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Sample records for activated stat3 pias3

  1. SHP2, SOCS3 and PIAS3 Expression Patterns in Medulloblastomas: Relevance to STAT3 Activation and Resveratrol-Suppressed STAT3 Signaling.

    PubMed

    Li, Cong; Li, Hong; Zhang, Peng; Yu, Li-Jun; Huang, Tian-Miao; Song, Xue; Kong, Qing-You; Dong, Jian-Li; Li, Pei-Nan; Liu, Jia

    2016-12-27

    Activated STAT3 signaling is critical for human medulloblastoma cells. SHP2, SOCS3 and PIAS3 are known as the negative regulators of STAT3 signaling, while their relevance to frequent STAT3 activation in medulloblastomas remains unknown. Tissue microarrays were constructed with 17 tumor-surrounding noncancerous brain tissues and 61 cases of the classic medulloblastomas, 44 the large-cell medulloblastomas, and 15 nodular medulloblastomas, which were used for immunohistochemical profiling of STAT3, SHP2, SOCS3 and PIAS3 expression patterns and the frequencies of STAT3 nuclear translocation. Three human medulloblastoma cell lines (Daoy, UW228-2 and UW228-3) were cultured with and without 100 μM resveratrol supplementation. The influences of resveratrol in SHP2, SOCS3 and PIAS3 expression and SOCS3 knockdown in STAT3 activation were analyzed using multiple experimental approaches. SHP2, SOCS3 and PIAS3 levels are reduced in medulloblastomas in vivo and in vitro, of which PIAS3 downregulation is more reversely correlated with STAT3 activation. In resveratrol-suppressed medulloblastoma cells with STAT3 downregulation and decreased incidence of STAT3 nuclear translocation, PIAS3 is upregulated, the SHP2 level remains unchanged and SOCS3 is downregulated. SOCS3 proteins are accumulated in the distal ends of axon-like processes of resveratrol-differentiated medulloblastoma cells. Knockdown of SOCS3 expression by siRNA neither influences cell proliferation nor STAT3 activation or resveratrol sensitivity but inhibits resveratrol-induced axon-like process formation. Our results suggest that (1) the overall reduction of SHP2, SOCS3 and PIAS3 in medulloblastoma tissues and cell lines; (2) the more inverse relevance of PIAS3 expression with STAT3 activation; (3) the favorable prognostic values of PIAS3 for medulloblastomas and (4) the involvement of SOCS3 in resveratrol-promoted axon regeneration of medulloblastoma cells.

  2. SHP2, SOCS3 and PIAS3 Expression Patterns in Medulloblastomas: Relevance to STAT3 Activation and Resveratrol-Suppressed STAT3 Signaling

    PubMed Central

    Li, Cong; Li, Hong; Zhang, Peng; Yu, Li-Jun; Huang, Tian-Miao; Song, Xue; Kong, Qing-You; Dong, Jian-Li; Li, Pei-Nan; Liu, Jia

    2016-01-01

    Background: Activated STAT3 signaling is critical for human medulloblastoma cells. SHP2, SOCS3 and PIAS3 are known as the negative regulators of STAT3 signaling, while their relevance to frequent STAT3 activation in medulloblastomas remains unknown. Methods: Tissue microarrays were constructed with 17 tumor-surrounding noncancerous brain tissues and 61 cases of the classic medulloblastomas, 44 the large-cell medulloblastomas, and 15 nodular medulloblastomas, which were used for immunohistochemical profiling of STAT3, SHP2, SOCS3 and PIAS3 expression patterns and the frequencies of STAT3 nuclear translocation. Three human medulloblastoma cell lines (Daoy, UW228-2 and UW228-3) were cultured with and without 100 μM resveratrol supplementation. The influences of resveratrol in SHP2, SOCS3 and PIAS3 expression and SOCS3 knockdown in STAT3 activation were analyzed using multiple experimental approaches. Results: SHP2, SOCS3 and PIAS3 levels are reduced in medulloblastomas in vivo and in vitro, of which PIAS3 downregulation is more reversely correlated with STAT3 activation. In resveratrol-suppressed medulloblastoma cells with STAT3 downregulation and decreased incidence of STAT3 nuclear translocation, PIAS3 is upregulated, the SHP2 level remains unchanged and SOCS3 is downregulated. SOCS3 proteins are accumulated in the distal ends of axon-like processes of resveratrol-differentiated medulloblastoma cells. Knockdown of SOCS3 expression by siRNA neither influences cell proliferation nor STAT3 activation or resveratrol sensitivity but inhibits resveratrol-induced axon-like process formation. Conclusion: Our results suggest that (1) the overall reduction of SHP2, SOCS3 and PIAS3 in medulloblastoma tissues and cell lines; (2) the more inverse relevance of PIAS3 expression with STAT3 activation; (3) the favorable prognostic values of PIAS3 for medulloblastomas and (4) the involvement of SOCS3 in resveratrol-promoted axon regeneration of medulloblastoma cells. PMID:28035977

  3. TRIM8 regulates stemness in glioblastoma through PIAS3-STAT3.

    PubMed

    Zhang, Changming; Mukherjee, Subhas; Tucker-Burden, Carol; Ross, James L; Chau, Monica J; Kong, Jun; Brat, Daniel J

    2017-03-01

    Glioblastoma (GBM) is the most malignant form of primary brain tumor, and GBM stem-like cells (GSCs) contribute to the rapid growth, therapeutic resistance, and clinical recurrence of these fatal tumors. STAT3 signaling supports the maintenance and proliferation of GSCs, yet regulatory mechanisms are not completely understood. Here, we report that tri-partite motif-containing protein 8 (TRIM8) activates STAT3 signaling to maintain stemness and self-renewing capabilities of GSCs. TRIM8 (also known as 'glioblastoma-expressed ring finger protein') is expressed equally in GBM and normal brain tissues, despite its hemizygous deletion in the large majority of GBMs, and its expression is highly correlated with stem cell markers. Experimental knockdown of TRIM8 reduced GSC self-renewal and expression of SOX2, NESTIN, and p-STAT3, and promoted glial differentiation. Overexpression of TRIM8 led to higher expression of p-STAT3, c-MYC, SOX2, NESTIN, and CD133, and enhanced GSC self-renewal. We found that TRIM8 activates STAT3 by suppressing the expression of PIAS3, an inhibitor of STAT3, most likely through E3-mediated ubiquitination and proteasomal degradation. Interestingly, we also found that STAT3 activation upregulates TRIM8, providing a mechanism for normalized TRIM8 expression in the setting of hemizygous gene deletion. These data demonstrate that bidirectional TRIM8-STAT3 signaling regulates stemness in GSC.

  4. Protein inhibitor of activated STAT3 (PIAS3) protein promotes SUMOylation and nuclear sequestration of the intracellular domain of ErbB4 protein.

    PubMed

    Sundvall, Maria; Korhonen, Anna; Vaparanta, Katri; Anckar, Julius; Halkilahti, Kalle; Salah, Zaidoun; Aqeilan, Rami I; Palvimo, Jorma J; Sistonen, Lea; Elenius, Klaus

    2012-06-29

    ErbB4 is a receptor tyrosine kinase implicated in the development and homeostasis of the heart, central nervous system, and mammary gland. Cleavable isoforms of ErbB4 release a soluble intracellular domain (ICD) that can translocate to the nucleus and function as a transcriptional coregulator. In search of regulatory mechanisms of ErbB4 ICD function, we identified PIAS3 as a novel interaction partner of ErbB4 ICD. In keeping with the small ubiquitin-like modifier (SUMO) E3 ligase function of protein inhibitor of activated STAT (PIAS) proteins, we showed that the ErbB4 ICD is modified by SUMO, and that PIAS3 stimulates the SUMOylation. Upon overexpression of PIAS3, the ErbB4 ICD generated from the full-length receptor accumulated into the nucleus in a manner that was dependent on the functional nuclear localization signal of ErbB4. In the nucleus, ErbB4 colocalized with PIAS3 and SUMO-1 in promyelocytic leukemia nuclear bodies, nuclear domains involved in regulation of transcription. Accordingly, PIAS3 overexpression had an effect on the transcriptional coregulatory activity of ErbB4, repressing its ability to coactivate transcription with Yes-associated protein. Finally, knockdown of PIAS3 with siRNA partially rescued the inhibitory effect of the ErbB4 ICD on differentiation of MDA-MB-468 breast cancer and HC11 mammary epithelial cells. Our findings illustrate that PIAS3 is a novel regulator of ErbB4 receptor tyrosine kinase, controlling its nuclear sequestration and function.

  5. Protein Inhibitor of Activated STAT3 (PIAS3) Protein Promotes SUMOylation and Nuclear Sequestration of the Intracellular Domain of ErbB4 Protein*

    PubMed Central

    Sundvall, Maria; Korhonen, Anna; Vaparanta, Katri; Anckar, Julius; Halkilahti, Kalle; Salah, Zaidoun; Aqeilan, Rami I.; Palvimo, Jorma J.; Sistonen, Lea; Elenius, Klaus

    2012-01-01

    ErbB4 is a receptor tyrosine kinase implicated in the development and homeostasis of the heart, central nervous system, and mammary gland. Cleavable isoforms of ErbB4 release a soluble intracellular domain (ICD) that can translocate to the nucleus and function as a transcriptional coregulator. In search of regulatory mechanisms of ErbB4 ICD function, we identified PIAS3 as a novel interaction partner of ErbB4 ICD. In keeping with the small ubiquitin-like modifier (SUMO) E3 ligase function of protein inhibitor of activated STAT (PIAS) proteins, we showed that the ErbB4 ICD is modified by SUMO, and that PIAS3 stimulates the SUMOylation. Upon overexpression of PIAS3, the ErbB4 ICD generated from the full-length receptor accumulated into the nucleus in a manner that was dependent on the functional nuclear localization signal of ErbB4. In the nucleus, ErbB4 colocalized with PIAS3 and SUMO-1 in promyelocytic leukemia nuclear bodies, nuclear domains involved in regulation of transcription. Accordingly, PIAS3 overexpression had an effect on the transcriptional coregulatory activity of ErbB4, repressing its ability to coactivate transcription with Yes-associated protein. Finally, knockdown of PIAS3 with siRNA partially rescued the inhibitory effect of the ErbB4 ICD on differentiation of MDA-MB-468 breast cancer and HC11 mammary epithelial cells. Our findings illustrate that PIAS3 is a novel regulator of ErbB4 receptor tyrosine kinase, controlling its nuclear sequestration and function. PMID:22584572

  6. Protein inhibitor of activated STAT3 inhibits adipogenic gene expression

    SciTech Connect

    Deng Jianbei; Hua Kunjie; Caveney, Erica J.; Takahashi, Nobuyuki; Harp, Joyce B. . E-mail: jharp@unc.edu

    2006-01-20

    Protein inhibitor of activated STAT3 (PIAS3), a cytokine-induced repressor of signal transducer and activator of transcription 3 (STAT3) and a modulator of a broad array of nuclear proteins, is expressed in white adipose tissue, but its role in adipogenesis is not known. Here, we determined that PIAS3 was constitutively expressed in 3T3-L1 cells at all stages of adipogenesis. However, it translocated from the nucleus to the cytoplasm 4 days after induction of differentiation by isobutylmethylxanthine, dexamethasone, and insulin (MDI). In ob/ob mice, PIAS3 expression was increased in white adipose tissue depots compared to lean mice and was found in the cytoplasm of adipocytes. Overexpression of PIAS3 in differentiating preadipocytes, which localized primarily to the nucleus, inhibited mRNA level gene expression of adipogenic transcription factors C/EBP{alpha} and PPAR{gamma}, as well as their downstream target genes aP2 and adiponectin. PIAS3 also inhibited C/EBP{alpha} promoter activation mediated specifically by insulin, but not dexamethasone or isobutylmethylxanthine. Taken together, these data suggest that PIAS3 may play an inhibitory role in adipogenesis by modulating insulin-activated transcriptional activation events. Increased PIAS3 expression in adipose tissue may play a role in the metabolic disturbances of obesity.

  7. STAT3 Activities and Energy Metabolism: Dangerous Liaisons

    PubMed Central

    Camporeale, Annalisa; Demaria, Marco; Monteleone, Emanuele; Giorgi, Carlotta; Wieckowski, Mariusz R.; Pinton, Paolo; Poli, Valeria

    2014-01-01

    STAT3 mediates cytokine and growth factor receptor signalling, becoming transcriptionally active upon tyrosine 705 phosphorylation (Y-P). Constitutively Y-P STAT3 is observed in many tumors that become addicted to its activity, and STAT3 transcriptional activation is required for tumor transformation downstream of several oncogenes. We have recently demonstrated that constitutively active STAT3 drives a metabolic switch towards aerobic glycolysis through the transcriptional induction of Hif-1α and the down-regulation of mitochondrial activity, in both MEF cells expressing constitutively active STAT3 (Stat3C/C) and STAT3-addicted tumor cells. This novel metabolic function is likely involved in mediating pre-oncogenic features in the primary Stat3C/C MEFs such as resistance to apoptosis and senescence and rapid proliferation. Moreover, it strongly contributes to the ability of primary Stat3C/C MEFs to undergo malignant transformation upon spontaneous immortalization, a feature that may explain the well known causative link between STAT3 constitutive activity and tumor transformation under chronic inflammatory conditions. Taken together with the recently uncovered role of STAT3 in regulating energy metabolism from within the mitochondrion when phosphorylated on Ser 727, these data place STAT3 at the center of a hub regulating energy metabolism under different conditions, in most cases promoting cell survival, proliferation and malignant transformation even though with distinct mechanisms. PMID:25089666

  8. Somatic mutations activating STAT3 in human inflammatory hepatocellular adenomas

    PubMed Central

    Pilati, Camilla; Amessou, Mohamed; Bihl, Michel P.; Balabaud, Charles; Van Nhieu, Jeanne Tran; Paradis, Valérie; Nault, Jean Charles; Izard, Tina; Bioulac-Sage, Paulette; Couchy, Gabrielle; Poussin, Karine

    2011-01-01

    Inflammatory hepatocellular adenomas (IHCAs) are benign liver tumors. 60% of these tumors have IL-6 signal transducer (IL6ST; gp130) mutations that activate interleukin 6 (IL-6) signaling. Here, we report that 12% of IHCA subsets lacking IL6ST mutations harbor somatic signal transducer and activator of transcription 3 (STAT3) mutations (6/49). Most of these mutations are amino acid substitutions in the SH2 domain that directs STAT3 dimerization. In contrast to wild-type STAT3, IHCA STAT3 mutants constitutively activated the IL-6 signaling pathway independent of ligand in hepatocellular cells. Indeed, the IHCA STAT3 Y640 mutant homodimerized independent of IL-6 and was hypersensitive to IL-6 stimulation. This was associated with phosphorylation of tyrosine 705, a residue required for IL-6–induced STAT3 activation. Silencing or inhibiting the tyrosine kinases JAK1 or Src, which phosphorylate STAT3, impaired constitutive activity of IHCA STAT3 mutants in hepatocellular cells. Thus, we identified for the first time somatic STAT3 mutations in human tumors, revealing a new mechanism of recurrent STAT3 activation and underscoring the role of the IL-6–STAT3 pathway in benign hepatocellular tumorigenesis. PMID:21690253

  9. Morin inhibits STAT3 tyrosine 705 phosphorylation in tumor cells through activation of protein tyrosine phosphatase SHP1.

    PubMed

    Gupta, Subash C; Phromnoi, Kanokkarn; Aggarwal, Bharat B

    2013-04-01

    The major goal of cancer drug discovery is to find an agent that is safe and affordable, yet effective against cancer. Here we show that morin (3,5,7,2',4'-pentahydroxyflavone) has potential against cancer cells through suppression of the signal transducer and activator of transcription 3 (STAT3) pathway, which is closely linked to the transformation, survival, proliferation, and metastasis of cancer. We found that morin completely suppressed inducible and constitutively activated STAT3 and blocked the nuclear translocation of STAT3 and its DNA binding in multiple myeloma and head and neck squamous carcinoma cells. Morin inhibited activated Src, JAK-1, and JAK-2, all of which are linked to STAT3 activation, while up-regulating a protein inhibitor of activated STAT3, PIAS3. Pervanadate reversed the effects of morin on STAT3 phosphorylation, indicating the role of a protein tyrosine phosphatase. Furthermore, morin induced SHP1 expression at both the mRNA and protein levels, and silencing of SHP1 abrogated the effect of morin on STAT3 phosphorylation, indicating that morin mediates its effects on STAT3 through SHP1. Suppression of STAT3 correlated with the down-regulation of various gene products linked to tumor survival, proliferation, and angiogenesis and led to sensitization of tumor cells to thalidomide and bortezomib. Comparing the activities of morin with those of four structurally related flavonols demonstrated the importance of hydroxyl groups in the B ring in inhibiting STAT3 activation. These findings suggest that morin suppresses the STAT3 pathway, leading to the down-regulation of STAT3-dependent gene expression and chemosensitization of tumor cells. Copyright © 2012 Elsevier Inc. All rights reserved.

  10. Protein Inhibitor of Activated STAT3 Suppresses Oxidized LDL-induced Cell Responses during Atherosclerosis in Apolipoprotein E-deficient Mice

    PubMed Central

    Wang, Rong; Zhang, Yanjin; Xu, Liran; Lin, Yan; Yang, Xiaofeng; Bai, Liang; Chen, Yulong; Zhao, Sihai; Fan, Jianglin; Cheng, Xianwu; Liu, Enqi

    2016-01-01

    Atherosclerosis is a serious public health concern. Excessive inflammatory responses of vascular cells are considered a pivotal pathogenesis mechanism underlying atherosclerosis development. It is known that Janus kinase/signal transducer and activator of transcription 3 (JAK/STAT3) signalling plays an important role in atherosclerosis progression. Protein inhibitor of activated STAT3 (PIAS3) is the key negative regulator of JAK/STAT3 signalling. However, its effect on atherogenesis is unknown. Here, we observed that PIAS3 levels are reduced in atherosclerotic lesions and that PIAS3 expression decreases in conjunction with increases in interleukin-6 expression and atherosclerosis severity. Oxidized low-density lipoprotein (ox-LDL), an atherogenic stimulus, reduced PIAS3 expression, an effect that may be attributed to nitric oxide synthesis upregulation. In turn, PIAS3 overexpression effectively suppressed ox-LDL-induced inflammation, lipid accumulation and vascular smooth muscle cell proliferation. These results indicate that PIAS3 is a critical repressor of atherosclerosis progression. The findings of this study have contributed to our understanding on the pathogenesis of atherosclerosis and have provided us with a potential target through which we can inhibit atherosclerosis-related cellular responses. PMID:27845432

  11. Activation of Hepatic STAT3 Maintains Pulmonary Defense during Endotoxemia

    PubMed Central

    Hilliard, Kristie L.; Allen, Eri; Traber, Katrina E.; Kim, Yuri; Wasserman, Gregory A.; Jones, Matthew R.; Mizgerd, Joseph P.

    2015-01-01

    Pneumonia and infection-induced sepsis are worldwide public health concerns. Both pathologies elicit systemic inflammation and induce a robust acute-phase response (APR). Although APR activation is well regarded as a hallmark of infection, the direct contributions of liver activation to pulmonary defense during sepsis remain unclear. By targeting STAT3-dependent acute-phase changes in the liver, we evaluated the role of liver STAT3 activity in promoting host defense in the context of sepsis and pneumonia. We employed a two-hit endotoxemia/pneumonia model, whereby administration of 18 h of intraperitoneal lipopolysaccharide (LPS; 5 mg/kg of body weight) was followed by intratracheal Escherichia coli (106 CFU) in wild-type mice or those lacking hepatocyte STAT3 (hepSTAT3−/−). Pneumonia alone (without endotoxemia) was effectively controlled in the absence of liver STAT3. Following endotoxemia and pneumonia, however, hepSTAT3−/− mice, with significantly reduced levels of circulating and airspace acute-phase proteins, exhibited significantly elevated lung and blood bacterial burdens and mortality. These data suggested that STAT3-dependent liver responses are necessary to promote host defense. While neither recruited airspace neutrophils nor lung injury was altered in endotoxemic hepSTAT3−/− mice, alveolar macrophage reactive oxygen species generation was significantly decreased. Additionally, bronchoalveolar lavage fluid from this group of hepSTAT3−/− mice allowed greater bacterial growth ex vivo. These results suggest that hepatic STAT3 activation promotes both cellular and humoral lung defenses. Taken together, induction of liver STAT3-dependent gene expression programs is essential to countering the deleterious consequences of sepsis on pneumonia susceptibility. PMID:26216424

  12. Inhibition of STAT3 activation by KT-18618 via the disruption of the interaction between JAK3 and STAT3.

    PubMed

    Shin, Dae-Seop; Jung, Seung Nam; Yun, Jieun; Lee, Chang Woo; Han, Dong Cho; Kim, Bumtae; Min, Yong Ki; Kang, Nam Sook; Kwon, Byoung-Mog

    2014-05-01

    The constitutive activation of STAT3 in human cancers causes the abnormal proliferation and survival of cancer cells, and thus, STAT3 is a therapeutic target of antitumor drugs. We screened a small-molecule library of 8600 synthetic compounds from the "Korea Chemical Bank" to identify inhibit STAT3 activity using a cell-based luciferase assay system. KT-18618 ((Z)-N-(4-chlorophenyl)-N-methyl-2-[1,3,3,3,-tetrafluoro-2-(thiophen-2-yl)prop-1-enyloxy]-acetamide) was selected as a novel inhibitor of the JAK/STAT3 pathway. KT-18618 inhibited STAT3 phosphorylation and the expression of STAT3-regulated genes. The inhibition of STAT3 phosphorylation led to the apoptosis of MDA-MB-468 cells. We postulated that the inhibition of the JAK family of proteins or c-Src inhibited STAT3 phosphorylation. Interestingly, the phosphorylation of these kinases was only mildly inhibited, but the phosphorylation of STAT3 was completely inhibited. This result implies that the inhibition of STAT3 phosphorylation by KT-18618 is an independent event that occurs through the phosphorylation of upstream kinases. Co-immunoprecipitation experiments revealed that KT-18618 inhibited the JAK3-STAT3 interaction. Moreover, JAK3 molecules were captured by biotinylated KT-18618, implying that KT-18618 bound to JAK3 molecules. Additionally, 1μM KT-18618 inhibited JAK3 kinase activity by approximately 28% in an in vitro kinase assay. From these results, we suggest that KT-18618 binds to JAK3 molecules and disrupts the JAK3-STAT3 interaction, which leads to the inhibition of STAT3 phosphorylation. KT-18618 is the first inhibitor of the JAK3-STAT3 interaction.

  13. Urokinase induces activation of STAT3 in lung epithelial cells.

    PubMed

    Shetty, Sreerama; Rao, Gadiparthi N; Cines, Douglas B; Bdeir, Khalil

    2006-10-01

    Urokinase-type plasminogen activator (uPA) is a serine protease that plays a major role in diverse physiological and pathological processes. Studies from our laboratory have shown that exposure of human lung epithelial cells to uPA induces proliferation. To understand uPA mitogenic signaling events, we sought to elucidate its effects on tyrosine phosphorylation in a human bronchial epithelial cell line (Beas2B). uPA induced tyrosine phosphorylation of several proteins in a time-dependent manner. One of these proteins was identified as the 91-kDa signal transduction activator transcription (Stat)3 moiety. Tyrosine phosphorylation of Stat3 by uPA was time dependent. uPA induced Stat3-DNA binding activity in a time-dependent manner. uPA-induced Stat3 activation does not require uPA catalytic activity, as the uPA amino-terminal fragment alone was as potent as active two-chain uPA (tcuPA) in causing this effect. Single-chain uPA likewise induced tyrosine phosphorylation of Stat3 to a similar extent as intact tcuPA. Plasmin did not alter uPA-induced Stat3 activation. Furthermore, transfection of Beas2B cells with dominant-negative Stat3 blocked uPA-induced DNA synthesis. These results reveal for the first time that the uPA-uPAR interaction leads to activation of Stat3, independent of its catalytic activity but dependent on its interaction with its receptor, uPAR, leading to DNA synthesis in lung epithelial cells.

  14. Activation of epithelial STAT3 regulates intestinal homeostasis.

    PubMed

    Neufert, Clemens; Pickert, Geethanjali; Zheng, Yan; Wittkopf, Nadine; Warntjen, Moritz; Nikolaev, Alexei; Ouyang, Wenjun; Neurath, Markus F; Becker, Christoph

    2010-02-15

    The intestinal epithelium that lines the mucosal surface along the GI-tract is a key player for the intestinal homeostasis of the healthy individual. In case of a mucosal damage or a barrier defect as seen in patients with inflammatory bowel disease, the balance is disturbed, and translocation of intestinal microbes to the submucosa is facilitated. We recently demonstrated a pivotal role of STAT3 activation in intestinal epithelial cells (IEC) for the restoration of the balance at the mucosal surface of the gut in an experimental colitis model. STAT3 was rapidly induced in intestinal epithelial cells upon challenge of mice in both experimental colitis and intestinal wound healing models. STAT3 activation was found to be dispensable in the steady-state conditions but was important for efficient regeneration of the epithelium in response to injury. Here, we extend our previous findings by showing epithelial STAT3 activation in human patients suffering from IBD and provide additional insights how the activation of epithelial STAT3 by IL-22 regulates intestinal homeostasis and mucosal wound healing. We also demonstrate that antibody-mediated neutralization of IL-22 has little impact on the development of experimental colitis in mice, but significantly delays recovery from colitis. Thus, our data suggest that targeting the STAT3 signaling pathway in IEC is a promising therapeutic approach in situations when the intestinal homeostasis is disturbed, e.g., as seen in Crohn's disease or Ulcerative colitis.

  15. Mutations in the linker domain affect phospho-STAT3 function and suggest targets for interrupting STAT3 activity.

    PubMed

    Mertens, Claudia; Haripal, Bhagwattie; Klinge, Sebastian; Darnell, James E

    2015-12-01

    Crystallography of the cores of phosphotyrosine-activated dimers of STAT1 (132-713) and STAT3 (127-722) bound to a similar double-stranded deoxyoligonucleotide established the domain structure of the STATs and the structural basis for activation through tyrosine phosphorylation and dimerization. We reported earlier that mutants in the linker domain of STAT1 that connect the DNA-binding domain and SH2 domain can prevent transcriptional activation. Because of the pervasive importance of persistently activated STAT3 in many human cancers and the difficulty of finding useful drug candidates aimed at disrupting the pY interchange in active STAT3 dimers, we have examined effects of an array of mutants in the STAT3 linker domain. We have found several STAT3 linker domain mutants to have profound effects of inhibiting STAT3 transcriptional activation. From these results, we propose (i) there is definite functional interaction of the linker both with the DNA binding domain and with the SH2 domain, and (ii) these putative contacts provide potential new targets for small molecule-induced pSTAT3 inhibition.

  16. Mutations in the linker domain affect phospho-STAT3 function and suggest targets for interrupting STAT3 activity

    PubMed Central

    Mertens, Claudia; Haripal, Bhagwattie; Klinge, Sebastian; Darnell, James E.

    2015-01-01

    Crystallography of the cores of phosphotyrosine-activated dimers of STAT1 (132–713) and STAT3 (127–722) bound to a similar double-stranded deoxyoligonucleotide established the domain structure of the STATs and the structural basis for activation through tyrosine phosphorylation and dimerization. We reported earlier that mutants in the linker domain of STAT1 that connect the DNA-binding domain and SH2 domain can prevent transcriptional activation. Because of the pervasive importance of persistently activated STAT3 in many human cancers and the difficulty of finding useful drug candidates aimed at disrupting the pY interchange in active STAT3 dimers, we have examined effects of an array of mutants in the STAT3 linker domain. We have found several STAT3 linker domain mutants to have profound effects of inhibiting STAT3 transcriptional activation. From these results, we propose (i) there is definite functional interaction of the linker both with the DNA binding domain and with the SH2 domain, and (ii) these putative contacts provide potential new targets for small molecule-induced pSTAT3 inhibition. PMID:26553978

  17. Resveratrol inhibits Src and Stat3 signaling and induces the apoptosis of malignant cells containing activated Stat3 protein.

    PubMed

    Kotha, Anupama; Sekharam, Madhavi; Cilenti, Lucia; Siddiquee, Khandaker; Khaled, Annette; Zervos, Antonis S; Carter, Bradford; Turkson, James; Jove, Richard

    2006-03-01

    Resveratrol is a naturally occurring phytoalexin with antioxidant and antiinflammatory properties. Recent studies suggest that resveratrol possesses anticancer effects, although its mechanism of action is not well understood. We now show that resveratrol inhibits Src tyrosine kinase activity and thereby blocks constitutive signal transducer and activator of transcription 3 (Stat3) protein activation in malignant cells. Analyses of resveratrol-treated malignant cells harboring constitutively-active Stat3 reveal irreversible cell cycle arrest of v-Src-transformed mouse fibroblasts (NIH3T3/v-Src), human breast (MDA-MB-231), pancreatic (Panc-1), and prostate carcinoma (DU145) cell lines at the G0-G1 phase or at the S phase of human breast cancer (MDA-MB-468) and pancreatic cancer (Colo-357) cells, and loss of viability due to apoptosis. By contrast, cells treated with resveratrol, but lacking aberrant Stat3 activity, show reversible growth arrest and minimal loss of viability. Moreover, in malignant cells harboring constitutively-active Stat3, including human prostate cancer DU145 cells and v-Src-transformed mouse fibroblasts (NIH3T3/v-Src), resveratrol treatment represses Stat3-regulated cyclin D1 as well as Bcl-xL and Mcl-1 genes, suggesting that the antitumor cell activity of resveratrol is in part due to the blockade of Stat3-mediated dysregulation of growth and survival pathways. Our study is among the first to identify Src-Stat3 signaling as a target of resveratrol, further defining the mechanism of antitumor cell activity of resveratrol and raising its potential application in tumors with an activated Stat3 profile.

  18. Heme Mediated STAT3 Activation in Severe Malaria

    PubMed Central

    Liu, Mingli; Amodu, Audu S.; Pitts, Sidney; Patrickson, John; Hibbert, Jacqueline M.; Battle, Monica; Ofori-Acquah, Solomon F.; Stiles, Jonathan K.

    2012-01-01

    Background The mortality of severe malaria [cerebral malaria (CM), severe malaria anemia (SMA), acute lung injury (ALI) and acute respiratory distress syndrome (ARDS)] remains high despite the availability associated with adequate treatments. Recent studies in our laboratory and others have revealed a hitherto unknown correlation between chemokine CXCL10/CXCR3, Heme/HO-1 and STAT3 and cerebral malaria severity and mortality. Although Heme/HO-1 and CXCL10/CXCR3 interactions are directly involved in the pathogenesis of CM and fatal disease, the mechanism dictating how Heme/HO-1 and CXCL10/CXCR3 are expressed and regulated under these conditions is still unknown. We therefore tested the hypothesis that these factors share common signaling pathways and may be mutually regulated. Methods We first clarified the roles of Heme/HO-1, CXCL10/CXCR3 and STAT3 in CM pathogenesis utilizing a well established experimental cerebral malaria mouse (ECM, P. berghei ANKA) model. Then, we further determined the mechanisms how STAT3 regulates HO-1 and CXCL10 as well as mutual regulation among them in CRL-2581, a murine endothelial cell line. Results The results demonstrate that (1) STAT3 is activated by P. berghei ANKA (PBA) infection in vivo and Heme in vitro. (2) Heme up-regulates HO-1 and CXCL10 production through STAT3 pathway, and regulates CXCL10 at the transcriptional level in vitro. (3) HO-1 transcription is positively regulated by CXCL10. (4) HO-1 regulates STAT3 signaling. Conclusion Our data indicate that Heme/HO-1, CXCL10/CXCR3 and STAT3 molecules as well as related signaling pathways play very important roles in the pathogenesis of severe malaria. We conclude that these factors are mutually regulated and provide new opportunities to develop potential novel therapeutic targets that could be used to supplement traditional prophylactics and treatments for malaria and improve clinical outcomes while reducing malaria mortality. Our ultimate goal is to develop novel therapies

  19. HIV-1 gp120 activates the STAT3/interleukin-6 axis in primary human monocyte-derived dendritic cells.

    PubMed

    Del Cornò, Manuela; Donninelli, Gloria; Varano, Barbara; Da Sacco, Letizia; Masotti, Andrea; Gessani, Sandra

    2014-10-01

    Dendritic cells (DCs) are fundamental for the initiation of immune responses and are important players in AIDS immunopathogenesis. The modulation of DC functional activities represents a strategic mechanism for HIV-1 to evade immune surveillance. Impairment of DC function may result from bystander effects of HIV-1 envelope proteins independently of direct HIV-1 infection. In this study, we report that exposure of immature monocyte-derived DCs (MDDCs) to HIV-1 R5 gp120 resulted in the CCR5-dependent production of interleukin-6 (IL-6) via mitogen-activated protein kinase (MAPK)/NF-κB pathways. IL-6 in turn activated STAT3 by an autocrine loop. Concomitantly, gp120 promoted an early activation of STAT3 that further contributed to IL-6 induction. This activation paralleled a concomitant upregulation of the STAT3 inhibitor PIAS3. Notably, STAT3/IL-6 pathway activation was not affected by the CCR5-specific ligand CCL4. These results identify STAT3 as a key signaling intermediate activated by gp120 in MDDCs and highlight the existence of a virus-induced dysregulation of the IL-6/STAT3 axis. HIV-1 gp120 signaling through STAT3 may provide an explanation for the impairment of DC function observed upon HIV exposure. This study provides new evidence for the molecular mechanisms and signaling pathways triggered by HIV-1 gp120 in human DCs in the absence of productive infection, emphasizing a role of aberrant signaling in early virus-host interaction, contributing to viral pathogenesis. We identified STAT3 as a key component in the gp120-mediated signaling cascade involving MAPK and NF-κB components and ultimately leading to IL-6 secretion. STAT3 now is recognized as a key regulator of DC functions. Thus, the identification of this transcription factor as a signaling molecule mediating some of gp120's biological effects unveils a new mechanism by which HIV-1 may deregulate DC functions and contribute to AIDS pathogenesis. Copyright © 2014, American Society for Microbiology

  20. Activating STAT3 Alpha for Promoting Healing of Neurons

    NASA Technical Reports Server (NTRS)

    Conway, Greg

    2008-01-01

    A method of promoting healing of injured or diseased neurons involves pharmacological activation of the STAT3 alpha protein. Usually, injured or diseased neurons heal incompletely or not at all for two reasons: (1) they are susceptible to apoptosis (cell death); and (2) they fail to engage in axogenesis that is, they fail to re-extend their axons to their original targets (e.g., muscles or other neurons) because of insufficiency of compounds, denoted neurotrophic factors, needed to stimulate such extension. The present method (see figure) of treatment takes advantage of prior research findings to the effect that the STAT3 alpha protein has anti-apoptotic and pro-axogenic properties.

  1. PIAS3 induction of PRB sumoylation represses PRB transactivation by destabilizing its retention in the nucleus

    PubMed Central

    Man, Jiang-Hong; Li, Hui-Yan; Zhang, Pei-Jing; Zhou, Tao; He, Kun; Pan, Xin; Liang, Bing; Li, Ai-Ling; Zhao, Jie; Gong, Wei-Li; Jin, Bao-Feng; Xia, Qing; Yu, Ming; Shen, Bei-Fen; Zhang, Xue-Min

    2006-01-01

    Progesterone receptor (PR) plays a critical role in cell proliferation and differentiation, and its transcriptional activity is known to be modulated by cofactor proteins. In the present study, we demonstrated that in the presence of progesterone, protein inhibitor of activated STAT-3 (PIAS3) significantly inhibited the PR transcriptional activity and the expression of progesterone-responsive genes. Reduction of endogenous PIAS3 by PIAS3 small-interfering RNA enhanced PR transactivation in a ligand-dependent manner. PIAS3 interacted with PR both in vitro and in vivo and the interaction was enhanced by progesterone. Furthermore, our findings suggested that PIAS3 strongly induced PRB sumoylation at three sites, Lys-7, Lys-388 and Lys-531. In addition, novel roles in PRB nuclear retention and transactivation were identified for these sites. Our data also suggested that PIAS3 was recruited in a largely hormone-dependent manner in response to a progesterone-responsive promoter. Finally, we demonstrated that PIAS3 inhibited the DNA-binding activity of PR and influenced its nuclear export as well as PR transactivation. Taken together, these data strongly suggested that PIAS3 played an important physiological role in PR function. PMID:17020914

  2. CNTF protects neurons from hypoxic injury through the activation of STAT3pTyr705.

    PubMed

    Gu, Ying Li; Gao, Guan Qun; Ma, Ning; Ye, Lin Lin; Zhang, Li Wei; Gao, Xu; Zhang, Zhuo Bo

    2016-12-01

    The aim of the present study was to investigate whether ciliary neurotrophic factor (CNTF) plays its neuroprotective role following hypoxic injury through the activation of signal transducer and activator of transcription 3 (STAT3) signaling. Firstly, to determine whether CNTF exerts its effects via STAT3 following hypoxic injury, cultured neurons from the cerebral cortex of mice were prepared and a neuronal model of hypoxia was then established. The neurons exposed to hypoxia were then pre-treated with CNTF and transfected with small interference RNA (siRNA) targeting STAT3 (STAT3 siRNA) using polybrene, or with STAT3Tyr705 mutant or STAT3Ser727 mutant using an electroporation system. The survival, proliferation and neurite outgrowth of the neurons subjected to different treatments were also determined. RT-qPCR and western blot analysis were employed to examine the expression levels of STAT3, p-STAT3Tyr705 and p-STAT3Ser727 following treatment with CNTF and other treatments. Our results revealed that treatment with CNTF: i) protected neurons from hypoxic injury by promoting survival and neurite growth; ii) induced a significant increase in the levels of STAT3, STAT3pTyr705 and the STAT3pTyr705/STAT3 ratio; it did not however, significantly affect the levels of STAT3pSer727 in the hypoxic cerebral cortex neurons. Transfection of the hypoxic neurons pre-treated with CNTF with STAT3 siRNA or STAT3Tyr705 neutralized the protective effects exerted by CNTF. The findings of our study thus demonstrate that CNTF protects neurons from hypoxic injury through the activation of STAT3pTyr705.

  3. PASD1 promotes STAT3 activity and tumor growth by inhibiting TC45-mediated dephosphorylation of STAT3 in the nucleus.

    PubMed

    Xu, Zhi-Sheng; Zhang, Hong-Xia; Zhang, Yu-Long; Liu, Tian-Tian; Ran, Yong; Chen, Liu-Ting; Wang, Yan-Yi; Shu, Hong-Bing

    2016-06-01

    Activation of the transcription factor signal transducer and activator of transcription 3 (STAT3) is tightly regulated during various physiological processes, such as cell proliferation, survival, and differentiation, and aberrant STAT3 activation results in tumorigenesis. In this study, we identified the cancer/testis antigen PASD1 as a positive regulator of STAT3 activity. Overexpression of PASD1 activated STAT3 and potentiated IL-6-induced activation of STAT3, whereas knockdown of PASD1 had opposite effects. Endogenous coimmunoprecipitation experiments indicated that PASD1 interacted with STAT3 in the nucleus. Overexpression of PASD1 enhanced both basal and IL-6-induced STAT3 phosphorylation at Y705, whereas knockdown of PASD1 had opposite effects. Mechanistically, PASD1 competed with TC45, a nuclear protein tyrosine phosphatase, to associate with STAT3, thus inhibited TC45-mediated dephosphorylation of STAT3. Consistently, knockdown of PASD1 inhibited expression of many pro-oncogenic genes, leading to suppression of cell proliferation, anchorage-independent growth, cell migration, and tumor growth in nude mice. Our findings demonstrate that PASD1 serves as a critical nuclear positive regulator of STAT3-mediated gene expression and tumorigenesis.

  4. Herpesvirus saimiri STP A11 protein interacts with STAT3 and stimulates its transcriptional activity.

    PubMed

    Park, Junsoo; Seo, Taegun; Jung, Jun; Choe, Joonho

    2004-07-16

    Herpesvirus saimiri (HVS) is an oncogenic gamma-2 herpesvirus that causes lymphoma in New World primates. HVS can be further divided into subgroups A, B, and C, based on sequence divergence. Saimiri transforming protein (STP) is coded for by the first open reading frame at the left end of the HVS genome and is responsible for its oncogenic potential. Here we show that STP A11 binds to signal transducers and activators of transcription 3 (STAT3), stimulates STAT3 phosphorylation, and activates STAT3-dependent transcription. STP A11 recruited c-Src kinase to phosphorylate STAT3 protein, and co-expression of STP A11 with c-Src dramatically increased STAT3 phosphorylation. We found that the amino terminal domain of STP A11 is required for both STAT3 interaction and activation, and that physical interaction is required for STAT3 activation.

  5. Mechanisms of STAT3 activation in the liver of FXR knockout mice

    PubMed Central

    Li, Guodong; Zhu, Yan; Tawfik, Ossama; Kong, Bo; Williams, Jessica A.; Zhan, Le; Kassel, Karen M.; Luyendyk, James P.; Wang, Li

    2013-01-01

    Farnesoid X receptor (FXR, Nr1h4) is a ligand-activated transcription factor belonging to the nuclear receptor superfamily. FXR is essential in maintaining bile acid (BA) homeostasis, and FXR−/− mice develop cholestasis, inflammation, and spontaneous liver tumors. The signal transducer and activator of transcription 3 (STAT3) is well known to regulate liver growth, and STAT3 is feedback inhibited by its target gene, the suppressor of cytokine signaling 3 (SOCS3). Strong activation of STAT3 was detected in FXR−/− mouse livers. However, the mechanism of STAT3 activation with FXR deficiency remains elusive. Wild-type (WT) and FXR−/− mice were used to detect STAT3 pathway activation in the liver. In vivo BA feeding or deprivation was used to determine the role of BAs in STAT3 activation, and in vitro molecular approaches were used to determine the direct transcriptional regulation of SOCS3 by FXR. STAT3 was activated in FXR−/− but not WT mice. BA feeding increased, but deprivation by cholestyramine reduced, serum inflammatory markers and STAT3 activation. Furthermore, the Socs3 gene was determined as a direct FXR target gene. The elevated BAs and inflammation, along with reduced SOCS3, collectively contribute to the activation of the STAT3 signaling pathway in the liver of FXR−/− mice. This study suggests that the constitutive activation of STAT3 may be a mechanism of liver carcinogenesis in FXR−/− mice. PMID:24091600

  6. Activated Rac1 requires gp130 for Stat3 activation, cell proliferation and migration

    SciTech Connect

    Arulanandam, Rozanne; Geletu, Mulu; Feracci, Helene; Raptis, Leda

    2010-03-10

    Rac1 (Rac) is a member of the Rho family of small GTPases which controls cell migration by regulating the organization of actin filaments. Previous results suggested that mutationally activated forms of the Rho GTPases can activate the Signal Transducer and Activator of Transcription-3 (Stat3), but the exact mechanism is a matter of controversy. We recently demonstrated that Stat3 activity of cultured cells increases dramatically following E-cadherin engagement. To better understand this pathway, we now compared Stat3 activity levels in mouse HC11 cells before and after expression of the mutationally activated Rac1 (Rac{sup V12}), at different cell densities. The results revealed for the first time a dramatic increase in protein levels and activity of both the endogenous Rac and Rac{sup V12} with cell density, which was due to inhibition of proteasomal degradation. In addition, Rac{sup V12}-expressing cells had higher Stat3, tyrosine-705 phosphorylation and activity levels at all densities, indicating that Rac{sup V12} is able to activate Stat3. Further examination of the mechanism of Stat3 activation showed that Rac{sup V12} expression caused a surge in mRNA of Interleukin-6 (IL6) family cytokines, known potent Stat3 activators. Knockdown of gp130, the common subunit of this family reduced Stat3 activity, indicating that these cytokines may be responsible for the Stat3 activation by Rac{sup V12}. The upregulation of IL6 family cytokines was required for cell migration and proliferation induced by Rac{sup V12}, as shown by gp130 knockdown experiments, thus demonstrating that the gp130/Stat3 axis represents an essential effector of activated Rac for the regulation of key cellular functions.

  7. Epstein-Barr virus-derived EBNA2 regulates STAT3 activation

    SciTech Connect

    Muromoto, Ryuta; Ikeda, Osamu; Okabe, Kanako; Togi, Sumihito; Kamitani, Shinya; Fujimuro, Masahiro; Harada, Shizuko; Oritani, Kenji; Matsuda, Tadashi

    2009-01-16

    The Epstein-Barr virus (EBV)-encoded latency protein EBNA2 is a nuclear transcriptional activator that is essential for EBV-induced cellular transformation. Here, we show that EBNA2 interacts with STAT3, a signal transducer for an interleukin-6 family cytokine, and enhances the transcriptional activity of STAT3 by influencing its DNA-binding activity. Furthermore, EBNA2 cooperatively acts on STAT3 activation with LMP1. These data demonstrate that EBNA2 acts as a transcriptional coactivator of STAT3.

  8. Activation status of the JAK/STAT3 pathway in mantle cell lymphoma.

    PubMed

    Yared, Marwan A; Khoury, Joseph D; Medeiros, L Jeffrey; Rassidakis, George Z; Lai, Raymond

    2005-08-01

    Signal transducer and activator of transcription 3 (STAT3) is oncogenic, and we previously found evidence of constitutive STAT3 activation in a relatively small number of frozen mantle cell lymphoma (MCL) cell tumors. To comprehensively survey the activation and phosphorylation status of STAT3 in MCL and to assess if STAT3 activation in these tumors is due to cytokine stimulation by examining the phosphorylation and activation status of Janus kinase (JAK), the physiologic activator of STAT3. We evaluated 43 formalin-fixed, paraffin-embedded MCL tumors using immunohistochemistry and phospho-specific antibodies against STAT3 and JAK. There were 37 small cell and 6 blastoid cases. There was heterogeneous expression of phospho-STAT3 (pSTAT3), with 23 negative cases (53%), 12 weakly positive cases (28%), and 8 strongly positive cases (19%). JAK3 was the only member detectable in 3 MCL cell lines, and immunoprecipitation data showed a relatively low level of tyrosine phosphorylation of JAK3 in these cells. Using immunohistochemistry, phospho-JAK3 (pJAK3) was detectable in 18 (44%) of 41 MCL tumors examined, and pJAK3 expression correlated with that of pSTAT3 (P = .008). A notable exception to this correlation was seen in the blastoid variant, since 4 (67%) of 6 blastoid cases were pSTAT3 positive but pJAK3 negative. We have confirmed our previous finding that STAT3 is constitutively activated in MCL tumors, with an overall frequency of 47% in this series. STAT3 activation in the small cell but not the blastoid variant of MCL is likely mediated by JAK3.

  9. High glucose enhances progression of cholangiocarcinoma cells via STAT3 activation.

    PubMed

    Saengboonmee, Charupong; Seubwai, Wunchana; Pairojkul, Chawalit; Wongkham, Sopit

    2016-01-08

    Epidemiological studies have indicated diabetes mellitus (DM) as a risk of cholangiocarcinoma (CCA), however, the effects and mechanisms of high glucose on progression of CCA remain unclear. This study reports for the first time of the enhancing effects of high glucose on aggressive phenotypes of CCA cells via STAT3 activation. CCA cells cultured in high glucose media exerted significantly higher rates of cell proliferation, adhesion, migration and invasion than those cultured in normal glucose. The phosphokinase array revealed STAT3 as the dominant signal activated in response to high glucose. Increased nuclear STAT3, p-STAT3 and its downstream target proteins, cyclin D1, vimentin and MMP2, were shown to be underling mechanisms of high glucose stimulation. The link of high glucose and STAT3 activation was confirmed in tumor tissues from CCA patients with DM that exhibited higher STAT3 activation than those without DM. Moreover, the levels of STAT3 activation were correlated with the levels of blood glucose. Finally, decreasing the level of glucose or using a STAT3 inhibitor could reduce the effects of high glucose. These findings suggest that controlling blood glucose or using a STAT3 inhibitor as an alternative approach may improve the therapeutic outcome of CCA patients with DM.

  10. Activating germline mutations in STAT3 cause early-onset multi-organ autoimmune disease

    PubMed Central

    Caswell, Richard; Allen, Hana Lango; De Franco, Elisa; McDonald, Timothy J.; Rajala, Hanna; Ramelius, Anita; Barton, John; Heiskanen, Kaarina; Heiskanen-Kosma, Tarja; Kajosaari, Merja; Murphy, Nuala P.; Milenkovic, Tatjana; Seppänen, Mikko; Lernmark, Åke; Mustjoki, Satu; Otonkoski, Timo; Kere, Juha; Morgan, Noel G.; Ellard, Sian; Hattersley, Andrew T.

    2014-01-01

    Monogenic causes of autoimmunity give key insights to the complex regulation of the immune system. We report a new monogenic cause of autoimmunity resulting from de novo germline activating STAT3 mutations in 5 individuals with a spectrum of early-onset autoimmune disease including type 1 diabetes. These findings emphasise the critical role of STAT3 in autoimmune disease and contrast with the germline inactivating STAT3 mutations that result in Hyper IgE syndrome. PMID:25038750

  11. Activating germline mutations in STAT3 cause early-onset multi-organ autoimmune disease.

    PubMed

    Flanagan, Sarah E; Haapaniemi, Emma; Russell, Mark A; Caswell, Richard; Lango Allen, Hana; De Franco, Elisa; McDonald, Timothy J; Rajala, Hanna; Ramelius, Anita; Barton, John; Heiskanen, Kaarina; Heiskanen-Kosma, Tarja; Kajosaari, Merja; Murphy, Nuala P; Milenkovic, Tatjana; Seppänen, Mikko; Lernmark, Åke; Mustjoki, Satu; Otonkoski, Timo; Kere, Juha; Morgan, Noel G; Ellard, Sian; Hattersley, Andrew T

    2014-08-01

    Monogenic causes of autoimmunity provide key insights into the complex regulation of the immune system. We report a new monogenic cause of autoimmunity resulting from de novo germline activating STAT3 mutations in five individuals with a spectrum of early-onset autoimmune disease, including type 1 diabetes. These findings emphasize the critical role of STAT3 in autoimmune disease and contrast with the germline inactivating STAT3 mutations that result in hyper IgE syndrome.

  12. Early Activation of STAT3 Regulates Reactive Astrogliosis Induced by Diverse Forms of Neurotoxicity

    PubMed Central

    O'Callaghan, James P.; Kelly, Kimberly A.; VanGilder, Reyna L.; Sofroniew, Michael V.; Miller, Diane B.

