CD147 Is Highly Expressed On Peripheral Blood Neutrophils From Patients With Psoriasis And Induces Neutrophil Chemotaxis

ObjectiveTo investigate the expression of CD147 on peripheral blood neutrophils from patients with psoriasis and healthy volunteers and the effects of CD147siRNA transfection on chemotaxis of neutrophils.To further identify the role of CD147 in the mechanism of psoriasis and the chemotaxis process of neutrophils.To investigate the feasibility of using gene therapy of siRNA technology to treat psoriasis with CD 147 as a new target molecule.Methods1.Peripheral blood samples were obtained from 15 patients with psoriasis,which were confirmed by clinical manifestations and pathology; Normal control samples were taken from 15 healthy sex- and age-matched volunteers;Psoriasis Area and Severity Index(PASI) were applied to evaluate the severity and activity of the disease;Peripheral blood neutrophils were isolated by Ficoll density gradient centrifugation; Wright-Giemsa and trypan blue exclusion staining were performed to appraise their purity and activity,respectively;2.The expression of CD147 on peripheral blood neutrophils from patients with psoriasis and healthy volunteers were examined by Flow-cytometry and western blot,respectively;The non-parametric Spearman Correlation test was used for correlation analysis between the PASI of the patient and the expression of CD147.3.The human promyeloid cell line HL-60 treated by ATRA were iuduced to differentiate;The differentiated HL-60(dHL-60) cells were observed under the light microscope to find out the morphologic changes; Flow-cytometry was used to assay the expression of the differentiation marker CD11b;MTT was applied to detect the proliferation activity;4.dHL-60 cells were transfected with CD147siRNA1 and CD147siRNA2 using the Gene Pulser Electroprotocols;The blank and the non-specific siRNA were acted as controls;RT-PCR and Flow-cytometry were applied to assay the changed mRNA and protein levels of CD147;5.The effect of neutrophil chemotaxis by down-regulated CD147 expression was assayed by transwell system.Results1.Flow cytometry and western blot analysis demonstrated that the expression of CD147 on neutrophils from psoriasis patients were significantly higher than that from the normal controls(P＜0.001);. Moreover,there was a significant positive correlation between PASI and CD147-MFImean(P＜0.05);2.After treated with ATRA for 96 h,the cytoplasm to nucleus ratios of HL-60 cells were decreased and there was a gradual increase in chromatin condensation and nuclear segmentation;flow cytometry showed the expression of CD11b was gradually increased during differentiation (P＜0.001);the proliferation activity of dHL-60 cell was inhibited significantly(P＜0.001);3.CD147siRNA1 and CD147siRNA2 were transfected into dHL-60 cells,respectively;the expression of CD147 was significantly inhibited in dHL-60 cells at 24 h after transfected with CD147siRNA1(P ＜0.001) and restored after 48h(P＞0.05);4.After transfected with CD147siRNA1 for 24 h,dHL-60 cell exhibited a significantly reduced chemotactic response to both fMLP and IL-8(P＜0.001)Conclusion1.The dHL-60 model were reproduced to act as a substitute of neutrophil;A transient transfected method of neutrophils with siRNA was established;2.The expression of CD147 on peripheral blood neutrophils from psoriasis patients were highly increased and that CD147 took an important role in the chemotaxis process of neutrophils.