劉洋馮新民陳濤黃澤楠張亮

間充質(zhì)干細(xì)胞衰老機(jī)制的研究進(jìn)展

劉洋1,2馮新民2陳濤2黃澤楠2張亮2

間充質(zhì)干細(xì)胞（MSCs）具有來源廣泛、組織修復(fù)能力強(qiáng)、自我更新能力強(qiáng)及多向分化等特征，是組織工程學(xué)上理想的種子細(xì)胞，在骨科損傷與修復(fù)重建領(lǐng)域具有廣闊的應(yīng)用前景。但隨著年齡增長(zhǎng)或體外傳代次數(shù)的增加，MSCs同樣面臨衰老問題，從而影響MSCs的臨床應(yīng)用及治療效果。因此闡明MSCs衰老的機(jī)制并尋求抗衰老的策略問題是目前亟待解決的問題。本文主要從端粒酶與端粒、氧化應(yīng)激損傷、沉默信息調(diào)節(jié)蛋白、高糖狀態(tài)、Wnt/β-catenin通路激活及DNA損傷等方面對(duì)MSCs衰老機(jī)制作一綜述，并思考應(yīng)對(duì)MSCs衰老策略，以達(dá)到延緩甚至逆轉(zhuǎn)其衰老的目的。

間質(zhì)干細(xì)胞； 衰老； 老化； 沉默信息調(diào)節(jié)蛋白； 抗衰老

作者單位：116044 大連醫(yī)科大學(xué)研究生院1；225001 揚(yáng)州大學(xué)臨床醫(yī)學(xué)院2

干細(xì)胞是一種具有自我更新及多向分化潛能的未成熟細(xì)胞，根據(jù)來源不同一般可分為胚胎干細(xì)胞、成體干細(xì)胞及多能誘導(dǎo)干細(xì)胞。在成體干細(xì)胞中間充質(zhì)干細(xì)胞（mesenchymal stem cells，MSCs）被視為最有力競(jìng)爭(zhēng)者，MSCs來源于發(fā)育早期的中胚層和外胚層，目前已從骨髓、脂肪、肌肉、羊水、臍帶血等組織中分離出該細(xì)胞；同時(shí)以MSCs為基礎(chǔ)的生物學(xué)治療是多種退變性疾病及損傷修復(fù)重建的研究熱點(diǎn)[1-2]。雖然MSCs在體內(nèi)分布廣泛，但數(shù)量很少，骨髓中每十萬個(gè)有核細(xì)胞才有1個(gè)MSCs。為獲得足夠數(shù)量的MSCs為臨床所用，MSCs體外擴(kuò)增培養(yǎng)是不可或缺的環(huán)節(jié)。正常動(dòng)物細(xì)胞在體內(nèi)外生長(zhǎng)過程中的分裂次數(shù)存在一個(gè)被稱為Hay fl ick極限的“極限值”，當(dāng)細(xì)胞分裂次數(shù)達(dá)到Hay fl ick極限，細(xì)胞就會(huì)發(fā)生衰老，而衰老細(xì)胞最終會(huì)通過啟動(dòng)細(xì)胞凋亡而死亡[3]。衰老是機(jī)體細(xì)胞、組織和器官在功能上出現(xiàn)逐步弱化的生理過程，主要表現(xiàn)為自身抵抗能力降低及對(duì)外界環(huán)境刺激反應(yīng)能力降低，從而誘發(fā)疾病的易感性并最終引起死亡[4]。

MSCs在擴(kuò)增培養(yǎng)過程中同樣會(huì)發(fā)生衰老及凋亡，從而導(dǎo)致其生物學(xué)功能下降，甚至出現(xiàn)功能失調(diào)，從而限制了MSCs的臨床應(yīng)用[5]。衰老是一種復(fù)雜的生命過程，是多種因素共同參與的結(jié)果。MSCs的衰老機(jī)制尚未完全明確，近年來研究主要集中在端粒與端粒酶、氧化應(yīng)激損傷、SIRT家族蛋白、高糖狀態(tài)、Wnt/β-catenin通路激活及DNA損傷等方面[5-6]。本文就MSCs的衰老機(jī)制及抗衰老策略等方面的研究進(jìn)展作一綜述。

一、MSCs的衰老機(jī)制

（一）端粒與端粒酶

端粒是位于真核細(xì)胞線狀染色體末端的、由短小高度重復(fù)非轉(zhuǎn)錄DNA序列（TTAGGG）和一些結(jié)構(gòu)蛋白構(gòu)成的特殊結(jié)構(gòu)，在染色體定位、復(fù)制、保護(hù)和控制細(xì)胞生長(zhǎng)及壽命方面具有重要作用[6]。端粒的長(zhǎng)度是實(shí)現(xiàn)其功能的前提，DNA每次復(fù)制，端粒都會(huì)丟失約20～300 bp，隨MSCs分裂次數(shù)增多端粒逐漸縮短，當(dāng)端粒短縮到無法繼續(xù)DNA復(fù)制及不能保證染色體穩(wěn)定性時(shí)就會(huì)誘發(fā)MSCs衰老的發(fā)生，導(dǎo)致細(xì)胞分裂停滯并無法繼續(xù)復(fù)制，最終發(fā)生凋亡[5,7]。

