干细胞样U87胶质瘤细胞向内皮细胞转分化及VEGFR2人源化单克隆抗体的干预作用

目的建立干细胞样U87胶质瘤细胞向内皮细胞的转分化模型,初步评价抗VEGFR2人源化单克隆抗体对转分化的影响。方法体外诱导培养获得干细胞样U87胶质瘤细胞;流式细胞仪检测CD133+细胞含量;定量PCR检测CD133、Nestin以及VEGFR2表达;观察干细胞样U87细胞的致瘤能力、肿瘤生长速度和CD31表达情况;观察干细胞样U87细胞在Matrigel上血管状结构形成和CD31、CD34、vW F表达情况;建立干细胞样U87细胞向内皮细胞的转分化模型,评价抗VEGFR2人源单抗对转分化的抑制作用。结果获得干细胞球样的U87细胞,其肿瘤生长更快,且血管增多,接种在Matrigel上形成血管状结构,CD31、CD34、vW F表达增加;抗VEGFR2人源单抗使血管状结构和CD31、CD34、vW F表达均减少。结论成功建立干细胞样U8细胞向内皮细胞转分化模型,VEGFR2人源化单抗对转分化有抑制作用。     

CD133阳性肝癌细胞周期分析

目的:免疫磁珠分选法分选肝癌细胞系SMMC7721中CD133+细胞,并检测CD133+和CD133-细胞周期.方法:采用免疫磁珠分选法分选肝癌细胞系SMMC7721中CD133+和CD133-细胞,乙醇固定,PI染色,流式细胞仪(FCM)检测细胞周期.结果:CD133+细胞在SMMC7721中所占比例为0.9%,FCM检测显示SMMC7721细胞株中CD133.细胞周期处于G0/G1期的比例为85.3%,而CDl33.细胞G0/G1期的比例相对较低,为61.6%,未分选细胞居中,为68.9%.结论:人肝癌细胞系SMMC7721中CD133+亚群,比例约为0.9%,且CD133.细胞多处于静止期,具有一般干细胞的特性.     

Effects of temozolomide chemotherapy on CD133+ cells in glioma U251

目的 探讨化疗耐药与肿瘤干细胞之间的关系.方法 分析替莫唑胺(TMZ)作用前后胶质瘤U251中CD133蛋白表达及阳性细胞数量的变化.通过MTT实验评估U251细胞对TMZ的敏感性.利用免疫荧光染色和Western blot检测TMZ作用前后U251细胞中CD133、巢蛋白(nestin)、神经元特异性烯醇化酶(NSE)、神经胶质细胞纤维酸性蛋白(GFAP)等蛋白表达.结果 U251细胞中未加入TMZ组的CD133+细胞比例约为0.060±0.003,明显低于加入TMZ组的0.337±0.012(P＜0.05).在TMZ作用后,CD133和nestin表达明显增加,而GFAP和NSE表达明显减少(P＜0.05).结论 胶质瘤U251细胞株中存在CD133+细胞,大部分具有脑肿瘤干细胞特性;在TMZ作用后,这类细胞比例增加,提示其与化疗耐药有关.     

免疫磁珠分选肾癌干细胞及端粒酶活性在肾癌干细胞中的表达

目的 探讨肾癌干细胞端粒酶活性的改变.方法 利用无血清悬浮培养的方法培养富集肾癌干细胞,流式细胞分析技术检测其CD133的表达,用免疫磁珠分选系统分离CD133+及CD133-细胞,以正常肾组织细胞作为阴性对照,采用TRAP实时定量的方法分别检测肾癌CD133+及CD133-细胞端粒酶活性.结果 CD133+细胞和CD133-端粒酶活性分别为(20.54士2.45)和(20.69±2.53)Ct值,而正常肾组织没有检测出端粒酶活性(P<0.05).结论 肾癌干细胞端粒酶活性明显增高.     

