Biomedical subjects

Anne Nicholson-Weller

Publications and source records attributed to Anne Nicholson-Weller.

7 recordsLinked to original sources

PubMed2005

Complement C3a enhances CXCL12 (SDF-1)-mediated chemotaxis of bone marrow hematopoietic cells independently of C3a receptor.

Complement C3a promotes CXCL12-induced migration and engraftment of human and murine hemopoietic progenitor cells, suggesting a cross-influence between anaphylatoxin and chemokine axes. Here we have explored the underlying mechanism(s) of complement anaphylatoxin and chemokine cooperation. In addition to C3a, C3a-desArg and C4a but not C5a, are potent enhancers of CXCL12-induced chemotaxis of human and murine bone marrow (BM) stem/progenitor cells and B lineage cells. C3a enhancement of chemotaxis is chemokine specific because it is also observed for chemotaxis to CCL19 but not to CXCL13. The potentiating effect of C3a on CXCL12 is independent of the classical C3a receptor (C3aR). First, human BM CD34(+) and B lineage cells do not express C3aR by flow cytometry. Second, the competitive C3aR inhibitor SB290157 does not affect C3a-mediated enhancement of CXCL12-induced chemotaxis. Third, enhancement of chemotaxis of hemopoietic cells is also mediated by C3a-desArg, which does not bind to C3aR. Finally, C3a enhances CXCL12-induced chemotaxis of BM cells from C3aR knockout mice similar to BM cells from wild-type mice. Subsequent studies revealed that C3a increased the binding affinity of CXCL12 to human CXCR4(+)/C3aR(-), REH pro-B cells, which is compatible with a direct interaction between C3a and CXCL12. BM stromal cells were able to generate C3a, C3a-desArg, C4a, as well as CXCL12, suggesting that this pathway could function in vivo. Taken together, we demonstrate a C3a-CXCL12 interaction independent of the C3aR, which may provide a mechanism to modulate the function of CXCL12 in the BM microenvironment.

Anaphylatoxins↗

PubMed2005

Phagocytosis of Salmonella montevideo by human neutrophils: immune adherence increases phagocytosis, whereas the bacterial surface determines the route of intracellular processing.

Complement-opsonized particles become immune adherent to complement receptor 1 (CR1 or CD35) on human erythrocytes, allowing particles to be ingested by phagocytes in the liver and the spleen. We investigated the role that immune adherence plays in the uptake and killing of Salmonella montevideo by human neutrophils. Exposure to serum induced loss of flagella and facilitated immune adherence, which was followed by more-efficient phagocytosis and killing, compared with that after exposure to serum-opsonized, free bacteria. One correlate of bacterial killing is the fusion of phagosomes with lysosomes, which can be monitored by Lyso-Tracker or lysosomal-associated membrane protein 2 colocalization with the intracellular bacteria. At 5 min, phagolysosmal fusion was significantly faster for immune-adherent bacteria than for non-immune-adherent bacteria, but, by 35 min, the difference between the 2 groups was minimal. Immune adherence also facilitated the ingestion of antibody complement-opsonized fluorescent microspheres, but, unlike bacteria, most internalized microspheres failed to fuse with lysosomes. However, addition of lipopolysaccharide, a Toll-like receptor ligand, to microspheres directed their intracellular trafficking, resulting in rapid lysosomal fusion. Thus, immune adherence facilitates phagocytosis, but the route of intracellular processing depends on the molecular nature of the target and is independent of host complement and antibody.

Bacterial Adhesion↗

PubMed2005

Staphylococcus aureus strains that express serotype 5 or serotype 8 capsular polysaccharides differ in virulence.

Most isolates of Staphylococcus aureus produce a serotype 5 (CP5) or 8 (CP8) capsular polysaccharide. To investigate whether CP5 and CP8 differ in their biological properties, we created isogenic mutants of S. aureus Reynolds that expressed CP5, CP8, or no capsule. Biochemical analyses of CP5 and CP8 purified from the isogenic S. aureus strains were consistent with published structures. The degree of O acetylation of each polysaccharide was similar, but CP5 showed a greater degree of N acetylation. Mice challenged with the CP5(+) strain showed a significantly higher bacteremia level than mice challenged with the CP8(+) strain. Similarly, the CP5(+) strain survived preferentially in the bloodstream and kidneys of infected mice challenged with a mixed inoculum containing both strains. The enhanced virulence of the CP5(+) strain in vivo correlated with its greater resistance to in vitro killing in whole mouse blood. Likewise, in vitro opsonophagocytic killing assays with human neutrophils and sera revealed greater survival of the Reynolds (CP5) strain, even though the kinetics of opsonization by C3b and iC3b was similar for both the CP5(+) and CP8(+) strains. Electron micrographs demonstrated C3 molecules on the cell wall beneath the capsule layer for both serotype 5 and 8 strains. Purified CP5 and CP8 stimulated a modest oxidative burst in human neutrophils but failed to activate the alternative complement pathway. These results indicate that CP5 and CP8 differ in a number of biological properties, and these differences likely contribute to the relative virulence of serotype 5 and 8 S. aureus in vivo.

Animals↗

PubMed2004

A role for lipid rafts in C1q-triggered O2- generation by human neutrophils.

