IL-13/STAT6 signaling plays a critical role in the epithelial-mesenchymal transition of colorectal cancer cells.
ABSTRACT: Colorectal cancer (CRC) is one of the most common causes of cancer-related death worldwide due to the distant metastases. Compelling evidence has reported that epithelial-mesenchymal transition (EMT) is involved in promoting cancer invasion and metastasis. However, the precise molecular events that initiate this complex EMT process remain poorly understood. Here, we showed that the pleiotropic cytokine interleukin-13 (IL-13) could induce an aggressive phenotype displaying EMT by enhancing the expression of EMT-promoting factor ZEB1. Importantly, STAT6 signaling inhibitor and STAT6 knockdown significantly reversed IL-13-induced EMT and ZEB1 induction in CRC cells, whereas ectopic STAT6 expression in STAT6null CRC cell line markedly promoted EMT in the present of IL-13. ChIP-PCR and Luciferase assays revealed that activated STAT6 directly bound to the promoter of ZEB1. Otherwise, we found IL-13 also up-regulated the stem cell markers (nanog, CD44, CD133 and CD166) and promoted cell migration and invasion through STAT6 pathway. We also found that siRNA-mediated knockdown of IL-13R?1 could reverse IL-13-induced ZEB1 and EMT changes by preventing STAT6 signaling. Finally, we demonstrated positive correlation between IL-13R?1 and ZEB1 at mRNA levels in human CRC samples. Taken together, our findings first demonstrated that IL-13/IL-13R?1/STAT6/ZEB1 pathway plays a critical role in promoting EMT and aggressiveness of CRC.
Project description:We herein investigated the role of the STAT signaling cascade in the production of pro-inflammatory cytokines and cisplatin ototoxicity. A significant hearing impairment caused by cisplatin injection was observed in Balb/c (wild type, WT) and STAT4(-/-), but not in STAT6(-/-) mice. Moreover, the expression levels of the protein and mRNA of pro-inflammatory cytokines, including TNF-?, IL-1?, and IL-6, were markedly increased in the serum and cochlea of WT and STAT4(-/-), but not STAT6(-/-) mice. Organotypic culture revealed that the shape of stereocilia bundles and arrays of sensory hair cell layers in the organ of Corti from STAT6(-/-) mice were intact after treatment with cisplatin, whereas those from WT and STAT4(-/-) mice were highly distorted and disarrayed after the treatment. Cisplatin induced the phosphorylation of STAT6 in HEI-OC1 auditory cells, and the knockdown of STAT6 by STAT6-specific siRNA significantly protected HEI-OC1 auditory cells from cisplatin-induced cell death and inhibited pro-inflammatory cytokine production. We further demonstrated that IL-4 and IL-13 induced by cisplatin modulated the phosphorylation of STAT6 by binding with IL-4 receptor alpha and IL-13R?1. These findings suggest that STAT6 signaling plays a pivotal role in cisplatin-mediated pro-inflammatory cytokine production and ototoxicity.
Project description:This study demonstrates that 24?h following viral vector-based vaccination IL-13R?2 functions as a master sensor on conventional dendritic cells (cDCs), abetted by high protein stability coupled with minimal mRNA expression, to rapidly regulate DC mediated IL-13 responses at the lung mucosae, unlike IL-13R?1. Under low IL-13, IL-13R?2 performs as a primary signalling receptor, whilst under high IL-13, acts to sequester IL-13 to maintain homeostasis, both in a STAT3-dependent manner. Likewise, we show that viral vector-derived IL-13 levels at the vaccination site can induce differential STAT3/STAT6 paradigms in lung cDC, that can get regulated collaboratively or independently by TGF-?1 and IFN-?. Specifically, low IL-13 responses associated with recombinant Fowlpox virus (rFPV) is regulated by early IL-13R?2, correlated with STAT3/TGF-?1 expression. Whilst, high IL-13 responses, associated with recombinant Modified Vaccinia Ankara (rMVA) is regulated in an IL-13R?1/STAT6 dependent manner associated with IFN-?R expression bias. Different viral vaccine vectors have previously been shown to induce unique adaptive immune outcomes. Taken together current observations suggest that IL-13R?2-driven STAT3/STAT6 equilibrium at the cDC level may play an important role in governing the efficacy of vector-based vaccines. These new insights have high potential to be exploited to improve recombinant viral vector-based vaccine design, according to the pathogen of interest and/or therapies against IL-13 associated disease conditions.