端粒長(zhǎng)度主要靠端粒酶維持，端粒酶是一種可延長(zhǎng)短縮端粒長(zhǎng)度的逆轉(zhuǎn)錄酶，在保持染色體穩(wěn)定和細(xì)胞活性上扮演重要角色[7-8]。Wang等[9]發(fā)現(xiàn)端粒酶的活性與機(jī)體年齡相關(guān)，隨年齡增長(zhǎng)MSCs端粒酶活性逐漸降低。He等[8]通過比較鼠及人MSCs發(fā)現(xiàn)，人MSCs不會(huì)在長(zhǎng)期培養(yǎng)中自發(fā)啟動(dòng)轉(zhuǎn)化從而逃脫細(xì)胞衰老，這主要是由于其端粒較鼠MSCs明顯縮短，因此提出當(dāng)使用人MSCs作為模型評(píng)估基于人MSC的細(xì)胞治療安全性時(shí)需謹(jǐn)慎考慮。Zhao等[10]將人端粒酶逆轉(zhuǎn)錄酶應(yīng)用于大鼠骨髓MSCs，下調(diào)人端粒酶逆轉(zhuǎn)錄酶表達(dá)可短縮端粒長(zhǎng)度及端粒酶活性，并抑制細(xì)胞增殖，促進(jìn)細(xì)胞凋亡；而過表達(dá)人端粒酶逆轉(zhuǎn)錄酶可增加端粒長(zhǎng)度及端粒酶活性，從而促進(jìn)細(xì)胞增殖及減少細(xì)胞凋亡；同時(shí)發(fā)現(xiàn)人端粒酶逆轉(zhuǎn)錄酶是通過PI3K/AKT信號(hào)通路實(shí)現(xiàn)對(duì)細(xì)胞的調(diào)節(jié)。

（二）氧化應(yīng)激損傷

細(xì)胞正常新陳代謝過程中產(chǎn)生活性氧（reactive oxygen species，ROS），低濃度ROS是細(xì)胞增殖分化的必備條件；但病理?xiàng)l件下引發(fā)的氧化應(yīng)激損傷增加并產(chǎn)生大量ROS，高濃度ROS具有很強(qiáng)細(xì)胞毒性作用及誘導(dǎo)細(xì)胞損傷的能力[11-12]。研究發(fā)現(xiàn)凋亡細(xì)胞中ROS表達(dá)較正常細(xì)胞明顯增加[13]，且伴隨年齡[14]及體外傳代次數(shù)[15]MSCs產(chǎn)生的ROS逐漸增加，過量ROS或外源性H2O2可損傷MSCs的增殖分化能力。Ko等[16]發(fā)現(xiàn)亞致死量ROS及電離輻射可導(dǎo)致人臍帶來源MSCs的DNA損傷，出現(xiàn)DNA合成減少及細(xì)胞增殖減慢，從而導(dǎo)致細(xì)胞衰老。Borodkin等[17]通過外源性H2O2誘發(fā)人子宮內(nèi)膜來源MSCs發(fā)生衰老，而抑制p38/MAPK信號(hào)通路可部分減輕H2O2誘發(fā)的細(xì)胞衰老。Zhou等[18]發(fā)現(xiàn)亞致死劑量H2O2可使骨髓MSCs細(xì)胞周期停滯于G0/G1期，同時(shí)細(xì)胞成骨分化能力明顯抑制，而抗氧化劑褪黑素以濃度依賴性方式促進(jìn)增殖細(xì)胞進(jìn)入S期，同時(shí)通過抑制p38/MAPK信號(hào)通路減輕MSCs衰老。因此氧化應(yīng)激因素可能通過ROS及p38/MAPK信號(hào)通路誘發(fā)MSCs衰老的發(fā)生。

（三）SIRT家族蛋白

沉默信息調(diào)節(jié)蛋白（silent information regulator protein，Sirtuin）是一類在進(jìn)化過程中非常保守，依賴于煙酰胺腺嘌呤二核苷酸（NAD+）的組蛋白去乙?；概cADP-核糖基轉(zhuǎn)移酶的蛋白酶，在機(jī)體病理生理過程中有重要調(diào)控作用，參與調(diào)控機(jī)體炎癥反應(yīng)、能量代謝及衰老等過程[19-20]。

