CD10-positive stromal cells in gastric carcinoma: correlation with invasion and metastasis

BACKGROUND: CD10 is a cell surface metalloproteinase expressed by a variety of normal cell types, including lymphoid precursor cells, germinal center B lymphocytes and some epithelial cells. Although accumulating data indicate that CD10 expression by stromal cells is involved in colorectal carcinogenesis and it is a novel prognostic factor in breast carcinoma, CD10-positive stromal cells and their correlation with invasion and metastasis have not been studied in gastric carcinoma. The aim of this study was to immunohistochemically investigate the rate of CD10 production in the stromal cells in our gastric carcinoma collection and clarify its correlation with invasion and metastasis. METHODS: One hundred and sixteen cases of gastric carcinoma were analyzed immunohistochemically using a monoclonal CD10 antibody (clone 56C6). RESULTS: The expression of CD10 by stromal cells was significantly higher in the primary gastric carcinomas than in normal and dysplasia mucosas (P = 0.014). More frequent expression of CD10 by stromal cells was detected in differentiated carcinoma than in undifferentiated carcinoma (P < 0.001). CD10 expression by the stromal cells was associated with depth of invasion and lymph node metastasis (P < 0.05). Stromal CD10 expression was lower in gastric carcinoma without vessel invasion than in those with vessel invasion (P = 0.001). However, no association was observed between stromal CD10 expression and TNM stage. In differentiated carcinoma, stromal CD10 expression was associated with the depth of invasion, lymph node metastasis, vessel invasion and TNM stage (P < 0.01). CONCLUSION: These results indicate that stromal cells expressing CD10 may play an important role in gastric carcinogenesis. CD10 expression by stromal cells seems to promote invasion and metastasis of differentiated gastric carcinoma.     

Cisplatin selects for CD133+ cells in lung cancer cells

Objective Platinum-based chemotherapy is the first-line treatment for non-small cell lung cancer,but the chemoresistance of tumor cells continues to be a considerable challenge in the management of NSCLCs,leading to recurrence of most patients.CD133(prominin-1)is a five-transmembrane glycoprotein,and recent evidence suggests that CD133+cells are the cause of drug resistance and tumor recurrence.In this study,the correlation between cisplatin and CD133+cells was investigated systematically.Methods Four lung cancer cell lines,including A549,H460,801D and H1299,were treated with different concentrations of cisplatin.Cell viability was determined by MTT assay.Sphere-forming assay was performed to detect the capability of sphere-forming.CD133+cells was detected by BD FACScaliber flow cytometer.Results The results showed that cisplatin could increase the number of CD133+cells in both time-and dose-dependent manner.The enrichment would weaken but the proportion of CD133+cells was still higher than the basic level as incubation time extended after cisplatin was withdrawn.Compared with adherent culture,the proportion of CD133+cells was higher when the cells were maintained suspension culture.The proportion of CD133+cells significantly increased when cisplatin was provided in suspension culture.Conclusion These results revealed that cisplatin induces the enrichment of CD133+cells and CD133 is a new therapeutic target.Our data partially explained drug resistance to second-line chemotherapy in cisplatin-treated patients with NSCLCs.     

Characterization of the conversion between CD133+ and CD133- cells in colon cancer SW620 cell line

The state of cancer stem cells (CSC) under reversible fluctuations, which has been revealed in breast cancer cells most recently, suggests that subpopulations with distinct phenotypes and functions within cancer cells can undergo inter-conversion. To investigate the possibility in colon cancer cells, we employed CD133 as the CSC marker, and characterized CD133 expression pattern and the biological features of the CD133⁺ and CD133^- subsets. Flow cytometry revealed that CD133 was bimodally expressed in SW620 cells among eight colon cancer cell lines. The CD133⁺ clonal SW620 cells displayed a differential gene expression profile, higher cellular reactive oxygen species (ROS), enhanced tumorigenesis and resistance to 5-fluorouracil. The conversion in term of the CD133 phenotype of the sorted cells was observed in vitro and in vivo. The fraction of the CD133⁺ cells decreased from 99% to 80% in the sorted CD133⁺ population while rising from 5 to 10% in the sorted CD133^- population during the first 20-day cultivation and then stayed almost unchanged. A fraction (about 20%) of the CD133⁺ clonal cells lost their CD133 marker while about 10% of the CD133^- clonal cells acquired the CD133 marker. 5-Azacytidine enhanced the fraction of the CD133⁺ cells in both of the CD133⁺ and CD133^- clonal cells. Our data demonstrate that CD133 expression is dynamic and reversible, and reveal the inter-conversion between the CD133⁺ and the CD133^- SW620 cells, suggesting that the CD133 phenotype of SW620 cell population is retained by the conversion between the two cell subsets.     

