title: Cloning and expression of recombinant equine interleukin-3 and its effect on sulfidoleukotriene and cytokine production by equine peripheral blood leukocytes.
abstract: Interleukin-3 is a growth and differentiation factor for various hematopoietic cells. IL-3 also enhances stimulus-dependent release of mediators and cytokine production by mature basophils. Function of IL-3 has not been studied in horses because of lack of horse-specific reagents. Our aim was to produce recombinant equine IL-3 and test its effect on sulfidoleukotriene and cytokine production by equine peripheral blood leukocytes (PBL). Equine IL-3 was cloned, expressed in E. coli and purified. PBL of 19 healthy and 20 insect bite hypersensitivity (IBH)-affected horses were stimulated with Culicoides nubeculosus extract with or without IL-3. Sulfidoleukotriene (sLT) production was measured in supernatants by ELISA and mRNA expression of IL-4, IL-13 and thymic stromal lymphopoietin (TSLP) assessed in cell lysate by quantitative real-time PCR. Recombinant equine IL-3 (req-IL-3) had a dose dependent effect on sLT production by stimulated equine PBL and significantly increased IL-4, IL-13 and TSLP expression compared to non-primed cells. IL-3 priming significantly increased Culicoides-induced sLT production in IBH-affected but not in non-affected horses and was particularly effective in young IBH-affected horses (≤ 3 years). A functionally active recombinant equine IL-3 has been produced which will be useful for future immunological studies in horses. It will also allow improving the sensitivity of cellular in vitro tests for allergy diagnosis in horses.
authors.affiliation: Department of Clinical Research-VPH, Vetsuisse Faculty, University of Bern, Switzerland; Center Pigmod, Institute of Animal Physiology and Genetics, Academy of Sciences of Czech Republic, Libechov, Czech Republic.
authors.forename: Jozef
authors.initials: J
authors.lastname: Janda
authors.affiliation: ImmunoTools, Friesoythe, Germany.
authors.forename: Melissa
authors.initials: M
authors.lastname: Lehmann
authors.affiliation: ImmunoTools, Friesoythe, Germany.
authors.forename: Werner
authors.initials: W
authors.lastname: Luttmann
authors.affiliation: Department of Clinical Research-VPH, Vetsuisse Faculty, University of Bern, Switzerland. Electronic address: eliane.marti@vetsuisse.unibe.ch.
authors.forename: Eliane
authors.initials: E
authors.lastname: Marti
mesh_terms: D000375:Aging; D000818:Animals; D002519:Ceratopogonidae; D003001:Cloning, Molecular; D016207:Cytokines / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects*; D006736:Horses / Q000378:metabolism*; D006967:Hypersensitivity / Q000276:immunology / Q000662:veterinary; D007299:Insect Bites and Stings / Q000276:immunology / Q000662:veterinary; D007377:Interleukin-3 / Q000494:pharmacology*; D007962:Leukocytes / Q000187:drug effects / Q000378:metabolism; D015289:Leukotrienes / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins
chemical_list: D016207:Cytokines; D007377:Interleukin-3; D015289:Leukotrienes; D011994:Recombinant Proteins
keywords: Cytokines; Horse; Interleukin-3; Sulphidoleukotriene release