    2014-01-01

    Astrogliosis, a cellular response characterized by astrocytic hypertrophy and accumulation of GFAP, is a hallmark of all types of central nervous system (CNS) injuries. Potential signaling mechanisms driving the conversion of astrocytes into “reactive” phenotypes differ with respect to the injury models employed and can be complicated by factors such as disruption of the blood-brain barrier (BBB). As denervation tools, neurotoxicants have the advantage of selective targeting of brain regions and cell types, often with sparing of the BBB. Previously, we found that neuroinflammation and activation of the JAK2-STAT3 pathway in astrocytes precedes up regulation of GFAP in the MPTP mouse model of dopaminergic neurotoxicity. Here we show that multiple mechanistically distinct mouse models of neurotoxicity (MPTP, AMP, METH, MDA, MDMA, KA, TMT) engender the same neuroinflammatory and STAT3 activation responses in specific regions of the brain targeted by each neurotoxicant. The STAT3 effects seen for TMT in the mouse could be generalized to the rat, demonstrating cross-species validity for STAT3 activation. Pharmacological antagonists of the neurotoxic effects blocked neuroinflammatory responses, pSTAT3tyr705 and GFAP induction, indicating that damage to neuronal targets instigated astrogliosis. Selective deletion of STAT3 from astrocytes in STAT3 conditional knockout mice markedly attenuated MPTP-induced astrogliosis. Monitoring STAT3 translocation in GFAP-positive cells indicated that effects of MPTP, METH and KA on pSTAT3tyr705 were localized to astrocytes. These findings strongly implicate the STAT3 pathway in astrocytes as a broadly triggered signaling pathway for astrogliosis. We also observed, however, that the acute neuroinflammatory response to the known inflammogen, LPS, can activate STAT3 in CNS tissue without inducing classical signs of astrogliosis. Thus, acute phase neuroinflammatory responses and neurotoxicity-induced astrogliosis both signal through

  13. Epidermal hyperplasia induced by Raf-MAPK signaling requires Stat3 activation.

    PubMed

    Tarutani, Masahito; Nakajima, Kimiko; Takaishi, Mikiro; Ohko, Kentaro; Sano, Shigetoshi

    2013-11-01

    Raf is one of the downstream effectors of Ras GTPases, and plays a key role in regulating cell proliferation and differentiation through the activation of MAPK. We have previously demonstrated that temporal induction of Raf in the epidermis of K14-Raf:ER transgenic mice results in epidermal hyperplasia resembling squamous cell carcinoma and psoriasis. It has been demonstrated that epidermal Stat3 activation is required for psoriasis development, since keratinocyte-specific Stat3 activation in a mouse model elicits a psoriasis-like phenotype, which is reversed by inhibition of Stat3 signaling. The aim of this study was whether Stat3 signaling is involved in Raf-MAPK-dependent epidermal hyperplasia. K14-Raf:ER transgenic mice, in which the 4-hydroxytamoxifen (4OHT)-responsive mutant estrogen receptor ligand binding domain-Raf fusion gene is expressed under control of the keratin 14 promoter, were mated with epidermis-specific Stat3 null mice (K5-Cre.Stat3(flox/flox)). K5-Cre.Stat3(flox/flox) mice were used to define the impact of Stat3 deficiency on Raf-induced epidermal hyperplasia. Over-expression of Raf by 4OHT treatment in K14-Raf:ER;K5-Cre.Stat3(flox/flox) mice greatly attenuated the epidermal hyperplasia and dermal cell infiltrates compared with K14-Raf:ER;K5-Cre.Stat3(flox/WT) mice. Also, up-regulation of psoriasis-associated cytokine profiles, including VEGF, was inhibited in the skin from K14-Raf:ER;K5-Cre.Stat3(flox/flox) mice following 4OHT treatment. These results clearly indicate that Raf-MAPK-dependent psoriatic-like epidermal hyperplasia requires Stat3 signaling in keratinocytes. Copyright © 2013 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.

  14. Genomic and Transcriptomic Alterations Associated with STAT3 Activation in Head and Neck Cancer

    PubMed Central

    Peyser, Noah D.; Pendleton, Kelsey; Gooding, William E.; Lui, Vivian W. Y.; Johnson, Daniel E.; Grandis, Jennifer R.

    2016-01-01

    Background Hyperactivation of STAT3 via constitutive phosphorylation of tyrosine 705 (Y705) is common in most human cancers, including head and neck squamous carcinoma (HNSCC). STAT3 is rarely mutated in cancer and the (epi)genetic alterations that lead to STAT3 activation are incompletely understood. Here we used an unbiased approach to identify genomic and epigenomic changes associated with pSTAT3(Y705) expression using data generated by The Cancer Genome Atlas (TCGA). Methods and Findings Mutation, mRNA expression, promoter methylation, and copy number alteration data were extracted from TCGA and examined in the context of pSTAT3(Y705) protein expression. mRNA expression levels of 1279 genes were found to be associated with pSTAT3(705) expression. Association of pSTAT3(Y705) expression with caspase-8 mRNA expression was validated by immunoblot analysis in HNSCC cells. Mutation, promoter hypermethylation, and copy number alteration of any gene were not significantly associated with increased pSTAT3(Y705) protein expression. Conclusions These cumulative results suggest that unbiased approaches may be useful in identifying the molecular underpinnings of oncogenic signaling, including STAT3 activation, in HNSCC. Larger datasets will likely be necessary to elucidate signaling consequences of infrequent alterations. PMID:27855189

  15. Death-associated protein kinase controls STAT3 activity in intestinal epithelial cells.

    PubMed

    Chakilam, Saritha; Gandesiri, Muktheshwar; Rau, Tilman T; Agaimy, Abbas; Vijayalakshmi, Mahadevan; Ivanovska, Jelena; Wirtz, Ralph M; Schulze-Luehrmann, Jan; Benderska, Natalya; Wittkopf, Nadine; Chellappan, Ajithavalli; Ruemmele, Petra; Vieth, Michael; Rave-Fränk, Margret; Christiansen, Hans; Hartmann, Arndt; Neufert, Clemens; Atreya, Raja; Becker, Christoph; Steinberg, Pablo; Schneider-Stock, Regine

    2013-03-01

    The TNF-IL-6-STAT3 pathway plays a crucial role in promoting ulcerative colitis-associated carcinoma (UCC). To date, the negative regulation of STAT3 is poorly understood. Interestingly, intestinal epithelial cells of UCC in comparison to ulcerative colitis show high expression levels of anti-inflammatory death-associated protein kinase (DAPK) and low levels of pSTAT3. Accordingly, epithelial DAPK expression was enhanced in STAT3(IEC-KO) mice. To unravel a possible regulatory mechanism, we used an in vitro TNF-treated intestinal epithelial cell model. We identified a new function of DAPK in suppressing TNF-induced STAT3 activation as DAPK siRNA knockdown and treatment with a DAPK inhibitor potentiated STAT3 activation, IL-6 mRNA expression, and secretion. DAPK attenuated STAT3 activity directly by physical interaction shown in three-dimensional structural modeling. This model suggests that DAPK-induced conformational changes in the STAT3 dimer masked its nuclear localization signal. Alternatively, pharmacological inactivation of STAT3 led to an increase in DAPK mRNA and protein levels. Chromatin immunoprecipitation showed that STAT3 restricted DAPK expression by promoter binding, thereby reinforcing its own activation by inducing IL-6. This novel negative regulation principle might balance TNF-induced inflammation and seems to play an important role in the inflammation-associated transformation process as confirmed in an AOM+DSS colon carcinogenesis mouse model. DAPK as a negative regulator of STAT3 emerges as therapeutic option in the treatment of ulcerative colitis and UCC. Copyright © 2013 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  16. Nuclear protein I{kappa}B-{zeta} inhibits the activity of STAT3

    SciTech Connect

    Wu, Zhihao; Zhang, Xiaoai; Yang, Juntao; Wu, Guangzhou; Zhang, Ying; Yuan, Yanzhi; Jin, Chaozhi; Chang, Zhijie; Wang, Jian; Yang, Xiaoming; He, Fuchu

    2009-09-18

    STAT3 (Signal transducer and activator of transcription 3) is a key transcription factor of the JAK-STAT (Janus kinase/signal transducer and activator of transcription) pathway that regulates cell proliferation and apoptosis. Activation of STAT3 is under tight regulation, and yet the different signaling pathways and the mechanisms that regulate its activity remain to be elucidated. Using a yeast two-hybrid screening, we have identified a nuclear protein I{kappa}B-{zeta} that interacts in a novel way with STAT3. This physical interaction was further confirmed by co-immunoprecipitation assays. The interaction regions were mapped to the coiled-coil domain of STAT3 and the C-terminal of I{kappa}B-{zeta}. Overexpression of I{kappa}B-{zeta} inhibited the transcriptional activity of STAT3. It also suppressed cell growth and induced cell apoptosis in SRC-simulated cells, which is partially mediated by down-regulation of expression of a known STAT3 target gene, MCL1. Our results suggest that I{kappa}B-{zeta} is a negative regulator of STAT3, and demonstrate a novel mechanism in which a component of the NF-{kappa}B signaling pathway inhibits the activation of STAT3.

  17. Activation of the Notch1/STAT3/Twist signaling axis promotes gastric cancer progression.

    PubMed

    Hsu, Kai-Wen; Hsieh, Rong-Hong; Huang, Kuo-Hung; Fen-Yau Li, Anna; Chi, Chin-Wen; Wang, Tzu-Yin; Tseng, Min-Jen; Wu, Kou-Juey; Yeh, Tien-Shun

    2012-08-01

    Gastric carcinoma is one of the most common malignancies and a lethal cancer in the world. Notch signaling and transcription factors STAT3 (signal transducer and activator of transcription 3) and Twist regulate tumor development and are critical regulators of gastric cancer progression. Herein, the relationship among Notch, STAT3 and Twist pathways in the control of gastric cancer progression was studied. We found that Twist and phosphorylated STAT3 levels were promoted by the activated Notch1 receptor in human stomach adenocarcinoma SC-M1, embryonic kidney HEK293 and erythroleukemia K562 cells. Notch1 signaling dramatically induced Twist promoter activity through a C promoter binding factor-1-independent manner and STAT3 phosphorylation. Overexpression of Notch1 receptor intracellular domain (N1IC) enhanced the interaction between nuclear STAT3 and Twist promoter in cells. Gastric cancer progression of SC-M1 cells was promoted by N1IC through STAT3 phosphorylation and Twist expression including colony formation, migration and invasion. STAT3 regulated gastric cancer progression of SC-M1 cells via Twist. N1IC also elevated the progression of other gastric cancer cells such as AGS and KATO III cells through STAT3 and Twist. The N1IC-promoted tumor growth and lung metastasis of SC-M1 cells in mice were suppressed by the STAT3 inhibitor JSI-124 and Twist knockdown. Furthermore, Notch1 and Notch ligand Jagged1 expressions were significantly associated with phosphorylated STAT3 and Twist levels in gastric cancer tissues of patients. Taken together, these results suggest that Notch1/STAT3/Twist signaling axis is involved in progression of human gastric cancer and modulation of this cascade has potential for the targeted combination therapy.

  18. Mouse mammary tumors display Stat3 activation dependent on leukemia inhibitory factor signaling

    PubMed Central

    Quaglino, Ana; Schere-Levy, Carolina; Romorini, Leonardo; Meiss, Roberto P; Kordon, Edith C

    2007-01-01

    Introduction It has been demonstrated that leukemia inhibitory factor (LIF) induces epithelium apoptosis through Stat3 activation during mouse mammary gland involution. In contrast, it has been shown that this transcription factor is commonly activated in breast cancer cells, although what causes this effect remains unknown. Here we have tested the hypothesis that locally produced LIF can be responsible for Stat3 activation in mouse mammary tumors. Methods The studies were performed in different tumorigenic and non-tumorigenic mammary cells. The expression of LIF and LIF receptor was tested by RT-PCR analysis. In tumors, LIF and Stat3 proteins were analyzed by immunohistochemistry, whereas Stat3 and extracellular signal-regulated kinase (ERK)1/2 expression and phosphorylation were studied by Western blot analysis. A LIF-specific blocking antibody was used to determine whether this cytokine was responsible for Stat3 phosphorylation induced by conditioned medium. Specific pharmacological inhibitors (PD98059 and Stat3ip) that affect ERK1/2 and Stat3 activation were used to study their involvement in LIF-induced effects. To analyze cell survival, assays with crystal violet were performed. Results High levels of LIF expression and activated Stat3 were found in mammary tumors growing in vivo and in their primary cultures. We found a single mouse mammary tumor cell line, LM3, that showed low levels of activated Stat3. Incidentally, these cells also showed very little expression of LIF receptor. This suggested that autocrine/paracrine LIF would be responsible for Stat3 activation in mouse mammary tumors. This hypothesis was confirmed by the ability of conditioned medium of mammary tumor primary cultures to induce Stat3 phosphorylation, activity that was prevented by pretreatment with LIF-blocking antibody. Besides, we found that LIF increased tumor cell viability. Interestingly, blocking Stat3 activation enhanced this effect in mammary tumor cells. Conclusion LIF is

  19. Stat3 Activation in Murine Colitis Induced by Enterotoxigenic Bacteroides fragilis

    PubMed Central

    Wick, Elizabeth C.; Rabizadeh, Shervin; Albesiano, Emilia; Wu, XinQun; Wu, Shaoguang; Chan, June; Rhee, Ki-Jong; Ortega, Guillermo; Huso, David L.; Pardoll, Drew; Housseau, Franck; Sears, Cynthia L.

    2014-01-01

    Background Enterotoxigenic Bacteroides fragilis (ETBF), a molecular subclass of the common human commensal, B. fragilis, has been associated with inflammatory bowel disease. ETBF colitis is characterized by the activation of Stat3 and a Th17 immune response in the colonic mucosa. This study was designed to investigate the time course and cellular distribution of Stat3 activation in ETBF-colonized mice. Methods C57BL/6 wild-type, C57BL/6Stat3ΔIEC, or Rag-1 mice were inoculated with saline, nontoxigenic B. fragilis or ETBF. Histologic diagnosis and mucosal Stat activation (immunohistochemistry, Western blot, and/or electrophorectic mobility shift assay) were evaluated over time (6–24 h, 1–7 d, and 1–18 mo after inoculation). Mucosal permeability was evaluated at 16 hours, 1 day, and 3 days. Mucosal immune responses were evaluated at 1 week, and 12 and 18 months. Results ETBF induced rapid-onset colitis that persisted for up to 1 year. Stat3 activation (pStat3) was noted in the mucosal immune cells within 16 hours, with colonic epithelial cell activation evident at 24 hours after inoculation. ETBF-induced increased mucosal permeability was first observed at 24 hours after inoculation, after which the initial immune cell pStat3 activation was noted. Immune cell pStat3 was present in the absence of epithelial pStat3 (C57BL/ 6Stat3ΔIEC). Epithelial pStat3 was present in the absence of T and B cells (Rag-1 mice). pStat3 persisted in the epithelial and immune cells for 1 year, characterized by isolated pStat3-positive cell clusters, with varying intensity distributed through the proximal and distal colon. Similarly, mucosal Th17 immune responses persisted for up to 1 year. Loss of fecal ETBF colonization was associated with the loss of mucosal pStat3 and Th17 immune responses. Conclusions ETBF rapidly induces immune cell pStat3, which is independent of epithelial pStat3. This occurs before ETBF-induced mucosal permeability, suggesting that ETBF, likely through B

  20. At High Levels, Constitutively Activated STAT3 Induces Apoptosis of Chronic Lymphocytic Leukemia Cells.

    PubMed

    Rozovski, Uri; Harris, David M; Li, Ping; Liu, Zhiming; Wu, Ji Yuan; Grgurevic, Srdana; Faderl, Stefan; Ferrajoli, Alessandra; Wierda, William G; Martinez, Matthew; Verstovsek, Srdan; Keating, Michael J; Estrov, Zeev

    2016-05-15

    In chronic lymphocytic leukemia (CLL), the increment in PBLs is slower than the expected increment calculated from the cells' proliferation rate, suggesting that cellular proliferation and apoptosis are concurrent. Exploring this phenomenon, we found overexpression of caspase-3, higher cleaved poly (ADP-ribose) polymerase levels (p < 0.007), and a higher apoptosis rate in cells from patients with high counts compared with cells from patients with low counts. Although we previously found that STAT3 protects CLL cells from apoptosis, STAT3 levels were significantly higher in cells from patients with high counts than in cells from patients with low counts. Furthermore, overexpression of STAT3 did not protect the cells. Rather, it upregulated caspase-3 and induced apoptosis. Remarkably, putative STAT3 binding sites were identified in the caspase-3 promoter, and a luciferase assay, chromatin immunoprecipitation, and an EMSA revealed that STAT3 activated caspase-3 However, caspase-3 levels increased only when STAT3 levels were sufficiently high. Using chromatin immunoprecipitation and EMSA, we found that STAT3 binds with low affinity to the caspase-3 promoter, suggesting that at high levels, STAT3 activates proapoptotic mechanisms and induces apoptosis in CLL cells.

  1. High-Content pSTAT3/1 Imaging Assays to Screen for Selective Inhibitors of STAT3 Pathway Activation in Head and Neck Cancer Cell Lines

    PubMed Central

    Sen, Malabika; Hua, Yun; Camarco, Daniel; Shun, Tong Ying; Lazo, John S.; Grandis, Jennifer R.

    2014-01-01

    Abstract The oncogenic transcription factor signal transducer and activator of transcription 3 (STAT3) is hyperactivated in most cancers and represents a plausible therapeutic target. In the absence of STAT3-selective small-molecule inhibitors, we sought to develop pSTAT3/1 high-content imaging (HCS) assays to screen for selective inhibitors of STAT3 pathway activation in head and neck squamous cell carcinomas (HNSCC) tumor cell lines. Based on the expression of the interleukin-6 (IL-6)Rα and gp130 subunits of the IL-6 receptor complex and STAT3, we selected the Cal33 HNSCC cell line as our model. After developing image acquisition and analysis procedures, we rigorously investigated the cytokine activation responses to optimize the dynamic ranges of both assays and demonstrated that the pan-Janus kinase inhibitor pyridone 6 nonselectively inhibited pSTAT3 and pSTAT1 activation with 50% inhibition concentrations of 7.19±4.08 and 16.38±8.45 nM, respectively. The optimized pSTAT3 HCS assay performed very well in a pilot screen of 1,726 compounds from the Library of Pharmacologically Active Compounds and the National Institutes of Health clinical collection sets, and we identified 51 inhibitors of IL-6-induced pSTAT3 activation. However, only three of the primary HCS actives selectively inhibited STAT3 compared with STAT1. Our follow-up studies indicated that the nonselective inhibition of cytokine induced pSTAT3 and pSTAT1 activation by G-alpha stimulatory subunit-coupled G-protein-coupled receptor agonists, and forskolin was likely due to cyclic adenosine monophosphate-mediated up-regulation of suppressors of cytokine signaling 3. Azelastine, an H1 receptor antagonist approved for the treatment of seasonal allergic rhinitis, nonallergic vasomotor rhinitis, and ocular conjunctivitis, was subsequently confirmed as a selective inhibitor of IL-6-induced pSTAT3 activation that also reduced the growth of HNSCC cell lines. These data illustrate the power of a chemical

  2. Insulin stimulates the serine phosphorylation of the signal transducer and activator of transcription (STAT3) isoform.

    PubMed

    Ceresa, B P; Pessin, J E

    1996-05-24

    Insulin stimulation of Chinese hamster ovary cells expressing the human insulin receptor and differentiated 3T3L1 adipocytes resulted in a time-dependent reduction in the SDS-polyacrylamide gel electrophoretic mobility of STAT3. The decreased STAT3 mobility initially occurred by 2 min and was quantitative by 5 min. In addition, the change in STAT3 mobility was concentration-dependent and was detectable at 0.3 nm insulin with maximal effect between 1 and 3 nm. Although both these cell types also express the STAT1 alpha, STAT1 beta, STAT5, and STAT6 isoforms, only STAT3 was observed to undergo an insulin-dependent reduction in mobility. Immuno-precipitation of STAT1 and STAT3 from 32P-labeled cells demonstrated that only STAT3 was phosphorylated in response to insulin whereas phosphoamino acid analysis indicated that this phosphorylation event occurred exclusively on serine residues. Furthermore, treatment of cell extracts with alkaline phosphatase reversed the insulin-stimulated decrease in STAT3 mobility. Together, these data demonstrate that insulin is a specific activator of STAT3 serine phosphorylation without affecting the other STAT isoforms.

  3. Stat3 links activated keratinocytes and immunocytes required for development of psoriasis in a novel transgenic mouse model.

    PubMed

    Sano, Shigetoshi; Chan, Keith Syson; Carbajal, Steve; Clifford, John; Peavey, Mary; Kiguchi, Kaoru; Itami, Satoshi; Nickoloff, Brian J; DiGiovanni, John

    2005-01-01

    Here we report that epidermal keratinocytes in psoriatic lesions are characterized by activated Stat3. Transgenic mice with keratinocytes expressing a constitutively active Stat3 (K5.Stat3C mice) develop a skin phenotype either spontaneously, or in response to wounding, that closely resembles psoriasis. Keratinocytes from K5.Stat3C mice show upregulation of several molecules linked to the pathogenesis of psoriasis. In addition, the development of psoriatic lesions in K5.Stat3C mice requires cooperation between Stat3 activation in keratinocytes and activated T cells. Finally, abrogation of Stat3 function by a decoy oligonucleotide inhibits the onset and reverses established psoriatic lesions in K5.Stat3C mice. Thus, targeting Stat3 may be potentially therapeutic in the treatment of psoriasis.

  4. Maximal activation of transcription by Stat1 and Stat3 requires both tyrosine and serine phosphorylation.

    PubMed

    Wen, Z; Zhong, Z; Darnell, J E

    1995-07-28

    Stat1 and Stat3 are latent transcriptional factors activated initially through phosphorylation on single tyrosine residues induced by cytokine and growth factor occupation of cell surface receptors. Here we show that phosphorylation on a single serine (residue 727) in each protein is also required for maximal transcriptional activity. Both cytokines and growth factors are capable of inducing the serine phosphorylation of Stat1 and Stat3. These experiments show that gene activation by Stat1 and Stat3, which obligatorily require tyrosine phosphorylation to become active, also depends for maximal activation on one or more of the many serine kinases.

  5. Activation of Stat3 transcription factor by Herpesvirus saimiri STP-A oncoprotein.

    PubMed

    Chung, Young-Hwa; Cho, Nam-hyuk; Garcia, Maria Ines; Lee, Sun-Hwa; Feng, Pinghui; Jung, Jae U

    2004-06-01

    The saimiri transforming protein (STP) oncogene of Herpesvirus saimiri subgroup A strain 11 (STP-A11) is not required for viral replication but is required for lymphoid cell immortalization in culture and lymphoma induction in primates. We previously showed that STP-A11 interacts with cellular Src kinase through its SH2 binding motif and that this interaction elicits Src signal transduction. Here we demonstrate that STP-A11 interacts with signal transducer and activator of transcription 3 (Stat3) independently of Src association and that the amino-terminal short proline-rich motif of STP-A11 and the central linker region of Stat3 are necessary for their interaction. STP-A11 formed a triple complex with Src kinase and Stat3 where Src kinase phosphorylated Stat3, resulting in the nuclear localization and transcriptional activation of Stat3. Consequently, the constitutively active Stat3 induced by STP-A11 elicited cellular signal transduction, which ultimately induced cell survival and proliferation upon serum deprivation. Furthermore, this activity was strongly correlated with the induction of Fos, cyclin D1, and Bcl-XL expression. These results demonstrate that STP-A11 independently targets two important cellular signaling molecules, Src and Stat3, and that these proteins cooperate efficiently to induce STP-A11-mediated transformation.

  6. Endothelial STAT3 Activation Increases Vascular Leakage Through Downregulating Tight Junction Proteins: Implications for Diabetic Retinopathy.

    PubMed

    Yun, Jang-Hyuk; Park, Sung Wook; Kim, Kyung-Jin; Bae, Jong-Sup; Lee, Eun Hui; Paek, Sun Ha; Kim, Seung U; Ye, Sangkyu; Kim, Jeong-Hun; Cho, Chung-Hyun

    2017-05-01

    Vascular inflammation is characteristic feature of diabetic retinopathy. In diabetic retina, a variety of the pro-inflammatory cytokines are elevated and involved in endothelial dysfunction. STAT3 transcription factor has been implicated in mediating cytokine signaling during vascular inflammation. However, whether and how STAT3 is involved in the direct regulation of the endothelial permeability is currently undefined. Our studies revealed that IL-6-induced STAT3 activation increases retinal endothelial permeability and vascular leakage in retinas of mice through the reduced expression of the tight junction proteins ZO-1 and occludin. In a co-culture model with microglia and endothelial cells under a high glucose condition, the microglia-derived IL-6 induced STAT3 activation in the retinal endothelial cells, leading to increasing endothelial permeability. In addition, IL-6-induced STAT3 activation was independent of ROS generation in the retinal endothelial cells. Moreover, we demonstrated that STAT3 activation downregulates the ZO-1 and occludin levels and increases the endothelial permeability through the induction of VEGF production in retinal endothelial cells. These results suggest the potential importance of IL-6/STAT3 signaling in regulating endothelial permeability and provide a therapeutic target to prevent the pathology of diabetic retinopathy. J. Cell. Physiol. 232: 1123-1134, 2017. © 2016 Wiley Periodicals, Inc.

  7. JAB1 regulates unphosphorylated STAT3 DNA-binding activity through protein–protein interaction in human colon cancer cells

    SciTech Connect

    Nishimoto, Arata; Kugimiya, Naruji; Hosoyama, Toru; Enoki, Tadahiko; Li, Tao-Sheng; Hamano, Kimikazu

    2013-08-30

    Highlights: •JAB1 interacted with unphosphorylated STAT3 in the nucleus. •JAB1 knockdown tended to increase nuclear STAT3 expression. •JAB1 knockdown significantly decreased unphosphorylated STAT3 DNA-binding activity. •JAB1 knockdown significantly decreased MDR1, NANOG, and VEGF expressions. •Nuclear JAB1, but not nuclear STAT3, correlated with STAT3 DNA-binding activity. -- Abstract: Recent studies have revealed that unphosphorylated STAT3 forms a dimer, translocates to the nucleus, binds to the STAT3 binding site, and activates the transcription of STAT3 target genes, thereby playing an important role in oncogenesis in addition to phosphorylated STAT3. Among signaling steps of unphosphorylated STAT3, nuclear translocation and target DNA-binding are the critical steps for its activation. Therefore, elucidating the regulatory mechanism of these signaling steps of unphosphorylated STAT3 is a potential step in the discovery of a novel cancer drug. However, the mechanism of unphosphorylated STAT3 binding to the promoter of target genes remains unclear. In this study, we focused on Jun activation domain-binding protein 1 (JAB1) as a candidate protein that regulates unphosphorylated STAT3 DNA-binding activity. Initially, we observed that both unphosphorylated STAT3 and JAB1 existed in the nucleus of human colon cancer cell line COLO205 at the basal state (no cytokine stimulation). On the other hand, phosphorylated STAT3 did not exist in the nucleus of COLO205 cells at the basal state. Immunoprecipitation using nuclear extract of COLO205 cells revealed that JAB1 interacted with unphosphorylated STAT3. To investigate the effect of JAB1 on unphosphorylated STAT3 activity, RNAi studies were performed. Although JAB1 knockdown tended to increase nuclear STAT3 expression, it significantly decreased unphosphorylated STAT3 DNA-binding activity. Subsequently, JAB1 knockdown significantly decreased the expression levels of MDR1, NANOG, and VEGF, which are STAT3 target

  8. Lactococcus lactis KR-050L inhibit IL-6/STAT3 activation.

    PubMed

    Hwang, J T; Jang, H-J; Kim, J H; Park, C S; Kim, Y; Lim, C-H; Lee, S W; Rho, M-C

    2017-05-01

    The purpose of this study was to investigate IL-6/STAT3 inhibitory activity using lactic acid bacteria (LABs) isolated from Gajuknamu kimchi. Six LABs were isolated from Gajuknamu kimchi and identified through 16S rRNA sequencing. Among them, the culture broth of Lactococcus lactis KR-050L inhibited IL-6-induced STAT3 luciferase activity. Fifteen compounds were isolated from the EtOAc extract of culture broth though column chromatography and preparative high-performance liquid chromatography, and they were identified as 2,5-diketopipperazine structures by spectroscopic analyses (MS, (1) H- and (13) C-NMR). They also showed inhibitory activities on IL-6-induced STAT3 activation, and showed the different in activity according to the presence of a phenylalanine residue, hydroxyl groups and isometric structure. The six new LABs isolated from Gajuknamu kimchi, and Lc. lactis KR-050L was selected as candidate IL-6/STAT3 inhibitors. The activity levels of 15 2,5-DKPs isolated from Lc. lactis KR-050L were verified. This study constitutes the first attempt to isolate various LABs from Gajuknamu kimchi and to discover IL-6/STAT3 inhibitors in the EtOAc extract of Lc. lactis KR-050L culture broth. Moreover, our data provide useful biochemical information regarding the commercialization of Lc. lactis isolated from Gajuknamu kimchi as an approach to use functional foods for the treatment of various diseases via IL-6/STAT3 activation. © 2017 The Society for Applied Microbiology.

  9. Lung Adenocarcinomas and Lung Cancer Cell Lines Show Association of MMP-1 Expression With STAT3 Activation1

    PubMed Central

    Schütz, Alexander; Röser, Katrin; Klitzsch, Jana; Lieder, Franziska; Aberger, Fritz; Gruber, Wolfgang; Mueller, Kristina M.; Pupyshev, Alexander; Moriggl, Richard; Friedrich, Karlheinz

    2015-01-01

    Signal transducer and activator of transcription 3 (STAT3) is constitutively activated in the majority of lung cancer. This study aims at defining connections between STAT3 function and the malignant properties of non–small cell lung carcinoma (NSCLC) cells. To address possible mechanisms by which STAT3 influences invasiveness, the expression of matrix metalloproteinase-1 (MMP-1) was analyzed and correlated with the STAT3 activity status. Studies on both surgical biopsies and on lung cancer cell lines revealed a coincidence of STAT3 activation and strong expression of MMP-1. MMP-1 and tyrosine-phosphorylated activated STAT3 were found co-localized in cancer tissues, most pronounced in tumor fronts, and in particular in adenocarcinomas. STAT3 activity was constitutive, although to different degrees, in the lung cancer cell lines investigated. Three cell lines (BEN, KNS62, and A549) were identified in which STAT3 activitation was inducible by Interleukin-6 (IL-6). In A549 cells, STAT3 activity enhanced the level of MMP-1 mRNA and stimulated transcription from the MMP-1 promoter in IL-6–stimulated A549 cells. STAT3 specificity of this effect was confirmed by STAT3 knockdown through RNA interference. Our results link aberrant activity of STAT3 in lung cancer cells to malignant tumor progression through up-regulation of expression of invasiveness-associated MMPs. PMID:25926075

  10. STAT3 activation in circulating monocytes contributes to neovascular age-related macular degeneration

    PubMed Central

    Chen, Mei; Lechner, Judith; Zhao, Jiawu; Toth, Levente; Hogg, Ruth; Silvestri, Giuliana; Kissenpfennig, Adrien; Chakravarthy, Usha; Xu, Heping

    2016-01-01

    Infiltrating macrophages are critically involved in pathogenic angiogenesis such as neovascular age-related macular degeneration (nAMD). Macrophages originate from circulating monocytes and three subtypes of monocyte exist in humans: classical (CD14+CD16-), non-classical (CD14-CD16+) and intermediate (CD14+CD16+) monocytes. The aim of this study was to investigate the role of circulating monocyte in neovascular age-related macular degeneration (nAMD). Flow cytometry analysis showed that the intermediate monocytes from nAMD patients expressed higher levels of CX3CR1 and HLA-DR compared to those from controls. Monocytes from nAMD patients expressed higher levels of phosphorylated Signal Transducer and Activator of Transcription 3 (pSTAT3), and produced higher amount of VEGF. In the mouse model of choroidal neovascularization (CNV), pSTAT3 expression was increased in the retina and RPE/choroid, and 49.24% of infiltrating macrophages express pSTAT3. Genetic deletion of the Suppressor of Cytokine Signalling 3 (SOCS3) in myeloid cells in the LysM-Cre+/-:SOCS3fl/fl mice resulted in spontaneous STAT3 activation and accelerated CNV formation. Inhibition of STAT3 activation using a small peptide LLL12 suppressed laser-induced CNV. Our results suggest that monocytes, in particular the intermediate subset of monocytes are activated in nAMD patients. STAT3 activation in circulating monocytes may contribute to the development of choroidal neovascularisation in AMD. PMID:27009107

  11. The Role of Stat3 Activation in Androgen Receptor Signaling and Prostate Cancer

    DTIC Science & Technology

    2006-07-01

    Naruto, M., and Kishimoto, T. Molecular cloning of APRF, a novel IFN-stimulated gene factor 3 p91- related transcrip- tion factor involved in the gp130...independent tumor cells. The molecular mechanism characterizing prostate cancer progression from androgen-dependence to androgenindependence is incompletely...consequence of Stat3 activation in prostate cancer cell growth and to determine the molecular basis of Stat3 interactions with androgen receptor signaling

  12. Inhibition of Stat3 activation in the endometrium prevents implantation: a nonsteroidal approach to contraception.

    PubMed

    Catalano, Rob D; Johnson, Martin H; Campbell, Elizabeth A; Charnock-Jones, D Stephen; Smith, Stephen K; Sharkey, Andrew M

    2005-06-14

    Activation of the receptors for leukemia inhibitory factor (LIF) and IL-11 is essential for embryo attachment and decidualization in mice. Both receptors induce activation of the Stat family of signal transducers via the Jak/Stat pathway. Here, we aimed to establish whether activation of Stat3 in maternal endometrium is essential for successful implantation. Functional blockade of Stat3 before implantation, by injection into the uterine lumen of a cell-permeable Stat3 peptide inhibitor, reduced embryo implantation specifically by 70% (P < 0.001). Stat3 is phosphorylated in the luminal epithelium (LE) in response to LIF, and this phosphorylation was significantly reduced both in vitro and in vivo by the Stat3 inhibitor. The inhibitor also blocked induction by LIF of several LIF-regulated genes in the LE including Irg1, which has been shown previously to be essential for implantation. Successful implantation is therefore dependent on phosphorylation and activation of Stat3 in the endometrium before implantation. This finding provides a target for contraceptive development, based on selective blockade of signal transduction pathways essential for implantation. This study demonstrates that cell-permeable peptide inhibitors can be used effectively to target intracellular signaling pathways in the uterine LE.

  13. STATs profiling reveals predominantly-activated STAT3 in cholangiocarcinoma genesis and progression.

    PubMed

    Dokduang, Hasaya; Techasen, Anchalee; Namwat, Nisana; Khuntikeo, Narong; Pairojkul, Chawalit; Murakami, Yoshinori; Loilome, Watcharin; Yongvanit, Puangrat

    2014-10-01

    We investigated the aberrant expression of the STAT family in humans and liver fluke (Opisthorchis viverrini, Ov)-induced hamster cholangiocarcinoma (CCA) tissues. The expression and phosphorylation of STAT1, STAT2, STAT3, STAT4, STAT5a, STAT5b and STAT6 in human hamster CCA tissues were immunohistochemistry-profiled. Localizations of STAT5 in macrophages and lipopolysaccharide (LPS)-induced macrophage-conditioned media mediated STAT3 activation in CCA cells were demonstrated. The expressions of STAT 1-4 and 6 were detected in the cytoplasm of hyperplastic bile ducts and tumor cells, whereas STAT5a and STAT5b were observed in macrophages and connective tissues surrounding tumor, respectively. The expressions of STAT3 and STAT5b were significantly observed in tumors with a poorer histological differentiation. STAT3 expression was significantly associated with shorter survival of CCA patients and was predominately activated in CCA cell lines. In the CCA-hamsters, STATs expression was gradually increased along the carcinogenesis, especially at 30 days post-infection in which the inflammatory response was markedly observed, showing the correlation between the inflammation and STATs activation. Moreover, LPS-induced macrophage-conditioned media could mediate STAT3 activation in CCA cells. STAT3 is the major STAT, which plays roles in the inflammation that contributes to CCA carcinogenesis and progression and may serve as a marker for a poor prognosis of CCA. © 2014 Japanese Society of Hepato-Biliary-Pancreatic Surgery.

  14. STAT3-Activated GM-CSFRα Translocates to the Nucleus and Protects CLL Cells from Apoptosis

    PubMed Central

    Li, Ping; Harris, David; Liu, Zhiming; Rozovski, Uri; Ferrajoli, Alessandra; Wang, Yongtao; Bueso-Ramos, Carlos; Hazan-Halevy, Inbal; Grgurevic, Srdana; Wierda, William; Burger, Jan; O'Brien, Susan; Faderl, Stefan; Keating, Michael; Estrov, Zeev

    2014-01-01

    Here it was determined that Chronic Lymphocytic Leukemia (CLL) cells express the α-subunit but not the β-subunit of the granulocyte-macrophage colony-stimulating factor receptor (GM-CSFR/CSF3R). GM-CSFRα was detected on the surface, in the cytosol, and the nucleus of CLL cells via confocal microscopy, cell fractionation, and GM-CSFRα antibody epitope mapping. Because STAT3 is frequently activated in CLL and the GM-CSFRα promoter harbors putative STAT3 consensus binding sites, MM1 cells were transfected with truncated forms of the GM-CSFRα promoter, then stimulated with IL-6 to activate STAT3 to identify STAT3 binding sites. Chromatin immunoprecipitation (ChIP) and an electoromobility shift assay (EMSA) confirmed STAT3 occupancy to those promoter regions in both IL-6 stimulated MM1 and CLL cells. Transfection of MM1 cells with STAT3 siRNA or CLL cells with STAT3 shRNA significantly down-regulated GM-CSFRα mRNA and protein levels. RNA transcripts, involved in regulating cell-survival pathways, and the proteins KAP1 (TRIM28) and ISG15 co-immunoprecipitated with GM-CSFRα. GM-CSFRα-bound KAP1 enhanced the transcriptional activity of STAT3, whereas ISG15 inhibited the NF-κB pathway. Nevertheless, overexpression of GM-CSFRα protected MM1 cells from dexamethasone-induced apoptosis, and GM-CSFRα knockdown induced apoptosis in CLL cells, suggesting that GM-CSFRα provides a ligand-independent survival advantage. PMID:24836891

  15. Prevention of Hypovolemic Circulatory Collapse by IL-6 Activated Stat3

    PubMed Central

    Tsimelzon, Anna I.; Mastrangelo, Mary-Ann A.; Hilsenbeck, Susan G.; Poli, Valeria; Tweardy, David J.

    2008-01-01

    Half of trauma deaths are attributable to hypovolemic circulatory collapse (HCC). We established a model of HCC in rats involving minor trauma plus severe hemorrhagic shock (HS). HCC in this model was accompanied by a 50% reduction in peak acceleration of aortic blood flow and cardiomyocyte apoptosis. HCC and apoptosis increased with increasing duration of hypotension. Apoptosis required resuscitation, which provided an opportunity to intervene therapeutically. Administration of IL-6 completely reversed HCC, prevented cardiac dysfunction and cardiomyocyte apoptosis, reduced mortality 5-fold and activated intracardiac signal transducer and activator of transcription (STAT) 3. Pre-treatment of rats with a selective inhibitor of Stat3, T40214, reduced the IL-6-mediated increase in cardiac Stat3 activity, blocked successful resuscitation by IL-6 and reversed IL-6-mediated protection from cardiac apoptosis. The hearts of mice deficient in the naturally occurring dominant negative isoform of Stat3, Stat3β, were completely resistant to HS-induced apoptosis. Microarray analysis of hearts focusing on apoptosis related genes revealed that expression of 29% of apoptosis related genes was altered in HS vs. sham rats. IL-6 treatment normalized the expression of these genes, while T40214 pretreatment prevented IL-6-mediated normalization. Thus, cardiac dysfunction, cardiomyocyte apoptosis and induction of apoptosis pathway genes are important components of HCC; IL-6 administration prevented HCC by blocking cardiomyocyte apoptosis and induction of apoptosis pathway genes via Stat3 and warrants further study as a resuscitation adjuvant for prevention of HCC and death in trauma patients. PMID:18270592

  16. ERp57 modulates STAT3 activity in radioresistant laryngeal cancer cells and serves as a prognostic marker for laryngeal cancer.

    PubMed

    Choe, Min Ho; Min, Joong Won; Jeon, Hong Bae; Cho, Dong-Hyung; Oh, Jeong Su; Lee, Hyun Gyu; Hwang, Sang-Gu; An, Sungkwan; Han, Young-Hoon; Kim, Jae-Sung

    2015-02-20

    Although targeting radioresistant tumor cells is essential for enhancing the efficacy of radiotherapy, the signals activated in resistant tumors are still unclear. This study shows that ERp57 contributes to radioresistance of laryngeal cancer by activating STAT3. Increased ERp57 was associated with the radioresistant phenotype of laryngeal cancer cells. Interestingly, increased interaction between ERp57 and STAT3 was observed in radioresistant cells, compared to the control cells. This physical complex is required for the activation of STAT3 in the radioresistant cells. Among STAT3-regulatory genes, Mcl-1 was predominantly regulated by ERp57. Inhibition of STAT3 activity with a chemical inhibitor or siRNA-mediated depletion of Mcl-1 sensitized radioresistant cells to irradiation, suggesting that the ERp57-STAT3-Mcl-1 axis regulates radioresistance of laryngeal cancer cells. Furthermore, we observed a positive correlation between ERp57 and phosphorylated STAT3 or Mcl-1 and in vivo interactions between ERp57 and STAT3 in human laryngeal cancer. Importantly, we also found that increased ERp57-STAT3 complex was associated with poor prognosis in human laryngeal cancer, indicating the prognostic role of ERp57-STAT3 regulation. Overall, our data suggest that ERp57-STAT3 regulation functions in radioresistance of laryngeal cancer, and targeting the ERp57-STAT3 pathway might be important for enhancing the efficacy of radiotherapy in human laryngeal cancer.

  17. Ultraviolet light induces Stat3 activation in human keratinocytes and fibroblasts through reactive oxygen species and DNA damage.

    PubMed

    Bito, Toshinori; Sumita, Naoko; Masaki, Taro; Shirakawa, Toshiro; Ueda, Masato; Yoshiki, Ryutaro; Tokura, Yoshiki; Nishigori, Chikako

    2010-07-01

    Stat3 is activated by the outer stressors, such as ultraviolet (UV) exposure. In this study, we investigated the Stat3 response to UV irradiation in human epidermal keratinocytes and dermal fibroblasts. Results indicated that UVB and UVC differentially activate Stat3 in these cells. The UV-induced Stat3 activation was mediated by both reactive oxygen species (ROS) and DNA damage, and the dominancy of ROS and DNA damage to activate Stat3 depended on the wavelength of UV. By using fibroblasts from a patient with xeroderma pigmentosum A (XP-A) and those transfected with human XPA gene, we found that UVB activates Stat3 via both ROS and DNA damage, while UVC does so mainly via DNA damage. The present data suggest that Stat3 activation in UV-exposed human skin is one of the initial events where DNA damage and ROS are involved.

  18. Prevention of Trauma and Hemorrhagic Shock-Mediated Liver Apoptosis by Activation of Stat3α

    PubMed Central

    Moran, Ana; Akcan Arikan, Ayse; Mastrangelo, Mary-Ann A.; Wu, Yong; Yu, Bi; Poli, Valeria; Tweardy, David J.

    2008-01-01

    Trauma is a major cause of mortality in the United States. Death among those surviving the initial insult is caused by multiple organ failure (MOF) with the liver among the organs most frequently affected. We previously demonstrated in rodents that trauma complicated by hemorrhagic shock (trauma/HS) results in liver injury that can be prevented by IL-6 administration at the start of resuscitation; however, the contribution of the severity of HS to the extent of liver injury, whether or not resuscitation is required and the mechanism for the IL-6 protective effect have not been reported. In the experiments reported here, we demonstrated that the extent of liver apoptosis induced by trauma/HS depends on the duration of hypotension and requires resuscitation. We established that IL-6 administration at the start of resuscitation is capable of completely reversing liver apoptosis and is associated with increased Stat3 activation. Microarray analysis of the livers showed that the main effect of IL-6 was to normalize the trauma/HS-induced apoptosis transcriptome. Pharmacological inhibition of Stat3 activity within the liver blocked the ability of IL-6 to prevent liver apoptosis and to normalize the trauma/HS- induced liver apoptosis transcriptome. Genetic deletion of a Stat3β, a naturally occurring, dominant-negative isoform of the Stat3, attenuated trauma/HS-induced liver apoptosis, confirming a role for Stat3, especially Stat3α, in preventing trauma/HS-mediated liver apoptosis. Thus, trauma/HS-induced liver apoptosis depends on the duration of hypotension and requires resuscitation. IL-6 administration at the start of resuscitation reverses HS-induced liver apoptosis, through activation of Stat3α, which normalizes the trauma/HS-induced liver apoptosis transcriptome. PMID:18997875

  19. An Activating Mutation in STAT3 Results in Neonatal Diabetes Through Reduced Insulin Synthesis.

    PubMed

    Velayos, Teresa; Martínez, Rosa; Alonso, Milagros; Garcia-Etxebarria, Koldo; Aguayo, Anibal; Camarero, Cristina; Urrutia, Inés; Martínez de LaPiscina, Idoia; Barrio, Raquel; Santin, Izortze; Castaño, Luis

    2017-04-01

    Neonatal diabetes mellitus (NDM) is a rare form of diabetes diagnosed within the first 6 months of life. Genetic studies have allowed the identification of several genes linked to the development of NDM; however, genetic causes for ∼20% of the cases remain to be clarified. Most cases of NDM involve isolated diabetes, but sometimes NDM appears in association with other pathological conditions, including autoimmune diseases. Recent reports have linked activating mutations in STAT3 with early-onset autoimmune disorders that include diabetes of autoimmune origin, but the functional impact of STAT3-activating mutations have not been characterized at the pancreatic β-cell level. By using whole-exome sequencing, we identified a novel missense mutation in the binding domain of the STAT3 protein in a patient with NDM. The functional analyses showed that the mutation results in an aberrant activation of STAT3, leading to deleterious downstream effects in pancreatic β-cells. The identified mutation leads to hyperinhibition of the transcription factor Isl-1 and, consequently, to a decrease in insulin expression. These findings represent the first functional indication of a direct link between an NDM-linked activating mutation in STAT3 and pancreatic β-cell dysfunction.

  20. MicroRNA-424 impairs ubiquitination to activate STAT3 and promote prostate tumor progression

    PubMed Central

    Dallavalle, Cecilia; Civenni, Gianluca; Merulla, Jessica; Ostano, Paola; Mello-Grand, Maurizia; Rossi, Simona; Losa, Marco; D’Ambrosio, Gioacchino; Sessa, Fausto; Thalmann, George N.; Zitella, Andrea; Chiorino, Giovanna; Catapano, Carlo V.

    2016-01-01

    Mutations and deletions in components of ubiquitin ligase complexes that lead to alterations in protein turnover are important mechanisms in driving tumorigenesis. Here we describe an alternative mechanism involving upregulation of the microRNA miR-424 that leads to impaired ubiquitination and degradation of oncogenic transcription factors in prostate cancers. We found that miR-424 targets the E3 ubiquitin ligase COP1 and identified STAT3 as a key substrate of COP1 in promoting tumorigenic and cancer stem-like properties in prostate epithelial cells. Altered protein turnover due to impaired COP1 function led to accumulation and enhanced basal and cytokine-induced activity of STAT3. We further determined that loss of the ETS factor ESE3/EHF is the initial event that triggers the deregulation of the miR-424/COP1/STAT3 axis. COP1 silencing and STAT3 activation were effectively reverted by blocking of miR-424, suggesting a possible strategy to attack this key node of tumorigenesis in ESE3/EHF–deficient tumors. These results establish miR-424 as an oncogenic effector linked to noncanonical activation of STAT3 and as a potential therapeutic target. PMID:27820701

  1. Stat3 Activation in Urothelial Stem Cells Leads to Direct Progression to Invasive Bladder Cancer

    PubMed Central

    Ho, Philip Levy; Lay, Erica Julianne; Jian, Weiguo; Parra, Diana; Chan, Keith Syson

    2012-01-01

    Two subtypes of human bladder cancer, noninvasive papillary and muscle-invasive cancer, develop through independent pathologic and molecular pathways. Human invasive bladder cancer frequently develops without prior clinical evidence of a noninvasive tumor stage. However, an animal model that recapitulates this unique clinical progression of invasive bladder cancer has not yet been developed. In this study, we created a novel transgenic mouse model of invasive bladder cancer by targeting an active dimerized form of Stat3 to the basal cells of bladder epithelium. When exposed to the carcinogen nitrosamine, Stat3-transgenic mice developed invasive cancer directly from carcinoma in situ (CIS), bypassing the noninvasive papillary tumor stage. Remarkably, invasive bladder cancer driven by active Stat3 was predominantly composed of stem cells, which were characterized by cytokeratin 14 (CK14) staining and enhanced tumor sphere-forming ability. Active Stat3 was also shown to localize to the nucleus of human invasive bladder cancers that were primarily composed of CK14+ stem cells. Together, our findings show that Stat3-induced stem cell expansion plays a critical role in the unique clinical progression of invasive bladder cancer through the CIS pathway. PMID:22532166

  2. MicroRNA-424 impairs ubiquitination to activate STAT3 and promote prostate tumor progression.

    PubMed

    Dallavalle, Cecilia; Albino, Domenico; Civenni, Gianluca; Merulla, Jessica; Ostano, Paola; Mello-Grand, Maurizia; Rossi, Simona; Losa, Marco; D'Ambrosio, Gioacchino; Sessa, Fausto; Thalmann, George N; Garcia-Escudero, Ramon; Zitella, Andrea; Chiorino, Giovanna; Catapano, Carlo V; Carbone, Giuseppina M

    2016-12-01

    Mutations and deletions in components of ubiquitin ligase complexes that lead to alterations in protein turnover are important mechanisms in driving tumorigenesis. Here we describe an alternative mechanism involving upregulation of the microRNA miR-424 that leads to impaired ubiquitination and degradation of oncogenic transcription factors in prostate cancers. We found that miR-424 targets the E3 ubiquitin ligase COP1 and identified STAT3 as a key substrate of COP1 in promoting tumorigenic and cancer stem-like properties in prostate epithelial cells. Altered protein turnover due to impaired COP1 function led to accumulation and enhanced basal and cytokine-induced activity of STAT3. We further determined that loss of the ETS factor ESE3/EHF is the initial event that triggers the deregulation of the miR-424/COP1/STAT3 axis. COP1 silencing and STAT3 activation were effectively reverted by blocking of miR-424, suggesting a possible strategy to attack this key node of tumorigenesis in ESE3/EHF-deficient tumors. These results establish miR-424 as an oncogenic effector linked to noncanonical activation of STAT3 and as a potential therapeutic target.

  3. Signal transducer and activator of transcription 3 (STAT3) homologue in turbot (Scophthalmus maximus): molecular characterization and expression analysis.

