- Research
- Open access
- Published:
Interleukin-21 receptor signaling promotes metabolic dysfunction-associated steatohepatitis-driven hepatocellular carcinoma by inducing immunosuppressive IgA+ B cells
Molecular Cancer volume 23, Article number: 95 (2024)
Abstract
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.
Background
Hepatocellular carcinoma (HCC) is the most common malignancy, with the fifth highest incidence and third highest mortality among malignant tumors around the world. Viral infections, alcohol consumption and obesity are the main causes of HCC. With recent advances in the prevention and treatment of hepatitis B virus (HBV) and hepatitis C virus (HCV) infection, the burden of HCC due to viral hepatitis is declining; however, the prevalence of HCC caused by metabolic dysfunction-associated steatotic liver disease (MASLD) [1], previously known as non-alcoholic fatty liver disease (NAFLD), is clearly increasing [2, 3]. Globally, a quarter of the population has NAFLD [2], and approximately 20% of these individuals harbor non-alcoholic steatohepatitis (NASH), now referred to as metabolic dysfunction-associated steatohepatitis (MASH) [1]. MASH, whose key features are steatosis, fibrosis and chronic inflammation [4, 5], can progress to cirrhosis and further to HCC [3]. Generally, MASH is the liver manifestation of metabolic syndrome and is associated with obesity, insulin resistance and dyslipidemia [2, 6], although lean MASH patients that doesn’t fit this definition can be also diagnosed. Although the percentage of patients diagnosed annually with MASH-associated HCC is still relatively low, MASH will inevitably become the most common etiology of HCC in the near future due to the rapidly increasing incidence of obesity and diabetes globally [7]. Thus, more extensive exploration on the tumorigenesis of MASH-associated HCC is urgently needed to develop efficient therapeutic strategies to prevent the progression from MASH to HCC.
Emerging investigations imply that MASH-driven HCC development is accompanied by the accumulation of immune cells within the tumor microenvironment, playing an important role in initiating, maintaining, or exacerbating the transition from MASH to HCC [8]. Dysregulation of immune surveillance is proposed to be one of the new mechanisms that have been implicated in MASH-driven HCC [2]. For instance, Kupffer cells, the liver resident macrophages, represent a first-line defense force in the liver; however, they have been shown to lack effective turnover in MASH [9] and are likely to turn into tumor-associated macrophages (TAMs) to fuel a tumor-promoting inflammatory response under the fibrotic and steatotic tumor environment [10]. In addition, other innate immune cells, such as dendritic cells (DCs) [11] and natural killer (NK) cells [12] may also participate in the process of MASH-driven HCC. Notably, adaptive CD8+ T cells play a central role in the hepatocarcinogenesis in the context of MASLD. Recently, IL-15 produced in the hepatic microenvironment is revealed to downregulate FOXO1 in CD8+ T cells, enabling them to acquire a resident character by upregulating CXCR6, thus making CXCR6+PD1+CD8+ T cells to be capable of eliciting auto-aggressive killing of liver cells to trigger MASH and the transition to HCC [13]. Furthermore, anti–PD-1 therapy fails to reduce the tumor burden in preclinical models of MASH-related HCC and indeed results in the accumulation of CXCR6+PD1+CD8+ T cells [14], suggesting that these cells are likely responsible for the lack of responsiveness of MASH-related HCC to immune checkpoint inhibitors. Compared to CD8+ T cells, naïve CD4+ T cells are more vulnerable to the MASLD microenvironment, where they display higher mortality rate due to the oxidative stress–related cytotoxic effects exerted by free fatty acids [15]. Hence, the loss of CD4+ T cells leads to failure of cancer surveillance in MASH-induced HCC [15]. However, regulatory T (Treg) cells, a subset of CD4+ T cells, are increased in the liver of MASH-induced HCC and promote carcinogenesis by supporting an immunosuppressive microenvironment [16]. Growing evidences clarify the contribution of B cells to MASH as well as MASH-to-HCC transition, and we have also demonstrated that inflammation-induced immunosuppressive IgA+ B cells dismantle anti-cancer immunity by suppressing cytotoxic CD8+ T lymphocyte (CTL) activation in MASH-driven HCC [17]. However, the mechanism of producing of IgA+ B cells is still yet to be explored.
Interleukin-21 (IL-21), a member of the γ chain (γc) cytokine family, is produced mainly by T cells and natural killer T (NKT) cells [18]. Its private receptor, IL-21 receptor (IL-21R), which activates Janus kinase (JAK)-signal transducers and activators of transcription (STAT) signaling upon ligand binding, is expressed by multiple immune cell subsets including, but not limited to, B cells, T cells, NK cells, macrophages and DCs [18, 19]. IL-21/IL-21R signaling plays critical role in immune responses and has been implicated in the regulation of inflammation in various acute and chronic inflammatory diseases, such as cancer [20]. However, the role of IL-21/IL-21R in cancer development remains controversial and has not been extensively investigated in faithful in vivo models. IL-21 was originally demonstrated to be a growth and survival factor in human myeloma cell lines, which is mediated through the activation of the JAK1/STAT3 signaling [21]. In addition, IL-21 has been demonstrated to be immunosuppressive because of its ability to induce IL-10. However, a large number of reports have shown that IL-21 promotes tumor clearance, rather than tumor survival, suggesting that IL-21 is a promising immunotherapeutic agent for cancer treatment [18, 19]. Although limited studies have showed that IL-21/IL-21R is involved in the development of HBV-related diseases, including HCC [22,23,24,25,26]. All of these studies are in vitro or xenograft mouse models studies, lacking of models for spontaneous tumorigenesis in vivo, which is important for studying mechanisms relying on a complicated tumor microenvironment and chronic inflammation. Thus, more extensive exploration for the function of IL-21/IL-21R in HCC tumorigenesis is required. Importantly, IL-21/IL-21R plays a pivotal role in the production of IgA+ B cells [27,28,29,30,31], which are identified as immunosuppressive cells in MASH-driven HCC [17], indicating that IL-21/IL-21R signaling may be involved in the tumorigenesis of MASH-driven HCC. However, the function and underlying mechanism of IL-21/IL-21R in MASH-driven hepatocarcinogenesis are completely unknown.
