胃癌干细胞的分离鉴定以及干细胞特性研究

背景:肿瘤干细胞(CSCs)已成为当前肿瘤研究的热点之一,开展相关研究的首要问题是CSCs的分离和鉴定,悬浮培养法是分离CSCs的重要方法。目的:分离、鉴定胃癌干细胞(GCSCs)并评价其干细胞特性。方法:人胃癌细胞株MKN45、MGC803、SGC7901以含表皮生长因子(EGF)、碱性成纤维细胞生长因子(bFGF)的无血清培养基悬浮培养。对培养得到的第三代肿瘤球,以集落形成实验检测其克隆形成能力,免疫荧光法检测GCSCs标记物,细胞划痕实验和Transwell小室细胞侵袭实验检测其迁移、侵袭能力,CCK-8实验检测其对顺铂的耐药性。结果:培养得到的肿瘤球高表达干细胞标记物CD44、CD54,低表达胃壁细胞标记物H+/K+-ATPase,克隆形成能力、划痕愈合速度和Transwell小室穿膜细胞数明显大于或多于普通胃癌细胞,差异有统计学意义(P0.05)。顺铂对肿瘤球的50%抑制浓度明显高于普通胃癌细胞。结论:应用无血清培养基悬浮培养法能成功获得GCSCs富集的肿瘤球;肿瘤球具有较强的自我更新、增殖、迁移、侵袭能力和多向分化潜能,对化疗药物更易产生耐药性。     

Gastrointestinal cancer stem cells as targets for innovative immunotherapy

The role of cancer stem cells in gastrointestinal cancer-associated death has been widely recognized. Gastrointestinal cancer stem cells(GCSCs) are considered to be responsible for tumor initiation, growth, resistance to cytotoxic therapies,recurrence and metastasis due to their unique properties. These properties make the current therapeutic trials against GCSCs ineffective. Moreover, recent studies have shown that targeting stem cell surface markers or stemness associated pathways might have an additional off-target effect on the immune system.Recent advances in oncology and precision medicine have opened alternative therapeutic strategies in the form of cancer immunotherapy. This approach differs from classical anti-cancer therapy through its mechanism of action involving the activation and use of a functional immune system against tumor cells, instead of aiming physically destruction of cancer cells through radio-or chemotherapy. New immunological approaches for GCSCs targeting involve the use of different immune cells and various immune mechanisms like targeting specific surface antigens, using innate immune cells like the natural killer and T cells, T-cell chimeric antigen receptor technology, dendritic cell vaccine, or immune checkpoint inhibitors. In this respect, better understandings of immune regulatory mechanisms that govern anti-tumor response bring new hope in obtaining long-term remission for cancer therapy.     

Stem cells in gastric cancer

Gastric cancer(GC) is one of the leading causes of cancerrelated mortality worldwide.Cancer stem cells(CSCs),which were first identified in acute myeloid leukemia and subsequently in a large array of solid tumors,play important roles in cancer initiation,dissemination and recurrence.CSCs are often transformed tissue-specific stem cells or de-differentiated transit amplifying progenitor cells.Several populations of multipotent gastric stem cells(GSCs) that reside in the stomach have been determined to regulate physiological tissue renewal and injury repair.These populations include the Villin+ and Lgr5+ GSCs in the antrum,the Troy+ chief cells in the corpus,and the Sox2+ GSCs that are found in both the antrum and the corpus.The disruption of tumor suppressors in Villin+ or Lgr5+ GSCs leads to GC in mouse models.In addition to residing GSCs,bone marrow-derived cells can initiate GC in a mouse model of chronic Helicobacter infection.Furthermore,expression of the cell surface markers CD133 or CD44 defines gastric CSCs in mouse models and in human primary GC tissues and cell lines.Targeted elimination of CSCs effectively reduces tumor size and grade in mouse models.In summary,the recent identification of normal GSCs and gastric CSCs has greatly improved our understanding of the molecular and cellular etiology of GC and will aid in the development of effective therapies to treat patients.     

