Construction of genetically modified Lactococcus lactis that produces bioactive anti-interleukin-4 single-chain fragment variable.

Abstract

Interleukin 4 (IL-4) is a cytokine that induces T-cell differentiation and the production of antibodies from B cells, and plays a crucial role in the allergic response. Therefore, development of a therapeutic approach against IL-4 signaling is expected to prevent or control Th2-related allergic diseases. IL-4 single-chain fragment variable (scFv), which is a recombinant protein consisting of the Fv region of an IL-4 antibody connected to a flexible peptide linker, is expected to be an inhibitor of IL-4 signaling. In this study, recombinant IL-4 scFv was produced by genetically modified lactic acid bacteria (gmLAB); this system is gaining attention as a type of microbial therapeutics. Recombinant gene expression was confirmed with western blotting, and the IL-4 recognition ability of IL-4 scFv produced by gmLAB was examined with an enzyme-linked immunosorbent assay. The macrophage cell line, Raw264.7, and peritoneal macrophages isolated from C57BL/6 mice were employed for an in vitro IL-4 signaling inhibition assay. IL-4 stimulation increased the mRNA expression of arginase-1, a biomarker of IL-4 signaling in macrophages, but arginase-1 expression was suppressed by IL-4 scFv produced by gmLAB, indicating that IL-4 scFv has IL-4 signaling inhibitory activity. gmLAB that produces bioactive IL-4 scFv that was constructed in this study could be an attractive approach for treating allergic disorders.