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Self-renewal programs in leukemia stem cells (LSCs) predict poor prognosis in patients with acute myeloid leukemia (AML). We identify CD4 + T cell-derived interleukin (IL)-21 as an important negative regulator of self-renewal of LSCs. IL-21/IL-21R signaling favors asymmetric cell division and differentiation in LSCs through the activation of p38-MAPK signaling, resulting in reduced LSC numbers and significantly prolonged survival in murine AML models. In human AML, serum IL-21 at diagnosis is identified as an independent positive prognostic biomarker for outcome and correlates with improved survival and higher complete remission rates in patients that underwent high-dose chemotherapy. IL-21 treatment inhibits primary LSC function and enhances the effect of cytarabine and CD70 CAR T cell treatment on LSCs in vitro. Low-dose IL-21 treatment prolongs the survival of AML mice in syngeneic and xenograft experiments. Therefore, promoting IL-21/IL-21R signaling on LSCs may be an approach to reduce stemness and increase differentiation in AML., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

IL-21/IL-21R signaling dysregulation is linked to multiple chronic intestinal inflammatory disorders in humans and animal models of human diseases. In addition to its critical requirement for the generation and development of germinal center B cells, IL-21/IL-21R signaling can also regulate the effector functions of a variety of T-cell subsets. The antibody-mediated abrogation of IL-21/IL-21R signaling led to the impaired expression of IFN-γ by mucosal CD4+ T cells from human subjects with colitis, suggesting an IL-21/IL-21R-triggered positive feedback loop of the TH1 immune response in the colon. Despite recent advances in our understanding of the mechanisms underpinning the regulation of proinflammatory immune responses by the IL-21/IL-21R signaling axis, it remains unclear how this pathway or its downstream molecules contribute to inflammation during bacterial-induced colitis. This study found that IL-21 enhances the surface expression of IL-12Rβ2, but not IL-12Rβ1, in CD4+ T cells, leading to TH1 differentiation and stability. Consistently, these findings also point to an indispensable role of the IL-12Rβ2 signaling axis in promoting proinflammatory immune responses during Citrobacter rodentium-induced colitis. Genetic deletion of the IL-12Rβ2 signaling pathway led to the attenuation of C. rodentium-induced colitis in vivo. The genetic deletion of the IL-12Rβ2 signaling pathway did not alter the host's ability to respond adequately to C. rodentium infection or the ability of Il12rb2-/- mice to express antigen-specific cytokines (IFN-γ, IL-17A). IL-21 is a pleiotropic cytokine exerting a wide range of immunomodulatory functions in multiple tissues, and its direct targeting may result in undesirable off-target consequences. These findings highlight the possibility for targeted manipulations of signaling cascades downstream of main regulators of proinflammatory responses to control invading pathogens while preserving the integrity of host immune responses. A better understanding of the novel mechanisms by which IL-21/IL-21R signaling regulates bacterial-induced colitis will provide insights into the development of new therapeutic and preventive strategies to harness IL-21/IL-21R signaling or its downstream molecules to treat infectious colitis., Competing Interests: Conflict of interest statement. None declared., (© The Author(s) 2024. Published by Oxford University Press on behalf of Society for Leukocyte Biology.)

Background: Dysregulation of immune surveillance is tightly linked to the development of metabolic dysfunction-associated steatohepatitis (MASH)-driven hepatocellular carcinoma (HCC); however, its underlying mechanisms remain unclear. Herein, we aimed to determine the role of interleukin-21 receptor (IL-21R) in MASH-driven HCC., Methods: The clinical significance of IL-21R was assessed in human HCC specimens using immunohistochemistry staining. Furthermore, the expression of IL-21R in mice was assessed in the STAM model. Thereafter, two different MASH-driven HCC mouse models were applied between IL-21R-deficient mice and wild type controls to explore the role of IL-21R in MASH-driven HCC. To further elucidate the potential mechanisms by which IL-21R affected MASH-driven HCC, whole transcriptome sequencing, flow cytometry and adoptive lymphocyte transfer were performed. Finally, flow cytometry, enzyme-linked immunosorbent assay, immunofluorescent staining, chromatin immunoprecipitation assay and western blotting were conducted to explore the mechanism by which IL-21R induced IgA + B cells., Results: HCC patients with high IL-21R expression exhibited poor relapse-free survival, advanced TNM stage and severe steatosis. Additionally, IL-21R was demonstrated to be upregulated in mouse liver tumors. Particularly, ablation of IL-21R impeded MASH-driven hepatocarcinogenesis with dramatically reduction of lipid accumulation. Moreover, cytotoxic CD8 + T lymphocyte activation was enhanced in the absence of IL-21R due to the reduction of immunosuppressive IgA + B cells. Mechanistically, the IL-21R-STAT1-c-Jun/c-Fos regulatory axis was activated in MASH-driven HCC and thus promoted the transcription of Igha, resulting in the induction of IgA + B cells., Conclusions: IL-21R plays a cancer-promoting role by inducing IgA + B cells in MASH-driven hepatocarcinogenesis. Targeting IL-21R signaling represents a potential therapeutic strategy for cancer therapy., (© 2024. The Author(s).)

IL-21 is a cytokine with versatile antitumor and pro-tumorigenic activities. It is mainly produced by CD4+ T cells and B cells are one of its pivotal targets. In this study, we assessed and compared the expression of IL-21 by CD4+ T cells and the IL-21 receptor (IL-21R) on B cells in the peripheral blood of women with breast cancer and healthy individuals. Blood samples were taken from both patients and controls. Mononuclear cells were seperated using Ficoll-Hypaque density gradient centrifugation. These isolated cells were then stained with either anti-CD19/anti-IL-21R or anti-CD4/anti-IL-21 antibodies and analyzed using flow cytometry. The results showed that there was no significant difference in the percentage of IL-21R+ B cells and IL-21+CD4+ T cells between patients and controls. However, the percentage of CD4+ T cells decreased significantly in patients with breast cancer (P=0.003). This decline was observed from the early stage and before lymph node (LN) involvement. In comparison to the control group, IL-21R+ B cells were relatively lower in patients with stages I+II and those with fewer than 4 involved LNs. The intensity of IL-21 expression in T cells was associated with HER2 expression (P=0.029). Furthermore, we found that the majority of IL-21R+ B cells exhibited a naïve phenotype and most of IL-21+CD4+ T cells did not produce IFN-γ or IL-17. In conclusion, breast cancer from the early stages leads to a significant reduction in the proportion of peripheral CD4+ T cells. However, we did not find a significant change in IL-21 and its receptor expression during disease progression.

Strategies to improve T cell therapy efficacy in solid tumors such as hepatocellular carcinoma (HCC) are urgently needed. The common cytokine receptor γ chain (γ c ) family cytokines such as IL-2, IL-7, IL-15 and IL-21 play fundamental roles in T cell development, differentiation and effector phases. This study aims to determine the combination effects of IL-21 in T cell therapy against HCC and investigate optimized strategies to utilize the effect of IL-21 signal in T cell therapy. The antitumor function of AFP-specific T cell receptor-engineered T cells (TCR-T) was augmented by exogenous IL-21 in vitro and in vivo. IL-21 enhanced proliferation capacity, promoted memory differentiation, downregulated PD-1 expression and alleviated apoptosis in TCR-T after activation. A novel engineered IL-21 receptor was established, and TCR-T armed with the novel engineered IL-21 receptors (IL-21R-TCR-T) showed upregulated phosphorylated STAT3 expression without exogenous IL-21 ligand. IL-21R-TCR-T showed better proliferation upon activation and superior antitumor function in vitro and in vivo. IL-21R-TCR-T exhibited a less differentiated, exhausted and apoptotic phenotype than conventional TCR-T upon repetitive tumor antigen stimulation. The novel IL-21 receptor in our study programs powerful TCR-T and can avoid side effects induced by IL-21 systemic utilization. The novel IL-21 receptor creates new opportunities for next-generation TCR-T against HCC., (© 2024. The Author(s).)

Aims: Aimed to identify a new susceptibility gene associated with primary biliary cholangitis (PBC) in Chinese Han and investigate the possible mechanism of that gene in PBC., Methods: A total of 466 PBC and 694 healthy controls (HC) were included in our study, and genotyping GTF2I gene variants by Sequenom. CD19 + B cells were isolated for Chromatin immunoprecipitation sequencing (ChIP-seq). Additionally, MEME-ChIP was utilized to perform searches for known motifs and de novo motif discovery. The GTF2I ChIP-seq of hematopoietic cell line (K562) results were obtained from ENCODE (GSE176987, GSE177691). The Genomic HyperBrowser was used to determine overlap and hierarchal clustering between ours and ENCODE datasets., Results: The frequency of the rs117026326 variant T allele was significantly higher in PBC patients than that in HC (20.26% compared with 13.89%, Pc = 1.09E-04). Furthermore, we observed an elevated proportion of GTF2I binding site located in the upstream and 5' UTR of genes in PBC in comparison with HC. Additionally, an in-depth analysis of IL21R region revealed that GTF2I might bind to the IL21R promoter to regulate the expression of the IL21R, with four peaks of GTF2I binding sites, including three increased binding sites in upstream, one increased binding site in 5' UTR. Motif analysis by MEME-ChIP uncovered five significant motifs. A significant overlap between our ChIP and GSE176987, GSE17769 were found by the Genomic HyperBroswer., Conclusions: Our study confirmed that GTF2I was associated with PBC in Chinese Han. Furthermore, our gene function analysis indicated that IL21R may be the target gene regulated by GTF2I., (© 2023. Asian Pacific Association for the Study of the Liver.)