The biology of cancer stem cells and its clinical implication in hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is a highly malignant tumor with limited treatment options in its advanced state. The molecular mechanisms underlying HCC remain unclear because of the complexity of its multi-step development process. Cancer stem cells (CSCs) are defined as a small population of cells within a tumor that possess the capability for self-renewal and the generation of heterogeneous lineages of cancer cells. To date, there have been two theories concerning the mechanism of carcinogenesis, i.e., the stochastic (clonal evolution) model and the hierarchical (cancer stem cell-driven) model. The concept of the CSC has been established over the past decade, and the roles of CSCs in the carcinogenic processes of various cancers, including HCC, have been emphasized. Previous experimental and clinical evidence indicated the existence of liver CSCs; however, the potential mechanistic links between liver CSCs and the development of HCC in humans are not fully understood. Although definitive cell surface markers for liver CSCs have not yet been found, several putative markers have been identified, which allow the prospective isolation of CSCs from HCC. The identification and characterization of CSCs in HCC is essential for a better understanding of tumor initiation or progression in relation to signaling pathways. These markers could be used along with clinical parameters for the prediction of chemoresistance, radioresistance, metastasis and survival and may represent potential targets for the development of new molecular therapies against HCC. This review describes the current evidence for the existence and function of liver CSCs and discuss the clinical implications of CSCs in patients demonstrating resistance to conventional anti-cancer therapies, as well as clinical outcomes. Such data may provide a future perspective for targeted therapy in HCC.     

消化系肿瘤干细胞研究进展

近年来,肿瘤干细胞（CSCs）已成为肿瘤研究的热点。CSCs是一类具有自我更新和分化潜能的细胞．参与肿瘤的发生、发展、转移和复发。迄今,已发现包括食管癌、胃癌、结肠癌、肝癌、胰腺癌等在内的多种消化系CSCs。本文就消化系CSCs的研究进展作一综述,以期为消化系肿瘤的治疗提供新的策略。     

Liver cancer stem cell markers: Progression and therapeutic implications

Cancer stem cells(CSCs) are a small subpopulation in cancer, have been proposed to be cancer-initiating cells, and have been shown to be responsible for chemotherapy resistance and cancer recurrence. The identification of CSC subpopulations inside a tumor presents a new understanding of cancer development because it implies that tumors can only be eradicated by targeting CSCs. Although advances in liver cancer detection and treatment have increased the possibility of curing the disease at early stages, unfortunately, most patients will relapse and succumb to their disease. Strategies aimed at efficiently targeting liver CSCs are becoming important for monitoring the progress of liver cancer therapy and for evaluating new therapeutic approaches. Herein, we provide a critical discussion of biological markers described in the literature regarding liver cancer stem cells and the potential of these markers to serve as therapeutic targets.     

Novel therapeutic target for cancer stem cells in hepatocellular carcinoma

Cancer is a disease of genetic and epigenetic alterations, which are emphasized as the central mechanisms of tumor progression in the multi-stepwise model. Discovery of rare subpopulations of cancer stem cells (CSCs) has created a new focus in cancer research. The heterogeneity of tumors can be explained with the help of CSCs supported by anti-apoptotic signaling. CSCs mimic normal adult stem cells by demonstrating unique characteristics of self-renewal and pluripotency, and the critical role for tumor growth and resistance to anti-cancer therapy. We found that CD13 was a surface marker for CSCs in human liver cancer cell lines and clinical samples, and that CD13+ CSCs were associated with a hypoxic marker in clinical hepatocellular carcinoma (HCC) sample, suggesting that CD13+ CSCs have the critical role in tumor growth and resistance to anti-cancer therapy in liver cancers. In this review article, we update recent findings regarding the involvement of CSCs, especially in HCC.     

Cancer stem cells: Involvement in pancreatic cancer pathogenesis and perspectives on cancer therapeutics

Pancreatic cancer is one of the most aggressive and lethal malignancies. Despite remarkable progress in understanding pancreatic carcinogenesis at the molecular level, as well as progress in new therapeutic approaches, pancreatic cancer remains a disease with a dismal prognosis. Among the mechanisms responsible for drug resistance, the most relevant are changes in individual genes or signaling pathways and the presence of highly resistant cancer stem cells (CSCs). In pancreatic cancer, CSCs represent 0.2%-0.8% of pancreatic cancer cells and are considered to be responsible for tumor growth, invasion, metastasis and recurrence. CSCs have been extensively studied as of late to identify specific surface markers to ensure reliable sorting and for signaling pathways identified to play a pivotal role in CSC self-renewal. Involvement of CSCs in pancreatic cancer pathogenesis has also highlighted these cells as the preferential targets for therapy. The present review is an update of the results in two main fields of research in pancreatic cancer, pathogenesis and therapy, focused on the narrow perspective of CSCs.     