    PubMed

    Wang, Na; Yang, Chang-Geng; Sun, Zhong-Zhi; Wang, Xian-Li; Chen, Song-Lin

    2011-01-01

    Signal transducer and activator of transcription 3 (STAT3) acts as an important mediator in multiple biological processes induced by different cytokines. So far, little information is available in fish STAT3. In this study, turbot (Scophthalmus maximus) STAT3 gene was cloned and characterized for the first time. The turbot STAT3 full-length cDNA consists of 2355 nucleotides encoding a polypeptide of 784 amino acids with four conserved domains including STAT_int, STAT_alpha, STAT_bind and SH2 domain. The phylogenetic tree showed that turbot STAT3 shared the closest relationship with mandarin fish (Siniperca chuatsi) STAT3. The autoactivation experiment in yeast proved that turbot STAT3 was a strong transcription factor. The quantitative RT-PCR experiment indicated that Stat3 mRNA was expressed in widespread tissues with the highest expression levels in the liver. And the further expression patterns analysis revealed that turbot Stat3 expression levels were increased in liver, spleen, kidney of fish infected with Vibrio anguillarum and liver of fish infected with LCDV. Meantime, hepcidin, one of STAT3 target gene, was also up-regulated in liver of fish infected with two pathogens. These results suggested that turbot Stat3 may involved in the immune defense process as a transcription factor.

  4. Acetylation directs survivin nuclear localization to repress STAT3 oncogenic activity.

    PubMed

    Wang, Haijuan; Holloway, Michael P; Ma, Li; Cooper, Zachary A; Riolo, Matthew; Samkari, Ayman; Elenitoba-Johnson, Kojo S J; Chin, Y Eugene; Altura, Rachel A

    2010-11-12

    The multiple functions of the oncofetal protein survivin are dependent on its selective expression patterns within immunochemically distinct subcellular pools. The mechanism by which survivin localizes to these compartments, however, is only partly understood. Here we show that nuclear accumulation of survivin is promoted by CREB-binding protein (CBP)-dependent acetylation on lysine 129 (129K, Lys-129). We demonstrate a mechanism by which survivin acetylation at this position results in its homodimerization, while deacetylation promotes the formation of survivin monomers that heterodimerize with CRM1 and facilitate its nuclear export. Using proteomic analysis, we identified the oncogenic transcription factor STAT3 as a binding partner of nuclear survivin. We show that acetylated survivin binds to the N-terminal transcriptional activation domain of the STAT3 dimer and represses STAT3 transactivation of target gene promoters. Using multiplex PCR and DNA sequencing, we identified a single-nucleotide polymorphism (A → G) at Lys-129 that exists as a homozygous mutation in a neuroblastoma cell line and corresponds with a defect in survivin nuclear localization. Our results demonstrate that the dynamic equilibrium between survivin acetylation and deacetylation at amino acid 129 determines its interaction with CRM1, its subsequent subcellular localization, and its ability to inhibit STAT3 transactivation, providing a potential route for therapeutic intervention in STAT3-dependent tumors.

  5. ATF3 activates Stat3 phosphorylation through inhibition of p53 expression in skin cancer cells.

    PubMed

    Hao, Zhen-Feng; Ao, Jun-Hong; Zhang, Jie; Su, You-Ming; Yang, Rong-Ya

    2013-01-01

    ATF3, a member of the ATF/CREB family of transcription factors, has been found to be selectively induced by calcineurin/NFAT inhibition and to enhance keratinocyte tumor formation, although the precise role of ATF3 in human skin cancer and possible mechanisms remain unknown. In this study, clinical analysis of 30 skin cancer patients and 30 normal donors revealed that ATF3 was accumulated in skin cancer tissues. Functional assays demonstrated that ATF3 significantly promoted skin cancer cell proliferation. Mechanically, ATF3 activated Stat3 phosphorylation in skin cancer cell through regulation of p53 expression. Moreover, the promotion effect of ATF3 on skin cancer cell proliferation was dependent on the p53-Stat3 signaling cascade. Together, the results indicate that ATF3 might promote skin cancer cell proliferation and enhance skin keratinocyte tumor development through inhibiting p53 expression and then activating Stat3 phosphorylation.

  6. AG490 inhibits NFATc1 expression and STAT3 activation during RANKL induced osteoclastogenesis

    SciTech Connect

    Li, Chang-hong; Zhao, Jin-xia; Sun, Lin; Yao, Zhong-qiang; Deng, Xiao-li; Liu, Rui; Liu, Xiang-yuan

    2013-06-14

    Highlights: •AG490 inhibits RANKL-induced osteoclastogenesis in RAW264.7 cells. •AG490 affects cell proliferation and cell cycle distribution. •AG490 reduces NFATc1 expression during RANKL-induced osteoclastogenesis. •AG490 disrupts the activation of RANKL-mediated JAK2/STAT3 signaling pathway. •STAT3 depletion partly mimics the effect of AG490 on RANKL-induced osteoclastogenesis. -- Abstract: Commonly, JAK/STAT relays cytokine signals for cell activation and proliferation, and recent studies have shown that the elevated expression of JAK/STAT is associated with the immune rejection of allografts and the inflammatory processes of autoimmune disease. However, the role which JAK2/STAT3 signaling plays in the receptor activator of nuclear factor-κB ligand (RANKL)-mediated osteoclastogenesis is unknown. In this study, we investigated the effects of AG490, specific JAK2 inhibitor, on osteoclast differentiation in vitro. AG490 significantly inhibited osteoclastogenesis in murine osteoclast precursor cell line RAW264.7 induced by RANKL. AG490 suppressed cell proliferation and delayed the G1 to S cell cycle transition. Furthermore, AG490 also suppressed the expression of nuclear factor of activated T cells (NFAT) c1 but not c-Fos in RAW264.7. Subsequently, we investigated various intracellular signaling components associated with osteoclastogenesis. AG490 had no effects on RANKL-induced activation of Akt, ERK1/2. Interestingly, AG490 partly inhibited RANKL-induced phosphorylation of Ser{sup 727} in STAT3. Additionally, down-regulation of STAT3 using siRNA resulted in suppression of TRAP, RANK and NFATc1 expression. In conclusion, we demonstrated that AG490 inhibited RANKL-induced osteoclastogenesis by suppressing NFATc1 production and cell proliferation via the STAT3 pathway. These results suggest that inhibition of JAK2 may be useful for the treatment of bone diseases characterized by excessive osteoclastogenesis.

  7. Signal transducer and activator of transcription 3 (Stat3) expression and activation in rat uterus during early pregnancy.

    PubMed

    Teng, Chun-Bo; Diao, Hong-Lu; Ma, Hong; Cong, Jing; Yu, Hao; Ma, Xing-Hong; Xu, Li-Bin; Yang, Zeng-Ming

    2004-08-01

    Signal transducer and activator of transcription 3 (Stat3), a member of the Stat family, is specifically activated during mouse embryo implantation. The aim of this study was to investigate the expression, activation and regulation of Stat3 in rat uterus during early pregnancy, pseudopregnancy, delayed implantation and artificial decidualization. Stat3 mRNA was highly expressed in the luminal epithelium on day 5 and in the luminal epithelium and underlying stromal cells at implantation sites on day 6 of pregnancy. There was a strong level of Stat3 protein expression and phosphorylation in the stromal cells near the lumen and in the luminal epithelium on day 5 of pregnancy, which was similar to day 5 of pseudopregnancy. In the afternoon of day 6, the strong level of Stat3 phosphorylation was detected only in the luminal epithelium. Stat3 was highly expressed and activated in the decidual cells from days 7 to 9 of pregnancy and under artificial decidualization in the present study. Our results suggest that the strong level of Stat3 activation in the luminal epithelium and underlying stromal cells during the pre-implantation period may be important for establishing uterine receptivity as in mice, and the high level of Stat3 expression and activation in decidual cells may play a role during decidualization.

  8. Sphingosine-1-phosphate lyase downregulation promotes colon carcinogenesis through STAT3-activated microRNAs.

    PubMed

    Degagné, Emilie; Pandurangan, Ashok; Bandhuvula, Padmavathi; Kumar, Ashok; Eltanawy, Abeer; Zhang, Meng; Yoshinaga, Yuko; Nefedov, Mikhail; de Jong, Pieter J; Fong, Loren G; Young, Stephen G; Bittman, Robert; Ahmedi, Yasmin; Saba, Julie D

    2014-12-01

    Growing evidence supports a link between inflammation and cancer; however, mediators of the transition between inflammation and carcinogenesis remain incompletely understood. Sphingosine-1-phosphate (S1P) lyase (SPL) irreversibly degrades the bioactive sphingolipid S1P and is highly expressed in enterocytes but downregulated in colon cancer. Here, we investigated the role of SPL in colitis-associated cancer (CAC). We generated mice with intestinal epithelium-specific Sgpl1 deletion and chemically induced colitis and tumor formation in these animals. Compared with control animals, mice lacking intestinal SPL exhibited greater disease activity, colon shortening, cytokine levels, S1P accumulation, tumors, STAT3 activation, STAT3-activated microRNAs (miRNAs), and suppression of miR-targeted anti-oncogene products. This phenotype was attenuated by STAT3 inhibition. In fibroblasts, silencing SPL promoted tumorigenic transformation through a pathway involving extracellular transport of S1P through S1P transporter spinster homolog 2 (SPNS2), S1P receptor activation, JAK2/STAT3-dependent miR-181b-1 induction, and silencing of miR-181b-1 target cylindromatosis (CYLD). Colon biopsies from patients with inflammatory bowel disease revealed enhanced S1P and STAT3 signaling. In mice with chemical-induced CAC, oral administration of plant-type sphingolipids called sphingadienes increased colonic SPL levels and reduced S1P levels, STAT3 signaling, cytokine levels, and tumorigenesis, indicating that SPL prevents transformation and carcinogenesis. Together, our results suggest that dietary sphingolipids can augment or prevent colon cancer, depending upon whether they are metabolized to S1P or promote S1P metabolism through the actions of SPL.

  9. Androgen-STAT3 activation may contribute to gender disparity in human simply renal cysts.

    PubMed

    Liu, Min; Xu, Yun-Fei; Feng, Yuan; Zhai, Wei; Che, Jian-Ping; Xia, Sheng-Qiang; Wang, Guang-Chun; Zheng, Jun-Hua

    2013-01-01

    Simple renal cysts (SRC) are a common urological disease mostly in elderly, however the male-to-female ratio was 2.81. Androgen receptor (AR) activation was initially proposed as a vital signaling pathway in prostate cancer and consequent signal transducer and activator of transcription 3 (STAT3)-AR complex led an important putative mechanism by which prostate cells are sensitized with growth factor signals. However, in SRC disease, no related study emerged. 30 patients with SRC and 20 age-matched healthy controls were recruited. Puncture biopsy was performed to acquire cyst-adjacent kidney tissue and normal kidney tissues were from healthy kidney donor who received living-related donor nephrectomy. The expression of STAT3 and androgen receptor was determined by immunohistochemical staining and western blotting. The in-vitro effect of androgen on human HK-2 (an immortalized proximal tubule epithelial cell line from normal adult human kidney) cells' STAT3 expression was analyzed as well. Activated STAT3 was strongly expressed in tubular epithelial cells from kidneys of SRC patients, while it was barely found in normal kidneys. Meanwhile, the androgen receptor positive cyst epithelial cells and adjacent normal renal tubule cells were observed in kidneys from SRC patients, however, AR was weakly expressed in normal healthy male kidneys, statistically significant differences existed. In-vitro experiment demonstrated that when treated with exogenous added androgen, the expression level of STAT3 in HK-2 cells was significantly elevated. Our data raised the possible novel evidence that androgen-STAT3 activation might contribute to gender disparity in human SRC disease and clarification the esoteric mechanisms will provide us attractive therapy target for cystic kidney disease.

  10. Sphingosine-1-phosphate lyase downregulation promotes colon carcinogenesis through STAT3-activated microRNAs

    PubMed Central

    Degagné, Emilie; Pandurangan, Ashok; Bandhuvula, Padmavathi; Kumar, Ashok; Eltanawy, Abeer; Zhang, Meng; Yoshinaga, Yuko; Nefedov, Mikhail; de Jong, Pieter J.; Fong, Loren G.; Young, Stephen G.; Bittman, Robert; Ahmedi, Yasmin; Saba, Julie D.

    2014-01-01

    Growing evidence supports a link between inflammation and cancer; however, mediators of the transition between inflammation and carcinogenesis remain incompletely understood. Sphingosine-1-phosphate (S1P) lyase (SPL) irreversibly degrades the bioactive sphingolipid S1P and is highly expressed in enterocytes but downregulated in colon cancer. Here, we investigated the role of SPL in colitis-associated cancer (CAC). We generated mice with intestinal epithelium-specific Sgpl1 deletion and chemically induced colitis and tumor formation in these animals. Compared with control animals, mice lacking intestinal SPL exhibited greater disease activity, colon shortening, cytokine levels, S1P accumulation, tumors, STAT3 activation, STAT3-activated microRNAs (miRNAs), and suppression of miR-targeted anti-oncogene products. This phenotype was attenuated by STAT3 inhibition. In fibroblasts, silencing SPL promoted tumorigenic transformation through a pathway involving extracellular transport of S1P through S1P transporter spinster homolog 2 (SPNS2), S1P receptor activation, JAK2/STAT3-dependent miR-181b-1 induction, and silencing of miR-181b-1 target cylindromatosis (CYLD). Colon biopsies from patients with inflammatory bowel disease revealed enhanced S1P and STAT3 signaling. In mice with chemical-induced CAC, oral administration of plant-type sphingolipids called sphingadienes increased colonic SPL levels and reduced S1P levels, STAT3 signaling, cytokine levels, and tumorigenesis, indicating that SPL prevents transformation and carcinogenesis. Together, our results suggest that dietary sphingolipids can augment or prevent colon cancer, depending upon whether they are metabolized to S1P or promote S1P metabolism through the actions of SPL. PMID:25347472

  11. Evaluation of quantitative assays for the identification of direct signal transducer and activator of transcription 3 (STAT3) inhibitors

    PubMed Central

    Furtek, Steffanie L.; Matheson, Christopher J.; Backos, Donald S.; Reigan, Philip

    2016-01-01

    In many forms of cancer the signal transducer and activator of transcription 3 (STAT3) transcription factor remains constitutively active, driving cancer survival and progression. The critical role of STAT3 in tumorigenesis has prompted a campaign of drug discovery programs to identify small molecules that disrupt the function of STAT3, with more recent efforts focusing on direct STAT3 inhibition. There are two target binding sites for direct STAT3 inhibitors: the SH2 dimerization domain and the DNA-binding domain. An in vitro fluorescence polarization assay, using recombinant STAT3 protein, has successfully identified compounds that target the SH2 domain; however, no assay has been reported to identify inhibitors that bind the DNA-binding domain. The lack of such a quantitative assay has limited the identification and development of STAT3 DNA-binding domain inhibitors. Here, we report a modified DNA-binding ELISA to incorporate recombinant STAT3 protein to evaluate small molecules that prevent STAT3-DNA binding. The concomitant use of the ELISA and fluorescence polarization assay enables the classification of direct STAT3 inhibitors by their site of action. Our data provide further support that niclosamide inhibits STAT3 through interaction with the DNA-binding domain. Furthermore, the ELISA can support medicinal chemistry efforts by identifying DNA-binding domain inhibitors and allowing the determination of an IC50 value, supporting the ranking of inhibitors and development of structure-activity relationships. Therefore, we propose a tandem evaluation approach to identify small molecules that target the SH2 domain or the DNA-binding domain of STAT3, which allows for quantitative evaluation of candidate STAT3 inhibitors. PMID:27793003

  12. Evaluation of quantitative assays for the identification of direct signal transducer and activator of transcription 3 (STAT3) inhibitors.

    PubMed

    Furtek, Steffanie L; Matheson, Christopher J; Backos, Donald S; Reigan, Philip

    2016-11-22

    In many forms of cancer the signal transducer and activator of transcription 3 (STAT3) transcription factor remains constitutively active, driving cancer survival and progression. The critical role of STAT3 in tumorigenesis has prompted a campaign of drug discovery programs to identify small molecules that disrupt the function of STAT3, with more recent efforts focusing on direct STAT3 inhibition. There are two target binding sites for direct STAT3 inhibitors: the SH2 dimerization domain and the DNA-binding domain. An in vitro fluorescence polarization assay, using recombinant STAT3 protein, has successfully identified compounds that target the SH2 domain; however, no assay has been reported to identify inhibitors that bind the DNA-binding domain. The lack of such a quantitative assay has limited the identification and development of STAT3 DNA-binding domain inhibitors. Here, we report a modified DNA-binding ELISA to incorporate recombinant STAT3 protein to evaluate small molecules that prevent STAT3-DNA binding. The concomitant use of the ELISA and fluorescence polarization assay enables the classification of direct STAT3 inhibitors by their site of action. Our data provide further support that niclosamide inhibits STAT3 through interaction with the DNA-binding domain. Furthermore, the ELISA can support medicinal chemistry efforts by identifying DNA-binding domain inhibitors and allowing the determination of an IC50 value, supporting the ranking of inhibitors and development of structure-activity relationships. Therefore, we propose a tandem evaluation approach to identify small molecules that target the SH2 domain or the DNA-binding domain of STAT3, which allows for quantitative evaluation of candidate STAT3 inhibitors.

  13. Both endogenous and exogenous testosterone decrease myocardial STAT3 activation and SOCS3 expression after acute ischemia and reperfusion.

    PubMed

    Wang, Meijing; Wang, Yue; Abarbanell, Aaron; Tan, Jiangjing; Weil, Brent; Herrmann, Jeremy; Meldrum, Daniel R

    2009-08-01

    Signal transducer and activator of transduction 3 (STAT3) pathway has been shown to be cardioprotective. We observed decreased STAT3/suppressor of cytokine signaling 3 (SOCS3) in male hearts, which was associated with worse postischemic myocardial function compared with females. However, it is unclear whether this downregulation of myocardial STAT3/SOCS3 is due to testosterone in males. We hypothesized that after ischemia/reperfusion (I/R), (1) endogenous testosterone decreases myocardial STAT3 and SOCS3 in males, and (2) administration of exogenous testosterone reduces myocardial STAT3/SOCS3 in female and castrated male hearts. To study this, hearts from I/R injury (Langendorff) were homogenized and assessed for phosphorylated-STAT3 (p-STAT3), total-STAT3 (T-STAT3), SOCS3, and GAPDH by Western blot. We grouped age-matched adult males, females, castrated males, males with androgen receptor blocker-flutamide implantation, females, and castrated males with chronic (3-week) 5alpha-dihydrotestosterone (DHT) release pellet implantation or acute (5-minute) testosterone infusion (ATI) before ischemia (n = 5-9 per group). Castration or flutamide treatment significantly increased SOCS3 expression in male hearts after I/R. However, only castration increased myocardial STAT3 activation. Notably, DHT replacement or ATI decreased markedly myocardial STAT3/SOCS3 in castrated males and females subjected to I/R. These results suggest that endogenous and exogenous testosterone decrease myocardial STAT3 activation and SOCS3 expression after I/R. This represents the initial demonstration of testosterone-downregulated STAT3/SOCS3 signaling in myocardium.

  14. [STAT3 activation by hypoxia in in vitro models of cervix cancer and endothelial cells].

    PubMed

    Ortega, Óscar; Ondo-Méndez, Alejandro; Garzón, Ruth

    2017-01-24

    The biological behavior of cancer cells is influenced by the tumor microenvironment in which they develop. In this context, stressor stimuli such as hypoxia are considered critical for tumor development and therapeutic management. Cellular response to various stimuli is evidenced in the activation of intracellular signaling pathways such as JAK/STAT, which is one of the most important for its effects in differentiation and cell proliferation. To evaluate the condition of the JAK/STAT pathway through the expression/activation of the STAT3 protein in cervix cancer cells (HeLa) and endothelial cells (EA.hy926) subjected to hypoxia. Cell lines were subjected to physical (1% O2) or chemical (deferoxamine, DFO, 100 μM) hypoxia for 2, 6 and 24 hours. Changes in the expression and activation of STAT3, and its subcellular localization by indirect immunofluorescence, were determined by western blot. Hypoxia was evidenced by the activation and translocation to the nucleus of HIF-1. Neither physical nor chemical hypoxia altered STAT3 expression, but it did affect its activation, as seen in its phosphorylation and translocation to the nucleus in the two models under study. The present study highlights the importance of hypoxia as a stimulus that modifies the activation of the STAT3 protein in HeLa and EA.hy926 cells, which makes it an important factor in the design of therapeutic strategies against cancer.

  15. Hippocalcin Is Required for Astrocytic Differentiation through Activation of Stat3 in Hippocampal Neural Precursor Cells

    PubMed Central

    Kang, Min-Jeong; Park, Shin-Young; Han, Joong-Soo

    2016-01-01

    Hippocalcin (Hpca) is a neuronal calcium sensor protein expressed in the mammalian brain. However, its function in neural stem/precursor cells has not yet been studied. Here, we clarify the function of Hpca in astrocytic differentiation in hippocampal neural precursor cells (HNPCs). When we overexpressed Hpca in HNPCs in the presence or absence of bFGF, expression levels of nerve-growth factors such as neurotrophin-3 (NT-3), neurotrophin-4/5 (NT-4/5), and brain-derived neurotrophic factor (BDNF), together with the proneural basic helix loop helix (bHLH) transcription factors NeuroD and neurogenin 1 (Ngn1), increased significantly. In addition, there was an increase in the number of cells expressing glial fibrillary acidic protein (GFAP), an astrocyte marker, and in branch outgrowth, indicating astrocytic differentiation of the HNPCs. Downregulation of Hpca by transfection with Hpca siRNA reduced expression of NT-3, NT-4/5, BDNF, NeuroD, and Ngn1 as well as levels of GFAP protein. Furthermore, overexpression of Hpca increased the phosphorylation of STAT3 (Ser727), and this effect was abolished by treatment with a STAT3 inhibitor (S3I-201), suggesting that STAT3 (Ser727) activation is involved in Hpca-mediated astrocytic differentiation. As expected, treatment with Stat3 siRNA or STAT3 inhibitor caused a complete inhibition of astrogliogenesis induced by Hpca overexpression. Taken together, this is the first report to show that Hpca, acting through Stat3, has an important role in the expression of neurotrophins and proneural bHLH transcription factors, and that it is an essential regulator of astrocytic differentiation and branch outgrowth in HNPCs. PMID:27840601

  16. An inhibitor of cholesterol absorption displays anti-myeloma activity by targeting the JAK2-STAT3 signaling pathway

    PubMed Central

    Zhu, Jingyu; Mao, Hongwu; Lin, Xu; Zhang, Zubin; Cao, Biyin; Zeng, Yuanying; Mao, Xinliang

    2016-01-01

    The activated JAK2-STAT3 signaling pathway is a high risk factor for multiple myeloma (MM), a fatal malignancy of plasma cells. In the present study, SC09, a potential inhibitor of cholesterol absorption, was identified in a STAT3-targeted drug screen. SC09 suppressed the activation of STAT3 in a time-course and concentration-dependent manner but did not affect its family members STAT1 and STAT5. SC09 inhibited STAT3 transcriptional activity and downregulated the expression of STAT3-regulated genes. Further studies showed that SC09 selectively inhibited JAK2 activation but not other kinases including c-Src, ERK, p38 and mTOR that are all associated with STAT3 activation. Moreover, SC09 obviously induced MM cell death in vitro and delayed MM tumor growth in vivo. SC09-induced MM cell death was dependent on the endogenous STAT3 status, and this effect could be attenuated by enforced expression of STAT3. All the results collectively indicated that SC09 blocks the JAK2-STAT3 signaling thus displaying anti-MM activity. Given its well tolerance and anti-MM potency, SC09 is credited for further investigation as a promising drug for MM treatment. PMID:27705908

  17. STAT3 activation and infiltration of eosinophil granulocytes in mycosis fungoides.

    PubMed

    Fredholm, Simon; Gjerdrum, Lise Mette R; Willerslev-Olsen, Andreas; Petersen, David L; Nielsen, Inger Ø; Kauczok, Claudia-S; Wobser, Marion; Ralfkiaer, Ulrik; Bonefeld, Charlotte M; Wasik, Mariusz A; Krejsgaard, Thorbjørn; Geisler, Carsten; Ralfkiaer, Elisabeth; Gniadecki, Robert; Woetmann, Anders; Odum, Niels

    2014-10-01

    Eosinophil granulocytes have been implicated in anticancer immunity but recent data indicate that eosinophils can also promote cancer. Herein, we studied eosinophils in skin lesions from 43 patients with mycosis fungoides (MF). The presence of eosinophils correlated with disease stage: 78% of patients with advanced disease displayed eosinophil infiltration, whereas this was only seen in 11% of patients with patches (p<0.01), and in 48% of those with plaque disease. Importantly, 72% of patients with positive staining for phospho-signal-transducer-and-activator-of-transcription (pY-STAT3) in malignant T-cells also stained positively for eosinophils, whereas this was only observed in 28% of pY-STAT3-negative patients (p<0.01). Notably, malignant T-cells expressed eosinophilic activation and trafficking factors: High-mobility group BOX-1 protein (HMGB1) and interleukin 5 (IL5). STAT3 siRNA profoundly inhibited IL5 but not HMGB1 expression. In conclusion, these data suggest that malignant T-cells orchestrate accumulation and activation of eosinophils supporting the notion of STAT3 being a putative target for therapy.

  18. STAT3 participates in transcriptional activation of the C-reactive protein gene by interleukin-6.

    PubMed

    Zhang, D; Sun, M; Samols, D; Kushner, I

    1996-04-19

    Interleukin-6 (IL-6) is the major cytokine inducing transcription of human C-reactive protein (CRP) during the acute phase response. STAT (signal transducers and activators of transcription) family members, recently shown to be important mediators of the effects of many cytokines including IL-6, generally induce their effects by binding to palindromic sequences with TT(N)5AA motifs. We report an IL-6 responsive element in the proximal region of the human CRP 5'-flanking region that bears a TT(N)4AA motif, which we have termed CRP acute phase response element (CRP-APRE). In Hep3B cells, IL-6 but not interferon-gamma was capable of activating CAT constructs driven by the CRP promoter containing CRP-APRE. Overexpressed STAT3 was able to transactivate CRP-chloramphenicol acetyltransferase constructs through the CRP-APRE and was able to enhance endogenous CRP mRNA accumulation in response to IL-6. STAT3 (or an antigenically related molecule) bound to the CRP-APRE in response to IL-6. Overexpression of STAT3 in the presence of IL-6 was capable of inducing expression of a construct consisting of the CRP-APRE and a minimal thymidine kinase promoter lacking a C/EBP site. Taken together, these findings indicate that STAT3 participates in the transcriptional activation of CRP in response to IL-6.

  19. Autoimmunity, hypogammaglobulinemia, lymphoproliferation, and mycobacterial disease in patients with activating mutations in STAT3

    PubMed Central

    Haapaniemi, Emma M.; Kaustio, Meri; Rajala, Hanna L. M.; van Adrichem, Arjan J.; Kainulainen, Leena; Glumoff, Virpi; Doffinger, Rainer; Kuusanmäki, Heikki; Heiskanen-Kosma, Tarja; Trotta, Luca; Chiang, Samuel; Kulmala, Petri; Eldfors, Samuli; Katainen, Riku; Siitonen, Sanna; Karjalainen-Lindsberg, Marja-Liisa; Kovanen, Panu E.; Otonkoski, Timo; Porkka, Kimmo; Heiskanen, Kaarina; Hänninen, Arno; Bryceson, Yenan T.; Uusitalo-Seppälä, Raija; Saarela, Janna; Seppänen, Mikko; Kere, Juha

    2015-01-01

    The signal transducer and activator of transcription (STAT) family of transcription factors orchestrate hematopoietic cell differentiation. Recently, mutations in STAT1, STAT5B, and STAT3 have been linked to development of immunodysregulation polyendocrinopathy enteropathy X-linked–like syndrome. Here, we immunologically characterized 3 patients with de novo activating mutations in the DNA binding or dimerization domains of STAT3 (p.K392R, p.M394T, and p.K658N, respectively). The patients displayed multiorgan autoimmunity, lymphoproliferation, and delayed-onset mycobacterial disease. Immunologically, we noted hypogammaglobulinemia with terminal B-cell maturation arrest, dendritic cell deficiency, peripheral eosinopenia, increased double-negative (CD4−CD8−) T cells, and decreased natural killer, T helper 17, and regulatory T-cell numbers. Notably, the patient harboring the K392R mutation developed T-cell large granular lymphocytic leukemia at age 14 years. Our results broaden the spectrum of phenotypes caused by activating STAT3 mutations, highlight the role of STAT3 in the development and differentiation of multiple immune cell lineages, and strengthen the link between the STAT family of transcription factors and autoimmunity. PMID:25349174

  20. Autoimmunity, hypogammaglobulinemia, lymphoproliferation, and mycobacterial disease in patients with activating mutations in STAT3.

    PubMed

    Haapaniemi, Emma M; Kaustio, Meri; Rajala, Hanna L M; van Adrichem, Arjan J; Kainulainen, Leena; Glumoff, Virpi; Doffinger, Rainer; Kuusanmäki, Heikki; Heiskanen-Kosma, Tarja; Trotta, Luca; Chiang, Samuel; Kulmala, Petri; Eldfors, Samuli; Katainen, Riku; Siitonen, Sanna; Karjalainen-Lindsberg, Marja-Liisa; Kovanen, Panu E; Otonkoski, Timo; Porkka, Kimmo; Heiskanen, Kaarina; Hänninen, Arno; Bryceson, Yenan T; Uusitalo-Seppälä, Raija; Saarela, Janna; Seppänen, Mikko; Mustjoki, Satu; Kere, Juha

    2015-01-22

    The signal transducer and activator of transcription (STAT) family of transcription factors orchestrate hematopoietic cell differentiation. Recently, mutations in STAT1, STAT5B, and STAT3 have been linked to development of immunodysregulation polyendocrinopathy enteropathy X-linked-like syndrome. Here, we immunologically characterized 3 patients with de novo activating mutations in the DNA binding or dimerization domains of STAT3 (p.K392R, p.M394T, and p.K658N, respectively). The patients displayed multiorgan autoimmunity, lymphoproliferation, and delayed-onset mycobacterial disease. Immunologically, we noted hypogammaglobulinemia with terminal B-cell maturation arrest, dendritic cell deficiency, peripheral eosinopenia, increased double-negative (CD4(-)CD8(-)) T cells, and decreased natural killer, T helper 17, and regulatory T-cell numbers. Notably, the patient harboring the K392R mutation developed T-cell large granular lymphocytic leukemia at age 14 years. Our results broaden the spectrum of phenotypes caused by activating STAT3 mutations, highlight the role of STAT3 in the development and differentiation of multiple immune cell lineages, and strengthen the link between the STAT family of transcription factors and autoimmunity.

  1. Folic acid mediates activation of the pro-oncogene STAT3 via the Folate Receptor alpha.

    PubMed

    Hansen, Mariann F; Greibe, Eva; Skovbjerg, Signe; Rohde, Sarah; Kristensen, Anders C M; Jensen, Trine R; Stentoft, Charlotte; Kjær, Karina H; Kronborg, Camilla S; Martensen, Pia M

    2015-07-01

    The signal transducer and activator of transcription 3 (STAT3) is a well-described pro-oncogene found constitutively activated in several cancer types. Folates are B vitamins that, when taken up by cells through the Reduced Folate Carrier (RFC), are essential for normal cell growth and replication. Many cancer cells overexpress a glycophosphatidylinositol (GPI)-anchored Folate Receptor α (FRα). The function of FRα in cancer cells is still poorly described, and it has been suggested that transport of folate is not its primary function in these cells. We show here that folic acid and folinic acid can activate STAT3 through FRα in a Janus Kinase (JAK)-dependent manner, and we demonstrate that gp130 functions as a transducing receptor for this signalling. Moreover, folic acid can promote dose dependent cell proliferation in FRα-positive HeLa cells, but not in FRα-negative HEK293 cells. After folic acid treatment of HeLa cells, up-regulation of the STAT3 responsive genes Cyclin A2 and Vascular Endothelial Growth Factor (VEGF) were verified by qRT-PCR. The identification of this FRα-STAT3 signal transduction pathway activated by folic and folinic acid contributes to the understanding of the involvement of folic acid in preventing neural tube defects as well as in tumour growth. Previously, the role of folates in these diseases has been attributed to their roles as one-carbon unit donors following endocytosis into the cell. Our finding that folic acid can activate STAT3 via FRα adds complexity to the established roles of B9 vitamins in cancer and neural tube defects.

  2. Zinc Chloride Transiently Maintains Mouse Embryonic Stem Cell Pluripotency by Activating Stat3 Signaling.

    PubMed

    Hu, Jing; Yang, Zhiyong; Wang, Jinbo; Yu, Jia; Guo, Jing; Liu, Shiying; Qian, Chunmei; Song, Liwen; Wu, Yi; Cheng, Jiajing

    2016-01-01

    An improved understanding of the pluripotency maintenance of embryonic stem (ES) cells is important for investigations of early embryo development and for cell replacement therapy, but the mechanism behind pluripotency is still incompletely understood. Recent findings show that zinc, an essential trace element in humans, is critically involved in regulating various signaling pathways and genes expression. However, its role in ES cell fate determination remains to be further explored. Here we showed that 2μM zinc chloride (ZnCl2) transiently maintained mouse ES cell pluripotency in vitro. The cultured mouse ES cells remained undifferentiated under 2μM ZnCl2 treatment in leukemia inhibitory factor (LIF) withdrawal, retinoic acid (RA) or embryoid bodies (EBs) differentiation assays. In addition, ZnCl2 increased pluripotency genes expression and inhibited differentiation genes expression. Further mechanistic studies revealed that ZnCl2 transiently activated signal transducers and activators of transcription 3 (Stat3) signaling through promoting Stat3 phosphorylation. Inhibition of Stat3 signaling abrogated the effects of ZnCl2 on mouse ES cell pluripotency. Taken together, this study demonstrated a critical role of zinc in the pluripotency maintenance of mouse ES cells, as well as an important regulator of Stat3 signaling.

  3. 14-3-3ζ Interacts with Stat3 and Regulates Its Constitutive Activation in Multiple Myeloma Cells

    PubMed Central

    Li, Wenliang; Xiong, Qian; Yang, Mingkun; Zheng, Peng; Li, Chongyang; Pei, Jianfeng; Ge, Feng

    2012-01-01

    The 14-3-3 proteins are a family of regulatory signaling molecules that interact with other proteins in a phosphorylation-dependent manner and function as adapter or scaffold proteins in signal transduction pathways. One family member, 14-3-3ζ, is believed to function in cell signaling, cycle control, and apoptotic death. A systematic proteomic analysis done in our laboratory has identified signal transducers and activators of transcription 3 (Stat3) as a novel 14-3-3ζ interacting protein. Following our initial finding, in this study, we provide evidence that 14-3-3ζ interacts physically with Stat3. We further demonstrate that phosphorylation of Stat3 at Ser727 is vital for 14-3-3ζ interaction and mutation of Ser727 to Alanine abolished 14-3-3ζ/Stat3 association. Inhibition of 14-3-3ζ protein expression in U266 cells inhibited Stat3 Ser727 phosphorylation and nuclear translocation, and decreased both Stat3 DNA binding and transcriptional activity. Moreover, 14-3-3ζ is involved in the regulation of protein kinase C (PKC) activity and 14-3-3ζ binding to Stat3 protects Ser727 dephosphorylation from protein phosphatase 2A (PP2A). Taken together, our findings support the model that multiple signaling events impinge on Stat3 and that 14-3-3ζ serves as an essential coordinator for different pathways to regulate Stat3 activation and function in MM cells. PMID:22279540

  4. Inhibition of STAT3 activity delays obesity-induced thyroid carcinogenesis in a mouse model.

    PubMed

    Park, Jeong Won; Han, Cho Rong; Zhao, Li; Willingham, Mark C; Cheng, Sheue-yann

    2016-01-01

    Compelling epidemiologic studies indicate that obesity is a risk factor for many human cancers, including thyroid cancer. In recent decades, the incidence of thyroid cancer has dramatically increased along with a marked rise in obesity prevalence. We previously demonstrated that a high fat diet (HFD) effectively induced the obese phenotype in a mouse model of thyroid cancer (Thrb(PV/PV)Pten(+/-) mice). Moreover, HFD activates the STAT3 signal pathway to promote more aggressive tumor phenotypes. The aim of the present study was to evaluate the effect of S3I-201, a specific inhibitor of STAT3 activity, on HFD-induced aggressive cancer progression in the mouse model of thyroid cancer. WT and Thrb(PV/PV)Pten(+/-) mice were treated with HFD together with S3I-201 or vehicle-only as controls. We assessed the effects of S3I-201 on HFD-induced thyroid cancer progression, the leptin-JAK2-STAT3 signaling pathway, and key regulators of epithelial-mesenchymal transition (EMT). S3I-201 effectively inhibited HFD-induced aberrant activation of STAT3 and its downstream targets to markedly inhibit thyroid tumor growth and to prolong survival. Decreased protein levels of cyclins D1 and B1, cyclin dependent kinase 4 (CDK4), CDK6, and phosphorylated retinoblastoma protein led to the inhibition of tumor cell proliferation in S3I-201-treated Thrb(PV/PV)Pten(+/-) mice. Reduced occurrence of vascular invasion and blocking of anaplasia and lung metastasis in thyroid tumors of S3I-201-treated Thrb(PV/PV)Pten(+/-) mice were mediated via decreased expression of vimentin and matrix metalloproteinases, two key effectors of EMT. The present findings suggest that inhibition of the STAT3 activity would be a novel treatment strategy for obesity-induced thyroid cancer. © 2016 Society for Endocrinology.

  5. Enhanced Transcriptional Activity and Mitochondrial Localization of STAT3 Co-induce Axon Regrowth in the Adult Central Nervous System.

    PubMed

    Luo, Xueting; Ribeiro, Marcio; Bray, Eric R; Lee, Do-Hun; Yungher, Benjamin J; Mehta, Saloni T; Thakor, Kinjal A; Diaz, Francisca; Lee, Jae K; Moraes, Carlos T; Bixby, John L; Lemmon, Vance P; Park, Kevin K

    2016-04-12

    Signal transducer and activator of transcription 3 (STAT3) is a transcription factor central to axon regrowth with an enigmatic ability to act in different subcellular regions independently of its transcriptional roles. However, its roles in mature CNS neurons remain unclear. Here, we show that along with nuclear translocation, STAT3 translocates to mitochondria in mature CNS neurons upon cytokine stimulation. Loss- and gain-of-function studies using knockout mice and viral expression of various STAT3 mutants demonstrate that STAT3's transcriptional function is indispensable for CNS axon regrowth, whereas mitochondrial STAT3 enhances bioenergetics and further potentiates regrowth. STAT3's localization, functions, and growth-promoting effects are regulated by mitogen-activated protein kinase kinase (MEK), an effect further enhanced by Pten deletion, leading to extensive axon regrowth in the mouse optic pathway and spinal cord. These results highlight CNS neuronal dependence on STAT3 transcriptional activity, with mitochondrial STAT3 providing ancillary roles, and illustrate a critical contribution for MEK in enhancing diverse STAT3 functions and axon regrowth. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  6. Photodynamic therapy activated signaling from epidermal growth factor receptor and STAT3

    PubMed Central

    Edmonds, Christine; Hagan, Sarah; Gallagher-Colombo, Shannon M.; Busch, Theresa M.; Cengel, Keith A.

    2012-01-01

    Patients with serosal (pleural or peritoneal) spread of malignancy have few definitive treatment options and consequently have a very poor prognosis. We have previously shown that photodynamic therapy (PDT) can be an effective treatment for these patients, but that the therapeutic index is relatively narrow. Here, we test the hypothesis that EGFR and STAT3 activation increase survival following PDT, and that inhibiting these pathways leads to increased PDT-mediated direct cellular cytotoxicity by examining BPD-PDT in OvCa and NSCLC cells. We found that BPD-mediated PDT stimulated EGFR tyrosine phosphorylation and nuclear translocation, and that EGFR inhibition by erlotinib resulted in reduction of PDT-mediated EGFR activation and nuclear translocation. Nuclear translocation and PDT-mediated activation of EGFR were also observed in response to BPD-mediated PDT in multiple cell lines, including OvCa, NSCLC and head and neck cancer cells, and was observed to occur in response to porfimer sodium-mediated PDT. In addition, we found that PDT stimulates nuclear translocation of STAT3 and STAT3/EGFR association and that inhibiting STAT3 signaling prior to PDT leads to increased PDT cytotoxicity. Finally, we found that inhibition of EGFR signaling leads to increased PDT cytotoxicity through a mechanism that involves increased apoptotic cell death. Taken together, these results demonstrate that PDT stimulates the nuclear accumulation of both EGFR and STAT3 and that targeting these survival pathways is a potentially promising strategy that could be adapted for clinical trials of PDT for patients with serosal spread of malignancy. PMID:22986230

  7. STAT3-mediated SMAD3 activation underlies Oncostatin M-induced Senescence.

    PubMed

    Bryson, Benjamin L; Junk, Damian J; Cipriano, Rocky; Jackson, Mark W

    2017-02-16

    Cytokines in the developing tumor microenvironment (TME) can drive transformation and subsequent progression toward metastasis. Elevated levels of the Interleukin-6 (IL-6) family cytokine Oncostatin M (OSM) in the breast TME correlate with aggressive, metastatic cancers, increased tumor recurrence, and poor patient prognosis. Paradoxically, OSM engages a tumor-suppressive, Signal Transducer and Activator of Transcription 3 (STAT3)-dependent senescence response in normal and non-transformed human mammary epithelial cells (HMEC). Here, we identify a novel link between OSM-activated STAT3 signaling and the Transforming Growth Factor-β (TGF-β) signaling pathway that engages senescence in HMEC. Inhibition of functional TGF-β/SMAD signaling by expressing a dominant-negative TGF-β receptor, treating with a TGF-β receptor inhibitor, or suppressing SMAD3 expression using a SMAD3-shRNA prevented OSM-induced senescence. OSM promoted a protein complex involving activated-STAT3 and SMAD3, induced the nuclear localization of SMAD3, and enhanced SMAD3-mediated transcription responsible for senescence. In contrast, expression of MYC (c-MYC) from a constitutive promoter abrogated senescence and strikingly, cooperated with OSM to promote a transformed phenotype, epithelial-mesenchymal transition (EMT), and invasiveness. Our findings suggest that a novel STAT3/SMAD3-signaling axis is required for OSM-mediated senescence that is coopted during the transformation process to confer aggressive cancer cell properties. Understanding how developing cancer cells bypass OSM/STAT3/SMAD3-mediated senescence may help identify novel targets for future "pro-senescence" therapies aiming to reengage this hidden tumor-suppressive response.

  8. Leukemia inhibitory factor promotes tumor growth and metastasis in human osteosarcoma via activating STAT3.

    PubMed

    Liu, Bin; Lu, Yi; Li, Jinzhi; Liu, Yanping; Liu, Jian; Wang, Weiguo

    2015-10-01

    The leukemia inhibitory factor (LIF) has been demonstrated to be an oncogene and participated in multiple procedures during the initiation and progression of many human malignancies. However, the role of LIF in osteosarcoma is still largely unknown. Here, we performed a series of in vitro and in vivo experiments to investigate the expression and biological functions of LIF in osteosarcoma. Compared to that in the non-cancerous tissues, LIF was significantly overexpressed in a panel of 68 osteosarcoma samples (p < 0.0001). Moreover, the overexpression of LIF was significantly correlated with advanced tumor stage, larger tumor size, and shorter overall survival. In addition, knockdown of LIF notably suppressed the proliferation and invasion of osteosarcoma via blocking the STAT3 signal pathway; in contrast, treatment with the recombinant LIF protein significantly promoted the growth and invasion of osteosarcoma through enhancing the phosphorylation of STAT3, which can be partially neutralized by the STAT3 inhibitor, HO-3867. In conclusion, we demonstrated that LIF was frequently overexpressed in osteosarcoma, which could promote the growth and invasion through activating the STAT3 pathway. Our findings proposed that LIF might be a potent therapeutic target for osteosarcoma. © 2015 APMIS. Published by John Wiley & Sons Ltd.

  9. DRUG-REPOSITIONING SCREENING IDENTIFIED PIPERLONGUMINE AS A DIRECT STAT3 INHIBITOR WITH POTENT ACTIVITY AGAINST BREAST CANCER

    PubMed Central

    Bharadwaj, Uddalak; Eckols, T. Kris; Kolosov, Mikhail; Kasembeli, Moses M.; Adam, Abel; Torres, David; Zhang, Xiaomei; Dobrolecki, Lacey E.; Wei, Wei; Lewis, Michael T.; Dave, Bhuvanesh; Chang, Jenny C.; Landis, Melissa D.; Creighton, Chad J.; Mancini, Michael A.; Tweardy, David J.

    2014-01-01

    Signal transducer and activator of transcription (STAT) 3 regulates many cardinal features of cancer including cancer cell growth, apoptosis resistance, DNA damage response, metastasis, immune escape, tumor angiogenesis, the Warburg effect, and oncogene addiction and has been validated as a drug target for cancer therapy. Several strategies have been employed to identify agents that target Stat3 in breast cancer but none has yet entered into clinical use. We used a high-throughput fluorescence microscopy search strategy to identify compounds in a drug-repositioning library (Prestwick library) that block ligand-induced nuclear translocation of Stat3 and identified piperlongumine (PL), a natural product isolated from the fruit of the pepper Piper longum. Piperlongumine inhibited Stat3 nuclear translocation, inhibited ligand-induced and constitutive Stat3 phosphorylation, and modulated expression of multiple Stat3-regulated genes. Surface plasmon resonance assay revealed that piperlongumine directly inhibited binding of Stat3 to its phosphotyrosyl (pY) peptide ligand. Phosphoprotein antibody array analysis revealed that PL does not modulate kinases known to activate Stat3 such as JAKs, Src kinase family members, or RTKs. PL inhibited anchorage-independent and anchorage-dependent growth of multiple breast cancer cell lines having increased pStat3 or total Stat3, and induced apoptosis. PL also inhibited mammosphere formation by tumor cells from patient derived xenografts (PDX). PL’s anti-tumorigenic function was causally linked to its Stat3-inhibitory effect. PL was non-toxic in mice up to a dose of 30 mg/Kg/day for 14 days and caused regression of breast cancer cell line xenografts in nude mice. Thus, PL represents a promising new agent for rapid entry into the clinic for use in treating breast cancer, as well as other cancers in which Stat3 plays a role. PMID:24681959

  10. Drug-repositioning screening identified piperlongumine as a direct STAT3 inhibitor with potent activity against breast cancer.

    PubMed

    Bharadwaj, U; Eckols, T K; Kolosov, M; Kasembeli, M M; Adam, A; Torres, D; Zhang, X; Dobrolecki, L E; Wei, W; Lewis, M T; Dave, B; Chang, J C; Landis, M D; Creighton, C J; Mancini, M A; Tweardy, D J

    2015-03-12

    Signal transducer and activator of transcription (STAT) 3 regulates many cardinal features of cancer including cancer cell growth, apoptosis resistance, DNA damage response, metastasis, immune escape, tumor angiogenesis, the Warburg effect and oncogene addiction and has been validated as a drug target for cancer therapy. Several strategies have been used to identify agents that target Stat3 in breast cancer but none has yet entered into clinical use. We used a high-throughput fluorescence microscopy search strategy to identify compounds in a drug-repositioning library (Prestwick library) that block ligand-induced nuclear translocation of Stat3 and identified piperlongumine (PL), a natural product isolated from the fruit of the pepper Piper longum. PL inhibited Stat3 nuclear translocation, inhibited ligand-induced and constitutive Stat3 phosphorylation, and modulated expression of multiple Stat3-regulated genes. Surface plasmon resonance assay revealed that PL directly inhibited binding of Stat3 to its phosphotyrosyl peptide ligand. Phosphoprotein antibody array analysis revealed that PL does not modulate kinases known to activate Stat3 such as Janus kinases, Src kinase family members or receptor tyrosine kinases. PL inhibited anchorage-independent and anchorage-dependent growth of multiple breast cancer cell lines having increased pStat3 or total Stat3, and induced apoptosis. PL also inhibited mammosphere formation by tumor cells from patient-derived xenografts. PL's antitumorigenic function was causally linked to its Stat3-inhibitory effect. PL was non-toxic in mice up to a dose of 30 mg/kg/day for 14 days and caused regression of breast cancer cell line xenografts in nude mice. Thus, PL represents a promising new agent for rapid entry into the clinic for use in treating breast cancer, as well as other cancers in which Stat3 has a role.

  11. Involvement of fish signal transducer and activator of transcription 3 (STAT3) in SGIV replication and virus induced paraptosis.

    PubMed

    Huang, Xiaohong; Huang, Youhua; Yang, Ying; Wei, Shina; Qin, Qiwei

    2014-12-01

    Signal transducer and activator of transcription 3 (STAT3) is an important transcription factor which plays crucial roles in immune regulation, inflammation, cell proliferation, transformation, and other physiological processes of the organism. In this study, a novel STAT3 gene from orange spotted grouper (Ec-STAT3) was cloned and characterized. Bioinformatic analysis revealed that full-length of Ec-STAT3 was 3105-bp long and contained a 280-bp 5'UTR, a 470-bp 3'UTR, and a 2355-bp open reading frame (ORF) that encoded a 784-amino acid peptide. The deduced protein of Ec-STAT3 showed 98% identity to that of turbot (Scophthalmus maximus). Amino acid alignment showed that Ec-STAT3 contained four conserved domains, including a protein interaction domain, a coiled coil domain, a DNA binding domain, and an SH2 domain. Quantitative real-time PCR analysis showed that the highest expression level was detected in the liver, followed by skin and spleen. After injection with Singapore grouper iridovirus (SGIV), the transcript of Ec-STAT3 in spleen was increased significantly. To further explore the function of Ec-STAT3, we investigated the roles of Ec-STAT3 in SGIV infection in vitro. Immune fluorescence analysis indicated that SGIV infection altered the distribution of phosphorylated Ec-STAT3 in nucleus, and a small part of phosphorylated Ec-STAT3 was associated with virus assembly sites, suggesting that Ec-STAT3 might be important for SGIV infection. Using STAT3 specific inhibitor, S3I-201, we found that inhibition of Ec-STAT3 activation decreased the SGIV replication significantly. Moreover, inhibition of Ec-STAT3 activation obviously altered SGIV infection induced cell cycle arrest and the expression of pro-survival genes, including Bcl-2, Bcl-xL and Bax inhibitor. Together, our results firstly demonstrated the critical roles of fish STAT3 in DNA virus replication and virus induced paraptosis, but also provided new insights into the mechanism of iridovirus pathogenesis.

  12. Enhanced Stat3 activation in POMC neurons provokes negative feedback inhibition of leptin and insulin signaling in obesity.

