搜索结果: 'methocult media formulations for mouse hematopoietic cells serum containing'

Normal stem cells and cancer stem cells (CSCs) share similar properties,in that both have the capacity to self-renew and differentiate into multiple cell types. In both the normal stem cell and cancer stem cell fields,there has been a great need for a universal marker that can effectively identify and isolate these rare populations of cells in order to characterize them and use this information for research and therapeutic purposes. Currently,it would appear that certain isoenzymes of the aldehyde dehydrogenase (ALDH) superfamily may be able to fulfill this role as a marker for both normal and cancer stem cells. ALDH has been identified as an important enzyme in the protection of normal hematopoietic stem cells,and is now also widely used as a marker to identify and isolate various types of normal stem cells and CSCs. In addition,emerging evidence suggests that ALDH1 is not only a marker for stem cells,but may also play important functional roles related to self-protection,differentiation,and expansion. This comprehensive review discusses the role that ALDH plays in normal stem cells and CSCs,with focus on ALDH1 and ALDH3A1. Discrepancies in the functional themes between cell types and future perspectives for therapeutic applications will also be discussed. View Publication

The aetiology of human fibrolamellar hepatocellular carcinomas (hFL-HCCs),cancers occurring increasingly in children to young adults,is poorly understood. We present a transplantable tumour line,maintained in immune-compromised mice,and validate it as a bona fide model of hFL-HCCs by multiple methods. RNA-seq analysis confirms the presence of a fusion transcript (DNAJB1-PRKACA) characteristic of hFL-HCC tumours. The hFL-HCC tumour line is highly enriched for cancer stem cells as indicated by limited dilution tumourigenicity assays,spheroid formation and flow cytometry. Immunohistochemistry on the hFL-HCC model,with parallel studies on 27 primary hFL-HCC tumours,provides robust evidence for expression of endodermal stem cell traits. Transcriptomic analyses of the tumour line and of multiple,normal hepatic lineage stages reveal a gene signature for hFL-HCCs closely resembling that of biliary tree stem cells--newly discovered precursors for liver and pancreas. This model offers unprecedented opportunities to investigate mechanisms underlying hFL-HCCs pathogenesis and potential therapies. View Publication

Transformed,oncogenic precursors,possessing both defining neural-stem-cell properties and the ability to initiate intracerebral tumours,have been identified in human brain cancers. Here we report that bone morphogenetic proteins (BMPs),amongst which BMP4 elicits the strongest effect,trigger a significant reduction in the stem-like,tumour-initiating precursors of human glioblastomas (GBMs). Transient in vitro exposure to BMP4 abolishes the capacity of transplanted GBM cells to establish intracerebral GBMs. Most importantly,in vivo delivery of BMP4 effectively blocks the tumour growth and associated mortality that occur in 100% of mice after intracerebral grafting of human GBM cells. We demonstrate that BMPs activate their cognate receptors (BMPRs) and trigger the Smad signalling cascade in cells isolated from human glioblastomas (GBMs). This is followed by a reduction in proliferation,and increased expression of markers of neural differentiation,with no effect on cell viability. The concomitant reduction in clonogenic ability,in the size of the CD133+ population and in the growth kinetics of GBM cells indicates that BMP4 reduces the tumour-initiating cell pool of GBMs. These findings show that the BMP-BMPR signalling system--which controls the activity of normal brain stem cells--may also act as a key inhibitory regulator of tumour-initiating,stem-like cells from GBMs and the results also identify BMP4 as a novel,non-cytotoxic therapeutic effector,which may be used to prevent growth and recurrence of GBMs in humans. View Publication

