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A role for Mac-1 (CDIIb/CD18) in immune complex-stimulated neutrophil function in vivo: Mac-1 deficiency abrogates sustained Fcgamma receptor-dependent neutrophil adhesion and complement-dependent proteinuria in acute glomerulonephritis.

Tang T, Rosenkranz A, Assmann KJ, Goodman MJ, Gutierrez-Ramos JC, Carroll MC, Cotran RS, Mayadas TN - J. Exp. Med. (1997)

Bottom Line: In vitro studies showed that spreading of Mac-1- PMNs to IC-coated dishes was equivalent to that of wild-type PMNs at 5-12 min but was markedly reduced thereafter, and was associated with an inability of mutant neutrophils to redistribute filamentous actin.Complement C3-deficient mice also had significantly decreased proteinuria compared to wild-type mice.Since Mac-1 on PMNs is the principal ligand for ic3b, an absence of Mac-1 interaction with C3 probably contributed to the abrogation of proteinuria in Mac-1- mice.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Brigham and Women's Hospital, Boston, Massachusetts 02115, USA.

ABSTRACT
Mac-1 (alphambeta2), a leukocyte adhesion receptor, has been shown in vitro to functionally interact with Fcgamma receptors to facilitate immune complex (IC)-stimulated polymorphonuclear neutrophil (PMN) functions. To investigate the relevance of Mac-1-FcgammaR interactions in IC-mediated injury in vivo, we induced a model of Fc-dependent anti-glomerular basement membrane (GBM) nephritis in wild-type and Mac-1-deficient mice by the intravenous injection of anti-GBM antibody. The initial glomerular PMN accumulation was equivalent in Mac-1 and wild-type mice, but thereafter increased in wild-type and decreased in mutant mice. The absence of Mac-1 interactions with obvious ligands, intercellular adhesion molecule 1 (ICAM-1), and C3 complement, is not responsible for the decrease in neutrophil accumulation in Mac-1- deficient mice since glomerular PMN accumulation in mice deficient in these ligands was comparable to those in wild-type mice. In vitro studies showed that spreading of Mac-1- PMNs to IC-coated dishes was equivalent to that of wild-type PMNs at 5-12 min but was markedly reduced thereafter, and was associated with an inability of mutant neutrophils to redistribute filamentous actin. This suggests that in vivo, Mac-1 is not required for the initiation of Fc-mediated PMN recruitment but that Mac-1-FcgammaR interactions are required for filamentous actin reorganization leading to sustained PMN adhesion, and this represents the first demonstration of the relevance of Mac-1-FcgammaR interactions in vivo. PMN-dependent proteinuria, maximal in wild-type mice at 8 h, was absent in Mac-1 mutant mice at all time points. Complement C3-deficient mice also had significantly decreased proteinuria compared to wild-type mice. Since Mac-1 on PMNs is the principal ligand for ic3b, an absence of Mac-1 interaction with C3 probably contributed to the abrogation of proteinuria in Mac-1- mice.

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PMN spreading on  ICs. Samples analyzed in Fig. 4  were also analyzed for PMN  spreading on ICs (A) or BSA alone  (B) and parallel samples were analyzed for F-actin distribution  (C). The spreading in Mac-1–  cells (diamonds) and wild-type  cells (squares) was equivalent at 5  min, but was dramatically reduced at 25 and 40 min in   cells. Representative Wright Giemsa–stained slides from the 40  min time point are shown. Spreading on BSA was significantly different between wild-type and   PMNs at 5 min and remained so  at 40 minutes. The difference in  profiles of PMN spreading on  ICs and BSA alone indicates the  Mac-1 effects on PMN spreading  that are IC-specific. *P <0.005,  #P <0.05 compared to wild-type  cells. (C) Wild-type (WT) and  Mac-1– cells adherent to ICs  for 5 and 40 min were stained  with rhodamine phalloidin to  identify F-actin. At 5 min, cells  of both genotypes initiated actin  polymerization at the periphery  of the cells. By 40 min, wild-type PMNs had developed an actin-rich surface with a punctate  cytoplasmic pattern and had visible projections. In contrast, mutant cells had not progressed to  this stage and several of them had  rounded up (arrow). Original  magnification: ×473.
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Figure 5: PMN spreading on ICs. Samples analyzed in Fig. 4 were also analyzed for PMN spreading on ICs (A) or BSA alone (B) and parallel samples were analyzed for F-actin distribution (C). The spreading in Mac-1– cells (diamonds) and wild-type cells (squares) was equivalent at 5 min, but was dramatically reduced at 25 and 40 min in cells. Representative Wright Giemsa–stained slides from the 40 min time point are shown. Spreading on BSA was significantly different between wild-type and PMNs at 5 min and remained so at 40 minutes. The difference in profiles of PMN spreading on ICs and BSA alone indicates the Mac-1 effects on PMN spreading that are IC-specific. *P <0.005, #P <0.05 compared to wild-type cells. (C) Wild-type (WT) and Mac-1– cells adherent to ICs for 5 and 40 min were stained with rhodamine phalloidin to identify F-actin. At 5 min, cells of both genotypes initiated actin polymerization at the periphery of the cells. By 40 min, wild-type PMNs had developed an actin-rich surface with a punctate cytoplasmic pattern and had visible projections. In contrast, mutant cells had not progressed to this stage and several of them had rounded up (arrow). Original magnification: ×473.

