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Induction of Apoptosis Over Time in DLD‐1 Colon Cancer Cells with Doxorubicin
Author(s) -
Kondaveeti Sriharsha,
Clack Beatrice
Publication year - 2015
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.29.1_supplement.569.12
Subject(s) - apoptosis , doxorubicin , glyceraldehyde 3 phosphate dehydrogenase , microbiology and biotechnology , gene expression , cancer research , programmed cell death , biology , colorectal cancer , cancer cell , complementary dna , real time polymerase chain reaction , gene , cancer , chemistry , chemotherapy , biochemistry , genetics
Doxorubicin (DOX) is a common chemotherapy drug used to treat a wide variety of cancer types. The drug is known as an intercalating agent that binds to DNA, inhibiting biosynthesis and ultimately resulting in apoptosis, or programmed cell death in treated cells. The exact molecular mechanisms are complex and remain unclear. This project aims to elucidate the sequence of events that occur in DOX treated cancer cells leading up to apoptosis in human colorectal adenocarcinoma (DLD‐1 ATCC ® CCL‐221 ™) . Cells were cultured in complete medium, synchronized through serum starvation, and treated with DOX at 4 hours intervals up to 24 hours. Both untreated and treated cells at each time point then underwent tests to measure cell survival rate and caspase 3/7 activity, which correlates to levels of apoptotic activity. The total RNA from the remaining untreated and treated cells was extracted and reverse transcribed into cDNA. cas 3, 7, 8, 9 and 10 expression levels were determined using real time PCR (RT‐PCR) all of which were normalized to levels of glyceraldehyde 3‐phosphate dehydrogenase (GAPDH) expression. RT² Profiler™ PCR Array plates (SA Biosciences) were used to test the expression levels of 84 apoptosis related gene products throughout the different treatment times. Results show that DOX induces extrinsic pathway due to levels of cas10 and cas8 gene expression being elevated. We present a timeline for the gene expression of 84 apoptosis related genes.