肿瘤干细胞标志物CD133、CD44、OCT-4在小细胞肺癌中的表达及其临床意义

目的：探讨肿瘤干细胞标志物CD133、CD44、OCT-4与小细胞肺癌临床病理特征之间的相关性及临床意义。方法应用免疫荧光技术检测小细胞肺癌细胞株NCI-H82中肿瘤干细胞标志物CD133、CD44、OCT-4的表达;同时应用免疫组织化学法检测79例小细胞肺癌组织中CD133、CD44、OCT-4的表达。结果在小细胞肺癌细胞株NCI-H82中,CD133和CD 44的荧光信号为阳性表达,OCT-4的荧光信号为阴性表达。小细胞肺癌组织中CD133和CD44表达与肿瘤直径、淋巴结转移和临床分期相关,差异具有统计学意义（P＜0.05）。小细胞肺癌组织中OCT-4为阴性表达。结论 CD133和CD44可能是小细胞肺癌肿瘤干细胞的标志物,对小细胞肺癌的诊断和治疗有一定的临床意义。     

Lenti-TK介导间充质干细胞对鼻咽癌CD133+干细胞的靶向迁移及杀伤作用

目的： 观察慢病毒-胸苷激酶（lentivirus-thymidine kinase,Lenti-TK)/间充质干细胞(mesenchymal stem cell,MSC）对鼻咽癌CD133+干细胞的靶向迁移及杀伤作用。 方法： 构建包含TK基因的重组慢病毒表达载体Lenti-TK,感染MSC后得到Lenti-TK-MSC,RT-PCR及Western blotting检测Lenti-TK-MSC中HA-TK的表达。免疫磁珠法从鼻咽癌5-8F细胞中分选CD133+细胞;Transwell小室迁移实验检测Lenti-TK-MSC对CD133+5-8F细胞的趋向性;Lenti-TK-MSC联合更昔洛韦（ganciclovir,Lenti-TK-MSC/GCV）与CD133+5-8F细胞共培养,CCK-8试剂盒检测其对细胞的杀伤作用和旁观者效应。 结果： 成功构建重组慢病毒载体Lenti-TK,其滴度为1×108UT/ml,Lenti-TK（MOI=50）感染MSC 72 h时,感染效率达（95.1±01）%。Lenti-TK-MSC迁移至CD133+5-8F细胞组的细胞数明显多于CD133-5-8F细胞组、未分选5-8F细胞组\[（83.0±8.7） vs （29.6±53）、（38.3±5.2）,P=0.000\]。Lenti-TK-MSC/GCV处理组与单独GCV处理组、Lenti-TK-MSC/GCV条件培养液（即Lenti-TK-MSC加入1 mg/L GCV培养48 h的培养上清）处理组相比,CD133+5-8F细胞的存活率明显降低\[（37.2±2.3）% vs （98.5±3.1）%、（83.8±34）%,P=0.000\]。Lenti-TK-MSC数量达到混合细胞总数（Lenti-TK-MSC和CD133+5-8F细胞)的20%时,CD133+5-8F细胞存活率为（68.2±2.3）%,表现出明显的旁观者杀伤效应。 结论： Lenti-TK感染后MSC对鼻咽癌CD133+5-8F细胞具有靶向迁移及杀伤作用。     

CD133与肿瘤干细胞研究进展

越来越多的证据表明肿瘤中存在肿瘤干细胞(cancer stem cells),并且其与肿瘤的增殖、转移、复发和对放化疗不敏感关系密切.因此,肿瘤治疗应当针对肿瘤干细胞,通过特异表面标记分选肿瘤干细胞是研究其生长特点的前提.近年来,CD133为研究最多的在于细胞(stem cell)和多种组织肿瘤干细胞表面独立表达的特异标记分子.通过CD133可以分选干细胞、前体细胞和肿瘤干细胞.众多研究表明,CD133~+肿瘤细胞与肿瘤的自我更新、分化潜能、信号传导调控、药物耐受、复发和预后等均有相关性.CD133~+细胞有望在干细胞相关疾病的治疗和肿瘤靶向治疗中发挥巨大作用.     

