Document Type : Research Paper
Department of Stem Cells and Regenerative Medicine, Institute of Medical Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran
Background: Tumor cells proliferation and apoptosis inhibition are the mechanisms through which the Colorectal Cancer (CRC) progression, metastasis and chemoresistance are promoted pathologically, offering clinical advantages for characterizing their molecular regulators.
Objectives: In this study, to unravel the role of PIWIL2 as a potential CRC oncogenic regulator, we examined the effect of its overexpression on proliferation, apoptosis and colony formation of SW480 colon cancer cell line.
Material and Methods: Established SW480-P (overexpression of PIWIL2) and SW480-control (SW480-empty vector) cell lines were cultured in DMEM containing 10% FBS with 1% penicillin-streptomycin. The total DNA and RNA was extracted for further experiments. Real-Time PCR and western blotting assay were performed to measure the differential expression of proliferation associated genes including the expression of cell cycle and anti-apoptotic genes as well as Ki-67 and PIWIL2 in both cell lines. Cell proliferation was determined using MTT assay, doubling time assay and the colony formation rate of transfected cells was measured with the 2D colony formation assay.
Results: At the molecular level, PIWIL2 overexpression was associated with significant up-regulation of cyclin D1, STAT3, BCL2-L1, BCL2-L2 and Ki-67 genes. MTT and doubling time assay showed that PIWIL2 expression induced time-related effects on proliferation rate of SW480 cells. Moreover, SW480-P cells had markedly greater capacity to form colonies.
Conclusions: PIWIL2 plays important roles to promote cancer cell proliferation and colonization via the cell cycle acceleration and inhibition of apoptosis, the mechanisms through which this gene seems to contribute to CRC development, metastasis and chemoresistance, hence potentially highlighting PIWIL2 targeted therapy as a valuable tool for CRC treatment.