Short Communication: Nonprogressive HIV-1 Infection Is Associated with Expansion of IL-21R Expressing Class-Switched Memory B Cells.

HIV perturbs the functionality of B cells resulting in defective humoral responses. As efficient humoral immune responses are important in controlling HIV-disease progression, we characterized the memory B cell population for its subsets and their activation (CD38 expression) and functional [interleukin (IL)-21R expression] profile in individuals with nonprogressive [long-term nonprogressors (LTNPs), N = 16] and progressive HIV disease (progressors, N = 16) along with 10 HIV uninfected healthy controls (HCs). The frequencies of total memory B cells were similar in HCs and HIV-infected individuals, whereas the frequencies of unswitched memory B (UMB) cells and CD38+ UMB cells were significantly higher in progressors than LTNPs and HCs (p < .03). LTNPs showed higher frequencies of class-switched memory B (SMB) cells and IL-21R expressing SMB cells than seen in progressors (p = .019), which were similar to that seen in HCs. The %UMB cells correlated inversely (p = .0002, r = -0.6053) and %SMB cells correlated positively (p = .0005, r = 0.5804) with CD4 count. IL-21/IL-21R interaction is required for class switching of B cells and differentiation into antibody-secreting plasma cells. The higher expression of IL-21R on class SMB cells from LTNPs might be resulting in efficient plasma cell differentiation and the functional humoral immune response that might be responsible for mounting efficient antibody response against the encountered infections. The more insights in this area might be required to further understand the role of IL-21R expressing class SMB cells in HIV infection.

CpG oligodeoxynucleotide CpG-685 upregulates functional interleukin-21 receptor on chronic lymphocytic leukemia B cells through an NF-κB mediated pathway.

CpG oligodeoxynucleotides (ODNs) upregulate the interleukin-21 receptor (IL21R) and enhance IL-21-mediated cytotoxicity in chronic lymphocytic leukemia (CLL) B cells. We demonstrate that treatment of CLL B cells with the ODN CpG-685 leads to increased IL21R expression, and that this increased expression enhances the effects of IL-21 treatment as evidenced by increased phosphorylation of JAK1, STAT1, and STAT3, as compared to IL-21 treatment without prior CpG stimulation. Induction of IL21R by CpG-685 also enhanced IL-21-mediated cytotoxicity. The mechanism by which CpG ODNs upregulate IL21R has not been elucidated, although IL21R regulation in T cells has been shown to be linked to T cell receptor-induced Sp1 binding to the IL21R promoter. Here, we demonstrate that luciferase reporter constructs containing the Sp1 binding site have increased basal luciferase activity compared to constructs lacking the Sp1 binding site, but fail to increase luciferase activity with CpG-685 stimulation in CLL B cells. By treating CLL cells with an NF-κB inhibitor, we inhibit the CpG ODN-mediated induction of IL21R, thus demonstrating that CpG-685 upregulates IL21R through an NF-κB mediated pathway. These findings suggest an alternative mechanism for induction of IL-21 receptor in CLL B cells and provide a basis for creation of future combination therapies.

Upregulated interleukin-21 receptor on B cells associated with the downregulation of IgE in patients with allergic rhinitis.

Allergic diseases, such as allergic rhinitis, caused by an immunoglobulin E (IgE)-mediated reaction to specific allergens are a common chronic condition worldwide. Interleukin-21 (IL-21), a type I cytokine that is produced by T cells, exerts regulatory effects on a variety of immune cells. In our previous study, we found that serum levels of IL-21 were significantly decreased in patients with severe atopic dermatitis, suggesting that IL-21 might play a role in allergic reactions. In this study, we investigated the role of IL-21/IL-21 receptor (IL-21R) in patients with allergic rhinitis. Our results demonstrated that there was no difference in IL-21 serum levels between allergic rhinitis patients and controls. However, allergic patients had significantly increased expression of IL-21R on naive and memory B cells. IL-21R was upregulated through stimulation by the combination of CD40 ligand (CD40L) and IL-4. IL-21 alone neither induced nor inhibited IgE secretion from CD40L-stimulated B cells. However, IL-21 inhibited IgE secretion of B cells that were induced by the combination of CD40L and IL-4 in allergic patients. Moreover, a negative correlation between the expression of IL-21R and serum levels of IgE was found in patients with allergy. These results suggest that the role of IL-21 in an ongoing allergic reaction is to downregulate the IgE level by binding to IL-21R on B cells, which increases the expression in allergic patients.

Potential role of myeloid cell/eosinophil-derived IL-17 in LPS-induced endotoxin shock.