SIRT家族包括7個(gè)成員SIRT1-7，SIRTl首先被發(fā)現(xiàn)，同時(shí)也是目前研究較為透徹的一員，SIRTl可調(diào)控衰老、代謝和應(yīng)激反應(yīng)等過程，其對(duì)MSCs抗衰老功能已被許多學(xué)者肯定[19-20]。MSCs中SIRT1蛋白水平及活性隨其體外傳代次數(shù)及年齡增加而下降[21]，SIRT1水平在維持MSCs細(xì)胞增殖及分化中發(fā)揮重要作用[22-23]。Kim等[24]發(fā)現(xiàn)高糖可增加脂肪MSCs中miR-486表達(dá)并抑制SIRT1表達(dá)，從而促進(jìn)細(xì)胞衰老。Simic等[23]研究發(fā)現(xiàn)，選擇性敲除SIRT1的骨髓MSCs早期出現(xiàn)細(xì)胞生長(zhǎng)減緩，逐漸進(jìn)入細(xì)胞停滯期及出現(xiàn)細(xì)胞衰老加速等情況，同時(shí)衰老細(xì)胞處于S期的細(xì)胞明顯減少；且衰老細(xì)胞中p16明顯增加，p21表達(dá)無明顯變化；而敲除脂肪來源MSCs的SIRT1基因則發(fā)現(xiàn)p16和p21表達(dá)均有所升高，說明SIRT1對(duì)不同組織來源的MSCs衰老的影響機(jī)制也是不同的。Wang等[25]發(fā)現(xiàn)適當(dāng)濃度的白藜蘆醇可增加臍帶來源MSCs的活力及增殖能力，減輕細(xì)胞衰老，促進(jìn)SIRT1表達(dá)而抑制p53及p16表達(dá)，因此認(rèn)為白藜蘆醇通過劑量依賴方式經(jīng)SIRT1信號(hào)通路調(diào)節(jié)臍帶來源MSCs的自我更新能力及向神經(jīng)細(xì)胞分化能力。Ma等[26]發(fā)現(xiàn)SIRT1延緩MSCs衰老的機(jī)制與煙酰胺磷酸核糖轉(zhuǎn)移酶（nicotinamide phosphoribosyltransferase，Nampt）相關(guān)，Nampt為NAD+合成限速酶，是決定細(xì)胞內(nèi)NAD+含量的關(guān)鍵因素。Son等[27]證實(shí)通過恢復(fù)線粒體NAD+水平可延緩MSCs復(fù)制時(shí)產(chǎn)生的衰老，從而延長(zhǎng)MSCs壽命，因此維持線粒體NAD+水平對(duì)逆轉(zhuǎn)MSCs衰老至關(guān)重要。SIRT1及SIRT2功能的發(fā)揮依賴于NAD+的濃度，NAD+作為中心紐帶，將Nampt與SIRT1緊密相連，三者共同構(gòu)成哺乳動(dòng)物的衰老調(diào)控網(wǎng)絡(luò)。

SIRT家族中除SITR1及SIRT2外，SIRT3及SIRT6同樣具有抗衰老功能。Wang等[28]研究發(fā)現(xiàn)嚴(yán)重的氧化應(yīng)激導(dǎo)致骨髓MSCs的SIRT3表達(dá)水平下降，而SIRT3過表達(dá)可通過激活錳超氧化物歧化酶（MnSOD）和過氧化氫酶（Catalase CAT）表達(dá)，從而改善氧化應(yīng)激對(duì)MSCs造成的損傷，減少細(xì)胞凋亡。SIRT6通過影響DNA損傷修復(fù)過程來維持基因組穩(wěn)定，其功能缺陷將誘發(fā)衰老發(fā)生[29-31]。Lee等[32]研究SIRT6對(duì)骨髓MSCs影響時(shí)發(fā)現(xiàn)，SIRT6、衰老相關(guān)-β-半乳糖苷酶及p16表達(dá)隨年齡增加而增加，而細(xì)胞增殖及遷移能力則減弱；敲除SIRT6基因后，細(xì)胞增殖、遷移及抗氧化應(yīng)激能力受損，細(xì)胞加速衰老，提示SIRT6可調(diào)節(jié)MSCs衰老。

SIRT家族中多個(gè)成員均具有抗衰老功能，明確SIRT家族抗衰老機(jī)制可把握MSCs衰老過程中的關(guān)鍵環(huán)節(jié)，并為解決體外培養(yǎng)MSCs所面臨的衰老問題提供可行方法。

（四）高糖狀態(tài)

由于葡萄糖不僅是細(xì)胞代謝的重要能量來源，同時(shí)也是重要代謝產(chǎn)物，因此體內(nèi)外微環(huán)境中葡萄糖濃度顯著影響細(xì)胞的增殖、分化、衰老及凋亡[33]。Kim等[24]發(fā)現(xiàn)高糖環(huán)境可增加脂肪MSCs中miR-486表達(dá)并抑制SIRT1表達(dá)，從而促進(jìn)細(xì)胞衰老。Lei等[34]進(jìn)一步研究發(fā)現(xiàn)SIRT1抑制劑白藜蘆醇可保護(hù)由高糖環(huán)境誘導(dǎo)的MSCs衰老。孫超等[35]通過比較微環(huán)境中不同濃度葡萄糖對(duì)骨髓MSCs增殖周期、凋亡及衰老的影響，結(jié)果發(fā)現(xiàn)高糖和細(xì)胞蛋白質(zhì)O位氮乙酰葡糖胺（O-GlcNAc）糖基化修飾可抑制細(xì)胞增殖，阻滯細(xì)胞周期，促進(jìn)細(xì)胞凋亡及衰老，因此認(rèn)為高糖誘導(dǎo)的蛋白質(zhì)O-GlcNAc糖基化水平升高可能是其抑制MSCs衰老的機(jī)制之一。

