Modulation of the Interleukin-21 Pathway with Interleukin-4 Distinguishes Common Variable Immunodeficiency Patients with More Non-infectious Clinical Complications.

PURPOSE: Common variable immunodeficiency (CVID) is characterized by hypogammaglobulinemia and clinical manifestations such as infections, autoimmunity, and malignancy. We sought to determine if responsiveness to interleukin-21 (IL-21), a key cytokine for B cell differentiation, correlates with distinct clinical phenotypes in CVID. METHODS: CVID subjects were recruited through the Canadian Primary Immunodeficiency Evaluative Survey registry. Peripheral blood mononuclear cells were cultured with anti-CD40 ± interferon-gamma, interleukin-4 (IL-4), IL-21, and/or IL-4+IL-21. B cell subpopulations and IgG production were determined at baseline and day 7 by flow cytometry and ELISA. Clinical complications were compared using contingency tables. RESULTS: CVID subjects exhibited decreased CD27+ B cells and IgG production after 7 days of stimulation with anti-CD40+IL-21 (p < 0.05). In a subset of subjects [CVID responders (R)], the addition of IL-4 led to significant increases in CD27+ B cells and IgG (p < 0.05). In CVID non-responders (NR), CD27+ B cells and IgG remained lower despite the addition of IL-4. CVID NR experienced significantly more non-infectious clinical complications of CVID than R [OR 8.8, 95% confidence interval (CI) 1.6 to 48.13]. Previous studies reported that CVID subjects with ≤ 2% class-switched memory B cells were more at risk of these complications, but no significant association was found among this cohort of subjects [OR 3.5, CI 0.9 to 13.3]. In fact, 34.6% of CVID NR had > 2% class-switched memory B cells at baseline. CONCLUSIONS: The IL-4 and IL-21 in vitro assays distinguish two groups of CVID subjects and can be used with baseline B cell subpopulation phenotyping to better identify patients experiencing more vs. fewer clinical non-infectious complications and potentially to modulate therapy.

Corrigendum: Semaphorin 4C: A Novel Component of B-Cell Polarization in Th2-Driven Immune Responses.

[This corrects the article on p. 558 in vol. 7, PMID: 28003812.].

Peripherally Generated Foxp3+ Regulatory T Cells Mediate the Immunomodulatory Effects of IVIg in Allergic Airways Disease.

IVIg is widely used as an immunomodulatory therapy. We have recently demonstrated that IVIg protects against airway hyperresponsiveness (AHR) and inflammation in mouse models of allergic airways disease (AAD), associated with induction of Foxp3+ regulatory T cells (Treg). Using mice carrying a DTR/EGFP transgene under the control of the Foxp3 promoter (DEREG mice), we demonstrate in this study that IVIg generates a de novo population of peripheral Treg (pTreg) in the absence of endogenous Treg. IVIg-generated pTreg were sufficient for inhibition of OVA-induced AHR in an Ag-driven murine model of AAD. In the absence of endogenous Treg, IVIg failed to confer protection against AHR and airway inflammation. Adoptive transfer of purified IVIg-generated pTreg prior to Ag challenge effectively prevented airway inflammation and AHR in an Ag-specific manner. Microarray gene expression profiling of IVIg-generated pTreg revealed upregulation of genes associated with cell cycle, chromatin, cytoskeleton/motility, immunity, and apoptosis. These data demonstrate the importance of Treg in regulating AAD and show that IVIg-generated pTreg are necessary and sufficient for inhibition of allergen-induced AAD. The ability of IVIg to generate pure populations of highly Ag-specific pTreg represents a new avenue to study pTreg, the cross-talk between humoral and cellular immunity, and regulation of the inflammatory response to Ags.

Semaphorin 4C Protects against Allergic Inflammation: Requirement of Regulatory CD138+ Plasma Cells.

