Bystander effects and compartmental stress response to X-ray irradiation in L929 cells

Abstract

Bystander effects are indirect consequences of radiation and many other stress factors. They occur in cells that are not directly exposed to these factors, but receive signals from affected cells either by gap junctions or by molecules released in the medium. Characterizing these effects and deciphering the underlying mechanisms involved in radiation-induced bystander effects are relevant for cancer radiotherapy and radioprotection. At doses of X-ray radiation 0.5 and 1 Gy, we detected bystander effects as increased numbers of micronuclei shortly after the treatment, through medium transfer and by co-cultures. Interestingly, bystander cells did not exhibit long-term adverse changes in viability. Evaluation of several compartmental stress markers (CHOP, BiP, mtHsp60, cytHsp70) by qRT-PCR did not reveal expression changes at transcriptional level. We investigated the involvement of ROS and NO in this process by addition of specific scavengers of these molecules, DMSO or c-PTIO in the transferred medium. This approach proved that ROS but not NO is involved in the induction of lesions in the acceptor cells. These results indicate that L929 cells are susceptible to stress effects of radiation-induced bystander signaling.

Intracellular and Intercellular Signalling Mechanisms following DNA Damage Are Modulated By PINK1

“…Cells were directly exposed to 0–3 Gy of X-ray generated by a medical linear accelerator (Primus Mevatron 2D, 6 MV, Siemens, Germany), at a dose rate of 1.85 Gy/min [ 23 ], or treated with bleomycin sulfate (BLM, Sigma-Aldrich, USA) for 1 hour at 5–40 μ g/mL followed by 3 washes with culture medium [ 22 ].…”

“…Cytochalasin B was added 20 h before analysis at 3 μ g/mL to MEF cells and 6 μ g/mL to SH-SY5Y. For staining, cells were washed in PBS, fixed in acetic acid : methanol (1 : 9), and stained with Acridine Orange at 10 μ g/mL for 10 minutes followed by washing in PBS and mounting [ 23 ]. Representative images for the conditions employed in this study are presented in Supplementary Figure S3 .…”

“…The messenger molecules include ROS, nitric oxide, and cytokines as well as DNA and RNA molecules. These nontreated cells respond to signals produced by directly treated cells through a signalling process that has been termed the bystander effect (BE) [ 19 – 23 ]. The endpoints that demonstrate a BE address similar DNA damage and cell survival-cell death as in the case of AR.…”