（五）Wnt/β-catenin通路激活

Wnt/β-catenin信號(hào)通路在調(diào)節(jié)細(xì)胞增殖、分化及衰老等過程中發(fā)揮重要作用[36]。Zhang等[37]和Kajla等[38]研究表明Wnt/β-catenin信號(hào)途徑是MSCs中ROS生成的主要催化劑，可增強(qiáng)氧化酶活性，ROS通過破壞氧化劑與抗氧化劑的平衡，使Wnt/β-catenin信號(hào)通路激活，導(dǎo)致MSCs衰老；清除ROS會(huì)使Wnt/β-catenin信號(hào)通路誘導(dǎo)的MSCs衰老減弱，并減少DNA損傷及其他衰老基因的表達(dá)。Gu等[39]對(duì)系統(tǒng)性紅斑狼瘡患者骨髓MSCs進(jìn)行研究，結(jié)果發(fā)現(xiàn)Wnt/β-catenin信號(hào)通路過表達(dá)通過DNA損傷和p53/p21信號(hào)途徑激活誘發(fā)骨髓MSCs衰老。Zhang等[37，40]研究同樣證實(shí)Wnt/β-catenin信號(hào)途徑通過促進(jìn)細(xì)胞內(nèi)ROS生成及p53/p21信號(hào)途徑誘發(fā)MSCs衰老，并認(rèn)為Wnt/β-catenin信號(hào)途徑的過度激活可能是MSCs衰老的主要調(diào)節(jié)因子。

（六）DNA損傷

很多情況下MSCs衰老的發(fā)生均涉及DNA損傷反應(yīng)（DNA damage response，DDR）通路的激活[41-43]。當(dāng)MSCs發(fā)生DNA損傷，快速修復(fù)機(jī)制將被啟動(dòng)促使細(xì)胞順利完成細(xì)胞周期，若修復(fù)失敗將導(dǎo)致細(xì)胞出現(xiàn)衰老甚至凋亡；另一方面DDR導(dǎo)致的基因組高度不穩(wěn)定性是MSCs衰老的重要機(jī)制之一[6,41]。體外長(zhǎng)期培養(yǎng)衰老的MSCs出現(xiàn)DNA損傷，持續(xù)的DNA損傷可抑制DNA合成，改變MSCs的形態(tài)特征，促進(jìn)衰老[44]。Yu等[45]研究證實(shí)累積性DNA損傷可產(chǎn)生內(nèi)源性干擾素β，同時(shí)刺激干擾素相關(guān)信號(hào)通路，而干擾素可放大DDR，激活p53信號(hào)通路，并通過縮短端粒長(zhǎng)度促進(jìn)MSCs衰老。這些結(jié)果說明DNA損傷及其相關(guān)信號(hào)通路在MSCs衰老過程中發(fā)揮重要作用。

二、應(yīng)對(duì)MSCs衰老策略的研究

在MSCs體外培養(yǎng)過程中，衰老的發(fā)生不可避免，為實(shí)現(xiàn)MSCs在臨床的廣泛應(yīng)用，需尋求應(yīng)對(duì)MSCs衰老的策略。通過查閱及分析文獻(xiàn)發(fā)現(xiàn)，一般可從延緩MSCs衰老速度、恢復(fù)或增強(qiáng)MSCs活性及基因改造MSCs等三方面著手。

（一）延緩MSCs衰老速度

在MSCs體外培養(yǎng)方式上，與傳統(tǒng)貼壁培養(yǎng)相比，三維培養(yǎng)更適合MSCs的生物學(xué)特性，可延緩其復(fù)制衰老速度。Shearier等[46]在低氧及常氧條件下采用二維及三維培養(yǎng)方式培養(yǎng)人MSCs，檢測(cè)細(xì)胞外基質(zhì)及干性基因表達(dá)情況，結(jié)果發(fā)現(xiàn)在低氧環(huán)境下三維培養(yǎng)的MSCs細(xì)胞外基質(zhì)表達(dá)更多，在長(zhǎng)期培養(yǎng)過程中干性基因表達(dá)維持更好，說明低氧及三維培養(yǎng)環(huán)境有助于延緩MSCs的衰老。

