Transfection of IL-10 expression vectors into endothelial cultures attenuates alpha4beta7-dependent lymphocyte adhesion mediated by MAdCAM-1

Abstract

Background: Enhanced expression of MAdCAM-1 (mucosal addressin cell adhesion molecule-1) is associated with the onset and progression of inflammatory bowel disease. The clinical significance of elevated MAdCAM-1 expression is supported by studies showing that immunoneutralization of MAdCAM-1, or its ligands reduce inflammation and mucosal damage in models of colitis. Interleukin-10 (IL-10) is an endogenous anti-inflammatory and immunomodulatory cytokine that has been shown to prevent inflammation and injury in several animal studies, however clinical IL-10 treatment remains insufficient because of difficulties in the route of IL-10 administration and its biological half-life. Here, we examined the ability of introducing an IL-10 expression vector into endothelial cultures to reduce responses to a proinflammatory cytokine, TNF-alpha

Methods: A human IL-10 expression vector was transfected into high endothelial venular ('HEV') cells (SVEC4-10); we then examined TNF-alpha induced lymphocyte adhesion to lymphatic endothelial cells and TNF-alpha induced expression of MAdCAM-1 and compared these responses to control monolayers.

Results: Transfection of the IL-10 vector into endothelial cultures significantly reduced TNF-alpha induced, MAdCAM-1 dependent lymphocyte adhesion (compared to non-transfected cells). IL-10 transfected endothelial cells expressed less than half (46 +/- 6.6%) of the MAdCAM-1 induced by TNF-alpha (set as 100%) in non-transfected (control) cells.

Conclusion: Our results suggest that gene therapy of the gut microvasculature with IL-10 vectors may be useful in the clinical treatment of IBD.

Figure 1

Secretion of human IL-10 concentration by transfected endothelial cells. There was no detectable human IL-10 signal in the non-transfected cell medium (control). However, the medium from the IL-10 transfected SVEC medium showed a large and significant increase in the IL-10 concentration (1209 ± 2 pg/ml) at 48 h after IL-10 gene transfection (n = 5). (# p < 0.05 from control). One-way ANOVA with Fisher's PLSD test.

Figure 2

IL-10 gene transfected SVEC is resistant to TNF-α induced MAdCAM-1 expression. TNF-α (1 ng/ml, 24 h) significantly increased expression of MAdCAM-1 and this was significantly blocked by IL-10 gene transfection. Alone, IL-10 gene transfer had no effect on MAdCAM-1 expression (n = 5). (* p < 0.05 from TNF-α, # p < 0.05 from control). One-way ANOVA with Fisher's PLSD test.

Figure 3

IL-10 gene transfer blocked TNF-α induced lymphocyte adhesion on SVEC. TNF-α stimulation (1 ng/ml, 24 h) significantly increased the adhesion of TK-1 lymphocytes to SVEC monolayers. IL-10 gene transfer significantly reduced TK-1 adhesion in response to TNF-α stimulation at 24 h. IL-10 gene transfer did not modify the basal level of lymphocyte adhesion to the endothelium without TNF-α treatment (n = 7). (* p < 0.05 from TNF-α, # p < 0.05 from control). One-way ANOVA with Fisher's PLSD test.