Antiviral activity of bovine type III interferon against bovine viral diarrhea virus is greatly reduced in bovine turbinate cells due to limited expression of IFN lambda receptor 1 (IL-28Rα).

Introduction: The antiviral activity of recombinant bovine interferon lambda 3 (bovIFN-λ3) against bovine viral diarrhea virus (BVDV) has been demonstrated in vitro in Madin-Darby bovine kidney cells (MDBK) and in vivo in cattle. However, anti-BVDV activity of bovIFN-λ3 has not been studied in bovine respiratory tract epithelial cells, supposedly a primary target of BVDV infection when entering the host by the oronasal route. Methods: Here we investigated the anti-BVDV activity of bovIFN-λ3 in bovine turbinate-derived primary epithelial cells (BTu) using BVDV infection and immunoperoxidase staining, TCID50, RT-qPCR, DNA and transcriptome sequencing, and transfection with plasmids containing the two subunits, IL-28Rα and IL-10Rß that constitute the bovIFN-λ3 receptor. Results: Our immunoperoxidase staining, RT-qPCR, and TCID50 results show that while BVDV was successfully cleared in MDBK cells treated with bovIFN-λ3 and bovIFN-α, only the latter, bovIFN-α, cleared BVDV in BTu cells. Preincubation of MDBK cells with bovIFN-λ3 before BVDV infection was needed to induce optimal antiviral state. Both cell types displayed intact type I and III IFN signaling pathways and expressed similar levels of IL-10Rß subunit of the type III IFN receptor. Sequencing of PCR amplicon of the IL-28Rα subunit revealed intact transmembrane domain and lack of single nucleotide polymorphisms (SNPs) in BTu cells. However, RT-qPCR and transcriptomic analyses showed a lower expression of IL-28Rα transcripts in BTu cells as compared to MDBK cells. Interestingly, transfection of BTu cells with a plasmid encoding IL-28Rα subunit, but not IL-10Rß subunit, established the bovIFN-λ3 sensitivity showing similar anti-BVDV activity to the response in MDBK cells. Conclusion: Our results demonstrate that the sensitivity of cells to bovIFN-λ3 depends not only on the quality but also of the quantity of the IL-28Rα subunit of the heterodimeric receptor. A reduction in IL-28Rα transcript expression was detected in BTu as compared to MDBK cells, despite the absence of spliced variants or SNPs. The establishment of bovIFN-λ3 induced anti-BVDV activity in BTu cells transfected with an IL-28Rα plasmid suggests that the level of expression of this receptor subunit is crucial for the specific antiviral activity of type III IFN in these cells.

Interleukin-22 Promotes Cell Proliferation to Combat Virus Infection in Human Intestinal Epithelial Cells.

Interferon lambdas (IFN-λs) are crucial to control virus infections at mucosal surfaces. Interleukin-22 (IL-22) was reported to help IFN-λ control rotavirus infection in the intestinal epithelium of mice either by aiding in the induction of interferon-stimulated genes (ISGs) or by increasing cell proliferation thereby clearing virally infected cells. We investigated whether IL-22 and IFN-λs exhibit similar synergistic effects in human intestinal epithelial cells (IECs) models. Our results showed that co-treatment of IL-22 and IFN-λ induced more phosphorylation of STAT1 than either cytokine used alone. However, this increased STAT1 activation did not translate to increased ISGs production or antiviral protection. Transcriptomics analysis revealed that despite sharing a common subunit (IL-10Rb) within their heterodimeric receptors and activating similar STATs, the signaling generated by IL-22 and IFN-λs is independent, with IFN-λ signaling inducing ISGs and IL-22 signaling inducing cell proliferation genes. Using human intestinal organoids, we confirmed that IL-22 increased the size of the organoids through increased cell proliferation and expression of the stem cell marker (OLFM4). These findings suggest that in human intestinal cells, IFN-λs and IL-22 act independently to clear virus infections. IFN-λs induce ISGs to control virus replication and spread, whereas IL-22 increases cell proliferation to eliminate infected cells and repair the damage epithelium. Although these two cytokines do not act synergistically, each plays a key function in the protection of human IECs.

Interleukin 27, like interferons, activates JAK-STAT signaling and promotes pro-inflammatory and antiviral states that interfere with dengue and chikungunya viruses replication in human macrophages.