Rotator cuff tear (RCT) is a common shoulder disorder related to pain and dysfunction. However, the pathological mechanism of RCT remains unclear. Thus, this study aims to investigate the molecular events in RCT synovium and identify possible target genes and pathways as determined by RNA sequencing (RNA-Seq). The synovial tissue was biopsied from 3 patients with RCT (RCT group) and 3 patients with shoulder instability (Control group) during arthroscopic surgery. Then, differentially expressed (DE) mRNAs, long non-coding RNAs (lncRNAs) and micro RNAs (miRNAs) were comprehensively profiled by RNA-Seq. Gene ontology (GO) enrichment, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, and competing endogenous RNA (ceRNA) network analysis were performed to identify the potential functions of these DE genes. 447 mRNAs, 103 lncRNAs and 15 miRNAs were identified differentially expressed. The DE mRNAs were highlighted in inflammatory pathway including up-regulated T cell costimulation, positive regulation of T cell activation, and T cell receptor signaling. Down-regulated fatty acid degradation pathway and 5'-AMP-activated protein kinase (AMPK) signaling in RCT group are also enriched. Validation assay showed that the expression of pro-inflammatory molecules including IL21R, CCR5, TNFSF11, and MMP11 was significantly increased in RCT group compared with Control group. CeRNA analysis further revealed lncRNA-miRNA-mRNA regulatory networks involving IL21R and TNFSF11 in RCT. Activated synovial inflammation is the remarkable event of RCT. Importantly, increased T cell activation and disordered fatty acid metabolism signaling might play a significant role. ceRNA networks involving IL21R and TNFSF11 identified could potentially control the progression of RCT. In conclusion, our findings could provide new evidence for the molecular mechanisms of RCT and might identify new therapeutic targets., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Background: We identified a homologue of IL-21R (LcIL-21R) in large yellow croaker (Larimichthys crocea, Lc). Our investigation focused on understanding the molecular structural features and immune function of LcIL-21R., Methods: We cloned the LcIL-21R gene from the genome of Larimichthys crocea by RT‒PCR, and the molecular and structural characteristics of LcIL-21R were analyzed by a series of protein analysis tools. We used real-time PCR to investigate the tissue distribution of LcIL-21R, and LcIL-21R gene expression regulation was also measured in head kidney leukocytes under trivalent bacterial vaccine or poly (I:C) stimulation., Results: The open reading frame (ORF) of the LcIL-21R gene is 1629 bp long and encodes a precursor protein of 542 amino acids (aa), with a 23-aa signal peptide and a 519-aa mature peptide containing four putative N-glycosylation sites. LcIL-21R has two fibronectin type III (FNIII)-like domains (D1 and D2), a transmembrane domain, and a cytoplasmic region. A conserved WSXWS motif was also found in the D2 domain. The predicted structure of the extracellular region of LcIL-21R (LcIL-21R-Ex) is highly similar to that of human IL-21R. LcIL-21R was constitutively expressed in all tissues examined, and LcIL-21R mRNA levels were increased in the head kidney and spleen upon inactivated trivalent bacterial vaccine or poly(I:C) stimulation., Conclusions: Our results suggest that LcIL-21R shares structural and functional properties with IL-21Rs found in other vertebrates, indicating its potential involvement in the IL-21-mediated immune response to pathogenic infections. These findings contribute to our understanding of the evolutionary conservation of IL-21 signaling and its role in the immune system., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

Background: Interleukin-21 (IL-21) is produced by various cell types inducing positive and negative effects in immunity against tumors., Objective: To investigate the expression of IL-21 by CD4+T and IL-21 receptor (IL-21R) by B lymphocytes isolated from breast-tumor draining lymph nodes (TDLNs)., Methods: Fresh lymph node samples were obtained from 45 patients with breast cancer. To assess IL-21 expression, mononuclear cells were briefly stimulated whereas IL-21R expression was assessed in unstimulated B cells. Cells were stained with antibodies for CD4, IL-21, CD19 and IL-21R and acquired by flow cytometry., Results: The frequency of IL-21+CD4+T cells did not show significant association with disease parameters. However, the geometric mean fluorescence intensity (gMFI) of IL-21 in CD4+T cells was significantly lower in patients with grade III tumor than grade I + II (P = 0.042). In non-involved LNs, the intensity of IL-21 was significantly higher in patients with stage II compared with stage III (P = 0.038) and correlated negatively with the number of involved LNs. The frequency of IL-21R+CD19+B cells was significantly higher in grade III than grade I + II (P = 0.037)., Conclusion: The higher intensity of IL-21 in CD4+T cells showed association with good prognosticators in breast cancer and warrants further investigation of the role played by IL-21 in immunity against breast cancer.

Mutations in the interleukin-21 receptor (IL-21R) gene are recently defined as primary immunodeficiency diseases. IL-21R defects result in combined immunodeficiency by affecting the functions of innate and adaptive immune system components.A six-year-old girl was admitted to our hospital with complaints of chronic diarrhea that started after the newborn period and generalized rash over the last three months. She had severe respiratory distress due to Cytomegalovirus (CMV) pneumonia requiring mechanical ventilation and was diagnosed as combined immunodeficiency at another hospital at the age of four. Her physical examination on admission revealed erythematous rash on cheeks, extremities, gluteal region, and lymph node enlargements in cervical, axillary, and inguinal regions. CMV DNA and stool Cryptosporidium parvum were positive. Marginal zone lymphoma -negative for Epstein-Bar virus- was reported in the lymph node biopsy. Targeted next-generation sequencing Ion AmpliSeq™ primary immunodeficiency panel revealed a novel homozygous IL21R c.132delC (p.Ser45fs) mutation.This case is presented to emphasize that IL21R defects should be considered in the differential diagnosis of the patients with recurrent respiratory infections, chronic diarrhea, C. parvum infection, chronic liver disease, sclerosing cholangitis, and malignancy where early hematopoietic stem cell transplantation (HSCT) is life-saving. A total of eight cases with IL21R gene defects have been reported so far. The significance of this case is that it is the first case of malignancy among the published IL-21R deficient patients successfully treated with HSCT.

The IL-21/IL-21R interaction plays an important role in a variety of immune diseases; however, the roles and mechanisms in intracellular bacterial infection are not fully understood. In this study, we explored the effect of IL-21/IL-21R on chlamydial respiratory tract infection using a chlamydial respiratory infection model. The results showed that the mRNA expression of IL-21 and IL-21R was increased in Chlamydia muridarum -infected mice, which suggested that IL-21 and IL-21R were involved in host defense against C. muridarum lung infection. IL-21R -/- mice exhibited less body weight loss, a lower bacterial burden, and milder pathological changes in the lungs than wild-type (WT) mice during C. muridarum lung infection. The absolute number and activity of CD4 + T cells and the strength of Th1/Th17 responses in IL-21R -/- mice were significantly higher than those in WT mice after C. muridarum lung infection, but the Th2 response was weaker. Consistently, IL-21R -/- mice showed higher mRNA expression of Th1 transcription factors (T-bet/STAT4), IL-12p40, a Th17 transcription factor (STAT3), and IL-23. The mRNA expression of Th2 transcription factors (GATA3/STAT6), IL-4, IL-10, and TGF-β in IL-21R -/- mice was significantly lower than that in WT mice. Furthermore, the administration of recombinant mouse IL-21 aggravated chlamydial lung infection in C57BL/6 mice and reduced Th1 and Th17 responses following C. muridarum lung infection. These findings demonstrate that IL-21/IL-21R may aggravate chlamydial lung infection by inhibiting Th1 and Th17 responses., (Copyright © 2021 by The American Association of Immunologists, Inc.)

