CD133+ liver cancer stem cells modulate radioresistance in human hepatocellular carcinoma

CD133 is a cancer stem-cell (CSC) marker associated with radioresistance and chemoresistance in various cancers. In the present study, CD133-expressing liver cancer cells following radiation exposure showed higher activation of MAPK/PI3K signaling pathway and reduction in reactive oxygen species levels compared to CD133⁻ cells. The in vivo study with a xenograft model showed increased tumor formation in irradiated CD133⁺ cell-injected nude mice compared to the CD133⁻ group, suggesting that CD133 contributes to radioresistance in HCC. Therefore, CD133-expressing liver cancer cells have anti-apoptotic and radioresistance properties that may be useful to improve anti-cancer treatments, including chemotherapy/radiotherapy of HCC.     

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.     

乙醛脱氢酶1作为喉癌Hep-2细胞系肿瘤干细胞标志物的实验研究

目的 探讨乙醛脱氢酶1(ALDH1)在人喉癌Hep-2细胞系中的表达,观察ALDH1高表达细胞的体外生长特性,确定喉癌Hep-2细胞系肿瘤干细胞标志物.方法 采用荧光细胞化学染色和流式细胞术观察和检测喉癌Hep-2细胞系中的ALDH1的表达.采用流式分选术分离ALDH1高表达细胞,以四甲基偶氮唑蓝(MTT)法观察ALDH1不同亚群细胞的体外增殖能力,以流式细胞术检测ALDH1高表达细胞的体外分化能力.采用裸鼠成瘤实验比较不同亚群细胞的成瘤能力.结果 喉癌Hep-2细胞系中ALDH1呈不同程度的表达,其中ALDH1高表达细胞占2.9％±0.6％.ALDH1高表达细胞的体外增殖能力高于ALDH1低表达和未分选细胞.在含血清的培养液中,经过6d培养,ALDH1高表达细胞由最初分选后的94.2％±3.8％下降至分选前水平.ALDH1高表达细胞的成瘤能力显著高于其他亚群.结论 喉癌Hep-2细胞系中,ALDH1高表达细胞具有很强的体外分化能力、增殖能力和成瘤能力,可以作为Hep-2细胞系肿瘤干细胞的标志物之一.     

Autophagy contributes to the survival of CD133+ liver cancer stem cells in the hypoxic and nutrient-deprived tumor microenvironment

Liver cancer stem cells (LCSCs) can drive and maintain hepatocellular carcinoma (HCC) growth, metastasis, and recurrence. Therefore, they are potentially responsible for the poor prognosis of HCC. Oxygen and nutrient deficiencies are common characteristics of the tumor microenvironment. However, how LCSCs adapt to oxygen- and nutrient-deprived conditions is unclear. Here, we used immunofluorescent staining and flow cytometry analysis to show that CD133+ cells were significantly enriched after hypoxia and nutrient starvation (H/S) in the human HCC cell line Huh7. Sorted CD133+ cells showed higher survival, less apoptosis, and possess higher clonogenic ability under H/S compared to the CD133− population. Under H/S, electron microscopy revealed more advanced autophagic vesicles in CD133+ cells. Additionally, CD133+ cells had higher autophagy levels as measured by both RT-qPCR and Western blotting. CD133+ cells had more accumulated GFP-LC3 puncta, which can be detected by fluorescence microscopy. The autophagic inhibitor chloroquine (CQ) significantly increased apoptosis and decreased the clonogenic capacity of CD133+ cells under H/S. Pre-culturing in H/S enhanced the sphere-forming capacity of CD133+ cells. However, CQ significantly impaired this process. Therefore, autophagy is essential for LCSCs maintenance. CD133+ cells were also found to have a higher tumor-forming ability in vivo, which could be inhibited by CQ administration. Collectively, our results indicate that the involvement of autophagy in maintenance of CD133+ LCSCs under the oxygen- and nutrient-deprived conditions that are typical of the tumor microenvironment in HCC. Therefore, autophagy inhibitors may make LCSCs more sensitive to the tumor microenvironment and be useful in improving anti-cancer treatments.     

