Tumor-derived microparticles promote the progression of triple-negative breast cancer via PD-L1-associated immune suppression.

Membrane vesicles, including exosomes and microparticles (MPs), serve to package and transfer the cellular cargo during inter/extracellular communication, which is of great interest in cancer development, especially in the dissemination of signal transduction-associated traits from donor cells to recipient cells. Although increasing evidence suggests that microparticles (MPs) contribute to the development of cancer, their unique characteristics remain to be exploited. Here, we examined the secretion of MPs in tumor tissues from triple-negative breast cancer (TNBC) patients and found that the tumor cells could release MPs loaded with immune checkpoint molecular programmed cell death ligand 1 (PD-L1), especially in patients treated with traditional clinical interventions, such as chemotherapy and radiotherapy. These PD-L1-loading MPs contribute to the suppressive immune microenvironment, eventually resulting in the tumor progression in TNBC. Mechanically, we proved that PD-L1-loading MPs could suppress the activation and function of functional cluster of differentiation CD8+ T cells. Meanwhile, the PD-L1-loading MPs could mediate the differentiation of macrophages toward the immune-suppressive M2 phenotype via the activation of the TANK-binding kinase 1 (TBK1)/signal transducer and activator of transcription 6 (STAT6) signal and suppression of the serine-threonine kinase (AKT)/mammalian target of rapamycin (mTOR) signal. Given the increasing MP production induced by traditional clinical interventions, we further combined chemotherapy with the PD-L1 inhibitor atezolizumab (ATZ) to efficiently abrogate the immunosuppression caused by the PD-L1-loading MPs. Therefore, our study unveils the mechanism by which tumor cells systemically evade immune surveillance by releasing the PD-L1-loading MPs, and provides new insights into clinical TNBC immunotherapy.

Rab1A promotes IL-4R/JAK1/STAT6-dependent metastasis and determines JAK1 inhibitor sensitivity in non-small cell lung cancer.

Rab1A overexpression has been observed in several cancer types, however, its significance and the underlying mechanisms in non-small cell lung cancer (NSCLC) remain largely unexplored. This study demonstrated that Rab1A overexpression in NSCLC was significantly correlated to short survival and metastasis. Rab1A overexpression promoted cancer cell migration, invasion, and metastasis both in vitro and in vivo, by activating JAK1/STAT6 signaling through stabilizing IL-4Rα protein. Strikingly, high Rab1A level was associated with sensitivity to JAK1 inhibitor, and Rab1A overexpression rendered cancer cells vulnerable to JAK1-targeted agents. JAK1 inhibitor, Itacitinib adipate, dramatically inhibited high Rab1A NSCLC metastasis, in both cell line and patient derived xenograft models. Collectively, these findings demonstrated that Rab1A plays a critical role in the aggressive properties of NSCLC, revealing a unique mechanism by which it promotes metastasis. In addition, we found that Rab1A is a determinant of JAK1 inhibitor sensitivity, which could be explored for improving JAK1-targeted cancer therapy.

T-bet and STAT6 Coordinately Suppress the Development of IL-9-Mediated Atopic Dermatitis-Like Skin Inflammation in Mice.

T-bet and signal transducer and activator of transcription (STAT) 6 are critical factors for helper T-cell differentiation in humans and mice. Additionally, polymorphisms in TBX21 (T-bet) and STAT6 are associated with the susceptibility of allergic diseases. However, precise mechanisms of the reciprocal regulation between T-bet and STAT6 in allergy remain unclear. To determine the reciprocal regulation in vivo, we investigated the phenotype of T-bet/STAT6 double-deficient (T-bet-/- STAT6-/-) mice. Unexpectedly, T-bet-/- STAT6-/- mice but not T-bet-/- mice or STAT6-/- mice spontaneously developed severe dermatitis. Not only eosinophils and mast cells but also CD4+ T cells infiltrated into the skin of T-bet-/- STAT6-/- mice. Adoptive transfer of CD4+ T cells of T-bet-/- STAT6-/- mice into severe combined immunodeficient mice induced the accumulation of eosinophils and mast cells in the skin, whereas depletion of CD4+ T cells ameliorated the dermatitis in T-bet-/- STAT6-/- mice. Comprehensive transcriptome analyses revealed that IL-9 expression was enhanced in T-bet-/- STAT6-/- CD4+ T cells. Indeed, IL-9 neutralization ameliorated the dermatitis in T-bet-/- STAT6-/- mice. T-bet-/- STAT6-/- CD4+ T cells expressed functional thymic stromal lymphopoietin receptors and produced large amounts of IL-9 on thymic stromal lymphopoietin stimulation. These results indicate that T-bet and STAT6 coordinately suppress atopic dermatitis-like skin inflammation, possibly by inhibiting thymic stromal lymphopoietin-dependent IL-9 production in CD4+ T cells.