BACKGROUND: Primitive brain tumors are the leading cause of cancer-related death in children. Tumor cells with stem-like properties (TSCs),thought to account for tumorigenesis and therapeutic resistance,have been isolated from high-grade gliomas in adults. Whether TSCs are a common component of pediatric brain tumors and are of clinical relevance remains to be determined. METHODOLOGY/PRINCIPAL FINDINGS: Tumor cells with self-renewal properties were isolated with cell biology techniques from a majority of 55 pediatric brain tumors samples,regardless of their histopathologies and grades of malignancy (57% of embryonal tumors,57% of low-grade gliomas and neuro-glial tumors,70% of ependymomas,91% of high-grade gliomas). Most high-grade glioma-derived oncospheres (10/12) sustained long-term self-renewal akin to neural stem cells (textgreater7 self-renewals),whereas cells with limited renewing abilities akin to neural progenitors dominated in all other tumors. Regardless of tumor entities,the young age group was associated with self-renewal properties akin to neural stem cells (P = 0.05,chi-square test). Survival analysis of the cohort showed an association between isolation of cells with long-term self-renewal abilities and a higher patient mortality rate (P = 0.013,log-rank test). Sampling of low- and high-grade glioma cultures showed that self-renewing cells forming oncospheres shared a molecular profile comprising embryonic and neural stem cell markers. Further characterization performed on subsets of high-grade gliomas and one low-grade glioma culture showed combination of this profile with mesenchymal markers,the radio-chemoresistance of the cells and the formation of aggressive tumors after intracerebral grafting. CONCLUSIONS/SIGNIFICANCE: In brain tumors affecting adult patients,TSCs have been isolated only from high-grade gliomas. In contrast,our data show that tumor cells with stem cell-like or progenitor-like properties can be isolated from a wide range of histological sub-types and grades of pediatric brain tumors. They suggest that cellular mechanisms fueling tumor development differ between adult and pediatric brain tumors. View Publication

BACKGROUND Mechanisms of glioma invasion remain to be fully elucidated. Glioma cells within glioblastoma multiforme (GBM) range from well-differentiated tumor cells to less-differentiated brain tumor-initiating cells (BTICs). The β2-subunit of Na(+)/K(+)-ATPase,called the adhesion molecule on glia (AMOG),is highly expressed in normal glia but is thought to be universally downregulated in GBM. To test our hypothesis that expression of AMOG is heterogeneous in GBM and confers a less invasive phenotype,we compared it between BTICs and differentiated cells from patient-matched GBM and then tested GBM invasion in vitro after AMOG overexpression. METHODS Immunohistochemistry,immunoblotting,and real-time PCR were used to characterize AMOG protein and mRNA expression in tumor samples,BTICs,and differentiated cells. Matrigel invasion assay,scratch assay,and direct cell counting were used for testing in vitro invasion,migration,and proliferation,respectively. RESULTS While AMOG expression is heterogeneous in astrocytomas of grades II-IV,it is lost in most GBM. BTICs express higher levels of AMOG mRNA and protein compared with patient-matched differentiated tumor cells. Overexpression of AMOG decreased GBM cell and BTIC invasion without affecting migration or proliferation. Knockdown of AMOG expression in normal human astrocytes increased invasion. CONCLUSIONS AMOG expression inhibits GBM invasion. Its downregulation increases invasion in glial cells and may also represent an important step in BTIC differentiation. These data provide compelling evidence implicating the role of AMOG in glioma invasion and provide impetus for further investigation. View Publication

In the central nervous system,neural stem cells proliferate in the ventricular zone (VZ) and sequentially give rise to both neurons and glial cells in a temporally and spatially regulated manner,suggesting that stem cells may differ from one another in different brain regions and at different developmental stages. For the purpose of marking and purifying neural stem cells to ascertain whether such differences exist,we generated transgenic mice using promoters from Hes genes (pHes1 or pHes5) to drive expression of destabilized enhanced green fluorescent protein. In the developing brains of these transgenic mice,GFP expression was restricted to undifferentiated cells in the VZ,which could asymmetrically produce a Numb-positive neuronal daughter and a GFP-positive progenitor cell in clonal culture,indicating that they retain the capacity to self-renew. Our results suggest that pHes-EGFP transgenic mice can be used to explore similarities and differences among neural stem cells during development. View Publication

The detection of growth hormone (GH) and its receptor in germinal regions of the mammalian brain prompted our investigation of GH and its role in the regulation of endogenous neural precursor cell activity. Here we report that the addition of exogenous GH significantly increased the expansion rate in long-term neurosphere cultures derived from wild-type mice,while neurospheres derived from GH null mice exhibited a reduced expansion rate. We also detected a doubling in the frequency of large (i.e. stem cell-derived) colonies for up to 120 days following a 7-day intracerebroventricular infusion of GH suggesting the activation of endogenous stem cells. Moreover,gamma irradiation induced the ablation of normally quiescent stem cells in GH-infused mice,resulting in a decline in olfactory bulb neurogenesis. These results suggest that GH activates populations of resident stem and progenitor cells,and therefore may represent a novel therapeutic target for age-related neurodegeneration and associated cognitive decline. View Publication