CD133+ HCC cancer stem cells confer chemoresistance by preferential expression of the Akt/PKB survival pathway

The recent discovery of cancer stem cells (CSCs) has played a pivotal role in changing our view of carcinogenesis and chemotherapy. Based on this concept, CSCs are responsible for the formation and growth of neoplastic tissue and are naturally resistant to chemotherapy, explaining why traditional chemotherapies can initially shrink a tumor but fails to eradicate it in full, allowing eventual recurrence. Recently, we identified a CSC population in hepatocellular carcinoma (HCC) characterized by their CD133 phenotype. However, the molecular mechanism by which it escapes conventional therapies remains unknown. Here, we examined the sensitivity of these cells to chemotherapeutic agents (doxorubicin and fluorouracil) and the possible mechanistic pathway by which resistance may be regulated. Purified CD133+ HCC cells isolated from human HCC cell line and xenograft mouse models survived chemotherapy in increased proportions relative to most tumor cells which lack the CD133 phenotype; the underlying mechanism of which required the preferential expression of survival proteins involved in the Akt/PKB and Bcl-2 pathway. Treatment of CD133+ HCC cells with an AKT1 inhibitor, specific to the Akt/PKB pathway, significantly reduced the expression of the survival proteins that was normally expressed endogenously. In addition, treatment of unsorted HCC cells with both anticancer drugs in vitro significantly enriched the CD133+ subpopulation. In conclusion, our results show that CD133+ HCC cells contribute to chemoresistance through preferential activation of Akt/PKB and Bcl-2 cell survival response. Targeting of this specific survival signaling pathway in CD133+ HCC CSCs may provide a novel therapeutic model for the disease.     

CD133(+) and CD133(-) glioblastoma-derived cancer stem cells show differential growth characteristics and molecular profiles

Although glioblastomas show the same histologic phenotype, biological hallmarks such as growth and differentiation properties vary considerably between individual cases. To investigate whether different subtypes of glioblastomas might originate from different cells of origin, we cultured tumor cells from 22 glioblastomas under medium conditions favoring the growth of neural and cancer stem cells (CSC). Secondary glioblastoma (n = 7)-derived cells did not show any growth in the medium used, suggesting the absence of neural stem cell-like tumor cells. In contrast, 11/15 primary glioblastomas contained a significant CD133(+) subpopulation that displayed neurosphere-like, nonadherent growth and asymmetrical cell divisions yielding cells expressing markers characteristic for all three neural lineages. Four of 15 cell lines derived from primary glioblastomas grew adherently in vitro and were driven by CD133(-) tumor cells that fulfilled stem cell criteria. Both subtypes were similarly tumorigenic in nude mice in vivo. Clinically, CD133(-) glioblastomas were characterized by a lower proliferation index, whereas glial fibrillary acidic protein staining was similar. GeneArray analysis revealed 117 genes to be differentially expressed by these two subtypes. Together, our data provide first evidence that CD133(+) CSC maintain only a subset of primary glioblastomas. The remainder stems from previously unknown CD133(-) tumor cells with apparent stem cell-like properties but distinct molecular profiles and growth characteristics in vitro and in vivo.     