    PubMed

    Ernst, Marianne B; Wunderlich, Claudia M; Hess, Simon; Paehler, Moritz; Mesaros, Andrea; Koralov, Sergei B; Kleinridders, André; Husch, Andreas; Münzberg, Heike; Hampel, Brigitte; Alber, Jens; Kloppenburg, Peter; Brüning, Jens C; Wunderlich, F Thomas

    2009-09-16

    Leptin-stimulated Stat3 activation in proopiomelanocortin (POMC)-expressing neurons of the hypothalamus plays an important role in maintenance of energy homeostasis. While Stat3 activation in POMC neurons is required for POMC expression, the role of elevated basal Stat3 activation as present in the development of obesity has not been directly addressed. Here, we have generated and characterized mice expressing a constitutively active version of Stat3 (Stat3-C) in POMC neurons (Stat3-C(POMC) mice). On normal chow diet, these animals develop obesity as a result of hyperphagia and decreased POMC expression accompanied by central leptin and insulin resistance. This unexpected finding coincides with POMC-cell-specific, Stat3-mediated upregulation of SOCS3 expression inhibiting both leptin and insulin signaling as insulin-stimulated PIP(3) (phosphatidylinositol-3,4,5 triphosphate) formation and protein kinase B (AKT) activation in POMC neurons as well as with the fact that insulin's ability to hyperpolarize POMC neurons is largely reduced in POMC cells of Stat3-C(POMC) mice. These data indicate that constitutive Stat3 activation is not sufficient to promote POMC expression but requires simultaneous PI3K (phosphoinositide 3-kinase)-dependent release of FOXO1 repression. In contrast, upon exposure to a high-fat diet, food intake and body weight were unaltered in Stat3-C(POMC) mice compared with control mice. Taken together, these experiments directly demonstrate that enhanced basal Stat3 activation in POMC neurons as present in control mice upon high-fat feeding contributes to the development of hypothalamic leptin and insulin resistance.

  13. IL-6/STAT3 signaling pathway is activated in plasma cell mastitis.

    PubMed

    Liu, Yang; Zhang, Jian; Zhou, Yu-Hui; Jiang, Yi-Na; Zhang, Wei; Tang, Xiao-Jiang; Ren, Yu; Han, Shui-Ping; Liu, Pei-Jun; Xu, Jing; He, Jian-Jun

    2015-01-01

    Plasma cell mastitis (PCM), a particular type of mastitis, mainly occurs in females at nonpregnant and nonlactating stages. The infiltration of abundant plasma cells and lymphocytes is the hallmark of the disease. The incidence rate of PCM increased gradually and its pathogenesis remained unclear. In this study, we investigated the expression of IL-6/STAT3 signaling pathway, which is vital not only for the differentiation of plasma cells but also for survival of plasma cells and T lymphocytes, in 30 PCM cases, 10 acute mastitis cases and 10 normal breast tissues by immunohistochemical analysis. IL-6 level was significantly higher in PCM patients than in acute mastitis patients or normal group. The positive rate of IL-6 and p-STAT3 staining in PCM samples was 93.3% (28/30) and 70% (21/30), respectively, and there was a significant positive association between IL-6 and p-STAT3 staining (r=0.408, P=0.025). In PCM group, the rate of nipple retraction was 40% (12/30). Significantly higher IL-6 expression was found in PCM patients with nipple retraction than in other PCM patients. However, no significant difference in IL-6 or p-STAT3 staining was detected between PCM patients experiencing recurrence and other PCM patients. In addition, Bcl-2 level was higher in PCM patients than in acute mastitis patients or normal group, but there was no difference in Bcl-2 immunostaining between PCM patients experiencing recurrence and other PCM patients. These indicate that IL-6/STAT3 signaling is activated in PCM and may play an important role in the pathogenesis of PCM.

  14. STAT3 pathway regulates lung-derived brain metastasis initiating cell capacity through miR-21 activation.

    PubMed

    Singh, Mohini; Garg, Neha; Venugopal, Chitra; Hallett, Robin; Tokar, Tomas; McFarlane, Nicole; Mahendram, Sujeivan; Bakhshinyan, David; Manoranjan, Branavan; Vora, Parvez; Qazi, Maleeha; Arpin, Carolynn C; Page, Brent; Haftchenary, Sina; Rosa, David A; Lai, Ping-Shan; Gómez-Biagi, Rodolfo F; Ali, Ahmed M; Lewis, Andrew; Geletu, Mulu; Murty, Naresh K; Hassell, John A; Jurisica, Igor; Gunning, Patrick T; Singh, Sheila K

    2015-09-29

    Brain metastases (BM) represent the most common tumor to affect the adult central nervous system. Despite the increasing incidence of BM, likely due to consistently improving treatment of primary cancers, BM remain severely understudied. In this study, we utilized patient-derived stem cell lines from lung-to-brain metastases to examine the regulatory role of STAT3 in brain metastasis initiating cells (BMICs). Annotation of our previously described BMIC regulatory genes with protein-protein interaction network mapping identified STAT3 as a novel protein interactor. STAT3 knockdown showed a reduction in BMIC self-renewal and migration, and decreased tumor size in vivo. Screening of BMIC lines with a library of STAT3 inhibitors identified one inhibitor to significantly reduce tumor formation. Meta-analysis identified the oncomir microRNA-21 (miR-21) as a target of STAT3 activity. Inhibition of miR-21 displayed similar reductions in BMIC self-renewal and migration as STAT3 knockdown. Knockdown of STAT3 also reduced expression of known downstream targets of miR-21. Our studies have thus identified STAT3 and miR-21 as cooperative regulators of stemness, migration and tumor initiation in lung-derived BM. Therefore, STAT3 represents a potential therapeutic target in the treatment of lung-to-brain metastases.

  15. STAT3 Is Activated by JAK2 Independent of Key Oncogenic Driver Mutations in Non-Small Cell Lung Carcinoma

    PubMed Central

    Looyenga, Brendan D.; Hutchings, Danielle; Cherni, Irene; Kingsley, Chris

    2012-01-01

    Constitutive activation of STAT3 is a common feature in many solid tumors including non-small cell lung carcinoma (NSCLC). While activation of STAT3 is commonly achieved by somatic mutations to JAK2 in hematologic malignancies, similar mutations are not often found in solid tumors. Previous work has instead suggested that STAT3 activation in solid tumors is more commonly induced by hyperactive growth factor receptors or autocrine cytokine signaling. The interplay between STAT3 activation and other well-characterized oncogenic “driver” mutations in NSCLC has not been fully characterized, though constitutive STAT3 activation has been proposed to play an important role in resistance to various small-molecule therapies that target these oncogenes. In this study we demonstrate that STAT3 is constitutively activated in human NSCLC samples and in a variety of NSCLC lines independent of activating KRAS or tyrosine kinase mutations. We further show that genetic or pharmacologic inhibition of the gp130/JAK2 signaling pathway disrupts activation of STAT3. Interestingly, treatment of NSCLC cells with the JAK1/2 inhibitor ruxolitinib has no effect on cell proliferation and viability in two-dimensional culture, but inhibits growth in soft agar and xenograft assays. These data demonstrate that JAK2/STAT3 signaling operates independent of known driver mutations in NSCLC and plays critical roles in tumor cell behavior that may not be effectively inhibited by drugs that selectively target these driver mutations. PMID:22319590

  16. The Antiproliferative and Colony-suppressive Activities of STAT3 Inhibitors in Human Cancer Cells Is Compromised Under Hypoxic Conditions.

    PubMed

    Tian, Jilai; Xiao, Hui; Wu, Ruohan; Cao, Yang; Li, Chenglong; Xu, Ronald; Pierson, Christopher R; Finlay, Jonathan L; Yang, Fang; Gu, Ning; Lin, Jiayuh

    2017-02-01

    Constitutive activation of signal transducer and activator of transcription 3 (STAT3) has been indicated as a novel cancer drug target, since it plays an important role in diverse oncogenic processes including survival, cell proliferation and migration. Emerging STAT3 inhibitors have demonstrated efficacy in cancer cells and animal tumor models. It is well known that most solid tumors are characterized by hypoxia, but it is not clear if hypoxic conditions affect activity of STAT3 inhibitors. To examine this, two STAT3 inhibitors were tested to investigate their inhibitory efficacy in cancer cells grown under hypoxic conditions compared with those without hypoxia. Cell proliferation, colony formation and western blot assays were performed to examine the differences in the cell viability, proliferation and proteins in the STAT3 pathway. Under hypoxic conditions, the half-maximal inhibitory concentration values for both STAT3 inhibitors were increased compared to normoxic conditions in human pancreatic cancer, medulloblastoma and sarcoma cell lines. In addition, the ability of both STAT3 inhibitors to inhibit colony formation in pancreatic cancer, medulloblastoma and sarcoma cell lines was reduced under hypoxic conditions when compared to cells under normoxic conditions. Furthermore, there was an increase in phosphorylated STAT3 levels in cancer cells under hypoxic conditions, suggesting this may be one of the mechanisms of resistance. In summary, the results presented here provide a novel finding of STAT3 inhibitor activity under hypoxic conditions and indicate that under such low oxygen conditions, the anticancer efficacy of STAT3 inhibitors was indeed hampered. These results highlight the need to develop new therapeutic strategies to overcome the resistance of cancer cells to STAT3 inhibitors under hypoxic conditions. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  17. Push and release: TLR9 activation plus STAT3 blockade for systemic antitumor immunity.

    PubMed

    Kortylewski, Marcin; Kuo, Ya-Huei

    2014-01-01

    Proper immunostimulation ("push") and immune checkpoint blockade ("release") are both critical for the efficacy of anticancer immunotherapy. We have recently shown that activating Toll-like receptor 9 (TLR9) while specifically blocking signal transducer and activator of transcription 3 (STAT3) in leukemic cells enhances their immunogenicity, allowing for CD8(+) T cell-mediated tumor eradication. These findings underscore the therapeutic potential of such a "Push & Release" strategy against hematological malignancies.

  18. Placental STAT3 signaling is activated in women with polycystic ovary syndrome.

    PubMed

    Maliqueo, M; Sundström Poromaa, I; Vanky, E; Fornes, R; Benrick, A; Åkerud, H; Stridsklev, S; Labrie, F; Jansson, T; Stener-Victorin, E

    2015-03-01

    Does polycystic ovary syndrome (PCOS) in women without pregnancy complications affect placental signal transducer and activator of transcription 3 (STAT3) and mechanistic target of rapamycin (mTOR) signaling? Placental STAT3 signaling is activated but mTOR signaling is unaffected in PCOS. Women with PCOS have increased risk of poor pregnancy outcomes (e.g. restricted or accelerated fetal growth), indicating placental dysfunction. Placental STAT3 and mTOR pathways regulate placental function and indirectly affect fetal growth. In a case-control study, placental tissue and maternal blood were collected at delivery from 40 control pregnant women and 38 PCOS women with uncomplicated pregnancy. Women with PCOS were recruited at two medical centers and pregnant controls were recruited at one of these centers. Placental mRNA expression of genes encoding proteins related to steroid action, metabolic pathways and cytokines was analyzed by quantitative RT-PCR. Phosphorylated placental STAT3 (P-STAT3) and mTOR targets was measured by western blot. Levels of sex steroids in serum were determined by mass spectrometry. Placental P-STAT3 (Tyr-705) was increased in women with PCOS (P < 0.05) versus controls. Placental mTOR signaling was not affected in PCOS women when compared with controls. Circulating levels of androstenedione, androst-5-ene-3β, 17β-diol, testosterone, 5α-dihydrotestosterone and etiocholanolone glucuronide were higher and estradiol lower in women with PCOS than in controls (all P < 0.05). No correlation between sex steroid levels in serum and P-STAT3 was observed. Women with PCOS and pregnancy complications were excluded to avoid the confounding effects of placental pathologies, which could modify STAT3 and mTOR signaling. Moreover, 97.4% of women with PCOS in the study displayed oligoamenorrhea at diagnosis. Thus, the current findings could be restricted to PCOS women with the oligo-anovulatory phenotype without pregnancy complications. Phosphorylation of

  19. STAT3 activation in response to IL-6 is prolonged by the binding of IL-6 receptor to EGF receptor

    PubMed Central

    Wang, Yuxin; van Boxel-Dezaire, Anette H. H.; Cheon, HyeonJoo; Yang, Jinbo; Stark, George R.

    2013-01-01

    The activation of STAT3 by tyrosine phosphorylation, essential for normal development and for a normal inflammatory response to invading pathogens, is kept in check by negative regulators. Abnormal constitutive activation of STAT3, which contributes to the pathology of cancer and to chronic inflammatory diseases such as rheumatoid arthritis, occurs when negative regulation is not fully effective. SOCS3, the major negative regulator of STAT3, is induced by tyrosine-phosphorylated STAT3 and terminates STAT3 phosphorylation about 2 h after initial exposure of cells to members of the IL-6 family of cytokines by binding cooperatively to the common receptor subunit gp130 and JAKs 1 and 2. We show here that when the epidermal growth factor receptor (EGFR) is present and active, STAT3 is rephosphorylated about 4 h after exposure of cells to IL-6 or oncostatin M and remains active for many hours. Newly synthesized IL-6 drives association of the IL-6 receptor and gp130 with EGFR, leading to EGFR-dependent rephosphorylation of STAT3, which is not inhibited by the continued presence of SOCS3. This second wave of STAT3 activation supports sustained expression of a subset of IL-6-induced proteins, several of which play important roles in inflammation and cancer, in which both IL-6 secretion and EGFR levels are often elevated. PMID:24082147

  20. Sav1 Loss Induces Senescence and Stat3 Activation Coinciding with Tubulointerstitial Fibrosis.

    PubMed

    Leung, Janet Y; Wilson, Harper L; Voltzke, Kristin J; Williams, Lindsay A; Lee, Hyo Jin; Wobker, Sara E; Kim, William Y

    2017-06-15

    Tubulointerstitial fibrosis (TIF) is recognized as a final phenotypic manifestation in the transition from chronic kidney disease (CKD) to end-stage renal disease (ESRD). Here we show that conditional inactivation of Sav1 in the mouse renal epithelium resulted in upregulated expression of profibrotic genes and TIF. Loss of Sav1 induced Stat3 activation and a senescence-associated secretory phenotype (SASP) that coincided with the development of tubulointerstitial fibrosis. Treatment of mice with the YAP inhibitor verteporfin (VP) inhibited activation of genes associated with senescence, SASPs, and activation of Stat3 as well as impeded the development of fibrosis. Collectively, our studies offer novel insights into molecular events that are linked to fibrosis development from Sav1 loss and implicate VP as a potential pharmacological inhibitor to treat patients at risk for developing CKD and TIF. Copyright © 2017 American Society for Microbiology.

  1. Requirement of Stat3 but not Stat1 activation for epidermal growth factor receptor- mediated cell growth In vitro.

    PubMed Central

    Grandis, J R; Drenning, S D; Chakraborty, A; Zhou, M Y; Zeng, Q; Pitt, A S; Tweardy, D J

    1998-01-01

    Stimulation of epidermal growth factor receptor (EGFR) by ligand(s) leads to activation of signaling molecules including Stat1 and Stat3, two members of the signal transducers and activators of transcription (STAT) protein family. Activation of Stat1 and Stat3 was constitutive in transformed squamous epithelial cells, which produce elevated levels of TGF-alpha, and was enhanced by the addition of exogenous TGF-alpha. Targeting of Stat3 using antisense oligonucleotides directed against the translation initiation site, resulted in significant growth inhibition. In addition, cells stably transfected with dominant negative mutant Stat3 constructs failed to proliferate in vitro. In contrast, targeting of Stat1 using either antisense or dominant-negative strategies had no effect on cell growth. Thus, TGF-alpha/EGFR-mediated autocrine growth of transformed epithelial cells is dependent on activation of Stat3 but not Stat1. PMID:9769331

  2. T-cell growth transformation by herpesvirus saimiri is independent of STAT3 activation.

    PubMed

    Heck, Elke; Lengenfelder, Doris; Schmidt, Monika; Müller-Fleckenstein, Ingrid; Fleckenstein, Bernhard; Biesinger, Brigitte; Ensser, Armin

    2005-05-01

    Herpesvirus saimiri (saimirine herpesvirus 2) (HVS), a T-lymphotropic tumor virus, induces lymphoproliferative disease in several species of New World primates. In addition, strains of HVS subgroup C are able to transform T cells of Old World primates, including humans, to permanently growing T-cell lines. In concert with the Stp oncoprotein, the tyrosine kinase-interacting protein (Tip) of HVS C488 is required for T-cell transformation in vitro and lymphoma induction in vivo. Tip was previously shown to interact with the protein tyrosine kinase Lck. Constitutive activation of signal transducers and activators of transcription (STATs) has been associated with oncogenesis and has also been detected in HVS-transformed T-cell lines. Furthermore, Tip contains a putative consensus YXPQ binding motif for the SH2 (src homology 2) domains of STAT1 and STAT3. Tip tyrosine phosphorylation at this site was required for binding of STATs and induction of STAT-dependent transcription. Here we sought to address the relevance of STAT activation for transformation of human T cells by introducing a tyrosine-to-phenylalanine mutation in the YXPQ motif of Tip of HVS C488. Unexpectedly, the recombinant virus was still able to transform human T lymphocytes, but it had lost its capability to activate STAT3 as well as STAT1. This demonstrates that growth transformation by HVS is independent of STAT3 activation.

  3. HtrA2 suppresses autoimmune arthritis and regulates activation of STAT3

    PubMed Central

    Lee, Seung Hoon; Moon, Young-Mee; Seo, Hyeon-Beom; Kim, Se-Young; Kim, Eun-Kyung; Yi, Junyeong; Nam, Min-Kyung; Min, Jun-Ki; Park, Sung-Hwan; Rhim, Hyangshuk; Cho, Mi-La

    2016-01-01

    Rheumatoid arthritis (RA) is an autoimmune disease that is related to the induction of T helper (Th)17 cells, which secrete interleukin-17, and activation of the signal transducer and activator of transcription (STAT) 3. The expression of high-temperature requirement protein A (HtrA) 2, a serine protease involved in apoptosis, was decreased in RA patients nonresponsive to drug treatment of RA. The aim of this study was to determine whether overexpression of HtrA2 has a therapeutic effect on RA. Th17 differentiation, osteoclastogenesis, and lymphocyte activation are increased in motor neuron degeneration (mnd)2 mice, which lack HtrA2 activity because of a missense mutation (Ser276Cys) in the protease domain of HtrA2. The inhibitor of HtrA2 also increased Th17 differentiation. On the other hand, HtrA2 induced cleavage of STAT3 and overexpression of HtrA2 attenuated CIA in a mouse model. HtrA2 overexpression inhibited plaque development as well as the differentiation of Th17 in ApoE−/− mice after immunization with proteoglycans to induce a hyperlipidemia-based RA animal model. The therapeutic function of HtrA2 in inflammatory diseases is linked with Th17 development and the STAT3 pathway in splenocytes. These results suggest that HtrA2 participates in immunomodulatory activity where the upregulation of HtrA2 may shed light on therapeutic approaches to RA and hyperlipidemia. PMID:28008946

  4. Stat3 signaling is present and active during development of the central nervous system and eye of vertebrates.

    PubMed

    Yan, Ye; Bian, Wei; Xie, Zhiqin; Cao, Xinmin; Le Roux, Isabelle; Guillemot, Francois; Jing, Naihe

    2004-10-01

    Stat3, a member of the signal transducer and activator of transcription (STAT) family, plays a central role in mediating cell growth, differentiation, and survival signals. In this report, we show that Stat3 immunoreactivity was localized to specific regions in the developing mouse brain, neural tube, and eye from embryonic day 10.5 to postnatal day 0. The active form of Stat3 protein, which is phosphorylated on tyrosine 705 (pYStat3), was also found in the developing neural tube with more restricted distribution. An in ovo chick embryo electroporation assay showed that the endogenous chick Stat3 could drive consensus sis-inducible element-directed reporter gene expression. These results demonstrate that the active Stat3 protein is present and might play a role during the development of the central nervous system and eye.

  5. Signal Transducer and Activator of Transcription (STAT)-3 Activates Nuclear Factor (NF)-κB in Chronic Lymphocytic Leukemia Cells

    PubMed Central

    Liu, Zhiming; Hazan-Halevy, Inbal; Harris, David M.; Li, Ping; Ferrajoli, Alessandra; Faderl, Stefan; Keating, Michael J.; Estrov, Zeev

    2014-01-01

    Nuclear factor (NF)-κB plays a major role in the pathogenesis of B-cell neoplasms. A broad array of mostly extracellular stimuli has been reported to activate NF-κB, to various degrees, in chronic lymphocytic leukemia (CLL) cells. Because CLL cells harbor high levels of unphosphorylated (U) signal transducer and activator of transcription (STAT)-3 protein and U-STAT3 was reported to activate NF-κB, we sought to determine whether U-STAT3 activates NF-κB in CLL. Using the electrophoretic mobility shift assay (EMSA) we studied peripheral blood low-density cells from 15 patients with CLL and found that CLL cell nuclear extracts from all the samples bound to an NF-κB DNA probe, suggesting that NF-κB is constitutively activated in CLL. Immunoprecipitation studies showed that STAT3 bound NF-κB p65, and confocal microscopy studies detected U-STAT3/NF-κB complexes in the nuclei of CLL cells, thereby confirming these findings. Furthermore, infection of CLL cells with retroviral STAT3-shRNA attenuated the binding of NF-κB to DNA, as assessed by EMSA, and downregulated mRNA levels of NF-κB-regulated genes, as assessed by quantitative polymerase chain reaction. Taken together, our data suggest that U-STAT3 binds to the NF-κB p50/p65 dimers and that the U-STAT3/NF-κB complexes bind to DNA and activate NF-κB-regulated genes in CLL cells. PMID:21364020

  6. Stat3 inhibitor Stattic exhibits potent antitumor activity and induces chemo- and radio-sensitivity in nasopharyngeal carcinoma.

    PubMed

    Pan, Yunbao; Zhou, Fuling; Zhang, Ronghua; Claret, Francois X

    2013-01-01

    Nasopharyngeal carcinoma (NPC) is an Epstein-Barr virus-associated malignancy most common in East Asia, Africa and Alaska. Radiotherapy and cisplatin-based chemotherapy are the main treatment options. Unfortunately, disease response to concurrent chemoradiotherapy varies among patients with NPC, and many cases are resistant to cisplatin and radiotherapy. Signal transducer and activator of transcription 3 (Stat3) has been implicated in the development and progression of various solid tumors. In this study, we assessed the activation and expression of Stat3 in NPC cells. We found that Stat3 was activated and could be blocked by the small molecule inhibitor Stattic. The inhibition of Stat3 in NPC cells by Stattic decreased the expression of cyclin D1 in a dose- and time-dependent manner. Thus, Stattic was used to target Stat3 in NPC cell lines. We found that Stattic could inhibit cell viability and proliferation in NPC cells and significantly induced apoptosis. Additionally, Stat3 transfection attenuated, whereas Stat3 knockdown enhanced, the effects of Stattic upon cell viability inhibition and apoptosis induction. Furthermore, Stattic sensitized NPC cells to cisplatin and ionizing radiation (IR) by preventing cell proliferation and inducing apoptosis. Taken together, Stattic inhibit Stat3 and display antitumor effect in NPC, and enhanced chemosensitivity and radiosensitivity in NPC. Therefore, our findings provide the base for more rational approaches to treat NPC in the clinic.

  7. Sugiol inhibits STAT3 activity via regulation of transketolase and ROS-mediated ERK activation in DU145 prostate carcinoma cells.

    PubMed

    Jung, Seung-Nam; Shin, Dae-Seop; Kim, Hye-Nan; Jeon, Yoon Jung; Yun, Jieun; Lee, Yu-Jin; Kang, Jong Soon; Han, Dong Cho; Kwon, Byoung-Mog

    2015-09-01

    Signal transducer and activator of transcription 3 (STAT3) is constitutively activated in various human cancers and has been used as a therapeutic target for tumors. This study screened natural products to identify compounds that inhibit STAT3 activity using a STAT3-dependent luciferase reporter system. Sugiol was identified as a compound that decreased luciferase activity in a dose-dependent manner. Sugiol specifically inhibited STAT3 phosphorylation at Tyr-705 in DU145 prostate cells, and this inhibition was independent of the STAT3 upstream kinase. Sugiol induced cell cycle arrest and decreased the expression levels of STAT3 target genes, such as cyclin D1, cyclin A, and survivin. Notably, we observed that sugiol interacted with transketolase, an enzyme in central metabolism, and increased ROS levels leading to the activation of ERK, which inhibits STAT3 activity. The protein phosphatase MEG2 was also responsible for sugiol-induced STAT3 dephosphorylation. The inhibitory effect of sugiol on cell growth was confirmed using the DU145 mouse xenograft model. We propose that sugiol inhibits STAT3 activity through a mechanism that involves the inhibition of transketolase, which leads to increased ROS levels and MEG2 activation in DU145 cells. Therefore, sugiol is the first compound regulating STAT3 activity via modulation of cancer metabolic pathway and we suggest the use of sugiol as an inhibitor of the STAT3 pathway for the treatment of human solid tumors with activated STAT3. Copyright © 2015 Elsevier Inc. All rights reserved.

  8. A novel small molecule agent displays potent anti-myeloma activity by inhibiting the JAK2-STAT3 signaling pathway

    PubMed Central

    Zhu, Jingyu; Xu, Yujia; Wang, Siyu; Xu, Xin; Ji, Peng; Yu, Yang; Cao, Biyin; Han, Kunkun; Hou, Tingjun; Xu, Zhuan; Kong, Yan; Jiang, Gaofeng; Tang, Xiaowen; Qiao, Chunhua; Mao, Xinliang

    2016-01-01

    The oncogenic STAT3 signaling pathway is emerging as a promising target for the treatment of multiple myeloma (MM). In the present study, we identified a novel STAT3 inhibitor SC99 in a target-based high throughput screen. SC99 inhibited JAK2-STAT3 activation but had no effects on other transcription factors such as NF-κB, and kinases such as AKT, ERK, and c-Src that are in association with STAT3 signaling pathway. Furthermore, SC99 downregulated the expression of STAT3-modulated genes, including Bcl-2, Bcl-xL, VEGF, cyclin D2, and E2F-1. By inhibiting the STAT3 signaling, SC99 induced MM cell apoptosis which could be partly abolished by the ectopic expression of STAT3. Furthermore, SC99 displayed potent anti-MM activity in two independent MM xenograft models in nude mice. Oral administration of SC99 led to marked decrease of tumor growth within 10 days at a daily dosage of 30 mg/kg, but did not raise toxic effects. Taken together, this study identified a novel oral JAK2/STAT3 inhibitor that could be developed as an anti-myeloma agent. PMID:26814430

  9. Bioinformatic analyses reveal a distinct Notch activation induced by STAT3 phosphorylation in the mesenchymal subtype of glioblastoma.

    PubMed

    Cheng, Wen; Zhang, Chuanbao; Ren, Xiufang; Jiang, Yang; Han, Sheng; Liu, Yang; Cai, Jinquan; Li, Mingyang; Wang, Kuanyu; Liu, Yanwei; Hu, Huimin; Li, Qingbin; Yang, Pei; Bao, Zhaoshi; Wu, Anhua

    2017-01-01

    OBJECTIVE Glioblastoma (GBM) is the most common and lethal type of malignant glioma. The Cancer Genome Atlas divides the gene expression-based classification of GBM into classical, mesenchymal, neural, and proneural subtypes, which is important for understanding GBM etiology and for designing effective personalized therapy. Signal transducer and activator of transcription 3 (STAT3), a critical transcriptional activator in tumorigenesis, is persistently phosphorylated and associated with an unfavorable prognosis in GBM. Although a set of specific targets has been identified, there have been no systematic analyses of STAT3 signaling based on GBM subtype. METHODS This study compared STAT3-associated messenger RNA, protein, and microRNA expression profiles across different subtypes of GBM. RESULTS The analyses revealed a prominent role for STAT3 in the mesenchymal but not in other GBM subtypes, which can be reliably used to classify patients with mesenchymal GBM into 2 groups according to phosphorylated STAT3 expression level. Differentially expressed genes suggest an association between Notch and STAT3 signaling in the mesenchymal subtype. Their association was validated in the U87 cell, a malignant glioma cell line annotated as mesenchymal subtype. Specific associated proteins and microRNAs further profile the STAT3 signaling among GBM subtypes. CONCLUSIONS These findings suggest a prominent role for STAT3 signaling in mesenchymal GBM and highlight the importance of identifying signaling pathways that contribute to specific cancer subtypes.

  10. Smad7 enables STAT3 activation and promotes pluripotency independent of TGF-β signaling.

    PubMed

    Yu, Yi; Gu, Shuchen; Li, Wenjian; Sun, Chuang; Chen, Fenfang; Xiao, Mu; Wang, Lei; Xu, Dewei; Li, Ye; Ding, Chen; Xia, Zongping; Li, Yi; Ye, Sheng; Xu, Pinglong; Zhao, Bin; Qin, Jun; Chen, Ye-Guang; Lin, Xia; Feng, Xin-Hua

    2017-09-05

    Smad7 is a negative feedback product of TGF-β superfamily signaling and fine tunes a plethora of pleiotropic responses induced by TGF-β ligands. However, its noncanonical functions independent of TGF-β signaling remain to be elucidated. Here, we show that Smad7 activates signal transducers and activators of transcription 3 (STAT3) signaling in maintaining mouse embryonic stem cell pluripotency in a manner independent of the TGF-β receptors, yet dependent on the leukemia inhibitory factor (LIF) coreceptor glycoprotein 130 (gp130). Smad7 directly binds to the intracellular domain of gp130 and disrupts the SHP2-gp130 or SOCS3-gp130 complex, thereby amplifying STAT3 activation. Consequently, Smad7 facilitates LIF-mediated self-renewal of mouse ESCs and is also critical for induced pluripotent stem cell reprogramming. This finding illustrates an uncovered role of the Smad7-STAT3 interplay in maintaining cell pluripotency and also implicates a mechanism involving Smad7 underlying cytokine-dependent regulation of cancer and inflammation.

  11. Avicin D: A Protein Reactive Plant Isoprenoid Dephosphorylates Stat 3 by Regulating Both Kinase and Phosphatase Activities

    PubMed Central

    Haridas, Valsala; Nishimura, Goshi; Xu, Zhi-Xiang; Connolly, Fiona; Hanausek, Margaret; Walaszek, Zbigniew; Zoltaszek, Robert; Gutterman, Jordan U.

    2009-01-01

    Avicins, a class of electrophilic triterpenoids with pro-apoptotic, anti-inflammatory and antioxidant properties, have been shown to induce redox-dependant post-translational modification of cysteine residues to regulate protein function. Based on (a) the cross-talk that occurs between redox and phosphorylation processes, and (b) the role of Stat3 in the process of apoptosis and carcinogenesis, we chose to study the effects of avicins on the processes of phosphorylation/dephosphorylation in Stat3. Avicins dephosphorylate Stat3 in a variety of human tumor cell lines, leading to a decrease in the transcriptional activity of Stat3. The expression of Stat3-regulated proteins such as c-myc, cyclin D1, Bcl2, survivin and VEGF were reduced in response to avicin treatment. Underlying avicin-induced dephosphorylation of Stat3 was dephosphorylation of JAKs, as well as activation of protein phosphatase-1. Downregulation of both Stat3 activity and expression of Stat 3-controlled pro-survival proteins, contributes to the induction of apoptosis in avicin treated tumor cells. Based on the role of Stat3 in inflammation and wounding, and the in vivo inhibition of VEGF by avicins in a mouse skin carcinogenesis model, it is likely that avicin-induced inhibition of Stat3 activity results in the suppression of the pro-inflammatory and pro-oxidant stromal environment of tumors. Activation of PP-1, which also acts as a cellular economizer, combined with the redox regulation by avicins, can aid in redirecting metabolism from growth promoting anabolic to energy sparing pathways. PMID:19440292

  12. Inhibition of Signal Transducer and Activator of Transcription 3 (STAT3) reduces neonatal hypoxic-ischaemic brain damage.

    PubMed

    Hristova, Mariya; Rocha-Ferreira, Eridan; Fontana, Xavier; Thei, Laura; Buckle, Rheanan; Christou, Melina; Hompoonsup, Supanida; Gostelow, Naomi; Raivich, Gennadij; Peebles, Donald

    2016-03-01

    Hypoxic-ischaemic encephalopathy is a leading cause of child death, with high mortality and morbidity, including cerebral palsy, epilepsy and cognitive disabilities. Hypoxia-ischaemia (HI) strongly up-regulates Signal Transducer and Activator of Transcription 3 (STAT3) in the immature brain. Our aim was to establish whether STAT3 up-regulation is associated with neonatal HI-brain damage and evaluate the phosphorylated STAT3-contribution from different cell types in eliciting damage. We subjected postnatal day seven mice to unilateral carotid artery ligation followed by 60 min hypoxia. Neuronal STAT3-deletion reduced cell death, tissue loss, microglial and astroglial activation in all brain regions. Astroglia-specific STAT3-deletion also reduced cell death, tissue loss and microglial activation, although not as strongly as the deletion in neurons. Systemic pre-insult STAT3-blockade at tyrosine 705 (Y705) with JAK2-inhibitor WP1066 reduced microglial and astroglial activation to a more moderate degree, but in a pattern similar to the one produced by the cell-specific deletions. Our results suggest that STAT3 is a crucial factor in neonatal HI-brain damage and its removal in neurons or astrocytes, and, to some extent, inhibition of its phosphorylation via JAK2-blockade reduces inflammation and tissue loss. Overall, the protective effects of STAT3 inactivation make it a possible target for a therapeutic strategy in neonatal HI. Current data show that neuronal and astroglial STAT3 molecules are involved in the pathways underlying cell death, tissue loss and gliosis following neonatal hypoxia-ischaemia, but differ with respect to the target of their effect. Y705-phosphorylation contributes to hypoxic-ischaemic histopathology. Protective effects of STAT3 inactivation make it a possible target for a therapeutic strategy in neonatal hypoxia-ischaemia. © 2015 The Authors. Journal of Neurochemistry published by John Wiley & Sons Ltd on behalf of International Society for

  13. Atrazine promotes RM1 prostate cancer cell proliferation by activating STAT3 signaling.

    PubMed

    Hu, Kebang; Tian, Yong; Du, Yanwei; Huang, Liandi; Chen, Junyu; Li, Na; Liu, Wei; Liang, Zuowen; Zhao, Lijing

    2016-05-01

    Atrazine, a widely used pesticide, is frequently detected in soil and surface water, which alarms epidemiologists and medical professionals because of its potential deleterious effects on health. Indeed, atrazine is a potent endocrine disruptor that increases aromatase expression in some human cancer cell lines. Both animal and human studies have suggested that atrazine is possibly carcinogenic, although discrepant results have been reported. In this study, RM1 cells were used to explore the atrazine effects on prostate cancer. Proliferation, migration and invasion of RM1 cells were assessed by colony formation, wound-healing and invasion assays, respectively, after in vitro exposure to atrazine. In addition, an RM1 cell xenograft model was generated to evaluate the effects of atrazine in vivo. To explore the molecular mechanisms, qRT‑PCR, immunohistochemistry, and western blot analyses were employed to detect mRNA and protein levels of STAT3 signaling and cell cycle related proteins, including p53, p21, cyclin B1 and cyclin D1. Interestingly, RM1 cell proliferation was increased after treatment with atrazine, concomitantly with STAT3 signaling activation. These results suggest that atrazine promotes RM1 cell growth in vitro and in vivo by activating STAT3 signaling.

  14. Mesenchymal stem cells promote osteosarcoma cell survival and drug resistance through activation of STAT3

    PubMed Central

    Liu, Shen; Wang, Lei; Fan, Qiming; Hao, Yongqiang; Fan, Cunyi; Tang, Ting-Ting

    2016-01-01

    Increasing evidence suggests that the tumor microenvironment plays a key role in the development of drug resistant tumor cells. In this study, we tried to determine whether the mesenchymal stem cells (MSCs) in the tumor microenvironment contribute to the increased chemoresistance of osteosarcoma. We found that exposure of Saos-2 and U2-OS cells to MSCs conditioned medium (CM) increased the viable cells in the presence of therapeutic concentrations of doxorubicin or cisplatin. Meanwhile, the MSC CM-associated pro-proliferative effects were accompanied by reduced caspase 3/7 activity and Annexin V binding. We confirmed that STAT3 activation by IL-6 regulates MSCs-induced chemoresistance. Blockade of this signal re-sensitized drug-resistant Saos-2 cells to drug treatment. Using a osteosarcoma mouse model with co-injection of MSCs with Saos-2cells, we found that inhibition of STAT3 prolonged the survival time of tumor bearing mice by suppressing tumor growth and increasing the sensitivity of tumor cells to doxorubicin. Finally, we demonstrated that increased expression of p-STAT3, multidrug resistance protein (MRP) and P-glycoprotein (MDR-1) was associated with high chemotherapy resistance in clinical osteosarcoma samples. Collectively, our findings suggest that MSCs within the tumor microenvironment may represent a new target to enhance chemotherapeutic efficacy in osteosarcoma patients. PMID:27340780

  15. Nutritional control of IL-23/Th17-mediated autoimmune disease through HO-1/STAT3 activation

    PubMed Central

    Brück, Jürgen; Holstein, Julia; Glocova, Ivana; Seidel, Ursula; Geisel, Julia; Kanno, Toshio; Kumagai, Jin; Mato, Naoko; Sudowe, Stephan; Widmaier, Katja; Sinnberg, Tobias; Yazdi, Amir S.; Eberle, Franziska C.; Hirahara, Kiyoshi; Nakayama, Toshinori; Röcken, Martin; Ghoreschi, Kamran

    2017-01-01

    The nutritional curcumin (CUR) is beneficial in cell-mediated autoimmune diseases. The molecular mechanisms underlying this food-mediated silencing of inflammatory immune responses are poorly understood. By investigating antigen-specific immune responses we found that dietary CUR impairs the differentiation of Th1/Th17 cells in vivo during encephalomyelitis and instead promoted Th2 cells. In contrast, feeding CUR had no inhibitory effect on ovalbumin-induced airway inflammation. Mechanistically, we found that CUR induces an anti-inflammatory phenotype in dendritic cells (DC) with enhanced STAT3 phosphorylation and suppressed expression of Il12b and Il23a. On the molecular level CUR readily induced NRF2-sensitive heme oxygenase 1 (HO-1) mRNA and protein in LPS-activated DC. HO-1 enhanced STAT3 phosphorylation, which enriched to Il12b and Il23a loci and negatively regulated their transcription. These findings demonstrate the underlying mechanism through which a nutritional can interfere with the immune response. CUR silences IL-23/Th17-mediated pathology by enhancing HO-1/STAT3 interaction in DC. PMID:28290522

  16. Helicobacter pylori Activates IL-6-STAT3 Signaling in Human Gastric Cancer Cells: Potential Roles for Reactive Oxygen Species.

    PubMed

    Piao, Juan-Yu; Lee, Hee Geum; Kim, Su-Jung; Kim, Do-Hee; Han, Hyeong-Jun; Ngo, Hoang-Kieu-Chi; Park, Sin-Aye; Woo, Jeong-Hwa; Lee, Jeong-Sang; Na, Hye-Kyung; Cha, Young-Nam; Surh, Young-Joon

    2016-10-01

    Recent studies have shown that Helicobacter pylori (H. pylori) activates signal transducer and activator of transcription 3 (STAT3) that plays an important role in gastric carcinogenesis. However, the molecular mechanism underlying H. pylori-mediated STAT3 activation is still not fully understood. In this study, we investigated H. pylori-induced activation of STAT3 signaling in AGS human gastric cancer cells and the underlying mechanism. AGS cells were cocultured with H. pylori, and STAT3 activation was assessed by Western blot analysis, electrophoretic mobility shift assay and immunocytochemistry. To demonstrate the involvement of reactive oxygen species (ROS) in H. pylori-activated STAT3 signaling, the antioxidant N-acetylcysteine was utilized. The expression and production of interleukin-6 (IL-6) were measured by reverse-transcription polymerase chain reaction and enzyme-linked immunosorbent assay (ELISA), respectively. The interaction between IL-6 and IL-6 receptor (IL-6R) was determined by the immunoprecipitation assay. H. pylori activates STAT3 as evidenced by increases in phosphorylation on Tyr(705) , nuclear localization, DNA binding and transcriptional activity of this transcription factor. The nuclear translocation of STAT3 was also observed in H. pylori-inoculated mouse stomach. In the subsequent study, we found that H. pylori-induced STAT3 phosphorylation was dependent on IL-6. Notably, the increased IL-6 expression and the IL-6 and IL-6R binding were mediated by ROS produced as a consequence of H. pylori infection. H. pylori-induced STAT3 activation is mediated, at least in part, through ROS-induced upregulation of IL-6 expression. These findings provide a novel molecular mechanism responsible for H. pylori-induced gastritis and gastric carcinogenesis. © 2016 John Wiley & Sons Ltd.

  17. Berberine suppresses tumorigenicity and growth of nasopharyngeal carcinoma cells by inhibiting STAT3 activation induced by tumor associated fibroblasts

    PubMed Central

    2013-01-01

    Background Cortidis rhizoma (Huanglian) and its major therapeutic component, berberine, have drawn extensive attention in recent years for their anti-cancer properties. Growth inhibitory effects of berberine on multiple types of human cancer cells have been reported. Berberine inhibits invasion, induces cell cycle arrest and apoptosis in human cancer cells. The anti-inflammatory property of berberine, involving inhibition of Signal Transducer and Activator of Transcription 3 (STAT3) activation, has also been documented. Methods In this study, we have examined the effects of berberine on tumorigenicity and growth of nasopharyngeal carcinoma (NPC) cells and their relationship to STAT3 signaling using both in vivo and in vitro models. Results Berberine effectively inhibited the tumorigenicity and growth of an EBV-positive NPC cell line (C666-1) in athymic nude mice. Inhibition of tumorigenic growth of NPC cells in vivo was correlated with effective inhibition of STAT3 activation in NPC cells inside the tumor xenografts grown in nude mice. In vitro, berberine inhibited both constitutive and IL-6-induced STAT3 activation in NPC cells. Inhibition of STAT3 activation by berberine induced growth inhibition and apoptotic response in NPC cells. Tumor-associated fibroblasts were found to secret IL-6 and the conditioned medium harvested from the fibroblasts also induced STAT3 activation in NPC cells. Furthermore, STAT3 activation by conditioned medium of tumor-associated fibroblasts could be blocked by berberine or antibodies against IL-6 and IL-6R. Conclusions Our observation that berberine effectively inhibited activation of STAT3 induced by tumor-associated fibroblasts suggests a role of berberine in modulating the effects of tumor stroma on the growth of NPC cells. The effective inhibition of STAT3 activation in NPC cells by berberine supports its potential use in the treatment of NPC. PMID:24380387

  18. STAT3 activation in Th17 and Th22 cells controls IL-22-mediated epithelial host defense during infectious colitis.

    PubMed

    Backert, Ingo; Koralov, Sergei B; Wirtz, Stefan; Kitowski, Vera; Billmeier, Ulrike; Martini, Eva; Hofmann, Katharina; Hildner, Kai; Wittkopf, Nadine; Brecht, Katrin; Waldner, Maximilian; Rajewsky, Klaus; Neurath, Markus F; Becker, Christoph; Neufert, Clemens

    2014-10-01

    The Citrobacter rodentium model mimics the pathogenesis of infectious colitis and requires sequential contributions from different immune cell populations, including innate lymphoid cells (ILCs) and CD4(+) lymphocytes. In this study, we addressed the role of STAT3 activation in CD4(+) cells during host defense in mice against C. rodentium. In mice with defective STAT3 in CD4(+) cells (Stat3(ΔCD4)), the course of infection was unchanged during the innate lymphoid cell-dependent early phase, but significantly altered during the lymphocyte-dependent later phase. Stat3(ΔCD4) mice exhibited intestinal epithelial barrier defects, including downregulation of antimicrobial peptides, increased systemic distribution of bacteria, and prolonged reduction in the overall burden of C. rodentium infection. Immunomonitoring of lamina propria cells revealed loss of virtually all IL-22-producing CD4(+) lymphocytes, suggesting that STAT3 activation was required for IL-22 production not only in Th17 cells, but also in Th22 cells. Notably, the defective host defense against C. rodentium in Stat3(∆CD4) mice could be fully restored by specific overexpression of IL-22 through a minicircle vector-based technology. Moreover, expression of a constitutive active STAT3 in CD4(+) cells shaped strong intestinal epithelial barrier function in vitro and in vivo through IL-22, and it promoted protection from enteropathogenic bacteria. Thus, our work indicates a critical role of STAT3 activation in Th17 and Th22 cells for control of the IL-22-mediated host defense, and strategies expanding STAT3-activated CD4(+) lymphocytes may be considered as future therapeutic options for improving intestinal barrier function in infectious colitis. Copyright © 2014 by The American Association of Immunologists, Inc.

  19. STAT3 activation is required for the antiapoptotic effects of prolactin in cervical cancer cells.

    PubMed

    Ramírez de Arellano, Adrián; Lopez-Pulido, Edgar I; Martínez-Neri, Priscila A; Estrada Chávez, Ciro; González Lucano, Renee; Fafutis-Morris, Mary; Aguilar-Lemarroy, A; Muñoz-Valle, José F; Pereira-Suárez, Ana Laura

    2015-01-01

    Prolactin (PRL) has been implicated in the development of different types of cancer. However, signaling pathways might be activated depending on various forms of prolactin receptor (PRLR). JAK/STAT is an important pathway associated with PRL effects. The activation of JAK/STAT pathway might activate antiapoptotic genes that could importantly lead to progression of tumorigenesis. Recently, we have reported that PRL is associated with cell survival by inhibition of apoptosis and the precise activated signaling pathways for this process are still questioned. The purpose of this study was to evaluate the activation of different signaling pathways in response to PRL as well as to identify the induction of antiapoptotic genes. Cervical cancer cell lines HeLa, SiHa and C-33 A were stimulated with PRL (200 ng/mL) for 30 and 60 min and non stimulated cells were used to measure basal protein expression. Inhibition assays were performed by using Jak2 specific inhibitor AG490, either alone or in combination with PRL for 48 h. Western blot were carried out to evaluate protein induction of the different signaling pathways and antiapoptotic proteins. Significant effects were determined by using ANOVA test. STAT3 was significantly activated in cervical cancer lines in comparison with non-tumorigenic keratinocytes HaCaT. No significant differences were found when analyzing MAPK and PI3K signaling pathways. An increase of antiapoptotic genes Bcl-xl, Bcl-2, survivin and Mcl-1 was observed after stimulus with PRL; however, after inhibition with AG490, the induction of antiapoptotic genes was decreased. Our data suggests that STAT3 is an important signaling pathway activated by PRL in cervical cancer cells and it modulates the induction of antiapoptotic genes. Blocking STAT3 could represent a possible therapeutic strategy in cervical cancer.

  20. Activation of the GP130-STAT3 axis and its potential implications in nonalcoholic fatty liver disease

    PubMed Central

    Min, Hae-Ki; Mirshahi, Faridoddin; Verdianelli, Aurora; Pacana, Tommy; Patel, Vaishali; Park, Chun-Geon; Choi, Aejin; Lee, Jeong-Hoon; Park, Chung-Berm; Ren, Shunlin

    2015-01-01

    The status of the GP130-STAT3 signaling pathway in humans with nonalcoholic fatty liver disease (NAFLD) and its relevance to disease pathogenesis are unknown. The expression of the gp130-STAT3 axis and gp130 cytokine receptors were studied in subjects with varying phenotypes of NAFLD including nonalcoholic steatohepatitis (NASH) and compared with lean and weight-matched controls without NAFLD. Gp130 and its downstream signaling element (Tyk2 and STAT3) expression were inhibited in obese controls whereas they were increased in NAFLD. IL-6 levels were increased in NASH and correlated with gp130 expression (P < 0.01). Palmitate inhibited gp130-STAT3 expression and signaling. IL-6 and palmitate inhibited hepatic insulin signaling via STAT3-dependent and independent mechanisms, respectively. STAT3 overexpression reversed palmitate-induced lipotoxicity by increasing autophagy (ATG7) and decreasing endoplasmic reticulum stress. These data demonstrate that the STAT3 pathway is activated in NAFLD and can worsen insulin resistance while protecting against other lipotoxic mechanisms of disease pathogenesis. PMID:25747354

  1. CD45 Phosphatase Inhibits STAT3 Transcription Factor Activity in Myeloid Cells and Promotes Tumor-Associated Macrophage Differentiation.

    PubMed

    Kumar, Vinit; Cheng, Pingyan; Condamine, Thomas; Mony, Sridevi; Languino, Lucia R; McCaffrey, Judith C; Hockstein, Neil; Guarino, Michael; Masters, Gregory; Penman, Emily; Denstman, Fred; Xu, Xiaowei; Altieri, Dario C; Du, Hong; Yan, Cong; Gabrilovich, Dmitry I

    2016-02-16

    Recruitment of monocytic myeloid-derived suppressor cells (MDSCs) and differentiation of tumor-associated macrophages (TAMs) are the major factors contributing to tumor progression and metastasis. We demonstrated that differentiation of TAMs in tumor site from monocytic precursors was controlled by downregulation of the activity of the transcription factor STAT3. Decreased STAT3 activity was caused by hypoxia and affected all myeloid cells but was not observed in tumor cells. Upregulation of CD45 tyrosine phosphatase activity in MDSCs exposed to hypoxia in tumor site was responsible for downregulation of STAT3. This effect was mediated by the disruption of CD45 protein dimerization regulated by sialic acid. Thus, STAT3 has a unique function in the tumor environment in controlling the differentiation of MDSC into TAM, and its regulatory pathway could be a potential target for therapy.

  2. Eudesmane-type sesquiterpenoids from Salvia plebeia inhibit IL-6-induced STAT3 activation.

    PubMed

    Jang, Hyun-Jae; Oh, Hyun-Mee; Hwang, Joo Tae; Kim, Mi-Hwa; Lee, Soyoung; Jung, Kyungsook; Kim, Young-Ho; Lee, Seung Woong; Rho, Mun-Chual

    2016-10-01

    Seven eudesmane-type sesquiterpenoid lactones and the known plebeiolide C were isolated from an ethanol-soluble extract of the aerial parts of Salvia plebeia R. Br. Their structures were determined via NMR and MS, and their absolute configurations were elucidated using ECD, and X-ray crystallographic analysis, as well as the modified Mosher ester method. All isolates were evaluated for their inhibitory effects on IL6-induced STAT3 promoter activation in stably transfected Hep3B cells. Of these isolates, eudebeiolide D exhibited an inhibitory effect with the IC50 value of 1.1 μM.

