貢 波 徐志國(guó) 王紹紅 程紅玲 劉 超 閆銘杰
(浙江省臍帶血造血干細(xì)胞庫(kù),協(xié)和華東干細(xì)胞基因工程有限公司,湖州313000)
臍帶間充質(zhì)干細(xì)胞體外對(duì)臍帶血CD4+T淋巴細(xì)胞免疫調(diào)節(jié)作用的研究
貢 波 徐志國(guó) 王紹紅 程紅玲 劉 超 閆銘杰①
(浙江省臍帶血造血干細(xì)胞庫(kù),協(xié)和華東干細(xì)胞基因工程有限公司,湖州313000)
目的:研究臍帶間充質(zhì)干細(xì)胞(UCMSCs)在體外對(duì)異源臍血CD4+T淋巴細(xì)胞增殖、凋亡及向CD4+CD25+調(diào)節(jié)性T細(xì)胞(Treg)分化的免疫調(diào)控作用。方法:建立不同比例UCMSCs與植物血凝素(PHA)活化的臍血CD4+T淋巴細(xì)胞共培養(yǎng),或與Transwell共培養(yǎng)體系,檢測(cè)CD4+T淋巴細(xì)胞增殖情況、CD4+CD25+/CD4+比率的變化及調(diào)節(jié)性T淋巴細(xì)胞標(biāo)志基因Foxp3相對(duì)表達(dá)量的變化。同時(shí),建立UCMSCs與地塞米松(DXM)刺激的臍血CD4+T淋巴細(xì)胞共培養(yǎng),或與Transwell共培養(yǎng)體系,檢測(cè)CD4+T淋巴細(xì)胞凋亡情況。結(jié)果:PHA活化的臍血CD4+T淋巴細(xì)胞在與UCMSCs共培養(yǎng)3 d后,較無(wú)UCMSCs對(duì)照組相比,細(xì)胞數(shù)量明顯減少(P<0.05)且CD4+CD25+/CD4+T淋巴細(xì)胞的比率及Foxp3的相對(duì)表達(dá)量顯著增加(P<0.01),并隨UCMSCs數(shù)量增加,作用增強(qiáng)(P<0.05)。DXM誘導(dǎo)的臍血CD4+T淋巴細(xì)胞與UCMSCs共培養(yǎng)7 d后,細(xì)胞凋亡比率較無(wú)UCMSCs對(duì)照組顯著降低(P<0.01)。上述效應(yīng)在Transwell共培養(yǎng)中部分減弱但無(wú)法消除。結(jié)論:UCMSCs對(duì)臍血CD4+T細(xì)胞介導(dǎo)的免疫反應(yīng)具有負(fù)調(diào)節(jié)作用,其作用主要表現(xiàn)在對(duì)細(xì)胞增殖能力和分化的調(diào)控而不是促進(jìn)凋亡。
臍帶間充質(zhì)干細(xì)胞;臍血CD4+T細(xì)胞;CD4+CD25+調(diào)節(jié)性T細(xì)胞;增殖;分化;凋亡
間充質(zhì)干細(xì)胞(Mesenchymal stem cells,MSCs)來(lái)源于發(fā)育早期的中胚層,最初由Friedenstein等[1]從骨髓中發(fā)現(xiàn)。MSCs擁有的天然免疫調(diào)節(jié)作用,在體外表現(xiàn)出了非MHC限制性、劑量依賴(lài)性抑制T淋巴細(xì)胞反應(yīng)的特性[2],使其在器官移植、骨髓移植和自身免疫性疾病的防治中,發(fā)揮了重要作用[3,4]。
目前,自身免疫性疾病是繼心血管疾病、癌癥后威脅人類(lèi)健康的第三大殺手,已被列入我國(guó)十類(lèi)重大疾病之一。某些自身免疫性疾病,如重癥肌無(wú)力(Myasthenia gravis,MG)、皮肌炎(Dermatomyositis,DM)等,對(duì)嬰幼兒童造成的困擾遠(yuǎn)遠(yuǎn)超過(guò)成人。因此,研究MSCs對(duì)臍帶血來(lái)源T細(xì)胞的免疫調(diào)節(jié)功能,對(duì)新生兒和兒童自身免疫性疾病的臨床研究具有重要提示意義。
人臍帶間充質(zhì)干細(xì)胞(Human umbilical mesenchymal stem cells,hUCMSCs)與其他的成體干細(xì)胞相比,增殖分化能力更強(qiáng),已成為難治性疾病潛在的治療手段[5,6]。UCMSCs其在體外發(fā)揮抑制作用時(shí)沒(méi)有MHC限制性[7],能顯著降低移植物抗宿主病(Graft versus host disease,GVHD)反應(yīng),且同種異源性UCMSCs有不引起副反應(yīng)產(chǎn)生的特點(diǎn)[8]。hUCMSCs的培養(yǎng)及MSCs對(duì)成人外周血T淋巴細(xì)胞的調(diào)控研究已經(jīng)為眾多研究者報(bào)道[9-11],但有關(guān)hUCMSCs對(duì)臍帶血來(lái)源的免疫細(xì)胞調(diào)控作用的研究報(bào)道相對(duì)較少。