Mentions: To determine whether spreading of mutant PMNs on IC was compromised, we  determined the percentage of spread cells in the samples generated in Fig. 4. Initial spreading on ICs was the same for wild-type and Mac-1– cells at 5 min. The percentage of spread cells increased over time in wild-type cells (Fig. 5 A), consistent with the increased numbers of PMNs adherent to the IC at these later time points as seen in Fig. 4. On the other hand, the percentage of mutant PMNs spread on IC declined rapidly over time and was <10% of wild-type levels at 40 min (Fig. 5 A) and <0.03% at 120 min (data not shown). On BSA-coated surfaces, a difference in spreading was observed between wild-type and cells at 5 min and remained constant up to 40 min (Fig. 5 B). This is consistent with previous studies that indicate a role for Mac-1 in spreading on BSA (1). The equivalent spreading on ICs at 5 min versus the difference in spreading seen on BSA-coated surfaces at this time point again distinguishes Mac-1–mediated effects on these two substrates.


A role for Mac-1 (CDIIb/CD18) in immune complex-stimulated neutrophil function in vivo: Mac-1 deficiency abrogates sustained Fcgamma receptor-dependent neutrophil adhesion and complement-dependent proteinuria in acute glomerulonephritis.

Tang T, Rosenkranz A, Assmann KJ, Goodman MJ, Gutierrez-Ramos JC, Carroll MC, Cotran RS, Mayadas TN - J. Exp. Med. (1997)