Herein, we clarified the role of IL-21R in MASH-driven HCC and the underlying mechanism of producing IgA+ B cells. The results revealed that IL-21R played a cancer-promoting role by activating the IL-21R-STAT1-c-Jun/c-Fos regulatory axis, resulting in the production of immunosuppressive IgA+ B cells, and thus attenuated CTL activation in the tumorigenesis of MASH-driven HCC.
Methods
Human tissues
Human tumor tissue sections were obtained from 69 histologically confirmed HCC patients at The First Affiliated Hospital of Jinan University (JNU). The relevant characteristics of the 69 participants are summarized in Table S1. The study was approved by the Institutional Ethics Committee at The First Affiliated Hospital of JNU. Informed consent was obtained from all the participants.
Mouse studies
C57BL/6 control mice and IL-21R-deficient (Il21r−/−) mice were purchased from Biocytogen Pharmaceuticals (Beijing, China) and Jackson Laboratory (JAX), respectively. The genotypes of the Il21r−/− mice were identified according to the genotyping protocol on the website of JAX (https://www.jax.org/Protocol?stockNumber=019115&protocolID=25258). All mice were bred and maintained in filter-topped cages on autoclaved food and water at the JNU animal facility. All procedures for animal experiments were performed in accordance with the Guide for the Care and Use of Laboratory Animals (NIH publications Nos. 80–23, revised 1996) and according to the Animal Ethics Committee of JNU.
For the STAM model, Il21r−/− mice and wild type control mice were performed as previously described [32]. Briefly, male mouse pups were subcutaneously injected with 200 μg of streptozocin (STZ, #S0130, Sigma-Aldrich, St. Louis, MO, USA) at two days after birth and fed with high-fat diet (HFD) containing 60 kcal% fat (#D12492, Research Diets, New Brunswick, NJ, USA) at four weeks of age. The mice developed to MASH and HCC at eight and 20 weeks of age, respectively.
For the WD&High sugar solution&CCl4 model, Il21r−/− mice and wild type control mice were performed as previously described [33]. Briefly, four-weeks-old male mice were begun to feed with western diet (WD) containing 21.2% fat, 41% sucrose, and 1.25% cholesterol (#TD. 120528, Harlan, Madison, WI, USA) and a high sugar solution containing 23.1 g/L d-fructose (#F0127, Sigma-Aldrich, St. Louis, MO, USA) and 18.9 g/L d-glucose (#G8270, Sigma-Aldrich, St. Louis, MO, USA). Simultaneously, CCl4 dissolved in corn oil was injected intraperitoneally at the dose of 0.2 μL/g of body weight once per week. The mice developed to MASH and HCC at 12 and 24 weeks of age, respectively.
For adoptive lymphocyte transfer, male mice with HCC tumors were intravenously injected via the inner canthus veniplex with 200 μg anti-mouse CD20 antibody (#BE0356, Clone MB20-11, Bioxcell, Lebanon, NH, USA) per mouse to deplete B cells in vivo. Two days later, single-cell suspensions of spleens from either Il21r−/− mice or wild type control mice were incubated with biotin-labeled anti-mouse CD19 antibody (#115504, Clone 6D5, Biolegend, San Diego, CA, USA), and the CD19+ B cells were isolated from the suspensions by using Streptavidin Particles Plus-DM (#557812, BD Biosciences, San Diego, CA, USA). The isolated CD19+ B cells (5 × 106 cells per mouse) were randomly transferred into the mice previously treated with anti-mouse CD20 antibody by inner canthus veniplex injection. Mice were sacrificed seven weeks after lymphocyte transfer.
For antibody neutralization experiment, male mice with HCC tumors were injected intraperitoneally with 200 μg anti-mouse IL-21R antibody (#BE0258, Clone 4A9, Bioxcell, Lebanon, NH, USA) or its isotype control (#A2123, Clone 2A3, Selleckchem, Houston, TX, USA) twice a week for five weeks.
Analysis of gene expression
The expression levels of target genes were assessed by western blotting, immunohistochemistry (IHC) staining, immunofluorescent (IF) staining or real time quantitative RT-PCR (qRT-PCR). The antibodies used for western blotting, IHC and IF staining are listed in Table S2, and the primers used for qRT-PCR are listed in Table S3.
Chromatin immunoprecipitation (ChIP) assay
CD19+ B cells were isolated from splenocytes by using biotin-labeled anti-mouse CD19 antibody and Streptavidin Particles Plus-DM. The isolated CD19+ B cells (~ 3 × 107) were subsequently cross-linked with formaldehyde, and the reaction was stopped with glycine. The cross-linked cells were washed with cold 1 × phosphate-buffered saline (PBS) and the pellets were re-suspended with 300 μL of ChIP cell lysis buffer supplemented with protease inhibitor [34], and incubated on ice for 10 min. The lysed cells were centrifuged and the pellets were re-suspended with 300 μL of MNase digestion buffer [34], followed by digestion with MNase (#10011, Cell Signaling Technology, Danvers, MA, USA) (100 gel units/1 × 107 cells) at 37 °C for exactly 10 min, mixing by inversion every 2.5 min. After stopping digestion, the suspensions were centrifuged and the pellets were with re-suspended with 400 μL of ChIP dilution buffer [34] supplemented with protease inhibitor, followed by sonication (amplitude 2, processed time 30 s, pulse ON 5 s, pulse OFF 30 s). Thereafter, the chromatin complexes were immunoprecipitated using anti-c-Jun (#9165S, Cell Signaling Technology, Danvers, MA, USA), anti-c-Fos (#4384S, Cell Signaling Technology, Danvers, MA, USA) or the corresponding isotype-matched IgG overnight and then collected by incubation with Protein A/G Magnetic Beads (#88803, Thermo Fisher Pierce, Waltham, MA, USA) for two hours. The DNA–protein-beads complex was washed and then eluted. Subsequently, the DNA–protein cross-link was reversed by heating, and DNA was purified from the eluted solution and subjected to qRT-PCR using primers covering the activating protein 1 (AP-1) binding sites. The sequences of the primers are listed in Table S3.
