Quantitative expression analysis of the apoptosis-related genes BCL2, BAX and BCL2L12 in gastric adenocarcinoma cells following treatment with the anticancer drugs cisplatin, etoposide and taxol

The BCL2 family of proteins includes apoptosis-related molecules involved in normal physiology, as well as cancer pathology. Members of our team have discovered and cloned the novel gene BCL2L12, which codes for a protein member of the BCL2 family. The BCL2L12 expression has been studied extensively in various types of cancer and its important clinical value has been underlined. The main objective of this study is the relative quantification of the mRNA expression of the apoptosis-related genes BCL2, BAX and BCL2L12 in gastric cancer cells, following treatment with anticancer drugs. Gastric adenocarcinoma cells AGS were treated with various concentrations of the chemical substances cisplatin, etoposide and taxol for three time periods. Cell viability was examined by using the MTT assay. Total RNA was extracted and reverse transcribed into cDNA. A highly sensitive, quantitative real-time PCR method was developed based on the SYBR Green chemistry, for the proper mRNA quantification. GAPDH was used as a housekeeping gene. Relative quantification analysis was performed by using the comparative C-T method (2(T)(-DDC)). Treatment of AGS cells with 10 mu M cisplatin, 0.5 mu M etoposide and 10 nM taxol affected the BCL2, BAX and BCL2L12 mRNA levels, compared to the untreated cells. Cisplatin and etoposide induced a major down-regulation in the BCL2 mRNA levels after 72 h of treatment, while the BAX mRNA levels were slightly up-regulated. Moreover, taxol had an up-regulating effect on both BCL2 and BAX transcript levels after 48 h of incubation. Chemotherapy had a much smaller effect on the BCL2L12 expression levels, eventually characterised by a small down-regulation.