Bystander-induced Apoptosis and Premature Differentiation in Primary Urothelial Explants after Charged Particle Microbeam Irradiation

Abstract

The ureter primary explant technique was developed to study bystander effects under in vivo like conditions where stem and differentiated cells are present. Irradiation was performed with a 3He2+ charged particle microbeam available at the Gray Cancer Institute, with high (approximately 2 microns) precision. Tissue sections from porcine ureters were pre-irradiated with the microbeam at a single location with 10 3He2+ particles (5 MeV; LET 70 keV.micron-1). After irradiation, the tissue section was incubated for 7 days, thus allowing the explant outgrowth to form. Total cellular damage (total fraction of micronucleated and apoptotic cells) was measured according to morphological criteria. Apoptosis was also assessed using a 3'-OH DNA end-labelling technique. Premature differentiation was estimated using antibodies to uroplakin III, a specific marker of terminal urothelial differentiation. Results of our experiments demonstrated a significant bystander-induced differentiation and a less significant increase in apoptotic and micronucleated cells. A hypothesis based on the protective nature of the bystander effect is proposed.

Radiation-induced genomic instability and bystander effects: inter-related nontargeted effects of exposure to ionizing radiation

“…However, the fraction of micronucleated and apoptotic cells did not exceed 1-2% of the total number of the cells within the explant outgrowth and there was evidence for premature terminal differentiation that was interpreted as a protective effect. The potential to produce bystander-induced damage in this system seemed to depend on the proliferative status of the irradiated cell (Belyakov et al, 2002(Belyakov et al, , 2003.…”

Epigenetic changes and nontargeted radiation effects—Is there a link?

“…It can cause changes in gene expression, disruption of mitochondrial processes, cell cycle arrest, and apoptotic cell death [Amundson and Fornace, 2003;Belyakov et al, 2002Belyakov et al, , 2003Belyakov et al, , 2006Criswell et al, 2003;Fei and El-Deiry, …”

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

“…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).…”