Hepatic cancer stem cells and drug resistance: Relevance in targeted therapies for hepatocellular carcinoma

Hepatocellular carcinoma (HCC) is one of most common malignancies in the world. Systemic treatments for HCC, particularly for advanced stages, are limited by the drug resistance phenomenon which ultimately leads to therapy failure. Recent studies have indicated an association between drug resistance and the existence of the cancer stem cells (CSCs) as tumor initiating cells. The CSCs are resistant to conventional chemotherapies and might be related to the mechanisms of the ATP Binding Cassette (ABC) transporters and alterations in the CSCs signaling pathways. Therefore, to contribute to the development of new HCC treatments, further information on the characterization of CSCs, the modulation of the ABC transporters expression and function and the signaling pathway involved in the self renewal, initiation and maintenance of the cancer are required. The combination of transporters modulators/inhibitors with molecular targeted therapies may be a potent strategy to block the tumoral progression. This review summarizes the association of CSCs, drug resistance, ABC transporters activities and changes in signaling pathways as a guide for future molecular therapy for HCC.     

CD133:A cancer stem cells marker,is used in colorectal cancers

Colorectal cancer is one of the most common malignant tumors worldwide. A model of cancer development involving cancer stem cells has been put forward because it provides a possible explanation of tumor hierarchy. Cancer stem cells are characterized by their proliferation, tumorigenesis, differentiation, and selfrenewal capacities, and chemoradiotherapy resistance. Due to the role of cancer stem cells in tumor initiation and treatment failure, studies of cancer stem cell markers, such as CD133, have been of great interest. CD133, a five-transmembrane glycoprotein, is widely used as a marker to identify and isolate colorectal cancer stem cells. This marker has been investigated to better understand the characteristics and functions of cancer stem cells. Moreover, it can also be used to predict tumor progression, patient survival, chemoradiotherapy resistance and other clinical parameters. In this review, we discuss the use of CD133 in the identification of colorectal cancer stem cell, which is currently controversial. Although the function of CD133 is as yet unclear, we have discussed several possible functions and associated mechanisms that may partially explain the role of CD133 in colorectal cancers. In addition, we focus on the prognostic value of CD133 in colorectal cancers. Finally, we predict that CD133 may be used as a possible target for colorectal cancer treatment.     

Cancer stem cells as therapeutic targets of hepato‐biliary‐pancreatic cancers

Heterogeneity is one of the essential hallmarks of malignancies. Within bulk cancer cells, a striking variability differs in biological characteristics including the proliferation rate, cell–cell interaction, metastatic tendency and even sensitivity to anticancer therapies. Such diversity makes the investigation and treatment of the cancers complicated. Increasing evidence suggests this plasticity of cancers is a result of self‐renewing and differentiation of a small subpopulation of cancer cells with stem‐like properties, called cancer stem cells (CSCs). More importantly, CSCs are believed to be responsible for the resistance to conventional therapies and metastatic abilities in clinical practice. This review summarizes the molecular pathogenesis of hepato‐biliary‐pancreatic CSCs on the basis of the recent studies, and promising strategy of novel therapy targeting the signal transduction pathways or potentially epigenetic addictions of CSCs.     

Involvement of non-vascular stem cells in blood vessel formation

Blood vessels clearly act as conduits for blood flow, but recently the concept that they are also involved in organ maintenance, especially by providing a niche for organ-specific stem cells, has begun to emerge. Moreover, several lines of evidence suggest that hematopoietic stem cells can differentiate directly into cells composing blood vessels. Recently, cancer stem cells (CSCs) have also been assigned these roles in the cancer microenvironment. Although anti-angiogenic drugs have been developed and are utilized in the clinic for their anti-tumor activity, their suppressive effects on tumor growth have been disappointing. This may be caused by transferring drug resistance from CSCs to endothelial cells. It has been suggested that CSCs localize in the peri-vascular niche. Therefore, it is extremely important to know how the vascular niche maintains CSCs, as such knowledge may enable us to develop promising new approaches to cancer treatment.     