1.1 材料
1.1.1 主要試劑 ALyS505N-175培養(yǎng)基為日本細(xì)胞研究所產(chǎn)品;DMEM/F12培養(yǎng)基為Invitrogen產(chǎn)品;植物血凝素(Phytohemagglutinin,PHA)、地塞米松(Dexamethasone,DXM)為Sigma產(chǎn)品;胎牛血清(FBS)為Excell產(chǎn)品;淋巴細(xì)胞分離液(Ficoll,1.077 g/L)為深圳達(dá)科為生物技術(shù)股份有限公司生產(chǎn);CD4+T細(xì)胞分選試劑盒(CD4+T Cell Isolation Kit)為美天旎產(chǎn)品;絲裂霉素C(Mitomycin C,MMC)為Roche產(chǎn)品;RNA提取試劑盒(Ultrapure RNA Kit)為北京康為世紀(jì)生物科技有限公司生產(chǎn);逆轉(zhuǎn)錄試劑盒(ReverTra Ace qPCR RT Kit)、Real-time PCR試劑盒(SYBR Green Realtime PCR Master Mix-Plus-)為日本TOYOBO產(chǎn)品;CD73-PE抗體、CD90-PE抗體、CD105-FITC抗體、CD29-PE抗體、CD44-PE抗體、CD31-PE抗體、CD34-PE抗體、CD45-FITC抗體、CD4-FITC抗體、CD25-PE抗體、凋亡檢測(cè)試劑盒(Annexin V-FITC/PI)為美國(guó)BD產(chǎn)品。
1.1.2 樣本來(lái)源 實(shí)驗(yàn)?zāi)殠Ъ澳殠а獊?lái)源于浙江省臍帶血造血干細(xì)胞庫(kù)的捐獻(xiàn)標(biāo)本,HIV、HBV、HCV、CMV、梅毒螺旋體等檢測(cè)均為陰性。
1.2 實(shí)驗(yàn)方法
1.2.1 UCMSCs的分離培養(yǎng)及表面標(biāo)志檢測(cè) 將臍帶消毒后剪成2 mm3左右小塊,置于細(xì)胞培養(yǎng)瓶中,加入15 ml含10%胎牛血清的DMEM/F12培養(yǎng)液,5%CO2,37℃培養(yǎng)箱培養(yǎng)。取P3代細(xì)胞,將細(xì)胞密度調(diào)整為106個(gè)/ml,單抗(CD73-PE、CD90-PE、CD105-FITC、CD29-PE、CD44-PE、CD31-PE、CD34-PE、CD45-FITC)標(biāo)記,流式細(xì)胞儀檢測(cè)細(xì)胞免疫表型。
1.2.2 臍帶血CD4+T(UCB-CD4+T)淋巴細(xì)胞的獲取 采用Ficoll密度梯度離心法取得臍帶血單個(gè)核細(xì)胞(UCB-MNC)后,用含10%胎牛血清的ALyS505N-175培養(yǎng)基將UCB-MNC調(diào)整至106個(gè)/ml,置培養(yǎng)瓶中37℃孵育30 min,收集未貼壁細(xì)胞,按CD4+T Cell Isolation Kit說(shuō)明書(shū)操作,分選得到CD4+T淋巴細(xì)胞,流式細(xì)胞儀檢測(cè)CD4表達(dá)量。收集貼壁的抗原提呈細(xì)胞(Antigen-presenting cells,APCs),調(diào)整密度至104個(gè)/ml待用。
1.2.3 不同數(shù)量UCMSCs與PHA作用下的UCB-CD4+T細(xì)胞共培養(yǎng) 將達(dá)到80%融合的P3代UCMSCs分別以1×105個(gè)/孔和1×104/孔個(gè)兩種數(shù)量接種于24孔培養(yǎng)板或Transwell小室中,2 h后加入終濃度10 μg/ml的MMC去增殖處理3 h。將分選得到的臍帶血CD4+T淋巴細(xì)胞以1×106個(gè)/孔的數(shù)量接種至含UCMSCs的培養(yǎng)板中,同時(shí)每孔接種1×103個(gè)APCs,并加入終濃度為10 μg/ml的PHA。接觸共培養(yǎng)1×105、1×104UCMSCs設(shè)為C1、C2組,Transwell 1×105、1×104UCMSCs設(shè)為D1、D2組,臍帶血CD4+T細(xì)胞+PHA為對(duì)照組A,臍帶血CD4+T細(xì)胞+UCMSCs為對(duì)照組B,各組均設(shè)6個(gè)復(fù)孔。
