IgG3 anti-Kell allotypic variation results in differential antigen binding and phagocytosis.

BACKGROUND: Human immunoglobulin G (hIgG) includes four different subtypes (IgG1, IgG2, IgG3, and IgG4). Due to genetic variations, each IgG subtype contains different isoallotypes. It was previously shown that a Food and Drug Administration-approved monoclonal anti-IgG failed to recognize 2 of 15 recombinant, human IgG3 anti-Kell (K1) isoallotypes (rIgG3-03 and rIgG3-13) by indirect antiglobulin test (IAT). STUDY DESIGN AND METHODS: We expressed and purified 15 recombinant human rIgG3 anti-K1 isoallotypes and investigated their antigen binding and ability to induce phagocytosis using homozygous (KK) and heterozygous (Kk) K1-positive red blood cells (RBCs) by gel IAT, flow cytometry, and a monocyte monolayer assay (MMA) with peripheral blood monocytes and cultured inflammatory (M1) and anti-inflammatory (M2) macrophages. RESULTS: MMA results showed that differences in the Fc region of rIgG3 anti-K1 led to distinctive phagocytic activity with both monocytes and M1 macrophages. rIgG3-18 and rIgG3-19 showed an enhanced ability to induce phagocytosis. Differences in Fc regions also led to variations in the number of antibodies bound to KK RBCs. Despite the differences in phagocytic activity, all 15 rIgG3 clones are predicted to induce clinically significant hemolysis if K1-positive blood was transfused into patients. CONCLUSION: These results argue that antiglobulin reagents that fail to detect isoallotype rIgG3-03 or rIgG3-13 could present a transfusion risk or lack of detection of a potentially clinically significant anti-K1 in hemolytic disease of the fetus and newborn.

Isoagglutinin-reduced immunoglobulin retains efficacy in mouse models of immune thrombocytopenia and rheumatoid arthritis and is less likely to cause intravenous immunoglobulin-associated hemolysis.

BACKGROUND: Immunoglobulin therapy including intravenous immunoglobulin (IVIg) has been used as an effective treatment for autoimmune/inflammatory conditions with few side effects. However, high-dose IVIg (1-2 g/kg) has been recognized as a cause of hemolytic anemia in non-blood group O patients. Hemolysis when observed has been due to anti-A/anti-B isoagglutinins contained in the IVIg. Recently, an isoagglutinin-reduced IVIg, whereby the anti-A and anti-B titers have been reduced by immunoaffinity chromatography, has been introduced; however, whether this new product is as efficacious as nonreduced immunoglobulin (Ig) or will result in less IVIg-associated hemolysis has not been resolved. STUDY DESIGN AND METHODS: We used in vitro phagocytosis by monocytes and proinflammatory/anti-inflammatory macrophages, with isoagglutinin-reduced and -nonreduced Ig opsonized group A1 , B, and A1 B red blood cells, to estimate clinical significance of the IgG isoagglutinins. We also used immune thrombocytopenia (ITP) and rheumatoid arthritis (RA) mouse models to examine the in vivo efficacy of isoagglutinin-reduced versus -nonreduced Ig on the amelioration of the diseases. RESULTS: In contrast to nonreduced Ig, phagocytosis was largely absent when isoagglutinin-reduced Ig was used at a concentration equivalent to a patient receiving 2 g/kg. The in vivo efficacy of isoagglutinin-reduced versus nonreduced Ig on the amelioration of experimental ITP and RA was similar, indicating no loss of efficacy due to the chromatographic removal of isoagglutinins. CONCLUSION: Isoagglutinin-reduced Ig should have efficacy similar to nonreduced Ig and result in less IVIg-associated hemolysis.

Differential regulation of IgA+ B cells in vitro by stromal cells from distinctive anatomical compartments.

B cell development is regulated by stromal cells (SCs) that form a supportive microenvironment. These SCs along with other cell types produce cytokines, chemokines, and adhesion molecules that guide B cell commitment and differentiation. BM, spleen (Sp), and the gut lamina propria (LP) constitute distinctive anatomical compartments that support B cell differentiation. In order to characterize and compare the signals necessary to generate IgA+ B cells, we developed an in vitro system to co-culture gut LP, BM, or Sp-derived SCs with B lineage cells. Using this co-culture system, we found that gut LP SCs promote IgA+ B cell accumulation through the production of soluble stimulatory factors. In contrast to gut LP SCs, BM and splenic SCs were found to impair IgA+ B cell accumulation in vitro. Taken together, these observations provide new insights into how SCs derived from different anatomical locations shape IgA+ B cell responses.

Inflammation rapidly reorganizes mouse bone marrow B cells and their environment in conjunction with early IgM responses.

Systemic inflammation perturbs the bone marrow environment by evicting resident B cells and favoring granulopoiesis over lymphopoiesis. Despite these conditions, a subset of marrow B cell remains to become activated and produce potent acute immunoglobulin M (IgM) responses. This discrepancy is currently unresolved and a complete characterization of early perturbations in the B-cell niche has not been undertaken. Here, we show that within a few hours of challenging mice with adjuvant or cecal puncture, B cells accumulate in the bone marrow redistributed away from sinusoid vessels. This response correlates with enhanced sensitivity to CXC chemokine ligand 12 (CXCL12) but not CXCL13 or CC chemokine ligand 21. Concurrently, a number of B-cell survival and differentiation factors are elevated to produce a transiently supportive milieu. Disrupting homing dynamics with a CXC chemokine receptor 4 inhibitor reduced the formation of IgM-secreting cells. These data highlight the rapidity with which peripheral inflammation modifies the marrow compartment, and demonstrate that such modifications regulate acute IgM production within this organ. Furthermore, our study indicates that conversion to a state of emergency granulopoiesis is temporally delayed, allowing B cells opportunity to respond to antigen.