肿瘤干细胞的研究进展

肿瘤干细胞是指肿瘤组织中存在的一小群具有自我更新能力和多向分化潜能的细胞。肿瘤干细胞主宰着肿瘤分化、癌细胞增殖、自我更新及血管形成等诸多方面。如果能够有效抑制肿瘤干细胞,就可以从源头上阻断恶性肿瘤的发生、发展。本文对肿瘤干细胞的起源、肿瘤干细胞学说的发展、肿瘤干细胞与肿瘤异质性、转移和耐药的关系以及针对肿瘤干细胞的治疗策略等方面进行综述,为未来肿瘤的诊断和治疗提供新思路。     

A subpopulation of CD24⁺ cells in colon cancer cell lines possess stem cell characteristics

Cancer stem cells (CSCs) have been shown to contribute to the resistance and relapse in a range of cancer types such as breast cancer and glioma. However, colon cancer stem cells remain poorly characterized. Here we reported that CD24+ subpopulation in colon cancer cell lines HCT116 and SW480 exhibited cancer stem cell-like characteristics. Using flow cytometry candidate CSCs markers were selected after initial screening of known CSCs markers from other types of cancer on colon cancer cell lines HCT116, SW480 and HT29. CD24 was expressed in the minority of bulk cell population of HCT116 and SW480 cell lines. Moreover, functional tests demonstrated that CD24+ cells exhibited enhanced chemotherapy-resistance, self-renewal and tumorigenic capacity both in vitro and in vivo, compared to CD24- subpopulations. These results suggest that CD24+ subpopulation in colon cancer cell lines HCT116 and SW480 exhibits CSCs like characteristics, and represents a nice model to study and develop effective strategies to overcome chemo-resistance and relapse of colon cancer.     

Oncolytic viruses against cancer stem cells: A promising approach for gastrointestinal cancer

Gastrointestinal cancer has been one of the five most commonly diagnosed and leading causes of cancer mortality over the past few decades. Great progress in traditional therapies has been made, which prolonged survival in patients with early cancer, yet tumor relapse and drug resistance still occurred, which is explained by the cancer stem cell(CSC) theory. Oncolytic virotherapy has attracted increasing interest in cancer because of its ability to infect and lyse CSCs. This paper reviews the basic knowledge, CSC markers and therapeutics of gastrointestinal cancer(liver, gastric, colon and pancreatic cancer), as well as research advances and possible molecular mechanisms of various oncolytic viruses against gastrointestinal CSCs. This paper also summarizes the existing obstacles to oncolytic virotherapy and proposes several alternative suggestions to overcome the therapeutic limitations.     

Regulation of colon cancer recurrence and development of therapeutic strategies

Recurrence of colon cancer still remains a major issue which affects nearly 50% of patients treated by conventional therapeutics. Although the underlying causative factor(s) is not fully understood, development of drug-resistance has been associated with induction of cancer stem or stem-like cells (CSCs) which constitute a small sub-population of tumor cells known to be highly resistant to chemotherapy. In fact, the discovery of CSCs in a variety of tumors (including colon cancer) has changed the view of carcinogenesis and therapeutic strategies. Emerging reports have indicated that to improve patient outcomes, conventional anticancer therapies should be replaced with specifi c approaches targeting CSCs. Thus, therapeutic strategies that specifically target CSCs are being sought to reduce the risk of relapse and metastasis. In order to specifi cally target colon CSCs (while sparing somatic intestinal stem cells), it is critical to identify unique deregulated pathways responsible for self-renewal of CSCs and colon cancer recurrence. Colon CSCs present a unique opportunity to better understand the biology of solid tumors. Thus, a better understanding of the clinical signs and symptoms of colon cancer patients (under-going surgery or chemotherapy) during perioperative periods, along with the underlying regulatory events affecting the stem/progenitor cell self-renewal and differentiation of colon epithelial cells, is of immense importance. In this review we discuss the implication of clinical factors and the emerging role of CSCs during recurrence of colon cancer along with the development of new therapeutic strategies involving the use of natural agents.     