培養(yǎng)板置于37℃、5%CO2孵箱中連續(xù)培養(yǎng)3 d后,收集細(xì)胞PBS洗滌離心后,鏡下計(jì)數(shù)。各組收集3個(gè)孔的細(xì)胞,洗滌后加入CD4-FITC、CD25-PE抗體各20 μl室溫避光孵育30 min,PBS緩沖液混勻離心后,流式細(xì)胞儀檢測(cè)CD4+CD25+表達(dá)。
各組收集3個(gè)孔的細(xì)胞提取總RNA,反轉(zhuǎn)錄得到cDNA后,進(jìn)行Real-time PCR,檢測(cè)調(diào)節(jié)性T細(xì)胞標(biāo)志基因Foxp3表達(dá)量的變化,目標(biāo)基因Foxp3引物F:AACAGCACATTCCCAGAGTTCCT,R:CATTGAGTGTCCGCTGCTTCT;內(nèi)參基因GAPDH引物F:CTCTGCTCCTCCTGTTCGAC,R:ACGACCAAATCCGTTGACTC;在測(cè)定兩個(gè)基因的擴(kuò)增效率后,使用2-ΔΔCT法分析基因表達(dá)量的差異。
1.2.4 UCMSCs與DXM作用下UCB-CD4+T細(xì)胞共培養(yǎng) 將達(dá)到80%融合的P3代UCMSCs以5×104/孔的數(shù)量接種于24孔培養(yǎng)板或Transwell小室中,2 h后加入終濃度10 μg/ml的MMC增殖處理3 h。將分選得到的臍帶血CD4+T淋巴細(xì)胞以5×105個(gè)/孔的數(shù)量接種至含UCMSCs的培養(yǎng)板中,并加入終濃度為10-5mol/L的DXM。設(shè)置無(wú)MSCs對(duì)照組A,24孔板實(shí)驗(yàn)組B,Transwell實(shí)驗(yàn)組C,同時(shí)設(shè)置不含DXM對(duì)照組1,含DXM實(shí)驗(yàn)組2,各組均設(shè)3個(gè)復(fù)孔。
培養(yǎng)板置于37℃、5%CO2孵箱中連續(xù)培養(yǎng)7 d后,PBS洗滌離心后將細(xì)胞調(diào)整至106個(gè)/ml,取200 μl細(xì)胞懸液加入5 μl的AnnexinV-FITC混勻避光,室溫孵育15 min,加入5 μl的PI染色,補(bǔ)加200 μl PBS后流式檢測(cè)細(xì)胞凋亡。
2.1 UCMSCs的分離培養(yǎng)及流式細(xì)胞術(shù)檢測(cè) UCMSCs呈貼壁生長(zhǎng),流式結(jié)果顯示高表達(dá)CD73、CD90、CD105、CD29和CD44,極少表達(dá)CD31、 CD34、CD45,說(shuō)明細(xì)胞純度較好,見(jiàn)圖1。
2.2 共培養(yǎng)后細(xì)胞計(jì)數(shù) 在 PHA刺激下,可見(jiàn)細(xì)胞分裂活化聚集成團(tuán),形成明顯的細(xì)胞團(tuán)塊。第3天進(jìn)行細(xì)胞計(jì)數(shù),實(shí)驗(yàn)組細(xì)胞數(shù)量均少于對(duì)照組A,且UCMSCs數(shù)量越多,細(xì)胞數(shù)量越少,在相同UCMSCs數(shù)量的前提下,Transwell共培養(yǎng)組細(xì)胞數(shù)多于接觸共培養(yǎng)組。鏡下細(xì)胞狀態(tài)以一組實(shí)驗(yàn)為例見(jiàn)圖2,細(xì)胞數(shù)量統(tǒng)計(jì)見(jiàn)圖3。
2.3 流式測(cè)定CD4+CD25+細(xì)胞比例變化 臍血CD4+T細(xì)胞經(jīng)PHA活化后,CD4+CD25+/CD4+比率比活化前明顯升高,而經(jīng)過(guò)與UCMSCs共培養(yǎng)3 d后,雙陽(yáng)性比率提高更為顯著,效應(yīng)與UCMSCs數(shù)量呈正相關(guān),Transwell共培養(yǎng)組也呈現(xiàn)出相同趨勢(shì),但效應(yīng)較接觸共培養(yǎng)組弱。雙標(biāo)流式圖以一組實(shí)驗(yàn)為例見(jiàn)圖4,數(shù)據(jù)統(tǒng)計(jì)見(jiàn)圖5。
圖1 UCMSCs表型流式細(xì)胞術(shù)檢測(cè)結(jié)果Fig.1 UCMSCs surface marker detected by flow cytometryNote: Cells were labeled with antibodies against human antigens CD73,CD90,CD105,CD29,CD44,CD31,CD34,CD45,as analyzed by FACS.