Granzyme A-producing T helper cells are critical for acute graft-versus-host disease.

Acute graft-versus-host disease (aGVHD) can occur after hematopoietic cell transplant in patients undergoing treatment for hematological malignancies or inborn errors. Although CD4+ T helper (Th) cells play a major role in aGVHD, the mechanisms by which they contribute, particularly within the intestines, have remained elusive. We have identified a potentially novel subset of Th cells that accumulated in the intestines and produced the serine protease granzyme A (GrA). GrA+ Th cells were distinct from other Th lineages and exhibited a noncytolytic phenotype. In vitro, GrA+ Th cells differentiated in the presence of IL-4, IL-6, and IL-21 and were transcriptionally unique from cells cultured with either IL-4 or the IL-6/IL-21 combination alone. In vivo, both STAT3 and STAT6 were required for GrA+ Th cell differentiation and played roles in maintenance of the lineage identity. Importantly, GrA+ Th cells promoted aGVHD-associated morbidity and mortality and contributed to crypt destruction within intestines but were not required for the beneficial graft-versus-leukemia effect. Our data indicate that GrA+ Th cells represent a distinct Th subset and are critical mediators of aGVHD.

STAT6 Pathway Is Critical for the Induction and Function of Regulatory T Cells Induced by Mucosal B Cells.

B cells could convert naïve T cells into regulatory T cells (so-called Treg-of-B cells) which have the ability to treat animal models of inflammatory diseases, including allergic asthma, collagen-induced arthritis and colitis; however, the mechanisms of Treg-of-B cell generation remain unclear. In this study, we investigated the role of STAT6 in the generation of Treg-of-B (P) cells, which Treg cells were generated by Peyer's patch B cells (P stands for Peyer's patch). CD4+CD25- T cells from wild type, STAT6 knockout and IL-4 knockout mice were cocultured with wild type Peyer's patch B cells for Treg-of-B (P) cell generation. A murine asthmatic model was used to analyze the in vivo regulatory function of Treg-of-B (P) cells. The data demonstrated that STAT6 played a critical role in the generation of Treg-of-B (P) cells, which confirmed with STAT6-deficient T cells and the STAT6 inhibitor AS1517499. When STAT6 was lacking, Treg-of-B (P) cells exerted impaired suppressive ability with decreased LAG3 expression. Furthermore, Peyer's patch B cells played an essential role in regulatory T cell generation. In the absence of Peyer's patch B cells, T cells expressed decreased phosphorylated STAT6, which was followed by decreased LAG3 expression and impaired suppressive ability, suggesting that Peyer's patch B cells provided the critical signal to activate STAT6 phosphorylation in T cells. Moreover, STAT6 deficient Treg-of-B (P) cells could not alleviate inflammation in an animal model of asthma in vivo. IL-4 was downstream of phosphorylated STAT6 and maintained Treg-of-B (P) cell survival with increased expression of Bcl-2 and BclXL. We reported a novel finding that the STAT6-LAG3 signaling axis is important for the induction and function of Treg-of-B (P) cells.

Activation of the IL-4/STAT6 Signaling Pathway Promotes Lung Cancer Progression by Increasing M2 Myeloid Cells.