Whole transcriptome sequencing processing and analysis
Total RNA was extracted from the livers or tumors using TRI reagent (#T9424, Sigma-Aldrich, St. Louis, MO, USA), and was subsequently sent to BGI Tech Solutions Co., Ltd (Shenzhen, China) for whole transcriptome sequencing according to their standardized procedures.
For differential expression analysis, the sequencing data were performed using DESeq2, Q value ≤ 0.05 was used to judge the significance of expression difference. For Gene Ontology (GO) enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis, target genes were subjected to enrichment analysis using phyper, a function of R. The P-value was corrected using the Bonferroni method, and a corrected P-value ≤ 0.05 was taken as the threshold to define the significantly enriched GO or KEGG terms. Additionally, gene set enrichment analysis (GSEA) [35, 36] was performed to identify the hallmark gene sets according to the instruction. Heirarchical clustering was performed using Cluster 3.0 (Michiel de Hoon, Tokyo, Japan) and the data were visualized using Java TreeView (Eisen Lab, Berkeley, CA, USA).
Statistical analysis
Data were presented as mean ± SEM (standard error of the mean). The differences between two groups were analyzed by Student's t test. One-way analysis of variance (ANOVA) was used to compare three or more groups, while two-way ANOVA was used to compare the differences between groups that have been split on two independent categorical variables. All the statistical analyses were performed with GraphPad Prism 6.0 (GraphPad Software, Inc., San Diego, CA, USA). All the statistical tests were two-sided, and P < 0.05 was considered statistically significant.
Results
IL-21R is increased in MASH-driven HCC and associated with poor prognosis in human HCC patients
While the role of IL-21R signaling has been investigated in infectious and inflammatory diseases, unequivocal evidence about its possible contribution to tumor development in vivo is still poorly understood. To explore the role of IL-21R in HCC, we first assessed the protein level of IL-21R by using IHC staining (Fig. 1A). We found that patients with high IL-21R expression exhibited poor relapse-free survival, more advanced stage of HCC and more liver injury (Fig. 1B, C and Table S1). We further observed that there were 21 patients with different grades of steatosis according to the IHC results. Interestingly, most of these patients expressed high IL-21R (17/21 = 81%), and most of the patients with high IL-21R expression exhibited moderate to severe steatosis (15/17 = 88%), whereas most of the patients with low IL-21R expression displayed mild steatosis (3/4 = 75%, Fig. 1D and Table S4), suggesting that IL-21R might play an important role in MASH-related HCC. Moreover, we evaluated the impact of IL-21R in human HCC patients on relapse-free survival and overall survival using TCGA data. Consistently, we obtained similar results in two independent cohorts (Fig. S1 and Table S5).
Thereafter, we applied a well-established mouse model of MASH-driven HCC (Fig. S2A-E) to investigate the role of IL-21R in MASH-driven HCC. We found that both the mRNA (Fig. 1E) and protein (Fig. 1F and Fig. S2F) levels of IL-21R were upregulated in the liver tumors.
Taken together, these observations in human and mouse HCC samples suggest a potential cancer-promoting role of IL-21R in MASH-driven HCC.
IL-21R–deficient mice impede MASH-driven HCC development
To determine the role of IL-21R in MASH-driven HCC development, the STAM model was applied between IL-21R-deficient mice (Il21r−/−, Fig. S3A) and wild type (WT) controls. We found that ablation of IL-21R did not affect weight gain, liver damage, colon length and glucose tolerance (Fig. 2A-C and Fig. S3B, C). However, liver weight, HCC tumorigenesis and spleen weight were significantly reduced in Il21r−/− mice compared to WT controls (Fig. 2D-H and Fig. S3D). Consistent with the limited tumor burden, HCC tumors from IL-21R-deficient mice were also characterized by lower expression of proliferation markers, including cyclin D1 (Ccnd1), antigen identified by monoclonal antibody Ki67 (Mki67) and cyclin dependent kinase 4 (Cdk4) (Fig. S3E), implying reduced proliferative capacity of tumors in the absence of IL-21R, which was further confirmed by IHC staining for Ki67 (Fig. S3F). Nonetheless, ablation of IL-21R did not change the apoptotic cell death (Fig. S3G). We further confirmed that IL-21R-deficient mice impede MASH-driven HCC tumorigenesis in another mouse model of MASH-driven HCC (Fig. S4A-H).
Furthermore, IL-21R-deficient mice showed no obvious alterations in hepatocyte ballooning and Kupffer cells infiltration (Fig. 2I). Importantly, lipid accumulation was dramatically reduced in HCC tumors from Il21r−/− mice (Fig. 2I, J). Consistently, both triglyceride and cholesterol levels decreased in the serum and tumor tissue of Il21r−/− mice (Fig. 2K, L). Similar results were obtained using another mouse model of MASH-driven HCC (Fig. S4I-L).
Collectively, these data clearly demonstrate that IL-21R promotes MASH-driven HCC and affects lipid accumulation.
Ablation of IL-21R enhances cytotoxic CD8+ T lymphocyte activation in vivo
To elucidate the potential mechanisms by which IL-21R affects MASH-driven HCC, we first conducted whole transcriptome sequencing on isolated HCC tumors from Il21r−/− mice and WT controls. GO and KEGG pathway analyses revealed that differential genes were enriched in inflammation-, immune- and metabolism-related pathways (Fig. 3A, B). Further GSEA analysis showed that the Il21−/− group enriched gene sets related to inflammation and immune, whereas the WT group enriched in metabolism-related gene sets (Fig. 3C and Fig. S5A, B). The inflammation- and immune-related genes with differential expression were clustered (Fig. 3D) and most of them, including Cd44, Cd74, Tnf, were confirmed to be upregulated in the tumors of Il21r−/− mice (Fig. 3E). We also found that lipid-related genes decreased in the tumors of Il21r−/− mice, but no alterations of fibrosis-related genes (Fig. 3F and Fig. S5C), confirming the results of histology (Fig. 2I). Moreover, pro-inflammation cytokines, such as Il1b, Il12b and Il18, were upregulated in the tumor of Il21r−/− mice; whereas anti-inflammation cytokines, including Il10 and Il22 were decreased in the tumor of Il21r−/− mice (Fig. S5D).