下调CD133表达对肝癌干细胞上皮间质化和侵袭能力影响及机制探讨

目的 肿瘤干细胞是肿瘤复发和转移的原因.探讨CD133基因RNA干扰(RNA interference,RNAi)的重组质粒,对人肝癌细胞株MHCC-H的CD133基因表达、上皮间质化和侵袭能力的影响.方法 通过免疫磁珠分选MHCC-H细胞中的CD133阳性细胞,设计并合成特异性靶向CD133的小分子干扰RNA(small interfering RNA,siRNA)片段,并构建pSuper retro GFP-neo-siRNA-CD133表达质粒,将其转入MHCC-H-CD133+细胞,并设空白对照组、阳性对照组.通过G418筛选出稳定株.通过粘附实验、Boyden小室实验和软琼脂克隆形成实验观察各细胞株侵袭能力的变化.采用明胶酶法测定肝癌细胞的基质金属蛋白酶-2(matrix metalloproteinase-2,MMP 2)和MMP-9的含量,蛋白质印迹法检测上皮间质化相关基因的变化.结果 通过免疫磁珠分选后的MHCC-H细胞中CD133表达率为83.5％.qRT-PCR结果显示,shCD133组相对空白对照组的CD133表达量下降了80％,P＜0.05.蛋白质印迹法检测结果显示,shCD133组的CD133表达明显下降,约为空白对照组的12％和shNC组的10％,均P＜0.05.shCD133组的粘附能力为86.9±12.4,较空白对照组的568.5±53.2和shNC组的538.8±35.6明显下降,P＜0.05.Boyden小室实验发现,shCD133组穿膜细胞数为(80.6±11.4)个,较空白对照组的(228.5±33.2)个和shNC组的(230.8±32.9)个明显减少,P＜0.05.软琼脂克隆形成率的检测发现,shCD133的细胞克隆形成(60±5)个,比空白对照组的(178±23)个和shNC组的(168±25)个明显下降,P＜0.05.明胶酶法检测发现,shCD133组MMP-2和MMP-9相对活性为0.4±0.14和0.6±0.16,较shNC组的1.03±0.19和1.3±0.16明显下降,P＜0.05.蛋白质印迹法检测结果表明,shCD133组N-cadherin蛋白表达为36.3±4.5,Snail为53.6±6.7,Slug为41.63±5.6,Twist为39.4±3.9,均明显低于空白对照组的87.6±8.6、80.6±7.5、81.9±9.2和83.9±9.1,也明显低于shNC组的89.4±9.6、83.5±8.9、85.1±8.7和87.6±9.3,均P＜0.05;而shCD133组E-cadherin表达为88.4±9.2,较空白对照组的57.6±8.7和shNC组的53.9±8.9明显上调,P＜0.05.结论 沉默MHCC-H细胞CD133基因的表达能有效抑制其上皮间质化和侵袭能力,CD133基因可能成为肝癌基因治疗的有效靶点.     

FGF10/FGFR2 signal induces cell migration and invasion in pancreatic cancer

Pancreatic cancer has one of the highest mortalities among all malignancies and there is an urgent need for new therapy. This might be achieved by resolving the detailed biological mechanism, and in this study we examined how pancreatic cancer cells develop aggressive properties by focusing on signalling through the fibroblast growth factor (FGF)10 and FGF receptor (FGFR)2, which play important roles in pancreatic organogenesis. Immunostaining of pancreatic cancer tissues showed that FGFR2 was expressed in cancer cells, whereas FGF10 was expressed in stromal cells surrounding the cancer cells. Patients with high FGFR2 expression in cancer cells had a shorter survival time compared to those with low FGFR2 expression. Fibroblast growth factor 10 induced cell migration and invasion of CFPAC-1 and AsPC-1 pancreatic cancer cells through interaction with FGFR2-IIIb, a specific isoform of FGFR2. Fibroblast growth factor 10 also induced expression of mRNA for membrane type 1-matrix metalloproteinase (MT1-MMP) and transforming growth factor (TGF)-beta1, and increased secretion of TGF-beta1 protein from these cell lines. These data indicate that stromal FGF10 induces migration and invasion in pancreatic cancer cells through interaction with FGFR2, resulting in a poor prognosis. This suggests that FGF10/FGFR2 signalling is a promising target for new molecular therapy against pancreatic cancer.     

