存在APC基因缺失突變或失活[22]。APC基因突變點(diǎn)位于與Axin結(jié)合位點(diǎn)，產(chǎn)生截短APC蛋白，導(dǎo)致不能形成降解復(fù)合體降解β-catenin。APC功能缺失不僅導(dǎo)致β-catenin聚積，亦促進(jìn)β-catenin與TCF4形成 TCF/β-catenin復(fù)合物，激活Wnt信號(hào)轉(zhuǎn)導(dǎo)通路下游的靶基因。Yoshie等[23]用氣相色譜法比較表達(dá)正常APC蛋白和截短APC蛋白的SW480結(jié)直腸癌細(xì)胞代謝物含量，結(jié)果顯示，表達(dá)截短APC蛋白的SW480細(xì)胞的代謝產(chǎn)物含量明顯提高，提示APC突變通過(guò)改變能量代謝通路參與結(jié)直腸癌的發(fā)展。此外，Bellis等[24]的研究表明，APC突變的小鼠腺瘤細(xì)胞中，干細(xì)胞不對(duì)稱分裂發(fā)生改變，此為結(jié)直腸癌的發(fā)展提供了新線索。③Axin基因突變：結(jié)直腸癌細(xì)胞株中Axin表達(dá)缺失可導(dǎo)致β-catenin大量積聚，從而引起Wnt信號(hào)通路異?；钴S。Vermeulen等[21]通過(guò)研究克什米爾人群中結(jié)直腸癌患者的Axin基因突變模式發(fā)現(xiàn)，Axin1和Axin2基因突變參與了結(jié)直腸癌的發(fā)生和發(fā)展，并發(fā)現(xiàn)Axin2基因突變可能是克什米爾人群患結(jié)直腸癌的誘發(fā)因素。

2. Wnt信號(hào)通路與結(jié)直腸癌干細(xì)胞：腫瘤干細(xì)胞能夠自我更新以及維持分化潛能，其行為需依賴外部信號(hào)，如Wnt信號(hào)[25]。研究顯示，Wnt非經(jīng)典途徑可拮抗β-catenin依賴的轉(zhuǎn)錄，表明其有抗癌作用[26]。近年來(lái)，大量研究證實(shí)，Wnt信號(hào)、干細(xì)胞信號(hào)及以腫瘤干細(xì)胞行為間存在聯(lián)系，提示干細(xì)胞生物學(xué)與結(jié)直腸癌相關(guān)。

①腸道干細(xì)胞與結(jié)直腸癌干細(xì)胞：在自我更新以及多向分化潛能方面，腫瘤干細(xì)胞與干細(xì)胞具有相似的生物學(xué)特性。腫瘤干細(xì)胞學(xué)說(shuō)認(rèn)為，腫瘤干細(xì)胞可能源于正常干細(xì)胞的累積突變和(或)祖細(xì)胞通過(guò)基因突變重新獲得自我更新能力。腫瘤干細(xì)胞可能帶有正常干細(xì)胞/祖細(xì)胞的特異性表面標(biāo)記物，如結(jié)腸癌干細(xì)胞具有與結(jié)腸干細(xì)胞相同的標(biāo)記物如CD133、Lgr5。

②Wnt信號(hào)通路與結(jié)腸干細(xì)胞：正常人結(jié)腸由數(shù)百萬(wàn)個(gè)隱窩組成，每個(gè)隱窩含有2 000個(gè)細(xì)胞。干細(xì)胞位于隱窩基底部，每個(gè)隱窩約有5~10個(gè)干細(xì)胞[27-28]。肌成纖維細(xì)胞可通過(guò)調(diào)控Wnt信號(hào)對(duì)肝配蛋白B1(EFNB1)、EPHB2以及EPHB3作用，從而調(diào)節(jié)干細(xì)胞功能。Wnt信號(hào)通路亦對(duì)結(jié)腸干細(xì)胞自我更新的調(diào)控起重要作用[29-30]。此外，Lgr蛋白作為R-脊柱蛋白受體可介導(dǎo)Wnt信號(hào)與干細(xì)胞緊密聯(lián)系。通過(guò)抑制TCF4或給予Wnt拮抗劑Dickkopf-1(DDK1)阻斷Wnt信號(hào)，導(dǎo)致上皮細(xì)胞增殖能力減弱。