Interferons (IFNs) are a family of cytokines that activate the JAK-STAT signaling pathway to induce an antiviral state in cells. Interleukin 27 (IL-27) is a member of the IL-6 and/or IL-12 family that elicits both pro- and anti-inflammatory responses. Recent studies have reported that IL-27 also induces a robust antiviral response against diverse viruses, both in vitro and in vivo, suggesting that IFNs and IL-27 share many similarities at the functional level. However, it is still unknown how similar or different IFN- and IL-27-dependent signaling pathways are. To address this question, we conducted a comparative analysis of the transcriptomic profiles of human monocyte-derived macrophages (MDMs) exposed to IL-27 and those exposed to recombinant human IFN-α, IFN-γ, and IFN-λ. We utilized bioinformatics approaches to identify common differentially expressed genes between the different transcriptomes. To verify the accuracy of this approach, we used RT-qPCR, ELISA, flow cytometry, and microarrays data. We found that IFNs and IL-27 induce transcriptional changes in several genes, including those involved in JAK-STAT signaling, and induce shared pro-inflammatory and antiviral pathways in MDMs, leading to the common and unique expression of inflammatory factors and IFN-stimulated genes (ISGs)Importantly, the ability of IL-27 to induce those responses is independent of IFN induction and cellular lineage. Additionally, functional analysis demonstrated that like IFNs, IL-27-mediated response reduced chikungunya and dengue viruses replication in MDMs. In summary, IL-27 exhibits properties similar to those of all three types of human IFN, including the ability to stimulate a protective antiviral response. Given this similarity, we propose that IL-27 could be classified as a distinct type of IFN, possibly categorized as IFN-pi (IFN-π), the type V IFN (IFN-V).

Peritoneal B1 and B2 cells respond differently to LPS and IL-21 stimulation.

Peritoneal B cells can be divided into B1 cells (CD11b+CD19+) and B2 cells (CD11b-CD19+) based on CD11b expression. B1 cells play a crucial role in the innate immune response by producing natural antibodies and cytokines. B2 cells share similar traits with B1 cells, influenced by the peritoneal environment. However, the response of both B1 and B2 cells to the same stimuli in the peritoneum remains uncertain. We isolated peritoneal B1 and B2 cells from mice and assessed differences in Interleukin-10(IL-10) secretion, apoptosis, and surface molecule expression following exposure to LPS and Interleukin-21(IL-21). Our findings indicate that B1 cells are potent IL-10 producers, possessing surface molecules with an IgMhiCD43+CD21low profile, and exhibit a propensity for apoptosis in vitro. Conversely, B2 cells exhibit lower IL-10 production and surface markers characterized as IgMlowCD43-CD21hi, indicative of some resistance to apoptosis. LPS stimulates MAPK phosphorylation in B1 and B2 cells, causing IL-10 production. Furthermore, LPS inhibits peritoneal B2 cell apoptosis by enhancing Bcl-xL expression. Conversely, IL-21 has no impact on IL-10 production in these cells. Nevertheless, impeding STAT3 phosphorylation permits IL-21 to increase IL-10 production in peritoneal B cells. Moreover, IL-21 significantly raises apoptosis levels in these cells, a process independent of STAT3 phosphorylation and possibly linked to reduced Bcl-xL expression. This study elucidates the distinct functional and response profiles of B1 and B2 cells in the peritoneum to stimuli like LPS and IL-21, highlighting their differential roles in immunological responses and B cell diversity.

Interleukin-22 Alleviates Caerulein-Induced Acute Pancreatitis by Activating AKT/mTOR Pathway.

BACKGROUND: Acute pancreatitis (AP) is one of the most common acute abdominal disorders; due to the lack of specific treatment, the treatment of acute pancreatitis, especially serious acute pancreatitis (SAP), is difficult and challenging. We will observe the changes of Interleukin -22 levels in acute pancreatitis animal models, and explore the mechanism of Interleukin -22 in acute pancreatitis. OBJECTIVE: This study aims to assess the potential protective effect of Interleukin -22 on caerulein-induced acute pancreatitis and to explore its mechanism. METHODS: Blood levels of amylase and lipase and Interleukin -22 were assessed in mice with acute pancreatitis. In animal model and cell model of caerulein-induced acute pancreatitis, the mRNA levels of P62 and Beclin-1 were determined using PCR, and the protein expression of P62, LC3-II, mTOR, AKT, p-mTOR, and p-AKT were evaluated through Western blot analysis. RESULTS: Interleukin -22 administration reduced blood amylase and lipase levels and mitigated tissue damage in acute pancreatitis mice model. Interleukin -22 inhibited the relative mRNA levels of P62 and Beclin-1, and the Interleukin -22 group showed a decreased protein expression of LC3-II and P62 and the phosphorylation of the AKT/mTOR pathway. Furthermore, we obtained similar results in the cell model of acute pancreatitis. CONCLUSION: This study suggests that Interleukin -22 administration could alleviate pancreatic damage in caerulein-induced acute pancreatitis. This effect may result from the activation of the AKT/mTOR pathway, leading to the inhibition of autophagy. Consequently, Interleukin -22 shows potential as a treatment.

