CD81 Mediated Regulation of Synoviolin Expression in Synovial Sarcoma Cells

Abstract

Tetraspanin CD81, which is up-regulated in rheumatoid arthritis (RA) synoviocytes, is a molecule containing four transmembrane domains. We recently showed that small interfering RNA targeting CD81 (CD81 siRNA) ameliorated arthritis in rats. In that study, the expression of synoviolin was decreased in RA joints by CD81 siRNA. We also showed that CD81 siRNA decreased the expression of synoviolin and TNF- in SW982 synovial sarcoma cells, suggesting that overexpression of CD81 in synovial cells induces the expression of both synoviolin and TNF- in the same cells. Here we raised monoclonal antibodies (MAbs) against CD81 by immunization of the outer membrane region of CD81 (LEL) that was overexpressed as a recombinant protein.One of these MAbs stimulated CD81 on the membrane of SW982 and up-regulated the expression of synoviolin. We also defined the region of the synoviolin gene promoter that is essential for induction of synoviolin gene expression by using reporter gene system. This system might be useful for the screening of anti-RA medicine.

Identification of Specific Protein Markers of Rheumatoid Arthritis in Synovial Fluid and Serum

“…To establish anti-CD81 MAb, we previously overexpressed and purified the large outer membrane region of CD81 (LEL) 11,12) . Although this MAb recognizes the LEL region of CD81, we detected the whole molecule (26 kDa) and did not fi nd LEL or smaller molecules in western blotting under reducing conditions, indicating that CD81 exists in its entirety on the membranes of microsomes in RA synovial fl uid, and is not processed or degraded in RA joints.…”

Tetraspanin CD81 is expressed in human parotid cancer tissue and mediates cell proliferation

“…We used mouse anti-CD81 IgG1 monoclonal antibody (Clone 1-1 E10-1 F8) and mouse anti-CD81 IgM monoclonal antibody (Clone 1-1 C2-1 B3) as previously described [18,19]. The established parotid cancer cells were seeded in 96-well plates at 1×10 3 cells per well, and after 24 h, the cells were exposed to the anti-CD81 antibody (at 0, 1.0, 2.5, 5.0, and 10.0 µg/ml) for 1-4 d. Afterward, the cells were incubated with reagents from the Cell Counting Kit-8 (Dojindo Molecular Technology) for 4 h, and the absorbance was measured at 450 nm using a 96-well plate reader (Corona grating microplate reader SH-1000 Lab).…”