IL-17RA is a shared receptor subunit for several cytokines of the IL-17 family, including IL-17A, IL-17C, IL-17E (also called IL-25) and IL-17F. It has been shown that mice deficient in IL-17RA are more susceptible to sepsis than wild-type mice, suggesting that IL-17RA is important for host defense against sepsis. However, it is unclear which ligands for IL-17RA, such as IL-17A, IL-17C, IL-17E/IL-25 and/or IL-17F, are involved in the pathogenesis of sepsis. Therefore, we examined IL-17A, IL-17E/IL-25 and IL-17F for possible involvement in LPS-induced endotoxin shock. IL-17A-deficient mice, but not IL-25- or IL-17F-deficient mice, were resistant to LPS-induced endotoxin shock, as compared with wild-type mice. Nevertheless, studies using IL-6-deficient, IL-21Rα-deficient and Rag-2-deficient mice, revealed that neither IL-6 and IL-21, both of which are important for Th17 cell differentiation, nor Th17 cells were essential for the development of LPS-induced endotoxin shock, suggesting that IL-17A-producing cells other than Th17 cells were important in the setting. In this connection, IL-17A was produced by macrophages, DCs and eosinophils after LPS injection. Taken together, these findings indicate that IL-17A, but not IL-17F or IL-25, is crucial for LPS-induced endotoxin shock. In addition, macrophages, DCs and eosinophils, but not Th17 cells or γδ T cells, may be sources of IL-17A during LPS-induced endotoxin shock.

IL-21 enhances phagocytosis in mononuclear phagocyte cells: identification of spleen tyrosine kinase as a novel molecular target of IL-21.

The biological significance of the IL-21/IL-21R system in human monocytes/macrophages is not well documented, and the expression of IL-21R is unclear and has been disputed. In this study, we showed for the first time, to our knowledge, that human monocyte-like THP-1 cells expressed the two IL-21R components, CD132 (γc) and IL-21Rα, on their cell surface, as assessed by flow cytometry. Moreover, IL-21 was found to enhance FcR-mediated phagocytosis, but not endocytosis. The ability of IL-21 to enhance phagocytosis was not associated with an increased expression of both IL-21R components at the cell surface, and IL-21 did not act in synergy with IL-15. IL-21 activated spleen tyrosine kinase (Syk), as evidenced by its ability to increase Syk phosphorylation. Using a pharmacological approach to inhibit Syk activity, and an antisense technique to downregulate Syk protein expression, we demonstrated the importance of Syk in IL-21-induced phagocytosis. In addition, both CD132 and IL-21Rα were expressed on the cell surface of naive monocytes, as well as in GM-CSF-monocyte-derived macrophages. Moreover, IL-21 also induced phagocytosis in these cells. We conclude that IL-21 possesses important biological effects in mononuclear phagocyte cells and that Syk is a novel molecular target of IL-21 that was previously unknown. Therefore, future development of therapeutic strategies targeting the IL-21/IL-21R system should consider that monocyte and macrophage cell physiology may be affected by this system.

IL-21 promotes lupus-like disease in chronic graft-versus-host disease through both CD4 T cell- and B cell-intrinsic mechanisms.

T cell-driven B cell hyperactivity plays an essential role in driving autoimmune disease development in systemic lupus erythematosus. IL-21 is a member of the type I cytokine family with pleiotropic activities. It regulates B cell differentiation and function, promotes T follicular helper (T(FH)) cell and Th17 cell differentiation, and downregulates the induction of T regulatory cells. Although IL-21 has been implicated in systemic lupus erythematosus, the relative importance of IL-21R signaling in CD4(+) T cells versus B cells is not clear. To address this question, we took advantage of two induced models of lupus-like chronic graft-versus-host disease by using wild-type or IL-21R(-/-) mice as donors in the parent-into-F1 model and as hosts in the Bm12→B6 model. We show that IL-21R expression on donor CD4(+) T cells is essential for sustaining T(FH) cell number and subsequent help for B cells, resulting in autoantibody production and more severe lupus-like renal disease, but it does not alter the balance of Th17 cells and regulatory T cells. In contrast, IL-21R signaling on B cells is critical for the induction and maintenance of germinal centers, plasma cell differentiation, autoantibody production, and the development of renal disease. These results demonstrate that IL-21 promotes autoimmunity in chronic graft-versus-host disease through both CD4(+) T cell- and B cell-intrinsic mechanisms and suggest that IL-21 blockade may attenuate B cell hyperactivity, as well as the aberrant T(FH) cell pathway that contributes to lupus pathogenesis.

Heterogeneous expression and function of IL-21R and susceptibility to IL-21-mediated apoptosis in follicular lymphoma cells.