Interleukin-36gamma Mediates the In Vitro Activation of CD8+ T Cells from Patients Living with Chronic Human Immunodeficiency Virus-1 Infection.

Interleukin-36 (IL-36) signaling plays an important role in promoting CD8+ T cell-mediated antitumor immune responses. The role of IL-36 signaling in CD8+ T cells that are involved in host immune responses during human immunodeficiency virus-1 (HIV-1) infection has not been characterized. Sixty-one patients living with chronic HIV-1 infection and 23 controls were enrolled in this study. The levels of IL-36 cytokine family members were measured by enzyme-linked immunosorbent assay. Purified CD8+ T cells were stimulated with recombinant IL-36gamma (1 or 10 ng/mL). The expression of inhibitory receptors, the secretion of cytotoxic molecules and interferon-gamma, and the mRNA levels of apoptosis-related ligands were assessed to evaluate the effect of IL-36gamma on CD8+ T cell function in vitro. There were no significant differences in IL-36alpha, IL-36beta, or IL-36 receptor antagonist levels between patients living with chronic HIV-1 infection and controls. Plasma IL-36gamma levels were reduced in patients living with chronic HIV-1 infection. Perforin, granzyme B, and granulysin secretion, as well as tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and Fas ligand (FasL) mRNA expression, but not programmed death-1 (PD-1) or cytotoxic T lymphocyte-associated protein-4 (CTLA-4) expression was downregulated in CD8+ T cells from patients living with chronic HIV-1 infection. The addition of both 1 and 10 ng/mL IL-36gamma enhanced perforin, granzyme B, granulysin, and interferon-gamma secretion by CD8+ T cells without affecting PD-1/CTLA-4 or TRAIL/FasL mRNA expression in CD8+ T cells from patients living with chronic HIV-1 infection. The addition of 1 ng/mL IL-36gamma also promoted perforin and granzyme B secretion by HIV-1-specific CD8+ T cells from patients living with chronic HIV-1 infection. The reduced IL-36gamma levels in patients living with chronic HIV-1 infection might be insufficient for the activation of CD8+ T cells, leading to CD8+ T cell exhaustion.

Interleukin 21 Drives a Hypermetabolic State and CD4+ T-Cell-Associated Pathogenicity in Chronic Intestinal Inflammation.

BACKGROUND & AIMS: Incapacitated regulatory T cells (Tregs) contribute to immune-mediated diseases. Inflammatory Tregs are evident during human inflammatory bowel disease; however, mechanisms driving the development of these cells and their function are not well understood. Therefore, we investigated the role of cellular metabolism in Tregs relevant to gut homeostasis. METHODS: Using human Tregs, we performed mitochondrial ultrastructural studies via electron microscopy and confocal imaging, biochemical and protein analyses using proximity ligation assay, immunoblotting, mass cytometry and fluorescence-activated cell sorting, metabolomics, gene expression analysis, and real-time metabolic profiling utilizing the Seahorse XF analyzer. We used a Crohn's disease single-cell RNA sequencing dataset to infer the therapeutic relevance of targeting metabolic pathways in inflammatory Tregs. We examined the superior functionality of genetically modified Tregs in CD4+ T-cell-induced murine colitis models. RESULTS: Mitochondria-endoplasmic reticulum appositions, known to mediate pyruvate entry into mitochondria via voltage-dependent anion channel 1 (VDAC1), are abundant in Tregs. VDAC1 inhibition perturbed pyruvate metabolism, eliciting sensitization to other inflammatory signals reversible by membrane-permeable methyl pyruvate supplementation. Notably, interleukin (IL) 21 diminished mitochondria-endoplasmic reticulum appositions, resulting in enhanced enzymatic function of glycogen synthase kinase 3 ß, a putative negative regulator of VDAC1, and a hypermetabolic state that amplified Treg inflammatory response. Methyl pyruvate and glycogen synthase kinase 3 ß pharmacologic inhibitor (LY2090314) reversed IL21-induced metabolic rewiring and inflammatory state. Moreover, IL21-induced metabolic genes in Tregs in vitro were enriched in human Crohn's disease intestinal Tregs. Adoptively transferred Il21r-/- Tregs efficiently rescued murine colitis in contrast to wild-type Tregs. CONCLUSIONS: IL21 triggers metabolic dysfunction associated with Treg inflammatory response. Inhibiting IL21-induced metabolism in Tregs may mitigate CD4+ T-cell-driven chronic intestinal inflammation.

