Molecular cloning and in vivo evaluation of canine granulocyte-macrophage colony-stimulating factor

Abstract

Canine granulocyte-macrophage colony-stimulating factor (caGM-CSF) was cloned and expressed to allow further investigation of GM-CSF in a large animal model. The cDNA is 850 base pairs (bp) long and encodes a peptide of 144 amino acids. The nucleotide and amino acid sequence homology between caGM-CSF and human GM-CSF (hGM-CSF) is 80% and 70%, respectively. A mammalian expression vector pCMV/CAGM was constructed and used to transfect COS cells for expression of caGM-CSF. Supernatant from transfected COS cells enriched with caGM-CSF was shown to have significant stimulating activity in granulocyte-macrophage colony forming unit (CFU-GM) assays of canine marrow. caGM-CSF, expressed from bacteria, was used to treat seven dogs at varying doses twice daily subcutaneously (sc) for 14 to 16 days. Circulating blood neutrophils and monocytes increased significantly. The increase in circulating eosinophils was variable. Thrombocytopenia developed during administration of caGM-CSF but corrected rapidly after cessation of treatment. Evaluation of survival times of 51Cr-labeled autologous platelets suggested increased consumption as the primary reason for thrombocytopenia. A species-specific GM-CSF will be a useful tool for hematologic or immunologic studies in dogs.