Proteinase 3 expression on the neutrophils of patients with paroxysmal nocturnal hemoglobinuria.

Proteinase 3 (PR3) is released from neutrophils and regulates platelet activity, which is associated with cluster of differentiation (CD)177 antigen (NB1), a glycosylphosphatidylinositol-linked protein. In the present study, the effect of PR3 on thrombosis in paroxysmal nocturnal hemoglobinuria (PNH) and PNH-aplastic anemia (AA) syndrome was explored. The expression of PR3 and NB1 on CD59- neutrophils was detected by flow cytometry, immunofluorescence (IF), reverse transcription-quantitative polymerase chain reaction analysis and western blotting. Serum levels of PR3, proteinase-activated receptor 1 (PAR1) and D-Dimer were measured using ELISAs. The expression of PR3 and NB1 on the plasma membrane of CD59- neutrophils in patients with PNH/PNH-AA was significantly lower compared with their expression on CD59+ neutrophils in patients and controls (P=0.001). However, no correlation between PR3 and NB1 expression was identified. IF staining further demonstrated partially positive PR3 expression on CD59- neutrophils. The serum level of PR3 in patients was identified to be significantly decreased compared with healthy controls (P<0.0001), and significantly negatively correlated with PAR1 (r=-0.456; P=0.043) and D-Dimer (r=-0.503; P=0.028) levels. The mRNA and protein levels of PR3 on PNH clones did not change significantly compared with the control group. In conclusion, PR3 expression on the plasma membrane of neutrophils and in the serum of patients with PNH/PNH-AA decreased, which may result in increased PAR1 expression and increased clotting. The present study provides the basis for further study on platelets in PNH.