Whether full-length IL-38 acts as a promoter or inhibitor in activating rheumatoid arthritis fibroblast-like synoviocytes depends on IL-1ß.

AIM: IL-38 is a recently discovered inflammatory factor that belongs to the IL-1 family and has full-length and truncated forms. Clinical findings demonstrated that serum IL-38 levels in people with infectious and autoimmune diseases are significantly different from those in healthy people, but the form remains unclear. We are keenly interested in learning more about the regulatory role of full-length IL-38 in rheumatoid arthritis (RA), a classic autoimmune disease. METHODS: RA-fibroblast-like synoviocytes (RA-FLS) were isolated from six RA patients and stimulated with full-length IL-38 to observe IL-6 and IL-8 secretion. Then, the migration and invasion functions of FLS were assessed. Next, the protein expressions of the MAPK, NF-κB, and JAK pathways were evaluated. In addition, we examined the effect of full-length IL-38 on FLS functions in the presence of IL-1ß. The function of FLS affected by full-length IL-38 was also examined after blocking IL-1 and IL-36 receptors. RESULTS: The functions of FLS were activated after the cells were stimulated with full-length IL-38. IL-6 and IL-8 levels increased with an increase in the full-length IL-38 concentration, and full-length IL-38 induced the acceleration of FLS migration and invasion functions. In addition, the levels of proteins in the MAPK signaling pathway increased after stimulation with full-length IL-38 and depended on its concentration. However, when the FLS were stimulated by IL-38 and IL-1ß simultaneously, all experiments generated opposite results. Full-length IL-38 inhibited FLS function in the presence of IL-1ß. IL-1R and IL-36R blockers terminated all effects of full-length IL-38 on RA-FLS. CONCLUSION: Full-length IL-38 activates FLS functions and acts as a promoter in RA, whereas it inhibits FLS functions and acts as an inhibitor of RA in the presence of IL-1ß. The function of full-length IL-38 can be blocked by IL-1Ra and IL-36Ra.

Biological effects of IL-21 on immune cells and its potential for cancer treatment.

Interleukin-21 (IL-21), a member of the IL-2 cytokine family, is one of the most important effector and messenger molecules in the immune system. Produced by various immune cells, IL-21 has pleiotropic effects on innate and adaptive immune responses via regulation of natural killer, T, and B cells. An anti-tumor role of IL-21 has also been reported in the literature, as it may support cell proliferation or on the contrary induce growth arrest or apoptosis of the tumor cell. Anti-tumor effect of IL-21 enhances when combined with other agents that target tumor cells, immune regulatory circuits, or other immune-enhancing molecules. Therefore, understanding the biology of IL-21 in the tumor microenvironment (TME) and reducing its systemic toxic and side effects is crucial to ensure the maximum benefits of anti-tumor treatment strategies. In this review, we provide a comprehensive overview on the biological functions, roles in tumors, and the recent advances in preclinical and clinical research of IL-21 in tumor immunotherapy.

Interleukin-37 relieves PM2.5-triggered lung injury by inhibiting autophagy through the AKT/mTOR signaling pathway in vivo and in vitro.

Autophagy mediates PM2.5-related lung injury (LI) and is tightly linked to inflammation and apoptosis processes. IL-37 has been demonstrated to regulate autophagy. This research aimed to examine the involvement of IL-37 in the progression of PM2.5-related LI and assess whether autophagy serves as a mediator for its effects.To create a model of PM2.5-related LI, this research employed a nose-only PM2.5 exposure system and utilized both human IL-37 transgenic mice and wild-type mice. The hIL-37tg mice demonstrated remarkable reductions in pulmonary inflammation and pathological LI compared to the WT mice. Additionally, they exhibited activation of the AKT/mTOR signaling pathway, which served to regulate the levels of autophagy and apoptosis.Furthermore, in vitro experiments revealed a dose-dependent upregulation of autophagy and apoptotic proteins following exposure to PM2.5 DMSO extraction. Simultaneously, p-AKT and p-mTOR expression was found to decrease. However, pretreatment with IL-37 demonstrated a remarkable reduction in the levels of autophagy and apoptotic proteins, along with an elevation of p-AKT and p-mTOR. Interestingly, pretreatment with rapamycin, an autophagy inducer, weakened the therapeutic impact of IL-37. Conversely, the therapeutic impact of IL-37 was enhanced when treated with 3-MA, a potent autophagy inhibitor. Moreover, the inhibitory effect of IL-37 on autophagy was successfully reversed by administering AKT inhibitor MK2206. The findings suggest that IL-37 can inhibit both the inflammatory response and autophagy, leading to the alleviation of PM2.5-related LI. At the molecular level, IL-37 may exert its anti autophagy and anti apoptosis effects by activating the AKT/mTOR signaling pathway.