CD13: Waving the flag for a novel cancer stem cell target

Cancer stem cells (CSCs) are generally dormant or slowly cycling tumor cells that have the ability to reconstitute tumors. They are thought to be involved in tumor resistance to chemo/radiation therapy and tumor relapse and progression. However, neither their existence nor their identity within many cancers has been well defined. Here, we have demonstrated that CD13 is a marker for semiquiescent CSCs in human liver cancer cell lines and clinical samples and that targeting these cells might provide a way to treat this disease. CD13+ cells predominated in the G0 phase of the cell cycle and typically formed cellular clusters in cancer foci. Following treatment, these cells survived and were enriched along the fibrous capsule where liver cancers usually relapse. Mechanistically, CD13 reduced ROS-induced DNA damage after genotoxic chemo/radiation stress and protected cells from apoptosis. In mouse xenograft models, combination of a CD13 inhibitor and the genotoxic chemotherapeutic fluorouracil (5-FU) drastically reduced tumor volume compared with either agent alone. 5-FU inhibited CD90+ proliferating CSCs, some of which produce CD13+ semiquiescent CSCs, while CD13 inhibition suppressed the self-renewing and tumor-initiating ability of dormant CSCs. Therefore, combining a CD13 inhibitor with a ROS-inducing chemo/radiation therapy may improve the treatment of liver cancer.     

Glioma stem cells: turpis omen in nomen? (the evil in the name?)

High‐grade gliomas remain incurable and lethal. Through the availability of the stem‐like cells responsible for glioblastoma (GB) formation, expansion, resilience and recurrence, the discovery of glioma cancer stem cells (GCSCs) is revolutionizing this field. GCSCs provide an unprecedented opportunity to reproduce and study GB pathophysiology more accurately. This critically emphasizes our ability to unambiguously identify, isolate and investigate cells that do qualify as GCSCs, to use them as a potential model that is truly predictive of GBs and of their regulation and response to therapeutic agents. We review this concept against the background of key findings on somatic, neural and solid tumour stem cells (SCs), also taking into account the emerging phenomenon of phenotypic SC plasticity. We suggest that basic approaches in these areas can be imported into the GCSC field, so that the same functional method used to identify normal somatic SCs becomes the most appropriate to define GCSCs. This, combined with knowledge of the cellular and molecular basis of normal adult neurogenesis, promises to improve the identification of GCSCs and of selective markers, as well as the development of innovative, more specific and efficacious antiglioma strategies.     

Cancer stem cells in Helicobacter pylori infection and aging: Implications for gastric carcinogenesis

AIM: To demonstrated the combined effects of aging and carcinogen treatment on cancer stem/stem-like cells(CSCs) of gastric mucosa in an animal model. METHODS: In this study we investigated the effects of aging and Helicobacter pylori(H. pylori) inflammation as a model for inflammation induced carcinogenesis in human and rat gastric mucosa samples. In aging studies, we compared 4-mo old(young) with 22 mo(aged) old Fischer-344 rats. For human studies, gastric biop-sies and resection specimens representing normal mucosa or different stages of H. pylori gastritis and gastric adenocarcinomas were used for determining the expression of stem cell markers CD166, ALDH1 and LGR5. In addition we performed immunofluorescent double labeling for B-catenin and Lgr5 in both rat and human gastric tissues to examine the status of Wnt signaling in these cells. RESULTS: CSC markers ALDH1, LGR5, and CD166 were expressed in very low levels in normal human gastric mucosa or young rat gastric mucosa. In contrast, level of expression for all three markers significantly increased in H. pylori gastritis and gastric adenocarcinomas as well as in normal gastric mucosa in aged rats. We also observed cytoplasmic B-catenin staining in both aged rat and human H. pylori inflamed gastric mucosa, which were found to be colocalized with Lgr5 immunoreactive cells. The increased number of ALDH1, CD166 and LGR5 positive cells in H. pylori gastritis indicates that increased number of stem-like cells in gastric mucosa is an early event, and may constitute an important step in the progression to neoplasia. CONCLUSION: Our observation of the age-related increase in cancer stem/stem-like cells in the gastric mucosa may explain the increased incidence of gastric cancer during aging. Combination of aging and H. pylori infection may have additive effects in progression to neoplasia.     

Role of microRNA-regulated cancer stem cells in recurrent hepatocellular carcinoma

Among the most common cancers, hepatocellular carcinoma (HCC) has a high rate of tumor recurrence, tumor dormancy, and drug resistance after initial successful chemotherapy or radiotherapy. A small subset of cancer cells, cancer stem cells (CSCs), exhibit stem cell characteristics and are present in various cancers, including HCC. The dysregulation of microRNAs (miRNAs) often accompanies the occurrence and development of HCC. miRNAs can influence tumorigenesis, progression, recurrence, and drug resistance by regulating CSCs properties, which supports their clinical utility in managing and treating HCC. This review summarizes the regulatory effects of miRNAs on CSCs in HCC with a special focus on their impact on HCC recurrence.