Emerging evidence shows that signal transducer and activator of transcription 6 (STAT6) plays critical roles in tumor development. We previously found high-level expression of STAT6 in human lung adenocarcinoma and squamous cell carcinoma, specifically in infiltrated immune cells located in the lung interstitium. Nevertheless, the role of STAT6 signaling in lung carcinogenesis and lung cancer proliferation and its underlying mechanisms remain unclear. This study aimed to investigate the role of STAT6 and the interaction between STAT6 and the tumor microenvironment in pulmonary tumorigenesis. We established a murine model of primary lung carcinogenesis in STAT6-deficient (STAT6-/-) and STAT6 wild-type (WT) BALB/c mice using the carcinogen urethane. Two-month-old male mice were intraperitoneally injected with urethane (1 g/kg) dissolved in phosphate buffered saline (PBS). Primary tumors were monitored in vivo by positron emission tomography scanning. At 4, 6, and 9 months after urethane injection, lung tumors were harvested from the STAT6-/- and WT mice for analysis. Small interfering RNA was used to downregulate the expression of STAT6 in tumor cells. Fluorescence activated cell sorting analysis was used to analyze fluorescence-conjugated cell markers. Transwell assays were used in coculturing experiments. STAT6 protein expression was detected by Western blotting, immunohistochemistry, and immunofluorescence. STAT6 mRNA expression was detected by quantitative real time-polymerase chain reaction. Cell Counting Kit-8 and colony formation assays were performed to evaluate cell proliferation. We detected high expression of STAT6 in CD11b+ cells of lung carcinoma. Our results indicate that STAT6 deficiency inhibits carcinogen-induced tumor growth and improves prognosis. STAT6 deficiency also decreased the mobilization and differentiation of CD11b+ cells. STAT6 deficiency in CD11b+ cells but not tumor cells decreased interleukin (IL)-4 secretion and the differentiation of CD11b+ cells into M2 macrophage cells. In conclusion, our findings indicate that IL-4/STAT6 signaling in CD11b+ cells promotes lung cancer progression by triggering an IL-4 positive feedback loop and increasing M2 myeloid cells. STAT6 may be a new therapeutic target for the prevention and treatment of lung cancer.

IL-4 and IL-17A Cooperatively Promote Hydrogen Peroxide Production, Oxidative DNA Damage, and Upregulation of Dual Oxidase 2 in Human Colon and Pancreatic Cancer Cells.

Dual oxidase 2 (DUOX2) generates H2O2 that plays a critical role in both host defense and chronic inflammation. Previously, we demonstrated that the proinflammatory mediators IFN-γ and LPS enhance expression of DUOX2 and its maturation factor DUOXA2 through STAT1- and NF-κB‒mediated signaling in human pancreatic cancer cells. Using a panel of colon and pancreatic cancer cell lines, we now report the induction of DUOX2/DUOXA2 mRNA and protein expression by the TH2 cytokine IL-4. IL-4 activated STAT6 signaling that, when silenced, significantly decreased induction of DUOX2. Furthermore, the TH17 cytokine IL-17A combined synergistically with IL-4 to increase DUOX2 expression in both colon and pancreatic cancer cells mediated, at least in part, by signaling through NF-κB. The upregulation of DUOX2 was associated with a significant increase in the production of extracellular H2O2 and DNA damage-as indicated by the accumulation of 8-oxo-dG and γH2AX-which was suppressed by the NADPH oxidase inhibitor diphenylene iodonium and a DUOX2-specific small interfering RNA. The clinical relevance of these experiments is suggested by immunohistochemical, microarray, and quantitative RT-PCR studies of human colon and pancreatic tumors demonstrating significantly higher DUOX2, IL-4R, and IL-17RA expression in tumors than in adjacent normal tissues; in pancreatic adenocarcinoma, increased DUOX2 expression is adversely associated with overall patient survival. These data suggest a functional association between DUOX2-mediated H2O2 production and induced DNA damage in gastrointestinal malignancies.

ER-stress regulates macrophage polarization through pancreatic EIF-2alpha kinase.

During the process of NAFLD progression, ER-stress is activated in macrophages and induces the pro-inflammatory polarization of macrophage. As one of the three ER membrane resident proteins, pancreatic eIF-2alpha kinase (PERK) plays an important role in ER stress, but its participation in macrophage polarization is largely unknown. In this study, we found that the PA mediated ER-stress activation could induce M1-type polarization in macrophages, and this phenotype polarization could be inhibited by ER-stress inhibitor 4-PBA as well as GSK2656157, an inhibitor of PERK. Moreover, the knockdown of PERK altered the STAT1 and STAT6 pathways in macrophages, which then led to the M1-to-M2 phenotypic shift. In summary, we found that PERK could regulate the phenotypic polarization of macrophages. This finding may provide new insight into the suppression of pathological progression of fatty liver or liver ischemia reperfusion injury induced by M1-type macrophages.

Hierarchical control of interleukin 13 (IL-13) signals in lung fibroblasts by STAT6 and SOX11.

