Neutrophil Functions and Autoimmune Arthritis in the Absence of p190RhoGAP: Generation and Analysis of a Novel Null Mutation in Mice

Abstract

β2-integrins of neutrophils play a critical role in innate immune defense but they also participate in tissue destruction during autoimmune inflammation. p190RhoGAP, a regulator of Rho-family small GTPases, is required for integrin signal transduction in fibroblasts. Prior studies have also suggested a role for p190RhoGAP in β2 integrin signaling in neutrophils. To directly test that possibility, we have generated a novel targeted mutation completely disrupting the p190RhoGAP-encoding gene in mice. p190RhoGAP deficiency led to perinatal lethality and defective neural development, precluding the analysis of neutrophil functions in adult p190RhoGAP−/− animals. This was overcome by transplantation of fetal liver cells from p190RhoGAP−/− fetuses into lethally irradiated wild type recipients. Neutrophils from such p190RhoGAP−/− bone marrow chimeras developed normally and expressed normal levels of various cell surface receptors. Though p190RhoGAP−/− neutrophils showed moderate reduction of β2 integrin-mediated adherent activation, they showed mostly normal migration in β2-integrin-dependent in vitro and in vivo assays and normal β2 integrin-mediated killing of serum-opsonized S. aureus and E. coli. A neutrophil- and β2 integrin-dependent transgenic model of the effector phase of autoimmune arthritis also proceeded normally in p190RhoGAP−/− bone marrow chimeras. In contrast, all the above responses were completely blocked in CD18−/− neutrophils or CD18−/− bone marrow chimeras. These results suggest that p190RhoGAP likely does not play a major indispensable role in β2 integrin-mediated in vitro and in vivo neutrophil functions or the effector phase of experimental autoimmune arthritis.

Role of Rac GTPase activating proteins in regulation of NADPH oxidase in human neutrophils

“…The lack of effect upon immunodepletion of ARHGAP35 agrees with data obtained in mice with ARHGAP35-deficient hemopoetic system [62]. Apparently, membrane-associated RacGAPs exhibit selectivity in their interactions with the Nox2 complex.…”

The neglected terminators: Rho family GAPs in neutrophils

“…Rac/Rho Human (cell free) NADPH oxidase [6,9,11] Mouse No role in integrin-mediated neutrophil functions and in rheumatoid arthritis [12] Human β2 integrin activation [13] ARHGAP46/GMIP RhoA Mouse/human Azurophilic granule exocytosis. [14] ARHGAP15 Rac Mouse Migration, NADPH oxidase, phagocytosis, abdominal sepsis, retention in bone marrow [15,16] Human/cell culture Migration, actin polymerization [17] ARHGAP13/srGAP1 Cdc42 Mouse/human Neutrophil chemotaxis [18] BCR/ABR Rac/Cdc42 Mouse (double knockout) Acute inflammatory responses [19] BCR Rac/Cdc42 Human (cell free) NADPH oxidase [9] Mouse Septic shock NADPH oxidase [20] ARHGAP12 Rac Human/cell culture Actin polymerization [8] ARHGAP43/SH3BP1 Rac Human/cell culture Actin polymerization [8] ARHGAP21/ ARHGAP10…”

“…31,32 Although the regulatory function could be demonstrated both with human proteins and with murine cells, contribution of the protein to integrin activation differs in human and murine neutrophils. 12,13 In addition to phosphorylation, GAP activity can be altered in numerous other ways. 33 Neutrophils are characterized by a short lifetime (see Hellebrekers et al, in this issue); hence, expression-level regulation is not what one would typically expect.…”

Rac GTPase Activating Protein ARHGAP25 Regulates Leukocyte Transendothelial Migration in Mice

“…To reveal differences in body weight between wild-type (WT) and Arhgap25 2/2 mice, 5-wk-old animals (three males per genotype and two females per genotype) were weighed for 14 wk. Arhgap25 2/2 and control WT bone marrow chimeras were generated using bone marrow cells from adult donors as described previously (24,25). Arhgap25 2/2 bone marrow cell suspensions were injected i.v.…”

“…Efficiency of repopulation of the hematopoietic compartment by donor-derived cells was .98%, tested 4 wk after transplantation by flow cytometry: we tested the expression of CD45.2 (donor) allele in the granulocyte gate determined by Ly-6G staining, as described previously (24,25) (data not shown). Bone marrow chimeras were used 4-8 wk after transplantation.…”