In the light of enrichment of inflammation- and immune-related pathways, we next performed flow cytometry (Fig. S6) to explore the difference of immune microenvironment between Il21r−/− mice and WT controls. Compared with WT controls, Il21r−/− mice had remarkably fewer liver CD8+ T cells (Fig. 4A), but equal numbers of spleen CD8+ T cells (Fig. S7A). Nonetheless, compared to WT controls, Il21r−/− mice had more effector and degranulating CD8+ T cells in both liver and spleen (Fig. 4B-F and Fig. S7B), but had a similar percentage of Tim3+CD8+ exhausted T cells (Fig. S7C). Similar results were obtained using another MASH-driven HCC mouse model (Fig. S7D-F). Additionally, ablation of IL-21R had no effect on total CD4+ T cells (Fig. 4G and Fig. S8A), although the numbers of HCC-infiltrating Th1, Th17 and Treg cells increased in the absence of IL-21R (Fig. S8B-E), while having little effect on liver B cells (Fig. S9A).
These data indicate that MASH-driven HCC development is impeded in Il21r−/− mice mainly by enhanced cytotoxic CTL activation.
IL-21R-deficient mice have fewer IgA+ B cells, and thus impede MASH-driven hepatocarcinogenesis
IL-21R is expressed on a broad array of cell types including B cells and T cells. First, we checked which cells expressed IL-21R in MASH-driven HCC by using flow cytometry and immunofluorescence staining. Interestingly, we found that IL-21R was mostly expressed on CD19+ B cells (Fig. S9B, C). Subsequently, we applied an adoptive transfer experiment (Fig. 5A and Fig. S9D) to determine whether the B cells from Il21r−/− mice had any effect on HCC tumorigenesis. Indeed, adoptive transfer of B cells from IL-21R-deficient mice into WT mice during the HCC stage suppressed tumorigenesis, accompanied by splenomegaly and decreased lipid accumulation, but no change for the liver weight (Fig. 5A-E). Even though the percentage of CD8+ T cells decreased, both effector and degranulating CD8+ T cells significantly increased after adoptive transfer with B cells from Il21r−/− mice (Fig. 5F,G and Fig. S9E-H).
Next, we investigated how IL-21R affected B cells during MASH-driven HCC tumorigenesis. We noticed that the differential genes between Il21r−/− mice and WT controls were also enriched in intestinal immune network for IgA production according to the KEGG pathway analysis (Fig. 3B). Given that IgA+ B cells were proven to be immunosuppressive cells in our previous work [17], we next checked whether IL-21R regulated the emergence of IgA+ B cells. Compared with WT controls, Il21r−/− mice had significantly fewer liver and spleen IgA+ B cells (Fig. 5H and Fig. S10A, B). Moreover, the levels of serum IgA in Il21r−/− mice were significantly decreased (Fig. 5I). These data indicate that IL-21R-deficient mice have fewer IgA+ B cells and thus impede MASH-driven HCC development.
To further explore the mechanism by which IL-21R regulated IgA+ B cells, we first confirmed that ablation of IL-21R decreased the mRNA level of Igha in B cells (Fig. 6A). We thus further evaluated the transcriptional regulation of Igha. Five consensus AP-1 binding sites were predicted (referred to Site A, B, C, D and E, respectively) between 2 kb upstream of the transcription start site (TSS) and the TSS (Fig. S11A). AP-1 is an inducible transcription factor complex consisting of a group of transcription factors including Jun, Fos and ATF family proteins. ChIP assay confirmed the existence of a direct interaction between c-Jun/c-Fos and Site A, B, D of the Igha promoter in B cells from WT controls, but not in B cells from Il21r−/− mice (Fig. 6B), suggesting that c-Jun/c-Fos modulates the transcription of Igha by binding to the Site A, B, D. IL-21 has been shown to activate the JAK-STAT pathway, and STAT1 can interact with AP-1 to trigger transcriptional activation [37]. Thereafter, we further showed that IL-21R, phosphorylated STAT1, STAT1 and c-Jun were upregulated in B cells during MASH-driven heptocacinogenesis (Fig. S11B), and ablation of IL-21R decreased the protein levels of phosphorylated STAT1, c-Jun and c-Fos (Fig. 6C). Furthermore, STAT1 inhibitor blocked the elevation of c-Jun without change the expression of IL-21R, resulting in the downregulation of Igha mRNA level in B cells from mice at HCC stage (Fig. S11C, D). Importantly, blockade of IL-21R signaling with an IL-21R blocking antibody induced tumor regression by enhancing cytotoxic CTL activation (Fig. 6D-F and Fig. S12).
Taken together, we suggest an IL-21R-STAT1-c-Jun/c-Fos-IgA regulatory pathway (Fig. 6G): IL-21R is increased in MASH-driven HCC, resulting in upregulation of phosphorylated STAT1, which acts as a co-activator of c-Jun/c-Fos. The c-Jun/c-Fos complex binds to the promoter of Igha and promotes the transcription of Igha, resulting in the emergence of immunosuppressive IgA+ B cells, and thereby inhibits CTL activation and eventually facilitates MASH-driven HCC. Importantly, ablation of IL-21R can reduce the induction of IgA+ B cells, and thus enhances CTL activation and impedes MASH-driven HCC. These findings indicate that IL-21R promotes the tumorigenesis of MASH-driven HCC by inducing immunosuppressive IgA+ B cells and disrupting the IL-21R regulatory axis may be beneficial for MASH-driven HCC therapy.
Discussion
In the present study, we revealed that the IL-21R-STAT1-c-Jun/c-Fos-IgA regulatory pathway was activated during MASH-driven hepatocarcinogenesis, leading to the induction of immunosuppressive IgA+ B cells, and thereby inhibited CTL activation and eventually facilitated MASH-driven HCC in vivo. These findings support the immunosuppressive effect of IL-21/IL-21R in tumorigenesis. To our knowledge, this is the first attempt to circumstantiate the function of IL-21R in MASH-driven hepatocarcinogenesis, and to elucidate the mechanism by which IL-21R induces IgA+ B cells.