Genetic and epigenetic factors are considered to be critical for host-parasite interactions. There are limited data on the role of such factors during human infections with Ascaris lumbricoides . Here, we describe the potential role of genetic factors as determinants of the Th2 immune response to A. lumbricoides in Brazilian children. Stool samples were collected from the children to detect A. lumbricoides by microscopy and peripheral blood leukocytes (PBLs) were cultured in whole blood cultures for detection of cytokines (IL-5, IL-10, and IL-13) in vitro . Levels of anti- A. lumbricoides IgE and IgG4 were measured in plasma. DNA was extracted from PBLs and genotyped using Illumina 2.5 Human Omni Beadchip. Candidate genes associated with A. lumbricoides responses were identified and SNVs in these selected genes associated with the Th2 immune response to A. lumbricoides . Haplotype, gene expression, and epigenetic analyses were done to identify potential associations with Th2 immune responses. GWAS on samples from 1,189 children identified WSB1 as a candidate gene, and IL-21R was selected as a biologically relevant linked gene for further analysis. Variants in WSB1 and IL21R were associated with markers of Th2 immune responses: increased A. lumbricoides -specific IgE and IL-5/IL-13 by PBLs from infected compared to uninfected individuals. In infected children, WSB1 but not IL21R gene expression was suppressed and increased methylation was observed in the WSB1 promoter region. This is the first study to show an association between genetic variants in WSB1 and IL21R and Th2 immune responses during A. lumbricoides infections in children. WSB1 / IL21R pathways could provide a potential target for the treatment of Th2-mediated diseases., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Carneiro, da Silva, Queiroz, Veiga, Oliveira, Carneiro, Pires, da Silva, Sena, Belitardo, Nascimento, Silva, Marques, Costa, Alcantra-Neves, Barreto, Cooper and Figueiredo.)

The role of IL-21, produced mainly by Th17 cells and T follicular helper cells, has been intensively investigated in B cell differentiation and Ab class switch. However, how IL-21 regulates memory IgA + B cell development and memory IgA responses in the intestines is still not completely understood. In this study, we found the total IgA + B cells as well as CD38 + CD138 - IgA + memory B cells were significantly increased in intestinal lamina propria (LP) of TCRβxδ -/- mice after transfer of microbiota Ag-specific Th17 cells but not Th1 cells. Although IL-21R -/- mice or IL-17R -/- mice showed decreased Ag-specific memory IgA production in the intestines upon infection with Citrobacter rodentium , the percentage of IgA + CD38 + CD138 - memory B cells in Peyer's patches and LP was decreased only in IL-21R -/- mice, but not in IL-17R -/- mice, after reinfection with C. rodentium compared with wild-type mice. Blockade IL-21 in vivo suppressed intestinal C. rodentium -specific IgA production as well as IgA + CD38 + CD138 - memory B cells in Peyer's patches and LP. Furthermore, IL-21 significantly induced B cell IgA production in vitro, with the increased expression of genes related with class-switching and memory B cell development, including Aicda, Ski, Bmi1, and Klf2. Consistently, Aicda and Ski expression was decreased in B cells of IL-21R -/- mice after C. rodentium reinfection. In conclusion, our study demonstrated that IL-21 promotes intestinal memory IgA B cell development, possibly through upregulating differentiation-related and class switching-related genes, indicating a potential role of IL-21 in memory IgA + B cell responses in the intestines., (Copyright © 2020 by The American Association of Immunologists, Inc.)

IL-21 is the most recently discovered common gamma-chain cytokine that promotes persistent T-cell responses in chronic infections, autoimmunity and cancer. However, the therapeutic potential of inhibiting the IL-21-BATF signaling axis, particularly in transplant rejection, remains unclear. We used heart transplant models to examine the effects of IL-21 blockade in prevention of chronic cardiac allograft vasculopathy (CAV) using genetic knock-out and therapeutic approaches. Both wild-type C57BL/6 and IL-21-/- strains acutely rejected Balb/c skin grafts and once immunized with this skin graft, rejected Balb/c heart allografts in an accelerated fashion. However, when transplanted with heart grafts from the class-II major histocompatibility complex mutant, B6bm12 mice; wild-type recipients developed CAV, while IL-21-/- recipients were protected, even at day 100 post-transplant. Similarly, BATF-/- recipients, lacking the transcription factor BATF responsible for IL-21 production, did not develop CAV in B6-bm12 heart allografts. Strikingly, in a transient treatment protocol, the development of CAV in wild-type recipients of B6-bm12 hearts allografts was blocked by the administration of IL-21 receptor fusion protein (R-Fc). Thus, we demonstrate that CAV is regulated at least in part by IL-21 signaling and its blockade by genetic approaches or therapy with IL-21R-Fc prevents CAV in mice., Competing Interests: The authors have declared that no competing interests exist.

Pancreatic ductal adenocarcinoma (PDAC) displays a marked fibro-inflammatory microenvironment in which infiltrated immune cells fail to eliminate the tumor cells and often-rather paradoxically-promote tumor progression. Of special interest are tumor-promoting T cells that assume a Th17-like phenotype because their presence in PDAC tissue is associated with a poor prognosis. In that context, the role of IL-21, a major cytokine released by Th17-like cells, was assessed. In all tissue samples (n = 264) IL-21 + immune cells were detected by immunohistochemistry and high density of those cells was associated with poor prognosis. In the majority of patients (221/264), tumor cells expressed the receptor for IL-21 (IL-21R) and also a downstream target of IL-21, Blimp-1 (199/264). Blimp-1 expression closely correlated with IL-21R expression and multivariate analysis revealed that expression of both IL-21R and Blimp-1 was associated with shorter survival time of the patients. In vitro data using pancreatic tumor cells lines provided a possible explanation: IL-21 activated ERK and STAT3 pathways and upregulated Blimp-1. Moreover, IL-21 increased invasion of tumor cell lines in a Blimp-1-dependent manner. As an in vivo correlate, an avian xenograft model was used. Here again Blimp-1 expression was significantly upregulated in IL-21 stimulated tumor cells. In summary, our data showed an association of IL-21 + immune cell infiltration and IL-21 receptor expression in PDAC with poor survival, most likely due to an IL-21-mediated promotion of tumor cell invasion and enhanced colony formation, supporting the notion of the tumor-promoting abilities of the tumor microenvironment., Competing Interests: The authors declare that there is no conflict of interest regarding the publication of this paper.

Group 3 innate lymphoid cells (ILC3s) have dual roles in intestinal health, acting in both protective and pathogenic capacities, and importantly, modulations in this population of innate lymphoid cells have been implicated in inflammatory bowel disease. Further, subpopulations of ILC3s have been described as serving specific functions in maintaining homeostasis or responding to infection, and aberrant activation of one or more of these subpopulations could exacerbate inflammatory bowel disease. However, the signals that enforce the protective and pathogenic features of ILC3s are not fully elucidated. In this article, we show that IL-21, a cytokine primarily produced by CD4 T cells, acts on a subpopulation of intestinal ILC3s to promote a protective phenotype. IL-21 signaling does not affect the MHC class II-expressing ILC3 subset but promotes ILC3s that express Tbet and are poised to produce IL-22. Consistent with a protective phenotype, IL-21 deficiency dampens cytokine-induced IL-17A production. We show that exacerbated colitis develops in mice lacking the IL-21 receptor, in agreement with a protective role for IL-21 signaling on ILC3s. To our knowledge, these data reveal a novel role for IL-21 in shaping innate lymphoid cell responses in the intestine and provide one mechanism by which effector CD4 T cells can influence innate immunity., (Copyright © 2019 The Authors.)

