Improving Communication With Patients Learning by Doing
JAMA-JOURNAL OF THE AMERICAN MEDICAL ASSOCIATION
2013; 310 (21): 2257-2258
A piece of my mind. The road back to the bedside.
JAMA-the journal of the American Medical Association
2013; 310 (8): 799-800
A cascade of Ca2+/calmodulin-dependent protein kinases regulates the differentiation and functional activation of murine neutrophils
2008; 36 (7): 832-844
Heterogeneity of functional responses in differentiated myeloid cell lines reveals EPRO cells as a valid model of murine neutrophil functional activation
JOURNAL OF LEUKOCYTE BIOLOGY
2005; 77 (5): 669-679
The function of neutrophils as primary mediators of innate immunity depends on the activity of granule proteins and critical components of the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex. Expression of their cognate genes is regulated during neutrophil differentiation by a complex network of intracellular signaling pathways. In this study, we have investigated the role of two members of the calcium/calmodulin-dependent protein kinase (CaMK) signaling cascade, CaMK I-like kinase (CKLiK) and CaMKKalpha, in regulating neutrophil differentiation and functional activation.Mouse myeloid cell lines were used to examine the expression of a CaMK cascade in developing neutrophils and to examine the effects of constitutive activation vs inhibition of CaMKs on neutrophil maturation.Expression of CaMKKalpha was shown to increase during neutrophil differentiation in multiple cell lines, whereas expression of CKLiK increased as multipotent progenitors committed to promyelocytes, but then decreased as cells differentiated into mature neutrophils. Expression of constitutively active CKLiKs did not affect morphologic maturation, but caused dramatic decreases in both respiratory burst responses and chemotaxis. This loss of neutrophil function was accompanied by reduced secondary granule and gp91(phox) gene expression. The CaMK inhibitor KN-93 attenuated cytokine-stimulated proliferative responses in promyelocytic cell lines, and inhibited the respiratory burst. Similar data were observed with the CaMKKalpha inhibitor, STO-609.Overactivation of a cascade of CaMKs inhibits neutrophil maturation, suggesting that these kinases play an antagonistic role during neutrophil differentiation, but at least one CaMK is required for myeloid cell expansion and functional activation.
View details for DOI 10.1016/j.exphem.2008.02.009
View details for Web of Science ID 000257349400010
View details for PubMedID 18400360
Mature neutrophils display multiple functional responses upon activation that include chemotaxis, adhesion to and transmigration across endothelial cells, phagocytosis, and pathogen destruction via potent microbicidal enzymes and reactive oxygen species. We are using myeloid cell line models to investigate the signaling pathways that govern neutrophil functional activation. To facilitate these studies, we have performed a direct comparison of functional responses of human and murine myeloid cell line models upon neutrophil differentiation. Our results show that EPRO cells, promyelocytes that undergo complete neutrophil maturation, demonstrate a full spectrum of functional responses, including respiratory burst, chemotaxis toward two murine chemokines, and phagocytosis. We also extend previous studies of granulocyte-colony stimulating factor-induced 32Dcl3 cells, showing they demonstrate chemotaxis and phogocytosis but completely lack a respiratory burst as a result of the absent expression of a critical oxidase subunit, gp91(phox). Induced human leukemic NB4 and HL-60 cells display a respiratory burst and phagocytosis but have defective chemotaxis to multiple chemoattractants. We also tested each cell line for the ability to up-regulate cell-surface membrane-activated complex-1 (Mac-1) expression upon activation, a response mediating neutrophil adhesion and a surrogate marker for degranulation. We show that EPRO cells, but not 32Dcl3 or NB4, significantly increase Mac-1 surface expression upon functional activation. Together, these data show that EPRO and MPRO cells demonstrate complete, functional activation upon neutrophil differentiation, suggesting these promyelocytic models accurately reflect the functional capacity of mature murine neutrophils.
View details for DOI 10.1189/jlb.1004567
View details for Web of Science ID 000228875500008
View details for PubMedID 15673544