We have previously shown high relevance between EC cells and ovar

We have previously shown high relevance between EC cells and ovarian serous carcinoma patient samples at the miRNA level. Pluripotent EC cells can differentiate into cells representing all three germ layers and are con sidered the malignant equivalent of embryonic stem cells. Nullipotent EC cells can avoid differ entiation in vivo to generate poorly differentiated, highly malignant tumors. Comparison of ES cells with pluripotent and nullipotent EC cells can establish mechanisms required for functional malignant differentiation. The cells are so similar that EC cells are used as an easily cultured model of ES biology, reflect ing the difficulty of targeting CSCs without damaging non malignant stem cell populations. In this study we first used gene microarrays to assess upstream regulation of differentiation in murine EC and mES cells.

Our analysis describes aberrant regulation of differentiation in EC cells. Subsequently, we compared mEC genelists to our previously published primary versus recurrent tumor sample data. We described the presence of a cancer stemness p53 p21 regulatory mechanism in ovarian tumor samples. selleckchem Etizolam This mechanism is employed by primary disease and sup pressed in recurrent disease. Subsequently, we con ducted a meta analysis of our previously published human EC and tumor sample miRNA data. We report that cancer stemness signature miRNAs are more relevant to ovarian cancer than cancer stemness signature genes. We detail substantial recruitment of stemness signature miRNAs by recurrent disease. Thus recurrent tumors suppress and activate stemness signa ture genes and miRNAs respectively.

Our analysis indi cates that cancer read more here stemness mechanisms are specifically and differentially regulated in primary and recurrent ovarian malignancy, with obvious implications for treatment. Methods Cell Culture Murine ES and EC cells were pur chased from ATCC, cultured on murine irradiated fibro blasts in DMEM supplemented with 10% foetal bovine serum, 4 mM L glutamine and 100 U ml of penicillin streptomycin and spontaneously differentiated via removal of feeder layer. Human EC cells were retinoic acid differentiated as previously described. Tumor Samples Tumor sample data was previously published. Briefly, two cohorts of primary and recurrent samples were assessed. Cohort 1 contained 5 primary and recur rent serous papillary adenocarcinomas.

Cohort 2 contained 3 paired ovarian cancers from the same patient but with different histologies, papillary serous, mixed mullerian and clear cell carcinomas. Microarray Analysis RNA was isolated using the RNeasy kit as per manufacturers protocol. Digoxi genin UTP labelled cRNA was synthesized via the Chemiluminescent RT IVT Labelling Kit v2. 0 and hybridized to Mouse Genome Survey arrays as per manufacturers instructions.

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