  3. STAT3 activation in pressure-overloaded feline myocardium: role for integrins and the tyrosine kinase BMX.

    PubMed

    Willey, Christopher D; Palanisamy, Arun P; Johnston, Rebecca K; Mani, Santhosh K; Shiraishi, Hirokazu; Tuxworth, William J; Zile, Michael R; Balasubramanian, Sundaravadivel; Kuppuswamy, Dhandapani

    2008-06-27

    Growth, survival and cytoskeletal rearrangement of cardiomyocytes are critical for cardiac hypertrophy. Signal transducer and activator of transcription-3 (STAT3) activation is an important cardioprotective factor associated with cardiac hypertrophy. Although STAT3 activation has been reported via signaling through Janus Kinase 2 (JAK2) in several cardiac models of hypertrophy, the importance of other nonreceptor tyrosine kinases (NTKs) has not been explored. Utilizing an in vivo feline right ventricular pressure-overload (RVPO) model of hypertrophy, we demonstrate that in 48 h pressure-overload (PO) myocardium, STAT3 becomes phosphorylated and redistributed to detergent-insoluble fractions with no accompanying JAK2 activation. PO also caused increased levels of phosphorylated STAT3 in both cytoplasmic and nuclear fractions. To investigate the role of other NTKs, we used our established in vitro cell culture model of hypertrophy where adult feline cardiomyocytes are embedded three-dimensionally (3D) in type-I collagen and stimulated with an integrin binding peptide containing an Arg-Gly-Asp (RGD) motif that we have previously shown to recapitulate the focal adhesion complex (FAC) formation of 48 h RVPO. RGD stimulation of adult cardiomyocytes in vitro caused both STAT3 redistribution and activation that were accompanied by the activation and redistribution of c-Src and the TEC family kinase, BMX, but not JAK2. However, infection with dominant negative c-Src adenovirus was unable to block RGD-stimulated changes on either STAT3 or BMX. Further analysis in vivo in 48 h PO myocardium showed the presence of both STAT3 and BMX in the detergent-insoluble fraction with their complex formation and phosphorylation. Therefore, these studies indicate a novel mechanism of BMX-mediated STAT3 activation within a PO model of cardiac hypertrophy that might contribute to cardiomyocyte growth and survival.

  4. PTEN ameliorates autoimmune arthritis through down-regulating STAT3 activation with reciprocal balance of Th17 and Tregs

    PubMed Central

    Lee, Seung Hoon; Park, Jin-Sil; Byun, Jae-Kyung; Jhun, JooYeon; Jung, KyungAh; Seo, Hyeon-Beom; Moon, Young-Mee; Kim, Ho-Youn; Park, Sung-Hwan; Cho, Mi-La

    2016-01-01

    PTEN is a tyrosine phosphatase with significant function in inhibiting STAT3 activation. Recently, inactivation of STAT3 has been demonstrated as a therapeutic candidate for autoimmune arthritis. The expression of PTEN controlled by p53 regulates autoimmune arthritis through modulating the balance between Th17 and Treg. We hypothesized that PTEN regulated by p53 might reduce CIA severity and inflammatory response via inhibiting STAT3 activation. Our results revealed that PTEN could ameliorate experimental autoimmune arthritis by reducing STAT3 activity and Th17 differentiation. Systemic infusion of PTEN overexpression downregulated CIA severity. In addition, PTEN overexpression decreased the activation of T cells and modulated reciprocal differentiation of Th17 and Treg cells. We observed that PTEN expression downregulated by p53 deficiency induced the activation of STAT3. Loss of p53 exacerbated autoimmune arthritis and dysregulated the population of Th17 and Treg. These data suggest that induction of STAT3-modulatory activity of PTEN may be a therapeutic target for rheumatoid arthritis therapy. PMID:27708408

  5. PTEN ameliorates autoimmune arthritis through down-regulating STAT3 activation with reciprocal balance of Th17 and Tregs.

    PubMed

    Lee, Seung Hoon; Park, Jin-Sil; Byun, Jae-Kyung; Jhun, JooYeon; Jung, KyungAh; Seo, Hyeon-Beom; Moon, Young-Mee; Kim, Ho-Youn; Park, Sung-Hwan; Cho, Mi-La

    2016-10-06

    PTEN is a tyrosine phosphatase with significant function in inhibiting STAT3 activation. Recently, inactivation of STAT3 has been demonstrated as a therapeutic candidate for autoimmune arthritis. The expression of PTEN controlled by p53 regulates autoimmune arthritis through modulating the balance between Th17 and Treg. We hypothesized that PTEN regulated by p53 might reduce CIA severity and inflammatory response via inhibiting STAT3 activation. Our results revealed that PTEN could ameliorate experimental autoimmune arthritis by reducing STAT3 activity and Th17 differentiation. Systemic infusion of PTEN overexpression downregulated CIA severity. In addition, PTEN overexpression decreased the activation of T cells and modulated reciprocal differentiation of Th17 and Treg cells. We observed that PTEN expression downregulated by p53 deficiency induced the activation of STAT3. Loss of p53 exacerbated autoimmune arthritis and dysregulated the population of Th17 and Treg. These data suggest that induction of STAT3-modulatory activity of PTEN may be a therapeutic target for rheumatoid arthritis therapy.

  6. Glutamine activates STAT3 to control cancer cell proliferation independently of glutamine metabolism.

    PubMed

    Cacace, Andrea; Sboarina, Martina; Vazeille, Thibaut; Sonveaux, Pierre

    2017-04-01

    Cancer cells can use a variety of metabolic substrates to fulfill the bioenergetic and biosynthetic needs of their oncogenic program. Besides bioenergetics, cancer cell metabolism also directly influences genetic, epigenetic and signaling events associated with tumor progression. Many cancer cells are addicted to glutamine, and this addiction is observed in oxidative as well as in glycolytic cells. Although both oxidative and bioreductive glutamine metabolism can contribute to cancer progression and glutamine can further serve to generate peptides (including glutathione) and proteins, we report that glutamine promotes the proliferation of cancer cells independently of its use as a metabolic fuel or as a precursor of glutathione. Extracellular glutamine activates transcription factor signal transducer and activator of transcription 3 (STAT3), which is necessary and sufficient to mediate the proliferative effects of glutamine on glycolytic and oxidative cancer cells. Glutamine also activates transcription factors hypoxia-inducible factor-1, mammalian target of rapamycin and c-Myc, but these factors do not mediate the effects of glutamine on cancer cell proliferation. Our findings shed a new light on the anticancer effects of l-asparaginase that possesses glutaminase activity and converts glutamine into glutamate extracellularly. Conversely, cancer resistance to treatments that block glutamine metabolism could arise from glutamine-independent STAT3 reactivation.

  7. STAT3-Activating Cytokines: A Therapeutic Opportunity for Inflammatory Bowel Disease?

    PubMed Central

    Nguyen, Paul M.; Putoczki, Tracy L.

    2015-01-01

    The gastrointestinal tract is lined by a single layer of epithelial cells that secrete mucus toward the lumen, which collectively separates the immune sentinels in the underlying lamina propria from the intestinal microflora to prevent aberrant immune responses. Inflammatory bowel disease (IBD) describes a group of autoimmune diseases that arise from defects in epithelial barrier function and, as a consequence, aberrant production of inflammatory cytokines. Among these, interleukin (IL)-6, IL-11, and IL-22 are elevated in human IBD patients and corresponding mouse models and, through activation of the JAK/STAT3 pathway, can both propagate and ameliorate disease. In particular, cytokine-mediated activation of STAT3 in the epithelial lining cells affords cellular protection, survival, and proliferation, thereby affording therapeutic opportunities for the prevention and treatment of colitis. In this review, we focus on recent insights gained from therapeutic modulation of the activities of IL-6, IL-11, and IL-22 in models of IBD and advocate a cautionary approach with these cytokines to minimize their tumor-promoting activities on neoplastic epithelium. PMID:25760898

  8. Insulin modulates STAT3 protein activation and gene transcription in hepatic cells.

    PubMed

    Campos, S P; Wang, Y; Baumann, H

    1996-10-04

    Treatment of rat hepatoma cells with insulin attenuated the interleukin 6 (IL-6) stimulation of acute phase plasma protein genes. To identify the potential mechanism of this action, the influence of insulin on IL-6 signal transduction was determined. An insulin dose-dependent reduction in signal transducer and activator of transcription 3 (STAT3) gene transcription, mRNA accumulation, protein concentration, and IL-6-inducible DNA binding activity was detected. A reduction in the IL-6-activated sis-inducible element binding of STAT3 was observed within 4 h of insulin treatment, whereas a maximal 3-4-fold lower STAT protein concentration was measured after 8-24 h of insulin treatment. Insulin mediated a similar magnitude reduction in the amount of mRNA encoding the IL-6 receptor alpha subunit and IL-6 binding activity. These effects of insulin appear to contribute to the strongly suppressed transcriptional induction of the IL-6-responsive acute phase plasma protein genes.

  9. Predominant Activation of JAK/STAT3 Pathway by Interleukin-6 Is Implicated in Hepatocarcinogenesis.

    PubMed

    Jung, In Hye; Choi, Jeffrey Hyun-Kyu; Chung, Yong-Yoon; Lim, Ga-Lam; Park, Young-Nyun; Park, Seung Woo

    2015-07-01

    Chronic inflammation is an important process leading to tumorigenesis. Therefore, targeting and controlling inflammation can be a promising cancer therapy. Inflammation is often caused by a variety of inflammatory cytokine such as the interleukin (IL)-6, a pleiotrophic cytokine known to be involved in the tumorigenesis. In this study, an in vivo hepatic tumorigenesis model of zebrafish was generated to demonstrate a direct consequence of the human IL6 expression causing hepatocarcinogenesis. To do this, an elevated expression of the hIL6 gene was established to specifically target the zebrafish hepatocytes by transgenesis. Interestingly, the elevated hIL6 expression caused the chronic inflammation which results in a massive infiltration of inflammatory cells. This eventually resulted in the generation of various dysplastic lesions such as clear cell, small cell, and large cell changes, and also eosinophilic and basophilic foci of hepatocellular alteration. Hepatocellular carcinoma was then developed in the transgenic zebrafish. Molecular characterization revealed upregulation of the downstream components involved in the IL6-mediated signaling pathways, especially PI3K/Akt and JAK/STAT3 pathways. Further investigation indicated that PI3K was the most reactive to the infiltrated inflammatory cells and dysplasia with large cell change, whereas STAT3 was heavily activated in the region with dysplastic foci, suggesting that the JAK/STAT3 pathway was mainly implicated in the hepatic tumorigenesis in the current model. Our present study provides an in vivo evidence of the relationship between chronic inflammation and tumorigenesis and reinforces the pivotal role of IL6 in the inflammation-associated hepatocarcinogenesis.

  10. Pien Tze Huang suppresses IL-6-inducible STAT3 activation in human colon carcinoma cells through induction of SOCS3.

    PubMed

    Shen, Aling; Chen, Youqin; Hong, Fei; Lin, Jiumao; Wei, Lihui; Hong, Zhenfeng; Sferra, Thomas J; Peng, Jun

    2012-12-01

    IL-6/STAT3 is one of the most critical cellular signal transduction pathways known to malfunction in colorectal cancer (CRC). As a target gene of signal transducer and activator of transcription 3 (STAT3) signaling, suppressor of cytokine signaling 3 (SOCS3) can be quickly induced by interleukin-6 (IL-6) stimulation but it then strongly inhibits IL-6-mediated STAT3 activation, functioning as a negative feedback regulator of the IL-6/STAT3 pathway. Aberrant activation of STAT3 and/or reduced expression of SOCS are strongly correlated with carcinogenesis, which therefore becomes a promising target for the development of novel anticancer chemotherapies. Pien Tze Huang (PZH) is a well-known traditional Chinese formula that was first prescribed by a royal physician 450 years ago in the Ming Dynasty. It has been used in China and Southeast Asia for centuries as a folk remedy for various types of cancer including CRC. However, the precise mechanism of its antitumor activity remains largely unclear. In the present study, we found that PZH could significantly and dose-dependently inhibit IL-6-mediated increase of STAT3 phosphorylation levels and transcriptional activity in the human colon carcinoma HT-29 cells, resulting in the suppression of cell proliferation and the induction of apoptosis. In addition, PZH treatment profoundly inhibited IL-6-induced upregulation of cyclin D1 and Bcl-2, two key target genes of the STAT3 pathway. Moreover, PZH treatment increased the expression of SOCS3. These results suggest that PZH could effectively inhibit proliferation and promote apoptosis of human colon carcinoma cells via modulation of the IL-6/STAT3 signaling pathway and its target genes.

  11. Xanthohumol inhibits STAT3 activation pathway leading to growth suppression and apoptosis induction in human cholangiocarcinoma cells.

    PubMed

    Dokduang, Hasaya; Yongvanit, Puangrat; Namwat, Nisana; Pairojkul, Chawalit; Sangkhamanon, Sakkarn; Yageta, Mika Sakurai; Murakami, Yoshinori; Loilome, Watcharin

    2016-04-01

    STAT3 plays a significant role in the development of cholangiocarcinoma (CCA) associated with the liver fluke (Opisthorchis viverrini; Ov). Xanthohumol (XN), a prenylated flavonoid extracted from hops, has known anticancer activity and could potentially target STAT3. The present study determined the effect of XN on STAT3, as well as ascertained its usefulness against CCA. The CCA cell proliferation at 20 µM and 50 µM of XN was shown to inhibited, while 20 µM partially inhibited IL-6-induced STAT3 activation. At 50 µM, the inhibition was complete. The reduction in STAT3 activity at 20 and 50 µM was associated with a significant reduction of CCA cell growth and apoptosis. We also found that the administration of 50 µM XN orally in drinking water to nude mice inoculated with CCA led to a reduction in tumor growth in comparison with controls. In addition, apoptosis of cancer cells increased although there was no visible toxicity. The present study shows that XN can inhibit STAT3 activation both in vivo and in vitro due to suppression of the Akt-NFκB signaling pathway. XN should be considered as a possible therapeutic agent against CCA.

  12. Icaritin, a novel FASN inhibitor, exerts anti-melanoma activities through IGF-1R/STAT3 signaling

    PubMed Central

    Wu, Jinfeng; Du, Juan; Fu, Xiuqiong; Liu, Bin; Cao, Huihui; Li, Ting; Su, Tao; Xu, Jinhua; Tse, Anfernee Kai-Wing; Yu, Zhi-Ling

    2016-01-01

    Icaritin (IT) is a flavonoid isolated from Herba Epimedii. In this study, we evaluated the anti-melanoma activities of IT, and determined its cytotoxic mechanism. We found that IT exerted cytotoxicity to melanoma cells. Furthermore, IT induced melanoma cell apoptosis, which was accompanied with PARP cleavage. Mechanistically, IT suppressed p-STAT3 (tyr705) level in parallel with increases of p-STAT3 (ser727), p-ERK and p-AKT. IT significantly inhibited STAT3 nuclear translocation and reduced the levels of STAT3 -targeted genes. IT also inhibited IGF-1-induced STAT3 activation through down-regulation of total IGF-1R level. No dramatic changes in IGF-1R mRNA levels were observed in IT-treated cells, suggesting that IT acted primarily at a post-transcriptional level. Using molecular docking analysis, IT was identified as a novel fatty acid synthase (FASN) inhibitor. We found that IT reduced the level of total IGF-1R via FASN inhibition. In summary, we reported that IT exerted anti-melanoma activities, and these effects were partially due to inhibition of FASN/IGF-1R/STAT3 signaling. PMID:27323414

  13. Signal transducer and activator of transcription (STAT)-3 regulates microRNA gene expression in chronic lymphocytic leukemia cells

    PubMed Central

    2013-01-01

    Backgrounds Approximately 1,000 microRNAs (miRs) are present in the human genome; however, little is known about the regulation of miR transcription. Because miR levels are deregulated in chronic lymphocytic leukemia (CLL) and signal transducer and activator of transcription (STAT)-3 is constitutively activated in CLL, we sought to determine whether STAT3 affects the transcription of miR genes in CLL cells. Methods We used publically available data from the ENCODE project to identify putative STAT3 binding sites in the promoters of miR genes. Then we transfected CLL cells with STAT3-shRNA or with an empty vector, and to determine which miRs are differentially expressed, we used a miR microarray approach followed by validation of the microarray results for 6 miRs using quantitative real-time polymerase chain reaction (qRT-PCR). Results We identified putative STAT3 binding sites in 160 promoter regions of 200 miRs, including miR-21, miR-29, and miR-155, whose levels have been reported to be upregulated in CLL. Levels of 72 miRs were downregulated (n = 63) or upregulated (n = 9). qRT-PCR confirmed the array data in 5 of 6 miRs. Conclusions The presence of activated STAT3 has a profound effect on miR expression in CLL cells. PMID:23725032

  14. Salinomycin Abolished STAT3 and STAT1 Interactions and Reduced Telomerase Activity in Colorectal Cancer Cells.

    PubMed

    Chung, Seyung S; Adekoya, Debbie; Enenmoh, Ikechukwu; Clarke, Orette; Wang, Piwen; Sarkyssian, Marianna; Wu, Yong; Vadgama, Jaydutt V

    2017-02-01

    Colorectal cancer is the third leading cause of cancer-related mortality in most developed countries. This mortality is mainly due to the metastatic progression to the liver with frequent recurrence. Colorectal cancer remains a therapeutic challenge and this has intensified the search for new drug targets. In an effort to establish a novel targeted-therapy, we studied the molecular mechanisms of cancer stem cell inhibitor salinomycin. Co-immunoprecipitation was performed to examine STAT3-STAT1 protein interactions. Telomerase activity was measured by polymerase chain reaction (PCR) and ELISA assays. Apoptosis and cell stress arrays were analyzed to identify key proteins responding to salinomycin treatments. IL-6 and TNF-α induced STAT3 and STAT1 interactions, however the interactions were abolished by salinomycin challenge. Salinomycin reduced cancer stem cell phenotype and decreased telomerase activity of colorectal cancer cells. Our work uncovers a new mechanism through which salinomycin inhibits cancer stemness suggesting a novel targeted-therapy for metastatic colorectal cancer. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  15. Hes1 Desynchronizes Differentiation of Pluripotent Cells by Modulating STAT3 Activity

    PubMed Central

    Zhou, Xinzhi; Smith, Andrew JH; Waterhouse, Anna; Blin, Guillaume; Malaguti, Mattias; Lin, Chia-Yi; Osorno, Rodrigo; Chambers, Ian; Lowell, Sally

    2013-01-01

    Robust development of the early embryo may benefit from mechanisms that ensure that not all pluripotent cells differentiate at exactly the same time: such mechanisms would build flexibility into the process of lineage allocation. This idea is supported by the observation that pluripotent stem cells differentiate at different rates in vitro. We use a clonal commitment assay to confirm that pluripotent cells commit to differentiate asynchronously even under uniform differentiation conditions. Stochastic variability in expression of the Notch target gene Hes1 has previously been reported to influence neural versus mesodermal differentiation through modulation of Notch activity. Here we report that Hes1 also has an earlier role to delay exit from the pluripotent state into all lineages. The early function of Hes1 to delay differentiation can be explained by an ability of Hes1 to amplify STAT3 responsiveness in a cell-autonomous manner. Variability in Hes1 expression therefore helps to explain why STAT3 responsiveness varies between individual ES cells, and this in turn helps to explain why pluripotent cells commit to differentiate asynchronously. Stem Cells 2013;31:1511–1522 PMID:23649667

  16. Hes1 desynchronizes differentiation of pluripotent cells by modulating STAT3 activity.

    PubMed

    Zhou, Xinzhi; Smith, Andrew J H; Waterhouse, Anna; Blin, Guillaume; Malaguti, Mattias; Lin, Chia-Yi; Osorno, Rodrigo; Chambers, Ian; Lowell, Sally

    2013-08-01

    Robust development of the early embryo may benefit from mechanisms that ensure that not all pluripotent cells differentiate at exactly the same time: such mechanisms would build flexibility into the process of lineage allocation. This idea is supported by the observation that pluripotent stem cells differentiate at different rates in vitro. We use a clonal commitment assay to confirm that pluripotent cells commit to differentiate asynchronously even under uniform differentiation conditions. Stochastic variability in expression of the Notch target gene Hes1 has previously been reported to influence neural versus mesodermal differentiation through modulation of Notch activity. Here we report that Hes1 also has an earlier role to delay exit from the pluripotent state into all lineages. The early function of Hes1 to delay differentiation can be explained by an ability of Hes1 to amplify STAT3 responsiveness in a cell-autonomous manner. Variability in Hes1 expression therefore helps to explain why STAT3 responsiveness varies between individual ES cells, and this in turn helps to explain why pluripotent cells commit to differentiate asynchronously.

  17. Obox4-silencing-activated STAT3 and MPF/MAPK signaling accelerate nuclear membrane breakdown in mouse oocytes.

    PubMed

    Lee, Hyun-Seo; Kim, Kyeoung-Hwa; Kim, Eun-Young; Lee, Su-Yeon; Ko, Jung-Jae; Lee, Kyung-Ah

    2016-04-01

    Mouse oocytes begin to mature in vitro once liberated from ovarian follicles. Previously, we showed that oocyte-specific homeobox 4 (Obox4) is critical for maintaining the intact nuclear membrane of the germinal vesicle (GV) in oocytes and for completing meiosis at the metaphase I-II (MI-MII) transition. This study further examines the molecular mechanisms of OBOX4 in regulating GV nuclear membrane breakdown. Maturation-promoting factor (MPF) and MAPK are normally inactive in GV stage oocytes but were activated prematurely in arrested GV stage oocytes by 3-isobutyl-1-metyl-xanthine (IBMX) in vitro after Obox4 RNA interference (RNAi). Furthermore, signal transducer and activator of transcription 3 (STAT3) was significantly activated by Obox4 RNAi. We confirmed that this Obox4 RNAi-induced premature STAT3 and MPF/MAPK activation at the GV stage provoked subsequent GV breakdown (GVBD) despite the opposing force of high cAMP in the IBMX-supplemented medium to maintain intact GV. When cumulus-oocyte complexes were exposed to interferon α (IFNA), a STAT3 activator, oocytes matured and cumulus cells expanded to resume nuclear maturation in IBMX-supplemented medium, suggesting that STAT3 activation is sufficient for stimulating the continuation of meiosis. Using Stattic, a specific STAT3 inhibitor, we confirmed that GVBD involves STAT3 activation in Obox4-silenced oocytes. Based on these findings, we concluded that i) Obox4 is an important upstream regulator of MPF/MAPK and STAT3 signaling, and ii) Obox4 is a key regulator of the GV arrest mechanism in oocytes.

  18. Dimethylfumarate suppresses adipogenic differentiation in 3T3-L1 preadipocytes through inhibition of STAT3 activity.

    PubMed

    Kang, Hyeon-Ji; Seo, Hyun-Ae; Go, Younghoon; Oh, Chang Joo; Jeoung, Nam Ho; Park, Keun-Gyu; Lee, In-Kyu

    2013-01-01

    The excessive accumulation of adipocytes contributes to the development of obesity and obesity-related diseases. The interactions of several transcription factors, such as C/EBPβ, PPARγ, C/EBPα, Nrf2, and STAT3, are required for adipogenic differentiation. Dimethylfumarate (DMF), an immune modulator and antioxidant, may function as an inhibitor of STAT3 and an activator of Nrf2. This study examined whether DMF inhibits adipogenic differentiation of 3T3-L1 preadipocytes by inhibiting STAT3 or activating Nrf2. DMF suppressed 3T3-L1 preadipocyte differentiation to mature adipocytes in a dose-dependent manner as determined by Oil Red O staining. The mRNA and protein levels of adipogenic genes, including C/EBPβ, C/EBPα, PPARγ, SREBP-1c, FAS, and aP2, were significantly lower in DMF-treated 3T3-L1 preadipocytes. Suppression of adipogenic differentiation by DMF treatment resulted primarily from inhibition of the early stages of differentiation. DMF inhibits clonal expansion during adipogenic differentiation through induction of a G1 cell cycle arrest. Additionally, DMF regulates cell cycle-related proteins, such as p21, pRb, and cyclin D. DMF treatment markedly inhibited differentiation medium-induced STAT3 phosphorylation and inhibited STAT3 transcriptional activation of a reporter construct composed of four synthetic STAT3-response elements. Moreover, inhibition of endogenous Nrf2 activity using a dominant negative Nrf2 did not abolish the DMF-induced inhibition of adipogenic differentiation of 3T3-L1 preadipocytes. In summary, DMF is a negative regulator of adipogenic differentiation based on its regulation of adipogenic transcription factors and cell cycle proteins. This negative regulation by DMF is mediated by STAT3 inhibition, but is unlikely to involve Nrf2 activation.

  19. STAT3 signaling is activated preferentially in tumor-initiating cells in claudin-low models of human breast cancer.

    PubMed

    Wei, Wei; Tweardy, David J; Zhang, Mei; Zhang, Xiaomei; Landua, John; Petrovic, Ivana; Bu, Wen; Roarty, Kevin; Hilsenbeck, Susan G; Rosen, Jeffrey M; Lewis, Michael T

    2014-10-01

    In breast cancer, a subset of tumor-initiating cells (TIC) or "cancer stem cells" are thought to be responsible for tumor maintenance, treatment resistance, and disease recurrence. While current breast cancer stem cell markers (e.g., CD44(high) /CD24(low/neg) , ALDH positive) have allowed enrichment for such cells, they are not universally expressed and may actually identify distinct TIC subpopulations in the same tumor. Thus, additional markers of functional stem cells are needed. The STAT3 pathway is a critical regulator of the function of normal stem cells, and evidence is accumulating for its important role in breast cancer stem cells. However, due to the lack of a method for separating live cells based on their level of STAT3 activity, it remains unknown whether STAT3 functions in the cancer stem cells themselves, or in surrounding niche cells, or in both. To approach this question, we constructed a series of lentiviral fluorescent (enhanced green fluorescent protein, EGFP) reporters that enabled flow cytometric enrichment of cells differing in STAT3-mediated transcriptional activity, as well as in vivo/in situ localization of STAT3 responsive cells. Using in vivo claudin-low cell line xenograft models of human breast cancer, we found that STAT3 signaling reporter activity (EGFP(+) ) is associated with a subpopulation of cancer cells enriched for mammosphere-forming efficiency, as well as TIC function in limiting dilution transplantation assays compared to negative or unsorted populations. Our results support STAT3 signaling activity as another functional marker for human breast cancer stem cells thus making it an attractive therapeutic target for stem-cell-directed therapy in some breast cancer subtypes. © AlphaMed Press.

  20. Activation of Antitumorigenic Stat3beta in Breast Cancer by Splicing Redirection

    DTIC Science & Technology

    2013-07-01

    used a formulation of the compounds (vivo-morpholino) where a cationic dendrimer is coupled to the oligonucleotide instead of the peptide. This variant...rather than a direct block of canonical STAT3 targets. Conjugation of the STAT3β- inducing PMOs to a cell-penetrating dendrimer (vivo- morpholinos

  1. Activation of Antitumorigenic Stat3beta in Breast Cancer by Splicing Redirection

    DTIC Science & Technology

    2013-07-01

    therapy. Nat Rev Cancer 2: 740-749. 3. Levy DE, Lee CK (2002) What does Stat3 do? J Clin Invest 109: 1143-1148. 4. Akira S ( 2000 ) Roles of STAT3 defined...breast cancer cells. Cancer Res 57: 978-987. 17. Inghirami G, Chiarle R, Simmons WJ, Piva R, Schlessinger K, et al. (2005) New and old functions of

  2. STAT3 activation by IL-6 from mesenchymal stem cells promotes the proliferation and metastasis of osteosarcoma.

    PubMed

    Tu, Bing; Du, Lin; Fan, Qi-Ming; Tang, Ze; Tang, Ting-Ting

    2012-12-01

    We previously demonstrated that human mesenchymal stem cells (MSCs) promote the growth of osteosarcoma in the bone microenvironment. The aim of the present study was to further determine the effect of IL-6/STAT3 signaling on the progression of osteosarcoma. First, conditioned medium from MSCs was used to stimulate the growth of osteosarcoma cells (Saos-2) in vitro. We found that STAT3 was activated and that the activation could be blocked by an IL-6-neutralizing antibody. The inhibition of STAT3 in Saos-2 cells by siRNA or AG490 decreased cell proliferation, migration and invasion, down-regulated the mRNA expression of Cyclin D, Bcl-xL and Survivin and enhanced the apoptotic response. Furthermore, a nude mouse osteosarcoma model was established by injecting luciferase-labeled Saos-2 cells into the tibia, and the effect of STAT3 on tumor growth was determined by treating the mice with AG490. In vivo bioluminescence images showed that tumor growth was dramatically reduced in the AG490 group. In addition, STAT3 inhibition decreased the lung metastasis rate and prolonged the survival of these mice. After treatment with AG490, the protein levels of IL-6, p-STAT3 and PCNA were decreased, and the level of apoptosis in the tumor was increased. Altogether, these data indicate that MSCs in the bone microenvironment might promote the progression of osteosarcoma and protect tumor cells from drug-induced apoptosis through IL-6/STAT3 signaling. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

  3. IL-15 Activates the Jak3/STAT3 Signaling Pathway to Mediate Glucose Uptake in Skeletal Muscle Cells

    PubMed Central

    Krolopp, James E.; Thornton, Shantaé M.; Abbott, Marcia J.

    2016-01-01

    Myokines are specialized cytokines that are secreted from skeletal muscle (SKM) in response to metabolic stimuli, such as exercise. Interleukin-15 (IL-15) is a myokine with potential to reduce obesity and increase lean mass through induction of metabolic processes. It has been previously shown that IL-15 acts to increase glucose uptake in SKM cells. However, the downstream signals orchestrating the link between IL-15 signaling and glucose uptake have not been fully explored. Here we employed the mouse SKM C2C12 cell line to examine potential downstream targets of IL-15-induced alterations in glucose uptake. Following differentiation, C2C12 cells were treated overnight with 100 ng/ml of IL-15. Activation of factors associated with glucose metabolism (Akt and AMPK) and known downstream targets of IL-15 (Jak1, Jak3, STAT3, and STAT5) were assessed with IL-15 stimulation. IL-15 stimulated glucose uptake and GLUT4 translocation to the plasma membrane. IL-15 treatment had no effect on phospho-Akt, phospho-Akt substrates, phospho-AMPK, phospho-Jak1, or phospho-STAT5. However, with IL-15, phospho-Jak3 and phospho-STAT3 levels were increased along with increased interaction of Jak3 and STAT3. Additionally, IL-15 induced a translocation of phospho-STAT3 from the cytoplasm to the nucleus. We have evidence that a mediator of glucose uptake, HIF1α, expression was dependent on IL-15 induced STAT3 activation. Finally, upon inhibition of STAT3 the positive effects of IL-15 on glucose uptake and GLUT4 translocation were abolished. Taken together, we provide evidence for a novel signaling pathway for IL-15 acting through Jak3/STAT3 to regulate glucose metabolism. PMID:28066259

  4. IL-15 Activates the Jak3/STAT3 Signaling Pathway to Mediate Glucose Uptake in Skeletal Muscle Cells.

    PubMed

    Krolopp, James E; Thornton, Shantaé M; Abbott, Marcia J

    2016-01-01

    Myokines are specialized cytokines that are secreted from skeletal muscle (SKM) in response to metabolic stimuli, such as exercise. Interleukin-15 (IL-15) is a myokine with potential to reduce obesity and increase lean mass through induction of metabolic processes. It has been previously shown that IL-15 acts to increase glucose uptake in SKM cells. However, the downstream signals orchestrating the link between IL-15 signaling and glucose uptake have not been fully explored. Here we employed the mouse SKM C2C12 cell line to examine potential downstream targets of IL-15-induced alterations in glucose uptake. Following differentiation, C2C12 cells were treated overnight with 100 ng/ml of IL-15. Activation of factors associated with glucose metabolism (Akt and AMPK) and known downstream targets of IL-15 (Jak1, Jak3, STAT3, and STAT5) were assessed with IL-15 stimulation. IL-15 stimulated glucose uptake and GLUT4 translocation to the plasma membrane. IL-15 treatment had no effect on phospho-Akt, phospho-Akt substrates, phospho-AMPK, phospho-Jak1, or phospho-STAT5. However, with IL-15, phospho-Jak3 and phospho-STAT3 levels were increased along with increased interaction of Jak3 and STAT3. Additionally, IL-15 induced a translocation of phospho-STAT3 from the cytoplasm to the nucleus. We have evidence that a mediator of glucose uptake, HIF1α, expression was dependent on IL-15 induced STAT3 activation. Finally, upon inhibition of STAT3 the positive effects of IL-15 on glucose uptake and GLUT4 translocation were abolished. Taken together, we provide evidence for a novel signaling pathway for IL-15 acting through Jak3/STAT3 to regulate glucose metabolism.

  5. TLR9-Targeted STAT3 Silencing Abrogates Immunosuppressive Activity of Myeloid-Derived Suppressor Cells from Prostate Cancer Patients

    PubMed Central

    Hossain, Dewan M. S.; Pal, Sumanta K.; Moreira, Dayson; Duttagupta, Priyanka; Zhang, Qifang; Won, Haejung; Jones, Jeremy; D'Apuzzo, Massimo; Forman, Stephen; Kortylewski, Marcin

    2015-01-01

    Purpose Recent advances in immunotherapy of advanced human cancers underscored the need to address and eliminate tumor immune evasion. The myeloid-derived suppressor cells (MDSCs) are important inhibitors of T cell responses in solid tumors, such as prostate cancers. However, targeting MDSCs proved challenging due to their phenotypic heterogeneity. Experimental Design Myeloid cell populations were evaluated using flow cytometry on blood samples, functional assays and immunohistochemical/immunofluorescent stainings on tumor/tumor-draining lymph node specimens from healthy subjects, localized and metastatic castration-resistant prostate cancer patients. Results Here, we identify a population of Lin−CD15HICD33LO granulocytic MDSCs that accumulate in patients’ circulation during prostate cancer progression from localized to metastatic disease. The prostate cancer-associated MDSCs potently inhibit autologous CD8+ T cells proliferation and production of IFNγ and Granzyme-B. The circulating MDSCs have high levels of activated STAT3, which is a central immune checkpoint regulator. The granulocytic pSTAT3+ cells are also detectable in patients’ prostate tissues. We previously generated an original strategy to silence genes specifically in Toll-like Receptor-9 (TLR9) positive myeloid cells using CpG-siRNA conjugates. We demonstrate that human granulocytic MDSCs express TLR9 and rapidly internalize naked CpG-STAT3siRNA thereby silencing STAT3 expression. STAT3 blocking abrogates immunosuppressive effects of patients-derived MDSCs on effector CD8+ T cells. These effects depended on reduced expression and enzymatic activity of Arginase-1, a downstream STAT3 target gene and a potent T cell inhibitor. Conclusions Overall, we demonstrate the accumulation of granulocytic MDSCs with prostate cancer progression and the feasibility of using TLR9-targeted STAT3siRNA delivery strategy to alleviate MDSC-mediated immunosuppression. PMID:25967142

  6. Activation of the STAT3 Signaling Pathway Is Associated With Poor Survival in Diffuse Large B-Cell Lymphoma Treated With R-CHOP

    PubMed Central

    Huang, Xin; Meng, Bin; Iqbal, Javeed; Ding, B. Belinda; Perry, Anamarija M.; Cao, Wenfeng; Smith, Lynette M.; Bi, Chengfeng; Jiang, Chunsun; Greiner, Timothy C.; Weisenburger, Dennis D.; Rimsza, Lisa; Rosenwald, Andreas; Ott, German; Delabie, Jan; Campo, Elias; Braziel, Rita M.; Gascoyne, Randy D.; Cook, James R.; Tubbs, Raymond R.; Jaffe, Elaine S.; Armitage, James O.; Vose, Julie M.; Staudt, Louis M.; McKeithan, Timothy W.; Chan, Wing C.; Ye, B. Hilda; Fu, Kai

    2013-01-01

    Purpose We previously reported that constitutive STAT3 activation is a prominent feature of the activated B-cell subtype of diffuse large B-cell lymphomas (ABC-DLBCL). In this study, we investigated whether STAT3 activation can risk stratify patients with DLBCL. Patients and Methods By an immunohistochemical method, we investigated phosphotyrosine STAT3 (PY-STAT3) expression from 185 patients with DLBCL treated with R-CHOP (rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone). Cell line-based siRNA experiments were also performed to generate an 11-gene, PY-STAT3 activation signature, which was used to study a previously published cohort of 222 patients with DLBCL. The STAT3 activation status determined by these two methods and by STAT3 mRNA levels were then correlated with survival. Results PY-STAT3 was detected in 37% of DLBCL and enriched in ABC-DLBCL cases (P = .03). PY-STAT3 positivity significantly correlated with poor overall survival (OS; P = .01) and event-free survival (EFS; P = .006). Similar observations were made for high levels of STAT3 mRNA. In multivariable analysis, PY-STAT3 status (P = .02), International Prognostic Index (P = .02), and BCL2 expression (P = .046) were independent prognosticators of OS in this cohort. Among the cell-of-origin subgroups, PY-STAT3 was associated with poor EFS among non–germinal center B-cell DLBCL cases only (P = .027). Similarly, the 11-gene STAT3 activation signature correlated with poor survival in the entire DLBCL cohort (OS, P < .001; EFS, P < .001) as well as the ABC-DLBCL subgroup (OS, P = .029; EFS, P = .025). Conclusion STAT3 activation correlated with poor survival in patients with DLBCL treated with R-CHOP, especially those with tumors of the ABC-DLBCL subtype. PMID:24220563

  7. Regulation of STAT3 and NF-κB activations by S-nitrosylation in multiple myeloma.

    PubMed

    Kim, Jinsu; Choi, Seungho; Saxena, Nishant; Singh, Avtar K; Singh, Inderjit; Won, Je-Seong

    2017-05-01

    Numerous reports suggest that aberrant activations of STAT3 and NF-κB promote survival and proliferation of multiple myeloma (MM) cells. In the present report, we demonstrate that a synthetic S-nitrosothiol compound, S-nitroso-N-acetylcysteine (SNAC), inhibits proliferation and survival of multiple MM cells via S-nitrosylation-dependent inhibition of STAT3 and NF-κB. In human MM cells (e.g. U266, H929, and IM-9 cells), SNAC treatment increased S-nitrosylation of STAT3 and NF-κB and inhibited their activities. Consequently, SNAC treatment resulted in MM cell cycle arrest at G1/S check point and inhibited their proliferation. SNAC also decreased the expression of cell survival factors and increased the activities of caspases, thus increased sensitivity of MM cells to melphalan, a chemotherapeutic agent for MM. In U266 xenografted mice, SNAC treatment decreased the activity of STAT3 and reduced the growth of human CD138 positive cells (U266 cells) in the bone marrow and also reduced their production of human IgE into the serum. Taken together, these data document the S-nitrosylation mediated inhibition of MM cell proliferation and cell survival via inhibition of STAT3 and NF-κB pathways and its efficacy in animal model of MM.

  8. Differential Pro-Inflammatory Responses of Astrocytes and Microglia Involve STAT3 Activation in Response to 1800 MHz Radiofrequency Fields

    PubMed Central

    Lu, Yonghui; He, Mindi; Zhang, Yang; Xu, Shangcheng; Zhang, Lei; He, Yue; Chen, Chunhai; Liu, Chuan; Pi, Huifeng; Yu, Zhengping; Zhou, Zhou

    2014-01-01

    Microglia and astrocytes play important role in maintaining the homeostasis of central nervous system (CNS). Several CNS impacts have been postulated to be associated with radiofrequency (RF) electromagnetic fields exposure. Given the important role of inflammation in neural physiopathologic processes, we investigated the pro-inflammatory responses of microglia and astrocytes and the involved mechanism in response to RF fields. Microglial N9 and astroglial C8-D1A cells were exposed to 1800 MHz RF for different time with or without pretreatment with STAT3 inhibitor. Microglia and astrocytes were activated by RF exposure indicated by up-regulated CD11b and glial fibrillary acidic protein (GFAP). However, RF exposure induced differential pro-inflammatory responses in astrocytes and microglia, characterized by different expression and release profiles of IL-1β, TNF-α, IL-6, PGE2, nitric oxide (NO), inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX2). Moreover, the RF exposure activated STAT3 in microglia but not in astrocytes. Furthermore, the STAT3 inhibitor Stattic ameliorated the RF-induced release of pro-inflammatory cytokines in microglia but not in astrocytes. Our results demonstrated that RF exposure differentially induced pro-inflammatory responses in microglia and astrocytes, which involved differential activation of STAT3 in microglia and astrocytes. Our data provide novel insights into the potential mechanisms of the reported CNS impacts associated with mobile phone use and present STAT3 as a promising target to protect humans against increasing RF exposure. PMID:25275372

  9. Tocilizumab inhibits neuronal cell apoptosis and activates STAT3 in cerebral infarction rat model.

    PubMed

    Wang, Shaojun; Zhou, Jun; Kang, Weijie; Dong, Zhaoni; Wang, Hezuo

    2016-01-15

    Cerebral infarction is a severe hypoxic ischemic necrosis with accelerated neuronal cell apoptosis in the brain. As a monoclonal antibody against interleukin 6, tocilizumab (TCZ) is widely used in immune diseases, whose function in cerebral infarction has not been studied. This study aims to reveal the role of TCZ in regulating neuronal cell apoptosis in cerebral infarction. The cerebral infarction rat model was constructed by middle cerebral artery occlusion and treated with TCZ. Cell apoptosis in hippocampus and cortex of the brain was examined with TUNEL method. Rat neuronal cells cultured in oxygen-glucose deprivation (OGD) conditions and treated with TCZ were used to compare cell viability and apoptosis. Apoptosis-related factors including B-cell lymphoma extra large (Bcl-xL) and Caspase 3, as well as the phosphorylated signal transducer and activator of transcription 3 (p-STAT3) in brain cortex were analyzed from the protein level. Results indicated that TCZ treatment could significantly prevent the promoted cell apoptosis caused by cerebral infarction or OGD (P < 0.05 or P < 0.01). In brain cortex of the rat model, TCZ up-regulated Bcl-xL and down-regulated Caspase 3, consistent with the inhibited cell apoptosis. It also promoted tyrosine 705 phosphorylation of STAT3, which might be the potential regulatory mechanism of TCZ in neuronal cells. This study provided evidence for the protective role of TCZ against neuronal cell apoptosis in cerebral infarction. Based on these fundamental data, TCZ is a promising option for treating cerebral infarction, but further investigations on related mechanisms are still necessary.

  10. Tocilizumab inhibits neuronal cell apoptosis and activates STAT3 in cerebral infarction rat model

    PubMed Central

    Wang, Shaojun; Zhou, Jun; Kang, Weijie; Dong, Zhaoni; Wang, Hezuo

    2016-01-01

    Cerebral infarction is a severe hypoxic ischemic necrosis with accelerated neuronal cell apoptosis in the brain. As a monoclonal antibody against interleukin 6, tocilizumab (TCZ) is widely used in immune diseases, whose function in cerebral infarction has not been studied. This study aims to reveal the role of TCZ in regulating neuronal cell apoptosis in cerebral infarction. The cerebral infarction rat model was constructed by middle cerebral artery occlusion and treated with TCZ. Cell apoptosis in hippocampus and cortex of the brain was examined with TUNEL method. Rat neuronal cells cultured in oxygen-glucose deprivation (OGD) conditions and treated with TCZ were used to compare cell viability and apoptosis. Apoptosis-related factors including B-cell lymphoma extra large (Bcl-xL) and Caspase 3, as well as the phosphorylated signal transducer and activator of transcription 3 (p-STAT3) in brain cortex were analyzed from the protein level. Results indicated that TCZ treatment could significantly prevent the promoted cell apoptosis caused by cerebral infarction or OGD (P < 0.05 or P < 0.01). In brain cortex of the rat model, TCZ up-regulated Bcl-xL and down-regulated Caspase 3, consistent with the inhibited cell apoptosis. It also promoted tyrosine 705 phosphorylation of STAT3, which might be the potential regulatory mechanism of TCZ in neuronal cells. This study provided evidence for the protective role of TCZ against neuronal cell apoptosis in cerebral infarction. Based on these fundamental data, TCZ is a promising option for treating cerebral infarction, but further investigations on related mechanisms are still necessary. PMID:26773188

  11. NADPH oxidase and ERK1/2 are involved in cadmium induced-STAT3 activation in HepG2 cells.

    PubMed

    Souza, Verónica; Escobar, Ma Del Carmen; Bucio, Leticia; Hernández, Elizabeth; Gómez-Quiroz, Luis Enrique; Gutiérrez Ruiz, Ma Concepción

    2009-06-22

    The molecular mechanism of Cd-induced signal transduction is not well understood. The aims of this study were to determine the system that generates reactive oxygen species in response to Cd that contribute to intracellular signaling on the activation of the STAT3 pathway in HepG2 cells and to address the participation of STAT3 in the production of Hsp70. Cadmium induced a significant increase in STAT3 DNA-binding after 1h treatment. Serine phosphorylation of STAT3 was observed as a result of cadmium treatment while no tyrosine phosphorylation was detected. Cells were pretreated with inhibitors for several ROS generating systems, only diphenylen iodonium, an inhibitor of NADPH oxidase, decreased STAT3 activation. Cd induced 2.6-fold NADPH oxidase activity. Antioxidant treatment with pegylated-catalase reduced STAT3 activation. Cells were pretreated with different MAPK's inhibitors. ERK contributes in approximately 60%, and JNK in a small proportion, while p38 does not contribute in STAT3 activation. Cells were pretreated with a specific STAT3 peptide inhibitor that decreased the Cd-induced Hsp70 expression. Data suggest that STAT3 is phosphorylated at serine 727 by a Cd stress-activated signaling pathway inducing NADPH oxidase activity which produced ROS, leading ERK activation. MAPK promotes STAT3 phosphorylation that could induce a protective mechanism against Cd toxicity.

  12. STAT3 activation is associated with cerebrospinal fluid interleukin-10 (IL-10) in primary central nervous system diffuse large B cell lymphoma.

    PubMed

    Mizowaki, Takashi; Sasayama, Takashi; Tanaka, Kazuhiro; Mizukawa, Katsu; Takata, Kumi; Nakamizo, Satoshi; Tanaka, Hirotomo; Nagashima, Hiroaki; Nishihara, Masamitsu; Hirose, Takanori; Itoh, Tomoo; Kohmura, Eiji

    2015-09-01

    Signal transducers and activators of transcription 3 (STAT3) are activated by various cytokines and oncogenes; however, the activity and pathogenesis of STAT3 in diffuse large B cell lymphoma of the central nervous system have not been thoroughly elucidated. We investigated the phosphorylation levels of STAT3 in 40 specimens of primary central nervous system diffuse large B-cell lymphoma (PCNS DLBCL) and analyzed the association between phsopho-STAT3 (pSTAT3) expression and cerebrospinal fluid (CSF) concentration of interleukin-10 (IL-10) or IL-6. Immunohistochemistry and Western blot analysis revealed that most of the specimens in PCNS DLBCL expressed pSTST3 protein, and a strong phosphorylation levels of STAT3 was statistically associated with high CSF IL-10 levels, but not with CSF IL-6 levels. Next, we demonstrated that recombinant IL-10 and CSF containing IL-10 induced the phosphorylation of STAT3 in PCNS DLBCL cells. Furthermore, molecular subtype classified by Hans' algorithm was correlated with pSTAT3 expression levels and CSF IL-10 levels. These results suggest that the STAT3 activity is correlated with CSF IL-10 level, which is a useful marker for STAT3 activity in PCNS DLBCLs.

  13. Small-molecule targeting of signal transducer and activator of transcription (STAT) 3 to treat non-small cell lung cancer.

    PubMed

    Lewis, Katherine M; Bharadwaj, Uddalak; Eckols, T Kris; Kolosov, Mikhail; Kasembeli, Moses M; Fridley, Colleen; Siller, Ricardo; Tweardy, David J

    2015-11-01

    Lung cancer is the leading cause of cancer death in both men and women. Non-small cell lung cancer (NSCLC) has an overall 5-year survival rate of 15%. While aberrant STAT3 activation has previously been observed in NSCLC, the scope of its contribution is uncertain and agents that target STAT3 for treatment are not available clinically. We determined levels of activated STAT3 (STAT3 phosphorylated on Y705, pSTAT3) and the two major isoforms of STAT3 (α and β) in protein extracts of 8 NSCLC cell lines, as well as the effects of targeting STAT3 in vitro and in vivo in NSCLC cells using short hairpin (sh) RNA and two novel small-molecule STAT3 inhibitors, C188-9 and piperlongumine (PL). Levels of pSTAT3, STAT3α, and STATβ were increased in 7 of 8 NSCLC cell lines. Of note, levels of pSTAT3 were tightly correlated with levels of STAT3β, but not STAT3α. Targeting of STAT3 in A549 cells using shRNA decreased tSTAT3 by 75%; this was accompanied by a 47-78% reduction in anchorage-dependent and anchorage-independent growth and a 28-45% reduction in mRNA levels for anti-apoptotic STAT3 gene targets. C188-9 and PL (@30 μM) each reduced pSTAT3 levels in all NSCLC cell lines tested by ≥50%, reduced anti-apoptotic protein mRNA levels by 25-60%, and reduced both anchorage-dependent and anchorage-independent growth of NSCLC cell lines with IC50 values ranging from 3.06 to 52.44 μM and 0.86 to 11.66 μM, respectively. Treatment of nude mice bearing A549 tumor xenografts with C188-9 or PL blocked tumor growth and reduced levels of pSTAT3 and mRNA encoding anti-apoptotic proteins. STAT3 is essential for growth of NSCLC cell lines and tumors and its targeting using C188-9 or PL may be a useful strategy for treatment. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  14. EBV Latent Membrane Protein 1 Activates Akt, NFκB, and Stat3 in B Cell Lymphomas

    PubMed Central

    Shair, Kathy H. Y; Bendt, Katherine M; Edwards, Rachel H; Bedford, Elisabeth C; Nielsen, Judith N; Raab-Traub, Nancy

    2007-01-01

    Latent membrane protein 1 (LMP1) is the major oncoprotein of Epstein-Barr virus (EBV). In transgenic mice, LMP1 promotes increased lymphoma development by 12 mo of age. This study reveals that lymphoma develops in B-1a lymphocytes, a population that is associated with transformation in older mice. The lymphoma cells have deregulated cell cycle markers, and inhibitors of Akt, NFκB, and Stat3 block the enhanced viability of LMP1 transgenic lymphocytes and lymphoma cells in vitro. Lymphoma cells are independent of IL4/Stat6 signaling for survival and proliferation, but have constitutively activated Stat3 signaling. These same targets are also deregulated in wild-type B-1a lymphomas that arise spontaneously through age predisposition. These results suggest that Akt, NFκB, and Stat3 pathways may serve as effective targets in the treatment of EBV-associated B cell lymphomas. PMID:17997602

  15. SC-2001 Overcomes STAT3-mediated Sorafenib Resistance through RFX-1/SHP-1 Activation in Hepatocellular Carcinoma☆☆☆

    PubMed Central

    Su, Jung-Chen; Tseng, Ping-Hui; Wu, Szu-Hsien; Hsu, Cheng-Yi; Tai, Wei-Tien; Li, Yong-Shi; Chen, I-Ting; Liu, Chun-Yu; Chen, Kuen-Feng; Shiau, Chung-Wai

    2014-01-01

    Hepatocellular carcinoma is the fifth most common solid cancer worldwide. Sorafenib, a small multikinase inhibitor, is the only approved therapy for advanced HCC. The clinical benefit of sorafenib is offset by the acquisition of sorafenib resistance. Understanding of the molecular mechanism of STAT3 overexpression in sorafenib resistance is critical if the clinical benefits of this drug are to be improved. In this study, we explored our hypothesis that loss of RFX-1/SHP-1 and further increase of p-STAT3 as a result of sorafenib treatment induces sorafenib resistance as a cytoprotective response effect, thereby, limiting sorafenib sensitivity and efficiency. We found that knockdown of RFX-1 protected HCC cells against sorafenib-induced cell apoptosis and SHP-1 activity was required for the process. SC-2001, a molecule with similar structure to obatoclax, synergistically suppressed tumor growth when used in combination with sorafenib in vitro and overcame sorafenib resistance through up-regulating RFX-1 and SHP-1 resulting in tumor suppression and mediation of dephosphorylation of STAT3. In addition, sustained sorafenib treatment in HCC led to increased p-STAT3 which was a key mediator of sorafenib sensitivity. The combination of SC-2001 and sorafenib strongly inhibited tumor growth in both wild-type and sorafenib-resistant HCC cell bearing xenograft models. These results demonstrate that inactivation of RFX/SHP-1 induced by sustained sorafenib treatment confers sorafenib resistance to HCC through p-STAT3 up-regulation. These effects can be overcome by SC-2001 through RFX-1/SHP-1 dependent p-STAT3 suppression. In conclusion, the use of SC-2001 in combination with sorafenib may constitute a new strategy for HCC therapy. PMID:25047655

  16. Stimulation of the B-cell receptor activates the JAK2/STAT3 signaling pathway in chronic lymphocytic leukemia cells

    PubMed Central

    Rozovski, Uri; Wu, Ji Yuan; Harris, David M.; Liu, Zhiming; Li, Ping; Hazan-Halevy, Inbal; Ferrajoli, Alessandra; Burger, Jan A.; O’Brien, Susan; Jain, Nitin; Verstovsek, Srdan; Wierda, William G.; Keating, Michael J.