Interleukin-41 diminishes cigarette smoke-induced lung inflammation in mice.

BACKGROUND: Chronic obstructive pulmonary disease (COPD) and other inflammatory lung illnesses are markedly exacerbated by cigarette smoke (CS). The novel cytokine interleukin (IL)-41 has immunoregulatory effects, but data on its function in lung inflammation caused by CS are limited and inconclusive. Our study aimed to investigate the ability of IL-41 to protect against CS-induced lung inflammation in vivo. METHODS: In this model, mice were exposed to six cigarettes three times daily for 1 h, with 4-hour intervals between exposures, for 5 consecutive days. Mice received an intraperitoneal dose of IL-41 or a negative control 1 day prior to their initial exposure to CS. On day 6, mice were sacrificed to assess the impact of IL-41 on CS-induced lung inflammation. RESULTS: We found that IL-41 pre-treatment alleviated pulmonary inflammatory infiltration and lung tissue lesions. IL-41 pre-treatment also limited CS-induced weight loss, and resulted in lower numbers of macrophages in the bronchoalveolar lavage fluid and lower percentages of neutrophils and monocytes in the blood. Furthermore, it promoted the polarization of M2 macrophages rather than M1 macrophages, as determined by immunohistochemistry. Consistent with its effects on M2 polarization, pre-treatment with IL-41 was associated with higher levels of IL-10 in the bronchoalveolar lavage fluid and lung tissues of CS-exposed animals and lower production of tumor necrosis factor-α, IL-6 and IL-1ß in the serum and lung tissues. CONCLUSIONS: These findings suggest that IL-41 could be used therapeutically to treat CS-induced lung inflammatory disorders as it inhibits CS-induced pulmonary inflammation when administered in vivo in mice.

Synergism Between IL21 and Anti-PD-1 Combination Therapy is Underpinned by the Coordinated Reprogramming of the Immune Cellular Network in the Tumor Microenvironment.

T cell-stimulating cytokines and immune checkpoint inhibitors (ICI) are an ideal combination for increasing response rates of cancer immunotherapy. However, the results of clinical trials have not been satisfying. It is important to understand the mechanism of synergy between these two therapeutic modalities. Here, through integrated analysis of multiple single-cell RNA sequencing (scRNA-seq) datasets of human tumor-infiltrating immune cells, we demonstrate that IL21 is produced by tumor-associated T follicular helper cells and hyperactivated/exhausted CXCL13+CD4+ T cells in the human tumor microenvironment (TME). In the mouse model, the hyperactivated/exhausted CD4+ T cell-derived IL21 enhances the helper function of CD4+ T cells that boost CD8+ T cell-mediated immune responses during PD-1 blockade immunotherapy. In addition, we demonstrated that IL21's antitumor activity did not require T-cell trafficking. Using scRNA-seq analysis of the whole tumor-infiltrating immune cells, we demonstrated that IL21 treatment in combination with anti-PD-1 blockade synergistically drives tumor antigen-specific CD8+ T cells to undergo clonal expansion and differentiate toward the hyperactive/exhausted functional state in the TME. In addition, IL21 treatment and anti-PD-1 blockade synergistically promote dendritic cell (DC) activation and maturation to mature DC as well as monocyte to type 1 macrophage (M1) differentiation in the TME. Furthermore, the combined treatment reprograms the immune cellular network by reshaping cell-cell communication in the TME. Our study establishes unique mechanisms of synergy between IL21 and PD-1-based ICI in the TME through the coordinated promotion of type 1 immune responses. Significance: This study reveals how cytokine and checkpoint inhibitor therapy can be combined to increase the efficacy of cancer immunotherapy.

Anti-tumor Effect of Activated Canine B Cells With Interleukin-21 and Anti-B Cell Receptor.

