Abstract
[Objective] Intracellular reactive oxygen species (ROS) generated by X-irradiation cause DNA damage. Transcriptional factor NF-E2-related factor 2 (Nrf2) regulates the expression of multiple antioxidant enzymes, including NAD(P)H-quinone oxidoreductase l (NQO1) and heme oxygenase l (HO-1). Although hematopoietic stem cells (HSCs) are radio-sensitive cells and their intracellular antioxidant system is very important, little is known about those mechanisms. The present study investigated the response of both HO-l and NQO1 in X-irradiated human HSCs. Furthermore, the relationship between the expression of both molecules and the radio-sensitivity of HSCs was also examined.
[Material and Methods] This study was approved by the Committee of Medica1 Ethics of Hirosaki University Graduate School of Medicine and informed consent was obtained from the mothers. Placental/umbilical cord blood was collected after full-term delivery. CD34+ cells (HSCs) were purified using a magnetic cell sorting kit. Total RNA was extracted from the cells. The expression of HO-l and NQO1 mRNA were assayed by quantitative RT-PCR. Moreover, the clonogenic potential was assayed by methylcellulose culture with a combination of cytokines (GM-CSF, G-CSF, EPO, IL-3, SCF). After 14 days of culture, the colonies containing more than 50 cells were counted as colony-forming cells (CFC).
[Results and Discussion] The CFC counts derived from 2 Gy-irradiated HSCs decreased to 80% of the non-irradiated control. At the same time, the expression of HO-l and NQO1 in X-irradiated HSCs showed a significant increase in comparison to the non-irradiated control. Therefore, the present results suggest the possibility that the radio-sensitivity of human HSCs is associated with the expressions of HO-l and NQO1 mRNA. Ongoing experiments are now investigating whether the expressions of HO-1, NQO1 and other response genes are associated with radio-sensitivity and individual differences in HSCs.