    2014-01-01

    In chronic lymphocytic leukemia (CLL), stimulation of the B-cell receptor (BCR) triggers survival signals. Because in various cells activation of the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway provides cells with survival advantage, we wondered whether BCR stimulation activates the JAK/STAT pathway in CLL cells. To stimulate the BCR we incubated CLL cells with anti-IgM antibodies. Anti-IgM antibodies induced transient tyrosine phosphorylation and nuclear localization of phosphorylated (p) STAT3. Immunoprecipitation studies revealed that anti-JAK2 antibodies coimmunoprecipitated pSTAT3 and pJAK2 in IgM-stimulated but not unstimulated CLL cells, suggesting that activation of the BCR induces activation of JAK2, which phosphorylates STAT3. Incubation of CLL cells with the JAK1/2 inhibitor ruxolitinib inhibited IgM-induced STAT3 phosphorylation and induced apoptosis of IgM-stimulated but not unstimulated CLL cells in a dose- and time-dependent manner. Whether ruxolitinib treatment would benefit patients with CLL remains to be determined. PMID:24778152

  17. Stimulation of the B-cell receptor activates the JAK2/STAT3 signaling pathway in chronic lymphocytic leukemia cells.

    PubMed

    Rozovski, Uri; Wu, Ji Yuan; Harris, David M; Liu, Zhiming; Li, Ping; Hazan-Halevy, Inbal; Ferrajoli, Alessandra; Burger, Jan A; O'Brien, Susan; Jain, Nitin; Verstovsek, Srdan; Wierda, William G; Keating, Michael J; Estrov, Zeev

    2014-06-12

    In chronic lymphocytic leukemia (CLL), stimulation of the B-cell receptor (BCR) triggers survival signals. Because in various cells activation of the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) pathway provides cells with survival advantage, we wondered whether BCR stimulation activates the JAK/STAT pathway in CLL cells. To stimulate the BCR we incubated CLL cells with anti-IgM antibodies. Anti-IgM antibodies induced transient tyrosine phosphorylation and nuclear localization of phosphorylated (p) STAT3. Immunoprecipitation studies revealed that anti-JAK2 antibodies coimmunoprecipitated pSTAT3 and pJAK2 in IgM-stimulated but not unstimulated CLL cells, suggesting that activation of the BCR induces activation of JAK2, which phosphorylates STAT3. Incubation of CLL cells with the JAK1/2 inhibitor ruxolitinib inhibited IgM-induced STAT3 phosphorylation and induced apoptosis of IgM-stimulated but not unstimulated CLL cells in a dose- and time-dependent manner. Whether ruxolitinib treatment would benefit patients with CLL remains to be determined.

  18. Leptin activates chicken growth hormone promoter without chicken STAT3 in vitro.

    PubMed

    Murase, Daisuke; Namekawa, Shoko; Ohkubo, Takeshi

    2016-01-01

    Leptin is an adipocyte-derived hormone that not only regulates food intake and energy homeostasis but also induces growth hormone (GH) mRNA expression and release, thereby controlling growth and metabolism in mammals. The molecular mechanism of leptin-induced regulation of GH gene transcription is unclear. The current study investigated the effects of leptin on the chicken GH (cGH) promoter and the molecular mechanism underlying leptin-induced cGH gene expression in vitro. Leptin activated the cGH promoter in the presence of chPit-1α in CHO cells stably expressing the chicken leptin receptor. Promoter activation did not require STAT-binding elements in the cGH promoter or STAT3 activity. However, JAK2 activation was required for leptin-dependent activity. JAK2-dependent pathways include p42/44 MAPK and PI3K, and inhibition of these pathways partially blocked leptin-induced cGH gene transcription. Although CK2 directly activates JAK2, a CK2 inhibitor blocked leptin-dependent activation of the cGH gene without affecting JAK2 phosphorylation. The CK2 inhibitor suppressed Erk1/2 and Akt phosphorylation. Additional data implicate Src family kinases in leptin-dependent cGH gene activation. These results suggest that leptin activates the cGH gene in the presence of chPit-1α via several leptin-activated kinases. Although further study is required, we suggest that the leptin-induced JAK2/p42/44 MAPK and JAK2/PI3K cascades are activated by Src-meditated CK2, leading to CBP phosphorylation and interaction with chPit-1α, resulting in transactivation of the cGH promoter.

  19. Association of Toxicity of Sorafenib and Sunitinib for Human Keratinocytes with Inhibition of Signal Transduction and Activator of Transcription 3 (STAT3)

    PubMed Central

    Yamamoto, Kazuhiro; Mizumoto, Atsushi; Nishimura, Kohji; Uda, Atsushi; Mukai, Akira; Yamashita, Kazuhiko; Kume, Manabu; Makimoto, Hiroo; Bito, Toshinori; Nishigori, Chikako; Nakagawa, Tsutomu; Hirano, Takeshi; Hirai, Midori

    2014-01-01

    Hand–foot skin reaction is a most common multi-kinase inhibitor-related adverse event. This study aimed to examine whether the toxicity of sorafenib and sunitinib for human keratinocytes was associated with inhibiting signal transduction and activator of transcription 3 (STAT3). We studied whether STAT3 activity affects sorafenib- and sunitinib-induced cell growth inhibition in HaCaT cells by WST-8 assay. Stattic enhanced the cell-growth inhibitory and apoptotic effects of sorafenib and sunitinib. HaCaT cells transfected with constitutively-active STAT3 (STAT3C) were resistant to the sorafenib- and sunitinib-induced cell growth inhibition. STAT3 activity decreased after short-term treatment with sorafenib and sunitinib in a dose-dependent manner and recovered after long-term treatment with sorafenib and sunitinib at low doses. Moreover, the expression of survivin and bcl-2 decreased after treatment with sorafenib and sunitinib was concomitant with variations in STAT3 activity. Sorafenib-induced STAT3 inhibition was mediated by regulation via MAPK pathways in HaCaT cells, while sunitinib-induced STAT3 inhibition was not. Thus, STAT3 activation mediating apoptosis suppressors may be a key factor in sorafenib and sunitinib-induced keratinocyte cytotoxicity. PMID:25013907

  20. Interleukin-6-Specific Activation of the C/EBPδ Gene in Hepatocytes Is Mediated by Stat3 and Sp1

    PubMed Central

    Cantwell, Carrie A.; Sterneck, Esta; Johnson, Peter F.

    1998-01-01

    C/EBPδ (CCAAT/enhancer binding protein δ) has been implicated as a regulator of acute-phase response (APR) genes in hepatocytes. Its expression increases dramatically in liver during the APR and can be induced in hepatic cell lines by interleukin-6 (IL-6), an acute-phase mediator that activates transcription of many APR genes. Here we have investigated the mechanism by which C/EBPδ expression is regulated by IL-6 in hepatoma cells. C/EBPδ promoter sequences to −125 bp are sufficient for IL-6 inducibility of a reporter gene and include an APR element (APRE) that is essential for IL-6 responsiveness. DNA binding experiments and transactivation assays demonstrate that Stat3, but not Stat1, interacts with this APRE. Two Sp1 sites, one of which is adjacent to the APRE, are required for IL-6 induction and transactivation by Stat3. Thus, Stat3 and Sp1 function cooperatively to activate the C/EBPδ promoter. Replacement of the APRE with Stat binding elements (SBEs) from the ICAM-1 or C/EBPβ promoter, both of which recognize both Stat1 and Stat3, confers responsiveness to gamma interferon, a cytokine that selectively activates Stat1. Sequence comparisons suggest that the distinct Stat binding specificities of the C/EBPδ and C/EBPβ SBEs are determined primarily by a single base pair difference. Our findings indicate that the cytokine specificity of C/EBPδ gene expression is governed by the APRE sequence. PMID:9528783

  1. TNFR2 Stimulation Promotes Mitochondrial Fusion via Stat3- and NF-kB-Dependent Activation of OPA1 Expression.

    PubMed

    Nan, Jinliang; Hu, Hengxun; Sun, Yong; Zhu, Lianlian; Wang, Yingchao; Zhong, Zhiwei; Zhao, Jing; Zhang, Na; Wang, Ya; Wang, Yaping; Ye, Jian; Zhang, Ling; Hu, Xinyang; Zhu, Wei; Wang, Jian'an

    2017-08-04

    Mitochondria are important cellular organelles and play essential roles in maintaining cell structure and function. Emerging evidence indicates that in addition to having proinflammatory and proapoptotic effects, TNFα (tumor necrosis factor α) can, under certain circumstances, promote improvements in mitochondrial integrity and function, phenomena that can be ascribed to the existence of TNFR2 (TNFα receptor 2). The present study aimed to investigate whether and how TNFR2 activation mediates the effects of TNFα on mitochondria. Freshly isolated neonatal mouse cardiac myocytes treated with shRNA targeting TNFR1 were used to study the effects of TNFR2 activation on mitochondrial function. Neonatal mouse cardiac myocytes exhibited increases in mitochondrial fusion, a change that was associated with increases in mitochondrial membrane potential, intracellular ATP levels, and oxygen consumption capacity. Importantly, TNFR2 activation-induced increases in OPA1 (optic atrophy 1) protein expression were responsible for the above enhancements, and these changes could be attenuated using siRNA targeting OPA1. Moreover, both Stat3 and RelA bound to the promoter region of OPA1 and their interactions synergistically upregulated OPA1 expression at the transcriptional level. Stat3 acetylation at lysine 370 or lysine 383 played a key role in the ability of Stat3 to form a supercomplex with RelA. Meanwhile, p300 modulated Stat3 acetylation in HEK293T (human embryonic kidney 293T) cells, and p300-mediated Stat3/RelA interactions played an indispensable role in OPA1 upregulation. Finally, TNFR2 activation exerted beneficial effects on OPA1 expression in an in vivo transverse aortic constriction model, whereby TNFR1-knockout mice exhibited better outcomes than in mice with both TNFR1 and TNFR2 knocked out. TNFR2 activation protects cardiac myocytes against stress by upregulating OPA1 expression. This process was facilitated by p300-mediated Stat3 acetylation and Stat3/Rel

  2. The hydrolyzed products of iridoid glycoside with β-glucosidase treatment exert anti-proliferative effects through suppression of STAT3 activation and STAT3-regulated gene products in several human cancer cells.

    PubMed

    Hwang, Hyundu; Kim, Chulwon; Kim, Sung-Moo; Kim, Wan-Seok; Choi, Seung-Hoon; Chang, Il-Moo; Ahn, Kwang Seok

    2012-01-01

    Iridoids belong to a group of monoterpene compounds with cyclopentane ring and found as mostly the glycoside forms in nature. They act primarily as the defense substances and found in various medicinal plants. Although many iridoids exhibit anti-inflammatory and anticancer activities, their molecular targets/pathways are not fully understood. Here, the antiproliferative effect of the hydrolyzed-iridoid product (H-iridoid) form through the STAT3 signaling pathways on tumor cells was investigated. H-iridoids were obtained from five iridoid glycosides with β-glucosidase treatment. The effects of several H-iridoids on cell viability and cell proliferation in tumor cells were measured by the MTT assay. The phosphorylation levels of STAT3, its regulatory molecules, and apoptosis by H-geniposide treatment in DU145 cells were investigated by immunoblots and flow cytometry. No single iridoid glycoside exerted any cytotoxicity in the tumor cells, whereas H-iridoids had significant cytotoxic, antiproliferative, and STAT3 inhibitory effects and revealed different potencies depending on their chemical structures. Among the H-iridoids tested, H-geniposide inhibited constitutive STAT3 activation through inhibiting upstream JAK1 and c-Src. Consistent with STAT3 inactivation, H-geniposide downregulated the expressions of Bcl-2, Bcl-xL, survivin, and cyclin D1; this correlated with the accumulation of cells in the sub-G1 phase of the cell cycle and the induction of apoptosis. Our results indicate that the hydrolysis of the glycosidic bond from iridoid glycoside is required for exhibiting cytotoxicity in tumor cells. H-geniposide is the most potent agent and a novel blocker of STAT3 activation in DU145 cells.

  3. The Shc1 adaptor simultaneously balances Stat1 and Stat3 activity to promote breast cancer immune suppression

    PubMed Central

    Ahn, Ryuhjin; Sabourin, Valérie; Bolt, Alicia M.; Hébert, Steven; Totten, Stephanie; De Jay, Nicolas; Festa, Maria Carolina; Young, Yoon Kow; Im, Young Kyuen; Pawson, Tony; Koromilas, Antonis E.; Muller, William J.; Mann, Koren K.; Kleinman, Claudia L.; Ursini-Siegel, Josie

    2017-01-01

    Tyrosine kinase signalling within cancer cells is central to the establishment of an immunosuppressive microenvironment. Although tyrosine kinase inhibitors act, in part, to augment adaptive immunity, the increased heterogeneity and functional redundancy of the tyrosine kinome is a hurdle to achieving durable responses to immunotherapies. We previously identified the Shc1 (ShcA) scaffold, a central regulator of tyrosine kinase signalling, as essential for promoting breast cancer immune suppression. Herein we show that the ShcA pathway simultaneously activates STAT3 immunosuppressive signals and impairs STAT1-driven immune surveillance in breast cancer cells. Impaired Y239/Y240-ShcA phosphorylation selectively reduces STAT3 activation in breast tumours, profoundly sensitizing them to immune checkpoint inhibitors and tumour vaccines. Finally, the ability of diminished tyrosine kinase signalling to initiate STAT1-driven immune surveillance can be overcome by compensatory STAT3 hyperactivation in breast tumours. Our data indicate that inhibition of pY239/240-ShcA-dependent STAT3 signalling may represent an attractive therapeutic strategy to sensitize breast tumours to multiple immunotherapies. PMID:28276425

  4. Highly pathogenic avian influenza H5N1 virus delays apoptotic responses via activation of STAT3.

    PubMed

    Hui, Kenrie P Y; Li, Hung Sing; Cheung, Man Chun; Chan, Renee W Y; Yuen, Kit M; Mok, Chris K P; Nicholls, John M; Peiris, J S Malik; Chan, Michael C W

    2016-06-27

    Highly pathogenic avian influenza (HPAI) H5N1 virus continues to pose pandemic threat, but there is a lack of understanding of its pathogenesis. We compared the apoptotic responses triggered by HPAI H5N1 and low pathogenic H1N1 viruses using physiologically relevant respiratory epithelial cells. We demonstrated that H5N1 viruses delayed apoptosis in primary human bronchial and alveolar epithelial cells (AECs) compared to H1N1 virus. Both caspase-8 and -9 were activated by H5N1 and H1N1 viruses in AECs, while H5N1 differentially up-regulated TRAIL. H5N1-induced apoptosis was reduced by TRAIL receptor silencing. More importantly, STAT3 knock-down increased apoptosis by H5N1 infection suggesting that H5N1 virus delays apoptosis through activation of STAT3. Taken together, we demonstrate that STAT3 is involved in H5N1-delayed apoptosis compared to H1N1. Since delay in apoptosis prolongs the duration of virus replication and production of pro-inflammatory cytokines and TRAIL from H5N1-infected cells, which contribute to orchestrate cytokine storm and tissue damage, our results suggest that STAT3 may play a previously unsuspected role in H5N1 pathogenesis.

  5. Artocarpus altilis (Parkinson) Fosberg Extracts and Geranyl Dihydrochalcone Inhibit STAT3 Activity in Prostate Cancer DU145 Cells.

    PubMed

    Jeon, Yoon Jung; Jung, Seung-Nam; Chang, Hyeyoun; Yun, Jieun; Lee, Chang Woo; Lee, Joonku; Choi, Sangho; Nash, Oyekanmi; Han, Dong Cho; Kwon, Byoung-Mog

    2015-05-01

    Artocarpus altilis (Parkinson) Fosberg has traditionally been used in Indonesia for the treatment of liver cirrhosis, hypertension, and diabetes. In many other countries, it is used for the treatment of malaria, yellow fever, and dengue fever. It has been reported that A. altilis extracts have antiatherosclerotic and cytoprotective effects, but its molecular targets in tumor cells are not yet fully understood. The A. altilis extracts and the partially purified fraction have been shown to inhibit STAT3 activity and the phosphorylation of STAT3 in a dose-dependent manner. To identify the active components, a bioassay-guided isolation of the partially purified fraction resulted in the identification of a geranyl dihydrochalcone, CG901. Its chemical structure was established on the basis of spectroscopic evidence and comparison with published data. The partially purified fraction and the isolated a geranyl dihydrochalcone, CG901, down-regulated the expression of STAT3 target genes, induced apoptosis in DU145 prostate cancer cells via caspase-3 and PARP degradation, and inhibited tumor growth in human prostate tumor (DU145) xenograft initiation model. These results suggest that A. altilis could be a good natural source and that the isolated compound will be a potential lead molecule for developing novel therapeutics against STAT3-related diseases, including cancer and inflammation. Copyright © 2015 John Wiley & Sons, Ltd.

  6. Sox2 Cooperates with Inflammation-Mediated Stat3 Activation in the Malignant Transformation of Foregut Basal Progenitor Cells

    PubMed Central

    Liu, Kuancan; Jiang, Ming; Lu, Yun; Chen, Hao; Sun, Jun; Wu, Shaoping; Ku, Wei-Yao; Nakagawa, Hiroshi; Kita, Yoshiaki; Natsugoe, Shoji; Peters, Jeffrey H.; Rustgi, Anil; Onaitis, Mark W.; Kiernan, Amy; Chen, Xiaoxin; Que, Jianwen

    2013-01-01

    Summary Sox2 regulates the self-renewal of multiple types of stem cells. Recent studies suggest it also plays oncogenic roles in the formation of squamous carcinoma in several organs, including the esophagus where Sox2 is predominantly expressed in the basal progenitor cells of the stratified epithelium. Here, we use mouse genetic models to reveal a novel mechanism by which Sox2 cooperates with microenvironmental signals to malignantly transform epithelial progenitor cells. Conditional overexpression of Sox2 in basal cells expands the progenitor population in both the esophagus and forestomach. Significantly, carcinoma only develops in the forestomach where pathological progression correlates with inflammation and nuclear localization of Stat3 in progenitor cells. Importantly, co-overexpression of Sox2 and activated Stat3 (Stat3C) also transforms esophageal basal cells but not the differentiated suprabasal cells. These findings indicate basal stem/progenitor cells are the cells-of-origin of squamous carcinoma and that cooperation between Sox2 and microenvironment-activated Stat3 is required for Sox2-driven tumorigenesis. PMID:23472872

  7. The Shc1 adaptor simultaneously balances Stat1 and Stat3 activity to promote breast cancer immune suppression.

    PubMed

    Ahn, Ryuhjin; Sabourin, Valérie; Bolt, Alicia M; Hébert, Steven; Totten, Stephanie; De Jay, Nicolas; Festa, Maria Carolina; Young, Yoon Kow; Im, Young Kyuen; Pawson, Tony; Koromilas, Antonis E; Muller, William J; Mann, Koren K; Kleinman, Claudia L; Ursini-Siegel, Josie

    2017-03-09

    Tyrosine kinase signalling within cancer cells is central to the establishment of an immunosuppressive microenvironment. Although tyrosine kinase inhibitors act, in part, to augment adaptive immunity, the increased heterogeneity and functional redundancy of the tyrosine kinome is a hurdle to achieving durable responses to immunotherapies. We previously identified the Shc1 (ShcA) scaffold, a central regulator of tyrosine kinase signalling, as essential for promoting breast cancer immune suppression. Herein we show that the ShcA pathway simultaneously activates STAT3 immunosuppressive signals and impairs STAT1-driven immune surveillance in breast cancer cells. Impaired Y239/Y240-ShcA phosphorylation selectively reduces STAT3 activation in breast tumours, profoundly sensitizing them to immune checkpoint inhibitors and tumour vaccines. Finally, the ability of diminished tyrosine kinase signalling to initiate STAT1-driven immune surveillance can be overcome by compensatory STAT3 hyperactivation in breast tumours. Our data indicate that inhibition of pY239/240-ShcA-dependent STAT3 signalling may represent an attractive therapeutic strategy to sensitize breast tumours to multiple immunotherapies.

  8. Co-activation of STAT3 and YES-Associated Protein 1 (YAP1) Pathway in EGFR-Mutant NSCLC.

    PubMed

    Chaib, Imane; Karachaliou, Niki; Pilotto, Sara; Codony Servat, Jordi; Cai, Xueting; Li, Xuefei; Drozdowskyj, Ana; Servat, Carles Codony; Yang, Jie; Hu, Chunping; Cardona, Andres Felipe; Vivanco, Guillermo Lopez; Vergnenegre, Alain; Sanchez, Jose Miguel; Provencio, Mariano; de Marinis, Filipo; Passaro, Antonio; Carcereny, Enric; Reguart, Noemi; Campelo, Charo Garcia; Teixido, Christina; Sperduti, Isabella; Rodriguez, Sonia; Lazzari, Chiara; Verlicchi, Alberto; de Aguirre, Itziar; Queralt, Cristina; Wei, Jia; Estrada, Roger; Puig de la Bellacasa, Raimon; Ramirez, Jose Luis; Jacobson, Kirstine; Ditzel, Henrik J; Santarpia, Mariacarmela; Viteri, Santiago; Molina, Migual Angel; Zhou, Caicun; Cao, Peng; Ma, Patrick C; Bivona, Trever G; Rosell, Rafael

    2017-09-01

    The efficacy of epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) in EGFR-mutant non-small cell lung cancer (NSCLC) is limited by adaptive activation of cell survival signals. We hypothesized that both signal transducer and activator of transcription 3 (STAT3) and Src-YES-associated protein 1 (YAP1) signaling are dually activated during EGFR TKI treatment to limit therapeutic response. We used MTT and clonogenic assays, immunoblotting, and quantitative polymerase chain reaction to evaluate the efficacy of EGFR TKI alone and in combination with STAT3 and Src inhibition in three EGFR-mutant NSCLC cell lines. The Chou-Talalay method was used for the quantitative determination of drug interaction. We examined tumor growth inhibition in one EGFR-mutant NSCLC xenograft model (n = 4 mice per group). STAT3 and YAP1 expression was evaluated in tumors from 119 EGFR-mutant NSCLC patients (64 in an initial cohort and 55 in a validation cohort) by quantitative polymerase chain reaction. Kaplan-Meier and Cox regression analyses were used to assess the correlation between survival and gene expression. All statistical tests were two-sided. We discovered that lung cancer cells survive initial EGFR inhibitor treatment through activation of not only STAT3 but also Src-YAP1 signaling. Cotargeting EGFR, STAT3, and Src was synergistic in two EGFR-mutant NSCLC cell lines with a combination index of 0.59 (95% confidence interval [CI] = 0.54 to 0.63) for the PC-9 and 0.59 (95% CI = 0.54 to 0.63) for the H1975 cell line. High expression of STAT3 or YAP1 predicted worse progression-free survival (hazard ratio [HR] = 3.02, 95% CI = 1.54 to 5.93, P = .001, and HR = 2.57, 95% CI = 1.30 to 5.09, P = .007, respectively) in an initial cohort of 64 EGFR-mutant NSCLC patients treated with firstline EGFR TKIs. Similar results were observed in a validation cohort. Our study uncovers a coordinated signaling network centered on both STAT3 and Src-YAP signaling

  9. The Role of Stat3 Activation in Androgen Receptor Signaling and Prostate Cancer

    DTIC Science & Technology

    2005-07-01

    Lapidot oligonucleotides with the Stat3 siRNA identified in Z, Leeder JS, Freedman M, Cohen A, Gazit A, Levitzki A, this study in prostate cancer...3. Catlett-Falcone R, Landowski TH, Oshiro MM, Turkson J, stranded RNA directs the ATP-dependent cleavage of mRNA at Levitzki A, Savino R, Ciliberto G...Lee SO, Xiao X, Pflug B, Nelson JB, Gao AC. Levitzki A, Savino R, Ciliberto G, Moscinski L, Fernandez-Luna Stat3 enhances the growth of LNCaP human

  10. Antiproliferative effect of SOCS-1 through the suppression of STAT3 and p38 MAPK activation in gastric cancer cells.

    PubMed

    Souma, Yoshihito; Nishida, Toshirou; Serada, Satoshi; Iwahori, Kota; Takahashi, Tsuyoshi; Fujimoto, Minoru; Ripley, Barry; Nakajima, Kiyokazu; Miyazaki, Yasuaki; Mori, Masaki; Doki, Yuichiro; Sawa, Yoshiki; Naka, Tetsuji

    2012-09-15

    Inflammation is a crucial driving force in the development of gastric cancers (GCs). Accordingly, persistent activation of STAT3, a transcription factor pivotal in regulating both inflammation and oncogenesis, is often detected in GC, although its mechanism remains elusive. Suppressor of cytokine signaling-1 (SOCS-1) is a negative regulator of proinflammatory cytokine signaling and SOCS-1 gene methylation is frequently detected in various cancers including GC. However, the significance of SOCS-1 methylation in GC cells remains unexplored. Our study is undertaken to evaluate the role of SOCS-1 in GC cell proliferation and its effect on signaling pathways in GC cells. Among five GC cell lines, SOCS-1 gene was methylated in all cell lines and constitutive STAT3 phosphorylation with elevated endogenous IL-6 production was detected in two cell lines (NUGC-3 and AGS). Unexpectedly, anti-IL-6R antibody inhibited neither cell proliferation nor STAT3 phosphorylation in NUGC-3 and AGS. In contrast, enforced SOCS-1 expression by adenoviral vector (AdSOCS-1) markedly suppressed STAT3 phosphorylation and proliferation of NUGC-3 and AGS cells in vitro. Interestingly, the antiproliferative effect of SOCS-1 was attributable not only to the inhibition of STAT3 but also to that of p38 MAPK activity, and chemical inhibitors of JAK/STAT and p38 MAPK signaling effectively suppressed proliferation of these GC cells. Furthermore, treatment with AdSOCS-1 in vivo significantly suppressed GC proliferation in a xenograft model. These results suggest that SOCS-1 gene methylation is a critical step in the development of GC, and enforced expression of SOCS-1 may represent a novel therapeutic approach for the treatment of GC. Copyright © 2011 UICC.

  11. PIAS3 expression in human gastric carcinoma and its adjacent non-tumor tissues.

    PubMed

    Liu, Liang-ming; Yan, Ming-guo; Yang, Dao-hua; Sun, Wei-wei; Zhang, Ji-xiang

    2011-05-01

    PIAS3 is the endogenous inhibitor of STAT3, which has been implicated in the pathogenesis of many cancers. However, the effect of PIAS3 on human tumors remains elusive. The aim of this article is to investigate the expression of PIAS3 in gastric carcinoma and its adjacent non-tumor tissues. Samples were taken from 30 patients with gastric cancer, which included tumor or non-tumor tissues in the excised sections. The expression of PIAS3 protein was detected by immunocytochemistry, and that of mRNA by in situ hybridization. The results were semi-quantitative analyzed by using cell count and color depth to stage. The expression levels of PIAS3 protein and mRNA were significantly lower in gastric cancerous tissues than in its adjacent non-tumor tissues, and had a close relation with tumor size and differentiation, but not with age, gender and lymphatic metastasis in gastric carcinoma. The more large in size and poorly in differentiation, the more low PIAS3 expression was. Loss of PIAS3 expression may be an important characteristic of gastric cancer and suggest vicious degree of the tumor. Copyright © 2011 Elsevier Masson SAS. All rights reserved.

  12. Activation of signal transducer and activator of transcription 3 (STAT3) signaling in EGFR mutant non-small-cell lung cancer (NSCLC).

    PubMed

    Codony-Servat, Carles; Codony-Servat, Jordi; Karachaliou, Niki; Molina, Miguel Angel; Chaib, Imane; Ramirez, Jose Luis; de Los Llanos Gil, Maria; Solca, Flavio; Bivona, Trever G; Rosell, Rafael

    2017-07-18

    Gefitinib, erlotinib or afatinib are the current treatment for non-small-cell lung cancer (NSCLC) harboring an activating mutation of the epidermal growth factor receptor (EGFR), but less than 5% of patients achieve a complete response and the median progression-free survival is no longer than 12 months. Early adaptive resistance can occur as soon as two hours after starting treatment by activating signal transducer and activation of transcription 3 (STAT3) signaling. We investigated the activation of STAT3 in a panel of gefitinib-sensitive EGFR mutant cell lines, and gefitinib-resistant PC9 cell lines developed in our laboratory. Afatinib has great activity in gefitinib-sensitive as well as in gefitinib-resistant EGFR mutant NSCLC cell lines. However, afatinib therapy causes phosphorylation of STAT3 tyrosine 705 (pSTAT3Tyr705) and elevation of STAT3 and RANTES mRNA levels. The combination of afatinib with TPCA-1 (a STAT3 inhibitor) ablated pSTAT3Tyr705 and down-regulated STAT3 and RANTES mRNA levels with significant growth inhibitory effect in both gefitinib-sensitive and gefitinib-resistant EGFR mutant NSCLC cell lines. Aldehyde dehydrogenase positive (ALDH+) cells were still observed with the combination at the time that Hairy and Enhancer of Split 1 (HES1) mRNA expression was elevated following therapy. Although the combination of afatinib with STAT3 inhibition cannot eliminate the potential problem of a remnant cancer stem cell population, it represents a substantial advantage and opportunity to further prolong progression free survival and probably could increase the response rate in comparison to the current standard of single therapy.

  13. Hepatitis C virus and its protein NS4B activate the cancer-related STAT3 pathway via the endoplasmic reticulum overload response.

    PubMed

    Kong, Lingbao; Li, Shanshan; Yu, Xilan; Fang, Xiaonan; Xu, Ahui; Huang, Mingjie; Wu, Xiaoyu; Guo, Yunli; Guo, Fenglin; Xu, Jin

    2016-08-01

    Oxidative stress induces the activation of signal transducer and activator of transcription 3 (STAT3), which plays an important role in hepatocellular carcinoma (HCC). We have previously reported that hepatitis C virus (HCV) and its protein NS4B induce the production of reactive oxygen species (ROS) via the endoplasmic reticulum overload response (EOR) in human hepatocytes. Here, we found that NS4B and HCV induce STAT3 activation and stimulate the expression of cancer-related STAT3 target genes, including VEGF, c-myc, MMP-9 and Mcl-1, by EOR in human hepatocytes. Moreover, the cancer-related STAT3 pathway activated by NS4B and HCV via EOR were found to promote human hepatocyte viability. Taken together, these findings revealed that HCV NS4B might contribute to HCC by activating the EOR-mediated cancer-related STAT3 pathway, and this could provide novel insights into HCV-induced HCC.

  14. Signal transducer and activator of transcription 3 (Stat3) regulates host defense and protects mice against herpes simplex virus-1 (HSV-1) infection.

    PubMed

    Hsia, Hung-Ching; Stopford, Charles M; Zhang, Zhigang; Damania, Blossom; Baldwin, Albert S

    2016-12-13

    Signal transducer and activator of transcription 3 (STAT3) mediates cellular responses to multiple cytokines, governs gene expression, and regulates the development and activation of immune cells. STAT3 also modulates reactivation of latent herpes simplex virus-1 (HSV-1) in ganglia. However, it is unclear how STAT3 regulates the innate immune response during the early phase of HSV-1 lytic infection. Many cell types critical for the innate immunity are derived from the myeloid lineage. Therefore, in this study, we used myeloid-specific Stat3 knockout mice to investigate the role of STAT3 in the innate immune response against HSV-1. Our results demonstrate that Stat3 knockout bone marrow-derived macrophages (BMMs) expressed decreased levels of interferon-α (IFN-α) and interferon-stimulated genes (ISGs) upon HSV-1 infection. In vivo, knockout mice were more susceptible to HSV-1, as marked by higher viral loads and more significant weight loss. Splenic expression of IFN-α and ISGs was reduced in the absence of STAT3, indicating that STAT3 is required for optimal type I interferon response to HSV-1. Expression of TNF-α and IL-12, cytokines that have been shown to limit HSV-1 replication and pathogenesis, was also significantly lower in knockout mice. Interestingly, Stat3 knockout mice failed to expand the CD8(+) conventional DC (cDC) population upon HSV-1 infection, and this was accompanied by impaired NK and CD8 T cell activation. Collectively, our data demonstrate that myeloid-specific Stat3 deletion causes defects in multiple aspects of the immune system and that STAT3 has a protective role at the early stage of systemic HSV-1 infection.

  15. MicroRNA-199a-5p promotes tumour growth by dual-targeting PIAS3 and p27 in human osteosarcoma

    PubMed Central

    Wang, Chen; Ba, Ximing; Guo, Yu; Sun, Defang; Jiang, Haoyang; Li, Wentao; Huang, Zhen; Zhou, Guangxin; Wu, Sujia; Zhang, Junfeng; Chen, Jiangning

    2017-01-01

    Osteosarcoma (OS) is the most common primary bone malignancy and remains a leading cause of cancer-related deaths in adolescents. Emerging evidence indicates that microRNAs (miRNAs) are correlated with clinical and biological characteristics of OS. However, the involvement of miR-199a-5p in OS development remains unclear. In this study, we examined the function of miR-199a-5p in vitro and in vivo. The results showed that miR-199a-5p was significantly up-regulated in OS patient tissues and cells. The inhibition of miR-199a-5p led to a significant decrease in cell proliferation and tumour growth. We further demonstrated that miR-199a-5p could directly bind to the 3′UTRs of the mRNA of both PIAS3 and p27 and mediate a decrease in the protein levels of PIAS3 and p27, thereby stimulating STAT3 activation and cell cycle progression in OS cells. Rescue experiments of PIAS3 and p27 further revealed that PIAS3 and p27 were functional targets of miR-199a-5p. Moreover, enhancing the expressions of both PIAS3 and p27 using miR-199a-5p-targeted inhibitors in an OS xenograft model was shown to be a promising approach for OS clinical therapy. Our findings indicate that the pathway of miR-199a-5p targeting both PIAS3 and p27 is a possible mechanism that contributes to tumour growth in OS. PMID:28120918

  16. [ITF increases the transcriptional activity of ITF promoter via the JAK-STAT3 signal transduction pathway].

    PubMed

    Sun, Yong; LE, Juan; Chen, Baojun; Pan, Xiaofeng; Zhang, Fang

    2014-11-01

    To investigate the eff ect of intestinal trefoil factor (ITF) on the transcriptional activity of ITF promoter and to explore the regulatory mechanism of Janus kinase/signal transducers and activators of transcription (JAK/STAT) on ITF promoter. The 5' flanking sequence of the ITF gene was cloned from human whole blood genomic DNA by PCR. ITF promoter fragment was cloned and inserted into the pGL3-Basic vector to construct recombinant vector. ITF promoter vector was stimulated with ITF at various concentrations and the luciferase activity was measured. The JAK-STAT3 signal transduction pathway was then blocked by a specific inhibitor AG490 to determine the signal pathway involved in ITF promoter activity. Restriction endonuclease analysis and DNA sequencing confirmed that the recombinant plasmid, containing ITF promoter, was constructed successfully. After transient transfection, the activity of ITF promoter was increased significantly in the presence of ITF (P<0.05). Blockage of the JAK-STAT3 signal transduction pathway with AG490 significantly reduced the ITF promoter activity (P<0.05). ITF increases the transcriptional activity of ITF promoter via the JAK-STAT3 signal transduction pathway.

  17. A single polymorphic amino acid on Toxoplasma gondii kinase ROP16 determines the direct and strain-specific activation of Stat3

    PubMed Central

    Yamamoto, Masahiro; Standley, Daron M.; Takashima, Seiji; Saiga, Hiroyuki; Okuyama, Megumi; Kayama, Hisako; Kubo, Emi; Ito, Hiroshi; Takaura, Mutsumi; Matsuda, Tadashi; Soldati-Favre, Dominique

    2009-01-01

    Infection by Toxoplasma gondii down-regulates the host innate immune responses, such as proinflammatory cytokine production, in a Stat3-dependent manner. A forward genetic approach recently demonstrated that the type II strain fails to suppress immune responses because of a potential defect in a highly polymorphic parasite-derived kinase, ROP16. We generated ROP16-deficient type I parasites by reverse genetics and found a severe defect in parasite-induced Stat3 activation, culminating in enhanced production of interleukin (IL) 6 and IL-12 p40 in the infected macrophages. Furthermore, overexpression of ROP16 but not ROP18 in mammalian cells resulted in Stat3 phosphorylation and strong activation of Stat3-dependent promoters. In addition, kinase-inactive ROP16 failed to activate Stat3. Comparison of type I and type II ROP16 revealed that a single amino acid substitution in the kinase domain determined the strain difference in terms of Stat3 activation. Moreover, ROP16 bound Stat3 and directly induced phosphorylation of this transcription factor. These results formally establish an essential and direct requirement of ROP16 in parasite-induced Stat3 activation and the significance of a single amino acid replacement in the function of type II ROP16. PMID:19901082

  18. Staphylococcal enterotoxin A (SEA) stimulates STAT3 activation and IL-17 expression in cutaneous T-cell lymphoma

    PubMed Central

    Willerslev-Olsen, Andreas; Krejsgaard, Thorbjørn; Lindahl, Lise M.; Litvinov, Ivan V.; Fredholm, Simon; Petersen, David L.; Nastasi, Claudia; Gniadecki, Robert; Mongan, Nigel P.; Sasseville, Denis; Wasik, Mariusz A.; Bonefeld, Charlotte M.; Geisler, Carsten; Woetmann, Anders; Iversen, Lars; Kilian, Mogens; Koralov, Sergei B.

    2016-01-01

    Cutaneous T-cell lymphoma (CTCL) is characterized by proliferation of malignant T cells in a chronic inflammatory environment. With disease progression, bacteria colonize the compromised skin barrier and half of CTCL patients die of infection rather than from direct organ involvement by the malignancy. Clinical data indicate that bacteria play a direct role in disease progression, but little is known about the mechanisms involved. Here, we demonstrate that bacterial isolates containing staphylococcal enterotoxin A (SEA) from the affected skin of CTCL patients, as well as recombinant SEA, stimulate activation of signal transducer and activator of transcription 3 (STAT3) and upregulation of interleukin (IL)-17 in immortalized and primary patient–derived malignant and nonmalignant T cells. Importantly, SEA induces STAT3 activation and IL-17 expression in malignant T cells when cocultured with nonmalignant T cells, indicating an indirect mode of action. In accordance, malignant T cells expressing an SEA-nonresponsive T-cell receptor variable region β chain are nonresponsive to SEA in monoculture but display strong STAT3 activation and IL-17 expression in cocultures with SEA-responsive nonmalignant T cells. The response is induced via IL-2 receptor common γ chain cytokines and a Janus kinase 3 (JAK3)-dependent pathway in malignant T cells, and blocked by tofacitinib, a clinical-grade JAK3 inhibitor. In conclusion, we demonstrate that SEA induces cell cross talk–dependent activation of STAT3 and expression of IL-17 in malignant T cells, suggesting a mechanism whereby SEA-producing bacteria promote activation of an established oncogenic pathway previously implicated in carcinogenesis. PMID:26738536

  19. Alantolactone Induces Apoptosis in HepG2 Cells through GSH Depletion, Inhibition of STAT3 Activation, and Mitochondrial Dysfunction

    PubMed Central

    Khan, Muhammad; Li, Ting; Ahmad Khan, Muhammad Khalil; Rasul, Azhar; Nawaz, Faisal; Sun, Meiyan; Zheng, Yongchen; Ma, Tonghui

    2013-01-01

    Signal transducer and activator of transcription 3 (STAT3) constitutively expresses in human liver cancer cells and has been implicated in apoptosis resistance and tumorigenesis. Alantolactone, a sesquiterpene lactone, has been shown to possess anticancer activities in various cancer cell lines. In our previous report, we showed that alantolactone induced apoptosis in U87 glioblastoma cells via GSH depletion and ROS generation. However, the molecular mechanism of GSH depletion remained unexplored. The present study was conducted to envisage the molecular mechanism of alantolactone-induced apoptosis in HepG2 cells by focusing on the molecular mechanism of GSH depletion and its effect on STAT3 activation. We found that alantolactone induced apoptosis in HepG2 cells in a dose-dependent manner. This alantolactone-induced apoptosis was found to be associated with GSH depletion, inhibition of STAT3 activation, ROS generation, mitochondrial transmembrane potential dissipation, and increased Bax/Bcl-2 ratio and caspase-3 activation. This alantolactone-induced apoptosis and GSH depletion were effectively inhibited or abrogated by a thiol antioxidant, N-acetyl-L-cysteine (NAC). The data demonstrate clearly that intracellular GSH plays a central role in alantolactone-induced apoptosis in HepG2 cells. Thus, alantolactone may become a lead chemotherapeutic candidate for the treatment of liver cancer. PMID:23533997

  20. Spinal leptin contributes to the development of morphine antinociceptive tolerance by activating the STAT3-NMDA receptor pathway in rats.

    PubMed

    Hu, Fen; Cui, Yu; Guo, Ruixian; Chen, Jingfu; Guo, Runming; Shen, Ning; Hua, Xiaoxiao; Mo, Liqiu; Feng, Jianqiang

    2014-08-01

    Leptin, an adipokine synthesized mainly by non‑neuronal tissues, has been reported to contribute to the pathogenesis of neuropathic pain. It has been hypothesized that morphine tolerance and neuropathic pain share some common pathological mechanisms. The present study was designed to examine whether spinal leptin is implicated in the development of morphine antinociceptive tolerance, and whether spinal leptin induces the activation of signal transducer and activator of transcription 3 (STAT3) signaling pathway and the NR1 subunit of N‑methyl‑D‑aspartate (NMDA) receptor, in morphine antinociceptive tolerance in rats. The results demonstrated that intrathecal (i.t.) administration of a leptin antagonist (LA) prevented the development of morphine antinociceptive tolerance in rats. Further studies revealed that the levels of the spinal leptin and the leptin receptor (Ob‑R) were time‑dependently increased following chronic morphine treatment. Mechanistic examination indicated that chronic morphine triggered activation of the STAT3 pathway and an increase in the expression of the NR1 subunit of the NMDA receptor, which was ameliorated by i.t. administration of AG490 [a Janus kinase (JAK)‑STAT inhibitor]. The increased activation of STAT3 and the NR1 subunit was markedly attenuated by i.t. treatment with LA. In addition, the spinal administration of AG490 or MK‑801 (a non‑competitive NMDA receptor inhibitor) blocked the development of morphine antinociceptive tolerance. Taken together, these results have demonstrated, for the first time, to the best of our knowledge, that spinal leptin contributes to the development of morphine antinociceptive tolerance by activating the spinal STAT3‑NMDA receptor pathway.

  1. GITRL modulates the activities of p38 MAPK and STAT3 to promote Th17 cell differentiation in autoimmune arthritis.

    PubMed

    Tang, Xinyi; Tian, Jie; Ma, Jie; Wang, Jiemin; Qi, Chen; Rui, Ke; Wang, Yungang; Xu, Huaxi; Lu, Liwei; Wang, Shengjun

    2016-02-23

    The glucocorticoid-induced TNFR family-related protein (GITR) and its ligand play a critical role in the pathogenesis of autoimmune arthritis by enhancing the Th17 cell response, but their molecular mechanisms remain largely unclear. This study aims to define the role of p38 mitogen-activated protein kinases (MAPK) and signal transducer and activator of transcription 3 (STAT3) signaling in GITRL-induced Th17 cells in autoimmune arthritis. We found that the p38 phosphorylation was enhanced by GITRL in activated CD4+T cells, and the p38 inhibitor restrained the GITRL-induced Th17 cell expansion in a dose-dependent manner. Moreover, there was decreased STAT3 activity on Tyr705 and Ser727 with the p38 inhibitor in vitro. Notably, the p38 inhibitor could prevent GITRL-treated arthritis progression and markedly decrease the Th17 cell percentages. The phosphorylation of the Tyr705 site was significantly lower in the GITRL-treated CIA mice administrated with the p38 inhibitor. A significantly higher phosphorylation of p38 was detected in RA patients and had a positive relationship with the serum level of anti-cyclic citrullinated peptide (anti-CCP) antibody. Our findings have indicated that GITRL could promote Th17 cell differentiation by p38 MAPK and STAT3 signaling in autoimmune arthritis.

  2. JAK/STAT3 and Smad3 activities are required for the wound healing properties of Periplaneta americana extracts

    PubMed Central

    Song, Qin; Xie, Yuxin; Gou, Qiheng; Guo, Xiaoqiang; Yao, Qian; Gou, Xiaojun

    2017-01-01

    Periplaneta americana extracts (PAEs) play a crucial role in skin wound healing. However, their molecular effects and signaling pathways in regenerating tissues and cells are not clear. In this study, we refined the PAE from Periplaneta americana to investigate the mechanisms underlying skin wound healing. The human keratinocyte line HaCaT was selected and a mouse model of deep second-degree thermal burn was established for in vitro and in vivo studies, respectively. PAE treatment induced the proliferation and migration of HaCaT cells and wound healing in the burn model. Furthermore, the effects of PAE on wound healing were found to depend on the Janus-activated kinase/signal transducer and activator of transcription 3 (JAK/STAT3) pathway and Smad3 activities, according to western blot analysis and immunohistochemical (IHC) assays in vitro and in vivo. Pretreatment with a STAT3 inhibitor blocked the cell proliferation and migration induced by PAE. The results indicate the wound-healing function of PAE via enhanced JAK/STAT3 signaling and Smad3 activities. Our studies provide a theoretical basis underlying the role of PAE in cutaneous wound healing. PMID:28656220

  3. STAT3 and HIF1α cooperatively activate HIF1 target genes in MDA-MB-231 and RCC4 cells.

    PubMed

    Pawlus, M R; Wang, L; Hu, C-J

    2014-03-27

    Solid tumors often exhibit simultaneously inflammatory and hypoxic microenvironments. The 'signal transducer and activator of transcription-3' (STAT3)-mediated inflammatory response and the hypoxia-inducible factor (HIF)-mediated hypoxia response have been independently shown to promote tumorigenesis through the activation of HIF or STAT3 target genes and to be indicative of a poor prognosis in a variety of tumors. We report here for the first time that STAT3 is involved in the HIF1, but not HIF2-mediated hypoxic transcriptional response. We show that inhibiting STAT3 activity in MDA-MB-231 and RCC4 cells by a STAT3 inhibitor or STAT3 small interfering RNA significantly reduces the levels of HIF1, but not HIF2 target genes in spite of normal levels of hypoxia-inducible transcription factor 1α (HIF1α) and HIF2α protein. Mechanistically, STAT3 activates HIF1 target genes by binding to HIF1 target gene promoters, interacting with HIF1α protein and recruiting coactivators CREB binding protein (CBP) and p300, and RNA polymerase II (Pol II) to form enhanceosome complexes that contain HIF1α, STAT3, CBP, p300 and RNA Pol II on HIF1 target gene promoters. Functionally, the effect of STAT3 knockdown on proliferation, motility and clonogenic survival of tumor cells in vitro is phenocopied by HIF1α knockdown in hypoxic cells, whereas STAT3 knockdown in normoxic cells also reduces cell proliferation, motility and clonogenic survival. This indicates that STAT3 works with HIF1 to activate HIF1 target genes and to drive HIF1-depedent tumorigenesis under hypoxic conditions, but also has HIF-independent activity in normoxic and hypoxic cells. Identifying the role of STAT3 in the hypoxia response provides further data supporting the effectiveness of STAT3 inhibitors in solid tumor treatment owing to their usefulness in inhibiting both the STAT3 and HIF1 pro-tumorigenic signaling pathways in some cancer types.

  4. Gpr110 deficiency decelerates carcinogen-induced hepatocarcinogenesis via activation of the IL-6/STAT3 pathway

    PubMed Central

    Ma, Benting; Zhu, Junjie; Tan, Juan; Mao, Yulei; Tang, Lingyun; Shen, Chunling; Zhang, Hongxing; Kuang, Ying; Fei, Jian; Yang, Xiao; Wang, Zhugang

    2017-01-01

    Hepatocarcinogenesis is a complex process that includes pronounced necroinflammation, unregulated hepatocyte damage, subsequent extensive fibrosis, and carcinogenesis. GPR110 was an adhesion G protein-coupled receptor. Analysis of the expression pattern of Gpr110 in mice displayed that Gpr110 was expressed highly in liver, implicating the tissue compartments where Gpr110 could execute its functions, the role of Gpr110 in the physiological and pathological state of liver remains unclear. Based on a Gpr110 knockout mouse model, we evaluated the role of Gpr110 in hepatocarcinogenesis by using a carbon tetrachloride (CCl4)-induced liver injury and fibrosis model, as well as diethylnitrosamine (DEN) plus CCl4-induced liver cancer model. In this study, we found subdued chronic liver injury, reduced compensatory proliferation, lower liver fibrosis, but enhanced inflammation occurred in Gpr110-/- mice during CCl4 challenge. In addition, Gpr110-/- mice were resistant to liver tumorigenesis induced by DEN plus CCl4 injection. Molecular mechanisms underlying these differences correlated with augmented activation of the IL-6/STAT3 pathway, which exerted hepatoprotective effects during liver damage, fibrosis, and oncogenesis in Gpr110-/- mice. Furthermore, pharmacological inhibition of the activation of the IL-6/STAT3 pathway enhanced hepatic fibrosis and promoted DEN plus CCl4-induced carcinogenesis in Gpr110-/- mice. In summary, absence of Gpr110 decelerates liver fibrosis/cirrhosis progressing into tumorigenesis, due to strengthening activation of the IL-6/STAT3 pathway, leading to a weaker liver injury and fibrosis microenvironment. It is indicated that targeting Gpr110 and activating the IL-6/STAT3 pathway may be considered to be preventive methods for some cirrhosis transition.

