基质血管成分与体外扩增的脂肪来源干细胞促进移植脂肪成活的对比研究

目的:比较血管基质成分(st romal vascul ar fract i on,SVF)与体外扩增的脂肪来源干细胞(adi pose-deri ved st emcel l s,ASCs)对移植脂肪成活的促进作用。方法:用手术方法切取家兔腹股沟脂肪,进行ASCs体外分离培养;取第3代ASCs分别进行成脂、成骨诱导实验,CD29和CD31流式鉴定;制备SVF,进行CD29和CD31流式鉴定;脂肪移植裸鼠实验分为3组:SVF、ASCs、和空白对照组(DMEM/F12),每组4只裸鼠,沿背部脊柱两侧对称部位4个移植位点,每组共16个注射移植位点,每点0.3ml脂肪颗粒(adi pose granul e,AG)+0.2ml细胞成分;术后4m时取材称重、固定行HE染色观察移植脂肪组织结构,行CD31免疫组化染色观察新生血管及其密度。结果:家兔ASCs体外分离培养成功,为贴壁生长,第3代细胞形态均呈长梭形。成脂诱导实验油红O染色显示形成脂滴,成骨诱导实验茜素红染色显示形成钙化结节。流式鉴定显示SVF:CD29:17.0%,CD31:1.3%;ASCs:CD29:96.2%,CD31:3.8%。SVF、ASCs和空白对照组各组移植脂肪成活量分别为0.2096±0.0024g,0.1798±0.0033g,0.1350±0.0020g,两两比较差异均有统计学意义,P<0.05。HE染色显示SVF组移植脂肪成活较好,组织结构完整,脂滴大小均一,可见脂肪组织中间有丰富的血管存在;ASCs组移植脂肪成活尚可,组织结构尚完整,脂滴大小一般均一,可见脂肪组织中有结缔组织纤维间隔和新生血管形成;空白对照组结缔组织纤维间隔明显增多,脂滴大小不一,有少量较大空泡形成。各组移植脂肪新生血管密度分别32.6±2.1条/mm2,29.3±1.6条/mm2,23.3±1.9条/mm2,两两比较均有统计学意义,P<0.05。结论:新鲜的干细胞成分SVF比体外扩增的ASCs能更好的促进移植脂肪成活。     

从脂肪抽出物的脂类和液体部分提取脂肪来源干细胞的细胞生物学研究

目的 探索从抽吸物中脂质和液体部分分离、体外培养脂肪组织来源干细胞的新方法,并通过其生长动力学、形态学、分化能力、细胞衰老和表面标记物轮廓5个方面的特征进行鉴定比较.方法 抽脂术后抽吸物分解为脂质和液体部分.脂质和液体部分分别用酶消化法和直接离心过滤法分离、培养ASCs,观察其在体外培养的形态学和生物学特点;MTT比色法测细胞活性,统计学分析;流式细胞仪测定细胞周期;随机选取3、4、6、8代做丫啶橙染色检测细胞的衰老;用流式细胞仪、免疫组织化学染色法鉴定其表面分子表达;成脂、成骨定向诱导分化,油红O染色、茜素红染色定性.结果 从吸脂抽吸物的脂质和液体两部分中都能培养出大量的ASCs,分别为PLA和LAF,呈成纤维细胞样贴壁生长,MTY测定细胞活性及细胞周期研究发现PLA、LAF这两种细胞的活力与增殖能力是非常相似的;丫啶橙染色3、4、6、8代细胞无明显衰老;流式细胞仪检测显示干细胞标志的CD29、CIM4、CD34的表达均呈阳性;免疫化学染色发现Ⅷ因子、CD31、CD105、SMA表达阳性;成脂诱导分化2周后,细胞内可见有大量脂滴,油红O染色可见胞浆内有大量红染颗粒.成骨诱导2周后,细胞可见白色矿化钙盐沉积,茜素红染色可见成骨细胞红染.结论 本实验建立了一种自人体脂肪抽吸物中脂质和液体部分分离和培养ASCs的新方法,经济简便实用,从成人脂肪抽吸物液体部分中也可以分离得到大量的可为脂肪组织工程所利用的ASCs,其细胞量与脂质来源的ASCs的量基本相同.贴壁的LAF与PLA细胞在细胞的生长动力学、形态学、细胞衰老、表面标志物和分化能力等方面具有非常相似的特性,都具有很强的增殖活性且衰老率较低,能稳定表达干细胞表面标志并能实现定向成脂、成骨多向诱导分化.这种经过最小限度人工干预的ASCs可能是将来脂肪组织工程比较理想的种子细胞之一.     

