腫瘤相關(guān)巨噬細(xì)胞促進(jìn)腫瘤轉(zhuǎn)移的機(jī)制

杜勝霖 王艷林

腫瘤相關(guān)巨噬細(xì)胞促進(jìn)腫瘤轉(zhuǎn)移的機(jī)制

杜勝霖 王艷林★

浸入腫瘤微環(huán)境中的腫瘤相關(guān)巨噬細(xì)胞在不同因子的刺激下，可分化為具有抗腫瘤功能的M1型巨噬細(xì)胞和具有促腫瘤功能的M2型巨噬細(xì)胞。在大多數(shù)腫瘤中腫瘤相關(guān)巨噬細(xì)胞以M2型為主，它參與腫瘤新血管形成、基底膜破壞、細(xì)胞外基質(zhì)重塑、腫瘤細(xì)胞上皮間質(zhì)轉(zhuǎn)化等與腫瘤細(xì)胞侵襲轉(zhuǎn)移相關(guān)的過程，由此腫瘤相關(guān)巨噬細(xì)胞已經(jīng)成為腫瘤靶向治療的重要靶點(diǎn)。本文主要探討腫瘤相關(guān)巨噬細(xì)胞與腫瘤細(xì)胞的相互作用，以及其在促進(jìn)腫瘤轉(zhuǎn)移的各個(gè)環(huán)節(jié)中的分子機(jī)制。

腫瘤；轉(zhuǎn)移；腫瘤相關(guān)巨噬細(xì)胞

腫瘤細(xì)胞的侵襲與轉(zhuǎn)移是腫瘤患者高死亡率的重要原因，闡明其相關(guān)的分子機(jī)制并探索有效的防治手段，是當(dāng)前腫瘤基礎(chǔ)和臨床研究的重大課題。大量研究證實(shí)，處于腫瘤微環(huán)境中的炎性細(xì)胞在腫瘤的發(fā)生、生長(zhǎng)、轉(zhuǎn)移等方面均發(fā)揮促進(jìn)作用，而在這些炎性細(xì)胞中又以腫瘤相關(guān)巨噬細(xì)胞（Tumor-associated macrophage，TAM）起著最為主要的作用。目前認(rèn)為，腫瘤微環(huán)境中的巨噬細(xì)胞至少分以下兩種[1～2]：（1）經(jīng)典的M1型巨噬細(xì)胞，它具有有效的清除感染微生物和殺傷腫瘤細(xì)胞的能力，表型為IL12high，IL23high，IL10low；（2）具有腫瘤促進(jìn)作用的變異型M2巨噬細(xì)胞，它參與腫瘤間質(zhì)的形成、促進(jìn)腫瘤生長(zhǎng)、轉(zhuǎn)移和腫瘤血管形成，同時(shí)導(dǎo)致腫瘤免疫抑制，表型為IL12low，IL23low，IL10high。二者雖均來源于循環(huán)血液中的單核細(xì)胞，但功能截然相反。浸潤(rùn)到腫瘤微環(huán)境中的巨噬細(xì)胞在多種細(xì)胞因子的作用下絕大多數(shù)都表現(xiàn)為M2型。本文就M2型TAM在腫瘤轉(zhuǎn)移中的作用及其機(jī)制的研究進(jìn)展作一簡(jiǎn)要綜述。

1 TAM與腫瘤新血管的形成

腫瘤的遠(yuǎn)距離轉(zhuǎn)移有血循環(huán)轉(zhuǎn)移和淋巴轉(zhuǎn)移兩種主要途徑，豐富的腫瘤血管將為腫瘤細(xì)胞經(jīng)血循環(huán)轉(zhuǎn)移奠定基礎(chǔ)。當(dāng)腫瘤瘤體的快速生長(zhǎng)超過原有腫瘤血管的供血能力時(shí)，瘤體組織內(nèi)就會(huì)出現(xiàn)缺氧及營(yíng)養(yǎng)物質(zhì)匱乏狀況。此時(shí)腫瘤組織將大量合成新的腫瘤血管以滿足瘤組織對(duì)氧和營(yíng)養(yǎng)的需求。腫瘤血管與正常血管比較有以下特點(diǎn)：缺乏經(jīng)典的微動(dòng)脈-毛細(xì)血管-微靜脈結(jié)構(gòu)，高通透性，血管外形異常（呈扭曲形，膨脹形，囊形），周細(xì)胞（內(nèi)皮細(xì)胞的支持細(xì)胞）與內(nèi)皮細(xì)胞松散結(jié)合或周細(xì)胞缺失，基底膜異常增厚或完全消失。這種異常的血管結(jié)構(gòu)容易造成腫瘤微環(huán)境缺氧，低pH值以及高組織間隙液壓[3～5]，同時(shí)具有免疫殺傷活性的細(xì)胞毒性淋巴細(xì)胞難以進(jìn)入腫瘤微環(huán)境[4]。