CD133作为肺癌干细胞标记物的应用及其局限性

肺癌是临床上最常见的恶性肿瘤之一,尚缺乏预后较为理想的治疗方案。肿瘤干细胞学说认为肺癌干细胞是肺癌发生、发展、转移、复发、耐受放化疗以及肺癌细胞具有侵袭性的主要原因。近年来越来越多的机构利用CD133糖基化表位来鉴定、分离、提纯肺癌干细胞。然而,随着研究的深入,CD133作为肺癌干细胞标记物逐渐受到质疑。本文对近年来利用CD133作为肺癌干细胞表面标记物的应用及其局限性做一综述。     

肿瘤干细胞标记物CD133在鼻咽癌组织中的表达及其临床意义

目的:研究干细胞标记物CD133在鼻咽癌及其声带息肉中的表达及其临床意义.方法:纤维鼻咽镜取30例鼻咽癌组织和10例声带息肉.Trizol法提取的总RNA进行逆转录,实时荧光定量PCR检测组织中CD133的表达.并对鼻咽癌病人予诱导时间调节化疗+同步放、化疗进行治疗.用SPSS 13.0医学统计学软件对结果和疗效进行分析.结果:鼻咽癌组织中CD133表达高于声带息肉组织(P＜0.05),CD133在鼻咽癌组织中的表达与年龄、性别、病理分型以及疗效之间均无统计学关联.结论:CD133在鼻咽癌组织中明显阳性表达,与年龄、性别、病理学类型及疗效之间的联系,还需进一步扩大病例数加以研究.     

Aldehyde dehydrogenase 1, a functional marker for identifying cancer stem cells in human nasopharyngeal carcinoma

Aldehyde dehydrogenase 1 (ALDH1) activity has now been employed successfully as a cancer stem cells (CSCs) marker in various tumors. We wanted to investigate whether ALDH1 can be used as a putative CSCs marker and a powerful prognostic factor in nasopharyngeal carcinoma (NPC). Here, we isolated ALDH1-positive cells from human NPC cell lines (5-8F and CNE2) and found that ALDH1-positive cancer cells grew faster and had higher clone formation efficiency (0.435 ± 0.15; 0.431 ± 0.025 vs. 0.131 ± 0.007; 0.121 ± 0.126), differentiation capability and had higher migration (233.00 ± 5.29; 228.60 ± 9.34 vs. 123.20 ± 7.70 vs. 97.20 ± 6.61) than those of ALDH1-negative cancer cells in vitro. In addition, ALDH1- positive cancer cells formed significantly more tumor spheres. Our in vivo experiments showed that only 5 × 10³ ALDH1-positive NPC cells were required to induce tumors. Notably, ALDH1-positive cells are enriched in the side-population (SP) cells, and stem cells-like genes OCT4, BMI1, KLF4 and SOX2 are preferentially expressed in ALDH1-positive cells. Immunohistochemical results showed that the expression of ALDH1 correlated significantly with T classification (P = 0.011), N classification (P = 0.005), M classification (P = 0.002) and clinical stage (P = 0.001). Interestingly, ALDH1 expression in the tumor correlated significantly with epithelial–mesenchymal transition (EMT) markers including vimentin expression and loss of E-cadherin (P = 0.003 and 0.008, respectively). Furthermore, multivariate analyses showed that ALDH1 expression was an independent prognostic indicator (P = 0.032). Taken together, for the first time, we demonstrate that ALDH1 could be a novel stem cells marker and a valuable predictor of poor survival NPC.     

CD44+ CD24(-) prostate cells are early cancer progenitor/stem cells that provide a model for patients with poor prognosis

Recent evidence supports the hypothesis that cancer stem cells are responsible for tumour initiation and formation. Using flow cytometry, we isolated a population of CD44+CD24(-) prostate cells that display stem cell characteristics as well as gene expression patterns that predict overall survival in prostate cancer patients. CD44+CD24(-) cells form colonies in soft agar and form tumours in NOD/SCID mice when as few as 100 cells are injected. Furthermore, CD44+CD24(-) cells express genes known to be important in stem cell maintenance, such as BMI-1 and Oct-3/4. Moreover, we can maintain CD44+CD24(-) prostate stem-like cells as nonadherent spheres in serum-replacement media without substantially shifting gene expression. Addition of serum results in adherence to plastic and shifts gene expression patterns to resemble the differentiated parental cells. Thus, we propose that CD44+CD24(-) prostate cells are stem-like cells responsible for tumour initiation and we provide a genomic definition of these cells and the differentiated cells they give rise to. Furthermore, gene expression patterns of CD44+CD24(-) cells have a genomic signature that is predictive of poor patient prognosis. Therefore, CD44+CD24(-) LNCaP prostate cells offer an attractive model system to both explore the biology important to the maintenance and differentiation of prostate cancer stem cells as well as to develop the therapeutics, as the gene expression pattern in these cells is consistent with poor survival in prostate cancer patients.     