脐血间充质干细胞的分离培养鉴定及诱导分化研究

目的：分离培养人脐血间充质干细胞（human umbilical cord blood-derived mesenchymal stem cells,hUCB-MSC）,体外观察其生长特性,并在特定条件下诱导分化,探讨其成脂成骨分化能力.方法：采用沉降法和密度梯度离心结合贴壁培养法自脐血中分离间充质干细胞,倒置显微镜下观察其形态及生长情况;流式细胞仪分析细胞周期并检测细胞表面标志物;用茜素红染色和油红0染色分别鉴定其成骨成脂分化能力.结果：纯化的hUCB-MSC贴壁生长,呈均一梭形,具有较强的增值能力,流式细胞仪分析P3代hUCB-MSC稳定表达间充质干细胞表面抗原标志CD73,CD105和CD90等,不表达造血标志CD34和CD45;成骨诱导后3周后细胞茜素红染色阳性;成脂诱导3周后细胞油红0染色阳性.结论：本实验分离的hUCB-MSC具有较强的增殖能力,表达间充质干细胞的表面标记,具有成骨成脂分化潜能.     

CD204阴性人脂肪来源细胞体外成软骨潜能的初步研究

目的比较CD204+和CD204-两种人脂肪来源细胞的干细胞特性和体外诱导成软骨的能力。方法分离培养人脂肪来源细胞并进行流式细胞分选,分别获得CD204+和CD204-两种细胞。将这两种细胞培养扩增至第2代,观察其形态学特点;成脂、成骨定向诱导分化,油红O染色、茜素红染色定性;同时将两种细胞进行pellet成软骨诱导分化,比较体外诱导成软骨的能力,并以HE、Safranin O和Ⅱ型胶原免疫组化染色等进行鉴定。结果经流式细胞分选,脂肪来源细胞中CD204表达阳性率约为10%。分选后的CD204+和CD204-细胞均呈成纤维细胞样贴壁生长,但CD204-细胞具有更强的增殖能力。成脂诱导10 d后,两组细胞油红O染色均可见胞浆内有大量红染脂滴,但CD204+细胞具有更强的成脂潜能。成骨诱导2周后,两组细胞茜素红染色均可见钙结节红染,而CD204-细胞具有更强的成骨潜能。同时,细胞pellet成软骨诱导4周后,大体观可见CD204-细胞组pellet形成白色透明样软骨成分,而CD204+细胞组pellet外观微黄。HE和Ⅱ型胶原免疫组化染色均显示CD204-组pellet可见成熟软骨陷窝形成,Safranin O染色显示CD204-组软骨特异性细胞外基质沉积;而CD204+细胞组pellet则呈纤维化改变。结论脂肪组织来源的CD204-细胞具有干细胞的生物学特点,且有良好的成软骨潜能,可以成为软骨组织工程良好的种子细胞来源。     

