Enrichment of a rare subpopulation of miR-302-expressing glioma cells by serum deprivation

MiR-302-367 is a cluster of polycistronic microRNAs that are exclusively expressed in embryonic stem [ES] cells. The miR-302-367 promoter is functional during embryonic development but is turned off in later stages. Motivated by the cancer stem cell hypothesis, we explored the potential expression of miR-302 in brain tumor cell lines. In the present experimental study, we have tried to expand our knowledge on the expression pattern and functionality of miR302 cluster by quantifying its expression in a series of glioma [A-172, 1321N1, U87MG] and medulloblastoma [DAOY] cell lines. To further assess the functionality of miR-302 in these cell lines, we cloned its promoter core region upstream of the enhanced green fluorescent protein [EGFP] or luciferase encoding genes. Our data demonstrated a very low expression of miR-302 in glioma cell lines, compared with that of embryonal carcinoma cell line NT2 being used as a positive control. The expression of miR-302 promoter-EGFP construct in the aforementioned cell lines demonstrated GFP expression in a rare subpopulation of the cells. Serum deprivation led to the generation of tumorospheres, enrichment of miR-302 positive cells and upregulation of a number of pluripotency genes. Taken together, our data suggest that miR-302 could potentially be used as a novel putative cancer stem cell marker to identify and target cancer stem cells within tumor tissues

Overexpression of BMI1, a polycomb group repressor protein, in bladder tumors: a preliminary report

A polycomb group repressor protein named BMI1 represses the genes that induce cellular senescence and cell death, and it can contribute to cancer when improperly expressed. We aimed to evaluate expression of BMI1 gene in bladder tumors. Tissue specimens containing bladder tumor were evaluated and compared with intact tissues from tumor margins and normal bladder. These were 40 tumor specimens of patients with transitional cell carcinoma of the bladder, 20 tumor-free tissues taken from the margin of the tumors, and 8 specimens from patients without tumor. Specific primers for BMI1 and B2M [as an internal control] were used for reverse transcript polymerase chain reaction technique. The production and distribution of BMI1 protein was also examined by western blotting and immunohistochemistry techniques. Polymerase chain reaction generated a 683-bp product, corresponding to the expect size of BMI1 amplified region. The identity of the amplified fragment was then confirmed by direct DNA sequencing. The mean of expression of BMI1 detected in tumor tissues was significantly higher than that in intact tissues, and there was also a significant association between the mean of gene expression and the stage of malignancy [P<0.001]. The expression of BMI1 at protein level was further confirmed by western blotting and immunohistochemistry. BMI1 is a potent repressor of retinoblastoma and p53 pathways, and hence, elucidating its role in tumorigenesis is very important. We reported for the first time the expression of BMI1 and its correlation with incidence and progress of bladder tumors