Suppression of arthritic bone destruction by adenovirus-mediated csk gene transfer to synoviocytes and osteoclasts
Hiroshi Takayanagi, … , Kozo Nakamura, Sakae Tanaka
Hiroshi Takayanagi, … , Kozo Nakamura, Sakae Tanaka
Published July 15, 1999
Citation Information: J Clin Invest. 1999;104(2):137-146. https://doi.org/10.1172/JCI6093.
View: Text | PDF
Article

Suppression of arthritic bone destruction by adenovirus-mediated csk gene transfer to synoviocytes and osteoclasts

Abstract

Rheumatoid arthritis (RA) is characterized by a chronic inflammation of the synovial joints resulting from hyperplasia of synovial fibroblasts and infiltration of lymphocytes, macrophages, and plasma cells, all of which manifest signs of activation. Recent studies have revealed the essential role of osteoclasts in joint destruction in RA. Src family tyrosine kinases are implicated in various intracellular signaling pathways, including mitogenic response to growth factors in fibroblasts, activation of lymphocytes, and osteoclastic bone resorption. Therefore, inhibiting Src activity can be a good therapeutic strategy to prevent joint inflammation and destruction in RA. We constructed an adenovirus vector carrying the csk gene, which negatively regulates Src family tyrosine kinases. Csk overexpression in cultured rheumatoid synoviocytes remarkably suppressed Src kinase activity and reduced their proliferation rate and IL-6 production. Bone-resorbing activity of osteoclasts was strongly inhibited by Csk overexpression. Furthermore, local injection of the virus into rat ankle joints with adjuvant arthritis not only ameliorated inflammation but suppressed bone destruction. In conclusion, adenovirus-mediated direct transfer of the csk gene is useful in repressing bone destruction and inflammatory reactions, suggesting the involvement of Src family tyrosine kinases in arthritic joint breakdown and demonstrating the feasibility of intervention in the kinases for gene therapy in RA.

Authors

Hiroshi Takayanagi, Takuo Juji, Tsuyoshi Miyazaki, Hideharu Iizuka, Tokiharu Takahashi, Masashi Isshiki, Masato Okada, Yoshiya Tanaka, Yasuko Koshihara, Hiromi Oda, Takahide Kurokawa, Kozo Nakamura, Sakae Tanaka

×

Figure 2

Options: View larger image (or click on image) Download as PowerPoint
Csk expression and the kinase activity of synoviocytes. (a) Western blot...
Csk expression and the kinase activity of synoviocytes. (a) Western blot analysis shows that Csk virus induced Csk expression in human synovial cells in an moi-dependent manner without significant changes in expression of c-Src protein. Fold increase in Csk expression normalized by β-actin is shown below each blot. The results shown are from a single experiment typical of 3 experiments yielding identical results. (b) Immunofluorescence staining using anti-Csk antibody shows that WT virus–infected synoviocytes expressed a very small amount of Csk protein. (c) Diffuse cytoplasmic staining in Csk virus–infected (moi = 100) synoviocytes. (d) In vitro kinase assay, using enolase as a substrate demonstrated that kinase activity of c-Src, Fyn, and c-Yes was remarkably repressed in Csk-overexpressing synoviocytes, but the expression of each protein was unaffected by Csk expression. Relative kinase activity of each kinase is shown below. The results were derived from 3 separate experiments. Suppression on each kinase was statistically significant (*P < 0.05 Csk+ vs. Csk–).