Interleukin-21 (IL-21), an autocrine cytokine predominantly produced by follicular helper T (Tfh) and T helper 17 (Th17) cells, has been proven to play an important role in the immune system, for example, by promoting proliferation and the development of Tfh and Th17 cells, balancing helper T cell subsets, inducing B cell generation and differentiation into plasma cells, and enhancing the production of immunoglobulin. These effects are mainly mediated by activation of the JAK/STAT, MAPK and PI3K pathways. Some IL-21 target genes, such as B lymphocyte induced maturation protein-1 (Blimp-1), suppressor of cytokine signaling (SOCS), CXCR5 and Bcl-6, play important roles in the immune response. Therefore, IL-21 has been linked to autoimmune diseases. Indeed, IL-21 levels are increased in the peripheral blood and tissues of patients with systematic lupus erythematosus (SLE), rheumatoid arthritis (RA), type 1 diabetes (T1D), immune thrombocytopenia (ITP), primary Sjogren's syndrome (pSS), autoimmune thyroid disease (AITD) and psoriasis. This increased IL-21 even positively associates with Tfh cells, plasma cells, autoantibodies and disease activity in SLE and RA. Additionally, IL-21 has been utilized as a therapeutic target in SLE, RA, T1D and psoriatic mouse models. Profoundly, clinical trials have shown safety and improvement in RA patients. However, tolerance and long-term pharmacodynamics effects with low bioavailability have been found in SLE patients. Therefore, this review aims to summarize the latest progress on IL-21 function and its signaling pathway and discuss the role of IL-21 in the pathogenesis of and therapy for autoimmune diseases, with the hope of providing potential therapeutic and diagnostic strategies for clinical use., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Hepatitis B virus (HBV) infection is a major public health concern due to the infection often leads to chronic infection, liver cirrhosis and also liver cancer. The host immune response to HBV infection and also genetic background play significant role in outcome of infection. Single nucleotide polymorphisms (SNPs) are the most important kind of variation in genetic sequences that caused by point mutations. As cytokines have major roles in viral infections, it seems that cytokine gene polymorphisms are independently associated with response to viral infections. Interleukin 21 (IL-21) plays an influential role in both innate and adaptive immune responses. Its specific receptor, IL-21R, produced and located on the surface of T, B and natural killer (NK) cells and is critical for the proliferation and differentiation of these immune effector cells. Many studies confirmed that the IL-21 involved in response to viral infections. We aimed to investigate the association of G/T IL-21 (rs2055979) and C/T IL-21R (rs3093390) gene polymorphisms with chronic hepatitis B virus infection and HBV spontaneous clearance in Iranian population., Methods: In this study, blood samples were gathered from 320 patients with chronic HBV and 310 healthy controls and also 120 HBV spontaneous clearance individuals. Following genomic DNA extraction, genotypes of the selected SNPs determined by PCR and restriction fragment length polymorphism (RFLP) method. The results were analyzed by SPSS software using Chi-square, Logistic Regression, ANOVA and Independent Samples t-Test., Results: According to our results, in IL-21R (rs3093390 C/T) gene polymorphism, allele frequency of T is statistically different in the HBV spontaneous clearance group compared to chronic HBV cases. But there is no significant difference between G/T IL-21 (rs2055979) and C/T IL-21R (rs3093390) genotypes distribution in three groups. Also we found that higher serum aspartate transaminase (AST) level in HBV spontaneous clearance group is significantly associated with TT genotype of IL-21 (rs2055979) compared to GG genotype (P value = 0.006)., Discussion: Our results showed that T allele frequency in IL-21R (rs3093390 C/T) gene polymorphism could consider as a host genetic factor for HBV spontaneous clearance. Probably we can serve it as a potential prognostic genetic marker for spontaneous clearance of HBV infection., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Despite studies indicating the effects of IL-21 signaling in intestinal inflammation, its roles in intestinal homeostasis and infection are not yet clear. Here, we report potent effects of commensal microbiota on the phenotypic manifestations of IL-21 receptor deficiency. IL-21 is produced highly in the small intestine and appears to be critical for mounting an IgA response against atypical commensals such as segmented filamentous bacteria and Helicobacter, but not to the majority of commensals. In the presence of these atypical commensals, IL-21R-deficient mice exhibit reduced numbers of germinal center and IgA + B cells and expression of activation-induced cytidine deaminase in Peyer's patches as well as a significant decrease in small intestine IgA + plasmablasts and plasma cells, leading to higher bacterial burdens and subsequent expansion of Th17 and Treg cells. These microbiota-mediated secondary changes in turn enhance T cell responses to an oral antigen and strikingly dampen Citrobacter rodentium-induced immunopathology, demonstrating a complex interplay between IL-21-mediated mucosal immunity, microbiota, and pathogens.

We examined the unique contributions of the cytokines IL-21 and IL-4 on germinal center (GC) B cell initiation and subsequent maturation in a murine model system. Similar to other reports, we found T follicular helper cell expression of IL-21 begins prior to T follicular helper cell migration into the B cell follicle and precedes that of IL-4. Consistent with this timing, IL-21 signaling has a greater influence on the perifollicular pre-GC B cell transition to the intrafollicular stage. Notably, Bcl6 hi B cells can form in the combined absence of IL-21R- and STAT6-derived signals; however, these nascent GC B cells cease to proliferate and are more prone to apoptosis. When B cells lack either IL-21R or STAT6, aberrant GCs form atypical centroblasts and centrocytes that differ in their phenotypic maturation and costimulatory molecule expression. Thus, IL-4 and IL-21 play nonredundant roles in the phased progression of GC B cell development that can initiate in the combined absence of these cytokine signals., (Copyright © 2018 by The American Association of Immunologists, Inc.)

B cells are implicated in rheumatoid arthritis (RA) based on the presence of autoantibodies and the therapeutic response to B cell depletion. IL-21 has a significant role in B cell development and function. Here we assess B cell responses to IL-21 and the mechanisms responsible for altered IL-21R expression in RA. Flow cytometry of PBMC and cultured B cells was used to quantify protein and mRNA levels of IL-21R, IL-21 signaling through pSTAT3, specificity protein 1 (SP1) and to determine cytokine production (IL-6) and maturation status of B cells in RA and healthy control subjects. SP1 binding to the IL21R promoter region in B cells was assessed with ChIP-qPCR. We demonstrate an increase in IL-21R expression in total and memory B cells from RA subjects, which correlated with responsiveness to IL-21 stimulation. Stimulation of naïve RA B cells with IL-21 and CD40L resulted in an increase in differentiation into plasmablasts and an increase in IL-6 production in comparison to healthy controls, which was dose dependent on IL-21 stimulation. IL-21R expression on memory B cells in RA synovial fluid was comparable to peripheral blood making our study pertinent to understanding B cell responses in the joint and site of inflammation. We identified an increase in SP1 protein and mRNA in RA B cells and demonstrate an increase in binding of SP1 to the IL21R promoter region, which suggests a mechanism by which IL-21R expression is enhanced on B cells in RA. Taken together, our results indicate a mechanism by which IL-21 enhances B cell development and function in RA through an SP1 mediated increase in IL-21R expression on B cells.

The current study investigated the therapeutic effect of berberine (BBR), an alkaloid derivative against IL-21/IL-21R mediated phosphotidyl inositol 3 kinase/protein kinase B (PI3K/Akt) signaling in adjuvant induced arthritic fibroblast-like synoviocytes (AA-FLS) isolated from rats and IL-21 mediated osteoclastogenesis in bone-marrow derived monocytes (BMMs). BBR (15-45 µM) treatment attenuated the gene and protein levels of IL-21R complex. BBR suppressed the levels of IL-21 (20 ng/ml) mediated production of inflammatory cytokines such as: tumor necrosis factor alpha (TNFα), interleukin 1 beta (IL-1β), interleukin 6 (IL-6) and interleukin 23 (IL-23) in AA-FLS cells. Subsequently, BBR ameliorated the gene and protein expression levels of mechanistic target of rapamycin (mTOR), IL-23 and nuclear factor kappa B (NFκB) p65 through the inhibition of PI3K and upregulation of phosphatase and tensin homolog (PTEN) at the protein level. Furthermore, BBR also inhibited the phosphorylation of Akt and NFκB p65 in a dose dependant manner. LY294002 (20 µM) treatment suppressed the PI3K/Akt signaling and its downstream elements in AA-FLS cells. BBR also modulated IL-21 mediated osteoclastogenesis through the suppression of PI3K dependant nuclear factor of activated T-cells 1 (Nfatc1) induction. Moreover, BBR controlled the osteoclast differentiation via inhibition of various bone resorptive enzymes including: cathepsin K, matrix metalloproteinase 9 (MMP9) and tartarate acid phosphatase (TRAP). LY294002 also inhibited osteoclast formation via suppression of PI3K mediated Nfatc1 induction and other downstream elements. Overall, our findings suggest that BBR is a potential candidate for therapeutic targeting of IL-21/IL-21R mediated RA pathogenesis., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

The aim of this study was to evaluate the relationship between interleukin-21 (IL-21) and interleukin-21 receptor (IL-21R) polymorphisms and the response to peginterferon alfa (PEG-IFN α) therapy in HBeAg-positive chronic hepatitis B (CHB) patients.A total of 143 HBeAg-positive CHB patients treated for 48 weeks with PEG-IFN α and followed up for 24 weeks post-treatment were retrospectively evaluated. Genotypes analysis was performed for IL-21 polymorphisms rs907715, rs2221903, and IL-21R polymorphisms rs3093301 and rs2285452. Serum IL-21 levels were measured by enzyme-linked immunosorbent assay.The end of virological response (EVR) rate was 46.9% (67/143) at the end of treatment, the sustained virological response (SVR) rate was 43.4% (62/143) and the complete response (CR) rate was 32.1% (46/143) at 24 weeks post-treatment. Patients who carried IL-21 rs 2221903 genotype AA had a rather higher rate of EVR (response rate: 52.4%, odds ratio [OR] 0.42, 95% confidence interval [CI]: 0.19-0.91, P = .021), SVR (response rate: 47.6%, OR 0.43, 95% CI: 0.19-0.95, P = .028), and CR (response rate: 38.1%, OR 0.31, 95% CI: 0.12-0.79, P = .014) when compared to those had AG genotype. Meanwhile, IL-21rs 2221903 genotype AA was also independently associated with markedly reduced HBsAg levels (>1og10 IU/mL) after 24 weeks treatment and low HBsAg levels (<100 IU/mL) at the end of treatment. IL-21 rs907715 AG/GG genotype was independently associated with SVR (OR: 2.92, 95% CI: 0.98-8.6, P = .039; OR: 3.23, 95% CI: 1.0-10.4, P = .039). Patients with IL-21 rs907715 AG/GG genotype had higher serum IL-21 levels than those with rs907715 AA genotype (P = .021).IL-21 rs2221903 and rs907715 polymorphisms were significantly associated with the treatment response to PEG-IFN α among Chinese HBeAg-positive CHB patients.