在缺氧條件下，TAMs以及腫瘤相關(guān)粒細(xì)胞（TANs）通過產(chǎn)生多種細(xì)胞因子、生長(zhǎng)因子及蛋白酶，特別是血管內(nèi)皮生長(zhǎng)因子-A（VEGF-A），激動(dòng)素-2（PROK-2）和金屬基質(zhì)蛋白酶（MMPs），而對(duì)腫瘤血管生成發(fā)揮重要的促進(jìn)作用[6～7]。

缺氧可通過激活低氧誘導(dǎo)因子（hypoxia inducible factor-1α，HIF-1α）途徑促使腫瘤細(xì)胞釋放VEGF和血小板衍生生長(zhǎng)因子（PDGF），這些細(xì)胞因子可使原有腫瘤血管的內(nèi)皮細(xì)胞增殖、出芽并產(chǎn)生新的腫瘤血管。在血管生成素-1（Ang-1）和VEGF的介導(dǎo)下，TAM前體細(xì)胞和骨髓源性抑制細(xì)胞（MDSCs）被大量募集到缺氧的腫瘤微環(huán)境中，并在多種細(xì)胞因子的作用下，通過激活NF-κB，STAT-3和AP-1信號(hào)通路，合成和釋放能促血管形成的因子，如：CXCL-1，CXCL-8，VEGF等[8～9]。另外有實(shí)驗(yàn)證實(shí)，表面有Tie-2（Ang-1的受體蛋白）表達(dá)的巨噬細(xì)胞可產(chǎn)生多種促血管生成因子，包括VEGF-A、MMP-9、Cox-2、胸苷磷酸化酶和組織蛋白酶B（CTSB）等[10～12]，它們不僅可促使血管生成，更能保持新生血管的結(jié)構(gòu)和生存力[13]，其促血管生成能力可在腫瘤細(xì)胞源型的Ang-1的作用下更進(jìn)一步增強(qiáng)[11～12]。

2 TAM與腫瘤細(xì)胞的上皮間質(zhì)轉(zhuǎn)化

在過去十年里，越來越多的證據(jù)表明腫瘤細(xì)胞可通過上皮間質(zhì)轉(zhuǎn)化（EMT）獲得侵襲及遷移的能力[14～16]。這些轉(zhuǎn)化后的細(xì)胞失去上皮細(xì)胞固有的特征，如細(xì)胞之間的緊密連接，細(xì)胞的極性，而且細(xì)胞骨架發(fā)生重塑進(jìn)而獲得間質(zhì)細(xì)胞的遷移性和可塑性等特征[16]。這些表型的改變與EMT相關(guān)轉(zhuǎn)錄因子（如Snail，Zeb和twist家族等）的活化密切相關(guān)，它們直接影響與上皮細(xì)胞表型相關(guān)基因的表達(dá)，特別是通過抑制Cdh1基因，下調(diào)鈣粘蛋白E（E-cadherin）的表達(dá)，從而使上皮細(xì)胞間的緊密連接消失，由此促進(jìn)細(xì)胞的遷移能力[17～18]。

已有大量研究表明腫瘤組織中浸潤(rùn)的免疫細(xì)胞，包括巨噬細(xì)胞、中性粒細(xì)胞、淋巴細(xì)胞以及MDSCs，可分泌大量細(xì)胞因子和趨化因子促進(jìn)腫瘤細(xì)胞發(fā)生上皮間質(zhì)轉(zhuǎn)化[15，19～21]。這些因子通過激活腫瘤細(xì)胞多種信號(hào)通路，包括TGF-β、TNF-α、NF-κB、Notch、Wnt以及Hedgehog等，而啟動(dòng)上皮間質(zhì)轉(zhuǎn)化的過程[22～24]。特別是TAMs分泌的TNF-α能通過上調(diào)NF-κB而誘導(dǎo)Snail的轉(zhuǎn)錄抑制活性。Snail是一種含鋅指結(jié)構(gòu)的抑制性轉(zhuǎn)錄因子，能通過下調(diào)E-cadhrein的表達(dá)而促進(jìn)EMT過程。Snail蛋白高度不穩(wěn)定，很容易經(jīng)GSK-3β磷酸化途徑或SCFβ-Trep泛素化途徑降解。而TNF-α可通過活化CNS2（COP9 signalosome2）而抑制Snail與GSK-3β和SCFβ-Trep的結(jié)合進(jìn)而增強(qiáng)Snail的穩(wěn)定性[25]。另外TAMs分泌的IL-6和IL-23可通過活化STAT-3信號(hào)通路上調(diào)Twist家族基因的表達(dá)，從而下調(diào)E-cadhrein并降低細(xì)胞間的粘附作用[26～27]。