BACKGROUND/AIM: Recently, novel studies on the pivotal role of B cells in the tumor-microenvironment and anti-tumor immunity have been conducted. Additionally, Interleukin-21 (IL-21) and anti-B cell receptor (BCR) have been reported to stimulate B cells to secrete granzyme B, which exhibits cytotoxic effects on tumor cells. However, the direct anti-tumor effect of B cells is not yet fully understood in the veterinary field. This study is the first attempt in veterinary medicine to identify the immediate effect of B cells on tumor suppression and the underlying mechanisms involved. MATERIALS AND METHODS: Canine B cells were isolated from peripheral blood and activated with IL-21 and anti-B cell receptor (BCR). The canine leukemia cell line GL-1 was co-cultured with B cells, and the anti-tumor effect was confirmed by assessing the changes in cell viability and apoptotic ratio. RESULTS: When B cells were activated with IL-21 and anti-BCR, the secretion of granzyme B and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) increased. Simultaneously, the viability of GL-1 cells decreased, and the apoptotic ratio increased, particularly when co-cultured with activated B cells. CONCLUSION: The results demonstrated the direct anti-tumor effect of granzyme B-and TRAIL and its enhanced potential of B cells to inhibit tumor cell growth after activation with IL-21 and anti-BCR. This study is the first study dealing with immunomodulation in the canine tumor micro-environment.

Evaluation of exosomes encapsulated recombinant Interleukin-29 for its in vitro anticancer studies.

Exosomes have recently been considered ideal biotherapeutic nanocarriers that broaden the frontiers of current drug delivery systems to overcome the shortcomings associated with cytokine-based immunotherapy. Using this approach, the current study aimed to assess anti-proliferative activity of purified IL-29 and exosomes encapsulated IL-29. The IL-29+pET-28a construct was transformed into Rosetta 2(DE3) cells which was used for the large-scale production of IL-29. Exosomes isolated from H1HeLa, and SF-767 cells using Total Exosome Isolation reagent were loaded with IL-29 via sonication. Isolation of exosomes was validated using their core protein signature by western blotting and specific miRNA profiles by RT-PCR. The drug loading efficiency of exosomes derived from H1HeLa cells was higher than that of SF-767-derived exosomes. The drug release kinetics of IL-29 encapsulated exosomes exhibited stable release of the recombinant drug. Around 50% of all cancer cell lines survived when IL-29 was administered at a concentration of 20 µg/mL. A survival rate of less than 10% was observed when cells were treated with 20 µg/mL IL-29 loaded exosomes. It was concluded that IL-29 loaded exosomes had a more significant cytotoxic effect against cancer cells, which might be attributed to sustained drug release, improved half-life, superior targeting efficacy, capacity to harness endogenous intracellular trafficking pathways, and heightened biocompatibility of exosomes.

IL-35 inhibits adipogenesis via PPARγ-Wnt/ß-catenin signaling pathway by targeting Axin2.

Interleukin (IL)-35, a member of the IL-12 family, functions as an immunosuppressive cytokine that plays a crucial role in the regulation of immune-related disorders and inflammatory diseases. Adipose tissue, which is now recognized as an immune organ, is regulated by immunocytes through various signaling pathways, including the peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT/enhancer-binding protein α (C/EBPα) pathway and the Wnt/ß-actin pathway. However, there is limited research regarding the effects of IL-35 on adipogenesis. Our current findings indicated that IL-35 impedes the proliferation and promotes the cytotoxicity of 3T3-L1 preadipocytes. Furthermore, IL-35 inhibited the adipogenic differentiation, as well as suppressed triglyceride and lipid accumulation. Additionally, the expression of PPARγ and C/EBPα, two key regulators of adipogenesis, were both down-regulated with IL-35 treatment. In order to explicate the mechanisms underlying the effects of IL-35, we conducted an investigation into the expression of Axin2, an intracellular inhibitor of Wnt/ß-catenin signaling, in 3T3-L1 preadipocyte cells. Gene silencing of Axin2 through small interfering RNAs (siRNAs) enhanced PPARγ and C/EBPα expression while decreasing nuclear ß-catenin levels in the presence of IL-35. Furthermore, in IL-35-treated cells, Axin2 knockdown boosted adipogenic differentiation (as measured by increased Oil Red O staining). These findings imply that IL-35 regulates Axin2 expression and thereby plays an important role in adipocyte development.

IFNL4 Genotypes and Risk of Childhood Burkitt Lymphoma in East Africa.