人脂肪干细胞的培养及鉴定

目的:探讨人脂肪干细胞的分离、培养并鉴定。方法:采用胶原酶法消化人脂肪组织,培养原代脂肪干细胞,通过多种方法检测鉴定。结果:细胞贴壁生长,形态呈梭形,并呈旋涡状或放射状排列;细胞生长曲线为典型的"S"形,细胞倍增时间为26h。流式细胞仪检测结果显示CD29、CD105、CD44为阳性;CD45为阴性;成脂诱导分化的细胞经油红O染色为阳性,成骨诱导分化后茜素红染色亦为阳染。结论:通过获取人腹部脂肪组织进体外分离培养及鉴定,证实其为脂肪干细胞。     

不同组织来源间充质干细胞体外成骨分化能力的比较研究

目的比较成人脂肪间充质干细胞(ASCs)、脐带间充质干细胞(UC-MSCs)和成人骨髓间充质干细胞(BMSCs)的成骨能力,选择优势干细胞种类作为应用于骨组织工程治疗骨缺损的种子细胞。方法采用含10%胎牛血清的DMEM/Ham’s F-12培养液培养3种MSCs。取3种MSCs的P3代,通过CCK8方法检测其增殖能力;通过流式细胞仪进行鉴定;通过碱性磷酸酶(ALP)和茜素红染色检测骨分化蛋白ALP的分泌和矿化钙结节的沉积,并对钙结节进行定量分析;通过实时荧光定量PCR(RT-q PCR)方法检测骨再生相关基因的表达。结果 3种P3代MSCs在3~5d之间增殖均处于对数生长期;流式鉴定3种细胞的表面标志物阳性率:CD44、CD90和CD105均高于97%,阴性率:CD14、CD34和CD45均低于1%;ALP染色结果显示3种MSCs成骨诱导9d时,细胞内均表达ALP,茜素红染色结果显示成骨诱导18d时,均呈现较好的矿化能力,BMSCs和UC-MSCs成骨诱导后形成的钙结节无显著性差异;RT-q PCR结果显示3种MSCs成骨诱导组相比较于对照组,成骨再生相关基因Osterix、ALP、I型胶原(COL1)和骨钙素(OCN)均显著性高表达;3种MSCs成骨诱导9d时,UC-MSCs实验组的COLI基因表达显著性高于BMSCs,成骨诱导18d时,ASCs实验组的Osterix基因表达显著性高于BMSCs。结论 ASCs和UC-MSCs具有一定的成骨矿化能力,有望成为骨组织工程治疗骨缺损的种子细胞。     

左归丸和右归丸对大鼠脂肪干细胞增殖及成骨分化影响的比较

目的 比较左归丸、右归丸对大鼠脂肪干细胞( ASCs)增殖和成骨性分化的影响。方法 成年大鼠15只,随机分为3组,分别以左归丸(ZGW组)、右归丸(YGW组)和生理盐水(CON组)连续灌胃后制备含药血清。胶原酶消化法分离大鼠脂肪干细胞,成骨诱导21 d后,茜素红染色对细胞进行鉴定。分别采用CCK-8法、PNPP法测定加药诱导后ASCs的增殖能力及碱性磷酸酶表达差异;比色法检测细胞外钙离子浓度。Western Blot法测定加药诱导7d后的Runx2蛋白表达量。结果ASCs细胞呈长梭形生长,增殖旺盛;茜素红染色显示为阳性。CCK-8法检测结果显示,加药24 h及48 h时,YGW组OD值显著高于ZGW组及CON组(P<0.05);加药72 h时,ZGW组、YGW组OD值均显著高于CON组(P<0.05),而两个加药组间并无显著性差异(P > 0. 05)。PNPP检测结果显示,ZGW组、YGW组OD值均显著高于CON组(P<0.05);其中YGW组OD值显著高于ZGW组(P < 0. 05)。YGW组细胞外钙离子浓度显著升高(P < 0. 05 )。 Western Blot结果显示,YGW组Runx2蛋白表达显著上调(P<0.05)。结论 与滋肾阴法相比,温肾阳法在促进ASCs增殖及成骨分化方面均表现出明显优势。     