另外，研究表明藥物干預(yù)可延緩MSCs的衰老。Fehrer等[47]研究發(fā)現(xiàn)在培養(yǎng)基中加入維生素C或N-乙酰半胱氨酸（N-acetylcysteine）可延緩MSCs衰老。Kanehira等[48]研究發(fā)現(xiàn)溶血磷脂酸（lysophosphatidic acid，LPA）對(duì)膜磷脂的合成至關(guān)重要，使用LPA受體拮抗劑在體外培養(yǎng)中可減少因復(fù)制衰老導(dǎo)致的MSCs功能丟失。另外在衰老MSCs中發(fā)現(xiàn)存在組蛋白脫乙酰酶表達(dá)下降，因此組蛋白乙酰轉(zhuǎn)移酶抑制劑的應(yīng)用可延緩MSCs復(fù)制衰老的發(fā)生[49]。

（二）恢復(fù)或增強(qiáng)衰老MSCs的活性

在延緩MSCs衰老速度的同時(shí)可尋求恢復(fù)或增強(qiáng)衰老MSCs活性的方法。鄭晨曦等[50]研究發(fā)現(xiàn)維生素C能夠增強(qiáng)衰老個(gè)體來源的骨髓MSCs增殖能力，且這種作用可能是通過增強(qiáng)端粒酶活性實(shí)現(xiàn)的。Gharibi等[51]通過比較加入不同生長(zhǎng)因子培養(yǎng)基培養(yǎng)的人MSCs活性，結(jié)果發(fā)現(xiàn)與對(duì)照組相比，加入成纖維生長(zhǎng)因子、血小板衍生生長(zhǎng)因子、抗壞血酸和內(nèi)皮生長(zhǎng)因子的培養(yǎng)基均可增加復(fù)制速率且在MSCs發(fā)生衰老前顯著增加細(xì)胞倍增數(shù)目。Mohammadi等[52]進(jìn)一步研究發(fā)現(xiàn)，不同濃度的血小板裂解物可充當(dāng)生長(zhǎng)因子的安全來源并用于MSCs體外培養(yǎng)，且與傳統(tǒng)胎牛血清培養(yǎng)基相比，在長(zhǎng)期培養(yǎng)過程中可較好保持干細(xì)胞的生長(zhǎng)分化潛能。在傳統(tǒng)中藥方面，詹菲等[53]通過實(shí)驗(yàn)研究發(fā)現(xiàn)左歸丸能提高衰老大鼠MSCs減弱的增殖能力。

（三）基因改造MSCs

在MSCs體外培養(yǎng)及應(yīng)用中，其來源也是需考慮的重要因素。越來越多的文章報(bào)道通過誘導(dǎo)多能干細(xì)胞（pluripotent stem cells，iPSCs）來引導(dǎo)功能性MSCs，稱為誘導(dǎo)性間充質(zhì)干細(xì)胞（induced MSCs，iMSCs）。iPSCs是一個(gè)值得引起重視的MSCs的來源，因?yàn)樵谄渲鼐幊踢^程中，細(xì)胞經(jīng)歷再生，展現(xiàn)出更好的細(xì)胞活力劑增殖分化能力[54-55]。iPSCs能夠在維持穩(wěn)定二倍體核型、一致的基因表達(dá)和表面抗原譜的情況下容易地?cái)U(kuò)增至40代[56]，且能夠保持完整的生物學(xué)功能[57]。值得注意的是，iPSCs保留供體來源DNA的甲基化譜，同時(shí)獲得了不完全的免疫調(diào)節(jié)功能[58]，因此iPSCs真正投入體外培養(yǎng)及臨床應(yīng)用仍面臨著許多挑戰(zhàn)。

綜上所述，MSCs在參與組織損傷修復(fù)重建過程中發(fā)揮重要作用，以干細(xì)胞為基礎(chǔ)的生物學(xué)方法已成為治療熱點(diǎn)及發(fā)展方向，因此研究MSCs的衰老機(jī)制對(duì)推進(jìn)其在臨床的廣泛應(yīng)用極其重要。同時(shí)MSCs體外培養(yǎng)過程中會(huì)出現(xiàn)衰老現(xiàn)象，嚴(yán)重阻礙MSCs的臨床應(yīng)用，在充分了解MSCs衰老機(jī)制的前提下，尋求應(yīng)對(duì)衰老策略具有重要意義。但目前抗MSCs衰老的方法尚處于初級(jí)階段且僅是針對(duì)體外培養(yǎng)的MSCs進(jìn)行的實(shí)驗(yàn)研究，尚缺乏臨床試驗(yàn)的相關(guān)研究證據(jù)，無法明確這些方法是否適合臨床移植的MSCs使用，同時(shí)是否會(huì)對(duì)移植的MSCs臨床效果產(chǎn)生影響，以上均需待進(jìn)一步的深入研究。

1 Elabd C, Centeno CJ, Schultz JR, et al. Intra-discal injection of autologous, hypoxic cultured bone marrow-derived mesenchymal stem cells in fi ve patients with chronic lower back pain: a long-term safety and feasibility study[J]. J Transl Med, 2016, 14:253.

2 Pereira CL, Gon?alves RM, Peroglio M, et al. The effect of hyaluronanbased delivery of stromal cell-derived factor-1 on the recruitment of MSCs in degenerating intervertebral discs[J]. Biomaterials, 2014,35(28):8144-8153.

