Medium-mediated intercellular communication is involved in bystander responses of X-ray-irradiated normal human fibroblasts

Abstract

Although radiation-induced bystander effects have been demonstrated in a number of cell types, the studies have largely been performed using high linear energy transfer (LET) radiation, such as a-particles. The literature is contradictory on whether fibroblasts show bystander responses, especially after low LET radiation such as X-or c-rays and whether the same signal transmission pathways are involved. Herein, a novel transwell insert culture dish method is used to show that X-irradiation induces medium-mediated bystander effects in AGO1522 normal human fibroblasts. The frequency of micronuclei formation in unirradiated bystander cells increases from a background of about 6.5% to about 9-13% at all doses from 0.1 to 10 Gy to the irradiated cells. Induction of p21 Waf1 protein and foci of c-H 2 AX in bystander cells is also independent of dose to the irradiated cells above 0.1 Gy. In addition, levels of reactive oxygen species (ROS) were increased persistently in directly irradiated cells up to 60 h after irradiation and in bystander cells for 30 h. Adding Cu-Zn superoxide dismutase (SOD) and catalase to the medium decreases the formation of micronuclei and induction of p21 Waf1 and c-H 2 AX foci in bystander cells, suggesting oxidative metabolism plays a role in the signaling pathways in bystander cells. The results of clonogenic assay of bystander cells showed that survival of bystander cells decreases from 0 to 0.5 Gy, and then is independent of the dose to irradiated cells from 0.5 to 10 Gy. Unlike the response with p21 Waf1 expression, c-H 2 AX foci and micronuclei, adding SOD and catalase has no effect on the survival of bystander cells. The data suggest that irradiated cells release toxic factors other than ROS into the medium.

TNF-TNFR2/p75 Signaling Inhibits Early and Increases Delayed Nontargeted Effects in Bone Marrow-derived Endothelial Progenitor Cells

“…RBR effects are well documented in vitro in variety of cell cultures (8 -11). These responses have been shown by various methodologies, such as media transfer experiments (12,13), co-cultures of irradiated (IR) and nonirradiated (N-IR) cells (14,15), microbeam studies (16), and animal models in vivo (11). It has been proposed that RBR is mediated by an initiating event near the cell surface that activates and integrates numerous intracellular signaling pathways followed by activation of transcription factors and expression of genes that mediate RBR (7).…”

ATR-dependent radiation-induced γH2AX foci in bystander primary human astrocytes and glioma cells

“…Interestingly, only a fraction of bystander cells have elevated gH2AX foci level which was also shown by Sokolov et al (2005) as well as Yang et al (2005) and could suggest a subfraction of cells within a culture that is more vulnerable to factors produced by direct targeted cells. One hypothesis is a correlation with cell cycle distribution where especially S-phase cells could be at increased risk of generating DSB after contact with agents that would otherwise induce predominantly singlestrand breaks like H 2 O 2 .…”

“…Most interestingly, several cytokines can be induced by ROS and NO/NOS (Ayache et al, 2002;Hsu and Wen, 2002;Kosmidou et al, 2002;Hwang et al, 2004;Ryan et al, 2004) providing a possible link between different factors potentially involved in bystander signalling. End points used for the study of bystander effects in vitro have included micronuclei formation (Azzam et al, 2002;Kashino et al, 2004;Shao et al, 2004), gene mutations and genomic instability (Zhou et al, 2000), gene expression changes (Azzam et al, 2002;Yang et al, 2005), transformation (Sawant et al, 2001), proliferation (Gerashchenko and Howell, 2003), cell survival, apoptosis (Belyakov et al, 2002;Lyng et al, 2002), cell cycle arrest (Azzam et al, 2000) and most recently the induction of gH2AX foci in bystander cells (Hu et al, 2005;Sokolov et al, 2005;Yang et al, 2005).…”

A correlation of long term effects and radiation quality in the progeny of bystander cells after microbeam radiations: The experimental study of radiotherapy for cancer risk mitigation

“…However, there is a significant evidence suggesting that similar effects can also be seen in non-irradiated bystander cells, a phenomenon known as the non-targeted effects which include bystander effects, genomic instability and others [1][2][3][4][5][6][7][8][9]. Bystander effect refers to the phenomenon that the induction of biological effects in non-irradiated bystander cells whose nucleus has not been directly traversed by IR [10].…”