人脂肪细胞的可塑性

目的 探索体外培养环境下人成熟脂肪细胞的去分化现象,旨在挖掘其作为种子细胞的潜能,为组织工程研究开辟新思路.方法 自成人吸脂术后抽吸物提取成熟脂肪细胞及脂肪组织来源干细胞(adipose-derived stromal cells,ASCs),天花板贴壁培养法诱导成熟脂肪细胞去分化,观察细胞形态变化,获得去分化脂肪细胞(dedifferentiated adipocytes,DA).相同的条件下,MTT比色法比较DA、ASCs活性并绘制细胞生长曲线;流式细胞仪鉴定DA、ASCs表面分子的表达;油红O染色、茜素红染色、阿尔辛蓝染色分别鉴定DA、ASCs成脂分化、成骨、成软骨分化能力.结果 人成熟脂肪细胞在体外培养环境下能去分化为成纤维细胞状DA;MTT比色法测细胞活性:DA、ASCs均有很强的增殖能力,两者差异无统计学意义;流式细胞仪测定:DA、ASCs中HLA-ABC、CD29、CD44均为阳性,CD45、CD34、CD106均为阴性;成脂分化2周,油红O染色可见DA、ASCs内出现红色脂滴;成骨分化2周,茜素红染色可见DA、ASCs内红色钙盐沉积;成软骨分化2周,阿尔辛蓝染色可见DA、ASCs内软骨基质沉积.结论 成熟的脂肪细胞在体外培养条件下可成为DA,DA具有很强的增殖活性,表达部分干细胞特征性表面蛋白,有成骨、成软骨及强大的成脂分化能力,有望成为组织工程优秀的种子细胞.

Abstract：

Objective To explore the dedifferentiation phenomenon of human mature adipocytes cultured in vitro and to discuss the possibility of using dedifferentiation adipocytes ( DA ) as seed cells.Methods Mature adipocytes and ASCs were harvested from human fat aspirates. Mature adipocytes were cultured and induced to DA by ceiling adherent culture method. Cell morphology were observed during the whole process. Viabilities of DA and ASCs were compared by MTT chromatometry and cell growth curves were drawn based on it. Cell surface markers of DA and ASCs were detected by flow cytometry. The adipogenic,osteogenic and chondrogenic ability of DA and ASCs were assessed by oil red O staining,alizarin bordeaux staining and alcian blue staining, respectively. Regults Human mature adipocytes can dedifferentiate into fibroblast-shaped DA. MTT chromatometry assay demonstrated that DA and ASCs both had strong reproductive activity, with no significant difference between them. Flow cytometry assay demonstrated that both DA and ASCs expressed HLA-ABC, CD29 and CD44, while didn't express CD45,CD34 and CD106. After two weeks of adipogenic differentiation, lipid droplets could be displayed by oil red O staining in both DA and ASCs. After two weeks of osteogenic differentiation, calcium salts mineralization in DA and ASCs could be detected by alizarin bordeaux staining. After two weeks of chondrogenic differentiation, matrix of cartilage cells in DA and ASCs could be detected by alcian blue staining. Conclusions Mature adipocytes can be dedifferentiated into DA in vitro. DA has strong reproductive activity, as well as osteogenic, chondrogenic ability and strong adipogenic ability. It expresses some of the stem cell-related cell surface proteins and is a promising seed cell for adipose tissue engineering.     