  5. Activation of TLR4/STAT3 signaling in VTA contributes to the acquisition and maintenance of morphine-induced conditioned place preference.

    PubMed

    Chen, Jia-Xin; Huang, Kang-Mei; Liu, Meng; Jiang, Jin-Xiang; Liu, Jian-Peng; Zhang, Yu-Xiang; Yang, Chen; Xin, Wen-Jun; Zhang, Xue-Qin

    2017-09-29

    Morphine, commonly used to relieve the acute or chronic pain, has a high potential for addiction and exerts rewarding effects via a critical role for mesolimbic dopamine system. Studies suggest that addiction-related behavior is highly associated with inflammatory immune response, but the mechanisms are poorly understood. The present study showed that intra-VTA microinjection of TLR4 antagonist LPS-RS prevented the acquisition and maintenance, but not the expression, of morphine-induced CPP in rats. In addition, chronic morphine treatment significantly activated STAT3 on day 6 and 11 in VTA, and bilateral microinjection of STAT3 inhibitor S3I-201 into the VTA suppressed the acquisition and maintenance of morphine-induced CPP in rats. Furthermore, local knockout of STAT3 by injection of the AAV-Cre-GFP into the VTA area of STAT3(flox/flox) mice also significantly impaired the acquisition of morphine CPP. Importantly, the TLR4 expression is colocalized with p-STAT3-positive cell in VTA, and repeated injection of LPS-RS significantly attenuated the STAT3 activation in VTA induced by chronic morphine treatment. Collectively, these data suggest that TLR4/STAT3 signaling pathway in VTA might play a critical role in the acquisition and maintenance of morphine CPP, and provides new evidence that TLR4/STAT3 signaling pathway might be a potential target for treatment of morphine addiction. Copyright © 2017. Published by Elsevier B.V.

  6. WP1066, a small molecule inhibitor of the JAK/STAT3 pathway, inhibits ceramide glucosyltransferase activity.

    PubMed

    Tsurumaki, Hirotaka; Katano, Hikaru; Sato, Kousuke; Imai, Ryou; Niino, Satomi; Hirabayashi, Yoshio; Ichikawa, Shinichi

    2017-09-16

    WP1066 is a well-known inhibitor of the JAK/STAT3 signaling pathway. By a screen of known small molecule inhibitors of various enzymes and protein factors, we identified WP1066 as a ceramide glucosyltransferase inhibitor. Ceramide glucosyltransferase catalyzes the first glycosylation step during glycosphingolipid synthesis. We found that WP1066 inhibited the activity of ceramide glucosyltransferase with an IC50 of 7.2 μM, and that its action was independent of JAK/STAT3 pathway blockade. Moreover, the modes of inhibition of ceramide glucosyltransferase were uncompetitive with respect to both C6-NBD-cermide and UDP-glucose. Copyright © 2017 Elsevier Inc. All rights reserved.

  7. Dual inhibition of Janus and Src family kinases by novel indirubin derivative blocks constitutively-activated Stat3 signaling associated with apoptosis of human pancreatic cancer cells.

    PubMed

    Nam, Sangkil; Wen, Wei; Schroeder, Anne; Herrmann, Andreas; Yu, Hua; Cheng, Xinlai; Merz, Karl-Heinz; Eisenbrand, Gerhard; Li, Hongzhi; Yuan, Yate-Ching; Jove, Richard

    2013-06-01

    Constitutively-activated JAK/Stat3 or Src/Stat3 signaling plays a crucial role in tumor cell survival, proliferation, angiogenesis and immune suppression. Activated JAK/Stat3 or Src/Stat3 has been validated as a promising molecular target for cancer therapy. However, prolonged inhibition of Src family kinases (SFKs) leads to reactivation of signal transducer and activator of transcript 3 (Stat3) and tumor cell survival through altered JAK/Stat3 interaction. This compensatory feedback suggests that dual inhibition of Janus kinases (JAKs) and SFKs might be a promising strategy for targeting downstream Stat3 signaling in the clinic. In this study, we identify that the natural product derivative E738 is a novel dual inhibitor of JAKs and SFKs. The IC(50) values of E738 against recombinant JAKs and SFKs in vitro are in the ranges of 0.7-74.1 nM and 10.7-263.9 nM, respectively. We observed that phosphorylation of both Jak2 and Src was substantially inhibited in the submicromolar range by E738 in cultured human pancreatic tumor cells, followed by blockade of downstream Stat3 activation. E738 down-regulated expression of the Stat3 target proteins Mcl-1 and survivin, associated with induction of apoptosis. Computational models and molecular dynamics simulations of E738/Tyk2 or E738/Src in silico suggest that E738 inhibits both tyrosine kinase 2 (Tyk2) and Src as an ATP-competitive ligand. Moreover, the planar E738 molecule demonstrates a strong binding affinity in the compact ATP-binding site of Tyk2. In sum, E738 is the first dual inhibitor of JAKs and SFKs, followed by inhibition of Stat3 signaling. Thus, according to in vitro experiments, E738 is a promising new therapeutic agent for human pancreatic cancer treatment by blocking both oncogenic pathways simultaneously.

  8. HCMV Activates the IL-6-JAK-STAT3 Axis in HepG2 Cells and Primary Human Hepatocytes

    PubMed Central

    Kumar, Amit; Tripathy, Manoj K.; Herbein, Georges

    2013-01-01

    Objectives There has been increased interest in the possible role of human cytomegalovirus (HCMV) in carcinogenesis during the last decade. HCMV seroprevalence was enhanced in patients with hepatocellular carcinoma (HCC) but a possible relationship between HCC and HCMV infection remained to be assessed. The aim of this work was to investigate the pro-tumor influence of HCMV on primary human hepatocytes (PHH) and HepG2 cells. Methods Following infection of PHH and HepG2 cells by two different strains of HCMV, we measured the production of IL-6 in culture supernatants by ELISA and the protein levels of STAT3, pSTAT3, JAK, cyclin D1, survivin, p53, p21, and Mdm2 by western Blotting in infected and uninfected cells. Cell proliferation and transformation were investigated using Ki67Ag expression measurement and soft-agar colony formation assay respectively. Results Infection of HepG2 cells and PHH by HCMV resulted in the production of IL-6 and the subsequent activation of the IL-6R-JAK-STAT3 pathway. HCMV increased the expression of cyclin D1 and survivin. Cell proliferation was enhanced in HepG2 and PHH infected with HCMV, despite a paradoxical overexpression of p53 and p21. More importantly, we observed the formation of colonies in soft agar seeded with PHH infected with HCMV and when we challenged the HepG2 cultures to form tumorspheres, we found that the HCMV-infected cultures formed 2.5-fold more tumorspheres than uninfected cultures. Conclusion HCMV activated the IL-6-JAK-STAT3 pathway in PHH and HepG2 cells, favored cellular proliferation, induced PHH transformation and enhanced HepG2 tumorsphere formation. Our observations raise the possibility that HCMV infection might be involved in the genesis of hepatocellular carcinoma. PMID:23555719

  9. Myocyte-Derived Hsp90 Modulates Collagen Upregulation via Biphasic Activation of STAT-3 in Fibroblasts during Cardiac Hypertrophy.

    PubMed

    Datta, Ritwik; Bansal, Trisha; Rana, Santanu; Datta, Kaberi; Datta Chaudhuri, Ratul; Chawla-Sarkar, Mamta; Sarkar, Sagartirtha

    2017-03-15

    Signal transducer and activator of transcription 3 (STAT-3)-mediated signaling in relation to upregulated collagen expression in fibroblasts during cardiac hypertrophy is well defined. Our recent findings have identified heat shock protein 90 (Hsp90) to be a critical modulator of fibrotic signaling in cardiac fibroblasts in this disease milieu. The present study was therefore intended to analyze the role of Hsp90 in the STAT-3-mediated collagen upregulation process. Our data revealed a significant difference between in vivo and in vitro results, pointing to a possible involvement of myocyte-fibroblast cross talk in this process. Cardiomyocyte-targeted knockdown of Hsp90 in rats (Rattus norvegicus) in which the renal artery was ligated showed downregulated collagen synthesis. Furthermore, the results obtained with cardiac fibroblasts conditioned with Hsp90-inhibited hypertrophied myocyte supernatant pointed toward cardiomyocytes' role in the regulation of collagen expression in fibroblasts during hypertrophy. Our study also revealed a novel signaling mechanism where myocyte-derived Hsp90 orchestrates not only p65-mediated interleukin-6 (IL-6) synthesis but also its release in exosomal vesicles. Such myocyte-derived exosomes and myocyte-secreted IL-6 are responsible in unison for the biphasic activation of STAT-3 signaling in cardiac fibroblasts that culminates in excess collagen synthesis, leading to severely compromised cardiac function during cardiac hypertrophy.

  10. Crosstalk between signaling pathways of adrenoreceptors and signal transducers and activators of transcription 3 (STAT3) in heart.

    PubMed

    Gong, Kai-zheng; Zhang, Hui; Du, Jian-hai; Zhang, You-yi

    2007-02-01

    Recently, there have been important advancements in our understanding of the signaling mechanisms of adrenoreceptors (AR) and signal transducers and activators of transcription 3 (STAT3). While their crucial roles in the pathological processes of the heart are well established, accumulating evidence suggests there is a complex pattern of crosstalk between these 2 signaling pathways. Moreover, the potential for crosstalk occurs at multiple levels in each signaling cascade and involves receptor transactivation, G proteins, small GTPases, cyclic adenosine 3',5'-monophosphate/protein kinase A, protein kinase C, scaffold/adaptor proteins, protein tyrosine kinases, and mitogen-activated protein kinases. In addition, post-translational modification (eg acetylation) of STAT3 may provide a link between STAT3 and AR signaling. In particular, crosstalk between these 2 systems in the heart would appear to be dependent upon the species/tissue studied, developmental stage, and eliciting stimulus. This at least partly accounts for the epigenetic effects on biological function that is mediated by the 2 signaling pathways. Elucidation of these mechanisms will provide new targets in the development of novel clinical strategies for heart disorders.

  11. gp130-mediated Stat3 activation in enterocytes regulates cell survival and cell-cycle progression during colitis-associated tumorigenesis.

    PubMed

    Bollrath, Julia; Phesse, Toby J; von Burstin, Vivian A; Putoczki, Tracy; Bennecke, Moritz; Bateman, Trudie; Nebelsiek, Tim; Lundgren-May, Therese; Canli, Ozge; Schwitalla, Sarah; Matthews, Vance; Schmid, Roland M; Kirchner, Thomas; Arkan, Melek C; Ernst, Matthias; Greten, Florian R

    2009-02-03

    Although gastrointestinal cancers are frequently associated with chronic inflammation, the underlying molecular links have not been comprehensively deciphered. Using loss- and gain-of-function mice in a colitis-associated cancer model, we establish here a link comprising the gp130/Stat3 transcription factor signaling axis. Mutagen-induced tumor growth and multiplicity are reduced following intestinal epithelial cell (IEC)-specific Stat3 ablation, while its hyperactivation promotes tumor incidence and growth. Conversely, IEC-specific Stat3 deficiency enhances susceptibility to chemically induced epithelial damage and subsequent mucosal inflammation, while excessive Stat3 activation confers resistance to colitis. Stat3 has the capacity to mediate IL-6- and IL-11-dependent IEC survival and to promote proliferation through G1 and G2/M cell-cycle progression as the common tumor cell-autonomous mechanism that bridges chronic inflammation to tumor promotion.

  12. NANOG regulates epithelial-mesenchymal transition and chemoresistance through activation of the STAT3 pathway in epithelial ovarian cancer.

    PubMed

    Liu, Suqing; Sun, Jing; Cai, Bin; Xi, Xiaowei; Yang, Liu; Zhang, Zhenbo; Feng, Youji; Sun, Yunyan

    2016-07-01

    NANOG is a key transcription factor that is overexpressed and plays an important role in various cancers. Its overexpression is associated with highly tumorigenic, drug-resistant, and poor prognosis. However, the underlying mechanism of action of NANOG in ovarian cancer remains unclear. Epithelial-mesenchymal transition (EMT), which is a critical process in cancer invasion and metastasis, is also associated with drug resistance. We determined whether NANOG is associated with EMT and chemoresistance in epithelial ovarian cancer cells. NANOG expression was increased in epithelial ovarian cancer cells (HEY and SKOV3) compared with normal epithelial ovarian cells (Moody). Low expression of NANOG increased the expression of E-cadherin and decreased the expression of vimentin, β-catenin, and Snail. Furthermore, the cell migration and invasion abilities were decreased. The multidrug resistance genes MDR-1 and GST-π were also downregulated when NANOG was lowly expressed. The cells that were transfected with the si-NANOG plasmid were more sensitive to cisplatin compared with the cells that were transfected with empty vector. The data demonstrated that Stat3 was correlated with NANOG-mediated EMT and drug resistance. The silencing of Stat3 expression abrogated NANOG-mediated EMT changes and increased the sensitivity of the cells to chemotherapy. These results suggest that NANOG mediates EMT and drug resistance through activation of the Stat3 pathway in epithelial ovarian cancer.

  13. Upregulation of Vesicular Glutamate Transporter 2 and STAT3 Activation in the Spinal Cord of Mice Receiving 3,3'-Iminodipropionitrile.

    PubMed

    Ohgomori, Tomohiro; Yamasaki, Ryo; Kira, Jun-Ichi; Jinno, Shozo

    2017-09-30

    Chronic administration of 3,3'-iminodipropionitrile (IDPN) causes axonal impairment. Although controversy still remains, it has been suggested that IDPN intoxication mimics the axonopathy of amyotrophic lateral sclerosis (ALS). Interestingly, recent studies including our own showed that signal transducer and activator of transcription 3 (STAT3) in spinal α-motoneurons was activated in both IDPN-treated mice and SOD1 (G93A) mice, a genetic model of familial ALS. Because activation of STAT3 occurs in response to various stimuli, such as axonal injury, ischemia, and excessive glutamate, here we focused on a potential link between phosphorylated STAT3 (pSTAT3, an active form) and vesicular glutamate transporter 2 (VGluT2, a regulator of glutamate storage and release) in IDPN-treated mice and SOD1 (G93A) mice. Impairment of axonal transport was confirmed by western blot analysis: the expression levels of phosphorylated neurofilament H were elevated in both models. As shown in SOD1 (G93A) mice, the expression frequencies of VGluT2 in synaptophysin-positive (SYP)(+) presynaptic terminals around spinal α-motoneurons were significantly higher in IDPN-treated mice than in vehicle controls. The coverages of spinal α-motoneurons by VGluT2(+) presynaptic terminals were more elevated around pSTAT3(+) cells than around pSTAT3(-) cells in IDPN-treated mice and SOD1 (G93A) mice. Considering that excessive glutamate is shown to be involved in axonal impairment and STAT3 activation, the present results suggest that IDPN-induced upregulation of VGluT2 may result in an increase in glutamate, which might cause axonopathy and induction of pSTAT3. The link between upregulation of VGluT2 and activation of STAT3 via glutamate may represent a common pathological feature of IDPN-treated mice and SOD1 (G93A) mice.

  14. Phosphorylation status determines the opposing functions of Smad2/Smad3 as STAT3 cofactors in TH17 differentiation.

    PubMed

    Yoon, Jeong-Hwan; Sudo, Katsuko; Kuroda, Masahiko; Kato, Mitsuyasu; Lee, In-Kyu; Han, Jin Soo; Nakae, Susumu; Imamura, Takeshi; Kim, Juryun; Ju, Ji Hyeon; Kim, Dae-Kee; Matsuzaki, Koichi; Weinstein, Michael; Matsumoto, Isao; Sumida, Takayuki; Mamura, Mizuko

    2015-07-21

    Transforming growth factor-β (TGF-β) and interleukin-6 (IL-6) are the pivotal cytokines to induce IL-17-producing CD4(+) T helper cells (TH17); yet their signalling network remains largely unknown. Here we show that the highly homologous TGF-β receptor-regulated Smads (R-Smads): Smad2 and Smad3 oppositely modify STAT3-induced transcription of IL-17A and retinoic acid receptor-related orphan nuclear receptor, RORγt encoded by Rorc, by acting as a co-activator and co-repressor of STAT3, respectively. Smad2 linker phosphorylated by extracellular signal-regulated kinase (ERK) at the serine 255 residue interacts with STAT3 and p300 to transactivate, whereas carboxy-terminal unphosphorylated Smad3 interacts with STAT3 and protein inhibitor of activated STAT3 (PIAS3) to repress the Rorc and Il17a genes. Our work uncovers carboxy-terminal phosphorylation-independent noncanonical R-Smad-STAT3 signalling network in TH17 differentiation.

  15. A Single Amino Acid Substitution in the v-Eyk Intracellular Domain Results in Activation of Stat3 and Enhances Cellular Transformation

    PubMed Central

    Besser, Daniel; Bromberg, Jacqueline F.; Darnell, James E.; Hanafusa, Hidesaburo

    1999-01-01

    The receptor tyrosine kinase Eyk, a member of the Axl/Tyro3 subfamily, activates the STAT pathway and transforms cells when constitutively activated. Here, we compared the potentials of the intracellular domains of Eyk molecules derived from c-Eyk and v-Eyk to transform rat 3Y1 fibroblasts. The v-Eyk molecule induced higher numbers of transformants in soft agar and stronger activation of Stat3; levels of Stat1 activation by the two Eyk molecules were similar. A mutation in the sequence Y933VPL, present in c-Eyk, to the v-Eyk sequence Y933VPQ led to increased activation of Stat3 and increased transformation efficiency. However, altering another sequence, Y862VNT, present in both Eyk molecules to F862VNT markedly decreased transformation without impairing Stat3 activation. These results indicate that activation of Stat3 enhances transformation efficiency and cooperates with another pathway to induce transformation. PMID:9891073

  16. Early Secreted Antigenic Target of 6-kDa of Mycobacterium tuberculosis Stimulates IL-6 Production by Macrophages through Activation of STAT3

    PubMed Central

    Jung, Bock-Gie; Wang, Xisheng; Yi, Na; Ma, Justin; Turner, Joanne; Samten, Buka

    2017-01-01

    As early secreted antigenic target of 6 kDa (ESAT-6) of Mycobacterium tuberculosis (Mtb) is an essential virulence factor and macrophages are critical for tuberculosis infection and immunity, we studied ESAT-6 stimulated IL-6 production by macrophages. ESAT-6 stimulated significantly higher IL-6 secretion by murine bone marrow derived macrophages (BMDM) compared to culture filtrate protein 10 kDa (CFP10) and antigen 85A. Polymyxin B, an LPS blocker, did not affect ESAT-6 stimulated macrophage IL-6 production. ESAT-6 but not Pam3CSK4 induced IL-6 by TLR2 knockout BMDM. ESAT-6 induced phosphorylation and DNA binding of STAT3 and this was blocked by STAT3 inhibitors but not by rapamycin. STAT3 inhibitors suppressed ESAT-6-induced IL-6 transcription and secretion without affecting cell viability. This was confirmed by silencing STAT3 in macrophages. Blocking neither IL-6Rα/IL-6 nor IL-10 affected ESAT-6-induced STAT3 activation and IL-6 production. Infection of BMDM and human macrophages with Mtb with esat-6 deletion induced diminished STAT3 activation and reduced IL-6 production compared to wild type and esat-6 complemented Mtb strains. Administration of ESAT-6 but not CFP10 induced STAT3 phosphorylation and IL-6 expression in the mouse lungs, consistent with expression of ESAT-6, IL-6 and phosphorylated-STAT3 in Mtb-infected mouse lungs. We conclude that ESAT-6 stimulates macrophage IL-6 production through STAT3 activation. PMID:28106119

  17. EGFR and KRAS mutations do not enrich for the activation of IL-6, JAK1 or phosphorylated STAT3 in resected lung adenocarcinoma.

    PubMed

    Clay, Timothy D; Russell, Prudence A; Do, Hongdo; Sundararajan, Vijaya; Conron, Matthew; Wright, Gavin M; Solomon, Benjamin; Dobrovic, Alexander; McLachlan, Sue-Anne; Moore, Melissa M

    2017-09-06

    Resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) against EGFR mutant lung adenocarcinoma develops after a median of nine to thirteen months. Upregulation of the interleukin-6 (IL-6)/Janus kinase (JAK)/signal transducers and activators of transcription (STAT) pathway may be a potential source of resistance to EGFR TKIs. We undertook a detailed assessment of the IL-6/JAK1/phosphorylated STAT3 (pSTAT3) pathway in resected lung adenocarcinoma specimens, with special interest in whether the presence of an EGFR mutation enriched for pSTAT3 positivity. Tumours from 143 patients with resected lung adenocarcinoma were assessed. EGFR and KRAS mutation status were scanned for with high-resolution melting and confirmed by polymerase chain reaction. Immunohistochemisty (IHC) was performed for IL-6, gp130, JAK1 and pSTAT3. Two methods for assigning IHC positivity were assessed (the presence of any positivity, and the presence of positivity at an H score >40). We found statistically significant associations between IL-6, JAK1 and pSTAT3 measured by IHC, consistent with the activation of the pathway in clinical specimens. No relationship was demonstrated between members of this pathway and oncogenic mutations in EGFR or KRAS. However, a proportion of tumours with EGFR mutations showed staining for IL-6, JAK1 and pSTAT3. No correlations with clinicopathologic features or survival outcomes were found for IL-6, JAK1 or pSTAT3 staining. The presence of EGFR or KRAS mutations did not enrich for the activation of IL-6, JAK1 or pSTAT3. pSTAT3 may still play a role in resistance to EGFR TKIs in clinical practice.

  18. Sphingosine-1-phosphate enhances satellite cell activation in dystrophic muscles through a S1PR2/STAT3 signaling pathway.

    PubMed

    Loh, Kenneth C; Leong, Weng-In; Carlson, Morgan E; Oskouian, Babak; Kumar, Ashok; Fyrst, Henrik; Zhang, Meng; Proia, Richard L; Hoffman, Eric P; Saba, Julie D

    2012-01-01

    Sphingosine-1-phosphate (S1P) activates a widely expressed family of G protein-coupled receptors, serves as a muscle trophic factor and activates muscle stem cells called satellite cells (SCs) through unknown mechanisms. Here we show that muscle injury induces dynamic changes in S1P signaling and metabolism in vivo. These changes include early and profound induction of the gene encoding the S1P biosynthetic enzyme SphK1, followed by induction of the catabolic enzyme sphingosine phosphate lyase (SPL) 3 days later. These changes correlate with a transient increase in circulating S1P levels after muscle injury. We show a specific requirement for SphK1 to support efficient muscle regeneration and SC proliferation and differentiation. Mdx mice, which serve as a model for muscular dystrophy (MD), were found to be S1P-deficient and exhibited muscle SPL upregulation, suggesting that S1P catabolism is enhanced in dystrophic muscle. Pharmacological SPL inhibition increased muscle S1P levels, improved mdx muscle regeneration and enhanced SC proliferation via S1P receptor 2 (S1PR2)-dependent inhibition of Rac1, thereby activating Signal Transducer and Activator of Transcription 3 (STAT3), a central player in inflammatory signaling. STAT3 activation resulted in p21 and p27 downregulation in a S1PR2-dependent fashion in myoblasts. Our findings suggest that S1P promotes SC progression through the cell cycle by repression of cell cycle inhibitors via S1PR2/STAT3-dependent signaling and that SPL inhibition may provide a therapeutic strategy for MD.

  19. Differential activation of neuronal and glial STAT3 in the spinal cord of the SOD1(G93A) mouse model of amyotrophic lateral sclerosis.

    PubMed

    Ohgomori, Tomohiro; Yamasaki, Ryo; Takeuchi, Hideyuki; Kadomatsu, Kenji; Kira, Jun-Ichi; Jinno, Shozo

    2017-08-01

    Signal transducer and activator of transcription (STAT) proteins are activated by phosphorylation in the spinal cord of patients suffering from amyotrophic lateral sclerosis (ALS). The major scope of our study is a comprehensive histological characterization of the mechanisms underlying neuronal and glial STAT3 activation in the pathogenesis of ALS using SOD1(G93A) mice. We calculated the fold changes (FCs, ratios vs. appropriate controls) of the numerical densities of the following phosphorylated STAT3-positive (pSTAT3)(+) cells - choline acetyltransferase (ChAT)(+) α-motoneurons, ionized calcium-binding adapter molecule 1 (Iba1)(+) microglia, and S100β(+) astrocytes in SOD1(G93A) mice. The FCs of pSTAT3(+) microglia and pSTAT3(+) astrocytes were increased from 9 to 15 weeks of age and then plateaued until 21 weeks. In contrast, the FCs of pSTAT3(+) α-motoneurons peaked at 9 weeks and then decreased until 21 weeks. The immunoreactivity for nonphosphorylated neurofilament protein (SMI-32), a marker of axonal impairment, was decreased in pSTAT3(+) α-motoneurons compared with pSTAT3(-) α-motoneurons at 9 weeks of age. We then compared the following pharmacological models - the chronic administration of 3,3'-iminodipropionitrile (IDPN), which models axonal impairment, and the acute administration of lipopolysaccharide (LPS), which is a model of neuroinflammation. The FCs of pSTAT3(+) α-motoneurons were increased in IDPN-treated mice, while those of pSTAT3(+) microglia were increased in LPS-treated mice. The FCs of pSTAT3(+) astrocytes were higher in SOD1(G93A) mice at 9 weeks compared with IDPN- and LPS-treated mice. Our results indicate that axonopathy and neuroinflammation may trigger the respective activation of neuronal and glial STAT3, which is observed during ALS pathogenesis. © 2017 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

  20. Gemcitabine enhances cell invasion via activating HAb18G/CD147-EGFR-pSTAT3 signaling

    PubMed Central

    Wu, Xiao-Qing; Wu, Bo; Xu, Liang; Jiang, Jian-Li; Li, Ling; Chen, Zhi-Nan

    2016-01-01

    Pancreatic cancer, one of the most lethal cancers, has very poor 5-year survival partly due to gemcitabine resistance. Recently, it was reported that chemotherapeutic agents may act as stressors to induce adaptive responses and to promote chemoresistance in cancer cells. During long-term drug treatment, the minority of cancer cells survive and acquire an epithelial-mesenchymal transition phenotype with increased chemo-resistance and metastasis. However, the short-term response of most cancer cells remains unclear. This study aimed to investigate the short-term response of pancreatic cancer cells to gemcitabine stress and to explore the corresponding mechanism. Our results showed that gemcitabine treatment for 24 hours enhanced pancreatic cancer cell invasion. In gemcitabine-treated cells, HAb18G/CD147 was up-regulated; and HAb18G/CD147 down-regulation or inhibition attenuated gemcitabine-enhanced invasion. Mechanistically, HAb18G/CD147 promoted gemcitabine-enhanced invasion by activating the EGFR (epidermal growth factor receptor)-STAT3 (signal transducer and activator of transcription 3) signaling pathway. Inhibition of EGFR-STAT3 signaling counteracted gemcitabine-enhanced invasion, and which relied on HAb18G/CD147 levels. In pancreatic cancer tissues, EGFR was highly expressed and positively correlated with HAb18G/CD147. These data indicate that pancreatic cancer cells enhance cell invasion via activating HAb18G/CD147-EGFR-pSTAT3 signaling. Our findings suggest that inhibiting HAb18G/CD147 is a potential strategy for overcoming drug stress-associated resistance in pancreatic cancer. PMID:27556697

  1. Gemcitabine enhances cell invasion via activating HAb18G/CD147-EGFR-pSTAT3 signaling.

    PubMed

    Xu, Bao-Qing; Fu, Zhi-Guang; Meng, Yao; Wu, Xiao-Qing; Wu, Bo; Xu, Liang; Jiang, Jian-Li; Li, Ling; Chen, Zhi-Nan

    2016-09-20

    Pancreatic cancer, one of the most lethal cancers, has very poor 5-year survival partly due to gemcitabine resistance. Recently, it was reported that chemotherapeutic agents may act as stressors to induce adaptive responses and to promote chemoresistance in cancer cells. During long-term drug treatment, the minority of cancer cells survive and acquire an epithelial-mesenchymal transition phenotype with increased chemo-resistance and metastasis. However, the short-term response of most cancer cells remains unclear. This study aimed to investigate the short-term response of pancreatic cancer cells to gemcitabine stress and to explore the corresponding mechanism. Our results showed that gemcitabine treatment for 24 hours enhanced pancreatic cancer cell invasion. In gemcitabine-treated cells, HAb18G/CD147 was up-regulated; and HAb18G/CD147 down-regulation or inhibition attenuated gemcitabine-enhanced invasion. Mechanistically, HAb18G/CD147 promoted gemcitabine-enhanced invasion by activating the EGFR (epidermal growth factor receptor)-STAT3 (signal transducer and activator of transcription 3) signaling pathway. Inhibition of EGFR-STAT3 signaling counteracted gemcitabine-enhanced invasion, and which relied on HAb18G/CD147 levels. In pancreatic cancer tissues, EGFR was highly expressed and positively correlated with HAb18G/CD147. These data indicate that pancreatic cancer cells enhance cell invasion via activating HAb18G/CD147-EGFR-pSTAT3 signaling. Our findings suggest that inhibiting HAb18G/CD147 is a potential strategy for overcoming drug stress-associated resistance in pancreatic cancer.

  2. Mitochondrial-targeted Signal Transducer and Activator of Transcription 3 (STAT3) Protects against Ischemia-induced Changes in the Electron Transport Chain and the Generation of Reactive Oxygen Species*

    PubMed Central

    Szczepanek, Karol; Chen, Qun; Derecka, Marta; Salloum, Fadi N.; Zhang, Qifang; Szelag, Magdalena; Cichy, Joanna; Kukreja, Rakesh C.; Dulak, Jozef; Lesnefsky, Edward J.; Larner, Andrew C.

    2011-01-01

    Expression of the STAT3 transcription factor in the heart is cardioprotective and decreases the levels of reactive oxygen species. Recent studies indicate that a pool of STAT3 resides in the mitochondria where it is necessary for the maximal activity of complexes I and II of the electron transport chain. However, it has not been explored whether mitochondrial STAT3 modulates cardiac function under conditions of stress. Transgenic mice with cardiomyocyte-specific overexpression of mitochondria-targeted STAT3 with a mutation in the DNA-binding domain (MLS-STAT3E) were generated. We evaluated the role of mitochondrial STAT3 in the preservation of mitochondrial function during ischemia. Under conditions of ischemia heart mitochondria expressing MLS-STAT3E exhibited modest decreases in basal activities of complexes I and II of the electron transport chain. In contrast to WT hearts, complex I-dependent respiratory rates were protected against ischemic damage in MLS-STAT3E hearts. MLS-STAT3E prevented the release of cytochrome c into the cytosol during ischemia. In contrast to WT mitochondria, ischemia did not augment reactive oxygen species production in MLS-STAT3E mitochondria likely due to an MLS-STAT3E-mediated partial blockade of electron transport through complex I. Given the caveat of STAT3 overexpression, these results suggest a novel protective mechanism mediated by mitochondrial STAT3 that is independent of its canonical activity as a nuclear transcription factor. PMID:21715323

  3. Glucagon-like peptide-1 (GLP-1) induces M2 polarization of human macrophages via STAT3 activation.

    PubMed

    Shiraishi, Daisuke; Fujiwara, Yukio; Komohara, Yoshihiro; Mizuta, Hiroshi; Takeya, Motohiro

    2012-08-24

    It is known that glucagon-like peptide-1 (GLP-1) is a hormone secreted postprandially from the L-cells of the small intestine and regulates glucose homeostasis. GLP-1 is now used for the treatment of diabetes because of its beneficial role against insulin resistance. The GLP-1 receptor (GLP-1R) is expressed on many cell types, including macrophages, and GLP-1 suppresses the development of atherosclerosis by inhibiting macrophage function. However, there have so far been few studies that have investigated the significance of GLP-1/GLP-1R signaling in macrophage activation. In the present study, we examined the effect of GLP-1 and exenatide, a GLP-1R agonist, on human monocyte-derived macrophage (HMDM) activation. We found that GLP-1 induced signal transducer and activator of transcription 3 (STAT3) activation. Silencing of GLP-1R suppressed the GLP-1-induced STAT3 activation. In addition, alternatively activated (M2) macrophage-related molecules, such as IL-10, CD163, and CD204 in HMDM, were significantly upregulated by GLP-1. Furthermore, the co-culture of 3T3-L1 adipocytes with GLP-1-treated RAW 264.7 macrophages increased the secretion of adiponectin compared to co-culture of the 3T3-L1 adipocytes with untreated RAW 264.7 macrophages. Our results demonstrate that GLP-1 induces macrophage polarization toward the M2 phenotype, which may contribute to the protective effects of GLP-1 against diabetes and cardiovascular diseases.

  4. IL-22 suppresses the infection of porcine enteric coronaviruses and rotavirus by activating STAT3 signal pathway.

    PubMed

    Xue, Mei; Zhao, Jing; Ying, Lan; Fu, Fang; Li, Lin; Ma, Yanlong; Shi, Hongyan; Zhang, Jiaoer; Feng, Li; Liu, Pinghuang

    2017-06-01

    Interleukin-22 (IL-22), a member of the IL-10 superfamily, plays essential roles in fighting against mucosal microbial infection and maintaining mucosal barrier integrity within the intestine. However, little knowledge exists on the ability of porcine IL-22 (pIL-22) to fight against viral infection in the gut. In this study, we found that recombinant mature pIL-22 (mpIL-22) inhibited the infection of multiple diarrhea viruses, including alpha coronavirus, porcine epidemic diarrhea virus (PEDV), transmissible gastroenteritis virus (TGEV), and porcine rotavirus (PoRV), in the intestinal porcine epithelial cell line J2 (IPEC-J2) cells. mpIL-22 up-regulated the expression of the antimicrobial peptide beta-defensin (BD-2), cytokine IL-18 and IFN-λ. Furthermore, we found that mpIL-22 induced phosphorylation of STAT3 on Ser727 and Tyr705 in IPEC-J2 cells. Inhibition of STAT3 phosphorylation by S3I-201 abrogated the antiviral ability of mpIL-22 and the mpIL-22-induced expression of BD-2, IL-18, and IFN-λ. Together, mpIL-22 inhibited the infection of PoRV and enteric coronaviruses, and up-regulated the expression of antimicrobial genes in IPEC-J2, which were mediated by the activation of the STAT3 signal pathway. The significant antiviral activity of IL-22 to curtail multiple enteric diarrhea viruses in vitro suggests that pIL-22 could be a novel therapeutic against devastating viral diarrhea in piglets. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. Andrographolide causes apoptosis via inactivation of STAT3 and Akt and potentiates antitumor activity of gemcitabine in pancreatic cancer.

    PubMed

    Bao, Guo-Qing; Shen, Bai-Yong; Pan, Chun-Peng; Zhang, Ya-Jing; Shi, Min-Min; Peng, Cheng-Hong

    2013-09-12

    Gemcitabine is a first-line drug utilised in the chemotherapy of pancreatic cancer; however, this drug induces chemo-resistance and toxicity to normal tissue during treatment. Here, we firstly report that andrographolide (ANDRO) alone not only has anti-pancreatic cancer activity, but it also potentiates the anti-tumour activity of gemcitabine. Treatment with ANDRO alone inhibits proliferation of the pancreatic cancer cell lines in a dose- and time-dependent manner in vitro. Interestingly, ANDRO induces cell cycle arrest and apoptosis of pancreatic cancer cells by inhibiting STAT3 and Akt activation, upregulating the expression of p21(WAF1) and Bax, and downregulating the expression of cyclinD1, cyclinE, survivin, X-IAP and Bcl-2. Additionally, ANDRO combined with gemcitabine significantly induce stronger cell cycle arrest and more obvious apoptosis than each single treatment. The mechanistic study demonstrates that this synergistic effect is also dependent on the inhibition of STAT3 and Akt activations which subsequently regulates the pathways involved in the apoptosis and cell cycle arrest. Furthermore, both ANDRO alone and the combination treatments exhibit efficacious anti-tumour activity in vivo. Overall, our results provide solid evidence supporting that ANDRO alone or its combination with gemcitabine is a potential chemotherapeutic approach for treating human pancreatic cancer in clinical practice.

  6. Induction of metastatic potential by TrkB via activation of IL6/JAK2/STAT3 and PI3K/AKT signaling in breast cancer

    PubMed Central

    Kim, Min Soo; Lee, Won Sung; Jeong, Joon; Kim, Seong-Jin; Jin, Wook

    2015-01-01

    In metastatic breast cancers, the acquisition of metastatic ability, which leads to clinically incurable disease and poor survival, has been associated with acquisition of epithelial-mesenchymal transition (EMT) program and self-renewing trait (CSCs) via activation of PI3K/AKT and IL6/JAK2/STAT3 signaling pathways. We found that TrkB is a key regulator of PI3K/AKT and JAK/STAT signal pathway-mediated tumor metastasis and EMT program. Here, we demonstrated that TrkB activates AKT by directly binding to c-Src, leading to increased proliferation. Also, TrkB increases Twist-1 and Twist-2 expression through activation of JAK2/STAT3 by inducing c-Src-JAK2 complex formation. Furthermore, TrkB in the absence of c-Src binds directly to JAK2 and inhibits SOCS3-mediated JAK2 degradation, resulting in increased total JAK2 and STAT3 levels, which subsequently leads to JAK2/STAT3 activation and Twist-1 upregulation. Additionally, activation of the JAK2/STAT3 pathway via induction of IL-6 secretion by TrkB enables induction of activation of the EMT program via induction of STAT3 nuclear translocation. These observations suggest that TrkB is a promising target for future intervention strategies to prevent tumor metastasis, EMT program and self-renewing trait in breast cancer. PMID:26515594

  7. Physalin A exerts anti-tumor activity in non-small cell lung cancer cell lines by suppressing JAK/STAT3 signaling

    PubMed Central

    Loo, Jacky F.C.; Xia, Dajin; Gao, Sizhi P.; Ma, Zhongjun; Chen, Zhe

    2016-01-01

    The signal transducers and activators of transcription 3 (STAT3) signaling pathway plays critical roles in the pathogenesis and progression of various human cancers, including non-small cell lung cancer (NSCLC). In this study, we aimed to evaluate the therapeutic potential of physalin A, a bioactive withanolide derived from Physalis alkekengi var. francheti used in traditional Chinese medicine, was evaluated in human NSCLC cells. Its and determined whether it effect oninhibited both constitutive and induced STAT3 activity, through repressing the phosphorylation levels of JAK2 and JAK3, resulting in anti-proliferation and pro-apoptotic effects on NSCLC cells was also determined, and. theThe antitumor effects of physalin A were also validated usingin an in vivo mouse xenograft models of NSCLC cells. Physalin A had anti-proliferative and pro-apoptotic effects in NSCLC cells with constitutively activated STAT3; it also suppressed both constitutive and induced STAT3 activity by modulating the phosphorylation of JAK2 and JAK3. Furthermore, physalin A abrogated the nuclear translocation and transcriptional activity of STAT3, thereby decreasing the expression levels of STAT3, its target genes, such as Bcl-2 and XIAP. Knockdown of STAT3 expression by small interfering RNA (siRNA) significantly enhanced the pro-apoptotic effects of physalin A in NSCLC cells. Moreover, physalin A significantly suppressed tumor xenograft growth. Thus, as an inhibitor of JAK2/3-STAT3 signaling, physalin A, has potent anti-tumor activities, which may facilitate the development of a therapeutic strategy for treating NSCLC. PMID:26843613

  8. Noncanonical NF-κB Activation Mediates STAT3-stimulated IDO Upregulation in Myeloid-Derived Suppressor Cells in Breast Cancer

    PubMed Central

    Yu, Jinpu; Wang, Yue; Yan, Fang; Zhang, Peng; Li, Hui; Zhao, Hua; Yan, Cihui; Yan, Fan; Ren, Xiubao

    2016-01-01

    Immunotherapy exerts anticancer effects by activating competent immune effectors and inhibiting immunosuppressive cells, such as myeloid-derived suppressor cells (MDSCs). However, the mechanism underlying MDSCs-mediated immunosuppression in breast cancer is unclear. We have identified a poorly differentiated subset of MDSCs in breast cancer, which suppresses T-cell functions through STAT3-dependent IDO upregulation. Here, we aimed to investigate the mechanisms by which IDO expression is aberrant in MDSCs. We found increased STAT3 phosphorylation and NIK expression were correlated with upregulated IDO expression in MDSCs in human breast cancer. In mouse 4T1 mammary cancer model, blocking STAT3 signal significantly inhibited the activation of NF-κB and IDO expression in MDSCs, resulting in decrease of tumor growth and metastasis. We also induced MDSCs by co-culturing human CD33+ myeloid progenitors with MDA-MB-231 breast cancer cells. In these induced MDSCs, increased STAT3 activation was correlated with the activation of the noncanonical NF-κB pathway, including increased NIK protein level, phosphorylation of IKKα and p100 in cytoplasm, and RelB-p52 nuclear translocation. Blocking STAT3 activation significantly inhibited the accumulation of NIK and IDO expression in MDSCs. Knock-down of NIK by siRNA transfection in MDSCs suppressed IDO expression, but not STAT3 activation. Transcription factor assay and ChIP assay showed that RelB-p52 dimers directly bound to the IDO promoter, leading to IDO expression in MDSCs. These results suggest a STAT3-NF-κB-IDO pathway in breast cancer-derived MDSCs. Furthermore, IL-6 was found to stimulate STAT3-dependent NF-κB-mediated IDO upregulation in MDSCs. This study provides insights into understanding mechanisms through which MDSCs play an immunosuppressive role in breast cancer. PMID:25063873

  9. High Glucose-Mediated STAT3 Activation in Endometrial Cancer Is Inhibited by Metformin: Therapeutic Implications for Endometrial Cancer.

    PubMed

    Wallbillich, John J; Josyula, Srirama; Saini, Uksha; Zingarelli, Roman A; Dorayappan, Kalpana Deepa Priya; Riley, Maria K; Wanner, Ross A; Cohn, David E; Selvendiran, Karuppaiyah

    2017-01-01

    STAT3 is over-expressed in endometrial cancer, and diabetes is a risk factor for the development of type 1 endometrial cancer. We therefore investigated whether glucose concentrations influence STAT3 expression in type 1 endometrial cancer, and whether such STAT3 expression might be inhibited by metformin. In Ishikawa (grade 1) endometrial cancer cells subjected to media with low, normal, or high concentrations of glucose, expression of STAT3 and its target proteins was evaluated by real-time quantitative PCR (qPCR). Ishikawa cells were treated with metformin and assessed with cell proliferation, survival, migration, and ubiquitin assays, as well as Western blot and qPCR. Expression of apoptosis proteins was evaluated with Western blot in Ishikawa cells transfected with a STAT3 overexpression plasmid and treated with metformin. A xenograft tumor model was used for studying the in vivo efficacy of metformin. Expression of STAT3 and its target proteins was increased in Ishikawa cells cultured in high glucose media. In vitro, metformin inhibited cell proliferation, survival and migration but induced apoptosis. Metformin reduced expression levels of pSTAT3 ser727, total STAT3, and its associated cell survival and anti-apoptotic proteins. Additionally, metformin treatment was associated with increased degradation of pSTAT3 ser727. No change in apoptotic protein expression was noticed with STAT3 overexpression in Ishikawa cells. In vivo, metformin treatment led to a decrease in tumor weight as well as reductions of STAT3, pSTAT3 ser727, its target proteins. These results suggest that STAT3 expression in type 1 endometrial cancer is stimulated by a high glucose environment and inhibited by metformin.

  10. High Glucose-Mediated STAT3 Activation in Endometrial Cancer Is Inhibited by Metformin: Therapeutic Implications for Endometrial Cancer

    PubMed Central

    Wallbillich, John J.; Josyula, Srirama; Saini, Uksha; Zingarelli, Roman A.; Dorayappan, Kalpana Deepa Priya; Riley, Maria K.; Wanner, Ross A.; Cohn, David E.; Selvendiran, Karuppaiyah

    2017-01-01

    Objectives STAT3 is over-expressed in endometrial cancer, and diabetes is a risk factor for the development of type 1 endometrial cancer. We therefore investigated whether glucose concentrations influence STAT3 expression in type 1 endometrial cancer, and whether such STAT3 expression might be inhibited by metformin. Methods In Ishikawa (grade 1) endometrial cancer cells subjected to media with low, normal, or high concentrations of glucose, expression of STAT3 and its target proteins was evaluated by real-time quantitative PCR (qPCR). Ishikawa cells were treated with metformin and assessed with cell proliferation, survival, migration, and ubiquitin assays, as well as Western blot and qPCR. Expression of apoptosis proteins was evaluated with Western blot in Ishikawa cells transfected with a STAT3 overexpression plasmid and treated with metformin. A xenograft tumor model was used for studying the in vivo efficacy of metformin. Results Expression of STAT3 and its target proteins was increased in Ishikawa cells cultured in high glucose media. In vitro, metformin inhibited cell proliferation, survival and migration but induced apoptosis. Metformin reduced expression levels of pSTAT3 ser727, total STAT3, and its associated cell survival and anti-apoptotic proteins. Additionally, metformin treatment was associated with increased degradation of pSTAT3 ser727. No change in apoptotic protein expression was noticed with STAT3 overexpression in Ishikawa cells. In vivo, metformin treatment led to a decrease in tumor weight as well as reductions of STAT3, pSTAT3 ser727, its target proteins. Conclusions These results suggest that STAT3 expression in type 1 endometrial cancer is stimulated by a high glucose environment and inhibited by metformin. PMID:28114390

  11. Nucleophosmin/anaplastic lymphoma kinase (NPM/ALK) oncoprotein induces the T regulatory cell phenotype by activating STAT3

    PubMed Central

    Kasprzycka, Monika; Marzec, Michal; Liu, Xiaobin; Zhang, Qian; Wasik, Mariusz A.

    2006-01-01

    The mechanisms of malignant cell transformation mediated by the oncogenic, chimeric nucleophosmin/anaplastic lymphoma kinase (NPM/ALK) tyrosine kinase remain only partially understood. Here we report that the NPM/ALK-carrying T cell lymphoma (ALK+TCL) cells secrete IL-10 and TGF-β and express FoxP3, indicating their T regulatory (Treg) cell phenotype. The secreted IL-10 suppresses proliferation of normal immune, CD3/CD28-stimulated peripheral blood mononuclear cells and enhances viability of the ALK+TCL cells. The Treg phenotype of the affected cells is strictly dependent on NPM/ALK expression and function as demonstrated by transfection of the kinase into BaF3 cells and inhibition of its enzymatic activity and expression in ALK+TCL cells. NPM/ALK, in turn, induces the phenotype through activation of its key signal transmitter, signal transducer and activator of transcription 3 (STAT3). These findings identify a mechanism of NPM/ALK-mediated oncogenesis based on induction of the Treg phenotype of the transformed CD4+ T cells. These results also provide an additional rationale to therapeutically target the chimeric kinase and/or STAT3 in ALK+TCL. PMID:16766651

  12. Gp130-mediated STAT3 activation by S-propargyl-cysteine, an endogenous hydrogen sulfide initiator, prevents doxorubicin-induced cardiotoxicity

    PubMed Central

    Wu, J; Guo, W; Lin, S-Z; Wang, Z-J; Kan, J-T; Chen, S-Y; Zhu, Y-Z

    2016-01-01

    Doxorubicin (Dox) could trigger a large amount of apoptotic cells in the myocardium, which leads to dilated cardiomyopathy and heart failure. S-propargyl-cysteine (SPRC), a producing agent of endogenous hydrogen sulfide (H2S), possesses cardioprotective efficacy. However, the specific effect and mechanism of SPRC in Dox-induced cardiotoxicity remain elusive. Given gp130 with its main downstream signaling molecule, signal transducer and activator of transcription 3 (STAT3), is involved in cardiac myocyte survival and growth; the present study was performed to elucidate whether SPRC counteracts Dox-induced cardiotoxicity, and if so, whether the gp130/STAT3 pathway is involved in this cardioprotective activity. SPRC stimulated the activation of STAT3 via gp130-mediated transduction tunnel in vitro and in vivo. In Dox-stimulated cardiotoxicity, SPRC enhanced cell viability, restored expression of gp130/STAT3-regulated downstream genes, inhibited apoptosis and oxidative stress, and antagonized mitochondrial dysfunction and intracellular Ca2+ overload. Intriguingly, blockade of gp130/STAT3 signaling abrogated all these beneficial capacities of SPRC. Our findings present the first piece of evidence for the therapeutic properties of SPRC in alleviating Dox cardiotoxicity, which could be attributed to the activation of gp130-mediated STAT3 signaling. This will offer a novel molecular basis and therapeutic strategy of H2S donor for the treatment of heart failure. PMID:27537522

  13. Calnexin controls the STAT3-mediated transcriptional response to EGF.

    PubMed

    Lakkaraju, Asvin K K; van der Goot, F Gisou

    2013-08-08

    Calnexin is a well-characterized transmembrane chaperone involved in the folding of newly synthesized glycoproteins in the lumen of the endoplasmic reticulum (ER). Here, we reveal a previously unrecognized function of calnexin in regulating the transcriptional response downstream of epidermal growth factor receptor (EGF), the product of a well-known human oncogene. We find that cell stimulation with EGF leads to the caspase-8-dependent cleavage of the calnexin cytoplasmic domain, preferentially at ER-mitochondria interaction sites. The released fragment translocates into the nucleus, binds to PIAS3--a natural inhibitor of activated STAT3--and, thus, acts as an enhancer of the STAT3-mediated transcriptional response to EGF. Also, we reveal the unsuspected capacity of calnexin to sense ER stress and, in response, prevent the EGF-induced processing of its cytosolic domain. Thus, cells integrate the health status of the ER to determine the amplitude of their response to EGF. Copyright © 2013 Elsevier Inc. All rights reserved.

  14. HCS Campaign to Identify Selective Inhibitors of IL-6-Induced STAT3 Pathway Activation in Head and Neck Cancer Cell Lines

    PubMed Central

    Sen, Malabika; Hua, Yun; Camarco, Daniel P.; Shun, Tong Ying; Lazo, John S.; Wilson, Gabriela Mustata; Resnick, Lynn O.; LaPorte, Matthew G.; Wipf, Peter; Huryn, Donna M.; Grandis, Jennifer R.