3 Hayflick L. The limitedin vitrolifetime of human diploid cell strains[J]. Exp Cell Res, 1965, 37:614-636.

4 Oh J, Lee YD, Wagers AJ. Stem cell aging:mechanisms,regulators and therapeutic opportunities[J]. Nat Med, 2014, 20(8):870-880.

5 Benameur L, Charif N, Li YE, et al. Toward an understanding of mechanism of aging-induced oxidative stress in human mesenchymal stem cells[J]. Biomed Mater Eng, 2015, 25(1):S41-S46.

6 Trachana V, Petrakis S, Fotiadis Z, et al. Human mesenchymal stem cells with enhanced telomerase activity acquire resistance against oxidative stress-induced genomic damage[J]. Cytotherapy, 2017, pii:S1465-3249(17)30541-30548. [Epub ahead of print]

7 Carrillo J, Calvete O, Pintado-Berninches L, et al. Mutations in XLF/NHEJ1/cernunnos gene results in downregulation of telomerase genes expression and telomere shortening[J]. Hum Mol Genet, 2017,26(10):1900-1914.

8 He L, Zheng Y, Wan Y, et al. A shorter telomere is the key factor in preventing cultured human mesenchymal stem cells from senescence escape[J]. Histochem Cell Biol, 2014, 142(3):257-267.

9 Wang X, Zou X, Zhao J, et al. Site-specific characteristics of bone marrow mesenchymal stromal cells modify the effect of aging on the skeleton [J]. Rejuvenation Res, 2015, [Epub ahead of print]

10 Zhao Q, Wang XY, Yu XX, et al. Expression of human telomerase reverse transcriptase mediates the senescence of mesenchymal stem cells through the PI3K/AKT signaling pathway[J]. Int J Mol Med,2015, 36(3):857-864.

11 Choo KB, Tai L, Hymavathee KS, et al. Oxidative stress-induced premature senescence in Wharton's jelly-derived mesenchymal stem cells[J]. Int J Med Sci, 2014, 11(11):1201-1207.

12 Lee JH, Jung HK, Han YS, et al. Antioxidant effects of Cirsium setidens extract on oxidative stress in human mesenchymal stem cells[J]. Mol Med Rep, 2016, 14(4):3777-3784.

13 Jeong SG, Cho GW. Endogenous ROS levels are increased in replicative senescence in human bone marrow mesenchymal stromal cells[J]. Biochem Biophys Res Commun, 2015, 460(4):971-976.

14 Stolzing A, Jones E, Mcgonagle D, et al. Age-related changes in human bone marrow-derived mesenchymal stem cells: Consequences for cell therapies[J]. Mech Ageing Dev, 2008, 129(3):163-173.

15 Lee JS, Lee MO, Moon BH, et al. Senescent growth arrest in mesenchymal stem cells is bypassed by Wip1-Mediated downregulation of intrinsic stress signaling pathways[J]. Stem Cells,2009, 27(8):1963-1975.

16 Ko E, Lee KY, Hwang DS. Human umbilical cord Blood-Derived mesenchymal stem cells undergo cellular senescence in response to oxidative stress[J]. Stem Cells Dev, 2012, 21(11):1877-1886.

17 Borodkina A, Shatrova A, Abushik PA, et al. Interaction between ROS dependent DNA damage, mitochondria and p38 MAPK underlies senescence of human adult stem cells[J]. Aging, 2014, 6(6):481-495.

18 Zhou L, Chen X, Liu T, et al. Melatonin reverses H2O2-induced premature senescence in mesenchymal stem cells via the SIRT1-dependent pathway[J]. J Pineal Res, 2015, 59(2):190-205.

19 Larsen SA, Kassem M, Rattan SI. Glucose metabolite glyoxal induces senescence in telomerase-immortalized human mesenchymal stem cells[J]. Chem Cent J, 2012, 6(1):18.

20 Kitada M, Kume S, Takeda-Watanabe A, et al. Sirtuins and renal diseases: relationship with aging and diabetic nephropathy[J]. Clin Sci(Lond), 2013, 124(3):153-164.

21 Li Y, He X, Li Y, et al. Nicotinamide phosphoribosyltransferase(Nam pt)affects the lineage fate determination of mesenchymal stem cells:a possible cause for reduced osteogenesis and increased adipogenesis in older individuals[J]. J Bone Miner Res, 2011, 26(11):2656-2664.

22 Chen H, Liu X, Chen H, et al. Role of SIRT1 and AMPK in mesenchymal stem cells differentiation [J]. Ageing Res Rev, 2014,13:55-64.

23 Simic P, Zainabadi K, Bell E, et al. SIRT1 regulates differentiation of mesenchymal stem cells by deacetylating β-catenin[J]. EMBO Mol Med, 2013, 5(3):430-440.

24 Kim YJ, Hwang SH, Lee SY, et al. miR-486-5p induces replicative senescence of human adipose tissue-derived mesenchymal stem cells and its expression is controlled by high glucose[J]. Stem Cells Dev,2012, 21(10):1749-1760.