皮肤源祖细胞的培养、鉴定和体外诱导分化

目的探讨皮肤源祖细胞的体外培养、鉴定及定向诱导成脂、成骨的方法，为组织工程提供较理想的种子细胞。方法出生1～3d的sD大鼠幼鼠皮肤，以含表皮生长因子和成纤维细胞生长因子的培养基进行培养。观察细胞生长情况，描绘生长曲线；免疫荧光鉴定细胞表达Nestin和Fibronectin情况；将第3代细胞，分别用成脂和成骨诱导液培养14d，以油红O染色、茜素红染色和免疫荧光检测皮肤源祖细胞诱导成脂、成骨情况。结果细胞呈悬浮生长，迅速增殖形成克隆球团；细胞免疫荧光表达Nestin和Fibronectin；成脂诱导14d后，细胞内大量致密颗粒形成，油红O染色可见红色脂滴；成骨诱导14d后，茜素红染色可见暗红色钙盐沉积，骨桥蛋白表达阳性显示成骨细胞形成。结论皮肤源祖细胞具有干细胞的特性，能分化为成骨细胞和脂肪细胞。     

组织工程学黄韧带干细胞的筛选与鉴定

【摘要】 目的：通过Fibronectin介导进行差别粘附法筛选出人黄韧带干细胞并进行鉴定,探讨其作为组织工程种子细胞的可行性。方法：从椎间孔镜或椎间盘镜微创手术中获取10例人黄韧带标本,机械-酶消化法联合获取原代细胞,扩增第二代后接种于Fibronectin包被过的培养瓶。流式细胞仪检测细胞表面特异性标志物。免疫组化方法测定干细胞表达相关特异性蛋白。使用干细胞诱导培养基（实验组）向成骨、成脂、成软骨多向诱导分化,使用常规生长培养基培养者作为阴性对照（对照组）,分别行茜素红染色、油红O染色、阿利辛兰染色鉴定。PT-PCR检测成骨、成脂肪、成软骨基因表达。取同一患者的髂骨骨髓提取骨髓间充质干细胞,统计学分析黄韧带干细胞与骨髓间充质干细胞的细胞周期、增殖能力和干性基因表达差异。结果：原代细胞形态呈三角形或多角形,经Fibronectin介导粘附培养后形态比较均一,呈长梭形克隆群。流式细胞仪检测筛选后的黄韧带干细胞CD90、CD73阳性率＞96.8%,CD105阳性率＞95.9%,而stro-1为阴性。实验组茜素红染色呈强阳性,油红O染色可见大小不等脂滴呈红色,阿利辛兰染色大量蛋白聚糖聚集呈蓝色,对照组均为阴性。PT-PCR测定实验组成骨基因（OC、ALP、RUNX-2）、成脂肪基因（LPL、APP、PPAR2）、成软骨基因（COL Ⅱ、AGG、SOX9）显著上调,其中RUNX-2在对照组有低水平表达。免疫组化显示黄韧带干细胞和骨髓间充质干细胞均表达a-SMA、Ⅰ型胶原、纤连蛋白,但均不表达Ⅱ型胶原。两种干细胞均超过80%的细胞比例处于G0/G1期提示都有很强自我更新能力（P>0.05）。CCK-8检测不同时间点两种细胞的增殖能力,在第1～10天时OD值逐渐增加,第10～14天达稳定状态,二者有相似的增殖能力（P>0.05）。两种干细胞均表达干性基因NONAG、OCT-4、SOX2,组间差异无显著性（P>0.05）。结论：通过Fibronectin差别粘附筛选法可有效筛选纯化出黄韧带干细胞,其可向成骨、成脂、成软骨多向分化,为组织工程技术治疗退变椎间盘提供了新的种子细胞。     