圖2 共培養(yǎng)后各組CD4+細(xì)胞形態(tài)(×40)Fig.2 Morphology of CD4+ cells in each group after coculture(×40)
圖3 共培養(yǎng)3 d后細(xì)胞計(jì)數(shù)結(jié)果Fig.3 Results of cell count after coculture for 3 daysNote: Group C2,Group D1 vs Group A,Group C2 vs Group C1,*.P<0.05;Group C1 vs Group A,Group D1 vs Group C1,**.P<0.01.
2.4 Real-time PCR測(cè)定Foxp3表達(dá)量的變化 通過(guò)Real-time PCR發(fā)現(xiàn),各實(shí)驗(yàn)組目標(biāo)基因Foxp3相對(duì)表達(dá)量均明顯高于對(duì)照組A(P<0.01),且UCMSCs的數(shù)量越多Foxp3基因的相對(duì)表達(dá)量越高(P<0.01)。在較高濃度MSCs組中,接觸共培養(yǎng)與Transwell共培養(yǎng)也表現(xiàn)出了一定差異(P<0.05),PCR擴(kuò)增曲線(xiàn)及各組Foxp3基因相對(duì)表達(dá)量見(jiàn)圖6。
2.5 UCMSCs對(duì)DXM作用下臍血CD4+T細(xì)胞凋亡的影響 UCMSCs對(duì)臍血CD4+T細(xì)胞早期凋亡和晚期凋亡均有明顯抑制作用(P<0.01),在早期凋亡組中接觸共培養(yǎng)比Transwell共培養(yǎng)具有更為明顯的抑制能力(P<0.01)。凋亡流式圖以一組實(shí)驗(yàn)為例見(jiàn)圖7,凋亡率統(tǒng)計(jì)及分析見(jiàn)圖8。
圖4 共培養(yǎng)3 d后各組CD4CD25雙標(biāo)流式檢測(cè)結(jié)果Fig.4 Results of flow cytometry double marked assay in each group after coculture for 3 days
圖5 共培養(yǎng)3 d后各組細(xì)胞CD4+CD25+/CD4+比率(%)Fig.5 Rate of CD4+CD25+/CD4+ in each group after coculture for 3 days(%)Note: Group C2 vs Group A,Group D2 vs Group D1,*.P<0.05;Group C1 vs Group A,Group D1 vs Group A,Group C2 vs Group C1,Group D1 vs Group C1,**.P<0.01.
圖6 目標(biāo)基因Foxp3的PCR擴(kuò)增曲線(xiàn)及2-ΔΔCT法計(jì)算的各組相對(duì)表達(dá)量的變化Fig.6 PCR amplification curve of target gene Foxp3 and 2-ΔΔCT method to calculate change of relative expression of each groupNote: Group D1 vs Group C1,*.P<0.05;All groups vs Group A,Group C2 vs Group C1,Group D2 vs Group D1,**.P<0.01.
圖7 共培養(yǎng)7 d后各組流式凋亡檢測(cè)結(jié)果Fig.7 Results of apoptosis by flow cytometry assay in each group after coculture for 7 days
圖8 共培養(yǎng)7 d后各組凋亡率分析Fig.8 Apoptosis rate analysis in each group after coculture for 7 daysNote: Early apoptosis Group B2,Group C2 vs Group A2,Group C2 vs Group B2;Late apoptosis Group B2,Group C2 vs Group A2,**.P<0.01.
