Abstract
Medulloblastoma (MB) is the most common malignant pediatric brain tumor. Despite the progress of new treatments, the risk of recurrence, morbidity, and death remains significant and the long-term adverse effects in survivors are substantial. The fraction of cancer stem-like cells (CSCs) because of their self-renewal ability and multi-lineage differentiation potential is critical for tumor initiation, growth, and resistance to therapies. For the development of new CSC-targeted therapies, further in-depth studies are needed using enriched and stable MB-CSCs populations. This work, aimed at identifying the amount of CSCs in three available human cell lines (DAOY, D341, and D283), describes different approaches based on the expression of stemness markers. First, we explored potential differences in gene and protein expression patterns of specific stem cell markers. Then, in order to identify and discriminate undifferentiated from differentiated cells, MB cells were characterized using a physical characterization method based on a high-frequency dielectrophoresis approach. Finally, we compared their tumorigenic potential in vivo, through engrafting in nude mice. Concordantly, our findings identified the D283 human cell line as an ideal model of CSCs, providing important evidence on the use of a commercial human MB cell line for the development of new strategic CSC-targeting therapies.
Highlights
Around 300,000 children and adolescents worldwide are diagnosed with cancer each year [1,2,3,4].Among pediatric cancers, Medulloblastoma (MB) is the most common intracranial primitive neuroectodermal malignancy, with an incidence of 6 children per million under 9 years of age and less than 2 cases out of a million among 15–19 years of age [5,6]
Our results showed a significantly higher level of CD133 gene expression in D283 compared with D341 cells and its almost complete lack in the DAOY cell line (Figure 1A)
For an in-depth analysis of their stemness features, MB cell lines were stained for various markers of stemness/differentiation
Summary
Around 300,000 children and adolescents worldwide are diagnosed with cancer each year [1,2,3,4]. It has been hypothesized that only a small subset of tumor cells can initiate and support tumor growth These rare stem cells, called cancer stem cells (CSCs), possess tumorigenic ability with marked capacity for proliferation, self-renewal, and differentiation potential [16,17,18]. Given the relatively MB low incidence, the possibility of accessing patient-derived samples is extremely limited and few MB cell lines are available in central repositories making it more complex to study this tumor compared to others [24]. CSCs enrichment often requires particular culture media and presents several difficulties in the experimental protocols, resulting in a time-consuming process and suggesting the need for a better characterization of available cell lines. Since physical characterization was recently reported as complementary to cell biological features, to identify CSCs, we used high-frequency dielectrophoresis (HF-DEP) crossover frequency, a label-free, accurate, fast, and low-cost diagnostic technique that exploits the polarization and consequent motion of bio-particles in an applied electric fields [31]
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