Granulocyte Macrophage Colony Stimulating Factor

Neutralizing polyclonal antibodies to MuGM-CSF have been characterized (52), and well-characterized monoclonal anti-MuGM-CSF Abs (53,54) are now commercially available. Such Abs form the basis of enzyme-linked immunosorbent assays (ELISAs) for determination of immunoreactive MuGM-CSF levels and have been used to show specificity in MuGM-CSF bioassays. Although no studies attempting to neutralize basal levels of endogenously produced MuGM-CSF by passive immunization in vivo have been reported, Abs have been used to neutralize GM-CSF activity in disease models. The effect of GM-CSF pretreatment to aggravate lipopolysaccharide (LPS)-induced mortality and hepatic toxicity could be ameliorated by the administration of GM-CSF Abs (55). Administration of an anti-GM-CSF Ab attenuated the severity of arthritis in two murine arthritis models, one in which erosive arthritis is induced by bovine serum albumin (BSA) and IL-1 administration (56), and in one model of collagen-induced arthritis (57).

A competitive antagonist of HuGM-CSF has been developed named E21R, which is a ligand analog in which amino acid 21 is changed from glutamic acid to arginine (58). Owing to the high species specificity of GM-CSF, preclinical in vivo studies with the moiety were performed in baboons, administering E21R for up to 21 d (59) (Table 4). E21R resulted in a transient eosinophilia and neutrophilia and granulocyte infiltrates in lymph nodes and duodenal submucosa. The transient eosinophilia was unexpected but was also seen in patients receiving E21R on a phase 1 study (59), and so is an effect of this agent accurately predicted by the animal model.

Two mouse lines with absolute GM-CSF deficiency owing to targeted gene disruption have been independently generated (60,61); both lines show identical phenotypes. Baseline hematopoiesis is unperturbed despite GM-CSF deficiency (61), although reduced frequencies of marrow CFU-E sensitive to low EPO concentrations in vitro have recently been documented (31). During M. avium infection, GM-CSF-/- mice fail

Table 4

Animal Models of Reduced GM-CSF Levels or Signaling

Animal Method of reduced GM-CSF signaling Major phenotypic consequences Reference

Table 4

Animal Models of Reduced GM-CSF Levels or Signaling

Animal Method of reduced GM-CSF signaling Major phenotypic consequences Reference

Mouse

Passive immunization with anti-MuGM-CSF antibodies

i LPS-induced mortality i LPS hepatic toxicity

55

Mouse

Targeted disruption of GM-CSF gene

Normal basal hematopoiesis Pulmonary alveolar proteinosis i hematopoiesis during chronic M. avium infection i zymocel-induced hepatic granulomatous inflammation

60-62, 68

Mouse

Targeted disruption of IL-3/GM-CSF/IL-5 receptor pc subunit

Normal basal hematopoiesis except i eosinophil production Pulmonary alveolar proteinosis Failure to develop eosinophila to parasitic infections

68, 69

Baboon

Competitive peptide antagonist (E21R)

Transient eosinophilia and neutrophilia

59

Abbreviations: GM-CSF, granulocyte-macrophage colony-stimulating factor; LPS, lipopolysaccharide; IL, interleukin; Mu, murine; T, increased; 4-, decreased.

Abbreviations: GM-CSF, granulocyte-macrophage colony-stimulating factor; LPS, lipopolysaccharide; IL, interleukin; Mu, murine; T, increased; 4-, decreased.

to sustain hematopoietic cell production (62), suggesting a role for GM-CSF under emergency if not basal conditions of hematopoiesis. GM-CSF-/- mice have been exploited to examine the role of this factor in several models of inflammation; different effects have been seen in different models. Acute peritoneal inflammation after casein injection was normal in GM-CSF-/- mice (63). GM-CSF deficiency delayed zymocel-induced hepatic granuloma formation and impaired monocyte infiltration and proliferation, although macrophages within granulomata expressed markers suggesting normal activation (64). Normal activation of peritoneal macrophages was observed during L. monocytogenes infection (45). GM-CSF deficiency attenuated inflammation in a murine model of arthritis induced by BSA and IL-1 injection (56) and also in murine models of immune-mediated glomerulonephritis (65). GM-CSF-/- mice have moderately impaired reproductive capacity and reduced long-term survival (66).

GM-CSF-/- mice develop a striking pulmonary pathology with extensive peribronchial B-cell infiltrates and alveolar accumulation of surfactant phospholipid, protein, and intra-alveolar macrophages, a disorder resembling pulmonary alveolar proteinosis (60,61). The pathophysiology relates to impaired surfactant clearance and catabolism (216) and can be reversed by local GM-CSF expression (67), evidence collectively indicating a local defect in alveolar macrophages.

GM-CSF signaling is initiated by ligand binding to a heterodimeric receptor comprising a specific a-subunit (GM-CSFRa) and a p-subunit (IL-3/GM-CSF/IL-5Rpc) shared in common with the analogously heterodimeric IL-3 and IL-5 receptors. (In mice, but not in humans, there are two rather than one IL-3 receptor P-subunits.) GM-CSF deficiency has been mimicked by targeted disruption of the IL-3/GM-CSF/IL-5Rpc gene (68,69), and these mice develop a similar, but less severe, pulmonary pathology (70).

Additionally, they showed additional manifestations of defective IL-5 signaling such as low baseline eosinophil numbers (68) and impaired eosinophil response to Nip-postrongylus brasiliensis (68,69,71). In this cell-autonomous model of the pulmonary disease, bone marrow transplantation with wild-type hematopoietic cells reversed the pulmonary pathology (72), albeit not completely (73). IL-3/GM-CSF/IL-5RPc-deficient mice displayed an attenuated cutaneous reaction to Leishmania major (74).

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