The regulatory properties of B cells have been studied in autoimmune diseases; however, their role in allergic diseases is poorly understood. We demonstrate that Semaphorin 4C (Sema4C), an axonal guidance molecule, plays a crucial role in B cell regulatory function. Mice deficient in Sema4C exhibited increased airway inflammation after allergen exposure, with massive eosinophilic lung infiltrates and increased Th2 cytokines. This phenotype was reproduced by mixed bone marrow chimeric mice with Sema4C deficient only in B cells, indicating that B lymphocytes were the key cells affected by the absence of Sema4C expression in allergic inflammation. We determined that Sema4C-deficient CD19+CD138+ cells exhibited decreased IL-10 and increased IL-4 expression in vivo and in vitro. Adoptive transfer of Sema4c-/- CD19+CD138+ cells induced marked pulmonary inflammation, eosinophilia, and increased bronchoalveolar lavage fluid IL-4 and IL-5, whereas adoptive transfer of wild-type CD19+CD138+IL-10+ cells dramatically decreased allergic airway inflammation in wild-type and Sema4c-/- mice. This study identifies a novel pathway by which Th2-mediated immune responses are regulated. It highlights the importance of plasma cells as regulatory cells in allergic inflammation and suggests that CD138+ B cells contribute to cytokine balance and are important for maintenance of immune homeostasis in allergic airways disease. Furthermore, we demonstrate that Sema4C is critical for optimal regulatory cytokine production in CD138+ B cells.

Semaphorin 4C: A Novel Component of B-Cell Polarization in Th2-Driven Immune Responses.

BACKGROUND: Semaphorins are important molecules in embryonic development and multiple semaphorins have been identified as having key roles in immune regulation. To date, there is little known about Semaphorin 4C (Sema4C) in immune biology. We report for the first time that Sema4C is inducible in human and murine B-cells and may be important for normal B-cell development. METHODS: Human tonsillar B-cells were studied following activation via anti-CD40 antibodies in the presence or absence of representative Th1, Th2, and regulatory cytokines. Murine B-cells from WT and Sema4C-/- mice were similarly stimulated. B-cell phenotyping in WT and Sema4C mutant mice was performed by flow cytometry and lymphoid architecture was studied by immunohistochemistry. Sema4C expression and synapse formation were analyzed by confocal microscopy. RESULTS: Gene array studies performed on human tonsillar B-cells stimulated to produce IgE revealed that Sema4C was among the top genes expressed at 24 h, and the only semaphorin to be increased under Th2 conditions. Validation studies demonstrated that human and murine B-cells expressed Sema4C under similar conditions. Sema4C-/- mice had impaired maturation of B-cell follicles in spleens and associated decreases in follicular and marginal zone B-cells as well as impaired IgG and IgA production. In keeping with a potential role in maturation of B-cells, Sema4C was expressed predominantly on CD27+ human B-cells. Within 72 h of B-cell activation, Sema4C was localized to one pole in a synapse-like structure, in association with F-actin, B-cell receptor, and Plexin-B2. Cell polarization was impaired in Sema4C-/- mice. CONCLUSION: We have identified a novel immune semaphorin induced in human and murine B-cells under Th2 conditions. Sema4C appears to be a marker for human memory B-cells. It may be important for B-cell polarization and for the formation of normal splenic follicles.

CC and CXC chemokines induce airway smooth muscle proliferation and survival.

The increase in airway smooth muscle (ASM) mass is a major structural change in asthma. This increase has been attributed to ASM cell (ASMC) hyperplasia and hypertrophy. The distance between ASMC and the epithelium is reduced, suggesting migration of smooth muscle cells toward the epithelium. Recent studies have suggested a role of chemokines in ASMC migration toward the epithelium; however, chemokines have other biological effects. The objective of the current study is to test the hypothesis that chemokines (eotaxin, RANTES, IL-8, and MIP-1α) can directly influence ASMC mass by increasing the rate of proliferation or enhancing the survival of these cells. Human ASMCs were exposed to different concentrations of eotaxin, RANTES, IL-8, or MIP-1α. To test for proliferation, matched control and stimulated ASMC were pulsed with [(3)H]thymidine, or ASMCs were stained with BrdU and then analyzed with flow cytometry. Apoptosis was measured using Annexin V staining and flow cytometry. Expression of phosphorylated p42/p44 and MAPKs was assessed by Western blot. In a concentration-dependent manner, chemokines including eotaxin, RANTES, IL-8, and MIP-1α increased ASMC's [(3)H]thymidine incorporation and DNA synthesis. IL-8, eotaxin, and MIP-1α decreased the rate of apoptosis of ASMCs compared with the matched controls. A significant increase in phosphorylated p42/p44 MAPKs was seen after treating ASMCs with RANTES and eotaxin. Moreover, inhibition of p42/p44 MAPK phosphorylation reduced the level of chemokine-induced ASM proliferation. We conclude that chemokines might contribute to airway remodeling seen in asthma by enhancing the number and survival of ASMCs.