脂肪组织来源干细胞定向分化脂肪组织的体内外实验研究

目的 分离和培养脂肪组织来源干细胞（ASCs），鉴定其是否具有干细胞表面标志，研究携带GFP基因的ASCs向脂肪组织的体外定向诱导分化能力，同时判断种子细胞ASCs与Ⅰ型胶原支架混合培养后在体内构建组织工程化脂肪组织的可能性。方法 取GFP小鼠腹股沟部脂肪组织，使用酶消化法进行原代培养，流式细胞仪鉴定其表面干细胞标志，细胞传至第3代后使用脂肪分化培养基诱导2周，观察细胞形态及功能变化。将诱导分化后的细胞与支架材料混合培养后12h，将支架材料移植到裸鼠背部皮下，观察新生组织情况，并对新生组织使用HE及油红O染色进行鉴定。结果 原代培养的ASC形态类似于成纤维细胞，具有很强的增殖能力，能持续稳定表达表面干细胞标志。在脂肪分化培养基的作用下，胞浆内脂滴不断聚集，逐渐演变为成熟的脂肪细胞，油红O染色阳性。体内实验中在裸鼠皮下发现了0．5ml的新生组织块，常规病理及油红O染色均证实其为成熟脂肪组织块。结论 脂肪组织来源的干细胞ASC能在体外定向诱导分化为成熟脂肪细胞，且ASC能作用种子细胞与Ⅰ型胶原支架在体内成功构建脂肪组织。     

Cell biological study of cultured cells derived from the fattyoffluid portions of liposuction aspirates]

OBJECTIVE: To explore an approach to isolate and culture the Adipose derived stem cells (ASCs) from the fatty and the fluid portions of liposuction aspirates, and to investigate the growth kinetics, morphology, differentiation capability, cell senescence, surface marker profiles of the ASCs. METHODS: The liposuction aspirates were divided into fatty portion and liquid portion. ASCs were isolated from each portion by collagenase digestion and directly centrifugate and cultured to observe the morphology and biology characters in vitro. Cell activity was studied by MTT chromatometry and analyzed statistically. Cell cycle was detected by flow cytometry. Cells were randomly selected from the 3rd, 4th, 6th, 8th generation cells to dectect senescence of ASCs by acridine orange staining. The cell surface markers were detected by flow cytometry and immunohistochemistry. Adipogenic and osteogenic lineage differentiation of ASCs was assessed by Oil Red O and alizarin bordeaux staining respectively. RESULTS: A large amount of ASCs could be islated and cultured both from the fatty portion and the liquid portion, including PLA cells and LAF cells which had fibroblastic characters with strong viability and proliferative activity. The statistical result indicated that the cell activity of PLA cells and LAF cells was very similar. ASCs from passage 3, 4, 6, 8 didn't show insenecence. CD29, CD44, CD34, which were the markers of mesenchymal stem cells, vWF, CD31, CD105, SMA were all expressed in ASCs. Adipogenic differentiation of ASCs was assessed by Oil Red O staining after 2 weeks. The cells contained many lipid-filled droplets. After 2 weeks' osteogenic induction, cells were positively stained by alizarin Bordeaux. CONCLUSIONS: The method can isolate ASCs by directly centrifugate from the fatty and the fluid portions of human liposuction aspirates. The way of culture is convenient and economical. ASCs isolated from the liquid and fatty portions of liposuction aspirates show identical in cells numbers and quality. LAF cells and PLA cells have similar characters in growth dynamics, morphology, cell senescence, surface marker profiles and differentiation ability, etc. Expression of the cell surface marker of stem cells is also observed in ASCs. ASCs can differentiate into adipose and osteogenesis directionally. The results suggest that the ASCs, which are isolated with minimum intervention, may be the ideal seed cells for adipose tissue engineering in future.     

Adipose tissues differentiated by adipose-derived stem cells harvested from transgenic mice