替莫唑胺化疗对胶质瘤U251中CD133~+细胞的影响

目的探讨化疗耐药与肿瘤干细胞之间的关系。方法分析替莫唑胺(TMZ)作用前后胶质瘤U251中CD133蛋白表达及阳性细胞数量的变化。通过MTT实验评估U251细胞对TMZ的敏感性。利用免疫荧光染色和Western blot检测TMZ作用前后U251细胞中CD133、巢蛋白(nestin)、神经元特异性烯醇化酶(NSE)、神经胶质细胞纤维酸性蛋白(GFAP)等蛋白表达。结果U251细胞中未加入TMZ组的CD133+细胞比例约为0.060±0.003,明显低于加入TMZ组的0.337±0.012(P<0.05)。在TMZ作用后,CD133和nestin表达明显增加,而GFAP和NSE表达明显减少(P<0.05)。结论胶质瘤U251细胞株中存在CD133+细胞,大部分具有脑肿瘤干细胞特性;在TMZ作用后,这类细胞比例增加,提示其与化疗耐药有关。     

胶质瘤中脑肿瘤干细胞的分离、培养和鉴定

目的 分离、培养和鉴定胶质瘤中脑肿瘤干细胞(BTSC),探讨CD133作为BTSC特异性标记,用于BTSC分离的可行性.方法 9例胶质瘤临床标本,胶质瘤C6和U87细胞系,用直接培养和磁珠选择CD133+ 细胞二种方法分离BTSC.参照NSC培养条件,进行体外培养.观察细胞生长情况,利用免疫荧光染色检测细胞标记,包括Ncstin,神经元特异性烯醇化酶(NSE),胶质纤维酸性蛋白(GFAP)等;诱导细胞分化,验证该种细胞是否具有干细胞特性.结果 9例临床标本要中,有2例儿童恶性胶质瘤在培养过程中出现神经球,在成人胶质瘤中均未出现.C6和U87细胞中,均出现比较典型的神经球,二种分离方法结果相同.神经球细胞具有增殖能力,体外培养可以不断增大.标记检测显示这种细胞Nestin阳性,而NSE和GFAP阴性.经诱导后,可以分化为肜态与来源细胞相似的细胞,能够表达NSE和GFAP等多种表型细胞标记.结论 在胶质瘤中,存在具有NSC特性的干细胞,即BTSC,CD133是BTSC重要的表面标记,可以用于BTSC的分离.在胶质瘤细胞系中CD1331 细胞是BTSC.     

CD133对CD133^+/Sox2^+脑肿瘤干细胞增殖、侵袭及药物敏感性的影响

目的分析靶向下调CD133表达对CD133^+/Sox2^+脑肿瘤干细胞增殖、侵袭及药物敏感性的影响。方法用肿瘤微球法从脑胶质瘤细胞系U251中筛选出CD133和Sox2均为阳性的脑肿瘤干细胞,后设计并合成针对CD133的特异性shRNA片段,构建GV115-shCD133RNA重组质粒,将其转染CD133^+/Sox2^+U251细胞,设为实验组(GV115-shCD133RNA-CD133^+/Sox2^+U251细胞),并设立空白组(CD133^+/Sox2^+U251细胞)和对照组(GV115-shRNA-CD133^+/Sox2^+U251细胞)。用Celigo细胞计数实验和Transwell实验分别检测细胞增殖能力和侵袭能力。向3组细胞分别加入8个不同浓度的顺铂和替莫唑胺(200%,150%,100%,75%,50%,25%,12.5%和5%),其中100%顺铂和和替莫唑胺的质量浓度分别为4和18μg·mL^-1),用Celigo细胞计数实验检测细胞增殖能力的变化,以评估其对药物敏感性的影响。结果实验组、对照组和空白组的CD133 mRNA分别为0.24±0.01,1.00±0.03和1.08±0.04,侵袭细胞个数分别为(18.93±0.64),(66.82±2.89)和(69.43±3.22)个,实验组的上述指标与空白组和对照组比较,差异均有统计学意义(均P<0.01)。从第1～5天,实验组的细胞增殖能力均明显低于对照组和空白组,差异均有统计学意义(均P<0.001)。实验组、对照组和空白组的顺铂半抑制浓度(IC₅₀)分别为(1.06±0.11),(4.03±0.21)和(3.94±0.25)μg·mL^-1,替莫唑胺IC₅₀分别为(4.23±0.38),(18.06±2.13)和(17.48±2.05)μg·mL^-1,实验组对顺铂和替莫唑胺的药物敏感性与对照组和空白组比较,差异均有统计学意义(均P<0.01)。结论靶向下调CD133表达能有效抑制脑胶质瘤干细胞的增殖及侵袭能力,提高其对化疗药物的敏感性。     

