Diphenylene iodonium and apocynin reduce the translocation and level of p47phox in PBMCs of premature infants to inhibit reactive oxygen species production
ZHANG Lingping;DONG Wenbin;LI Qingping;KANG Lan;ZHANG Lianyu;LU Youying;ZHAI Xuesong;The First Affiliated Hospital of Sichuan Medical University;
Objective To observe the effects of NADPH oxidase inhibitor diphenylene iodonium( DPI) and apocynin on the generation of reactive oxygen species( ROS) induced by p47 phox and the mechanism of p47phox-induced ROS production under hyperoxic conditions. Methods Peripheral blood mononuclear cells( PBMCs) were isolated from the peripheral blood( 2 m L) of premature infants of less than 32 weeks without oxygen uptake. The isolated cells were divided into four groups,control group,hyperoxia group,hyperoxia and DPI group,hyperoxia and apocynin group. The control group was cultured in incubator with 50 m L / L CO2 at 37℃,and the other groups were cultured in 950 m L / L O2 and 50 m L / L CO2 mixed gas. After 48 hours,ROS was detected by Mitosox Red staining under a confocal laser scanning microscope;malondialdehyde( MDA) was measured by thiobarbituric acid colorimetry; the location and translocation rate of p47 phox was observed by immunofluorescence staining; the level of p47 phox protein was tested by Western blotting. Results Compared with the hyperoxia group,the remaining three groups showed significantly decreased ROS and MDA levels and reduced translocation rate and level of p47 phox. Compared with the control group,both the hyperoxia and DPI group and the hyperoxia and apocynin group were not significantly different in the above indexes. Conclusion DPI and apocynin can reduce hyperoxia-induced ROS production by decreasing the translocation and level of p47 phox.