Objective: To induce adipocyte differentiation in vitro by adipose-derived stromal cells (ASCs) harvested from transgenic mice with green fluorescent protein (GFP) and assess the possibility of constructing adipose tissues via attachment of ASCs to typeⅡcollagen scaffolds. Methods: Inguinal fat pads from GFP transgenic mice were digested by enzymes for isolation of ASCs (primary culture). After expansion to three passages of ASCs, the cells were incubated in an adipogenic medium for two weeks, and the adipocyte differentiation by ASCs in vitro was assessed by morphological observation and Oil Red O staining. Then they were attached to collagen scaffolds and co-cultured for 12 hours, followed by hypodermic implantation to the dorsal skin of nude mice for 2 months. The newly-formed tissues were detected by HE staining. Results: The cultured primary stem cells were fibroblast-like and showed active proliferation. After being incubated in an adipocyte differentiation medium, the lipid droplets in the cytoplasm accumulated gradually and finally developed into mature adipocytes, which showed positive in Oil Red O staining. A 0. 5-cm3 new tissue clot was found under the dorsal skin of the nude mice and it was confirmed as mature adipose tissues by fluorescent observation and HE staining. Conclusions: ASCs can successfully differentiate adipose tissues into mature adipocytes, which exhibit an adipocyte-like morphology and express as intracytoplasmic lipid droplets. It is an efficient model of adipose tissues engineered with ASCs and type I collagen scaffolds.     

低氧对脂肪间充质干细胞向雪旺细胞分化的影响

目的探讨低氧对脂肪间充质干细胞（ADMSCs）向雪旺细胞（SCs）分化能力的影响。方法分离培养SD大鼠ADMSCs并用流式细胞仪、茜素红染色、油红O染色鉴定。将成功分离的ADMSCs随机分为3组：常氧诱导组,在常氧条件下（5％CO2,21％O2,37℃）诱导;低氧处理＋常氧诱导组,低氧处理（5％CO2,0．5％O2,37℃）后在常氧条件下诱导;低氧诱导组,低氧条件下（5％CO2,0．5％O2,37℃）诱导。观察各组细胞形态,MTT法检测细胞增殖情况,免疫荧光染色和Westernblot检测SCs标志物GFAP和S-100的表达。结果细胞分离后,经流式细胞仪分析可见细胞表面CD44阳性、CD45阳性、CD90阳性,茜素红及油红O染色均为阳性。MTT法检测结果：低氧处理＋常氧诱导组A值为0．861±0．039,高于常氧诱导组0．837±0．017,差异具有统计学意义（P〈0．05）;低氧诱导组A值为0．931±0．041,均高于常氧诱导组和低氧处理＋常氧诱导组（P均〈0．05）。免疫荧光染色发现常氧诱导组和低氧处理＋常氧诱导组大量细胞GFAP和S-100表达阳性,低氧诱导组仅少量细胞S-100和GFAP表达阳性。Westernblot检测发现常氧诱导组S-100蛋白表达最高,低氧处理＋常氧诱导组GFAP蛋白表达最高,低氧诱导组S-100蛋白、GFAP蛋白表达均最低。结论低氧抑制ADMSCs向SCs的分化,低氧处理后的ADMSCs在常氧条件下仍可向SCs分化。     

肾癌中CD133＋细胞和CD133-细胞生物学特性差异的研究

目的：研究人肾癌细胞系786-O和OS-RC-2中CD133＋细胞和CD133-细胞的生物学特性差异。方法：应用流式细胞仪检测肾癌细胞系786-O和OS-RC-2中CD133的膜表达状况;以免疫磁珠法将CD133＋细胞和CD133-细胞分离纯化,比较两者在光镜下的形态、生长特点、体外增殖能力、长期分化能力及体外克隆形成率的差异;流式细胞术分析两者细胞周期分布的变化,采用MTT法分析两者对化疗药物顺铂（DDP）（10、20、50、80／Lg／ml）敏感性的差别。结果：786-O和0S-RC-2均有CD133的少量表达,表达率为（8．12±0．86）％、（6．83±0．20）％,CD133＋细胞和CD133-细胞相比具有较强的体外增殖、克隆形成及长期分化能力;细胞耐药性实验表明CD133＋细胞、CD133-细胞对50bcg／m1DDP敏感性差异有统计学意义（P〈0．05）。结论：肾癌细胞系中CD133＋细胞具有肾癌干细胞的部分特征,能否作为肾癌干细胞的表面标志还需要进一步的实验加以明确。