Cancer stem/progenitor cells are highly enriched in CD133+CD44+ population in hepatocellular carcinoma

Both our previous study and other reports have suggested that CD133, originally classified as a hematopoietic stem cell marker, could be used for enrichment of cancer stem cells (CSCs) in human hepatocellular carcinoma (HCC). It was also noted that not all of CD133⁺ cells were representative of CSCs. Further identification and characterization of CSCs or tumor‐initiating cells in HCC are necessary to better understand hepatocarcinogenesis. In present study, we demonstrated that CSC phenotype could be precisely defined by co‐expression of CD133 and CD44 cell surface markers. CD133⁺CD44⁺ HCC cells showed stem cell properties, including extensive proliferation, self‐renewal, and differentiation into the bulk of cancer cells. In vivo xenograft experiments revealed that, actually, the highly tumorigenic capacity of CD133⁺ cells as previously described was primarily attributed to CD133⁺CD44⁺ cell subpopulation, instead of their CD133⁺CD44^? counterparts. Moreover, cells double‐positive for CD133 and CD44 exhibited preferential expression of some stem cell‐associated genes and were more resistant to chemotherapeutic agents due to the upregulation of ATP‐binding cassette (ABC) superfamily transporters, including ABCB1, ABCC1, and ABCG2, further supporting these cells as HCC cell origin. Our findings suggest that CD133⁺CD44⁺ cells might represent true cancer stem/progenitor cells in HCC, which could allow a better understanding of HCC initiation and progression, as well as establish a precise target for the development of more effective therapies.     

CD133-positive tumor cell content is a predictor of early recurrence in colorectal cancer

Background

The aims of this study were to demonstrate the tumorigenicity of CD133+ colon cancer cells in vitro, analyze the correlations between spheroid formation and clinicopathologic variables, and screen for overexpressed genes in CD133+ colon cancer stem cells. Moreover, the aim of this study was to establish a living tumor tissue bank using surgically resected specimens.

Methods

Using LoVo cell line, we isolated CD133+ cells and performed clonogenic assay and animal experiment to test tumorigenicity of CD133+ cells. Twenty-nine surgical samples were freshly collected from 27 patients who received curative or palliative surgery, and the samples were mechanically and enzymatically dissociated into single cells.

Results

We confirmed the enhanced tumorigenicity of CD133+ cells isolated from LoVo cell line both in vitro and in vivo. Of these 29 samples, 8 (28%) contained >3% CD133+ cells. Sphere formation was significantly higher in samples from patients with lymphatic invasion than in those without lymphatic invasion [54.5% (6/11) vs. 12.5% (2/16); P=0.033] and in samples containing >3% of CD133+ cells than in those containing ≤3% of CD133+ cells [36.4% (4/11) vs. 0% (0/16); P=0.019].

Conclusions

These findings indicate that CD133 is a valid marker for identifying cancer stem cells from fresh surgically resected colorectal cancer tissues. Furthermore, we successfully established a living tumor tissue bank using surgically resected colorectal tissues with a viability of >70%.     

Clinical Significance of Cancer Stem Cell Markers CD133 and CXCR4 in Osteosarcomas

Objective: Osteosarcomas (OS) is one the most common primary bone malignancy in humans with the lungs metastasis in most cases. Metastasis and recurrence of OS is attributed to cancer stem cells (CSCs). Our study aimed to evaluate the clinical significance of CD133 and C-X-C chemokine receptor type 4 (CXCR4) as the frequently applied markers for CSCs in OS patients. Methods: In this cross-sectional, a total of 50 tissue samples from the patients with primary OS were immunohistochemically examined to detect the expression of CD133 and CXCR4. The associations of the relative expression and clinical significance of each marker were also evaluated. Results: High level expression of CD133 was detected in 26% of OS patient tissues. Of the 12 patients who showed lung metastasis, 5 cases showed high expression of CD133 with marginal trend correlation (P=0.06). No significant correlation was observed between CD133 expression and clinicopathological factors. Only 36% of cases showed CXCR4 expression which was not significantly correlated with gender, age, tumor size, necrosis, stage and metastasis (P>0.05). Clinically, patients with concomitant CD133/CXCR4 expression had significant association with lung metastasis (P=0.05). Conclusion: Our findings showed that concomitant expression of CSC markers CD133/CXCR4 might had a synergistic effect on the OS poor prognosis. These markers could be considered as potential therapeutic candidates of OS targeted therapy.     