Psoriasis is largely mediated by interleukin (IL)-23/T helper (Th) 17 axis, and IL-21 is a pleiotropic cytokine expressed by Th17 cells. Despite previously reported possible pathogenic roles of IL-21 in human psoriasis, we found that IL-21 receptor (IL-21R) signalling was not crucial for imiquimod-induced psoriatic inflammation, using IL-21R -/- mice. The severity of imiquimod-induced psoriatic manifestation and pro-inflammatory Th17 cytokine levels, IL-17A-producing γδ T cells and CD4+ T cells, and in vitro IL-17A production by γδ T cells after IL-23 stimulation was comparable between wild-type and IL-21R -/- mice. Collectively, IL-21R signalling was not critically involved in IMQ-induced psoriatic inflammation despite an increased IL-21 expression in the IMQ-treated mouse skin. Our data may represent the significant differences between human psoriasis and murine psoriasis model, and further studies using other models will be required to elucidate the role of IL-21 in psoriasis pathogenesis., (© 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Oxymatrine (OMT) as a type of alkaloids collected from Sophora flavescens Ait exerts some biological functions including anticancer properties. Here, we investigated the therapeutic effects of OMT in gastric cancer cells (HGC 27 and AGS). As a result, the exposure of gastric cancer (GC) cells to OMT contributed to the suppression of cell proliferation and invasion. Interleukin 21 receptor (IL-21R) was identified to be differentially expressed between OMT treatment group (4 mg/mL) and control group (0 mg/mL), and knockdown of IL-21R repressed cell proliferation and invasion via inactivation of the JAK2/STAT3 pathway. The rescue experiment showed that IL-21R overexpression attenuated the anti-tumor effects of OMT through activation of the JAK2/STAT3 pathway. Moreover, the expression of IL-21R was significantly upregulated in GC samples compared with the adjacent normal tissues and associated with overall survival (OS) and tumor recurrence of GC patients. Taken together, in this study, we evaluated the anti-tumor effects of OMT on GC by investigating proliferation and invasion ability changes, and our findings show that OMT exhibits effects via regulation of JAK/STAT signaling pathway. Through the mechanism study, we may enlighten the potential therapeutic target for treatment of GC.

In this study, we determined the role of IL-21R signaling in Mycobacterium tuberculosis infection, using IL-21R knockout (KO) mice. A total of 50% of M. tuberculosis H37Rv - infected IL-21R KO mice died in 6 mo compared with no deaths in infected wild type (WT) mice. M. tuberculosis - infected IL-21R KO mice had enhanced bacterial burden and reduced infiltration of Ag-specific T cells in lungs compared with M. tuberculosis - infected WT mice. Ag-specific T cells from the lungs of M. tuberculosis - infected IL-21R KO mice had increased expression of T cell inhibitory receptors, reduced expression of chemokine receptors, proliferated less, and produced less IFN- γ, compared with Ag-specific T cells from the lungs of M. tuberculosis - infected WT mice. T cells from M. tuberculosis - infected IL-21R KO mice were unable to induce optimal macrophage responses to M. tuberculosis. This may be due to a decrease in the Ag-specific T cell population. We also found that IL-21R signaling is associated with reduced expression of a transcriptional factor Eomesodermin and enhanced functional capacity of Ag-specific T cells of M. tuberculosis - infected mice. The sum of our findings suggests that IL-21R signaling is essential for the optimal control of M. tuberculosis infection., (Copyright © 2017 by The American Association of Immunologists, Inc.)

The role and significance of interleukin (IL)-21 in the development of sporadic CRC have not been well defined. The aim of this study is therefore to investigate the dynamics of the IL-21 along colorectal adenoma-carcinoma sequence and to evaluate the impact of IL-21 on clinicopathological parameters and CRC prognosis. The real-time PCR results showed that the level of IL-21 in adenomas (n=50) and sporadic CRC (n=50) were significantly higher than that in normal controls (n=18), which were predominately observed in the adenoma/CRC stroma. Analysis revealed that IL-21 level was correlated with the overall survival time in CRC patients. Double immunofluorescence observations confirmed that IL-21 positive cells were mostly natural killer cells and T lymphocytes in the tumor stroma. These results indicate that significant increased IL-21 expression present within the adenoma/CRC microenvironment might have a potential predicating significance for survival time in patients with CRC., (Copyright © 2017 Elsevier Inc. All rights reserved.)

T-cell tolerance is a major obstacle to successful cancer immunotherapy; thus, developing strategies to break immune tolerance is a high priority. Here we show that expression of the E3 ubiquitin ligase Grail is upregulated in CD8 + T cells that have infiltrated into transplanted lymphoma tumours, and Grail deficiency confers long-term tumour control. Importantly, therapeutic transfer of Grail-deficient CD8 + T cells is sufficient to repress established tumours. Mechanistically, loss of Grail enhances anti-tumour reactivity and functionality of CD8 + T cells. In addition, Grail-deficient CD8 + T cells have increased IL-21 receptor (IL-21R) expression and hyperresponsiveness to IL-21 signalling as Grail promotes IL-21R ubiquitination and degradation. Moreover, CD8 + T cells isolated from lymphoma patients express higher levels of Grail and lower levels of IL-21R, compared with CD8 + T cells from normal donors. Our data demonstrate that Grail is a crucial factor controlling CD8 + T-cell function and is a potential target to improve cytotoxic T-cell activity.Grail is an E3 ubiquitin ligase that inhibits T-cell receptor signalling in CD4 + T cells. Here the authors show Grail also limits IL-21 receptor expression and function in CD8 + T cells, is overactive in these cells in patients with lymphoma, and promotes tumour development in a lymphoma transplant mouse model.

In spite of newly emerging therapies and the improved survival of patients with non-Hodgkin lymphoma (NHL), relapses or primary refractory disease are commonly observed and associated with dismal prognosis. Although discovery of the anti-CD20 antibody rituximab has markedly improved outcomes in B-cell NHL, rituximab resistance remains an important obstacle to successful treatment of these tumors. To improve the efficacy of CD20-targeted therapy, we fused interleukin 21 (IL-21), which induces direct lymphoma cytotoxicity and activates immune effector cells, to the anti-CD20 antibody (αCD20-IL-21 fusokine). We observed substantially enhanced IL-21R-mediated signaling by the fusokine compared with native IL-21 at equimolar concentrations. Fusokine treatment led to direct apoptosis of lymphoma cell lines and primary tumors that otherwise were resistant to native IL-21 treatment. In addition to direct cytotoxicity, the fusokine enhanced NK cell activation, effector functions, and interferon γ production, resulting in greater antibody-dependent cell-mediated cytotoxicity compared with IL-21 and/or anti-CD20 antibody treatments. Further, the αCD20-IL-21 fusokine stabilizes IL-21 and prolongs its half-life. In vivo αCD20-IL-21 therapy resulted in a significant tumor control in the rituximab-resistant A20-huCD20 tumors. Collectively, the dual functional ability of the αCD20-IL-21 fusokine to induce direct apoptosis and activate immune effector cells may provide benefit over existing treatments for NHL., (© 2017 by The American Society of Hematology.)

Gene profiling studies have indicated that in vitro differentiated human megakaryocytes express the receptor for IL-21 (IL-21R), an immunostimulatory cytokine associated with inflammatory disorders and currently under evaluation in cancer therapy. The aim of this study was to investigate whether IL-21 modulates megakaryopoiesis. We first checked the expression of IL-21 receptor on human bone marrow and in vitro differentiated megakaryocytes. We then investigated the effect of IL-21 on the in vitro differentiation of human blood CD34 + progenitors into megakaryocytes. Finally, we analyzed the consequences of hydrodynamic transfection-mediated transient expression of IL-21, on megakaryopoiesis and thrombopoiesis in mice. The IL-21Rα chain was expressed in human bone marrow megakaryocytes and was progressively induced during in vitro differentiation of human peripheral CD34 + progenitors, while the signal transducing γ chain was down-regulated. Consistently, the STAT3 phosphorylation induced by IL-21 diminished during the later stages of megakaryocytic differentiation. In vitro , IL-21 increased the number of colony-forming unit megakaryocytes generated from CD34 + cells and the number of megakaryocytes differentiated from CD34 + progenitors in a JAK3- and STAT3-dependent manner. Forced expression of IL-21 in mice increased the density of bi-potent megakaryocyte progenitors and bone marrow megakaryocytes, and the platelet generation, but increased platelet clearance with a consequent reduction in blood cell counts. Our work suggests that IL-21 regulates megakaryocyte development and platelet homeostasis. Thus, IL-21 may link immune responses to physiological or pathological platelet-dependent processes., (Copyright© Ferrata Storti Foundation.)

