The role of interchain disulfide bond in a recombinant human interleukin-17A variant.

Interleukin-17A (IL-17A) is the prototype of IL-17 family and has been implicated in the pathogenesis of a variety of autoimmune diseases. Therefore its structural and functional properties are of great medical interest. During our research on a recombinant human IL-17A (rhIL-17A) variant, four isoforms were obtained when it was refolded. While isoforms 1 and 2 represented non-covalent dimers, isoforms 3 and 4 were determined to be covalent dimers. All four isoforms were structurally similar by Circular Dichroism and fluorescence spectroscopy studies, but differential scanning calorimetry demonstrated thermal stability in the order of isoform 1=isoform 2

Negative regulation of IL-17 production by OX40/OX40L interaction.

The T-cell cytokine IL-17 is implicated in multiple inflammatory diseases through its induction of several pro-inflammatory cytokines and chemokines in a broad range of cell targets. Production of IL-17 defines the Th17 subset of helper T-cells associated with protection against microorganisms, a profile best characterized in the murine system. Multiple regulators of Th17 cell differentiation and IL-17 production are reported, but the impact of OX40L is not described. OX40 ligand (OX40L) is an early-stage activator of T-cells through its interaction with CD134 (OX40) that is up-regulated on antigen challenged T-cells. Here, we show that OX40L suppresses IL-17 production by PHA-stimulated human PBMC and purified CD4 and CD8 cells. In agreement with prior reports, OX40L signaling through CD134 increased IFNgamma and IL-4, both of which are reported to inhibit the production of IL-17. OX40L suppression of IL-17 was completely reversed by a neutralizing IFNgamma antibody while there was no effect with a neutralizing IL-4 antibody. Moreover, OX40L also suppressed IL-17 in the presence of IL-23, an established inducer of IL-17 and differentiation factor for Th17 cells. Presuming mediation by IFNgamma, we evaluated expression of this cytokine in the presence of OX40L and IL-23. Surprisingly, IL-23 also induced IFNgamma by PHA-stimulated T-cells and this effect was enhanced in the presence of OX40L. Addition of the IFNgamma antibody not only reversed the OX40L suppression of IL-17 in the presence of IL-23, it markedly enhanced the level of IL-17. These results further establish IFNgamma as a primary modulator of IL-17 production in the human cells, much as in the murine system.