CD133 initiates tumors,induces epithelial-mesenchymal transition and increases metastasis in pancreatic cancer

CD133 has been implicated as a cancer stem cell (CSC) surface marker in several malignancies including pancreatic cancer. However, the functional role of this surface glycoprotein in the cancer stem cell remains elusive. In this study, we determined that CD133 overexpression induced “stemness” properties in MIA-PaCa2 cells along with increased tumorigenicity, tumor progression, and metastasis in vivo. Additionally, CD133 expression induced epithelial-mesenchymal transition (EMT) and increased in vitro invasion. EMT induction and increased invasiveness were mediated by NF-κB activation, as inhibition of NF-κB mitigated these effects. This study showed that CD133 expression contributes to pancreatic cancer “stemness,” tumorigenicity, EMT induction, invasion, and metastasis.     

CD133 negative glioma cells form tumors in nude rats and give rise to CD133 positive cells

CD133 is a cell surface marker expressed on progenitors of haematopoietic and endothelial cell lineages. Moreover, several studies have identified CD133 as a marker of brain tumor-initiating cells. In this study, human glioblastoma multiforme biopsies were engrafted intracerebrally into nude rats. The resulting tumors were serially passaged in vivo, and monitored by magnetic resonance imaging. CD133 expression was analyzed at various passages. Tumors initiated directly from the biopsies expressed little or no CD133, and showed no contrast enhancement suggesting an intact blood-brain barrier. During passaging, the tumors gradually displayed more contrast enhancement, increased angiogenesis and a shorter survival. Real-time qPCR and immunoblots showed that this was accompanied by increased CD133 expression. Primary biopsy spheroids and xenograft tumors were subsequently dissociated and flow sorted into CD133 negative and CD133 positive cell populations. Both populations incorporated BrdU in cell culture, and expressed the neural precursor marker nestin. Notably, CD133 negative cells derived from 6 different patients were tumorgenic when implanted into the rat brains. For 3 of these patients, analysis showed that the resulting tumors contained CD133 positive cells. In conclusion, we show that CD133 negative glioma cells are tumorgenic in nude rats, and that CD133 positive cells can be obtained from these tumors. Upon passaging of the tumors in vivo, CD133 expression is upregulated, coinciding with the onset of angiogenesis and a shorter survival. Thus, our findings do not suggest that CD133 expression is required for brain tumor initiation, but that it may be involved during brain tumor progression.     

miR-34a、CD133在胰腺癌中的表达及临床意义

目的:检测胰腺癌肿瘤组织中微小RNA-34a与CD133表达情况,并探究其临床意义。方法:选取2012年12月至2014年2月本院收治的胰腺癌患者74例,取胰腺癌肿瘤组织及癌旁组织。采用实时定量PCR（qRT-PCR）法检测胰腺癌肿瘤组织及癌旁组织中miR-34a表达水平,采用免疫组化法检测CD133表达水平。结果:胰腺癌肿瘤组织miR-34a表达水平低于癌旁组织（P＜0.05）,CD133 mRNA表达水平较癌旁组织显著升高（P＜0.05）;CD133在胰腺癌肿瘤组织中阳性表达率显著高于癌旁组织（P＜0.05）。miR-34a、CD133表达与胰腺癌肿瘤分化程度、肿瘤大小、淋巴结转移有关（P＜0.05）;miR-34a、CD133表达与胰腺癌患者年龄、性别无显著相关性（P＞0.05）。miR-34a低表达胰腺癌患者3年总生存率显著低于高表达患者（P＜0.05）,CD133高表达胰腺患者3年总生存率显著低于CD133低表达者（P＜0.05）。miR-34a、CD133表达、肿瘤大小和肿瘤分化程度是影响胰腺癌患者预后的独立危险因素。胰腺癌肿瘤组织miR-34a与CD133 mRNA相对表达水平显著负相关（P＜0.05）。结论:miR-34a在胰腺癌组织中低表达,CD133高表达,与患者肿瘤分化程度、肿瘤大小、临床分期有关,可能作为胰腺癌病情监测及术后预后的生物学指标。     