Bone marrow monocytes are primarily committed to osteoclast formation. It is, however, unknown whether potential primary alterations are specifically present in bone marrow monocytes from patients with multiple myeloma, smoldering myeloma or monoclonal gammopathy of undetermined significance. We analyzed the immunophenotypic and transcriptional profiles of bone marrow CD14 + monocytes in a cohort of patients with different types of monoclonal gammopathies to identify alterations involved in myeloma-enhanced osteoclastogenesis. The number of bone marrow CD14 + CD16 + cells was higher in patients with active myeloma than in those with smoldering myeloma or monoclonal gammopathy of undetermined significance. Interestingly, sorted bone marrow CD14 + CD16 + cells from myeloma patients were more pro-osteoclastogenic than CD14 + CD16-cells in cultures ex vivo Moreover, transcriptional analysis demonstrated that bone marrow CD14 + cells from patients with multiple myeloma (but neither monoclonal gammopathy of undetermined significance nor smoldering myeloma) significantly upregulated genes involved in osteoclast formation, including IL21R IL21R mRNA over-expression by bone marrow CD14 + cells was independent of the presence of interleukin-21. Consistently, interleukin-21 production by T cells as well as levels of interleukin-21 in the bone marrow were not significantly different among monoclonal gammopathies. Thereafter, we showed that IL21R over-expression in CD14 + cells increased osteoclast formation. Consistently, interleukin-21 receptor signaling inhibition by Janex 1 suppressed osteoclast differentiation from bone marrow CD14 + cells of myeloma patients. Our results indicate that bone marrow monocytes from multiple myeloma patients show distinct features compared to those from patients with indolent monoclonal gammopathies, supporting the role of IL21R over-expression by bone marrow CD14 + cells in enhanced osteoclast formation., (Copyright© Ferrata Storti Foundation.)

Several memory B-cell subclasses with distinct functions have been described, of which the most effective is the class-switched (CS) memory B-cell population. We have previously shown, using virus-like particles (VLPs), that the proliferative potential of these CS memory B cells is limited and they fail to re-enter germinal centres (GCs). However, VLP-specific memory B cells quickly differentiated into secondary plasma cells (PCs) with the virtue of elevated antibody production compared with primary PCs. Whereas the induction of VLP + memory B cells was strongly dependent on T helper cells, we were wondering whether re-stimulation of VLP + memory B cells and their differentiation into secondary PCs would also require T helper cells. Global absence of T helper cells led to strongly impaired memory B cell proliferation and PC differentiation. In contrast, lack of interleukin-21 receptor-dependent follicular T helper cells or CD40 ligand signalling strongly affected proliferation of memory B cells, but differentiation into mature secondary PCs exhibiting increased antibody production was essentially normal. This contrasts with primary B-cell responses, where a strong dependence on CD40 ligand but limited importance of interleukin-21 receptor was seen. Hence, T helper cell dependence differs between primary and secondary B-cell responses as well as between memory B-cell proliferation and PC differentiation., (© 2016 John Wiley & Sons Ltd.)

Circulating T follicular helper (Tfh) cells in the blood are counterparts to conventional Tfh cells in germinal centres. Similarly to conventional Tfh cells, circulating Tfh cells provide helpful signals for B cells. Circulating Tfh cells can be divided into three subpopulations, including Tfh17 (CXCR3-CCR6+), Tfh1 (CXCR3+CCR6-), and Tfh2 (CXCR3-CCR6-) cells, based on differences in CXCR3 and CCR6 expression. Recent studies have demonstrated that alterations in circulating Tfh cell subsets have significant effects on the progression of numerous autoimmune diseases. To address the role of circulating Tfh cells in psoriasis, we used flow cytometry to analyse frequencies of circulating Tfh cell subsets and levels of IL-21 receptor (IL-21R) expression on B cells in 32 patients with psoriasis vulgaris and 13 sex- and age-matched healthy controls. We observed dynamic changes in Tfh17 cell frequency and disease severity in 10 psoriasis patients during a 6-month treatment. Our results demonstrate that the frequency of Tfh17 cells was significantly increased in psoriasis patients and was positively correlated with psoriasis area and severity index (PASI) score, a marker of disease severity (P<0.05). During the follow-up period, the frequencies of Tfh17 cells decreased, and reductions in PASI scores were observed. Moreover, IL-21R expression on B cells was significantly increased in patients with psoriasis vulgaris (P<0.05). Interestingly, IL-21R levels on B cells were correlated with both PASI score and the frequency of Tfh17 cells (P<0.05). In conclusion, this study is the first investigation of the distribution of circulating Tfh subsets in psoriasis. These results deepen our understanding of the immune state of psoriasis patients. Our data also indicate that Tfh17 cells may not be good biomarkers for diagnosing psoriasis but may be potential biomarkers of disease severity in psoriasis., (Copyright © 2016. Published by Elsevier B.V.)

Innate γδ T cells expressing Vγ6 produce IL-17A at an early stage following infection with Mycobacterium bovis Bacillus Calmette-Guérin (BCG). In this study, we used IL-21 receptor knockout (IL-21R KO) mice and IL-21-producing recombinant BCG mice (rBCG-Ag85B-IL-21) to examine the role of IL-21 in the regulation of IL-17A-producing innate γδ T-cell response following BCG infection. IL-17A-producing Vγ6 + γδ T cells increased in the peritoneal cavity of IL-21R KO mice more than in wild type mice after BCG infection. In contrast, the number of IL-17A-producing Vγ6 + γδ T cells was significantly lower after inoculation with rBCG-Ag85B-IL-21 compared with control rBCG-Ag85B. Notably, exogenous IL-21 selectively induced apoptosis of IL-17A-producing Vγ6 + γδ T cells via Bim. Thus, these results suggest that IL-21 acts as a potent inhibitor of a IL-17A-producing γδ T-cell subset during BCG infection.

TGF-β together with IL-21 or IL-6 can drive the differentiation of naïve CD8(+) T cells into IL-17-producing CD8(+) T cells. These IL-17-producing CD8(+) T cells are termed Tc17 cells. Tc17 cells preserve plasticity under various conditions in vitro and in vivo. IFN-γ-producing CD8(+) T cells are termed Tc1 cells. However, Tc1 cells are considered relatively stable. In the present study, we show that the combination of TGF-β plus IL-21, but not IL-6, converts Tc1 cells into Tc17 cells; this conversion is associated with elevated RORα, RORγt, and Batf mRNA levels. These results indicate that Tc1 cells are skewed to the Tc17 cell phenotype under TGF-β plus IL-21-polarizing conditions. Furthermore, IL-6R is expressed on naïve, but not activated, CD8(+) T cells. In contrast, IL-21R is expressed on both naïve and activated CD8(+) T cells. Thus, differential expression profiles of IL-6R and IL-21R on naïve and activated CD8(+) T cells may be one mechanism by which TGF-β plus IL-21, but not IL-6, can drive activated CD8(+) T cells to differentiate into IL-17-producing cells. Taken together, these results provide a novel viewpoint for the plasticity of Tc1 cells., (Copyright © 2016 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.)

Follicular T helper (Tfh) cells are key regulators of the germinal center reaction and long-term humoral immunity. Tfh cell differentiation requires the sustained expression of the transcriptional repressor Bcl6; however, its regulation in CD4(+)T cells is incompletely understood. Here, we report that the transcriptional coactivator Bob1, encoded by thePou2af1gene, promotes Bcl6 expression and Tfh cell development. We found that Bob1 together with the octamer transcription factors Oct1/Oct2 can directly bind to and transactivate theBcl6andBtlapromoters. Mixed bone marrow chimeras revealed that Bob1 is required for the expression of normal levels of Bcl6 andBTLA, thereby controlling the pool size and composition of the Tfh compartment in a T cell-intrinsic manner. Our data indicate that T cell-expressed Bob1 is directly involved in Tfh cell differentiation and required for mounting normal T cell-dependent B-cell responses., (© 2016 The Authors. Published under the terms of the CC BY NC ND 4.0 license.)

Microsporidia, a latent opportunistic infection associated with mild inflammation, is characterized by a strong CD8 T cell response, which has been shown to be CD4 T cell dependent. In this manuscript, we demonstrate that CD4 help is provided via IL-21 production, a common γ-chain cytokine closely related to IL-2. The peak of IL-21 expression, observed during the acute infection, is associated with an elevated IL-21(+) CD4 T subset, and these cells bear a phenotypic resemblance to T follicular helper cells. We observed that, during per-oral microsporidial infection, IL-21 was critical for the generation of an optimal effector CD8 T cell immunity. Sharply decreased effector KLRG1(+) CD8 response was observed in IL-21R knockout mice, and although these cells exhibited reduced functional properties, they retained the ability to proliferate. The role of IL-21 in the generation of CD8 effectors was cell intrinsic, as stronger defects were observed in the IL-21-deficient compartment from the bone marrow chimeric mice (IL-21R knockout/wild-type). These findings are different from those reported for viral infections in which IL-21 has been primarily associated with the generation and maintenance of CD8 memory response. To the best of our knowledge, this report demonstrates a critical role for IL-21 in the generation of a primary effector CD8 T cell response to an infectious disease model., (Copyright © 2015 by The American Association of Immunologists, Inc.)