25 Wang XX, Ma SS, Meng N, et al. Resveratrol exerts Dosage-Dependent effects on the Self-Renewal and neural differentiation of hUC-MSCs[J]. Mol Cells, 2016, 39(5):418-425.

26 Ma C, Pi CC, Yang YE, et al. Nampt expression decreases Age-Related senescence in rat bone marrow mesenchymal stem cells by targeting Sirt1[J]. PLoS One, 2017, 12(1):e0170930.

27 Son MJ, Kwon Y, Son T, et al. Restoration of mitochondrial NAD(+)levels delays stem cell senescence and facilitates reprogramming of aged somatic cells[J]. Stem Cells, 2016, 34(12):2840-2851.

28 Wang XQ, Shao Y, Ma CY, et al. Decreased SIRT3 in aged human mesenchymal stromal/stem cells increases cellular susceptibility to oxidative stress[J]. J Cell Mol Med, 2014, 18(11):2298-2310.

29 Sun HL, Wu YR, Fu DJ, et al. SIRT6 regulates osteogenic differentiation of rat bone marrow mesenchymal stem cells partially via suppressing the nuclear factor-kappa B signaling pathway[J]. Stem Cells, 2014, 32(7):1943-1955.

30 Mostoslavsky R, Chua KF, Lombard DB, et al. Genomic instability and aging-like phenotype in the absence of mammalian SIRT6[J]. Cel,2006, 124(2):315-329.

31 Gertler AA, Cohen HY. SIRT6, a protein with many faces[J].Biogerontology, 2013, 14(6):629-639.

32 Lee N, Kim DK, Kim ES, et al. Comparative interactomes of SIRT6 and SIRT7:Implication of functional links to aging[J]. Proteomics,2014, 14(13/14):1610-1622.

33 Zhang B, Liu N, Gu B, et al. [Observing the effect of high glucose on proliferation of bone marrow stromal stem cells through Wnt/Β-catenin pathway][J]. Zhongguo yi xue ke xue yuan xue bao, 2014,36(4):389-393.

34 Lei LT, Chen JB, Zhao YL, et al. Resveratrol attenuates senescence of adipose-derived mesenchymal stem cells and restores their paracrine effects on promoting insulin secretion of INS-1 cells through Pim-1[J].Eur Rev Med Pharmacol Sci, 2016, 20(6):1203-1213.

35 孫超, 王言, 邢輝, 等. 葡萄糖及O-GlcNAc糖基化對(duì)人骨髓間充質(zhì)干細(xì)胞增殖、凋亡、衰老的影響[J]. 中國(guó)脊柱脊髓雜志, 2016,26(4):354-361.

36 Clevers H, Nusse R. Wnt/β-Catenin signaling and disease[J]. Cell,2012, 149(6):1192-1205.

37 Zhang DY, Pan Y, Zhang C, et al. Wnt/beta-catenin signaling induces the aging of mesenchymal stem cells through promoting the ROS production[J]. Mol Cell Biochem, 2013, 374(1/2):13-20.

38 Kajla S, Mondol AS, Nagasawa A, et al. A crucial role for Nox 1 in redox-dependent regulation of Wnt-beta-catenin signaling[J]. FASEB Journal, 2012, 26(5):2049-2059.

39 Gu Z, Tan W, Feng G, et al. Wnt/β-catenin signaling mediates the senescence of bone marrow-mesenchymal stem cells from systemic lupus erythematosus patients through the p53/p21 pathway[J]. Mol Cell Biochem, 2014, 387(1/2):27-37.

40 Zhang DY, Wang HJ, Tan YZ. Wnt/beta-catenin signaling induces the aging of mesenchymal stem cells through the DNA damage response and the p53/p21 Pathway[J]. PLoS One, 2011, 6(6):e21397.

41 Tichon A, Eitan E, Kurkalli BG, et al. Oxidative stress protection by novel telomerase activators in mesenchymal stem cells derived from healthy and diseased individuals[J]. Curr Mol Med, 2013,13(6):1010-1022.

42 Yahata T, Takanashi T, Muguruma Y, et al. Accumulation of oxidative DNA damage restricts the self-renewal capacity of human hematopoietic stem cells[J]. Blood, 2011, 118(11):2941-2950.

43 Alves H, Munoz-Najar U, De Wit J, et al. A Link between the accumulation of DNA damage and loss of multi-potency of human mesenchymal stromal cells[J]. J Cell Mol Med, 2010,14(12):2729-2738.

44 Minieri V, Saviozzi S, Gambarotta G, et al. Persistent DNA damageinduced premature senescence alters the functional features of human bone marrow mesenchymal stem cells[J]. J Cell Mol Med, 2015,19(4):734-743.

45 Yu QJ, Katlinskaya YV, Carbone CJ, et al. DNA-Damage-Induced type I interferon promotes senescence and inhibits stem cell function[J].Cell Rep, 2015, 11(5):785-797.