③Wnt信號(hào)通路與結(jié)腸癌干細(xì)胞：Wnt/β-catenin通路可調(diào)控腫瘤細(xì)胞的生長(zhǎng)。研究[29]指出，結(jié)腸癌細(xì)胞球可減少β-catenin在細(xì)胞膜黏附，提高β-catenin、cyclin D1以及c-myc的胞質(zhì)水平，同時(shí)下調(diào)Axin1和磷酸化β-catenin。β-catenin表達(dá)增多與TCF/LEF轉(zhuǎn)錄活化密切相關(guān)，采用RNA干擾技術(shù)使β-catenin基因沉默，可導(dǎo)致TCF/LEF降低，從而減少結(jié)腸癌細(xì)胞球形成。相反，上調(diào)c-myc和TCF/LEF下游效應(yīng)器可大幅增加結(jié)腸癌球囊細(xì)胞生成。Barker等[30]通過(guò)敲除小鼠Lgr5-Cre+腸隱窩干細(xì)胞的APC基因，發(fā)現(xiàn)3~5周后干細(xì)胞仍存留于隱窩底部，并形成肉眼可見(jiàn)的Lgr5+腺瘤，提示W(wǎng)nt信號(hào)轉(zhuǎn)導(dǎo)通路參與正常腸干細(xì)胞向腫瘤干細(xì)胞轉(zhuǎn)化。

五、結(jié)語(yǔ)

基于Wnt信號(hào)轉(zhuǎn)導(dǎo)通路是一條多環(huán)節(jié)、多作用位點(diǎn)的開放通路，其對(duì)結(jié)直腸癌的發(fā)生、發(fā)展尤為重要，因此可針對(duì)該信號(hào)途徑進(jìn)行分子靶向治療。然而，結(jié)直腸癌的發(fā)生是一個(gè)復(fù)雜的網(wǎng)絡(luò)工程，Wnt信號(hào)轉(zhuǎn)導(dǎo)通路的作用機(jī)制以及Wnt信號(hào)轉(zhuǎn)導(dǎo)通路與其他信號(hào)通路如STATs 、MAPKs、Hedgehog、BMP等的相互作用機(jī)制尚未完全明確，仍有待進(jìn)一步研究。

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(2015-01-28收稿；2015-02-05修回)

·病例分析與個(gè)案報(bào)道·

摘要結(jié)直腸癌是胃腸道最常見(jiàn)的惡性腫瘤之一，發(fā)病機(jī)制尚未完全明確，目前認(rèn)為是一個(gè)多步驟、多階段參與的疾病。Wnt信號(hào)轉(zhuǎn)導(dǎo)通路可調(diào)控細(xì)胞生長(zhǎng)、運(yùn)動(dòng)和分化，且在胚胎發(fā)育、腫瘤發(fā)生等過(guò)程中起重要作用。本文就Wnt信號(hào)轉(zhuǎn)導(dǎo)通路與結(jié)直腸癌的研究進(jìn)展作一綜述。

關(guān)鍵詞結(jié)直腸腫瘤；Wnt信號(hào)通路；β連環(huán)素；干細(xì)胞

Advances in Study on Wnt Signaling Transduction Pathway and Colorectal CancerLILingli1,CHENXiaoyan2,ZHURong2,ZHAOKui2.1ZunyiMedicalCollege,Zunyi,GuizhouProvince(563003);2DepartmentofGastroenterology,AffiliatedHospitalofZunyiMedicalCollege,Zunyi,GuizhouProvince

Correspondence to: ZHAO Kui, Email: kuizhao95868@msn.com

AbstractColorectal cancer is one of the most commonly seen gastrointestinal carcinomas and its pathogenesis has not yet been fully clarified. It is considered as a multi-step and multi-stage disease. Wnt signaling transduction pathway regulates cell growth, motility and differentiation, and plays a crucial role in the regulation of embryonic development and tumor genesis. This article reviewed the advances in study on Wnt signaling transduction pathway and colorectal cancer.

Key wordsColorectal Neoplasms;Wnt Signaling Pathway;beta Catenin;Stem Cells

通信作者*本文，Email: kuizhao95868@msn.com

DOI:10.3969/j.issn.1008-7125.2015.10.015