Project description:Interleukin (IL)-32?, a newly identified IL-32 isoform, has been reported to exert pro-inflammatory effects through the association with protein kinase C delta (PKC?). In this study, we further examined the effects of IL-32? on IL-13 and IL-13R?2 expression and the related mechanism in THP-1 cells. Upon stimulating IL-32?-expressing and non-expressing cells with phorbol 12-myristate 13-acetate (PMA), the previous microarray analysis showed that IL-13R?2 and IL-13 mRNA expression were significantly decreased by IL-32?. The protein expression of these factors was also confirmed to be down-regulated. The nuclear translocation of transcription factors STAT3 and STAT6, which are necessary for IL-13R?2 and IL-13 promoter activities, was suppressed by IL-32?. Additionally, a direct association was found between IL-32?, PKC?, and signal transducer and activator of transcription 3 (STAT3), but not STAT6, revealing that IL-32? might act mainly through STAT3 and indirectly affect STAT6. Moreover, the interaction of IL-32? with STAT3 requires PKC?, since blocking PKC? activity eliminated the interaction and consequently limited the inhibitory effect of IL-32? on STAT3 activity. Interfering with STAT3 or STAT6 binding by decoy oligodeoxynucleotides (ODNs) identified that IL-32? had additive effects with the STAT3 decoy ODN to suppress IL-13 and IL-13R?2 mRNA expression. Taken together, our data demonstrate the intracellular interaction of IL-32?, PKC?, and STAT3 to regulate IL-13 and IL-13R?2 synthesis, supporting the role of IL-32? as an inflammatory modulator.
Project description:There continues to be a major need for more effective inflammatory bowel disease (IBD) therapies. IL-13R?2 is a decoy receptor that binds the cytokine IL-13 with high affinity and diminishes its STAT6-mediated effector functions. Previously, we found that IL-13R?2 was necessary for IBD in mice deficient in the anti-inflammatory cytokine IL-10. Here, we tested for the first time a therapeutic antibody specifically targeting IL-13R?2. We also used the antibody and Il13ra2-/- mice to dissect the role of IL-13R?2 in IBD pathogenesis and recovery. Il13ra2-/- mice were modestly protected from induction of dextran sodium sulfate (DSS)-induced colitis. Following a 7-day recovery period, Il13ra2-/- mice or wild-type mice administered the IL-13R?2-neutralizing antibody had significantly improved colon health compared to control mice. Neutralizing IL-13R?2 to increase IL-13 bioavailability promoted resolution of IBD even if neutralization occurred only during recovery. To link our observations in mice to a large human cohort, we conducted a phenome-wide association study of a more active variant of IL-13 (R130Q) that has reduced affinity for IL-13R?2. Human subjects carrying R130Q reported a lower risk for Crohn's disease. Our findings endorse moving anti-IL-13R?2 into preclinical drug development with the goal of accelerating recovery and maintaining remission in Crohn's disease patients.
Project description:Macrophages coexpress both the interleukin (IL)-2R? chain (?(c)) and IL-13R?1. These receptor chains can heterodimerize with IL-4R? to form type I or type II IL-4 receptor complexes, respectively. We used macrophages derived from Il2rg and Il13ra1 knockout (KO) mice to evaluate the requirements for these receptor chains for induction of the alternative macrophage activation (AMA) pathway by IL-4 and IL-13. Absence of ?(c) significantly decreased activation of STAT6 by IL-4 but not IL-13. However, although activation of STAT6 by IL-4 was markedly reduced in ?(c) KO macrophages, it was not abolished, indicating that IL-4 can still signal through type II IL-4 receptors via the IL-13R?1 chain. IL-13 failed to activate STAT6 in macrophages derived from Il13ra1 KO mice; however, these cells remained fully responsive to IL-4. The inability of IL-13 but not IL-4 to signal in Il13ra1(-/-) macrophages correlated with the inability of IL-13 but not IL-4 to induce expression of genes such as Arg1, Retnla and Ccl11 that are characteristically expressed by alternatively activated macrophages. In addition, IL-13 but not IL-4 failed to induce membrane fusion and giant cell formation by Il13ra1 KO macrophages. These findings demonstrate that the IL-13R?1 chain is essential for induction of the AMA pathway by IL-13 but not IL-4.