調(diào)節(jié)性T細(xì)胞(Treg)是T細(xì)胞中的一個(gè)特殊亞群,它們能夠抑制免疫系統(tǒng)的激活,對(duì)維持自身抗原的耐受和免疫細(xì)胞群體的穩(wěn)態(tài)具有重要意義。CD4+CD25+Foxp3+Treg細(xì)胞來(lái)源于胸腺,對(duì)自身反應(yīng)性T細(xì)胞具有抑制作用,在GVHD的預(yù)防和治療中起到了至關(guān)重要的作用,因此,我們?cè)谘芯縐CMSCs對(duì)臍血淋巴細(xì)胞免疫調(diào)控的研究中,重點(diǎn)關(guān)注對(duì)CD4+CD25+Foxp3+Treg細(xì)胞的調(diào)控。為了避免其他亞群淋巴細(xì)胞的干擾,我們選用經(jīng)免疫磁珠分選純化過(guò)的CD4+T淋巴細(xì)胞用于本次研究。
有研究發(fā)現(xiàn),T細(xì)胞的增殖可被MSCs抑制,這種作用與多種趨化因子如CXCR3、VCAM-1和ICAM-1等的高表達(dá)相關(guān)[12],另外,抑制性分子PD-1及其配體在MSCs抑制T淋巴細(xì)胞增殖的過(guò)程中也發(fā)揮了一定作用[13]。還有報(bào)道表明,MSCs可促進(jìn)CD4+CD25+Foxp3+Treg細(xì)胞的產(chǎn)生[14]。MSCs表達(dá)Notch1配體Delta-like ligand(DLL)1、3、4及Jagged1、2,Notch1信號(hào)活化可誘導(dǎo)CD4+T淋巴細(xì)胞向Treg分化[15]。另一研究證實(shí),MSCs能夠分泌血紅素氧合酶(Hemeoxygenaes-1,HO-1),對(duì)高表達(dá)IL-10的Treg(Tr1)產(chǎn)生重要的誘導(dǎo)作用,但目前尚不清楚這一作用的明確機(jī)制[16]。
本研究顯示,與UCMSCs的共培養(yǎng)后由PHA刺激的臍血CD4+T淋巴細(xì)胞增殖受到了明顯抑制,且UCMSCs數(shù)量越多,這種抑制作用越強(qiáng),呈正相關(guān)。同時(shí),通過(guò)Transwell培養(yǎng)板將UCMSCs和臍血CD4+T淋巴細(xì)胞分隔共培養(yǎng),這種抑制增殖趨勢(shì)相似,但大大減弱了抑制增殖的效果,在高濃度UCMSCs存在時(shí)與接觸培養(yǎng)組呈顯著差異(P<0.01),提示細(xì)胞間的直接接觸,抑制了CD4+T淋巴細(xì)胞的增殖,并且起到了主要作用。通過(guò)凋亡實(shí)驗(yàn)我們發(fā)現(xiàn),UCMSCs對(duì)臍血CD4+T細(xì)胞早期凋亡和晚期凋亡均有明顯的抑制作用,其中兩種細(xì)胞接觸共培養(yǎng)比Transwell共培養(yǎng)具有更為明顯的抑制能力。另外,經(jīng)PHA活化的臍血CD4+T細(xì)胞與UCMSCs共培養(yǎng)3 d后,CD4+CD25+雙陽(yáng)性細(xì)胞比率顯著提高,與UCMSCs數(shù)量呈正相關(guān)。同時(shí),在較高濃度MSCs組中,Transwell共培養(yǎng)組雙陽(yáng)性比率低于接觸培養(yǎng)組(P<0.01),但也顯著高于對(duì)照組A(P<0.01),提示可溶性細(xì)胞因子也促進(jìn)了CD4+CD25+雙陽(yáng)性細(xì)胞的產(chǎn)生。通過(guò)對(duì)調(diào)節(jié)性T細(xì)胞的標(biāo)志性分子叉狀頭轉(zhuǎn)錄因子Foxp3的Real-time PCR檢測(cè),顯示各實(shí)驗(yàn)組相對(duì)于對(duì)照組A,F(xiàn)oxp3基因的表達(dá)量顯著上升(P<0.01),趨勢(shì)與流式CD4+CD25+雙陽(yáng)性細(xì)胞比率的升高基本一致。說(shuō)明UCMSCs的存在,確實(shí)提高了CD4+T細(xì)胞中CD4+CD25+調(diào)節(jié)性T細(xì)胞的比率。
UCMSCs對(duì)臍血CD4+T細(xì)胞存在免疫抑制效應(yīng),這種效應(yīng)一方面表現(xiàn)在顯著抑制臍血CD4+T細(xì)胞的增殖,從而減少免疫應(yīng)答的產(chǎn)生;另一方面,UCMSCs誘導(dǎo)了CD4+T細(xì)胞向CD4+CD25+Foxp3+調(diào)節(jié)性T細(xì)胞的分化,而CD4+CD25+Foxp3+調(diào)節(jié)性T細(xì)胞也具有負(fù)免疫調(diào)控的作用。這兩種調(diào)控效應(yīng)均與UCMSCs數(shù)量呈正相關(guān),并且通過(guò)細(xì)胞與細(xì)胞接觸及釋放可溶性細(xì)胞因子實(shí)現(xiàn)。另外,UCMSCs對(duì)臍血CD4+T細(xì)胞的抑凋亡作用,也說(shuō)明了免疫抑制效應(yīng)主要集中在對(duì)細(xì)胞功能和分化的調(diào)控而不是促進(jìn)凋亡。UCMSCs對(duì)臍血CD4+T細(xì)胞存在的免疫抑制效應(yīng),為新生兒和兒童自身免疫性疾病及GVHD的臨床研究,提供了新的思路。
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[收稿2016-08-04]