TNF-α/NF-κB/Snail信號(hào)通路對(duì)EMT的發(fā)生起關(guān)鍵性作用[28]，NF-κB可視為EMT最有效的誘導(dǎo)因子，TNF-α能激活NF-κB和Akt繼而抑制GSK-3β活性，由此增加細(xì)胞內(nèi)Snail的水平，進(jìn)入細(xì)胞核后，Snail將抑制E-cadherin基因的轉(zhuǎn)錄并下調(diào)其細(xì)胞內(nèi)水平。此外，Snail還能抑制某些腫瘤轉(zhuǎn)移抑制基因的表達(dá)，如黑色素瘤中的Raf激酶抑制蛋白和前列腺癌中的PTEN等[29]。Li等[30]的研究發(fā)現(xiàn)，miR488能通過下調(diào)細(xì)胞內(nèi)NF-κB的水平而抑制EMT，其機(jī)制可能與miR488靶向抑制轉(zhuǎn)錄因子SATB1有關(guān)。SATB1能激活A(yù)mphiregulin（一種自分泌的上皮生長(zhǎng)因子）基因的轉(zhuǎn)錄，后者與細(xì)胞膜表面的上皮生長(zhǎng)因子受體結(jié)合后，能經(jīng)PI3K/Akt信號(hào)通路上調(diào)NF-κB的表達(dá)水平由此促進(jìn)EMT。而SATB1 mRNA是miR488的直接靶點(diǎn)，在其3'-UTR內(nèi)含有miR488的識(shí)別和結(jié)合序列，miR488與之結(jié)合將加快SATB1 mRNA降解和抑制SATB1蛋白合成。下調(diào)miR488水平將增高SATB1活性，上調(diào)NF-κB表達(dá)水平和促進(jìn)EMT發(fā)生。此外還發(fā)現(xiàn)，NF-κB能結(jié)合到miR488基因的啟動(dòng)子序列上并抑制miR488表達(dá)，由此構(gòu)成一個(gè)EMT的正反饋環(huán)[30]。另外NF-κB還控制著多種編碼細(xì)胞因子（IL-1，IL-2，ILp6，TNF-α），趨化因子（IL-8，MIP-1α，MCP-1，RANTES，eotaxin），粘附分子（血管細(xì)胞粘附分子，E-cadhrein），生長(zhǎng)因子和酶類（環(huán)氧化酶2，誘導(dǎo)型氮氧化物合酶）基因的表達(dá)[31]，它們均能直接或間接促進(jìn)腫瘤EMT和轉(zhuǎn)移。

TAMs分泌的TGF-β是能誘導(dǎo)EMT的另一種炎性因子。TGF-β原本的生理功能是控制細(xì)胞的增殖和促炎癥反應(yīng)，然而腫瘤細(xì)胞通常由于基因突變使其喪失了促凋亡和抑制增殖的能力，而在特定的腫瘤微環(huán)境中TGF-β發(fā)揮著重要的促進(jìn)EMT的功能。突變的P57/Smad復(fù)合物能抑制TGF-β的抗腫瘤作用并活化其促腫瘤轉(zhuǎn)移的能力。TGF-β可通過Smad依賴性和Smad非依賴性兩種不同的信號(hào)途徑促進(jìn)EMT發(fā)生[32]。經(jīng)典的TGF-β/TGF-βR/Smad2信號(hào)通路可從表觀遺傳水平上改變EMT相關(guān)基因的表達(dá)[33]，非經(jīng)典的Smad非依賴性信號(hào)途徑則經(jīng)由G蛋白偶聯(lián)受體、MAPKs、PI3K及NF-κB信號(hào)通路作用于Snail和Slug等轉(zhuǎn)錄因子，由此下調(diào)E-cadhrein和ZO-1表達(dá)和上調(diào)Vimentin表達(dá)，進(jìn)而誘導(dǎo)腫瘤細(xì)胞發(fā)生EMT[34～35]。以上研究結(jié)果提示，TAMs分泌的TGF-β能通過多種途徑誘導(dǎo)EMT的發(fā)生。