从PC-3细胞中富集前列腺癌干细胞的实验研究

目的从PC-3人前列腺癌细胞中分离并富集前列腺癌干细胞。方法体外无血清悬浮培养PC-3细胞,通过传代更新、诱导分化验证PC-3悬浮成球细胞具有强增殖性、自我更新及分化潜能,实时荧光定量核酸扩增检测系统检测CD44和CD133mRNA的表达,流式细胞术检测CD44+CD133+细胞和侧群(SP)细胞比例。结果 PC-3成球细胞可以在无血清培养液(SFM)中生存并形成可以稳定传代的悬浮细胞球。PC-3成球细胞具有自我更新和分化能力,其前列腺癌干细胞标志物CD44和CD133mRNA的相对表达量分别是PC-3贴壁细胞的41.83倍和10.48倍(P<0.05)。PC-3悬浮成球细胞中CD44+CD133+细胞比率为13.94%,SP细胞含量为3.10%,均显著高于PC-3贴壁细胞的0.77%和0(P<0.05)。结论通过无血清悬浮聚球培养可以从PC-3细胞中简便、高效地富集前列腺癌干细胞。     

Octamer 4基因在非小细胞肺癌中的表达及临床意义

目的检测Octamer 4(Oct-4)基因在非小细胞肺癌(NSCLC)中的表达,探讨Oct-4在NSCLC发生、发展中的作用。方法采用流式细胞仪分选出肺癌细胞系A549中Oct-4+及Oct-4-的细胞群,用免疫荧光法检测分选后的细胞亚群中CD133的表达情况;采用体外侵袭实验检测Oct-4+细胞群和Oct-4-细胞群的侵袭能力。结果 1以标记Oct-4通过流式细胞仪分离后,成功获得肺癌细胞株A549中Oct-4+细胞群和Oct-4-细胞群。且Oct-4+细胞群中CD133的表达明显高于Oct-4-细胞群,差异有统计学意义(P<0.05)。2Oct-4+细胞群的侵袭细胞数显著高于Oct-4-细胞群,差异有统计学意义(P<0.05)。3Oct-4在NSCLC中的表达率为83%(50/60),且Oct-4+细胞群中CD133的表达明显高于Oct-4-细胞群,两者比较差异有统计学意义(P<0.05);且Oct-4的表达与NSCLC组织的分化程度、肿瘤的临床分期及是否已经发生淋巴结转移有关,差异有统计学意义(P<0.05)。结论干细胞标志物Oct-4在NSCLC干细胞亚群中呈高表达,可能与肺癌的发生、发展及转移密切相关,为肺癌治疗提供新的靶点。     

CD4^＋CD25^＋T细胞亚群在系统性红斑狼疮患者中的检测和意义

目的:探讨CD4+CD25+T细胞亚群在系统性红斑狼疮(SLE)发病中的作用.方法:采用流式细胞仪检测35例SLE患者(其中活动组20例)及18例健康对照组的外周血CD4+CD25+T细胞亚群的百分比及其CD4+CD25highT细胞与CD4+CD25intT细胞的比率,同时评估疾病活动指数(SLEDAI),分析两者的相关性.结果:活动组和非活动组SLE患者外周血总CD4+GD25+T细胞、CD4+CD25highT细胞和CD4+CD25intT细胞占CD4+T细胞的百分比与对照组相比,差异均无统计学意义(P>0.05),且与SLEDAI无相关性.活动组SLE患者外周血CD4+CD25highT细胞与CD4+CD25intT细胞的比率与非活动组及健康对照组相比较明显降低,差异有统计学意义(P<0.05),且与SLEDAI呈明显负相关.结论:活动性SLE患者外周血CD4+CD25highT细胞与CD4+CD25intT细胞的比率降低可能在SLE的发病中起到重要的作用.     

