Decreased mutation frequencies among immunoglobulin G variable region genes during viremic HIV-1 infection.

BACKGROUND/OBJECTIVE: HIV-1 infection is complicated by high rates of opportunistic infections against which specific antibodies contribute to immune defense. Antibody function depends on somatic hypermutation (SHM) of variable regions of immunoglobulin heavy chain genes (VH-D-J). We characterized the frequency of SHM in expressed IgG mRNA immunoglobulin transcripts from control and HIV-1-infected patients. DESIGN: We compared utilization of genes in the most prominent VH family (VH3) and mutation frequencies and patterns of cDNA from VH3-IgG genes from 10 seronegative control subjects and 21 patients with HIV-1 infection (6 without and 15 patients with detectable plasma viremia). METHODS: Unique IgG VH3 family cDNA sequences (n = 1,565) were PCR amplified, cloned, and sequenced from blood. Sequences were analyzed using online (Vbase) and in-house immunoglobulin alignment resources. RESULTS: Mutation frequencies in the antigen-binding hypervariable complementarity determining regions (CDR1/2) of IgG class-switched B cells were lower among viremic HIV-1-infected patients vs. controls for nucleotides (CDR1/2: 10±5% vs. 13.5±6%, p = 0.03) and amino acids (CDR: 20%±10 vs. 25%±12, p = 0.02) and in structural framework regions. Mutation patterns were similar among groups. The most common VH3 gene, VH3-23, was utilized less frequently among viremic HIV-1-infected patients (p = 0.03), and overall, mutation frequencies were decreased in nearly all VH3 genes compared with controls. CONCLUSIONS: B cells from HIV-1-infected patients show decreased mutation frequencies, especially in antigen-binding VH3 CDR genes, and selective defects in gene utilization. Similar mutation patterns suggest defects in the quantity, but not quality, of mutator activity. Lower levels of SHM in IgG class-switched B cells from HIV-1-infected patients may contribute to the increased risk of opportunistic infections and impaired humoral responses to preventative vaccines.

Species specialization in cytokine biology: is interleukin-4 central to the T(H)1-T(H)2 paradigm in swine?

The TH1-TH2 paradigm provides an elegant model of directed response to infectious pathogens. Developed in the mouse, the model has provided a framework for systematic and mechanistic studies of immune regulation, protective immunity, and vaccine development in swine. Interleukin-4 (IL-4) plays a central role in the paradigm as a regulatory molecule directing development of the TH2 phenotype, as a developmental cytokine essential for antibody production, and as a soluble diagnostic marker of the TH2 cell type. In contrast, while characterizing the biological properties of porcine IL-4, we discovered that it was not a stimulatory factor for porcine B cells. Rather, it blocked antibody and IL-6 secretion and suppressed antigen-stimulated proliferation of B cells. Inhibition was not reversed by treatment with IL-2 and IL-6 treatment. IL-4 did not stimulate T lymphocyte proliferation, but induced cell growth in lymphoblasts in a dose-dependent fashion. These results suggest that IL-4 plays a different role in pigs than in mice and humans, in which it stimulates B cells and is essential for antibody production. Furthermore, the functions of IL-4 in swine cannot be inferred from results in model systems such as the mouse. General models of disease resistance show substantial variation between pigs and mice at the cellular and molecular level. Advances in somatic cell technologies and animal engineering to enable gene knockouts in pigs, in combination with a continuously expanding immunological toolkit, promise an exciting future for pig immunology, detailed mechanistic elucidation of the TH1-TH2 paradigm, and an improved understanding of the role of IL-4 in porcine immunity to infectious disease.

Antibody-enhanced pneumococcal adherence requires IgA1 protease.

IgA, the major class of Ig in secretions, classically functions by interfering with microbial attachment to host tissues. Many mucosal pathogens, including Streptococcus pneumoniae, express an IgA1 protease that may circumvent the protective effects of this Ig subclass. Because these proteases are specific for human IgA1, we generated human mAbs to the major surface antigen of the pneumococcus, its capsular polysaccharide, and tested their effect in a colonization model of bacterial adherence to respiratory epithelial cells in culture. Rather than inhibiting adherence, type-specific IgA1 markedly enhanced bacterial attachment to host cells, but only when cleaved by IgA1 protease. Neither antibodies of protease-insensitive subclasses (IgA2 and IgG) nor those directed against heterologous capsules had such activity. The adherence-promoting properties of cleaved antibodies correlated with the cationic characteristics of their variable segments, suggesting that bound Fab fragments may neutralize the inhibitory effect of negatively charged capsules on adhesive interaction with host cells. Coating of pneumococci with anticapsular polysaccharide antibody unmasked the bacterial phosphorylcholine ligand, allowing for increased adherence mediated by binding to the platelet activating factor receptor on epithelial cells. In addition, our findings provide evidence for a novel function of bacterial IgA1 proteases. These enzymes may enable pathogens to subvert the antigen specificity of the humoral immune response to facilitate adhesive interactions and persistence on the mucosal surface.

Mucosal plasma cell repertoire during HIV-1 infection.

Impaired development of local Ab responses may predispose HIV-1-infected patients to an increased rate, severity, and duration of mucosal infections. We characterized the repertoire of Ig-producing cells in the intestinal effector compartment (the lamina propria) of HIV-1-infected (n = 29) and seronegative control (n = 27) subjects. The density of Ig-producing cells per area was similar in both groups. However, the proportions of IgA-producing cells were lower in both the duodenum and colon from HIV-1-infected patients compared with those of control subjects (p < 0.05), with compensatory increases in IgG-producing cells in the colon and IgM-producing cells in the duodenum. Similarly, among Abs in the lumen the proportions of IgA were also decreased and the proportions of IgG were increased among HIV-1-infected patients. On a molecular level, V(H) gene repertoire analyses by RT-PCR revealed comparable proportions of the V(H)3 family among duodenal IgA transcripts (50-53%) from both groups. V(H)3 expression was decreased only for IgM among patients with advanced HIV-1 disease (n = 6) compared with that of control subjects (n = 8) (48 +/- 8 vs 62 +/- 13%; p < 0.01). Moreover, the frequencies of individual IgM and IgA V(H)3 genes were comparable in each group, including rates of putative HIV-1 gp120-binding V(H)3 genes (V3-23, V3-30, V3-30/3-30.5). We conclude that, despite a decrement in local IgA producing cells, the density and molecular V(H) repertoire of mucosal plasma cells are relatively intact among patients with HIV-1 infection. These data suggest that HIV-1-infected patients use functional regulatory mechanisms to provide sufficient V(H) diversity and effective induction and differentiation of mucosal B cells.