Recombinant enolase of Candida albicans could induce the formation of neutrophil extracellular traps

Objective To investigate the effect of enolase of Candida albicans (C.albicans) on human neutrophils. Methods Molecular cloning method was applied to plasmids construcion, cloning, expression and recombinant enolase purification, and density gradient centrifugation was used to isolate the human neutrophils. The effects of recombinant enolase on human neutrophils were observed by DCFH-DA method, SYTOX green and immunofluorescence. The positive control (SC5314 group) was neutrophils stimulated by C.albicans SC5314 (neitrophil∶C.albicans = 5∶1) and the negative control was neutrophils without stimulation. One-Way ANOVA and Tukey′s test were applied for statistical analysis. Results Recombinant enolase, 46 kDa, was successfully cloned and purified. Recombinant enolase could stimulate neutrophils to produce the intracellular ROS, which has no significant difference compared with those induced by C.albicans SC5314 group at 30, 60 and 120 min. Sytox green quantitative and immunoflu-orescence observation of extracellular DNA showed that recombinant enolase also could induce human neutrophils to form NETs. NETs quantitative results showed that there was no significant difference between SC5314 group[ (12.41 ± 2.00) %] and enolase group[ (11.0 ± 2.5) %] at 2 h (F = 6.13, P= 0.0886) , but enolase group[ (22.2 ± 2.0) %) ]was significant lower than SC5314 group [ (31.4 ± 3.1) %] at 4 h (F = 9.745, P = 0.0370) . Conclusion Recombinant enolase of C.albicans could induce human neu-trophils to form NETs. Key words: Enolase; Candida albicans; Neutrophils; Extracellular traps