Application of inflammation-responsive promoter for an in vitro arthritis model.
OBJECTIVE
The application of inflammation-regulated therapeutic gene expression in arthritis conditions increases the efficiency of gene therapy by self-limiting the transgene. Incidentally, constitutive overexpression of transgenes typically leads to detrimental effects in disease conditions; therefore, regulation of expression is warranted. We undertook this study to validate a new gene therapy approach using a cell culture-based inflammation model and a novel self-limiting, inflammation-responsive promoter construct.


METHODS
We designed a self-limiting promoter construct that expresses an antiinflammatory gene (interleukin-4 [IL-4]) only in the presence of inflammation. Our construct featured a truncated promoter sequence of cyclooxygenase 2 (COX-2) upstream of the IL-4 gene. We triggered inflammation in vitro in articular chondrocytes by applying the inflammatory cytokines IL-1beta and tumor necrosis factor alpha (TNFalpha) together exogenously, and we studied the extent of IL-4 expression and its effect on the inflammatory cascade.


RESULTS
Using articular chondrocytes, we showed that our COX-2 promoter construct expressed IL-4 only in the presence of IL-1beta and TNFalpha. IL-4 expressed in the presence of IL-1beta and TNFalpha down-regulated a series of inflammation mediators, prostaglandins, and matrix metalloproteinases.


CONCLUSIONS
The use of this construct for the expression of antiinflammatory genes allows production of a therapeutic gene product that is controlled by the severity of the disease. The effectiveness of this promoter construct for combating inflammation makes it a suitable candidate for the development of a new local gene therapy strategy for the treatment of osteoarthritis, in which IL-1beta and TNFalpha trigger a signal cascade that elevates COX-2 levels.
Rachakonda
P Sivaramakrishna
PS
Freie Universität Berlin, Berlin, Germany.
Rai
Muhammad Farooq
MF
Schmidt
Michael F G
MF
D000818:Animals; D019902:Chondrocytes / Q000276:immunology*; D051546:Cyclooxygenase 2 / Q000276:immunology; D004195:Disease Models, Animal; D004285:Dogs; D015536:Down-Regulation; D005786:Gene Expression Regulation; D015316:Genetic Therapy / Q000379:methods*; D066298:In Vitro Techniques; D007249:Inflammation / Q000276:immunology*; D053583:Interleukin-1beta / Q000378:metabolism; D015847:Interleukin-4 / Q000096:biosynthesis*; D011401:Promoter Regions, Genetic; D019076:Transgenes; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology
D053583:Interleukin-1beta; D014409:Tumor Necrosis Factor-alpha; D015847:Interleukin-4; D051546:Cyclooxygenase 2