Interferon lambda 4 (IFN-λ4) is a novel type-III interferon that can be expressed only by carriers of the genetic variant rs368234815-dG within the first exon of the IFNL4 gene. Genetic inability to produce IFN-λ4 (in carriers of the rs368234815-TT/TT genotype) has been associated with improved clearance of hepatitis C virus (HCV) infection. The IFN-λ4-expressing rs368234815-dG allele (IFNL4-dG) is most common (up to 78%) in West sub-Saharan Africa (SSA), compared to 35% of Europeans and 5% of individuals from East Asia. The negative selection of IFNL4-dG outside Africa suggests that its retention in African populations could provide survival benefits, most likely in children. To explore this hypothesis, we conducted a comprehensive association analysis between IFNL4 genotypes and the risk of childhood Burkitt lymphoma (BL), a lethal infection-associated cancer most common in SSA. We used genetic, epidemiologic, and clinical data for 4,038 children from the Epidemiology of Burkitt Lymphoma in East African Children and Minors (EMBLEM) and the Malawi Infections and Childhood Cancer case-control studies. Generalized linear mixed models fit with the logit link controlling for age, sex, country, P. falciparum infection status, population stratification, and relatedness found no significant association between BL risk and 3 coding genetic variants within IFNL4 (rs368234815, rs117648444, and rs142981501) and their combinations. Because BL occurs in children 6-9 years of age who survived early childhood infections, our results suggest that additional studies should explore the associations of IFNL4-dG allele in younger children. This comprehensive study represents an important baseline in defining the health effects of IFN-λ4 in African populations.

Interleukin 26 Induces Macrophage IL-9 Expression in Rheumatoid Arthritis.

Rheumatoid arthritis (RA) is an autoimmune disease with chronic inflammation, bone erosion, and joint deformation. Synovial tissue in RA patients is full of proinflammatory cytokines and infiltrated immune cells, such as T help (Th) 9, Th17, macrophages, and osteoclasts. Recent reports emphasized a new member of the interleukin (IL)-10 family, IL-26, an inducer of IL-17A that is overexpressed in RA patients. Our previous works found that IL-26 inhibits osteoclastogenesis and conducts monocyte differentiation toward M1 macrophages. In this study, we aimed to clarify the effect of IL-26 on macrophages linking to Th9 and Th17 in IL-9 and IL-17 expression and downstream signal transduction. Murine and human macrophage cell lines and primary culture cells were used and stimulated by IL26. Cytokines expressions were evaluated by flow cytometry. Signal transduction and transcription factors expression were detected by Western blot and real time-PCR. Our results show that IL-26 and IL-9 colocalized in macrophage in RA synovium. IL-26 directly induces macrophage inflammatory cytokines IL-9 and IL-17A expression. IL-26 increases the IL-9 and IL-17A upstream mechanisms IRF4 and RelB expression. Moreover, the AKT-FoxO1 pathway is also activated by IL-26 in IL-9 and IL-17A expressing macrophage. Blockage of AKT phosphorylation enhances IL-26 stimulating IL-9-producing macrophage cells. In conclusion, our results support that IL-26 promotes IL-9- and IL-17-expressing macrophage and might initiate IL-9- and IL-17-related adaptive immunity in rheumatoid arthritis. Targeting IL-26 may a potential therapeutic strategy for rheumatoid arthritis or other IL-9 plus IL-17 dominant diseases.

Interleukin-37 ameliorates atherosclerosis by regulating autophagy-mediated endothelial cell apoptosis and inflammation.

Atherosclerosis is a lipid-driven chronic inflammatory disease. Endothelial dysfunction is the initiating factor of atherosclerosis. Although much work has been done on the antiatherosclerotic effects of interleukin-37 (IL-37), the exact mechanism is still not fully understood. The aim of this study was to investigate whether IL-37 attenuates atherosclerosis by protecting endothelial cells and to confirm whether autophagy plays a role in this effect. In apolipoprotein E knockout (ApoE-/-) mice fed with a high fat diet, IL-37 treatment significantly attenuated progression of atherosclerotic plaques, reduced endothelial cell apoptosis and inflammasome activation. Human umbilical vein endothelial cells (HUVECs) were treated with oxidized low-density lipoprotein (ox-LDL) to establish an endothelial dysfunction model. We observed that IL-37 alleviated ox-LDL-induced endothelial cell inflammation and dysfunction, as evidenced by decreased nod-like receptor pyrin domain-containing 3 (NLRP3) inflammasome activation, ROS production, apoptosis rate and secretion of inflammatory cytokines IL-1ß and TNF-α. Furthermore, IL-37 could activate autophagy in endothelial cells, which is characterized by the upregulation of LC3II/LC3I, the downregulation of p62 and an increase in autophagosomes. The autophagy inhibitor 3-Methyladenine (3-MA) dramatically reversed the promotion of autophagy and the protective effect of IL-37 against endothelial injury. Our data illustrate that IL-37 alleviated inflammation and apoptosis of atherosclerotic endothelial cells by enhancing autophagy. The current study provides new insights and promising therapeutic strategies for atherosclerosis.