Interleukin (IL)-13 is a signature cytokine of type 2 inflammation important for the pathogenesis of various diseases, including allergic diseases. Signal transducer and activator of transcription (STAT) 6 is a critical transcriptional factor for the IL-13 signals; however, it remains unknown how expression of the IL-13-induced genes is differentiated by the transcriptional machineries. In this study, we identified IL-13-induced transcriptional factors in lung fibroblasts using DNA microarrays in which SOX11 was included. Knockdown of SOX11 down-regulated expression of periostin and CCL26, both of which are known to be downstream molecules of IL-13, whereas enforced expression of SOX11 together with IL-13 stimulation enhanced expression of periostin. Moreover, we found that in DNA microarrays combining IL-13 induction and SOX11 knockdown there exist both SOX11-dependent and -independent molecules in IL-13-inducible molecules. In the former, many inflammation-related and fibrosis-related molecules, including periostin and CCL26, are involved. These results suggest that SOX11 acts as a trans-acting transcriptional factor downstream of STAT6 and that in lung fibroblasts the IL-13 signals are hierarchically controlled by STAT6 and SOX11.

Synergistic action of dual IGF1/R and MEK inhibition sensitizes childhood acute lymphoblastic leukemia (ALL) cells to cytotoxic agents and involves downregulation of STAT6 and PDAP1.

Heterogeneous upregulation of multiple prosurvival pathways underlies resistance to damage-induced apoptosis in acute lymphoblastic leukemia (ALL) cells despite normal p53 responses. Here, we show that the dual combination of insulin-like growth factor 1 (IGF1)/IGF1 receptor (IGF1/R) and mitogen-activated protein kinase/extracellular signal-regulated kinase (ERK) kinase (MEK) inhibition using AG1024 + U0126 can sensitize apoptosis-resistant ALL cells to ionizing radiation-induced DNA damage irrespective of effect of single pathway inhibition in vitro. This AG1024 + U0126 combination also significantly potentiates the ability of the core chemotherapy compounds vincristine, dexamethasone, and daunorubicin to kill ALL cells in vitro. Evidence of the synergistic action of AG1024 + U0126 in samples with variable basal levels of phosphorylated IGF1/Rß and ERK1/2 suggested additional targets of this drug combination. Consistent with this, gene expression profiling identified 32 "synergy genes" differentially targeted by IGF1/R + MEK inhibition and, among these, Signal transducer and activator of transcription 6 (STAT6) and platelet-derived growth factor-associated protein 1 (PDAP1) were the most differentially downregulated cluster. Pearson correlation analysesrevealed that STAT6 and PDAP1 display significant expression codependency and a common expression pattern linked with other key "synergy" genes, supporting their predicted role in an STAT6-ERK-nuclear factor kappa beta (NF-κB) network. Knockdown studies revealed that loss of STAT6, but not PDAP1, impinges on the cell cycle, causing reduced numbers of viable cells. In combination with daunorubicin, STAT6 loss has an additive effect on cell killing, whereas PDAP1 loss is synergistic, indicating an important role of PDAP1 in the cellular response to this anthracycline. Inhibition of STAT6 or PDAP1 may therefore represent a potential novel therapeutic strategy for resistant ALL by enhancing sensitivity to chemotherapy.

Antagonizing Integrin ß3 Increases Immunosuppression in Cancer.

Integrin ß3 is critical for tumor invasion, neoangiogenesis, and inflammation, making it a promising cancer target. However, preclinical and clinical data of integrin ß3 antagonists have demonstrated no benefit or worse outcomes. We hypothesized that integrin ß3 could affect tumor immunity and evaluated tumors in mice with deletion of integrin ß3 in macrophage lineage cells (ß3KOM). ß3KOM mice had increased melanoma and breast cancer growth with increased tumor-promoting M2 macrophages and decreased CD8(+) T cells. Integrin ß3 antagonist, cilengitide, also enhanced tumor growth and increased M2 function. We uncovered a negative feedback loop in M2 myeloid cells, wherein integrin ß3 signaling favored STAT1 activation, an M1-polarizing signal, and suppressed M2-polarizing STAT6 activation. Finally, disruption of CD8(+) T cells, macrophages, or macrophage integrin ß3 signaling blocked the tumor-promoting effects of integrin ß3 antagonism. These results suggest that effects of integrin ß3 therapies on immune cells should be considered to improve outcomes. Cancer Res; 76(12); 3484-95. ©2016 AACR.

