Abstract
IntroductionDiagnostic and therapeutic ionizing radiation (IR) is one of the well known long term risk factors of breast cancer. Extremely lethal consequences of IR causes double-strand breaks, which are mainly responsible for genomic instability, altered gene expression, and cell death.FindingsThis study evaluated the effect of matrix metalloproteinases-2 (MMP-2) gene silencing using MMP-2 shRNA expression plasmids (pMMP-2) on IR induced cytotoxicity and DNA damage by MTT, dead green, γH2AX and comet assays in human normal dermal fibroblasts (HDFs) and MCF-7 human breast cancer cells. IR has decreased the viability of HDFs and MCF-7 cells with increasing IR (2-10Gy). IR induced DNA damage in both HDFs and MCF-7 cells. However, pMMP-2 transfection has increased the viability of irradiated HDFs (10Gy) and significantly decreased the viability of irradiated MCF-7 cells (10Gy). Further, DNA damage in terms of γH2AX foci decreased with pMMP-2 transfection in irradiated HDFs (10Gy) and increased in irradiated MCF-7 cells (10Gy). In addition, MMP-2 gene silencing using pMMP-2 decreased comet tail length in irradiated HDFs but increased in irradiated MCF-7 cells.ConclusionsThe results conclude that pMMP-2 has protected HDFs and sensitized the MCF-7 cells from IR induced DNA damage. This differential response might be due to IR induced MMP-2 distinctive ROS generation in HDFs and MCF-7 cells.
Highlights
Diagnostic and therapeutic ionizing radiation (IR) is one of the well known long term risk factors of breast cancer
The results conclude that Plasmid matrix metalloproteinases-2 (MMP-2) shRNA (pMMP-2) has protected Human normal Dermal Fibroblasts (HDF) and sensitized the MCF-7 cells from IR induced DNA damage
Transfection efficacy of pMMP-2 in HDFs and MCF-7 cells To evaluate the efficacy of Matrix metalloproteinases (MMP)-2 shRNA construct, the monolayer of HDFs and MCF-7 cells were transiently transfected with MMP-2 shRNA for 48 h and expression was analyzed by western blotting analysis
Summary
Diagnostic and therapeutic ionizing radiation (IR) is one of the well known long term risk factors of breast cancer. The high dose of radiation induces early and late skin effects and secondary neoplasm during radiotherapy of breast cancer [3]. Normal or cancer tissue is composed of extracellular matrix (ECM) and cellular constituents mainly fibroblasts. Matrix metalloproteinases-2 (MMP-2) is a member of gelatinase sub-family plays a crucial role in ECM turnover. It degenerates the component of the basement membrane, alters the interstitial collagens I and III, native collagen IV. The aim of the present investigation is to study the effect of MMP-2 gene silencing by transfection of MMP-2 shRNA expression plasmids (pMMP-2on radiation-induced DNA damage in human normal dermal fibroblasts (HDFs) and MCF-7 cells
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