Calreticulin, a candidate C1q receptor, was shown recently to be present on the surface of human neutrophils in association with glycosylphosphatidylinositol (GPI) anchored proteins, particularly CD59. In this study, we show that antibodies to CD59, as well as to every other GPI-anchored protein tested, inhibited the C1q-triggered release of O(2)(-) from PMN. Methyl beta cyclodextrin (M beta CD) treatment of the cells to disrupt lipid rafts also prevented C1q-triggered O(2)(-) production. beta(2) integrin-dependent co-stimulation is required for O(2)(-) production from PMN, however M beta CD had no effect on LFA-1 or Mac-1-mediated adhesion, soluble iC3b binding to PMN, or spreading and migration, all of which suggested that PMN integrin function remained intact. Flow cytometric analysis of PMN treated with M beta CD showed upregulation of PMN granule-associated integrins and a corresponding increase in integrin activation-reporter epitopes, in contrast to the decreased expression of GPI-anchored antigens. These data support a model where lipid rafts and their associated GPI-anchored proteins are critical for C1q-triggered O(2)(-) production, consistent with a model where calreticulin serves as the C1q receptor for O(2)(-) production from PMN.

Antibodies↗

PubMed2003

Calreticulin is at the surface of circulating neutrophils and uses CD59 as an adaptor molecule.

Calreticulin, which has been proposed to be a C1q receptor on neutrophils, has neither a transmembrane domain nor a GPI-anchor attachment site and must utilize an adaptor molecule to attach to the plasma membrane. The expression of ecto-calreticulin on purified human neutrophils did not result from contamination by soluble or intracellular calreticulin released during cell fractionation because it was expressed on circulating neutrophils, and the expression did not increase significantly with neutrophil isolation. All neutrophils expressed calreticulin with a unimodal distribution. Treatment of neutrophils with either a cholesterol-binding agent or phosphatidylinositol-specific phospholipase C dramatically decreased ecto-calreticulin expression indicating that the adaptor molecule(s) are located in lipid rafts and have a GPI-anchor. Analysis for the co-expression of specific GPI-anchored proteins and ecto-calreticulin in cells that were deficient in specific GPI-anchored proteins, indicated that ecto-calreticulin was best associated with CD59. Calreticulin reciprocally immunoprecipited with CD59, which provided direct evidence that CD59 is an adaptor for ecto-calreticulin. Immunofluorescence and confocal microscopy demonstrated that ecto-calreticulin co-localized with a fraction of CD59 at the cell surface. Cross-linking ecto-calreticulin with antibodies induced a Ca2+ flux, which suggests that ecto-calreticulin is capable of signaling following ligand binding. Ecto-calreticulin has been associated with diverse cellular functions. An appreciation that the adaptors for ecto-calreticulin are GPI-anchored will provide a framework for understanding any common features underlying ecto-calreticulin ligation.

Adaptor Proteins, Vesicular Transport↗

PubMed2003

Critical role of the complement system in group B streptococcus-induced tumor necrosis factor alpha release.

Group B Streptococcus (GBS) is a major cause of newborn sepsis and meningitis and induces systemic release of tumor necrosis factor alpha (TNF-alpha), believed to play a role in morbidity and mortality. While previous studies have shown that GBS can induce TNF-alpha release from monocytes and macrophages, little is known about the potential modulating effect of plasma or serum on GBS-induced TNF-alpha release, and there are conflicting reports as to the host receptors involved. In a human whole-blood assay system, GBS type III COH-1 potently induced substantial monocyte TNF-alpha release in adult peripheral blood and, due to a higher concentration of monocytes, 10-fold-greater TNF-alpha release in newborn cord blood. Remarkably, GBS-induced TNF-alpha release from human monocytes was enhanced approximately 1000-fold by heat-labile serum components. Experiments employing C2-, C3-, or C7-depleted serum demonstrated that C3 activation via the alternative pathway is crucial for potent GBS-induced TNF-alpha release. Accordingly, whole blood from C3-deficient mice demonstrated significantly reduced GBS-induced TNF-alpha release. Preincubation with human serum enhanced the TNF-alpha-inducing activity of GBS in a C3- and factor B-dependent manner, implying deposition of complement components via the alternative pathway. GBS-induced TNF-alpha release was inhibited by monoclonal antibodies directed against each of the components of CR3 and CR4: the common integrin beta subunit CD18 and the alpha subunits CD11b (of CR3) and CD11c (of CR4). Blood derived from CR3 (CD11b/CD18)-deficient mice demonstrated a markedly diminished TNF-alpha response to GBS. We conclude that the ability of plasma and serum to greatly amplify GBS-induced TNF-alpha release reflects the activity of the alternative complement pathway that deposits fragments on GBS and thereby enhances CR3- and CR4-mediated monocyte activation.

Adult↗

PubMed2002

Expression and function of C1q receptors and C1q binding proteins at the cell surface.

C1q is the recognition unit of the first component of complement that binds not only IgG and IgM containing immune complexes, but also recognizes foreign structures such as the lipid A of endotoxin, and molecules expressed at the surface of apoptotic cells. In this review, the plasma membrane receptors and binding proteins for C1q are discussed and new data are presented on calreticulin expression on human peripheral blood cells. Although much is known about C1q receptors and binding molecules there are still many questions regarding their role in vivo.

Animals↗