Project description:CCAAT/enhancer-binding homologous protein (CHOP), a transcriptional regulator induced by endoplasmic reticulum stress (ER stress) is a pivotal factor in the ER stress-mediated apoptosis pathway. Previous studies have shown that CHOP is involved in the formation of fibrosis in a variety of tissues and is associated with alternative macrophage activation. The role of CHOP in the pathologic effects of liver fibrosis in schistosomiasis has not been reported, and underlying mechanisms remain unclear. This study is aimed at understanding the effect of CHOP on liver fibrosis induced by Schistosoma japonicum (S. japonicum) in vivo and clarifying its mechanism. C57BL/6 mice were infected with cercariae of S. japonicum through the abdominal skin. The liver fibrosis was examined. The level of IL-13 was observed. The expressions of CHOP, Krüppel-like factor 4 (KLF4), signal transducer and activator of transcription 6 (STAT6), phosphorylation STAT6, interleukin-13 receptor alpha 1 (IL-13R?1), and interleukin-4 receptor alpha (IL-4R?) were analysed. The eosinophilic granuloma and collagen deposition were found around the eggs in mice infected for 6 and 10 weeks. IL-13 in plasma and IL-13R?1 and IL-4R? in liver tissue were significantly increased. The phosphorylated STAT6 was enhanced while Krüppel-like factor 4 (KLF4) was decreased in liver tissue. The expression of CHOP and colocalization of CHOP and CD206 were increased. Overall, these results suggest that CHOP plays a critical role in hepatic fibrosis induced by S. japonicum, likely through promoting alternative activation of macrophages.
Project description:The IL-13R?1 signaling pathway and M2 macrophages play crucial roles in schistosome egg-induced hepatic fibrosis via the expression of pro-fibrotic molecules. This study aims to investigate the inhibitory effect and mechanism of action of corilagin on schistosome egg-induced hepatic fibrosis via the IL-13R?1 signaling pathway in M2 macrophages in vitro and in vivo. The mRNA and protein expression of IL-13R?1, PPAR?, KLF4, SOCS1, STAT6, p-STAT6, and TGF-? was measured in vitro with corilagin treatment after IL-13 stimulation and in vivo corilagin treatment after effectively killing the adult schistosomes in schistosome-infected mice. Histological analysis of liver tissue was assessed for the degree of hepatic fibrosis. The results revealed that corilagin significantly reduced the expression of PPAR?, KLF4, SOCS1, p-STAT6, and TGF-? compared with model group and praziquantel administration (p < 0.01 or p < 0.05) in vivo and in vitro, which indicated a strong inhibitory effect of corilagin on IL-13R?1 signaling pathway. As well, the inhibitory effect of corilagin showed a significant dose-dependence (p < 0.05). The area of fibrosis and distribution of M2 macrophages in mouse liver tissue were reduced significantly and dose-dependently with corilagin treatment compared to model group or praziquantel administration (p < 0.01 or p < 0.05), indicating that corilagin suppressed IL-13R?1 signaling pathway and M2 macrophage polarization effectively in vivo. Furthermore, the anti-fibrogenic effect persisted even when IL-13R?1 was up- or down-regulated in vitro. In conclusion, corilagin can suppress schistosome egg-induced hepatic fibrosis via inhibition of M2 macrophage polarization in the IL-13R?1 signaling pathway.