Interleukin-8 derived from local tissue-resident stromal cells promotes tumor cell invasion

The aim of this study is to evaluate the role of adipose tissue resident stromal cells on tumor cell invasion. Our data show that a subpopulation of adipose tissue derived stromal cells expressing Nestin, NG2, α‐smooth muscle actin and PDGFR‐α migrate toward the cancer cells. Microarray analysis revealed the upregulation of IL‐8 in the migrated cells. We demonstrated that stromal cell derived IL‐8 promote the invasion and the anchorage‐independent growth of cancer cells. We conclude that human breast cancer cells attract a subpopulation of stromal cells that secrete IL‐8 to promote tumor cell invasion in a paracrine fashion. © 2011 Wiley Periodicals, Inc.     

骨髓增生异常综合征中CD34+细胞CXCR4的表达及其与细胞迁移的相关性

  目的  探讨不同危险度骨髓增生异常综合征（myelodysplasticsymdromes,MDS）中骨髓CD34+细胞CXCR4的表达情况及其与细胞迁移率的相关性。   方法  收集40例骨髓增生异常综合征患者的骨髓标本,根据IPSS积分系统进行危险度分组。低危组20例:IPSS积分0~1.5分;高危组20例:IPSS积分≥1.5分;同时采集10例健康者的骨髓标本作为对照。分离纯化骨髓CD34+细胞,通过流式细胞术检测CXCR4膜蛋白的表达;研究SDF-1α趋化作用下CD34+细胞的迁移率及CD34+细胞对骨髓基质细胞的迁移率。   结果  高危组MDS患者CD34+细胞CXCR4的表达率明显高于低危组和正常对照组（P < 0.000 1）;低危组和正常对照组之间CXCR4的表达率无显著性差异（P>0.05）。高危组CD34+细胞对SDF-1α及骨髓基质细胞的迁移率显著高于低危组及正常组（均P < 0.000 1）,且其对骨髓基质细胞的迁移率与CXCR4的表达呈正相关（P=0.000 1）。   结论  高危组MDS患者CD34+细胞CXCR4的表达量及其对骨髓基质细胞的迁移率均明显高于低危组患者,且其迁移率随CXCR4表达量的增加而升高,不同风险组的MDS患者存在SDF-1及其受体CXCR4表达和功能上的差异,SDF-1及其受体CXCR4在MDS发病中具有重要作用。      

肝细胞性肝癌组织CD133+细胞肿瘤干细胞特性的研究

目的 越来越多的研究表明,肝癌细胞系中能检测到肿瘤干细胞(cancer stem cells,CSCs)的存在.本研究旨在探讨从肝细胞性肝癌(hepatocellular carcinoma,HCC)组织样本中分离获得的CD133+细胞是否具有CSCs特性.方法 将2014-02-01-2015-06-30广西医科大学附属肿瘤医院肝胆外科25例手术获得的新鲜HCC组织和对应癌旁组织,采用酶消化法分别消化成单个肝癌细胞和单个肝细胞,利用流式细胞术检测部分单个肝癌细胞和单个肝细胞CD133的表达率.用剩余的单个肝癌细胞进行原代培养,流式细胞术将培养获得的肝癌细胞分选为CD133+和CD133-细胞,通过平板克隆形成实验、肿瘤球形成实验和裸鼠移植瘤形成实验对比分析这两组细胞的CSCs特性.结果 25例HCC组织中CD133的表达率为3.8％～8.3％,平均值为(5.8±1.6)％,而癌旁组织CD133的表达率为0.1％～0.4％,平均值为(0.2±0.1)％,两者比较差异有统计学意义,t=17.12,P＜0.001.CD133+和CD133-细胞的平均克隆率分别为(25.2±0.8)％和(7.6±0.8)％,两者比较差异有统计学意义,t=81.95,P＜0.001.CD133+和CD133-细胞的平均成球率分别为(20.3±0.6)％和(12.5±1.4)％,两者比较差异有统计学意义,t=68.17,P＜0.001.CD133+细胞的裸鼠移植瘤形成能力明显高于CD133-细胞.结论 从HCC组织样本中分离获得的CD133+细胞具有明显的CSCs特性.     