Opposing Effects of Interleukin-36γ and Interleukin-38 on Trained Immunity.

Trained immunity is the process of long-term functional reprogramming (a de facto innate immune memory) of innate immune cells such as monocytes and macrophages after an exposure to pathogens, vaccines, or their ligands. The induction of trained immunity is mediated through epigenetic and metabolic mechanisms. Apart from exogenous stimuli, trained immunity can be induced by endogenous compounds such as oxidized LDL, urate, fumarate, but also cytokines including IL-1α and IL-1ß. Here, we show that also recombinant IL-36γ, a pro-inflammatory cytokine of the IL-1-family, is able to induce trained immunity in primary human monocytes, demonstrated by higher cytokine responses and an increase in cellular metabolic pathways both regulated by epigenetic histone modifications. These effects could be inhibited by the IL-36 receptor antagonist as well as by IL-38, an anti-inflammatory cytokine of the IL-1 family which shares its main receptor with IL-36 (IL-1R6). Further, we demonstrated that trained immunity induced by IL-36γ is mediated by NF-κB and mTOR signaling. The inhibitory effect of IL-38 on IL-36γ-induced trained immunity was confirmed in experiments using bone marrow of IL-38KO and WT mice. These results indicate that exposure to IL-36γ results in long-term pro-inflammatory changes in monocytes which can be inhibited by IL-38. Recombinant IL-38 could therefore potentially be used as a therapeutic intervention for diseases characterized by exacerbated trained immunity.

Relationship between IL-22 and IL-22BP in diabetic cognitive dysfunction.

BACKGROUND: CD4 + T helper (Th)22 cells play a regulatory role in autoimmune diseases such as type 1 diabetes mellitus. The Th22-related cytokine interleukin (IL)-22, the expression of which is increased in diabetes mellitus (DM), can act as a neurotrophic factor to protect neurons from apoptosis. Paradoxically, neuronal apoptosis and learning and memory decline occur in DM. In this study, we investigated the relationship between IL-22 and its receptors IL-22Rα1 and IL-22 binding protein (IL-22BP, a soluble inhibitor of IL-22) in diabetic encephalopathy (DE) and the effects of IL-22 on hippocampal neurons, learning and memory. METHODS: A C57BL/6 mouse model of diabetes was constructed by intraperitoneal injection of streptozotocin. The mice were randomly divided into 4 groups: the control group, diabetes group, diabetes + recombinantIL-22 (rIL-22) group and diabetes + IL-22BP group. The Morris water maze test was used to evaluate learning and memory, the expression of IL-22 was measured by ELISA, and Evans Blue staining was used to evaluate blood-brain barrier permeability. Quantitative reverse transcription polymerase chain reaction (RT-qPCR) was used to measure the mRNA expression of IL-22 and IL-22Rα1 in the hippocampus. The morphology and number of hippocampal neurons were assessed by Nissl staining, and TUNEL staining was used to detect hippocampal neuronal apoptosis. Immunofluorescence was used to analyze IL-22Rα1 expression and localization in hippocampus, and Western blotting was used to evaluate the expression of IL-22, IL-22Rα1, IL-22BP, and the apoptosis related proteins Caspase-3 and C-caspase-3. RESULTS: Compared with those in the control group, mice in the diabetes group showed cognitive decline; apoptosis of hippocampal neurons; increased expression of hippocampal Caspase-3, C-Caspase-3, IL-22, IL-22Rα1, and IL-22BP; and a decreased IL-22/IL-22BP ratio. Learning and memory were improved, neuronal apoptosis was attenuated, IL-22Rα1 expression and the IL-22/IL-22BP ratio were increased, and caspase-3 and C-caspase-3 expression was decreased in the rIL-22-treated group compared with the diabetes group. IL-22BP treatment aggravated diabetic cognitive dysfunction and pathological alterations in the hippocampus, decreased the IL-22/IL-22BP ratio, and increased the expression of caspase-3 and C-caspase-3 in mice with diabetes. CONCLUSION: A decrease in the IL-22/IL-22BP ratio plays an important role in diabetic cognitive dysfunction, and rIL-22 can effectively alleviate DE. Herein, we shed light on the interaction between IL-22 and IL-22BP as therapeutic targets for DM.