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Effect of Dishevelled 2 on the secretion of vascular endothelial growth factor in rheumatoid arthritis fibroblast-like synoviocytes

Objective 
To optimize the culture method for rheumatoid arthritis fibroblast-like synoviocytes in vitro, and observe the effect of Dishevelled (Dvl) 2 on vascular endothelial growth factor (VEGF) in rheumatoid arthritis fibroblast-like synoviocytes (RA-FLS). 
 
 
Methods 
Synovium from RA patients who underwent knee arthroplasties were cut into small piece, and RA-FLS were isolated and cultured in vitro using tissue block method. Dvl 2 lentivirus overexpressing plasmid was constructed and transfected into RA-FLS. Q-polymerase chain reaction (PCR) and enzyme linked immunosorbent assay (ELISA) were used to detect the mRNA and protein expression levels of VEGF. Then we used 10 ng/ml tumor necrosis factor(TNF)-α recombinant protein to stimulate the transfected RA-FLS. 24 h after stimulation, mRNA and protein expression of VEGF were detected again. Student's t test was used for two group analyses. 
 
 
Results 
RA-FLS was successfully isolated and cultured in vitro. The multiplicity of infection was 30 and was in conjunction with appropriate concentration of polybrene to promote transfection. Transfection efficiency could meet the test requirements. The mRNA of Dvl 2 increased for 79-fold than the control group. Compared with the control group, Dvl 2 could mildly inhibit RA-FLS secretion of VEGF. After TNF-α stimulation, Dvl 2 could significantly inhibit the VEGF's mRNA (2.15±0.10, 2.92±0.47 fold, t=-3.924, P=0.003) and protein [(285±100) pg/ml, (155±61) pg/ml, t=-2.714, P=0.022] expression compared with the control group. 
 
 
Conclusion 
Dvl 2 can inhibit the effect of TNF-α induced secretion of VEGF in RA-FLS. The specific mechanism needs further study. 
 
 
Key words: 
Arthritis, rheumatoid; Wnt signaling pathway; Synoviocytes; Vascular endothelial growth factor; Dishevelled 2