Expression, purification, and in vitro biological activities of recombinant bovine granulocyte-colony stimulating...

Neutrophils are essential components of the innate immune system and they play a critical role in the defense of host against bacterial and fungal infections. The colony stimulating factors are a class of glycoproteins that are required for proliferation, differentiation, and functional activation of hematopoietic progenitor cells. Granulocyte-colony stimulating factor (G-CSF) is a member of this regulatory family of cytokines that specifically stimulates proliferation and maturation of precursor cells in the bone marrow into fully differentiated and functional neutrophils. G-CSF also modulates the biological activities of mature neutrophils in circulation. A bovine G-CSF (bG-CSF) cDNA clone (previously isolated and sequenced in our laboratory) was expressed in Escherichia coli and the biological activities of the solubilized protein from purified inclusion bodies were examined. Flow cytometric analysis of membrane antigen density of neutrophils activated with bG-CSF revealed an upregulation in the expression of CD11a (>114%), CD11b (>148%), CD11c (>87%), and CD18 (>109%). Expression of L-selectin was decreased by more than 43%. There was no change, however, in the expression of CD14. These findings indicate that recombinant bG-CSF (rbG-CSF) expressed in E. coli is biologically active and exerts the same type of effects on neutrophils in vitro as those of human G-CSF (hG-CSF).