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。     

DeltaNp63 expression is associated with poor survival in ovarian cancer.

BACKGROUND: P63 belongs to the 'p53 family' whose role in cancer progression has been recently revisited in light of the plethora of splicing variants that are generated. We analyzed the expression of the full-length TAp63 gene and its dominant-negative form deltaNp63 in ovarian cancer biopsies to correlate their expression with clinical outcome. MATERIALS AND METHODS: Real-time RT-PCR analysis was used to determine the levels of TAp63 and deltaNp63 in 83 stage I and in 86 stage III ovarian cancer biopsies and in seven human ovarian cancer cell. RESULTS: TAp63 levels were comparable in stage I and stage III, but deltaNp63 levels increased 77-fold in stage III, independently of the p53 status. Patients with high deltaNp63 expression had the worst overall survival (OS); patients with a deltaNp63/TAp63 ratio >2 had a poor OS. Patients with a high deltaNp63/TAp63 ratio were those with a poor response to platinum-based therapy. CONCLUSIONS: Data indicate a role for deltaNp63 as a potential biomarker to predict patient's outcome and tumor progression in ovarian cancer. This would have particularly clinical relevance in ovarian cancer where the high rate of mortality reflects our lack of knowledge of molecular mechanisms underlying cell progression toward malignancy.     

Hypoploidy defines patients with poor prognosis in breast cancer

The clinicopathological features currently used in breast cancer prognosis often fail to characterize the clinical heterogeneity of the disease accurately. Our study is aimed to investigate the predictive value of DNA flow cytometry in breast cancer. Previously untreated breast carcinoma samples (584) were snap frozen for flow-cytometry. Tumors were classified into three DNA index (DI) categories: i) tumors showing a DI =0.96-1.15 (diploid and near-diploid); ii) tumors with a DI >or=1.16 (hyperdiploid, tetraploid, multiploid and/or those with more than one diploid population); and iii) tumors with a DI 相似文献   

The side population of ovarian cancer cells defines a heterogeneous compartment exhibiting stem cell characteristics

Cancer stem cells (CSC) are believed to be involved in tumor evasion of classical antitumor therapies and have thus become an attractive target for further improvement of anticancer strategies. However, the existence and identity of CSC are still a matter of controversy. In a systematic screen of 13 ovarian cancer cell lines we show that cells with stem cell properties are reliably detectable as a minor population, characterized by ABC transporter expression resulting in the side population (SP) phenotype. In different cell lines, either ABCG2 or ABCB1 was found to be responsible for this effect. Purified SP cells featured virtually all characteristics of bona fide CSC, including clonogenicity, asymmetric division and high tumorigenicity in vivo. Using in-depth phenotyping by multicolor flow cytometry, we found that among the investigated ovarian cancer cell lines the SP compartment exhibits tremendous heterogeneity and is composed of multiple phenotypically distinct subpopulations. Thus, our study confirms previous results showing that CSC are contained within the SP. However, the exact identity of the CSC is still disguised by the high complexity of the CSC-containing compartment. Further functional studies are needed to determine whether a single cellular subset can unambiguously be defined as CSC or whether multiple stem cell-like cells with different properties coexist. Moreover, the observed heterogeneity may reflect a high level of plasticity and likely influences tumor progression, escape from immune-surveillance and development of resistance to anticancer therapies and should therefore be considered in the development of new treatment strategies.     

