Critical role of neutrophil alkaline phosphatase in the antimicrobial function of neutrophils.
OBJECTIVE
To investigate the roles of neutrophil alkaline phosphatase (NAP) in the migration, reactive oxygen species (ROS) generation and apoptosis of neutrophils.


METHODS
NAP was overexpressed in neutrophil-like differentiated HL-60 cells via transfecting coding sequence of NAP by lentivirus. NAP overexpression in HL-60 cells was confirmed by the methods of quantitative RT-PCR and Western blotting. HL-60 cells were induced to differentiate into neutrophil-like cells by exposure to 1.5% dimethylsulfoxide (DMSO). The migration of neutrophil-like cells were detected by Transwell migration assay. ROS generation of neutrophil-like cells were determined by flow cytometry. Neutrophil-like cells continued to be cultured for 24h, and were then harvested for apoptosis and Western blotting.


RESULTS
After GFP-NAP infection by lentivirus, the expression of NAP was up-regulated in HL-60 cells. HL-60 cells were allowed to differentiate into neutrophil-like cells after 5-day exposure to 1.5% DMSO. Overexpression of NAP in neutrophil-like cells resulted in an increase in the number of migrated cells, intracellular ROS and cell apoptosis followed by a rise in the expression of Caspase 3, Caspase 9 and Bax, while those results were reversed in the NEG and CON group.


CONCLUSIONS
NAP might play a critical role in the anti-microbial function of neutrophils by promoting its migration and ROS generation, as well as accelerating apoptosis in neutrophils.
Center of Research Laboratory, Lianyungang Hospital Affiliated to Xuzhou Medical College, 182 Tongguan Road, Lianyungang 222001, China; Deparment of Clinical Laboratory Diagnostics,Kangda College of Nanjing Medical University, Lianyungang 222000, China.
Haining
H
Li
Center of Research Laboratory, Lianyungang Hospital Affiliated to Xuzhou Medical College, 182 Tongguan Road, Lianyungang 222001, China.
Yao
Y
Zhao
Center of Research Laboratory, Lianyungang Hospital Affiliated to Xuzhou Medical College, 182 Tongguan Road, Lianyungang 222001, China.
Wei
W
Li
Center of Research Laboratory, Lianyungang Hospital Affiliated to Xuzhou Medical College, 182 Tongguan Road, Lianyungang 222001, China.
Jin
J
Yang
Center of Research Laboratory, Lianyungang Hospital Affiliated to Xuzhou Medical College, 182 Tongguan Road, Lianyungang 222001, China. Electronic address: 18961326009@163.com.
Huiyi
H
Wu
D000469:Alkaline Phosphatase / Q000378:metabolism*; D017209:Apoptosis; D002634:Chemotaxis, Leukocyte; D006801:Humans; D009504:Neutrophils / Q000166:cytology / Q000201:enzymology* / Q000276:immunology
D000469:Alkaline Phosphatase
Alkaline phosphatase; Apoptosis; Migration; Neutrophil; Reactive oxygen species