Targeting CD133 antigen in cancer

Background: Much attention has been focused on CD133 as a marker of cancer cells with stem-cell-like ability. In the cancer stem cells (CSCs) model, only a small proportion of tumour cells are able to self-renew extensively, while the bulk of cells proceed to differentiate into committed heterogeneous clones. On the basis of the involvement of CSCs in tumourigenesis and treatment resistance, it is conceivable that only eradication of CSCs can lead to a cancer cure. Objective: To highlight the most recent evidence about the role of CD133 as a marker of CSCs in human tumours, and the therapeutic perspectives associated with its specific targeting. Methods: A literature search through Medline to locate published full articles using the following key words for selection: ‘CD133 and cancer targeting’, ‘CD133 and chemo resistance’, and ‘CD133 and molecular pathways’. Only studies in English are considered. Results/Conclusions: The role of CD133 as a marker of CSCs has been documented in several human neoplasms; its expression seems to predict unfavourable prognosis. Novel therapeutic strategies aimed at targeting molecular pathways critical for CD133⁺ CSCs survival are being examined.     

CD133 as a target for colon cancer

Introduction: Recent evidence based on cancer stem cell (CSC) models, is boosting the progress of translational research and providing relevant clinical implications in many tumour types, including colorectal cancer. The current failure of standard therapies is attributed to a small fraction of the primary cell population with stem-like characteristics, such as self-renewal and differentiation. Identification of CSCs is based on two different criteria of selection: stemness-selective conditions and direct isolation based on putative stem cell markers expression. CD133, a transmembrane glycoprotein, was associated with tumor-initiating cells derived from several histological variants of tumors, including colon.

Areas covered: In this review the current understandings about CD133 as putative marker of tumour-initiating cells in colorectal cancer (CRC) is described. The focus of the discussion is on the need for additional markers to better identify the cell population able to recapitulate the parental tumor in immunocompromised mice.

Expert opinion: Identification and characterization of CSCs represents a relevant issue to define innovative therapeutic approaches, overcoming the emergence of cancer cell clones capable of evading standard therapy.     

CD133 as a target for colon cancer

INTRODUCTION: Recent evidence based on cancer stem cell (CSC) models, is boosting the progress of translational research and providing relevant clinical implications in many tumour types, including colorectal cancer. The current failure of standard therapies is attributed to a small fraction of the primary cell population with stem-like characteristics, such as self-renewal and differentiation. Identification of CSCs is based on two different criteria of selection: stemness-selective conditions and direct isolation based on putative stem cell markers expression. CD133, a transmembrane glycoprotein, was associated with tumor-initiating cells derived from several histological variants of tumors, including colon. AREAS COVERED: In this review the current understandings about CD133 as putative marker of tumour-initiating cells in colorectal cancer (CRC) is described. The focus of the discussion is on the need for additional markers to better identify the cell population able to recapitulate the parental tumor in immunocompromised mice. EXPERT OPINION: Identification and characterization of CSCs represents a relevant issue to define innovative therapeutic approaches, overcoming the emergence of cancer cell clones capable of evading standard therapy.     

