Effect of 5-FU and MTX on the Expression of Drug-resistance Related Cancer Stem Cell Markers in Non-small Cell Lung Cancer Cells

Cancer stem cells (CSCs) are often characterized by the elevated expression of drug-resistance related stem-cell surface markers, such as CD133 and ABCG2. Recently, we reported that CSCs have a high level of expression of the IL-6 receptor (IL-6R). The purpose of this study was to investigate the effect of anticancer drugs on the expression of the drug resistance-related cancer stem cell markers, ABCG2, IL-6R, and CD133 in non-small cell lung cancer (NSCLC) cell lines. A549, H460, and H23 NSCLC cell lines were treated with the anticancer drugs 5-fluorouracil (5-FU; 25 microg/ml) and methotrexate (MTX; 50 microg/ml), and the expression of putative CSC markers was analyzed by fluorescent activated cell sorter (FACS) and the gene expression level of abcg2, il-6r and cd133 by reverse transcriptasepolymerase chain reaction (RT-PCR). We found that the fraction of ABCG2-positive(+) cells was significantly increased by treatment with both 5-FU and MTX in NSCLC cells, and the elevation of abcg2, il-6r and cd133 expressions in response to these drugs was also confirmed using RT-PCR. Also, the number of IL-6R(+) cells was increased by MTX in the 3 cell lines mentioned and increased by 5-FU in the H460 cell line. The number of CD133(+) cells was also significantly increased by both 5-FU and MTX treatment in all of the cell lines tested. These results indicate that 5-FU and MTX considerably enhance the expression of drug-resistance related CSC markers in NSCLC cell lines. Thus, we suggest that antimetabolite cancer drugs, such as 5-FU and MTX, can lead to the propagation of CSCs through altering the expression of CSC markers.

Determination of Oxyclozanide in Beef and Milk using High-Performance Liquid Chromatography System with UV Detector / 한국실험동물학회지

This study was developed and validated for the determination of oxyclozanide residue concentrations in beef and commercial milk, using high-performance liquid chromatography system. Oxyclozanide was successfully separated on a reverse phase column (Xbridge-C18, 4.6x250 mm, 5 microm) with a mobile phase composed of acetonitrile and 0.1% phosphoric acid (60:40, v/v%). This analytical procedure involved a deproteinization process using acetonitrile for beef and 2% formic acid in acetonitrile for commercial milk, dehydration by adding sodium sulfate to the liquid analytical sample, and a defatting process using n-hexane; after these steps, the extract was exposed to a stream of nitrogen dryness. The final extracted sample was dissolved in the mobile phase and filtered using a 0.45 microm syringe filter. This method had good selectivity and recovery (70.70+/-7.90-110.79+/-14.95%) from the matrices. The LOQs ranged from 9.7 to 9.8 microg/kg for beef and commercial milk. The recoveries met the standards set by the CODEX guideline.