Interleukin 21 (IL21) is a cytokine produced predominantly by cluster of differentiation 4 (CD4+) T-cells and natural killer T-cells. There exists evidence that IL21 is implicated in various immunological processes through its specific receptor (IL21R). However, the participation of IL21 in the pathogenesis of solid tumors is not fully conclusive. In the present study, we demonstrated that there was differential expression of IL21R in breast cancer cells using reverse transcription-polymerase chain reaction (RT-PCR), western blotting and sequence analysis. The expression of IL21R was stronger in MDA-231 cells, weaker in MCF7 but negative in ZR-75.1 cells. The invasion and migratory capacity of IL21R+ MDA-231 cells was enhanced by IL21 in a dose-dependent manner. After IL21R was knocked-down by siRNA gene silencing, the response of MDA-231 to treatment with IL21 was attenuated. We found that siRNA silencing of IL21R also spontaneously suppressed cell proliferation. However, IL21 had no additional effect on the proliferation of MDA-231 cells. We also found that IL21R was involved in signaling pathways of matrix metalloproteinases (MMPs), that are crucial for spreading and migration of metastatic MDA231 cells. In conclusion, we unveiled the roles of IL21R in breast cancer cells, which enhances our knowledge on immunological regulation of cancer cells through the axis of IL21 and its receptor., (Copyright© 2015, International Institute of Anticancer Research (Dr. John G. Delinasios), All rights reserved.)

Asthma is a T helper 2 (Th2)-cell-mediated disease; however, recent findings implicate Th17 and innate lymphoid cells also in regulating airway inflammation. Herein, we have demonstrated profound interleukin-21 (IL-21) production after house dust mite (HDM)-driven asthma by using T cell receptor (TCR) transgenic mice reactive to Dermatophagoides pteronyssinus 1 and an IL-21GFP reporter mouse. IL-21-producing cells in the mediastinal lymph node (mLN) bore characteristics of T follicular helper (Tfh) cells, whereas IL-21(+) cells in the lung did not express CXCR5 (a chemokine receptor expressed by Tfh cells) and were distinct from effector Th2 or Th17 cells. Il21r(-/-) mice developed reduced type 2 responses and the IL-21 receptor (IL-21R) enhanced Th2 cell function in a cell-intrinsic manner. Finally, administration of recombinant IL-21 and IL-25 synergistically promoted airway eosinophilia primarily via effects on CD4(+) lymphocytes. This highlights an important Th2-cell-amplifying function of IL-21-producing CD4(+) T cells in allergic airway inflammation., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Objective: Surgical hindlimb ischemia (HLI) in mice has become a valuable preclinical model to study peripheral arterial disease. We previously identified that the different phenotypic outcomes after HLI across inbred mouse strains is related to a region on the short arm of mouse chromosome 7. The gene coding the interleukin-21 receptor (IL-21R) lies at the peak of association in this region., Approach and Results: With quantitative real-time polymerase chain reaction, we found that a mouse strain with a greater ability to upregulate IL-21R after HLI had better perfusion recovery than a strain with no upregulation after HLI. Immunofluorescent staining of ischemic hindlimb tissue showed IL-21R expression on endothelial cells (ECs) from C57BL/6 mice. An EC-enriched fraction isolated from ischemic hindlimb muscle showed higher Il-21R levels than an EC-enriched fraction from nonischemic limbs. In vitro, human umbilical vein ECs showed elevated IL-21R expression after hypoxia and serum starvation. Under these conditions, IL-21 treatment increased cell viability, decreased cell apoptosis, and augmented tube formation. In vivo, either knockout Il21r or blocking IL-21 signaling by treating with IL-21R-Fc (fusion protein that blocks IL-21 binding to its receptor) in C57BL/6 mice resulted in less perfusion recovery after HLI. Both in vitro and in vivo modulation of the IL-21/IL-21R axis under hypoxic conditions resulted in increased signal transducer and activator of transcription 3 phosphorylation and a subsequent increase in the B-cell lymphoma leukemia-2/BCL-2-associated X protein ratio., Conclusion: Our data indicate that IL-21R upregulation and ligand activation in hypoxic ECs may help perfusion recovery by limiting/preventing apoptosis and favoring cell survival and angiogenesis through the signal transducer and activator of transcription 3 pathway., (© 2015 American Heart Association, Inc.)

To determine the potential role of interleukin-21 (IL-21) / IL-21 receptor (IL-21R) in the pathogenesis of autoimmune thyroid disease (AITD) mainly known as Graves' disease (GD) and Hashimoto's thyroiditis (HT). IL-21 and IL-21R of peripheral blood samples and/or thyroid tissues from AITD patients and healthy controls were analyzed by ELISA, quantitative real-time polymerase chain reaction (qRT-PCR), flow cytometry and immunohistochemistry. In vitro, the mRNA and protein of inflammatory cytokines of cultured peripheral blood mononuclear cells (PBMCs) upon recombinant human IL-21 (rhIL-21) stimulation were detected. There was an increased serum concentration of IL-21 in untreated GD and HT patients, and IL-21(+)CD3(+)CD8(-)T cells were significantly increased in PBMCs of HT patients compared with healthy volunteers. The IL-21 mRNA expression in PBMCs increased dramatically in GD and HT patients, and marked augmentations of IL-21 and IL-21R mRNA in thyroid tissues of HT patients were observed. Immunohistochemical staining confirmed the expression of IL-21R protein in HT thyroid cells and lymphocytes. In vitro, PBMCs from GD cultured with rhIL-21 induced increased IL-17A but decreased IL-4 production, while from HT stimulated by rhIL-21 induced augmented production of IFN-γ. In conclusion, the expression of IL-21 and IL-21R were up-regulated in AITD and may be involved in the pathogenesis of the disease through augmenting aberrant immune cascade., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Interleukin-21 (IL-21), produced predominantly by CD4+ T cells and natural killer T (NKT) cells, is a newly discovered member of the common γ-chain family of cytokines. It has been implicated in many immunological processes and has been linked to autoimmune diseases, allergies and other inflammatory diseases. In recent years, the role for IL-21 in the pathogenesis of cancer has also been extensively studied. In this review, we will discuss recent advances concerning the role of IL-21 in immunological processes and the pathogenesis of cancer., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

Interleukin-21 (IL-21) is a T cell-derived cytokine modulating T cell, B cell, and natural killer cell responses. To determine whether IL-21 contributes to pathologic processes, recombinant IL-21 receptor (R) fusion protein (rhIL-21R-Fc) was examined in mice models of autoimmune arthritis (collagen-induced arthritis). DBA/1J mice were immunized with chicken type II collagen and then treated intraperitoneally with rhIL-21R-Fc, which was initiated after the onset of arthritis symptoms in 20% of the cohort. The mice were assessed 3 times per week for signs of arthritis and histologic features as well as serum immunoglobulin. Cytokine messenger RNA levels in the spleen were also examined. STAT3 phosphorylation is dose dependently activated by IL-21 and inhibited by rhIL-21R-Fc in vitro using T cells. Treatment of DBA/1J mice with rhIL-21R-Fc reduced the clinical and histologic signs of CIA. The IL-17 and STAT3-expressing CD4(+) splenocytes dramatically decreased in the rhIL-21R-Fc treated mice. IL-21R-Fc treated mice also decreased the production of IgG, STAT3 phosphorylation, and plasma cell transcription factor (Blimp1). These findings demonstrate a pathogenic role of IL-21 in animal models of RA, suggesting IL-21 as a promising therapeutic target among human RA., (Copyright © 2014 European Federation of Immunological Societies. Published by Elsevier B.V. All rights reserved.)

Purpose of Review: This review highlights the recent identification of human interleukin-21 (IL-21) and interleukin-21 receptor (IL-21R) deficiencies as novel entities of primary immunodeficiency., Recent Findings: We recently described the first patients with IL-21R deficiency who had cryptosporidial infections associated with chronic cholangitis and liver disease. All IL-21R-deficient patients suffered from recurrent respiratory tract infections. Immunological work-up revealed impaired B cell proliferation and immunoglobulin class-switch, reduced T cell effector functions, and variable natural killer cell dysfunctions. Recently, these findings have been extended by the discovery of one patient with a mutation in the IL21 gene. This patient predominantly manifested with very early onset inflammatory bowel disease and recurrent respiratory infections. Laboratory examination showed reduced circulating B cells and impaired B cell class-switch., Summary: Human IL-21 and IL-21R deficiencies cause severe, primary immunodeficiency reminiscent of common variable immunodeficiency. Early diagnosis is critical to prevent life-threatening complications, such as secondary liver failure. In view of the critical role of IL-21 in controlling immune homeostasis, early hematopoietic stem cell transplantation might be considered as therapeutic intervention in affected children.

