Silibinin-Induced Apoptosis and Downregulation of MicroRNA-21 and MicroRNA-155 in MCF-7 Human Breast Cancer Cells / 한국유방암학회지

PURPOSE: MicroRNAs (miRNAs) have received much attention owing to their aberrant expression in various stages of cancer. In many biological processes, miRNAs negatively regulate gene expression, and may be useful in therapeutic strategies. The present study evaluated the effects of silibinin (silybin), a natural flavonoid, on miRNA expression and attempted to elucidate therapeutic targets in MCF-7 breast cancer cells. METHODS: The rates of cell proliferation and apoptosis were determined in silibinin-treated and untreated MCF-7 cells. Furthermore, the expression levels of miR-21 and miR-155 were measured in MCF-7 cells after incubation with silibinin (100 µg/mL), and the putative targets of the miRNAs within the apoptotic pathways were predicted using bioinformatic approaches. The expression levels of some of these targets were evaluated by quantitative reverse transcription polymerase chain reaction (qRT-PCR). RESULTS: Silibinin induced apoptosis in MCF-7 cells in a dose- and time-dependent manner. qRT-PCR analysis revealed a decrease in miR-21 and miR-155 expression levels in silibinin-treated cells relative to the levels in the untreated cells. Potential miR-21 and miR-155 targets within the apoptotic pathways, such as CASP-9, BID, APAF-1, CASP-3, CASP-8, and PDCD4, were predicted by in silico analysis. qRT-PCR analysis showed upregulation of some of these potential targets including caspase-9 (CASP-9) and BID after silibinin treatment for 48 hours. CONCLUSION: Our results suggest a correlation between the expression of miR-21 and miR-155, and MCF-7 cell proliferation. The antiproliferative activity of silibinin may partly be attributable to the downregulation of miR-21 and miR-155, and the upregulation of their apoptotic targets. Furthermore, the upregulation of CASP-9 and BID indicates that silibinin induces apoptosis through both the extrinsic and intrinsic pathways.

Comparing the effect of silybin and silybin advanced[TM] on viability and HER2 expression on the human breast cancer SKBR3 cell line by no serum starvation

The polyphenol silybin has anti-oxidant and anti-cancer properties. The poor bioavailability of some polyphenols [flavonoids, and terpenoids] can be improved by binding them to phosphatidylcholine [phytosome technology]. Many studies have focused on the most common phytosome, silybin-phosphatidylcholine, particularly for its hepatoprotective effects. However, in recent years, studies have also been conducted to determine its anti-cancer effect. Considering that the serum starvation should not be used for studies that are not focused on cell cycle arrest, we studied the effect of silybin-phosphatidylcholine from Silybin Advanced[TM] in 1:2 ratio [one part silybin bound to two parts phosphatidylcholine] on HER2 gene expression on the SKBR3 breast cancer cell line which were cultured in complete medium [not serum deprivation]. The results were compared with our previous study of silybin on HER2 expression on SKBR3 cells. An MTT test was used to determine concentrations for cell treatment, and the gene expression was defined by real-time RT-PCR. Outcomes showed significant concentration- and time-dependent cell growth inhibitory effects of silybin, and silybin-phosphatidylcholine and HER2 down regulation on SKBR3 cells. Silybin-phosphatidylcholine concentrations had a much larger inhibitory and HER2 down regulate effect on cell growth than the same silybin concentrations on SKBR3 cells

comparison of the effects of silybin and silybin-phosphatidylcholine on viability and ESR expression in human breast cancer T47D cell line

Silybin is a polyphenol with anti-oxidant and anti-cancer properties. The poor bioavailability of some polyphenols can be improved by binding to phosphatidylcholine. In recent years, studies have been conducted to evaluate the anti-cancer effect of silybin. We studied the effect of silybin and silybin-phosphatidylcholine on ESR1 and ESR2 gene expression and viability in the T47D breast cancer cell line. In this experimental study, a 3-[4,5-Dimethylthiazol-2-Yl]-2,5- Diphenyltetrazolium Bromide test [MTT test] was used to determine doses for cell treatment, and the gene expression was analyzed by real-time reverse transcriptase-polymerase chain reaction [real-time RT- PCR]. Significant dose- and time-dependent cell growth inhibitory effects of silybin and silybin-phosphatidylcholine along with ESR1 down-regulation were observed in T47D cells. In contrast to ESR1, the T47D cell line showed negligible ESR2 expression. This study suggests that silybin and silybin-phosphatidylcholine down-regulate ESR1 in ER+ breast cancers. Results also show that in the T47D cell line, silybinphosphatidylcholine has a much higher growth inhibitory effect and a more significant down-regulation of ESR1 compared with silybin

[Up regulation of CD82 gene in prostate cancer PC-3 cell line treated with silibinin]

Prostate cancer is one of the most common cancer in the developed countries. Most of cancer deaths are due to development of metastasis. Hence, prevention of metastasis is critical. Silibinin is a flavonoid component that inhibits cell proliferation and causes cell death of human prostate cancer. In this study, the expression of CD82 gene in PC-3 cells treated with escalating concentrations of silibinin was evaluated which can result in new view for prostate cancer therapy. In this study, PC-3 cells were treated with different concentrations of silibinin for 24h. The LD50 was determined. RNA was extracted by trizol, then cDNA was synthesized. Precise primers were designed for CD82 and GAPDH genes by specific software. Quantity of CD82 gene expression compare to GAPDH gene in different concentrations of silibilin was analyzed using very sensitive quantitative Real-time PCR. CD82 gene expression in PC-3 cells treated with 100, 150 and 200?g/ml of silibinin at 24h was increased by 1.97 +/- 0.26 [P<0.05], 3.00 +/- 0.26 and 3.43 +/- 0.43 [P<0.01], respectively. The results of quantitative Real-time PCR indicated that silibinin can probably decrease metastasis, by up-regulation of CD82 metastasis suppressor gene in PC-3 cells

[Serial analysis of gene expression and its applications]

Serial analysis of gene expression or SAGE is a powerful technique that can be used for global analysis of gene expression. The result of a SAGE experiment is reported as a SAGE library, which contains the counts of various short segments [tags] of the cDNA clones. Unlike many other techniques SAGE does not require prior knowledge of the genes of interest and provides qualitative and quantitative data of every transcribed sequence in a particular cell or tissue type. The SAGE method is based on the isolation of unique sequence tags from individual mRNAs and serial concatenation of tags into long DNA molecules for lump-sum sequencing. SAGE has been widely applied to analyzing gene expression in many biological and medical studies. Some variations of the original SAGE were derived to improve the utility of SAGE in certain conditions for example micro SAGE, mini SAGE, long SAGE, super SAGE, etc. In this review, we present an outline of the original method, the basic principle of SAGE, compare SAGE with microarray, some of the modified SAGE, discusses the advantages and limitations of SAGE as well as examples of applications