|Junker, David - SCHERING-PLOUGH,CA|
|Cochran, Mark - SCHERING-PLOUGH,CA|
Submitted to: Veterinary Immunology and Immunopathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: August 28, 2002
Publication Date: October 28, 2002
Citation: Solano Aguilar, G., Zarlenga, D.S., Beshah, E., Gasbarre, L.C., Junker, D., Cochran, M., Dawson, H.D., Urban Jr, J.F., Lunney, J.K. 2002. Limited effect of recombinant porcine interleukin 12 on porcine lymphocytes due to a low level of il-12 beta 2 receptor. Veterinary Immunology and Immunopathology. 89:133-148 (2002) Interpretive Summary: Pigs are exposed to infections as they move through different production facilities, from farrowing to nursery to grow-out areas. These changes and the admixing of pig populations provide opportunities for them to become exposed to numerous infectious diseases, requiring drug treatments. ARS scientists at Beltsville, MD through the BRDC (Biotechnology and Research development Corporation) and their collaborating pharmaceutical and breeding company partners, have look for new disease and vaccine therapeutics. In this effort we cloned and purified a new biotherapeutic, the expressed recombinant porcine cytokine interleukin-12 (rPoIL-12) and tested it for efficacy as a stimulant of pig immune system activities. Unexpectedly, rPoIL-12 has limited stimulating activities for isolated peripheral blood mononuclear cells isolated from whole blood. Detailed in vitro and in vivo studies proved that this ineffectiveness was due to the fact that rPoIL-12 does not upregulate IL-12 receptor expression; even though the same rPoIL-12 does stimulate bovine cell IL-12R expression. Thus, tests in swine have shown that a predicted disease biotherapeutic, IL-12, when given alone, does not exhibit immune stimulatory activities, and thus would not be a good alternate for disease treatments. This is essential information for pharmaceutical and vaccine companies as they search for the best immunostimulants for swine.
Technical Abstract: The cytokine interleukin-12 (IL-12) is a key molecule in the regulation of CD4+ T cell development and specifically potentiates T helper 1 responses in mouse and man. However, biological effects mediated by IL-12 have not been well defined in pigs. Herein, recombinant porcine IL-12 (rPoIL-12) was expressed in a swine poxvirus system as a biologically active heterodimer and used to stimulate bovine or swine lymphoblast cells. After 3 days of incubation, only bovine blasts were responsive to the rPoIL-12 treatment as monitored by cell proliferation in several independent trials. Similarly, i.m. administration of rPoIL-12 in the hind leg of 3-week-old pigs indicated a reduction in the number of interferon-gamma (IFN-gamma) producing lymphocytes isolated from inguinal lymph nodes. The porcine IL-12R beta 2 (IL-12Rbeta 2) sequence was cloned and results generated by reverse transcriptase polymerase chain reaction (RT-PCR) demonstrated that the expression of IL-12R on porcine blasts as measured by the relative levels of IL-12Rbeta2 mRNA was less than that in bovine blasts and are in agreement with the reduced proliferation response of swine blast cells to rPoIL-12 treatment. Real time PCR analysis demonstrated that after PBMC stimulation, bovine blasts had an 11-fold increase in IL-12beta2 mRNA levels while porcine blasts had almost no change. These data support a mechanism for IL-12 stimulation in swine inconsistent with that observed in conventional models.