The dynamic interplay between regulatory T cells (T(regs)) and effector T cells (T(effs)) governs the balance between tolerance and effector immune responses. Perturbations of T(reg) frequency and function or imbalances in T(reg)/T(eff) levels are associated with the development of autoimmunity. The factors that mediate these changes remain poorly understood and were investigated in this study in murine autoimmune arthritis. T(regs) displayed a stable phenotype in arthritic mice and were fully functional in in vitro suppression assays. However, their expansion was delayed relative to T(effs) (T follicular helper cells and Th17 cells) during the early stages of autoimmune reactivity. This imbalance is likely to have led to insufficient T(reg) control of T(effs) and induced autoimmunity. Moreover, a counterregulatory and probably IL-7-driven increase in thymic T(reg) production and recruitment to inflamed tissues was too slow for disease prevention. Increased T(eff) over T(reg) expansion was further aggravated by inflammation and lymphopenia. Both these conditions contribute to autoimmune pathogenesis and were accompanied by decreases in the availability of IL-2 and increases in levels of IL-21. IL-2 neutralization or supplementation was used to show that T(reg) expansion mainly depended on this cytokine. IL-21R(-/-) cells were used to demonstrate that IL-21 promoted the maintenance of T(effs). Thus, at inflammatory sites in experimental arthritis, a deficit in IL-2 hampers T(reg) proliferation, whereas exaggerated IL-21 levels overwhelm T(reg) control by supporting T(eff) expansion. This identifies IL-2 and IL-21 as targets for manipulation in therapies for autoimmunity., (Copyright © 2014 by The American Association of Immunologists, Inc.)

Background: Interleukin-21 (IL-21) has been implicated in the development of Th2-mediated immune responses; however, the exact role it plays in allergic diseases is not well understood., Objective: To elucidate the contribution of IL-21 receptor signalling to Th2-dependent immune responses in the lung., Methods: We compared allergic airway responses in wild-type BALB/c and Il21r-deficient mice exposed to local airway challenge with house dust mite (HDM)., Results: We demonstrate that IL-21R-deficiency reduces HDM-driven airway hyperresponsiveness (AHR) with only partial effects on airway inflammation. Concomitant with the reduction in AHR in Il21r-deficient mice, significant suppression was observed in protein levels of the Th2 cytokines IL-4, and IL-13. In contrast, IL-21R-deficiency was associated with an increase in PBS- and allergen-driven IgE levels, while IgG1 and IgG2a levels were decreased. Moreover, our results suggest that IL-21 may contribute to AHR through its ability to both directly induce Th2 cell survival and to impair regulatory T-cell suppression of Th2 cytokine production. Importantly, we show that IL-21-positive cells are increased in the bronchial mucosa of asthmatics compared with non-asthmatics., Conclusion: These results suggest that IL-21 plays an important role in the allergic diathesis by enhancing Th2 cytokine production through multiple mechanisms including the suppression of Treg inhibitory effects on Th2 cell cytokine production., (© 2014 John Wiley & Sons Ltd.)

Chronic graft-versus-host disease (cGVHD) is a leading cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation. Having shown that germinal center (GC) formation and immunoglobulin deposition are required for multiorgan system cGVHD and associated bronchiolitis obliterans syndrome (BOS) in a murine model, we hypothesized that T follicular helper (Tfh) cells are necessary for cGVHD by supporting GC formation and maintenance. We show that increased frequency of Tfh cells correlated with increased GC B cells, cGVHD, and BOS. Although administering a highly depletionary anti-CD20 monoclonal antibody (mAb) to mice with established cGVHD resulted in peripheral B-cell depletion, B cells remained in the lung, and BOS was not reversed. BOS could be treated by eliminating production of interleukin-21 (IL-21) by donor T cells or IL-21 receptor (IL-21R) signaling of donor B cells. Development of BOS was dependent upon T cells expressing the chemokine receptor CXCR5 to facilitate T-cell trafficking to secondary lymphoid organ follicles. Blocking mAbs for IL-21/IL-21R, inducible T-cell costimulator (ICOS)/ICOS ligand, and CD40L/CD40 hindered GC formation and cGVHD. These data provide novel insights into cGVHD pathogenesis, indicate a role for Tfh cells in these processes, and suggest a new line of therapy using mAbs targeting Tfh cells to reverse cGVHD., (© 2014 by The American Society of Hematology.)

Follicular helper T cells (Tfh), which play a pivotal role in B cell activation and differentiation in lymphoid structures, secrete IL-21 whose augmented secretion is a hallmark of several autoimmune diseases. To decipher the cellular and molecular interactions occurring in salivary glands of patients suffering from primary Sjögren's syndrome (pSS), we investigated whether salivary gland epithelial cells (SGECs) were capable to induce Tfh differentiation. Co-cultures of naïve CD4(+) T cells and SGECs from both patients with pSS and controls were performed. Here, we report that IL-6 and ICOSL expression by SGECs contributes to naïve CD4(+) T differentiation into Tfh cells, as evidenced by their acquisition of a specific phenotype, characterized by Bcl-6, ICOS and CXCR5 expression and IL-21 secretion, but also but by their main functional feature: the capacity to enhance B lymphocytes survival. We demonstrated an increase of serum IL-21 with systemic activity. Finally, we analyzed the potential occurrence of a genetic association between IL-21 or IL-21R gene polymorphisms and pSS or elevated IL-21 secretion. This study, which demonstrates a direct induction of Tfh differentiation by SGECs, emphasizes a yet unknown pathogenic role of SGECs and suggests that Tfh and IL-21 might be relevant biomarkers and/or therapeutic targets in primary Sjögren's syndrome., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Objective: The cytokine interleukin-21 (IL-21) can have both proinflammatory and immunosuppressive effects. The purpose of this study was to investigate the potential dual role of IL-21 in experimental arthritis in relation to Th17 cells., Methods: Antigen-induced arthritis (AIA) and chronic streptococcal cell wall (SCW) arthritis were induced in IL-21 receptor-deficient (IL-21R(-/-) ) and wild-type mice. Knee joints, synovial tissue, and serum were analyzed for arthritis pathology and inflammatory markers., Results: During AIA and chronic SCW arthritis, IL-21R deficiency protected against severe inflammation and joint destruction. This was accompanied by suppressed serum IgG1 levels and antigen-specific T cell responses. Levels of IL-17 were reduced during AIA, and synovial lymphocytes isolated during SCW arthritis for flow cytometry demonstrated that mainly IL-17+ interferon-γ (IFNγ)-positive T cells were reduced in IL-21R(-/-) mice. However, during the acute phases of SCW arthritis, significantly higher joint swelling scores were observed, consistent with enhanced tumor necrosis factor and IL-6 expression. Interestingly, IL-21R(-/-) mice were significantly less capable of up-regulating suppressor of cytokine signaling 1 (SOCS-1) and SOCS-3 messenger RNA. IL-21 stimulation also affected the Toll-like receptor 2 (TLR-2)/caspase recruitment domain 15 response to SCW fragments in vitro, indicating that impaired SOCS regulation in the absence of IL-21 signaling might contribute to the increased local activation during SCW arthritis., Conclusion: In contrast to the proinflammatory role of IL-21 in adaptive immunity, which drives IL-17+IFN+ cells and joint pathology during chronic experimental arthritis, IL-21 also has an important immunosuppressive role, presumably by inhibiting TLR signaling via SOCS-1 and SOCS-3. If this dual role of IL-21 in various immune processes is present in human disease, it could make IL-21 a difficult therapeutic target in rheumatoid arthritis., (Copyright © 2014 by the American College of Rheumatology.)

Interleukin (IL) -21 is produced by Natural Killer T (NKT) cells and CD4(+) T cells and is produced in response to virus infections, where IL-21 has been shown to be essential in adaptive immune responses. Cells from the innate immune system such as Natural Killer (NK) cells and macrophages are also important in immune protection against virus. These cells express the IL-21 receptor (IL-21R) and respond to IL-21 with increased cytotoxicity and cytokine production. Currently, however it is not known whether IL-21 plays a significant role in innate immune responses to virus infections. The purpose of this study was to investigate the role of IL-21 and IL-21R in the innate immune response to a virus infection. We used C57BL/6 wild type (WT) and IL-21R knock out (KO) mice in a murine vaginal Herpes Simplex Virus type 2 (HSV-2) infection model to show that IL-21 - IL-21R signalling is indeed important in innate immune responses against HSV-2. We found that the IL-21R was expressed in the vaginal epithelium in uninfected (u.i) WT mice, and expression increased early after HSV-2 infection. IL-21R KO mice exhibited increased vaginal viral titers on day 2 and 3 post infection (p.i.) and subsequently developed significantly higher disease scores and a lower survival rate compared to WT mice. In addition, WT mice infected with HSV-2 receiving intra-vaginal pre-treatment with murine recombinant IL-21 (mIL-21) had decreased vaginal viral titers on day 2 p.i., significantly lower disease scores, and a higher survival rate compared to infected untreated WT controls. Collectively our data demonstrate the novel finding that the IL-21R plays a critical role in regulating innate immune responses against HSV-2 infection.