OBJECTIVE: Interleukin (IL)-21, a member of the IL-2 family, has antitumor activity and is now being tested in non-Hodgkin's lymphoma in combination with anti-CD20 antibodies. IL-21 may either induce apoptosis or promote growth in different lymphoid malignancies. We therefore investigated the IL-21/IL-21R system in follicular lymphoma (FL) cells. MATERIALS AND METHODS: IL-21R expression was studied by reverse transcription polymerase chain reaction, immunofluorescence, and Western blot analyses. Apoptosis was measured by Annexin-V-propidium iodide staining. Signaling via IL-21R was studied using antibodies specific for phosphorylated Janus-activating kinase and signal transducers and activators of transcription proteins by Western Blot. RESULTS: IL-21R was found on primary FL cells in 15 of 15 cases at diagnosis and IL-21 increased apoptosis in 10 of 10 FL samples. However, cells from areas of diffuse growth in FL and from two diffuse lymphomas evolved from previous FL, showed low IL-21R expression. The latter were also resistant to IL-21-mediated apoptosis. Among lymphoma cell lines bearing the t(14;18) translocation, only 1 of 7 showed increased apoptosis in response to IL-21 stimulation. This cell line was IL-21R-positive, whereas five of six nonresponsive cell lines showed very low IL-21R expression. Intriguingly, one of the IL-21-resistant cell lines (DOHH2) expressed high levels of IL-21R. Treatment with IL-21 or IL-4 upregulated suppressor of cytokine signaling 3 gene expression in the IL-21-responsive cell line, but not in DOHH2 cells, which showed defective Janus-activating kinase/signal transducers and activators of transcription signaling in response to IL-21, in relationship to the lack of Janus-activating kinase 3 gene expression. CONCLUSION: These data indicate that low IL-21R expression or defective signal transduction downstream IL-21R may cause refractoriness to IL-21-mediated effects in some FL cells.

Regulation of interleukin-21 receptor expression and its signal transduction by WSB-2.

Interleukin-21 (IL-21) is a pleiotropic cytokine that regulates T-cell, B-cell, NK-cell, and myeloid-cell functions. IL-21 binds with its cognate receptor complex, which consists of the IL-21 receptor (IL-21R) and the common gamma chain (gammac) receptor subunit. We identified novel IL-21R-binding molecule, WD-40 repeats containing SOCS-box-2, WSB-2. WSB-2 associated with the membrane-proximal intracytoplasmic region of IL-21R, including box1 and box2. Overexpression study of WSB-2 showed the reduction of IL-21R expression and IL-21-induced signal transduction. On the other hand, small interfering RNA for WSB-2 enhanced the expression level of IL-21R and IL-21-induced STAT3 activation, indicating that WSB-2 negatively controls the receptor expression. This report provides the first evidence that WSB-2 is a regulator of IL-21R expression and IL-21-induced signal transduction.

Transcriptional activation of the interleukin-21 gene and its receptor gene by human T-cell leukemia virus type 1 Tax in human T-cells.

At the incipient stages of the development of adult T-cell leukemia, T-cells infected with human T-cell leukemia virus type 1 (HTLV-1) suffer disregulation in cell growth caused by aberrant expression of host genes by the HTLV-1 transactivator protein Tax (Tax1). Tax1-mediated growth promotion is thought to result from, at least in part, up-regulation of genes for growth factors and their receptors that induce T-cell growth. In the present study, we demonstrate that Tax1 transactivates the interleukin-21 (IL-21) and its receptor (IL-21R) genes in human T-cells. Introduction of Tax1 via recombinant adenoviruses induced expression of endogenous IL-21 and IL-21R. Isolated promoters of the IL-21 and IL-21R genes were activated by Tax1 in reporter assays, which further revealed that there were at least two Tax1-responsive elements in either the IL-21 promoter or the IL-21R promoter. Chromatin immunoprecipitation assay and gel mobility shift assay exhibited that the IL-21 promoter elements bound transcription factors AP-1 and NF-kappaB, and the IL-21R promoter elements were associated with AP-1 and interferon regulatory factor. Collectively, Tax1-dependent activation of these transcriptional factors presumably contributes to expression of the IL-21 gene and its receptor gene. The related virus HTLV-2 with Tax2 similar to Tax1 is known not to be pathogenic. Tax2 exhibited little, if any, or no induction of the IL-21 transcription in CD4+ T-cells, in contrast to Tax1. The study suggests insights into cytokine-dependent aberrant growth of HTLV-1-infected T-cells and the molecular basis of different pathogenicity between HTLV-1 and HTLV-2.

IL-21 induces apoptosis of antigen-specific CD8+ T lymphocytes.

IL-21, a member of the common gamma-chain family of cytokines, has pleiotropic effects on T, B, and NK cells. We found that IL-21 and the prototype common gamma-chain cytokine IL-2 can stimulate proliferation and cytokine secretion by Ag-specific rhesus monkey CD8+ T cells. However, unique among the members of this family of cytokines, we found that IL-21 drives these cells to apoptosis by down-regulation of Bcl-2. These findings suggest that IL-21 may play an important role in the contraction of CD8+ T cell responses.