ABSTRACT
Cyclin D1 is an important cell-cycle regulator that drives the cell cycle from the G1 phase to the S phase. Elevated nuclear cyclin D1 expression has been found in human tumors, including thyroid carcinoma. Protein production is known to require DNA amplification in each cell, but reports of such amplification have not been published. This study aimed to analyze the relationship between cyclin D1 protein production and chromosome 11 in cultured cells by means of dual staining with fluorescence in situ hybridaization (FISH) and immunostaining. In addition, we immunostained anaplastic thyroid carcinoma tissue. The results indicate that cyclin D1 is not related to chromosome 11 in cultured cells. Furthermore, tissue study showed that cyclin D1 is produced in the cytoplasm and in nuclei in various ratios.
Subject(s)
Carcinoma/genetics , Chromosomes, Human, Pair 11 , Cyclin D1/metabolism , Gene Expression Regulation, Neoplastic , Immunohistochemistry/methods , In Situ Hybridization, Fluorescence , Staining and Labeling/methods , Thyroid Neoplasms/genetics , Carcinoma/metabolism , Carcinoma/pathology , Cell Cycle , G1 Phase , Humans , S Phase , Thyroid Neoplasms/metabolism , Thyroid Neoplasms/pathology , Tumor Cells, CulturedABSTRACT
Various methods are available for the assessment of cell viability. Recently, interest has centered on methods using fluorescent dyes. In this work, we used a double-staining assay, involving the use of rhodamin 123, which stains the mitochondria of viable cells, and propidium iodide, which stains the nuclei of dead cells, to investigate their use in assessing the viability of cells. We added adriamycin to NIH 3T3 cells and double-stained the cells that adhered to the dish and those that were suspended in the culture solution, and observed the results over time. We found that nearly all the adherent cells were stained with rhodamin 123 alone. However, the suspended cells in the control group accounted for most of the double-stained cells, and when adriamycin was added, most were stained with PI alone.
Subject(s)
3T3 Cells/physiology , Cell Survival , Fluorescence , Staining and Labeling , 3T3 Cells/cytology , Animals , Coloring Agents , Doxorubicin/pharmacology , Evaluation Studies as Topic , Fluoresceins , Mice , Microscopy, Fluorescence , Rhodamine 123ABSTRACT
The acquisition of drug-resistance is a major problem for cancer patients undergoing chemotherapy. To clarify genetic alterations in cancer cells that develop drug-resistance, comparative genomic hybridization (CGH) was applied to esophageal squamous cell carcinoma cell lines (SH-1V1, SH-1V2, SH-1V4 and SH-1V8) and chemoresistance-related genes in altered chromosomal regions were evaluated. These cell lines were derived from the parental SH-1 cell line, after multiple steps of selection by an increasing exposure to vindesine. SH-1V8 cells were strongly resistant to vindesine. DNA copy number at 16p which includes the MRP (multidrug resistance related protein) gene was markedly increased in all cell lines examined. Increased DNA copy numbers were found at the regions of 5q31-32, 10q11.1-23, and 14q32-qter in SH-1V8 cells that acquired resistance to other drugs as well. Both SH-1V4 and SH-1V8 showed increased DNA copy numbers at 7q11.1-22, 16q12.1-qter, 19p13.2-13.3, 19q11-13.2 and 20q13.1-qter. The chromosomal region of 7q11.1-22 including MDR-1 (multidrug resistance-1) gene was highly amplified in SH-1V4 and SH-1V8. Amplification of the MRP region suggests the prerequisite of developing resistance to vindesine, and further amplification of MDR-1 may play a critical role in acquiring drug-resistance. Several unknown genes related to the induction of chemoresistance might be concealed in other altered chromosomal regions.