(編輯 倪 鵬)
Immunoregulation study of UCMSCs on UCB CD4+T lymphocytes in vitro
GONGBo,XUZhi-Guo,WANGShao-Hong,CHENGHong-Ling,LIUChao,YANMing-Jie.
ZhejiangProvinceUmbilicalCordBloodStemCellBank,EasternUnionStemCellGeneEngineeringCo.Ltd.,Huzhou313000,China
Objective:Immunoregulation study of umbilical mesenchymal stem cell (UCMSCs) on allogeneic umbilical cord blood(UCB) CD4+T lymphocytes,which proliferation,apoptosis and the differentiation to CD4+CD25+regulatory T cell (Treg) in vitro.Methods: Establishing on direct contact or transwell co-culture system,adopt in different proportion of UCMCs with phytohaemagglutinin (PHA)-activated UCB CD4+T lymphocytes were co-cultured.The proliferation of lymphocyte,percent of CD4+CD25+/CD4+and Foxp3 expression,regulatory T cell marker gene were measured.Apoptosis of CD4+T lymphocytes were observed in the direct contact or transwell coculture system of UCMSCs with desamethason(DXM)-stimulated UCB CD4+T lymphocytes.Results: The UCB CD4+T lymphocytes cocultured with UCMSCs with PHA-activating for 3 days,compared with the UCMSCs free control group,the amount of cells was reduced noticeably(P<0.05) and the percent of CD4+CD25+in CD4+T lymphocytes and Foxp3 expression significantly increased(P<0.01) in a dose dependent way(P<0.05).The UCB CD4+T lymphocytes cocultured with UCMSCs with DXM-inducing for 7 days,the apoptosis rate was significantly lower than that of the control group without UCMSCs (P<0.01).These effects were partially attenuated in transwell coculture but could not be eliminated.Conclusion: UCMSCs are negative effect on UCB CD4+T lymphocytes-mediated immunity effects,and mainly manifested in the regulation on cell proliferate ability and differentiation rather than promoting apoptosis.
Umbilical cord mesenchymal stem cells (UCMSCs);Umbilical cord blood CD4+T lymphocytes;CD4+CD25+regulatory T cell;Proliferation;Differentiation;Apoptosis
10.3969/j.issn.1000-484X.2017.02.012
貢 波(1985年-),男,碩士,中級(jí)工程師,主要從事臍帶間充質(zhì)干細(xì)胞分離培養(yǎng)、生物學(xué)特性及免疫調(diào)控功能的研究,E-mail:gongbo-1019@163.com。
及指導(dǎo)教師:徐志國(guó)(1979年-),男,碩士,副高級(jí)工程師,主要從事臍帶血造血干細(xì)胞庫(kù)的建設(shè)和質(zhì)量運(yùn)行、臍帶間充質(zhì)干細(xì)胞的臨床研究, E-mail:huadongstemcell@163.com。
R392.12
A
1000-484X(2017)02-0220-06
①中國(guó)醫(yī)學(xué)科學(xué)院血液學(xué)研究所,天津300020。