IL-21/IL-21R signaling has been implicated in the regulation of inflammation in various acute and chronic inflammatory diseases. The role of IL-21/IL-21R in cancer development remains obscure and has not been extensively investigated in faithful in vivo models. To date, the function and mechanism of IL-21/IL-21R during MASH-driven hepatocarcinogenesis have not yet been reported. Here, we presented the following evidences: (i) Assessment of the levels of IL-21R by using IHC staining revealed that elevated IL21R expression was correlated with advanced stage of HCC and poor survival in our cohort (n = 69), and the conclusion was validated in two additional independent cohorts (n = 80 and 365, respectively) according to the TCGA database. Importantly, HCC patients with high IL-21R expression exhibited severe steatosis. (ii) Both the mRNA and protein levels of IL-21R were elevated in liver tumors in a well-established mouse model of MASH-driven HCC, the STAM model (n = 52 in total). (iii) By using two different in vivo mouse models of MASH-driven HCC, the STAM model and the WD&High sugar solution&CCl4 model, we revealed that ablation of IL-21R obstructed MASH-driven HCC by enhancing CTL activation with large number of mice (n = 86 in total). (iv) Adoptive transfer experiment (n = 14 in total) demonstrated that adoptive transfer of B cells from Il21r−/− mice into wild type mice during the HCC stage suppressed MASH-driven hepatocarcinogenesis in recipient mice. (v) According to the results of flow cytometry, ELISA, western blotting and ChIP assay, we elucidated that ablation of IL-21R limited the induction of immunosuppressive IgA+ B cells via the IL-21R-STAT-1-c-Jun/c-Fos-IgA regulatory axis. (vi) Blockade of IL-21R signaling with an IL-21R blocking antibody induced tumor regression of MASH-driven HCC. Collectively, our data suggest that, perhaps in MASH-driven HCC, IL-21R plays a pro-tumorigenic inflammatory role and that blockade of IL-21R signaling will result in attenuated MASH-driven HCC development.
IL-21 was originally demonstrated to be a growth and survival factor in human myeloma cell lines, which was mediated through the activation of JAK1/STAT3 signaling [21]. However, since then, a large number of reports have shown that IL-21 promotes tumor clearance, rather than tumor survival [19]. Subsequently, the discovery of the immunosuppressive actions of IL-21 further suggested that IL-21 is a “double-edged sword”: IL-21 stimulation may lead to either the induction or suppression of immune responses [18]. Thus, both the stimulatory and suppressive effects of IL-21 must be considered during the clinical use of IL-21-related immunotherapeutic agents. Moreover, the biological effects of IL-21 are also influenced by the presence of other cytokines or signaling molecules in the tumor microenvironment [18]. As the private receptor of IL-21, IL-21R can also be either immunosuppressive or immunostimulatory depending on the environmental context. To date, only one publication attempted to address the anti-tumor role of IL-21R in HCC development [24], particularly utilizing xenograft mouse models. Since such models have clear advantages in terms of rapid and uniform tumor growth, they may be less perfect for studying mechanisms which rely on a complicated tumor microenvironment and chronic inflammation, a key component regulated by cytokines in cancer. In contrast to their conclusion, our study demonstrated that ablation of IL-21R impeded liver cancer development, particularly MASH-driven HCC, in two different mouse models of spontaneous MASH-driven HCC: the STAM model and the WD&High sugar solution&CCl4 model. Along with human data on poor survival, advanced tumor stages and severe steatosis in HCC patients with high IL21R expression, our work implies that IL-21R plays a cancer-promoting role in MASH-driven HCC. An essential component of this mechanism is mediated by immunosuppressive IgA+ B cells via the IL-21R-STAT-1-c-Jun/c-Fos-IgA regulatory axis.
B cells are well known as key mediators of humoral immune responses via the production of antibodies, and immunoglobulin A is one of the most abundantly produced antibody isotypes. It has been reported that immunoglobulin A and IgA+ B cells exert anti- or pro- tumor effect in different tumor types [17, 38,39,40,41,42]. Elevated levels of intratumoral IgA have been shown to be associated with poor outcomes in patients with bladder cancer [38], colorectal cancer [39] and melanoma [40]. Additionally, our colleagues discovered that IgA+ B cells, which expressed PD-L1, IL-10 and Fas-L, could suppress anti-tumor immunity in oxaliplatin-treated pancreatic cancer [41], one of the mechanisms responsible for chemotherapy tolerance. Thereafter, together with our colleague, we demonstrated that inflammation-induced immunosuppressive IgA+ B cells dismantle anti-cancer immunity by suppressing CTL activation during MASH-driven HCC [17]. Although there is more than sufficient evidence indicating that IgA+ B cells exert prominent immunosuppressive effects, the anti-tumor effect of IgA+ B cells has also been reported. For instance, in ovarian cancer, tumor antigen-specific and tumor antigen-independent IgA responses antagonized the growth of tumor by governing coordinated tumor cell, T cell and B cell responses [42]. With respect to the reasons for the induction of IgA+ B cells, emerging evidence indicates that a combination of host, environmental and tumor factors mediates IgA class switching through upregulating class switching-related genes. First, TGF-β, which is highly expressed in the tumor microenvironment, can activate the constant heavy chain α (Cα) gene promoter to trigger T-cell-independent IgA class switching [43, 44]. Second, fatty acids or microbial products can activate Toll like receptors, which promote the activation of IκB kinase (IKK) complex to trigger the translocation of IκB-free NF-κB to the nucleus, and initiate IgA class switching by binding to the activation-induced cytidine deaminase (AID) gene promoter [44,45,46]. Nevertheless, no specific transcriptional markers have been identified to exclusively define the IgA+ B cells.
