AP-009

To study the effects of CD147 on neutrophil on matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) production by fibroblast-like synoviocytes (FLS) and the invasive potential of FLS in rheumatoid arthritis(RA).

FLS were isolated from the synovial tissues which were resected from the patients with rheumatoid arthritis (RA) and ostarthritis (OA) during synovectomy, and CD14, CD90 on RA FLS were detected by FCM. HL-60 cells were differentiated into mature neutrophil by alltransretinoic acid (ATRA) and the degree of cell differentiation was detected through NBT reduction reaction. CD147 expression on the differentiated and undifferentiated HL-60 cells and FLS were detected by FCM. The release and activity of MMP-2 and MMP-9 were detected in the supernantents of HL-60 cells cocultured with RA FLS by Gelatin zymography. The invasive potential of RA FLS was detected by invasion assay. To further investigate the effect of CD147 on neutrophil on MMP production by FLS and the invasive potential of FLS in RA, antagonist peptide against EMMPRIN/CD147 (AP-9) was added to the coculture.

The experimental system of FLS separation and culture and HL-60 differentiation were established successfully. The isolated FLS of RA were characterized by negative CD14 and positive CD90. CD147 expression on the differentiated HL-60 cells was higher than that on the undifferentiated HL-60 cells, and CD147 expression on both of them were higher than that on RA FLS (P < 0.05). The expressions of Pro-MMP-2, MMP-2 produced by RA FLS were higher than that produced by OA FLS (P < 0.05). Pro-MMP-2, MMP-2, Pro-MMP-9 and MMP-9 were significantly increased when RA FLS were cocultured with the undifferentiated or differentiated HL-60 cells (P < 0.05), and Pro-MMP-2 and MMP-2 in differentiated HL-60 cells cocultured with RA FLS increased more than those in the undifferentiated HL-60 cells cocultured with RA FLS (P < 0.05). The increase was inhibited by AP-9 significantly (P < 0.05). Invasion assay showed that the invasive potential of RA FLS was higher than that of OA FLS (P < 0.05), and the undifferentiated or differentiated HL-60 cells increased the invasive potential of RA FLS (P < 0.05), which could be blocked by AP-9.

Neutrophil might increase the production of MMP-2 and MMP-9 and enhanced the invasion of RA FLS via CD147, which can be inhibited by HAb18G/CD147 antagonistic peptide (AP9). Our research on RA pathogenesis and the mechanism of cartilage destruction may give us some good ideas for RA therapy in future.