Novel Roles for Chloride Channels, Exchangers, and Regulators in Chronic Inflammatory Airway Diseases.

Chloride transport proteins play critical roles in inflammatory airway diseases, contributing to the detrimental aspects of mucus overproduction, mucus secretion, and airway constriction. However, they also play crucial roles in contributing to the innate immune properties of mucus and mucociliary clearance. In this review, we focus on the emerging novel roles for a chloride channel regulator (CLCA1), a calcium-activated chloride channel (TMEM16A), and two chloride exchangers (SLC26A4/pendrin and SLC26A9) in chronic inflammatory airway diseases.

In Situ Characterization of Splenic Brucella melitensis Reservoir Cells during the Chronic Phase of Infection in Susceptible Mice.

Brucella are facultative intracellular Gram-negative coccobacilli that chronically infect humans as well as domestic and wild-type mammals, and cause brucellosis. Alternatively activated macrophages (M2a) induced by IL-4/IL-13 via STAT6 signaling pathways have been frequently described as a favorable niche for long-term persistence of intracellular pathogens. Based on the observation that M2a-like macrophages are induced in the spleen during the chronic phase of B. abortus infection in mice and are strongly infected in vitro, it has been suggested that M2a macrophages could be a potential in vivo niche for Brucella. In order to test this hypothesis, we used a model in which infected cells can be observed directly in situ and where the differentiation of M2a macrophages is favored by the absence of an IL-12-dependent Th1 response. We performed an in situ analysis by fluorescent microscopy of the phenotype of B. melitensis infected spleen cells from intranasally infected IL-12p40-/- BALB/c mice and the impact of STAT6 deficiency on this phenotype. Most of the infected spleen cells contained high levels of lipids and expressed CD11c and CD205 dendritic cell markers and Arginase1, but were negative for the M2a markers Fizz1 or CD301. Furthermore, STAT6 deficiency had no effect on bacterial growth or the reservoir cell phenotype in vivo, leading us to conclude that, in our model, the infected cells were not Th2-induced M2a macrophages. This characterization of B. melitensis reservoir cells could provide a better understanding of Brucella persistence in the host and lead to the design of more efficient therapeutic strategies.

Induction of Interleukin-9-Producing Mucosal Mast Cells Promotes Susceptibility to IgE-Mediated Experimental Food Allergy.

Experimental IgE-mediated food allergy depends on intestinal anaphylaxis driven by interleukin-9 (IL-9). However, the primary cellular source of IL-9 and the mechanisms underlying the susceptibility to food-induced intestinal anaphylaxis remain unclear. Herein, we have reported the identification of multifunctional IL-9-producing mucosal mast cells (MMC9s) that can secrete prodigious amounts of IL-9 and IL-13 in response to IL-33, and mast cell protease-1 (MCPt-1) in response to antigen and IgE complex crosslinking, respectively. Repeated intragastric antigen challenge induced MMC9 development that required T cells, IL-4, and STAT6 transcription factor, but not IL-9 signals. Mice ablated of MMC9 induction failed to develop intestinal mastocytosis, which resulted in decreased food allergy symptoms that could be restored by adoptively transferred MMC9s. Finally, atopic patients that developed food allergy displayed increased intestinal expression of Il9- and MC-specific transcripts. Thus, the induction of MMC9s is a pivotal step to acquire the susceptibility to IgE-mediated food allergy.

[Advances in research of Toxoplasma gondii rhoptry protein ROP16].

ROP16 is one member of the rhoptrys protein family in Toxoplasma gondii. In its protein structure, there exists serine/threonine kinase domain, which is the important virulence factor in the invasion process of T. gondii. ROP16 can secretes into the nucleus of the host cells, and can phosphorylate the signal transducer and activator of transcription (STAT3/6) and interfere the signal transduction pathway in the host cell. In this paper, the structure and function, as well as the immunogenicity of ROP16 are summarized.

JAK3/STAT6 Stimulates Bone Marrow-Derived Fibroblast Activation in Renal Fibrosis.

