Dataset Information

Phenotypic characterization of human colorectal cancer stem cells.

ABSTRACT: Recent observations indicate that, in several types of human cancer, only a phenotypic subset of cancer cells within each tumor is capable of initiating tumor growth. This functional subset of cancer cells is operationally defined as the "cancer stem cell" (CSC) subset. Here we developed a CSC model for the study of human colorectal cancer (CRC). Solid CRC tissues, either primary tissues collected from surgical specimens or xenografts established in nonobese diabetic/severe combined immunodeficient (NOD/SCID) mice, were disaggregated into single-cell suspensions and analyzed by flow cytometry. Surface markers that displayed intratumor heterogeneous expression among epithelial cancer cells were selected for cell sorting and tumorigenicity experiments. Individual phenotypic cancer cell subsets were purified, and their tumor-initiating properties were investigated by injection in NOD/SCID mice. Our observations indicate that, in six of six human CRC tested, the ability to engraft in vivo in immunodeficient mice was restricted to a minority subpopulation of epithelial cell adhesion molecule (EpCAM)(high)/CD44+ epithelial cells. Tumors originated from EpCAM(high)/CD44+ cells maintained a differentiated phenotype and reproduced the full morphologic and phenotypic heterogeneity of their parental lesions. Analysis of the surface molecule repertoire of EpCAM(high)/CD44+ cells led to the identification of CD166 as an additional differentially expressed marker, useful for CSC isolation in three of three CRC tested. These results validate the stem cell working model in human CRC and provide a highly robust surface marker profile for CRC stem cell isolation.

SUBMITTER: Dalerba P

PROVIDER: S-EPMC1891215 | biostudies-literature | 2007 Jun

REPOSITORIES: biostudies-literature

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Phenotypic characterization of human colorectal cancer stem cells.

Dalerba Piero P Dylla Scott J SJ Park In-Kyung IK Liu Rui R Wang Xinhao X Cho Robert W RW Hoey Timothy T Gurney Austin A Huang Emina H EH Simeone Diane M DM Shelton Andrew A AA Parmiani Giorgio G Castelli Chiara C Clarke Michael F MF

Proceedings of the National Academy of Sciences of the United States of America 20070604 24

Recent observations indicate that, in several types of human cancer, only a phenotypic subset of cancer cells within each tumor is capable of initiating tumor growth. This functional subset of cancer cells is operationally defined as the "cancer stem cell" (CSC) subset. Here we developed a CSC model for the study of human colorectal cancer (CRC). Solid CRC tissues, either primary tissues collected from surgical specimens or xenografts established in nonobese diabetic/severe combined immunodefici ...[more]

PMID: 17548814

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Project description:BackgroundCancer stem cells exhibit close resemblance to normal stem cells in phenotype as well as function. Hence, studying normal stem cell behavior is important in understanding cancer pathogenesis. It has recently been shown that human breast stem cells can be enriched in suspension cultures as mammospheres. However, little is known about the behavior of these cells in long-term cultures. Since extensive self-renewal potential is the hallmark of stem cells, we undertook a detailed phenotypic and functional characterization of human mammospheres over long-term passages.MethodologySingle cell suspensions derived from human breast 'organoids' were seeded in ultra low attachment plates in serum free media. Resulting primary mammospheres after a week (termed T1 mammospheres) were subjected to passaging every 7th day leading to the generation of T2, T3, and T4 mammospheres.Principal findingsWe show that primary mammospheres contain a distinct side-population (SP) that displays a CD24(low)/CD44(low) phenotype, but fails to generate mammospheres. Instead, the mammosphere-initiating potential rests within the CD44(high)/CD24(low) cells, in keeping with the phenotype of breast cancer-initiating cells. In serial sphere formation assays we find that even though primary (T1) mammospheres show telomerase activity and fourth passage T4 spheres contain label-retaining cells, they fail to initiate new mammospheres beyond T5. With increasing passages, mammospheres showed an increase in smaller sized spheres, reduction in proliferation potential and sphere forming efficiency, and increased differentiation towards the myoepithelial lineage. Significantly, staining for senescence-associated beta-galactosidase activity revealed a dramatic increase in the number of senescent cells with passage, which might in part explain the inability to continuously generate mammospheres in culture.ConclusionsThus, the self-renewal potential of human breast stem cells is exhausted within five in vitro passages of mammospheres, suggesting the need for further improvisation in culture conditions for their long-term maintenance.

Project description:Stimulating meniscus regeneration using meniscal progenitor cells has been suggested as a promising new strategy. However, there is a lack of studies which decisively identify and characterize progenitor cell populations in human meniscus tissues. In this study, donor-matched progenitor cells were isolated via selective fibronectin adhesion from the avascular and vascular regions of the meniscus and chondroprogenitors from articular cartilage (n = 5). The mixed populations of cells from these regions were obtained by standard isolation techniques for comparison. The colony formation efficacy of avascular progenitors, vascular progenitors and chondroprogenitors was monitored using Cell-IQ® live cell imaging. Proliferation rates of progenitors were compared with their mixed population counterparts. Cell surface markers indicative of mesenchymal stromal cells profile and progenitor markers were characterized by flow cytometry in all populations. The fibrochondrogenic capacity was assessed via fibrochondrogenic differentiation and measuring GAG/DNA content and morphology. All meniscal progenitor and chondroprogenitor populations showed superior colony forming efficacy and faster proliferation rates compare to their mixed populations. Progenitor populations showed significantly higher positivity for CD49b and CD49c compared to their mixed population counterparts and chondroprogenitors had a higher positivity level of CD166 compared to mixed chondrocytes. GAG/DNA analysis demonstrated that progenitor cells generally produced more GAG than mixed populations. Our study demonstrates that the human meniscus contains meniscal progenitor populations in both the avascular and vascular regions. Meniscal progenitors derived from the vascular region exhibit enhanced proliferative and fibrochondrogenic characteristics compared to those from the avascular region; this may associate with the enhanced meniscal healing potential in the vascular region. These findings build on the body of evidence which suggests that meniscal progenitors represent an attractive cell therapy strategy for meniscal regeneration.

Dataset Information

Phenotypic characterization of human colorectal cancer stem cells.

Publications

Phenotypic characterization of human colorectal cancer stem cells.

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