PMN spreading on  ICs. Samples analyzed in Fig. 4  were also analyzed for PMN  spreading on ICs (A) or BSA alone  (B) and parallel samples were analyzed for F-actin distribution  (C). The spreading in Mac-1–  cells (diamonds) and wild-type  cells (squares) was equivalent at 5  min, but was dramatically reduced at 25 and 40 min in   cells. Representative Wright Giemsa–stained slides from the 40  min time point are shown. Spreading on BSA was significantly different between wild-type and   PMNs at 5 min and remained so  at 40 minutes. The difference in  profiles of PMN spreading on  ICs and BSA alone indicates the  Mac-1 effects on PMN spreading  that are IC-specific. *P <0.005,  #P <0.05 compared to wild-type  cells. (C) Wild-type (WT) and  Mac-1– cells adherent to ICs  for 5 and 40 min were stained  with rhodamine phalloidin to  identify F-actin. At 5 min, cells  of both genotypes initiated actin  polymerization at the periphery  of the cells. By 40 min, wild-type PMNs had developed an actin-rich surface with a punctate  cytoplasmic pattern and had visible projections. In contrast, mutant cells had not progressed to  this stage and several of them had  rounded up (arrow). Original  magnification: ×473.
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Figure 5: PMN spreading on ICs. Samples analyzed in Fig. 4 were also analyzed for PMN spreading on ICs (A) or BSA alone (B) and parallel samples were analyzed for F-actin distribution (C). The spreading in Mac-1– cells (diamonds) and wild-type cells (squares) was equivalent at 5 min, but was dramatically reduced at 25 and 40 min in cells. Representative Wright Giemsa–stained slides from the 40 min time point are shown. Spreading on BSA was significantly different between wild-type and PMNs at 5 min and remained so at 40 minutes. The difference in profiles of PMN spreading on ICs and BSA alone indicates the Mac-1 effects on PMN spreading that are IC-specific. *P <0.005, #P <0.05 compared to wild-type cells. (C) Wild-type (WT) and Mac-1– cells adherent to ICs for 5 and 40 min were stained with rhodamine phalloidin to identify F-actin. At 5 min, cells of both genotypes initiated actin polymerization at the periphery of the cells. By 40 min, wild-type PMNs had developed an actin-rich surface with a punctate cytoplasmic pattern and had visible projections. In contrast, mutant cells had not progressed to this stage and several of them had rounded up (arrow). Original magnification: ×473.
Mentions: To determine whether spreading of mutant PMNs on IC was compromised, we  determined the percentage of spread cells in the samples generated in Fig. 4. Initial spreading on ICs was the same for wild-type and Mac-1– cells at 5 min. The percentage of spread cells increased over time in wild-type cells (Fig. 5 A), consistent with the increased numbers of PMNs adherent to the IC at these later time points as seen in Fig. 4. On the other hand, the percentage of mutant PMNs spread on IC declined rapidly over time and was <10% of wild-type levels at 40 min (Fig. 5 A) and <0.03% at 120 min (data not shown). On BSA-coated surfaces, a difference in spreading was observed between wild-type and cells at 5 min and remained constant up to 40 min (Fig. 5 B). This is consistent with previous studies that indicate a role for Mac-1 in spreading on BSA (1). The equivalent spreading on ICs at 5 min versus the difference in spreading seen on BSA-coated surfaces at this time point again distinguishes Mac-1–mediated effects on these two substrates.

Bottom Line: In vitro studies showed that spreading of Mac-1- PMNs to IC-coated dishes was equivalent to that of wild-type PMNs at 5-12 min but was markedly reduced thereafter, and was associated with an inability of mutant neutrophils to redistribute filamentous actin.Complement C3-deficient mice also had significantly decreased proteinuria compared to wild-type mice.Since Mac-1 on PMNs is the principal ligand for ic3b, an absence of Mac-1 interaction with C3 probably contributed to the abrogation of proteinuria in Mac-1- mice.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Brigham and Women's Hospital, Boston, Massachusetts 02115, USA.

ABSTRACT
Mac-1 (alphambeta2), a leukocyte adhesion receptor, has been shown in vitro to functionally interact with Fcgamma receptors to facilitate immune complex (IC)-stimulated polymorphonuclear neutrophil (PMN) functions. To investigate the relevance of Mac-1-FcgammaR interactions in IC-mediated injury in vivo, we induced a model of Fc-dependent anti-glomerular basement membrane (GBM) nephritis in wild-type and Mac-1-deficient mice by the intravenous injection of anti-GBM antibody. The initial glomerular PMN accumulation was equivalent in Mac-1 and wild-type mice, but thereafter increased in wild-type and decreased in mutant mice. The absence of Mac-1 interactions with obvious ligands, intercellular adhesion molecule 1 (ICAM-1), and C3 complement, is not responsible for the decrease in neutrophil accumulation in Mac-1- deficient mice since glomerular PMN accumulation in mice deficient in these ligands was comparable to those in wild-type mice. In vitro studies showed that spreading of Mac-1- PMNs to IC-coated dishes was equivalent to that of wild-type PMNs at 5-12 min but was markedly reduced thereafter, and was associated with an inability of mutant neutrophils to redistribute filamentous actin. This suggests that in vivo, Mac-1 is not required for the initiation of Fc-mediated PMN recruitment but that Mac-1-FcgammaR interactions are required for filamentous actin reorganization leading to sustained PMN adhesion, and this represents the first demonstration of the relevance of Mac-1-FcgammaR interactions in vivo. PMN-dependent proteinuria, maximal in wild-type mice at 8 h, was absent in Mac-1 mutant mice at all time points. Complement C3-deficient mice also had significantly decreased proteinuria compared to wild-type mice. Since Mac-1 on PMNs is the principal ligand for ic3b, an absence of Mac-1 interaction with C3 probably contributed to the abrogation of proteinuria in Mac-1- mice.

Show MeSH
Related in: MedlinePlus