Characteristics of human CD34(+) cells exposed to ionizing radiation under cytokine-free conditions

To clarify the mechanisms underlying radiation-induced hematopoietic stem cell death, we investigated the effects of excessive ionizing radiation on the clonogenic potential of CD34(+) cells obtained from human umbilical cord blood under cytokine-free conditions. The CD34(+) cells were X-ray–irradiated (up to 2 Gy) and were cultured for 0–48 h under cytokine-free conditions. At various time-points, the CD34(+) cells were investigated for survival, clonogenic potential and the generation of mitochondrial superoxide. At 12 h after X-ray irradiation, the number of viable cells had decreased to ∼70–80% compared with the 0-h non-irradiated control, whereas the clonogenic potential in the X-ray–irradiated cells had decreased to ∼50%–60% compared with the 0-h non-irradiated control. Furthermore, significant generation of mitochondrial superoxide was observed at 6 h, and reached a maximum value between 12 and 24 h after X-ray irradiation. However, no significant differences were observed between non-irradiated and X-ray–irradiated cells in terms of the generation of reactive oxygen species or in the intracellular mitochondrial contents. In addition, a cDNA microarray analysis showed that the majority of the altered genes in the CD34(+) cells at 6 h after X-ray irradiation were apoptosis-related genes. These results suggest the possibility that the elimination of the clonogenic potentials of CD34(+) cells involves the generation of mitochondrial superoxide induced by ionizing radiation.