It is thought that IL-21/IL-21R plays a pivotal role in IgA production. When combined with IL-4 and anti-CD40, IL-21 can induce class switch recombination to IgA and differentiation of IgA+ B cells in human peripheral blood mononuclear cells (PBMCs) [47]. Additionally, IL-21 ensures TGF-β-induced IgA isotype expression in mouse Peyer’s patches [27]. Moreover, it’s widely explored that IL-21 promotes intestinal IgA+ B cell production [28,29,30,31], possibly through upregulating differentiation-related and class switching–related genes such as Aicda, Ski and Prdm1 [30, 31]. Additionally, in the presence of atypical commensals such as segmented filamentous bacteria and Helicobacter, IL-21R-deficient mice exhibit reduced IgA+ B cells in the germinal center, Peyer’s patches and small intestine [29]. However, the relationship between IL-21/IL-21R and IgA in liver or tumor microenvironment is unclear. Consist with the mouse [29] and human clinical data [48], here we presented that IL-21R-deficient mice exhibited reduced numbers of spleen or liver/tumor IgA+ B cells and soluble IgA during MASH-driven hepatocarcinogenesis. Importantly, we elucidated the mechanism in which IL-21R activated IgA via the IL-21R-STAT-1-c-Jun/c-Fos-IgA regulatory axis. Moreover, we demonstrated that AP-1, mainly composed of c-Jun and c-Fos, was the right transcription factor for identifying IgA+ B cells.
To our knowledge, the current study provides the most comprehensive investigation regarding the cancer-promoting role of IL-21R by exerting immunosuppressive characteristics of B cells in MASH-driven hepatocarcinogenesis to date, implying that targeting IL-21R signaling represents a potential therapeutic strategy for cancer therapy. However, our study is not without limitations. Firstly, B cell-specific knockout of IL-21R mice cannot be generated by crossing Il21r flox/flox mice with CD19-cre or Mb1-cre mice in this study because all of these genes are located on chromosome seven. Secondly, there are no gross defects in the development of the immune system in Il21r−/− mice [49] or human [48]. However, certain patients with loss-of-function mutations in the IL-21R gene suffer from recurrent respiratory and gastrointestinal infections, additionally have cryptosporidiosis, leading to secondary cholangitis and liver disease according to some case reports [48, 50, 51]. Therefore, precision therapy based on IL-21R deficiency should be considered in both experiment and clinic in the future. Thirdly, according to our IHC staining result and the data from The Human Protein Atlas (https://www.proteinatlas.org/), IL-21R also expressed in the cytoplasma and membrane of tumor cells in human liver cancer. The role and mechanism of IL-21R in HCC cells are still yet to be explored.
Conclusions
In conclusion, our findings explore the cancer-promoting role of IL-21R in MASH-driven hepatocarcinogenesis and elucidate the mechanism by which IL-21R activates IgA via the IL-21R-STAT-1-c-Jun/c-Fos-IgA regulatory axis. Thus, targeting IL-21R signaling represents a potential therapeutic strategy for cancer therapy.
Availability of data and materials
Whole transcriptome sequencing data in this study are available in the Sequence Read Archive (SRA) database under accession number PRJNA1035974. All data supporting the conclusions of this article are included within the article and its additional files. Any other information is available from the corresponding authors upon reasonable request.
Abbreviations
- HCC:
-
Hepatocellular carcinoma
- MASLD:
-
Metabolic dysfunction-associated steatotic liver disease
- MASH:
-
Metabolic dysfunction-associated steatohepatitis
- NASH:
-
Non-alcoholic steatohepatitis
- NAFLD:
-
Non-alcoholic fatty liver disease
- IL-21:
-
Interleukin-21
- IL-21R:
-
Interleukin-21 receptor
- CTL:
-
CD8+ T lymphocyte
- JAK:
-
Janus kinase
- STAT:
-
Signal transducers and activators of transcription
- AP-1:
-
Activating protein 1
- TSS:
-
Transcription start site
- HBV:
-
Hepatitis B virus
- HCV:
-
Hepatitis C virus
- HFD:
-
High-fat diet
- WD:
-
Western diet
- DMEM:
-
Dulbecco's modified Eagle's medium
- FBS:
-
Fetal bovine serum
- PBS:
-
Phosphate-buffered saline
- qRT-PCR:
-
Real time quantitative RT-PCR
- HE:
-
Haematoxylin and eosin
- IHC:
-
Immunohistochemistry
- IF:
-
Immunofluorescent
- SR:
-
Sirius Red
- ORO:
-
Oil Red O
- ChIP:
-
Chromatin immunoprecipitation
- ELISA:
-
Enzyme-linked immunosorbent assay
- GO:
-
Gene Ontology
- KEGG:
-
Kyoto Encyclopedia of Genes and Genomes
- GSEA:
-
Gene Set Enrichment Analysis
- SEM:
-
Standard error of the mean
- ANOVA:
-
Analysis of variance
References
Rinella ME, Lazarus JV, Ratziu V, Francque SM, Sanyal AJ, Kanwal F, et al. A multisociety Delphi consensus statement on new fatty liver disease nomenclature. J Hepatol. 2023;79:1542–56.
Huang DQ, El-Serag HB, Loomba R. Global epidemiology of NAFLD-related HCC: trends, predictions, risk factors and prevention. Nat Rev Gastroenterol Hepatol. 2021;18:223–38.
Tan DJH, Ng CH, Lin SY, Pan XH, Tay P, Lim WH, et al. Clinical characteristics, surveillance, treatment allocation, and outcomes of non-alcoholic fatty liver disease-related hepatocellular carcinoma: a systematic review and meta-analysis. Lancet Oncol. 2022;23:521–30.
Febbraio MA, Reibe S, Shalapour S, Ooi GJ, Watt MJ, Karin M. Preclinical Models for Studying NASH-Driven HCC: How Useful Are They? Cell Metab. 2019;29:18–26.
Anstee QM, Reeves HL, Kotsiliti E, Govaere O, Heikenwalder M. From NASH to HCC: current concepts and future challenges. Nat Rev Gastroenterol Hepatol. 2019;16:411–28.
Kasper P, Martin A, Lang S, Kutting F, Goeser T, Demir M, et al. NAFLD and cardiovascular diseases: a clinical review. Clin Res Cardiol. 2021;110:921–37.
Ioannou GN. Epidemiology and risk-stratification of NAFLD-associated HCC. J Hepatol. 2021;75:1476–84.