3 TAM與腫瘤組織基底膜破壞和細(xì)胞外基質(zhì)重塑

腫瘤組織基底膜的破壞以及細(xì)胞外基質(zhì)的重塑為腫瘤細(xì)胞的轉(zhuǎn)移所必須，這一過程需要能降解細(xì)胞外基質(zhì)蛋白的絲氨酸蛋白酶、基質(zhì)金屬蛋白酶和半胱氨酸組織蛋白酶等的參與，它們的異常高表達(dá)與多種腫瘤患者的低生存率和不良預(yù)后有直接關(guān)系。上述蛋白水解酶可由腫瘤細(xì)胞分泌，但更多的是由腫瘤間質(zhì)的成纖維細(xì)胞和浸潤(rùn)的免疫細(xì)胞所分泌。

Mohamed等[36]對(duì)乳腺癌的研究發(fā)現(xiàn)，腫瘤細(xì)胞產(chǎn)生的IL-6能增加TAMs分泌蛋白水解酶CTSB（一種半胱氨酸蛋白酶），MMP-2和 MMP-9的能力，它們可直接作用于細(xì)胞間的連接，改變細(xì)胞外基質(zhì)（ECM）成分并破壞基底膜，用抗IL-6中和抗體可阻斷IL-6的上述功能。Wang等[37]對(duì)原發(fā)性肺癌的研究發(fā)現(xiàn)，肺癌組織中TAM高表達(dá)并分泌MMP-9和尿激酶型纖溶酶原激活物（uPA），二者均能促進(jìn)細(xì)胞外基質(zhì)降解。用無血清培養(yǎng)基培養(yǎng)從肺癌組織中分離的TAM后，獲得條件化的培養(yǎng)上清，再將這種上清用于SPC-A1、H460和A549肺癌細(xì)胞株的培養(yǎng)，結(jié)果發(fā)現(xiàn)，上述TAM條件化的培養(yǎng)上清能顯著性增加腫瘤細(xì)胞的侵襲和轉(zhuǎn)移能力，用抗u-PA和抗MMP-9抗體能阻斷這一過程。

4 結(jié)語

TAM在介導(dǎo)腫瘤侵襲和轉(zhuǎn)移過程中的關(guān)鍵性作用，使得TAM成為腫瘤靶向治療的重要靶點(diǎn)，通過誘導(dǎo)TAM發(fā)生M1表型轉(zhuǎn)化，或抑制TAM中與促腫瘤血管生成、促EMT發(fā)生和促細(xì)胞外基質(zhì)降解的細(xì)胞因子的合成與釋放，可望達(dá)到有效抑制腫瘤生長(zhǎng)和防止腫瘤細(xì)胞侵襲和轉(zhuǎn)移的目的。

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The metastasis-promoting roles of tumor-associated macrophages

DU Shenglin, WANG Yanlin★
(Three Gorges University Medical College, Institute of Molecular Biology, Hubei, Yichang 443002, China)

Under the stimulation of different cytokines, tumor-associated macrophages (TAM) immersed into the tumor microenvironment can be differentiated into M1 macrophage with anti-tumor function and M2 macrophage with tumor-promoting function. M2 macrophage is the major type of TAM in the majority of tumors. It is involved in the multiple metastasis-related processes, such as tumor angiogenesis, breakdown of the basement membrane, remodeling of the extracellular matrix and tumor cell's epithelial-mesenchymal transition. So that TAM has become an important target for tumor targeting therapy. This article mainly discusses the interaction between TAM and tumor cells and its underlying molecular mechanisms in promoting tumor metastasis.

Tumor; Metastasis; Tumor-associated macrophage

國(guó)家自然科學(xué)基金資助項(xiàng)目（30973445）

三峽大學(xué)醫(yī)學(xué)院，三峽大學(xué)分子生物學(xué)研究所，湖北，宜昌 443002

★通訊作者：王艷林，E-mail: fzswangyl@ctgu.edu.cn