Browsing by Subject "serum starvation"

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Open Access
Analysis of CHRNA5 expression in breast cancer cell lines in response to serum starvation and estrogen treatment
(2013) Açıkgöz, Azer Aylin
Breast cancer is a complex disease that can be classified into distinct molecular subtypes including Basal, Luminal A, Luminal B and HER2 positive. These molecular subtypes mainly differ in their hormone receptor expression and response to treatment. This makes the discovery of new molecular markers for further classification important. Cholinergic nicotinic acetylcholine receptors are ion channels involved in smoking behavior, neurodegenerative diseases and cancer. Cholinergic nicotinic receptor alpha 5 (CHRNA5) has been associated with nicotine addiction and recently with lung cancer yet its importance in breast cancer remains relatively unexplored. In the present study, a panel of 10 breast cancer cell lines were used for quantification of isoform-specific CHRNA5 expression using qPCR. Changes in CHRNA5 expression in response to serum starvation and estrogen treatment were assessed. qPCR showed that CHRNA5 was alternatively spliced, with at least five different isoforms in breast cancer cell lines. qPCR analysis for CHRNA5 expression in serum treated and serum starved cells were analyzed after outlier detection and exclusion; and statistical tests included ANCOVA using geometric mean of TPT1 and SDHA, as reference genes. Our results demonstrated that, CHRNA5 expression differed between different subtypes of breast cancer cell lines. CHRNA5 expression significantly responded to serum starvation in ZR75-1 and MDA-MB-157 cell lines, isoform specifically. Isoform expression of CHRNA5 exhibited significant alterations upon estrogen treatment in a dose and time-dependent manner. Expression of 1000bp variant, isoform2 and isoform3 of CHRNA5 significantly increased upon E2 treatment and total CHRNA5 and isoform2 CHRNA5 increased in expression at 24 hours when compared with 12 hours of treatment. Our findings show that CHRNA5 has multiple isoforms in breast cancer, with potential to be modulated by serum starvation and estrogen treatment in a cell-specific manner.
Open Access
Investigation of the effects of nicotine on the expression profile of SW620 colon adenocarcinoma cells using a functional genomics approach
(2009) Kaya, Onur
Colon cancer is the third most common form of cancer with approximately 655,000 deaths worldwide annually and the second principal cause of cancer-related death in the Western world. Studies focusing on genomic instability and cell culture in recent years have shown that there is a statistically significant link between tobacco smoking and colorectal cancer. Although nicotine is one of the most potent chemical in tobacco, it was not studied extensively in colorectal cancers. Nicotine works as an agonist of nicotinic acetylcholine receptors and modulates the intracellular calcium concentrations hence deregulating multiple signal transduction pathways (e.g., PI3K/AKT, MAPK, mTOR). It has been shown that nicotine accelerates cell proliferation while it increases cell migration, metastasis and angiogenesis, and inhibits apoptosis in lung and gastric cancers. The aim of this study was to give more insight into the association between nicotine and colon cancer by investigating the gene expression profiles of SW620 colon adenocarcinoma cells under 48h 1µM nicotine treatment at different serum levels to reflect molecular response to growth factor-induced and –depleted conditions (10% FBS or 0.1% FBS). We used multiple approaches including cell culture techniques, microarray technology, and gene-network analysis to assess the effects of nicotine on cell proliferation and transcriptome profile. Furthermore, the selected genes that are involved in cell cycle and apoptosis were used to confirm and evaluate the transcriptome analysis results with real time qRT-PCR and Western Blot techniques. In this project, our findings indicated that serum starvation of SW620 colon adenocarcinoma cell line resulted in decreased cell proliferation, which could be rescued by 1µM nicotine via deregulation of multiple pathways including cell cycle, apoptosis, Ca2+ signaling, and ribosomal protein expression. This study implicated that nicotine-, thus acetylcholine-mediated signaling may have an important role in tumor development and metastasis.