Expressão, caracterização e sinalização da citocina recombinante IL-17A de Bos taurus

Abstract

A Bos taurus interleukin 17A is a proinflammatory cytokine, which acts primarily in the defense of the host organism against extracellular, fungal and viral bacterial infections. Its inflammatory action is due to the signaling to the expression of other proinflammatory cytokines that generate a high level of recruitment of polymorphonuclear cells to the site of infection. The objective of the study was to express, characterize and evaluate the signaling of Bos taurus RIL-17A. Cloning was performed using pAE vector, and an expression in Rosetta ™ strain of Escherichia coli. A protein was expressed as expected molecular mass of approximately 15 kDa, and its characterization was performed by Western Blot using anti-6xHis monoclonal antibody (mAB). Mouse and bovine splenocytes, bovine peripheral blood mononuclear cells (PBMCs) and bovine kidney lineage cells (MDBK) were stimulated with 10μg of rIL-17A and cultured for 24 hours. As cells were collected and mRNA extracted for cDNA synthesis. This material was used to evaluate recombinant protein signaling through the detection of transcription levels of IL-17, IFNγ, IL-4 and IL-2 mRNA by the qPCR technique. rIL-17A showed no toxic effect on cells and maintained its biologically active activity, elevating IL-17 transcription levels in all as stimulated cells. An immune system control response can also be observed, since levels of IFNγ, IL-4 and IL-2 transcripts have also been elevated, albeit in a discrete manner. Further studies are needed to establish a rIL-17A as an immunobiological and potential adjuvant vaccine for cattle and other species.