MicroRNA-218 inhibits cell invasion and migration of pancreatic cancer via regulating ROBO1

miRNA-218 is a highlighted tumor suppressor and its underlying role in tumor progression is still unknown. Here, we restored the expression of miRNA-218 in pancreatic cancer to clarify the function and potent downstream pathway of miRNA-218. The expressions of both miRNA-218 and its potent target gene ROBO1 were revealed by RT-PCR and western blotting analysis. Transfection of miRNA-218 precursor mimics and luciferase assay were performed to elucidate the regulation mechanism between miRNA-218 and ROBO1. Cells, stably expressing miRNA-218 followed by forced expression of mutant ROBO1, were established through co-transfections of both lentivirus vector and plasmid vector. The cell migration and invasion abilities were evaluated by migration assay and invasion assay respectively. An increased expression of ROBO1 was revealed in cell BxPC-3-LN compared with cell BxPC-3. Elevated expression of miRNA-218 would suppress the expression of ROBO1 via complementary binding to a specific region within 3′UTR of ROBO1 mRNA (sites 971–978) in pancreatic cancer cells. Stably restoring the expression of miRNA-218 in pancreatic cancer significantly downregulated the expression of ROBO1 and effectively inhibited cell migration and invasion. Forced expression of mutant ROBO1 could reverse the repression effects of miRNA-218 on cell migration and invasion. Consequently, miRNA-218 acted as a tumor suppressor in pancreatic cancer by inhibiting cell invasion and migration. ROBO1 was a functional target of miRNA-218’s downstream pathway involving in cell invasion and migration of pancreatic cancer.     

RNA aptamers targeting cancer stem cell marker CD133

The monoclonal antibody against the AC133 epitope of CD133 has been widely used as a cell surface marker of cancer stem cells in several different cancer types. Here, we describe the isolation and characterisation of two RNA aptamers, including the smallest described 15 nucleotide RNA aptamer, which specifically recognise the AC133 epitope and the CD133 protein with high sensitivity. As well, both these aptamers show superior tumour penetration and retention when compared to the AC133 antibody in a 3-D tumour sphere model. These novel CD133 aptamers will aid future development of cancer stem cell targeted therapeutics and molecular imaging.     

抗胰腺癌干细胞单克隆抗体的筛选和鉴定

目的：从抗胰腺癌干细胞单抗库中筛选、鉴定识别胰腺癌干细胞的功能性单抗,为胰腺癌干细胞靶向治疗提供候选抗体药物。方法：无血清悬浮培养及PKH26染色确定胰腺癌HPAC细胞系中肿瘤干细胞的存在。流式细胞术检测HPAC的干细胞标志物CD133在球体细胞中的阳性比例,检测20株杂交瘤单抗在HPAC亲本和球体细胞中的阳性表达。双色荧光标记流式细胞术检测CD133和单抗在HPAC亲本和球体细胞中的共表达比例;无血清悬浮培养法观察单抗15E9对HPAC成球细胞自我更新的影响。CCK一8法检测单抗15E9对HPAC细胞增殖和耐药的影响。结果：HPAC细胞能在无血清培养基中存活、增殖并形成细胞球,成球率为4．8％±0．6％。HPAC球体细胞中CD133’细胞的比例较亲本细胞提高至11．6倍。20株候选杂交瘤单抗中有3株单抗能识别HPAC球体细胞中CD133’细胞,其中单抗15E9共染比例为3．5％,并能显著抑制HPAC细胞的成球,抑制率达到30．4％。单抗15E9联合吉西他滨能显著抑制HPAC球体细胞的增殖,联合组和对照组Ic50分别为30．8nmol／L和58．1nmol／L。结论：本研究成功筛选出1株杂交瘤单抗可以识别胰腺癌干细胞,并且可识别CD133＋的胰腺癌干细胞;体外功能显示该抗体具有抑制HPAC干细胞的自我更新能力,抗体干预后显著降低HPAC耐药能力,可能是潜在的胰腺癌干细胞的靶向治疗抗体药物。