Pinter M, Pinato DJ, Ramadori P, Heikenwalder M. NASH and Hepatocellular Carcinoma: Immunology and Immunotherapy. Clin Cancer Res. 2023;29:513–20.
Tran S, Baba I, Poupel L, Dussaud S, Moreau M, Gelineau A, et al. Impaired Kupffer Cell Self-Renewal Alters the Liver Response to Lipid Overload during Non-alcoholic Steatohepatitis. Immunity. 2020;53:627–640 e625.
Kohlhepp MS, Liu H, Tacke F, Guillot A. The contradictory roles of macrophages in non-alcoholic fatty liver disease and primary liver cancer-Challenges and opportunities. Front Mol Biosci. 2023;10:1129831.
Deczkowska A, David E, Ramadori P, Pfister D, Safran M, Li B, et al. XCR1(+) type 1 conventional dendritic cells drive liver pathology in non-alcoholic steatohepatitis. Nat Med. 2021;27:1043–54.
Martinez-Chantar ML, Delgado TC, Beraza N. Revisiting the role of natural killer cells in non-alcoholic fatty liver disease. Front Immunol. 2021;12:640869.
Dudek M, Pfister D, Donakonda S, Filpe P, Schneider A, Laschinger M, et al. Auto-aggressive CXCR6(+) CD8 T cells cause liver immune pathology in NASH. Nature. 2021;592:444–9.
Pfister D, Nunez NG, Pinyol R, Govaere O, Pinter M, Szydlowska M, et al. NASH limits anti-tumour surveillance in immunotherapy-treated HCC. Nature. 2021;592:450–6.
Ma C, Kesarwala AH, Eggert T, Medina-Echeverz J, Kleiner DE, Jin P, et al. NAFLD causes selective CD4(+) T lymphocyte loss and promotes hepatocarcinogenesis. Nature. 2016;531:253–7.
Wang H, Zhang H, Wang Y, Brown ZJ, Xia Y, Huang Z, et al. Regulatory T-cell and neutrophil extracellular trap interaction contributes to carcinogenesis in non-alcoholic steatohepatitis. J Hepatol. 2021;75:1271–83.
Shalapour S, Lin XJ, Bastian IN, Brain J, Burt AD, Aksenov AA, et al. Inflammation-induced IgA+ cells dismantle anti-liver cancer immunity. Nature. 2017;551:340–5.
Leonard WJ, Wan CK. IL-21 Signaling in Immunity. F1000Res. 2016;5:F1000.
Mehta DS, Wurster AL, Grusby MJ. Biology of IL-21 and the IL-21 receptor. Immunol Rev. 2004;202:84–95.
Davis MR, Zhu Z, Hansen DM, Bai Q, Fang Y. The role of IL-21 in immunity and cancer. Cancer Lett. 2015;358:107–14.
Brenne AT, Ro TB, Waage A, Sundan A, Borset M, Hjorth-Hansen H. Interleukin-21 is a growth and survival factor for human myeloma cells. Blood. 2002;99:3756–62.
Chen HM, Liu HL, Yang YC, Cheng XL, Wang YF, Xing FF, et al. Serum IL-21 levels associated with chronic hepatitis B and hepatitis B-related liver failure. Exp Ther Med. 2014;7:1013–9.
Pan Q, Yu Y, Tang Z, Xi M, Jiang H, Xun Y, et al. Increased levels of IL-21 responses are associated with the severity of liver injury in patients with chronic active hepatitis B. J Viral Hepatitis. 2014;21:e78–88.
Zheng X, Zhou Y, Yi X, Chen C, Wen C, Ye G, et al. IL-21 receptor signaling is essential for control of hepatocellular carcinoma growth and immunological memory for tumor challenge. Oncoimmunology. 2018;7:e1500673.
Jin Y, Sun Z, Geng J, Yang L, Song Z, Song H, et al. IL-21 reinvigorates exhausted natural killer cells in patients with HBV-associated hepatocellular carcinoma in STAT1-depedent pathway. Int Immunopharmacol. 2019;70:1–8.
Zhao C, Wu X, Chen J, Qian G. The therapeutic effect of IL-21 combined with IFN-gamma inducing CD4(+)CXCR5(+)CD57(+)T cells differentiation on hepatocellular carcinoma. J Adv Res. 2022;36:89–99.
Seo GY, Youn J, Kim PH. IL-21 ensures TGF-beta 1-induced IgA isotype expression in mouse Peyer’s patches. J Leukoc Biol. 2009;85:744–50.
Cao AT, Yao S, Gong B, Nurieva RI, Elson CO, Cong Y. Interleukin (IL)-21 promotes intestinal IgA response to microbiota. Mucosal Immunol. 2015;8:1072–82.
Cho H, Jaime H, de Oliveira RP, Kang B, Spolski R, Vaziri T, et al. Defective IgA response to atypical intestinal commensals in IL-21 receptor deficiency reshapes immune cell homeostasis and mucosal immunity. Mucosal Immunol. 2019;12:85–96.
Hashiguchi M, Kashiwakura Y, Kanno Y, Kojima H, Kobata T. IL-21 and IL-5 coordinately induce surface IgA(+) cells. Immunol Lett. 2020;224:21–7.
Huang X, Yang W, Yao S, Bilotta AJ, Lu Y, Zhou Z, et al. IL-21 Promotes Intestinal Memory IgA Responses. J Immunol. 2020;205:1944–52.
Fujii M, Shibazaki Y, Wakamatsu K, Honda Y, Kawauchi Y, Suzuki K, et al. A murine model for non-alcoholic steatohepatitis showing evidence of association between diabetes and hepatocellular carcinoma. Med Mol Morphol. 2013;46:141–52.
Tsuchida T, Lee YA, Fujiwara N, Ybanez M, Allen B, Martins S, et al. A simple diet- and chemical-induced murine NASH model with rapid progression of steatohepatitis, fibrosis and liver cancer. J Hepatol. 2018;69:385–95.
Yamakawa T, Itakura K. Chromatin immunoprecipitation assay using micrococcal nucleases in mammalian cells. J Vis Exp. 2019;147. PMID: 31132037.