Glioblastoma multiforme is the most malignant type of primary brain tumor with a poor prognosis. These tumors consist of a heterogeneous population of malignant cells,including well-differentiated tumor cells and less differentiated cells with stem cell properties. These cancer stem cells,known as brain tumor initiating cells,likely contribute to glioma recurrence,as they are highly invasive,mobile,resistant to radiation and chemotherapy,and have the capacity to self-renew. Glioblastoma tumor cells release excitotoxic levels of glutamate,which may be a key process in the death of peritumoral neurons,formation of necrosis,local inflammation,and glioma-related seizures. Moreover,elevated glutamate levels in the tumor may act in paracrine and autocrine manner to activate glutamate receptors on glioblastoma tumor cells,resulting in proliferation and invasion. Using a previously described culturing condition that selectively promotes the growth of brain tumor initiating cells,which express the stem cell markers nestin and SOX-2,we characterize the expression of α-amino-3-hydroxy-5-methyl-4-isozolepropionic acid (AMPA)-type glutamate receptor subunits in brain tumor initiating cells derived from glioblastomas. Here we show for the first time that glioblastoma brain tumor initiating cells express high concentrations of functional calcium-permeable AMPA receptors,compared to the differentiated tumor cultures consisting of non-stem cells. Up-regulated calcium-permeable AMPA receptor expression was confirmed by immunoblotting,immunocytochemistry,and intracellular calcium imaging in response to specific agonists. Our findings raise the possibility that glutamate secretion in the GBM tumor microenvironment may stimulate brain tumor derived cancer stem cells. View Publication

Induced pluripotent stem cells (iPSC) and their differentiated derivatives offer a unique source of human primary cells for toxicity screens. Here,we report on the comparative cytotoxicity of 80 compounds (neurotoxicants,developmental neurotoxicants,and environmental compounds) in iPSC as well as isogenic iPSC-derived neural stem cells (NSC),neurons,and astrocytes. All compounds were tested over a 24-h period at 10 and 100$\$,in duplicate,with cytotoxicity measured using the MTT assay. Of the 80 compounds tested,50 induced significant cytotoxicity in at least one cell type; per cell type,32,38,46,and 41 induced significant cytotoxicity in iPSC,NSC,neurons,and astrocytes,respectively. Four compounds (valinomycin,3,3',5,5'-tetrabromobisphenol,deltamethrin,and triphenyl phosphate) were cytotoxic in all four cell types. Retesting these compounds at 1,10,and 100$\$ using the same exposure protocol yielded consistent results as compared with the primary screen. Using rotenone,we extended the testing to seven additional iPSC lines of both genders; no substantial difference in the extent of cytotoxicity was detected among the cell lines. Finally,the cytotoxicity assay was simplified by measuring luciferase activity using lineage-specific luciferase reporter iPSC lines which were generated from the parental iPSC line. This article is part of a Special Issue entitled SI: PSC and the brain. View Publication

Premature proliferative arrest in benign or early-stage tumors induced by oncoproteins,chromosomal instability,or DNA damage is associated with p53/p21 activation,culminating in either senescence or apoptosis,depending on cell context. Growth hormone (GH) elicits direct peripheral metabolic actions as well as growth effects mediated by insulin-like growth factor 1 (IGF1). Locally produced peripheral tissue GH,in contrast to circulating pituitary-derived endocrine GH,has been proposed to be both proapoptotic and prooncogenic. Pituitary adenomas expressing and secreting GH are invariably benign and exhibit DNA damage and a senescent phenotype. We therefore tested effects of nutlin-induced p53-mediated senescence in rat and human pituitary cells. We show that DNA damage senescence induced by nutlin triggers the p53/p21 senescent pathway,with subsequent marked induction of intracellular pituitary GH in vitro. In contrast,GH is not induced in cells devoid of p53. Furthermore we show that p53 binds specific GH promoter motifs and enhances GH transcription and secretion in senescent pituitary adenoma cells and also in nonpituitary (human breast and colon) cells. In vivo,treatment with nutlin results in up-regulation of both p53 and GH in the pituitary gland,as well as increased GH expression in nonpituitary tissues (lung and liver). Intracrine GH acts in pituitary cells as an apoptosis switch for p53-mediated senescence,likely protecting the pituitary adenoma from progression to malignancy. Unlike in the pituitary,in nonpituitary cells GH exerts antiapoptotic properties. Thus,the results show that GH is a direct p53 transcriptional target and fulfills criteria as a p53 target gene. Induced GH is a readily measurable cell marker for p53-mediated cellular senescence. View Publication