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.