Mootha VK, Lindgren CM, Eriksson KF, Subramanian A, Sihag S, Lehar J, et al. PGC-1alpha-responsive genes involved in oxidative phosphorylation are coordinately downregulated in human diabetes. Nat Genet. 2003;34:267–73.
Subramanian A, Tamayo P, Mootha VK, Mukherjee S, Ebert BL, Gillette MA, et al. Gene set enrichment analysis: a knowledge-based approach for interpreting genome-wide expression profiles. Proc Natl Acad Sci USA. 2005;102:15545–50.
Xu W, Comhair SA, Zheng S, Chu SC, Marks-Konczalik J, Moss J, et al. STAT-1 and c-Fos interaction in nitric oxide synthase-2 gene activation. Am J Physiol Lung Cell Mol Physiol. 2003;285:L137–148.
Welinder C, Jirstrom K, Lehn S, Nodin B, Marko-Varga G, Blixt O, et al. Intra-tumour IgA1 is common in cancer and is correlated with poor prognosis in bladder cancer. Heliyon. 2016;2:e00143.
Liu R, Lu Z, Gu J, Liu J, Huang E, Liu X, et al. MicroRNAs 15A and 16–1 Activate Signaling Pathways That Mediate Chemotaxis of Immune Regulatory B cells to Colorectal Tumors. Gastroenterology. 2018;154:637–651 e637.
Bosisio FM, Wilmott JS, Volders N, Mercier M, Wouters J, Stas M, et al. Plasma cells in primary melanoma. Prognostic significance and possible role of IgA. Mod Pathol. 2016;29:347–58.
Shalapour S, Font-Burgada J, Di Caro G, Zhong Z, Sanchez-Lopez E, Dhar D, et al. Immunosuppressive plasma cells impede T-cell-dependent immunogenic chemotherapy. Nature. 2015;521:94–8.
Biswas S, Mandal G, Payne KK, Anadon CM, Gatenbee CD, Chaurio RA, et al. IgA transcytosis and antigen recognition govern ovarian cancer immunity. Nature. 2021;591:464–70.
Coffman RL, Lebman DA, Shrader B. Transforming growth factor beta specifically enhances IgA production by lipopolysaccharide-stimulated murine B lymphocytes. J Exp Med. 1989;170:1039–44.
Cerutti A. The regulation of IgA class switching. Nat Rev Immunol. 2008;8:421–34.
Kunisawa J, Hashimoto E, Inoue A, Nagasawa R, Suzuki Y, Ishikawa I, et al. Regulation of intestinal IgA responses by dietary palmitic acid and its metabolism. J Immunol. 2014;193:1666–71.
Kim M, Qie Y, Park J, Kim CH. Gut Microbial Metabolites Fuel Host Antibody Responses. Cell Host Microbe. 2016;20:202–14.
Borte S, Pan-Hammarstrom Q, Liu C, Sack U, Borte M, Wagner U, et al. Interleukin-21 restores immunoglobulin production ex vivo in patients with common variable immunodeficiency and selective IgA deficiency. Blood. 2009;114:4089–98.
Kotlarz D, Zietara N, Uzel G, Weidemann T, Braun CJ, Diestelhorst J, et al. Loss-of-function mutations in the IL-21 receptor gene cause a primary immunodeficiency syndrome. J Exp Med. 2013;210:433–43.
Kasaian MT, Whitters MJ, Carter LL, Lowe LD, Jussif JM, Deng B, et al. IL-21 limits NK cell responses and promotes antigen-specific T cell activation: a mediator of the transition from innate to adaptive immunity. Immunity. 2002;16:559–69.
Stepensky P, Keller B, Abuzaitoun O, Shaag A, Yaacov B, Unger S, et al. Extending the clinical and immunological phenotype of human interleukin-21 receptor deficiency. Haematologica. 2015;100:e72–76.
Erman B, Bilic I, Hirschmugl T, Salzer E, Cagdas D, Esenboga S, et al. Combined immunodeficiency with CD4 lymphopenia and sclerosing cholangitis caused by a novel loss-of-function mutation affecting IL21R. Haematologica. 2015;100:e216–219.
Acknowledgements
We gratefully acknowledge the constructive suggestion from Dr. Michael Karin of University of California San Diego. We also thank Dr. Xiaowei Li of Jinan University for technical support.
Funding
This project is supported by grants from the National Natural Science Foundation of China (82372797, 32030036 and 31830021), Tip-top Scientific and Technical Innovative Youth Talents of Guangdong special support program (2019TQ05Y110), Pearl River S & T Nova Program of Guangzhou, China (201906010085), the National Key Research and Development Program of China (2020YFA0803502) and the 111 Project (B16021).
Author information
Authors and Affiliations
Contributions
Y.X., Y.H. and Z.Y.L. contributed equally to this work. Y.X., Y.H. and Z.Y.L. designed the study, performed the experiments and analyzed the data. W.J. and N.X.S. performed the experiments. Y.L. provided human samples and interpreted clinical data. G.C. advised on the interpretation of data. Z.Y. advised on the conception and design of the study. X.J.L. supervised the project, conceptualized and designed the study, performed the experiments, analyzed and interpreted the data, and wrote the manuscript. All the authors vouch for the respective data and analysis, reviewed and approved the final version and agreed to publish the manuscript.
Corresponding authors
Ethics declarations
Ethics approval and consent to participate
The human study was approved by the Institutional Ethics Committee at The First Affiliated Hospital of JNU. All procedures for animal experiments were performed according to the Animal Ethics Committee of Jinan University (No. 20211214–05).
Consent for publication
Not applicable.
Competing interests
The authors declare no competing interests.
Additional information
Publisher’s Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary Information
Rights and permissions
Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.
About this article
Cite this article
Xie, Y., Huang, Y., Li, ZY. et al. Interleukin-21 receptor signaling promotes metabolic dysfunction-associated steatohepatitis-driven hepatocellular carcinoma by inducing immunosuppressive IgA+ B cells. Mol Cancer 23, 95 (2024). https://doi.org/10.1186/s12943-024-02001-2
Received:
Accepted:
Published:
DOI: https://doi.org/10.1186/s12943-024-02001-2