Expression of equine interleukin-18 by baculovirus expression system and its biologic activity.

The research article is about a study where the coding sequence for equine interleukin-18 (IL-18) was cloned and expressed through a recombinant baculovirus. The biological activity of the resulting recombinant equine IL-18 was also confirmed.

Cloning and Expression of Equine Interleukin-18

• The study started with the cloning of the cDNA of equine interleukin-18 (IL-18), a protein that plays a role in immune responses.
• A recombinant baculovirus named AcEIL-18 was created to express this cloned IL-18.
• The production of the recombinant equine IL-18 by AcEIL-18 in infected insect cells was confirmed by testing methods such as sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting.

Production and Analysis of IL-18 Protein

• The infected insect cells secreted a precursor version of IL-18, which has a molecular weight of 24 kilo daltons (kDa), into the surrounding culture medium.
• Crucially, mature equine IL-18 with a weight of about 18 kDa was also detected through Western blotting. This was verified without the co-expression of caspase-1, an enzyme often required for IL-18 activation.

Demonstration of Biological Activity

• The researchers then sought to verify if the produced recombinant IL-18 retained its biological function.
• This was done by testing whether the culture supernatant from AcEIL-18 infected cells could induce interferon-gamma gene expression in equine peripheral mononuclear cells.
• This is significant because interferon-gamma is a key molecule in immune response and is stimulated by IL-18. Successful stimulation by the produced recombinant IL-18 would confirm its biological activity.
• When combined with recombinant human interleukin-12, the infected cell supernatant showed a significant effect, suggesting that the equine IL-18 expressed in this study retained its biological function even without additional treatment.