Project description:Intestinal tumorigenesis in the ApcMin/+ model is initiated by aberrant activation of Wnt pathway. Increased IL-4 expression in human colorectal cancer tissue and growth of colon cancer cell lines implied that IL-4-induced Stat6-mediated tumorigenic signaling likely contributes to intestinal tumor progression in ApcMin/+ mice. Stat6 also appears to promote expansion of myeloid-derived suppressor cells (MDSCs) cells. MDSCs promote polyp formation in the ApcMin/+ model. Hence, Stat6 could have a broad role in coordinating both polyp cell proliferation and MDSC expansion. We found that IL-4-induced Stat6-mediated proliferation of intestinal epithelial cells is augmented by platelet-derived growth factor-BB, a tumor-promoting growth factor. To determine whether polyp progression in ApcMin/+ mice is dependent on Stat6 signaling, we disrupted Stat6 in this model. Total polyps in the small intestine were fewer in ApcMin/+ mice lacking Stat6. Furthermore, proliferation of polyp epithelial cells was reduced, indicating that Stat6 in part controlled polyp formation. Stat6 also promoted expansion of MDSCs in the spleen and lamina propria of ApcMin/+ mice, implying regulation of antitumor T-cell response. More CD8 cells and reduced PD-1 expression on CD4 cells correlated with reduced polyps. In addition, a strong CD8-mediated cytotoxic response led to killing of tumor cells in Stat6-deficient ApcMin/+ mice. Therefore, these findings show that Stat6 has an oncogenic role in intestinal tumorigenesis by promoting polyp cell proliferation and immunosuppressive mediators, and preventing an active cytotoxic process.
Project description:In this study, we examined the role IL-13 receptor alpha 1 (IL-13R?1) plays in macrophage differentiation and function. The findings indicate that IL-13R?1 is expressed on the M2 but not on the M1 subset of macrophages and specifically heterodimerizes with the IL-4R? chain to form a type II receptor, which controls the differentiation and function of these cells. Indeed, BM cells from IL-13R?1(+/+) and IL-13R?1(-/-) mice yield equivalent numbers of macrophages when cultured under M2 polarizing conditions. However, IL-13R?1(-/-) BM cells yield a much higher number of macrophages than IL-13R?1(+/+) BM cells when the differentiation is carried out under M1-polarizing conditions. Further analyses indicated that macrophages that express IL-13R?1 also display surface markers associated with an M2 phenotype. In addition, the IL-13R?1(+) macrophages were highly efficient in phagocytizing zymosan bioparticles both in vitro and in vivo, and supported differentiation of naïve T cells to a Th2 phenotype. Finally, when stimulated by IL-13, a cytokine that uses the heteroreceptor, the cells were able to phosphorylate STAT6 efficiently. These previously unrecognized findings indicate that IL-13R?1 serves as a marker for M2 macrophages and the resulting heteroreceptor influences both their differentiation and function.
Project description:Lysosomes play important roles in multiple aspects of physiology, but the problem of how the transcription of lysosomal genes is coordinated remains incompletely understood. The goal of this study was to illuminate the physiological contexts in which lysosomal genes are coordinately regulated and to identify transcription factors involved in this control.As transcription factors and their target genes are often co-regulated, we performed meta-analyses of array-based expression data to identify regulators whose mRNA profiles are highly correlated with those of a core set of lysosomal genes. Among the ~50 transcription factors that rank highest by this measure, 65% are involved in differentiation or development, and 22% have been implicated in interferon signaling. The most strongly correlated candidate was Stat6, a factor commonly activated by interleukin-4 (IL-4) or IL-13. Publicly available chromatin immunoprecipitation (ChIP) data from alternatively activated mouse macrophages show that lysosomal genes are overrepresented among Stat6-bound targets. Quantification of RNA from wild-type and Stat6-deficient cells indicates that Stat6 promotes the expression of over 100 lysosomal genes, including hydrolases, subunits of the vacuolar H? ATPase and trafficking factors. While IL-4 inhibits and activates different sets of lysosomal genes, Stat6 mediates only the activating effects of IL-4, by promoting increased expression and by neutralizing undefined inhibitory signals induced by IL-4.The current data establish Stat6 as a broadly acting regulator of lysosomal gene expression in mouse macrophages. Other regulators whose expression correlates with lysosomal genes suggest that lysosome function is frequently re-programmed during differentiation, development and interferon signaling.