Renal fibrosis is a final common manifestation of CKD resulting in progressive loss of kidney function. Bone marrow-derived fibroblast precursors contribute significantly to the pathogenesis of renal fibrosis. However, the signaling mechanisms underlying the activation of bone marrow-derived fibroblast precursors in the kidney are not fully understood. In this study, we investigated the role of the Janus kinase 3 (JAK3)/signal transducer and activator of transcription (STAT6) signaling pathway in the activation of bone marrow-derived fibroblasts. In cultured mouse monocytes, IL-4 or IL-13 activated STAT6 and induced expression of α-smooth muscle actin and extracellular matrix proteins (fibronectin and collagen I), which was abolished by a JAK3 inhibitor (CP690,550) in a dose-dependent manner or blocked in the absence of STAT6. In vivo, STAT6 was activated in interstitial cells of the obstructed kidney, an effect that was abolished by CP690,550. Mice treated with CP690,550 accumulated fewer bone marrow-derived fibroblasts in the obstructed kidneys compared with vehicle-treated mice. Treatment with CP690,550 also significantly reduced myofibroblast transformation, matrix protein expression, fibrosis development, and apoptosis in obstructed kidneys. Furthermore, STAT6-deficient mice accumulated fewer bone marrow-derived fibroblasts in the obstructed kidneys, produced less extracellular matrix protein, and developed much less fibrosis. Finally, wild-type mice engrafted with STAT6(-/-) bone marrow cells displayed fewer bone marrow-derived fibroblasts in the obstructed kidneys and showed less severe renal fibrosis compared with wild-type mice engrafted with STAT6(+/+) bone marrow cells. Our results demonstrate that JAK3/STAT6 has an important role in bone marrow-derived fibroblast activation, extracellular matrix production, and interstitial fibrosis development.

Cross talk between histone deacetylase 4 and STAT6 in the transcriptional regulation of arginase 1 during mouse dendritic cell differentiation.

l-Arginine and l-arginine-metabolizing enzymes play important roles in the biology of some types of myeloid cells, including macrophage and myeloid-derived suppressor cells. In this study, we found evidence that arginase 1 (Arg1) is required for the differentiation of mouse dendritic cells (DCs). Expression of Arg1 was robustly induced during monocyte-derived DC differentiation. Ectopic expression of Arg1 significantly promoted monocytic DC differentiation in a granulocyte-macrophage colony-stimulating factor culture system and also facilitated the differentiation of CD8α(+) conventional DCs in the presence of Flt3 ligand. Knockdown of Arg1 reversed these effects. Mechanistic studies showed that the induced expression of Arg1 in differentiating DCs was caused by enhanced recruitment of histone deacetylase 4 (HDAC4) to the Arg1 promoter region, which led to a reduction in the acetylation of both the histone 3 and STAT6 proteins and subsequent transcriptional activation of Arg1. Further investigation identified a novel STAT6 binding site within the Arg1 promoter that mediated its regulation by STAT6 and HDAC4. These observations suggest that the cross talk between HDAC4 and STAT6 is an important regulatory mechanism of Arg1 transcription in DCs. Moreover, overexpression of Arg1 clearly abrogated the ability of HDAC inhibitors to suppress DC differentiation. In conclusion, we show that Arg1 is a novel regulator of myeloid DC differentiation.

Transcriptional reversion of cardiac myocyte fate during mammalian cardiac regeneration.

RATIONALE: Neonatal mice have the capacity to regenerate their hearts in response to injury, but this potential is lost after the first week of life. The transcriptional changes that underpin mammalian cardiac regeneration have not been fully characterized at the molecular level. OBJECTIVE: The objectives of our study were to determine whether myocytes revert the transcriptional phenotype to a less differentiated state during regeneration and to systematically interrogate the transcriptional data to identify and validate potential regulators of this process. METHODS AND RESULTS: We derived a core transcriptional signature of injury-induced cardiac myocyte (CM) regeneration in mouse by comparing global transcriptional programs in a dynamic model of in vitro and in vivo CM differentiation, in vitro CM explant model, as well as a neonatal heart resection model. The regenerating mouse heart revealed a transcriptional reversion of CM differentiation processes, including reactivation of latent developmental programs similar to those observed during destabilization of a mature CM phenotype in the explant model. We identified potential upstream regulators of the core network, including interleukin 13, which induced CM cell cycle entry and STAT6/STAT3 signaling in vitro. We demonstrate that STAT3/periostin and STAT6 signaling are critical mediators of interleukin 13 signaling in CMs. These downstream signaling molecules are also modulated in the regenerating mouse heart. CONCLUSIONS: Our work reveals new insights into the transcriptional regulation of mammalian cardiac regeneration and provides the founding circuitry for identifying potential regulators for stimulating heart regeneration.