CD133: beyond a cancer stem cell biomarker

Abstract

CD133 (prominin-1), a pentaspan membrane glycoprotein, is one of the most well-characterized biomarkers used for the isolation of cancer stem cells (CSCs). The presence of CSCs is one of the main causes of tumour reversal and resilience. Accumulating evidence has shown that CD133 might be responsible for CSCs tumourigenesis, metastasis and chemoresistance. It is now understood that CD133 interacts with the Wnt/β-catenin and PI3K-Akt signalling pathways. Moreover, CD133 can upregulate the expression of the FLICE-like inhibitory protein (FLIP) in CD133-positive cells, inhibiting apoptosis. In addition, CD133 can increase angiogenesis by activating the Wnt signalling pathway and increasing the expression of vascular endothelial growth factor-A (VEGF-A) and interleukin-8. Therefore, CD133 could be considered to be an ‘Achilles’ heel’ for CSCs, because by inhibiting this protein, the signalling pathways that are involved in cell proliferation will also be inhibited. By understanding the molecular biology of CD133, we can not only isolate stem cells but can also utilise it as a therapeutic strategy. In this review, we summarise new insights into the fundamental cell biology of CD133 and discuss the involvement of CD133 in metastasis, metabolism, tumourigenesis, drug-resistance, apoptosis and autophagy.     

肿瘤干细胞标志物CD133和CD44在肾母细胞瘤中的表达及意义

目的 探讨肿瘤干细胞标志物CD133和CD44在肾母细胞瘤中的表达及其意义。方法 收集2010年1月至2017年10月在安徽省立医院未行放化疗而直接进行手术治疗的肾母细胞瘤患者手术切除组织标本,病理诊断同步进行免疫组织化学方法检测组织中CD133和CD44阳性信号强度和阳性细胞数目,计算免疫组织化学评分。同时收集临床病理参数,分析免疫组织化学评分与临床病理参数之间是否存在相关性。结果 共收集肾母细胞瘤患儿64例,其中男性30例,女性34例。年龄中位数为36个月,其中46例年龄≤36个月,18例年龄>36个月。肿瘤大小（最大直径）2～19 cm,中位数为6.5 cm,其中32例≤6.5 cm,另外32例>6.5 cm。58例表现为预后好的病理特征,6例表现为预后差的病理特征。根据NWTS的分期,24例为1期,26例为2期,4例为3期,8例为4期,2例为5期。免疫组织化学评分结果显示,44%（28/64）的标本为CD133阳性,13%（8/64）的标本为CD44阳性。CD133的免疫组织化学评分与肾母细胞瘤NWTS分期有正相关性（r=0.813,P<0.05）;在女童中CD133评分高于男童（P<0.05）;CD133评分与患者的年龄、肿瘤的大小及病理特征无相关性（P>0.05）。结论 CD133在肾母细胞瘤中高表达,其表达水平与肾母细胞瘤的发生及恶性程度存在关系。     

CD133在肝细胞性肝癌中的表达及其临床意义

目的 探讨肿瘤干细胞标志物CD133在肝细胞性肝癌(HCC)组织中的表达及其临床意义.方法 采用逆转录聚合酶链反应(RT-PCR)检测25例手术切除的HCC原发组织、癌旁组织中CD133 mRNA的表达,分析CD133的表达与临床参数的关系.结果 HCC原发组织和癌旁组织中CD133 mRNA表达的阴性率分别为72.0%和4.0%(P<0.01);原发组织CD133 mRNA的表达与肿瘤大小、甲胎蛋白(AFP)阳性表达有关(P<0.01),与性别、年龄及组织分化程度均无关.结论 CD133的表达与HCC的大小及AFP阳性表达有关,可能在HCC的诊断及判断复发中有一定价值.     

乳腺癌组织中CD133和VEGF的表达及意义

目的 研究CD133及血管内皮生长因子(VEGF)在乳腺癌组织中的表达及其意义.方法 采用免疫组织化学方法检测109例乳腺癌组织中的CD133及VEGF的表达,并对结果进行分析.结果 乳腺癌组织中CD133和VEGF蛋白的表达率分别是46.8%(51/109)和84.4%(92/109),其阳性表达率随肿瘤病理级别的升高而明显增加(P＜0.05).乳腺癌Basal-like亚群中CD133蛋白的阳性率表达高达69.2%,高于其他亚群(P＜0.05).CD133与VEGF表达密切相关(P＜0.01).结论 CD133与VEGF蛋白表达可能与乳腺癌肿瘤干细胞生长有关.