Impact of CD133 positive stem cell proportion on survival in patients with glioblastoma multiforme

Background

The aim of the study was to assess the impact of CD133-positive (CD133+) cancer stem cell proportions on treatment results of glioblastoma multiforme (GBM) patients.

Patients and methods

Patients with GBM (n = 42) received postoperative radiotherapy (± chemotherapy). Surgically excised GBM tissue sections were immunohistochemically examined for CD133 expression. The proportions of CD133+ GBM cells were determined (%). The proportion of CD133+ GBM stem cells was established by 2 independent researchers whose results were in good accordance (R = 0.8, p < 0.01). Additionally, CD133 expression levels were correlated with patients overall survival.

Results

The proportion of CD133+ cells varied between patients, being from 0.5% to 82%. Mean and median proportions of CD133+ cells of the entire study group were 33% ± 24% (mean ± SD) and 28%, respectively. Clinical data do not support the association between higher proportion of stem cells and the aggressiveness of GBM. Median survival time of the study group was 10.0 months (95% CI 9.0–11.0). The survival time clearly depended on the proportion of CD133+ cells (log rank test, p = 0.02). Median survival times for patients with low (< median) and high (≥ median) proportion of CD133+ cells were 9.0 months (95% CI 7.6–10.5) and 12.0 months (95% CI 9.3–14.7), respectively. In multivariate analysis, the proportion of CD133+ cells emerged as a significant independent predictor for longer overall survival (HR 2.0, 95% CI 1.0–3.8, p = 0.04).

Conclusions

In patients with higher stem cell proportion, significantly longer survival times after postoperative radiotherapy were achieved. Underlying reasons and possible higher sensitivity of GBM stem cells to fractionated radio-therapy should be clarified in further studies.     

Influence on biological behavior of colon cancer stem cells after RNA interfering CD133

Objective  

To observe the effects of blocking CD133 gene expression and activation on biological characteristic of the colon cancer stem cells.     

Brca1 breast tumors contain distinct CD44+/CD24- and CD133+ cells with cancer stem cell characteristics

Introduction

Whether cancer stem cells occur in BRCA1-associated breast cancer and contribute to therapeutic response is not known.

Methods

We generated and characterized 16 cell lines from five distinct Brca1deficient mouse mammary tumors with respect to their cancer stem cell characteristics.

Results

All cell lines derived from one tumor included increased numbers of CD44⁺/CD24^- cells, which were previously identified as human breast cancer stem cells. All cell lines derived from another mammary tumor exhibited low levels of CD44⁺/CD24^- cells, but they harbored 2% to 5.9% CD133⁺ cells, which were previously associated with cancer stem cells in other human and murine tumors. When plated in the absence of attachment without presorting, only those cell lines that were enriched in either stem cell marker formed spheroids, which were further enriched in cells expressing the respective cancer stem cell marker. In contrast, cells sorted for CD44⁺/CD24^- or CD133⁺ markers lost their stem cell phenotype when cultured in monolayers. As few as 50 to 100 CD44⁺/CD24^- or CD133⁺ sorted cells rapidly formed tumors in nonobese diabetic/severe combined immunodeficient mice, whereas 50-fold to 100-fold higher numbers of parental or stem cell depleted cells were required to form few, slow-growing tumors. Expression of stem cell associated genes, including Oct4, Notch1, Aldh1, Fgfr1, and Sox1, was increased in CD44⁺/CD24^- and CD133⁺ cells. In addition, cells sorted for cancer stem cell markers and spheroid-forming cells were significantly more resistant to DNA-damaging drugs than were parental or stem cell depleted populations, and they were sensitized to the drugs by the heat shock protein-90 inhibitor 17-DMAG (17-dimethylaminoethylamino-17-demethoxygeldanamycin hydrochloride).

Conclusion

Brca1-deficient mouse mammary tumors harbor heterogeneous cancer stem cell populations, and CD44⁺/CD24^- cells represent a population that correlates with human breast cancer stem cells.