46 Shearier E, Xing Q, Qian ZC, et al. Physiologically low Oxygen enhances biomolecule production and stemness of mesenchymal stem cell spheroids[J]. Tissue Eng Part C Methods, 2016, 22(4):360-369.

47 Fehrer C, Brunauer R, Laschober GA, et al. Reduced Oxygen tension attenuates differentiation capacity of human mesenchymal stem cells and prolongs their lifespan[J]. Aging Cell, 2007, 6(6):745-757.

48 Kanehira M, Kikuchi T, Ohkouchi SA, et al. Targeting lysophosphatidic acid signaling retards Culture-Associated senescence of human marrow stromal cells[J]. PLoS One, 2012, 7(2):e32185.

49 Jung JW, Lee S, Seo MS, et al. Histone deacetylase controls adult stem cell aging by balancing the expression of polycomb genes and jumonji domain containing 3[J]. Cellular and Molecular Life Sciences, 2010,67(7):1165-1176.

50 鄭晨曦, 隋秉東, 胡成虎, 等. 維生素C增強(qiáng)衰老個(gè)體來源的骨髓間充質(zhì)干細(xì)胞的增殖能力[J]. 南方醫(yī)科大學(xué)學(xué)報(bào), 2015,35(12):1689-1693.

51 Gharibi B, Hughes FJ. Effects of medium supplements on proliferation,differentiation potential, and in vitro expansion of mesenchymal stem cells[J]. Stem Cells Transl Med, 2012, 1(11):771-782.

52 Mohammadi S, Nikbakht M, Malek MA, et al. Human platelet lysate as a Xeno free alternative of fetal bovine serum for thein vitroexpansion of human mesenchymal stromal cells[J]. Int J Hematol Oncol Stem Cell Res, 2016, 10(3):161-171.

53 詹菲, 徐志偉, 黃進(jìn), 等. 左歸丸對(duì)衰老大鼠骨髓間充質(zhì)干細(xì)胞增殖的影響[J]. 中華中醫(yī)藥雜志, 2015, 30(7):2518-2521.

54 Sabapathy V, Kumar S. hiPSC-derived iMSCs: NextGen MSCs as an advanced therapeutically active cell resource for regenerative medicine[J]. J Cell Mol Med, 2016, 20(8):1571-1588.

55 Sequiera GL, Saravanan S, Dhingra S. Human-induced pluripotent stem cell-derived mesenchymal stem cells as an individual-specific and renewable source of adult stem cells [J]. Methods Mol Biol, 2017,1553:183-190.

56 Lian QZ, Zhang YE, Zhang JQ, et al. Functional mesenchymal stem cells derived from human induced pluripotent stem cells attenuate limb ischemia in mice[J]. Circulation, 2010, 121(9):1113-1123.

57 Wei H, Tan G, Manasi, et al. One-step derivation of cardiomyocytes and mesenchymal stem cells from human pluripotent stem cells[J].Stem Cell Res, 2012, 9(2):87-100.

58 Frobel J, Hemeda H, Lenz M, et al. Epigenetic rejuvenation of mesenchymal stromal cells derived from induced pluripotent stem cells[J]. Stem Cell Reports, 2014, 3(3):414-422.

Research progresses in aging mechanisms of mesenchymal stem cells

Liu Yang1,2, Feng Xinmin2,Chen Tao2, Huang Zenan2, Zhang Liang2.1Graduate School of Dalian Medical University, Dalian 116044, China;2Department of Orthopedics, Clinical Medical College of Yangzhou University,Yangzhou 225001, China.

Mesenchymal stem cells (MSCs) with self-renewal and multipotential differentiation are thought to be the ideal seed cell in the repair and regeneration of orthopedic injury and degenerative disease. However, the same as any other somatic cells, MSCs are also aging upon a long-term culture which will have a negative effect in the treatment of disease. Thus it is important to understand the mechanisms of aging and look for the solution to delay or reverse the aging of MSCs. This review discuss the aging mechanisms of MSCs mainly from the telomeres and telomerase system, oxidative stress, silent information regulator protein, high glucose, Wnt/β-catenin signaling pathways activation, DNA damage and so on. We look forward to finding the solution to achieve the goal of delaying or reversing the aging process of MSCs.

Mesenchymal stem cells; Aging; Silent information regulator protein; Anti-aging

Zhang Liang, Email:zhangliang6320@sina.com

2017-03-24）

（本文編輯:陳媛媛）

10.3877/cma.j.issn.2095-1221.2017.04.010

國(guó)家自然科學(xué)基金青年基金（81401830）；中國(guó)博士后科學(xué)基金（2015M571714）；江蘇省自然科學(xué)基金青年基金（BK 20140496）

張亮，Email：zhangliang6320@sina.com

劉洋,馮新民，陳濤，等.間充質(zhì)干細(xì)胞衰老機(jī)制的研究進(jìn)展[J/CD].中華細(xì)胞與干細(xì)胞雜志（電子版），2017，7（4）：242-246.