    2015-01-01

    Abstract Signal transducer and activator of transcription factor 3 (STAT3) is hyperactivated in head and neck squamous cell carcinomas (HNSCC). Cumulative evidence indicates that IL-6 production by HNSCC cells and/or stromal cells in the tumor microenvironment activates STAT3 and contributes to tumor progression and drug resistance. A library of 94,491 compounds from the Molecular Library Screening Center Network (MLSCN) was screened for the ability to inhibit interleukin-6 (IL-6)-induced pSTAT3 activation. For contractual reasons, the primary high-content screening (HCS) campaign was conducted over several months in 3 distinct phases; 1,068 (1.1%) primary HCS actives remained after cytotoxic or fluorescent outliers were eliminated. One thousand one hundred eighty-seven compounds were cherry-picked for confirmation; actives identified in the primary HCS and compounds selected by a structural similarity search of the remaining MLSCN library using hits identified in phases I and II of the screen. Actives were confirmed in pSTAT3 IC50 assays, and an IFNγ-induced pSTAT1 activation assay was used to prioritize selective inhibitors of STAT3 activation that would not inhibit STAT1 tumor suppressor functions. Two hundred three concentration-dependent inhibitors of IL-6-induced pSTAT3 activation were identified and 89 of these also produced IC50s against IFN-γ-induced pSTAT1 activation. Forty-nine compounds met our hit criteria: they reproducibly inhibited IL-6-induced pSTAT3 activation by ≥70% at 20 μM; their pSTAT3 activation IC50s were ≤25 μM; they were ≥2-fold selective for pSTAT3 inhibition over pSTAT1 inhibition; a cross target query of PubChem indicated that they were not biologically promiscuous; and they were ≥90% pure. Twenty-six chemically tractable hits that passed filters for nuisance compounds and had acceptable drug-like and ADME-Tox properties by computational evaluation were purchased for characterization. The hit structures were distributed

  15. Activation of the RAGE/STAT3 Pathway in the Dorsal Root Ganglion Contributes to the Persistent Pain Hypersensitivity Induced by Lumbar Disc Herniation.

    PubMed

    Zhang, Xin-Sheng; Li, Xiao; Luo, Hai-Jie; Huang, Zhu-Xi; Liu, Cui-Cui; Wan, Qing; Xu, Shu-Wei; Wu, Shao-Ling; Ke, Song-Jian; Ma, Chao

    2017-07-01

    Clinically, chronic low back pain and sciatica associated with lumbar disc herniation (LDH) is a common musculoskeletal disorder. Due to the unawareness of detailed mechanisms, it is difficult to get an effective therapy. The aim of the present study was to identify the role of the RAGE/STAT3 pathway in the dorsal root ganglion (DRG) on the formation and development of persistent pain hypersensitivity induced by LDH. Controlled animal study. University laboratory. After LDH induced by implantation of autologous nucleus pulposus (NP, harvested from animal tail) on the left L5 nerve root was established, mechanical thresholds and electrophysiological tests were conducted at relevant time points during an observation period of 28 days. Protein levels and localization of RAGE and p-STAT3 were performed by using Western blotting and immunohistochemistry, respectively. LDH induced persistent pain hypersensitivity, increased excitability of DRG neurons, and upregulated the expression of RAGE and p-STAT3 in the DRG. Consecutive injection of both RAGE antagonist FPS-ZM1 (i.t.) and STAT3 activity inhibitor S3I-201 (i.t.) inhibited the enhanced excitability of DRG neurons and mechanical allodynia induced by NP implantation. Furthermore, local knockdown of STAT3 by intrathecal injection of AAV-Cre-GFP into STAT3flox/flox mice markedly alleviated NP implantation-induced mechanical allodynia in mice. Importantly, the expression of p-STAT3 was colocalized with that of RAGE in the DRG and inhibition of RAGE with FPS-ZM1 prevented NP implantation-induced STAT3 activation. More underlying mechanism(s) of the role of the RAGE/STAT3 pathway on the formation and development of persistent pain hypersensitivity induced by LDH will be needed to be explored in future research. These findings suggest activation of the RAGE/STAT3 pathway plays a critical role in persistent pain induced by LDH, and this pathway may represent novel therapeutic targets for the treatment of LDH

  16. NOX1 is responsible for cell death through STAT3 activation in hyperoxia and is associated with the pathogenesis of Acute Respiratory Distress Syndrome

    PubMed Central

    Carnesecchi, Stephanie; Dunand-Sauthier, Isabelle; Zanetti, Filippo; Singovski, Grigory; Deffert, Christine; Donati, Yves; Cagarelli, Thomas; Pache, Jean-Claude; Krause, Karl-Heinz; Reith, Walter; Barazzone-Argiroffo, Constance

    2014-01-01

    Reactive oxygen species (ROS) contribute to alveolar cell death in Acute Respiratory Distress Syndrome (ARDS) and we previously demonstrated that NOX1-derived ROS contributed to hyperoxia-induced alveolar cell death in mice. The study investigates whether NOX1 expression is modulated in epithelial cells concomitantly to cell death and associated to STAT3 signaling in the exudative phase of ARDS. In addition, the role of STAT3 activation in NOX1-dependent epithelial cell death was confirmed by using a lung epithelial cell line and in mice exposed to hyperoxia. NOX1 expression, cell death and STAT3 staining were evaluated in the lungs of control and ARDS patients by immunohistochemistry. In parallel, a stable NOX1-silenced murine epithelial cell line (MLE12) and NOX1-deficient mice were used to characterize signalling pathways. In the present study, we show that NOX1 is detected in alveolar epithelial cells of ARDS patients in the exudative stage. In addition, increased alveolar epithelial cell death and phosphorylated STAT3 are observed in ARDS patients and associated with NOX1 expression. Phosphorylated STAT3 is also correlated with TUNEL staining. We also confirmed that NOX1-dependent STAT3 activation participates to alveolar epithelial cell death. Silencing and acute inhibition of NOX1 in MLE12 led to decreased cell death and cleaved-caspase 3 induced by hyperoxia. Additionally, hyperoxia-induced STAT3 phosphorylation is dependent on NOX1 expression and associated with cell death in MLE12 and mice. This study demonstrates that NOX1 is involved in human ARDS pathophysiology and is responsible for the damage occurring in alveolar epithelial cells at least in part via STAT3 signalling pathways. PMID:24551274

  17. New insights into the Shwachman-Diamond Syndrome-related haematological disorder: hyper-activation of mTOR and STAT3 in leukocytes

    PubMed Central

    Bezzerri, Valentino; Vella, Antonio; Calcaterra, Elisa; Finotti, Alessia; Gasparello, Jessica; Gambari, Roberto; Assael, Baroukh Maurice; Cipolli, Marco; Sorio, Claudio

    2016-01-01

    Shwachman-Diamond syndrome (SDS) is an inherited disease caused by mutations of a gene encoding for SBDS protein. So far little is known about SBDS exact function. SDS patients present several hematological disorders, including neutropenia and myelodysplastic syndrome (MDS), with increased risk of leukemic evolution. So far, the molecular mechanisms that underlie neutropenia, MDS and AML in SDS patients have been poorly investigated. STAT3 is a key regulator of several cellular processes including survival, differentiation and malignant transformation. Moreover, STAT3 has been reported to regulate neutrophil granulogenesis and to induce several kinds of leukemia and lymphoma. STAT3 activation is known to be regulated by mTOR, which in turn plays an important role in cellular growth and tumorigenesis. Here we show for the first time, to the best of our knowledge, that both EBV-immortalized B cells and primary leukocytes obtained from SDS patients present a constitutive hyper-activation of mTOR and STAT3 pathways. Interestingly, loss of SBDS expression is associated with this process. Importantly, rapamycin, a well-known mTOR inhibitor, is able to reduce STAT3 phosphorylation to basal levels in our experimental model. A novel therapeutic hypothesis targeting mTOR/STAT3 should represent a significant step forward into the SDS clinical practice. PMID:27658964

  18. Identification of canonical tyrosine-dependent and non-canonical tyrosine-independent STAT3 activation sites in the intracellular domain of the interleukin 23 receptor.

    PubMed

    Floss, Doreen M; Mrotzek, Simone; Klöcker, Tobias; Schröder, Jutta; Grötzinger, Joachim; Rose-John, Stefan; Scheller, Jürgen

    2013-07-05

    Signaling of interleukin 23 (IL-23) via the IL-23 receptor (IL-23R) and the shared IL-12 receptor β1 (IL-12Rβ1) controls innate and adaptive immune responses and is involved in the differentiation and expansion of IL-17-producing CD4(+) T helper (TH17) cells. Activation of signal transducer and activator of transcription 3 (STAT3) appears to be the major signaling pathway of IL-23, and STAT binding sites were predicted in the IL-23R but not in the IL-12Rβ1 chain. Using site-directed mutagenesis and deletion variants of the murine and human IL-23R, we showed that the predicted STAT binding sites (pYXXQ; including Tyr-504 and Tyr-626 in murine IL-23R and Tyr-484 and Tyr-611 in human IL-23R) mediated STAT3 activation. Furthermore, we identified two uncommon STAT3 binding/activation sites within the murine IL-23R. First, the murine IL-23R carried the Y(542)PNFQ sequence, which acts as an unusual Src homology 2 (SH2) domain-binding protein activation site of STAT3. Second, we identified a non-canonical, phosphotyrosine-independent STAT3 activation motif within the IL-23R. A third predicted site, Tyr-416 in murine and Tyr-397 in human IL-23R, is involved in the activation of PI3K/Akt and the MAPK pathway leading to STAT3-independent proliferation of Ba/F3 cells upon stimulation with IL-23. In contrast to IL-6-induced short term STAT3 phosphorylation, cellular activation by IL-23 resulted in a slower but long term STAT3 phosphorylation, indicating that the IL-23R might not be a major target of negative feedback inhibition by suppressor of cytokine signaling (SOCS) proteins. In summary, we characterized IL-23-dependent signal transduction with a focus on STAT3 phosphorylation and identified canonical tyrosine-dependent and non-canonical tyrosine-independent STAT3 activation sites in the IL-23R.

  19. Identification of Canonical Tyrosine-dependent and Non-canonical Tyrosine-independent STAT3 Activation Sites in the Intracellular Domain of the Interleukin 23 Receptor*

    PubMed Central

    Floss, Doreen M.; Mrotzek, Simone; Klöcker, Tobias; Schröder, Jutta; Grötzinger, Joachim; Rose-John, Stefan; Scheller, Jürgen

    2013-01-01

    Signaling of interleukin 23 (IL-23) via the IL-23 receptor (IL-23R) and the shared IL-12 receptor β1 (IL-12Rβ1) controls innate and adaptive immune responses and is involved in the differentiation and expansion of IL-17-producing CD4+ T helper (TH17) cells. Activation of signal transducer and activator of transcription 3 (STAT3) appears to be the major signaling pathway of IL-23, and STAT binding sites were predicted in the IL-23R but not in the IL-12Rβ1 chain. Using site-directed mutagenesis and deletion variants of the murine and human IL-23R, we showed that the predicted STAT binding sites (pYXXQ; including Tyr-504 and Tyr-626 in murine IL-23R and Tyr-484 and Tyr-611 in human IL-23R) mediated STAT3 activation. Furthermore, we identified two uncommon STAT3 binding/activation sites within the murine IL-23R. First, the murine IL-23R carried the Y542PNFQ sequence, which acts as an unusual Src homology 2 (SH2) domain-binding protein activation site of STAT3. Second, we identified a non-canonical, phosphotyrosine-independent STAT3 activation motif within the IL-23R. A third predicted site, Tyr-416 in murine and Tyr-397 in human IL-23R, is involved in the activation of PI3K/Akt and the MAPK pathway leading to STAT3-independent proliferation of Ba/F3 cells upon stimulation with IL-23. In contrast to IL-6-induced short term STAT3 phosphorylation, cellular activation by IL-23 resulted in a slower but long term STAT3 phosphorylation, indicating that the IL-23R might not be a major target of negative feedback inhibition by suppressor of cytokine signaling (SOCS) proteins. In summary, we characterized IL-23-dependent signal transduction with a focus on STAT3 phosphorylation and identified canonical tyrosine-dependent and non-canonical tyrosine-independent STAT3 activation sites in the IL-23R. PMID:23673666

  20. Polyunsaturated fatty acids promote the expansion of myeloid-derived suppressor cells by activating the JAK/STAT3 pathway.

    PubMed

    Yan, Dehong; Yang, Quan; Shi, Maohua; Zhong, Limei; Wu, Changyou; Meng, Tao; Yin, Huiyong; Zhou, Jie

    2013-11-01

    Polyunsaturated fatty acids (PUFAs) exert immunosuppressive effects that could prove beneficial in clinical therapies for certain autoimmune and inflammatory disorders. However, the mechanism of PUFA-mediated immunosuppression is far from understood. Here, we provide evidence that PUFAs enhance the accumulation of myeloid-derived suppressor cells (MDSCs), a negative immune regulator. PUFA-induced MDSCs have a more potent suppressive effect on T-cell responses than do control MDSCs. These observations were found both in cultured mouse bone marrow cells in vitro and in vivo in mice fed diets enriched in PUFAs. The enhanced suppressive activity of MDSCs by PUFAs administration was coupled with a dramatic induction of nicotinamide adenine dinucleo- tide phosphate oxidase subunit p47(phox) and was dependent on reactive oxygen species (ROS) production. Mechanistic studies revealed that PUFAs mediate its effects through JAK-STAT3 signaling. Inhibition of STAT3 phosphorylation by JAK inhibitor JSI-124 almost completely abrogated the effects of PUFAs on MDSCs. Moreover, the effects of PUFAs on MDSCs and the underlying mechanisms were confirmed in tumor-bearing mice. In summary, this study sheds new light on the immune modulatory role of PUFAs, and demonstrates that MDSCs expansion may mediate the effects of PUFAs on the immune system.

  1. Altered gp130 signalling ameliorates experimental colitis via myeloid cell-specific STAT3 activation and myeloid-derived suppressor cells

    PubMed Central

    Däbritz, Jan; Judd, Louise M.; Chalinor, Heather V.; Menheniott, Trevelyan R.; Giraud, Andrew S.

    2016-01-01

    STAT3 regulates the expansion of myeloid-derived suppressor cells (MDSCs) during inflammation, infection and cancer. Hyperactivation of STAT3 in gp130757F/F mice is associated with protection from experimental colitis. This study determined mechanisms for this protection and compared this to mice with myeloid-specific STAT3-deficiency (LysMcre/STAT3flox; gp130757F/F LysMcre/STAT3flox). Acute and chronic colitis was induced and colons were removed for histological, mRNA and protein analysis. Cell populations from spleen, mesenteric lymph node and colon were analyzed for different myeloid cell populations using flow cytometry. Functions of MDSCs and LPS-stimulated peritoneal macrophages were further characterized by in vitro and in vivo assays. Here we show that the resistance to experimental colitis in gp130757F/F mice is via myeloid-cell specific STAT3 activation, MDSC expansion and increased production of suppressive and protective cytokines. PMID:26848037

  2. IL-26 promotes the proliferation and survival of human gastric cancer cells by regulating the balance of STAT1 and STAT3 activation.

    PubMed

    You, Wei; Tang, Qiyun; Zhang, Chuanyong; Wu, Jindao; Gu, Chunrong; Wu, Zhengshan; Li, Xiangcheng

    2013-01-01

    Interleukin-26 (IL-26) is one of the cytokines secreted by Th17 cells whose role in human tumors remains unknown. Here, we investigated the expression and potential role of IL-26 in human gastric cancer (GC). The expression of IL-26 and related molecules such as IL-20R1, STAT1 and STAT3 was examined by real-time PCR and immunohistochemisty. The effects of IL-26 on cell proliferation and cisplatin-induced apoptosis were analyzed by BrdU cooperation assay and PI-Annexin V co-staining, respectively. Lentiviral mediated siRNA was used to explore its mechanism of action, and IL-26 related signaling was analyzed by western blotting. Human GC tissues showed increased levels of IL-26 and its related molecules and activation of STAT3 signaling, whereas STAT1 activation did not differ significantly between GC and normal gastric tissues. Moreover, IL-26 was primarily produced by Th17 and NK cells. IL-26 promoted the proliferation and survival of MKN45 and SGC-7901 gastric cancer cells in a dose-dependent manner. Furthermore, IL-20R2 and IL-10R1, which are two essential receptors for IL-26 signaling, were expressed in both cell lines. IL-26 activated STAT1 and STAT3 signaling; however, the upregulation of the expression of Bcl-2, Bcl-xl and c-myc indicated that the effect of IL-26 is mediated by STAT3 activation. Knockdown of STAT1 and STAT3 expression suggested that the proliferative and anti-apoptotic effects of IL-26 are mediated by the modulation of STAT1/STAT3 activation. In summary, elevated levels of IL-26 in human GC promote proliferation and survival by modulating STAT1/STAT3 signaling.

  3. Alternative Splicing of STAT3 Is Affected by RNA Editing.

    PubMed

    Goldberg, Lior; Abutbul-Amitai, Mor; Paret, Gideon; Nevo-Caspi, Yael

    2017-03-09

    A-to-I RNA editing, carried out by adenosine deaminase acting on RNA (ADAR) enzymes, is an epigenetic phenomenon of posttranscriptional modifications on pre-mRNA. RNA editing in intronic sequences may influence alternative splicing of flanking exons. We have previously shown that conditions that induce editing result in elevated expression of signal transducer and activator of transcription 3 (STAT3), preferentially the alternatively-spliced STAT3β isoform. Mechanisms regulating alternative splicing of STAT3 have not been elucidated. STAT3 undergoes A-to-I RNA editing in an intron residing in proximity to the alternatively spliced exon. We hypothesized that RNA editing plays a role in regulating alternative splicing toward STAT3β. In this study we extend our observation connecting RNA editing to the preferential induction of STAT3β expression. We study the involvement of ADAR1 in STAT3 editing and reveal the connection between editing and alternative splicing of STAT3. Deferoaxamine treatment caused the induction in STAT3 RNA editing and STAT3β expression. Silencing ADAR1 caused a decrease in STAT3 editing and expression with a preferential decrease in STAT3β. Cells transfected with a mutated minigene showed preferential splicing toward the STAT3β transcript. Editing in the STAT3 intron is performed by ADAR1 and affects STAT3 alternative splicing. These results suggest that RNA editing is one of the molecular mechanisms regulating the expression of STAT3β.

  4. The autophagy molecule Beclin 1 maintains persistent activity of NF-κB and Stat3 in HTLV-1-transformed T lymphocytes.

    PubMed

    Chen, Li; Liu, Dan; Zhang, Yang; Zhang, Huan; Cheng, Hua

    2015-10-02

    The retroviral oncoprotein Tax from human T cell leukemia virus type 1 (HTLV-1) induces persistent activation of IκB kinase (IKK)/NF-κB signaling, an essential step for initiating HTLV-1 oncogenesis. The regulation of the IKK/NF-κB signaling in HTLV-1-transformed T cells remains incompletely understood. In the present study, we showed that the autophagy molecule Beclin1 not only executed a cytoprotective function through induction of autophagy but also played a pivotal role in maintaining Tax-induced activation of two key survival factors, NF-κB and Stat3. Silencing Beclin1 in HTLV-1-transformed T cells resulted in diminished activities of NF-κB and Stat3 as well as impaired growth. In Beclin1-depleted cells, Tax failed to activate NF-κB and Stat3 at its full capacity. In addition, we showed that Beclin1 interacted with the catalytic subunits of IKK. Further, we observed that selective inhibition of IKK repressed the activities of both NF-κB and Stat3 in the context of HTLV-1-transformation of T cells. Our data, therefore, unveiled a key role of Beclin1 in maintaining persistent activities of both NF-κB and Stat3 in the pathogenesis of HTLV-1-mediated oncogenesis.

  5. The stem cell adjuvant with Exendin-4 repairs the heart after myocardial infarction via STAT3 activation

    PubMed Central

    Liu, Jianfeng; Wang, Haibin; Wang, Yan; Yin, Yujing; Du, Zhiyan; Liu, Zhiqiang; Yang, Junjie; Hu, Shunying; Wang, Changyong; Chen, Yundai

    2014-01-01

    The poor survival of cells in ischaemic myocardium is a major obstacle for stem cell therapy. Exendin-4 holds the potential of cardioprotective effect based on its pleiotropic activity. This study investigated whether Exendin-4 in conjunction with adipose-derived stem cells (ADSCs) could improve the stem cell survival and contribute to myocardial repairs after infarction. Myocardial infarction (MI) was induced by the left anterior descending artery ligation in adult male Sprague-Dawley rats. ADSCs carrying double-fusion reporter gene [firefly luciferase and monomeric red fluorescent protein (fluc-mRFP)] were quickly injected into border zone of MI in rats treated with or without Exendin-4. Exendin-4 enhanced the survival of transplanted ADSCs, as demonstrated by the longitudinal in vivo bioluminescence imaging. Moreover, ADSCs adjuvant with Exendin-4 decreased oxidative stress, apoptosis and fibrosis. They also improved myocardial viability and cardiac function and increased the differentiation rates of ADSCs into cardiomyocytes and vascular smooth muscle cells in vivo. Then, ADSCs were exposed to hydrogen peroxide/serum deprivation (H2O2/SD) to mimic the ischaemic environment in vitro. Results showed that Exendin-4 decreased the apoptosis and enhanced the paracrine effect of ADSCs. In addition, Exendin-4 activated signal transducers and activators of transcription 3 (STAT3) through the phosphorylation of Akt and ERK1/2. Furthermore, Exendin-4 increased the anti-apoptotic protein Bcl-2, but decreased the pro-apoptotic protein Bax of ADSCs. In conclusion, Exendin-4 could improve the survival and therapeutic efficacy of transplanted ADSCs through STAT3 activation via the phosphorylation of Akt and ERK1/2. This study suggests the potential application of Exendin-4 for stem cell–based heart regeneration. PMID:24779911

  6. In vivo and in vitro evidence that chronic activation of the hexosamine biosynthetic pathway interferes with leptin-dependent STAT3 phosphorylation

    PubMed Central

    Zimmerman, Arthur D.

    2015-01-01

    We previously reported that a 2-day peripheral infusion of glucosamine caused leptin resistance in rats, suggesting a role for the hexosamine biosynthetic pathway (HBP) in the development of leptin resistance. Here we tested leptin responsiveness in mice in which HBP activity was stimulated by offering 30% sucrose solution in addition to chow and water or by infusing glucosamine. Mice were leptin resistant after 33 days of access to sucrose. Resistance was associated with increased activity of the HBP and with phosphorylation of transcription factor signal transducer and activator of transcription-3 Tyr705 [pSTAT3(Y705)] but inhibition of suppressor of cytokine signaling 3 in the liver and hypothalamus. Intravenous infusion of glucosamine for 3 h stimulated pSTAT3(Y705) but prevented leptin-induced phosphorylation of STAT3(S727). In an in vitro system, glucose, glucosamine, and leptin each dose dependently increased O-linked β-N-acetylglucosamine (O-GlcNAc) protein and pSTAT3(Y705) in HepG2 cells. To test the effect of glucose on leptin responsiveness cells were incubated in 5.5 mM (LG) or 20 mM (HG) glucose for 18 h and were treated with 0 or 50 ng/ml leptin for 15 min. HG alone and LG + leptin produced similar increases in O-GlcNAc protein, glutamine fructose-6-phosphate amidotransferase (GFAT), and pSTAT3(Y705) compared with LG media. Leptin did not stimulate these proteins in HG cells, suggesting leptin resistance. Leptin-induced pSTAT3(S727) was prevented by HG media. Inhibition of GFAT with azaserine prevented LG + leptin and HG stimulation of pSTAT3. These data demonstrate development of leptin resistance in sucrose-drinking mice and provide new evidence of leptin-induced stimulation of the HBP. PMID:25568075

  7. Acquisition of resistance to trastuzumab in gastric cancer cells is associated with activation of IL-6/STAT3/Jagged-1/Notch positive feedback loop.

    PubMed

    Yang, Zhengyan; Guo, Liang; Liu, Dan; Sun, Limin; Chen, Hongyu; Deng, Que; Liu, Yanjun; Yu, Ming; Ma, Yuanfang; Guo, Ning; Shi, Ming

    2015-03-10

    In the present study, we demonstrate that prolonged treatment by trastuzumab induced resistance of NCI-N87 gastric cancer cells to trastuzumab. The resistant cells possessed typical characteristics of epithelial to mesenchymal transition (EMT)/cancer stem cells and acquired more invasive and metastatic potentials both in vitro and in vivo. Long term treatment with trastuzumab dramatically inhibited the phosphorylation of Akt, but triggered the activation of STAT3. The level of IL-6 was remarkably increased, implicating that the release of IL-6 that drives the STAT3 activation initiates the survival signaling transition. Furthermore, the Notch activities were significantly enhanced in the resistant cells, companied by upregulation of the Notch ligand Jagged-1 and the Notch responsive genes Hey1 and Hey2. Inhibiting the endogenous Notch pathway reduced the IL-6 expression and restored the sensitivities of the resistant cells to trastuzumab. Blocking of the STAT3 signaling abrogated IL-6-induced Jagged-1 expression, effectively inhibited the growth of the trastuzumab resistant cells, and enhanced the anti-tumor activities of trastuzumab in the resistant cells. These findings implicate that the IL-6/STAT3/Jagged-1/Notch axis may be a useful target and that combination of the Notch or STAT3 inhibitors with trastuzumab may prevent or delay clinical resistance and improve the efficacy of trastuzumab in gastric cancer.

  8. Signal transducer and activator of transcription (stat) binding sites but not stat3 are required for fasting-induced transcription of agouti-related protein messenger ribonucleic acid.

    PubMed

    Kaelin, Christopher B; Gong, Lijie; Xu, Allison Wanting; Yao, Fayi; Hockman, Kristin; Morton, Gregory J; Schwartz, Michael W; Barsh, Gregory S; MacKenzie, Robert G

    2006-10-01

    Energy homeostasis depends on the regulation of hypothalamic neurons by leptin, an adipocyte hormone whose circulating levels communicate body energy stores. Leptin activates the transcription factor signal transducer and activator of transcription 3 (Stat3) in hypothalamic neurons, including neuronal subtypes producing Agouti-related protein (Agrp), a neuropeptide that stimulates feeding. Previous studies have suggested a model in which high levels of Agrp transcription during fasting represent a default state that is actively repressed by phospho-Stat3 induced by leptin signaling in the fed state. We identify putative Stat3 binding elements in the Agrp promoter that have been highly conserved during vertebrate evolution. Using a reporter assay in transgenic mice that faithfully recapitulates normal regulation of Agrp, we show that these sites are required, but in a way opposite to that predicted by the existing model: mutation of the sites leads to a default state characterized by a low level of Agrp transcription and insensitivity to fasting. We also find that removing activatable Stat3 from Agrp neurons has no detectable effect on steady-state levels of Agrp mRNA in the fed or fasted state. These results suggest a new model for transcriptional regulation of orexigenic neuropeptides in which the default level of expression is low in the fed state, and transcriptional activation in response to fasting is mediated by factors other than Stat3.

  9. Inhibition of STAT3 activity re-activates anti-tumor immunity but fails to restore the immunogenicity of tumor cells in a B-cell lymphoma model

    PubMed Central

    Cao, Yang; Zhou, Xiaoxi; Zhou, Mi; Xu, Danmei; Ma, Quanfu; Zhang, Peilin; Huang, Xiaoyuan; Li, Qinlu; Ma, Ding; Zhou, Jianfeng

    2014-01-01

    A large number of patients with advanced lymphoma become refractory or relapse after initial treatment due to the persistence of minimal residual disease. Ideal immunotherapy strategy for eradicating the minimal residual disease of lymphoma and preventing the tendency to relapse need to be developed. Here, we use a mice model mimicked the disease entities of aggressive B-cell lymphoma dynamically to analyze the host anti-lymphoma immunity during the progression of lymphoma. We have shown that STAT3 activity was gradually enhanced in host immune effector cells with the progression of lymphoma. Inhibition of the STAT3 activity with a small molecule inhibitor was able to effectively enhance the function of both host innate and adaptive immunity, and thereby delayed the progression of lymphoma. Despite the therapeutic benefits were achieved by using of the STAT3 inhibitor, disrupting of STAT3 pathway did not prevent the eventual development of lymphoma due to the presence of point mutation of β2M, which controls immune recognition by T cells. Our findings highlight the complexity of the mechanism of immune evasion; therefore a detailed analysis of genes involved in the immune recognition process should be essential before an elegant immunotherapy strategy could be conducted. PMID:24915165

  10. Selenate Enhances STAT3 Transcriptional Activity in Endothelial Cells: Differential Actions of Selenate and Selenite on LIF Cytokine Signaling and Cell Viability

    PubMed Central

    Alturkmani, Hani J.; Zgheib, Carlos; Zouein, Fouad A.; Alshaaer, Nour Eddin F.; Kurdi, Mazen; Booz, George W.

    2012-01-01

    Sodium selenate may have utility in treating Alzheimer’s disease and diabetes; however, its impact on the associated proinflammatory cytokine signaling of endothelial cells has not been investigated. We report that treatment of human microvascular endothelial cells with sodium selenate at a pharmacological dose (100 μM) enhanced tyrosine phosphorylation of nuclear STAT3 on Y705 in response to IL-6-type cytokine, leukemia inhibitory factor (LIF), indicative of enhanced STAT3 activity. Accordingly, STAT3 nuclear binding to DNA was increased, as well as LIF-induced gene expression of chemokine (C-C motif) ligand 2 (CCL2). CCL2 plays a key role in inflammatory processes associated with neuronal degenerative and vascular diseases. The enhancing action of selenate on LIF-induced STAT3 Y705 phosphorylation was replicated by vanadate and a specific inhibitor of protein tyrosine phosphatase, non-receptor type 1 (PTP1B). Moreover, we observed that selenite, the cellular reduction bioproduct of selenate but not selenate itself, inhibited enzymatic activity of human recombinant PTP1B. Our findings support the conclusion that in human microvascular endothelial cells selenate has a vanadate-like effect in inhibiting PTP1B and enhancing proinflammatory STAT3 activation. These findings raise the possibility that beneficial actions of supranutritional levels of selenate for treating Alzheimer’s and diabetes may be offset by a proinflammatory action on endothelial cells. PMID:22366233

  11. Selenate enhances STAT3 transcriptional activity in endothelial cells: differential actions of selenate and selenite on LIF cytokine signaling and cell viability.

    PubMed

    Alturkmani, Hani J; Zgheib, Carlos; Zouein, Fouad A; Alshaaer, Nour Eddin F; Kurdi, Mazen; Booz, George W

    2012-04-01

    Sodium selenate may have utility in treating Alzheimer's disease and diabetes; however, its impact on the associated proinflammatory cytokine signaling of endothelial cells has not been investigated. We report that treatment of human microvascular endothelial cells with sodium selenate at a pharmacological dose (100 μM) enhanced tyrosine phosphorylation of nuclear STAT3 on Y705 in response to IL-6-type cytokine, leukemia inhibitory factor (LIF), indicative of enhanced STAT3 activity. Accordingly, STAT3 nuclear binding to DNA was increased, as well as LIF-induced gene expression of chemokine (C-C motif) ligand 2 (CCL2). CCL2 plays a key role in inflammatory processes associated with neuronal degenerative and vascular diseases. The enhancing action of selenate on LIF-induced STAT3 Y705 phosphorylation was replicated by vanadate and a specific inhibitor of protein tyrosine phosphatase, non-receptor type 1 (PTP1B). Moreover, we observed that selenite, the cellular reduction bioproduct of selenate but not selenate itself, inhibited enzymatic activity of human recombinant PTP1B. Our findings support the conclusion that in human microvascular endothelial cells selenate has a vanadate-like effect in inhibiting PTP1B and enhancing proinflammatory STAT3 activation. These findings raise the possibility that beneficial actions of supranutritional levels of selenate for treating Alzheimer's and diabetes may be offset by a proinflammatory action on endothelial cells.

  12. Differential STAT3 signaling in the heart

    PubMed Central

    Zgheib, Carlos; Zouein, Fouad A.; Kurdi, Mazen; Booz, George W.

    2012-01-01

    Multiple lines of evidence suggest that the transcription factor STAT3 is linked to a protective and reparative response in the heart. Thus, increasing duration or intensity of STAT3 activation ought to minimize damage and improve heart function under conditions of stress. Two recent studies using genetic mouse models, however, report findings that appear to refute this proposition. Unfortunately, studies often approach the question of the role of STAT3 in the heart from the perspective that all STAT3 signaling is equivalent, particularly when it comes to signaling by IL-6 type cytokines, which share the gp130 signaling protein. Moreover, STAT3 activation is typically equated with phosphorylation of a critical tyrosine residue. Yet, STAT3 transcriptional behavior is subject to modulation by serine phosphorylation, acetylation, and redox status of the cell. Unphosphorylated STAT3 is implicated in gene induction as well. Thus, how STAT3 is activated and also what other signaling events are occurring at the same time is likely to impact on the outcome ultimately linked to STAT3. Notably STAT3 may serve as a scaffold protein allowing it to interact with other singling pathways. In this context, canonical gp130 cytokine signaling may function to integrate STAT3 signaling with a protective PI3K/AKT signaling network via mutual involvement of JAK tyrosine kinases. Differences in the extent of integration may occur between those cytokines that signal through gp130 homodimers and those through heterodimers of gp130 with a receptor α chain. Signal integration may have importance not only for deciding the particular gene profile linked to STAT3, but for the newly described mitochondrial stabilization role of STAT3 as well. In addition, disruption of integrated gp130-related STAT3 signaling may occur under conditions of oxidative stress, which negatively impacts on JAK catalytic activity. For these reasons, understanding the importance of STAT3 signaling to heart function

  13. Interleukin-6 Knockout Prevents Angiotensin II Hypertension: Role of Renal Vasoconstriction and JAK2/STAT3 Activation

    PubMed Central

    Brands, Michael W.; Banes-Berceli, Amy K.L.; Inscho, Edward W.; Al-Azawi, Hind; Allen, Ashlyn J.; Labazi, Hicham

    2012-01-01

    Chronic angiotensin II (AngII) infusion stimulates IL-6 release, and we and others have shown that preventing the increase in IL-6 significantly attenuates AngII hypertension. This study measured renal blood flow (RBF) chronically, using Transonic flow probes in wildtype (WT) and IL-6 knockout (KO) mice, to determine the role of renal blood flow regulation in that response. AngII infusion at 200, 800, and 3600 ng/kg/min caused a dose-dependent decrease in renal blood flow in WT mice, and the response at 800 ng/kg/min was compared between WT and IL-6 KO mice. AngII infusion increased plasma IL-6 concentration in WT mice and increased MAP (19 hrs/day; DSI telemetry) from 113±4 to 149±4 mmHg (Δ 36 mmHg) over the 7-day infusion period, and that effect was blocked in IL-6 KO mice (119±7 to 126±7 mmHg). RBF decreased to an average of 61±8% of control over the 7-day period (control = 0.86±0.02 ml/min) in the WT mice; however, the average decrease to 72±6% of control (control = 0.88±0.02 ml/min) in the KO mice was not significantly different. There also was no difference in afferent arteriolar constriction by AngII in blood-perfused juxtamedullary nephrons in WT vs. KO mice. Phosphorylation of JAK2 and STAT3 in renal cortex homogenates increased significantly in AngII-infused WT mice, and that effect was prevented completely in AngII-infused IL-6 KO mice. These data suggest that IL-6-dependent activation of the renal JAK2/STAT3 pathway plays a role in AngII hypertension, but not by mediating the effect of AngII to decrease total renal blood flow. PMID:20921429

  14. Constitutive activation of signal transducer and activator of transcription 3 (STAT3) and nuclear factor κB signaling in glioblastoma cancer stem cells regulates the Notch pathway.

    PubMed

    Garner, Jo Meagan; Fan, Meiyun; Yang, Chuan He; Du, Ziyun; Sims, Michelle; Davidoff, Andrew M; Pfeffer, Lawrence M

    2013-09-06

    Malignant gliomas are locally aggressive, highly vascular tumors that have a dismal prognosis, and present therapies provide little improvement in the disease course and outcome. Many types of malignancies, including glioblastoma, originate from a population of cancer stem cells (CSCs) that are able to initiate and maintain tumors. Although CSCs only represent a small fraction of cells within a tumor, their high tumor-initiating capacity and therapeutic resistance drives tumorigenesis. Therefore, it is imperative to identify pathways associated with CSCs to devise strategies to selectively target them. In this study, we describe a novel relationship between glioblastoma CSCs and the Notch pathway, which involves the constitutive activation of STAT3 and NF-κB signaling. Glioma CSCs were isolated and maintained in vitro using an adherent culture system, and the biological properties were compared with the traditional cultures of CSCs grown as multicellular spheres under nonadherent culture conditions. Interestingly, both adherent and spheroid glioma CSCs show constitutive activation of the STAT3/NF-κB signaling pathway and up-regulation of STAT3- and NF-κB-dependent genes. Gene expression profiling also identified components of the Notch pathway as being deregulated in glioma CSCs, and the deregulated expression of these genes was sensitive to treatment with STAT3 and NF-κB inhibitors. This finding is particularly important because Notch signaling appears to play a key role in CSCs in a variety of cancers and controls cell fate determination, survival, proliferation, and the maintenance of stem cells. The constitutive activation of STAT3 and NF-κB signaling pathways that leads to the regulation of Notch pathway genes in glioma CSCs identifies novel therapeutic targets for the treatment of glioma.

  15. YC-1 enhances the anti-tumor activity of sorafenib through inhibition of signal transducer and activator of transcription 3 (STAT3) in hepatocellular carcinoma

    PubMed Central

    2014-01-01

    Background Traditional systemic chemotherapy does not provide survival benefits in patients with hepatocellular carcinoma (HCC). Molecular targeted therapy shows promise for HCC treatment, however, the duration of effectiveness for targeted therapies is finite and combination therapies offer the potential for improved effectiveness. Methods Sorafenib, a multikinase inhibitor, and YC-1, a soluble guanylyl cyclase (sGC) activator, were tested in HCC by proliferation assay, cell cycle analysis and western blot in vitro and orthotopic and ectopic HCC models in vivo. Results In vitro, combination of sorafenib and YC-1 synergistically inhibited proliferation and colony formation of HepG2, BEL-7402 and HCCLM3 cells. The combination also induced S cell cycle arrest and apoptosis, as observed by activated PARP and caspase 8. Sorafenib and YC-1 respectively suppressed the expression of phosphorylated STAT3 (p-STAT3) (Y705) in a dose- and time-dependent manner. Combination of sorafenib and YC-1 significantly inhibited the expression of p-STAT3 (Y705) (S727), p-ERK1/2, cyclin D1 and survivin and SHP-1 activity compared with sorafenib or YC-1 used alone in all tested HCC cell lines. In vivo, sorafenib-YC-1 combination significantly suppressed the growth of HepG2 tumor xenografts with decreased cell proliferation and increased apoptosis observed by PCNA and PARP. Similar results were also confirmed in a HCCLM3 orthotopic model. There was a reduction in CD31-positive blood vessels and reduced VEGF expression, which suggested a combinational effect of sorafenib and YC-1 on angiogenesis. The reduced expression of p-STAT3, cyclin D1 and survivin was also observed with the combination of sorafenib and YC-1. Conclusions Our data show that sorafenib-YC-1 combination is a novel potent therapeutic agent that can target the STAT3 signaling pathway to inhibit HCC tumor growth. PMID:24418169

  16. The inflammatory/cancer-related IL-6/STAT3/NF-κB positive feedback loop includes AUF1 and maintains the active state of breast myofibroblasts

    PubMed Central

    Hendrayani, Siti-Fauziah; Al-Harbi, Bothaina; Al-Ansari, Mysoon M.; Silva, Gabriela; Aboussekhra, Abdelilah

    2016-01-01

    The IL-6/STAT3/NF-κB positive feedback loop links inflammation to cancer and maintains cells at a transformed state. Similarly, cancer-associated myofibroblats remains active even in absence of cancer cells. However, the molecular basis of this sustained active state remains elusive. We have shown here that breast cancer cells and IL-6 persistently activate breast stromal fibroblasts through the stimulation of the positive IL-6/STAT3/NF-κB feedback loop. Transient neutralization of IL-6 in culture inhibited this signaling circuit and reverted myofibrobalsts to a normalized state, suggesting the implication of the IL-6 autocrine feedback loop as well. Importantly, the IL-6/STAT3/NF-κB pro-inflammatory circuit was also active in cancer-associated fibroblasts isolated from breast cancer patients. Transient inhibition of STAT3 by specific siRNA in active fibroblasts persistently reduced the level of the RNA binding protein AUF1, blocked the loop and normalized these cells. Moreover, we present clear evidence that AUF1 is also part of this positive feedback loop. Interestingly, treatment of breast myofibroblasts with caffeine, which has been previously shown to persistently inhibit active breast stromal fibroblasts, blocked the positive feedback loop through potent and sustained inhibition of STAT3, AKT, lin28B and AUF1. These results indicate that the IL-6/STAT3/NF-κB positive feedback loop includes AUF1 and is responsible for the sustained active status of cancer-associated fibroblasts. We have also shown that normalizing myofibroblasts, which could be of great therapeutic value, is possible through the inhibition of this procarcinogenic circuit. PMID:27248826

  17. The inflammatory/cancer-related IL-6/STAT3/NF-κB positive feedback loop includes AUF1 and maintains the active state of breast myofibroblasts.

    PubMed

    Hendrayani, Siti-Fauziah; Al-Harbi, Bothaina; Al-Ansari, Mysoon M; Silva, Gabriela; Aboussekhra, Abdelilah

    2016-07-05

    The IL-6/STAT3/NF-κB positive feedback loop links inflammation to cancer and maintains cells at a transformed state. Similarly, cancer-associated myofibroblats remains active even in absence of cancer cells. However, the molecular basis of this sustained active state remains elusive. We have shown here that breast cancer cells and IL-6 persistently activate breast stromal fibroblasts through the stimulation of the positive IL-6/STAT3/NF-κB feedback loop. Transient neutralization of IL-6 in culture inhibited this signaling circuit and reverted myofibrobalsts to a normalized state, suggesting the implication of the IL-6 autocrine feedback loop as well. Importantly, the IL-6/STAT3/NF-κB pro-inflammatory circuit was also active in cancer-associated fibroblasts isolated from breast cancer patients. Transient inhibition of STAT3 by specific siRNA in active fibroblasts persistently reduced the level of the RNA binding protein AUF1, blocked the loop and normalized these cells. Moreover, we present clear evidence that AUF1 is also part of this positive feedback loop. Interestingly, treatment of breast myofibroblasts with caffeine, which has been previously shown to persistently inhibit active breast stromal fibroblasts, blocked the positive feedback loop through potent and sustained inhibition of STAT3, AKT, lin28B and AUF1. These results indicate that the IL-6/STAT3/NF-κB positive feedback loop includes AUF1 and is responsible for the sustained active status of cancer-associated fibroblasts. We have also shown that normalizing myofibroblasts, which could be of great therapeutic value, is possible through the inhibition of this procarcinogenic circuit.

  18. Epstein-Barr Virus Latent Membrane Protein 2A (LMP2A) enhances IL-10 production through the activation of Bruton's tyrosine kinase and STAT3.

    PubMed

    Incrocci, Ryan; Barse, Levi; Stone, Amanda; Vagvala, Sai; Montesano, Michael; Subramaniam, Vijay; Swanson-Mungerson, Michelle

    2017-01-01

    Previous data demonstrate that Epstein-Barr Virus Latent Membrane Protein 2A (LMP2A) enhances IL-10 to promote the survival of LMP2A-expressing B cell lymphomas. Since STAT3 is an important regulator of IL-10 production, we hypothesized that LMP2A activates a signal transduction cascade that increases STAT3 phosphorylation to enhance IL-10. Using LMP2A-negative and -positive B cell lines, the data indicate that LMP2A requires the early signaling molecules of the Syk/RAS/PI3K pathway to increase IL-10. Additional studies indicate that the PI3K-regulated kinase, BTK, is responsible for phosphorylating STAT3, which ultimately mediates the LMP2A-dependent increase in IL-10. These data are the first to show that LMP2A signaling results in STAT3 phosphorylation in B cells through a PI3K/BTK-dependent pathway. With the use of BTK and STAT3 inhibitors to treat B cell lymphomas in clinical trials, these findings highlight the possibility of using new pharmaceutical approaches to treat EBV-associated lymphomas that express LMP2A. Copyright © 2016 Elsevier Inc. All rights reserved.

  19. Novel small molecular inhibitors disrupt the JAK/STAT3 and FAK signaling pathways and exhibit a potent antitumor activity in glioma cells.

    PubMed

    Swiatek-Machado, Karolina; Mieczkowski, Jakub; Ellert-Miklaszewska, Aleksandra; Swierk, Piotr; Fokt, Izabela; Szymanski, Slawomir; Skora, Stanislaw; Szeja, Wiesław; Grynkiewicz, Grzegorz; Lesyng, Bogdan; Priebe, Waldemar; Kaminska, Bozena

    2012-06-01

    JAK (Janus kinase)/STAT (signal transducers and activators of transcription) signaling is involved in the regulation of cell growth, differentiation and apoptosis. Constitutive activation of STATs, in particular STAT3, is observed in a large number of human tumors, including gliomas and may contribute to oncogenesis by stimulating cell proliferation and preventing apoptosis, thus it emerges as a promising target for anti-cancer therapy. To investigate the therapeutic potential of blocking STAT3 in glioma cells a set of small synthetic molecules - caffeic acid derivatives, structurally related to AG490 was screened for its ability to inhibit STAT3. Inhibitor 2 (E)-2-cyano-N-[(S)-1-phenylethyl]-3-(pyridin-2-yl)acrylamide was the most effective in inhibition of JAK/STAT3 signaling and at doses ≥ 25 μM significantly reduced the level of phosphorylated JAK1, JAK2 and STAT3 (at Tyr705) and downregulated the expression of known STAT3 targets. In treated cells we observed rapid detachment and rounding of cells associated with reduction of focal adhesion kinase phosphorylation and activity, followed by upregulation of phosphorylated p38, JNK and ERK1/2 levels. Accumulation of cells with fragmented DNA, increases of the cleaved caspase 3 and fragmented PARP levels were detected 24 h after the treatment suggesting ongoing apoptotic cell death. Three human malignant glioblastoma cell lines defective in tumor suppressors TP53 and/or PTEN were susceptible to inhibitor 2 that induced the programmed cell death. Global gene expression profiling revealed modulation of numerous genes in cells treated with inhibitor 2 revealing novel, potential JAK/STAT targets. Our study demonstrates that suitably modified caffeic acid molecules exhibit significant cytotoxic potential toward glioma cells.

  20. A novel 7-bromoindirubin with potent anticancer activity suppresses survival of human melanoma cells associated with inhibition of STAT3 and Akt signaling.

    PubMed

    Liu, Lucy; Kritsanida, Marina; Magiatis, Prokopios; Gaboriaud, Nicolas; Wang, Yan; Wu, Jun; Buettner, Ralf; Yang, Fan; Nam, Sangkil; Skaltsounis, Leandros; Jove, Richard

    2012-11-01

    STAT3 and Akt signaling have been validated as potential molecular targets for treatment of cancers including melanoma. These small molecule inhibitors of STAT3 or Akt signaling are promising for developing anti-melanoma therapeutic agents. MLS-2438, a novel 7-bromoindirubin, a derivative of the natural product indirubin, was synthesized with a bromo-group at the 7-position on one indole ring and a hydrophilic group at the 3'-position on the other indole ring. We tested the anticancer activity of MLS-2438 and investigated its mechanism of action in human melanoma cell lines. Here, we show that MLS-2438 inhibits viability and induces apoptosis of human melanoma cells associated with inhibition of STAT3 and Akt signaling. Several pro-apoptotic Bcl-2 family proteins are involved in the MLS-2438 mediated apoptosis. MLS-2438 inhibits Src kinase activity in vitro and phosphorylation of JAK2, Src, STAT3 and Akt in cultured cancer cells. In contrast to the decreased phosphorylation levels of JAK2, Src, STAT3 and Akt, phosphorylation levels of the MAPK (Erk1/2) signaling protein were not reduced in cells treated with MLS-2438. These results demonstrate that MLS-2438, a novel natural product derivative, is a Src inhibitor and potentially regulates kinase activity of JAK2 and Akt in cancer cells. Importantly, MLS-2438 suppressed tumor growth with low toxicity in a mouse xenograft model of human melanoma. Our findings support further development of MLS-2438 as a potential small-molecule therapeutic agent that targets both STAT3 and Akt signaling in human melanoma cells.

  1. A novel 7-bromoindirubin with potent anticancer activity suppresses survival of human melanoma cells associated with inhibition of STAT3 and Akt signaling

    PubMed Central

    Liu, Lucy; Kritsanida, Marina; Magiatis, Prokopios; Gaboriaud, Nicolas; Wang, Yan; Wu, Jun; Buettner, Ralf; Yang, Fan; Nam, Sangkil; Skaltsounis, Leandros; Jove, Richard

    2012-01-01

    STAT3 and Akt signaling have been validated as potential molecular targets for treatment of cancers including melanoma. These small molecule inhibitors of STAT3 or Akt signaling are promising for developing anti-melanoma therapeutic agents. MLS-2438, a novel 7-bromoindirubin, a derivative of the natural product indirubin, was synthesized with a bromo-group at the 7-position on one indole ring and a hydrophilic group at the 3′-position on the other indole ring. We tested the anticancer activity of MLS-2438 and investigated its mechanism of action in human melanoma cell lines. Here, we show that MLS-2438 inhibits viability and induces apoptosis of human melanoma cells associated with inhibition of STAT3 and Akt signaling. Several pro-apoptotic Bcl-2 family proteins are involved in the MLS-2438 mediated apoptosis. MLS-2438 inhibits Src kinase activity in vitro and phosphorylation of JAK2, Src, STAT3 and Akt in cultured cancer cells. In contrast to the decreased phosphorylation levels of JAK2, Src, STAT3 and Akt, phosphorylation levels of the MAPK (Erk1/2) signaling protein were not reduced in cells treated with MLS-2438. These results demonstrate that MLS-2438, a novel natural product derivative, is a Src inhibitor and potentially regulates kinase activity of JAK2 and Akt in cancer cells. Importantly, MLS-2438 suppressed tumor growth with low toxicity in a mouse xenograft model of human melanoma. Our findings support further development of MLS-2438 as a potential small-molecule therapeutic agent that targets both STAT3 and Akt signaling in human melanoma cells. PMID:22895078

  2. Sevoflurane postconditioning protects the myocardium against ischemia/reperfusion injury via activation of the JAK2–STAT3 pathway

    PubMed Central

    Wu, Jianjiang; Yu, Jin; Xie, Peng; Maimaitili, Yiliyaer; Wang, Jiang; Yang, Long; Ma, Haiping; Zhang, Xing; Yang, Yining

    2017-01-01

    Background Sevoflurane postconditioning (S-post) has similar cardioprotective effects as ischemic preconditioning. However, the underlying mechanism of S-post has not been fully elucidated. Janus kinase signaling transduction/transcription activator (JAK2–STAT3) plays an important role in cardioprotection. The purpose of this study was to determine whether the cardioprotective effects of S-post are associated with activation of the JAK2–STAT3 signal pathway. Methods An adult male Sprague–Dawley (SD) rat model of myocardial ischemia/reperfusion (I/R) injury was established using the Langendorff isolated heart perfusion apparatus. At the beginning of reperfusion, 2.4% sevoflurane alone or in combination with AG490 (a JAK2 selective inhibitor) was used as a postconditioning treatment. The cardiac function indicators, myocardial infarct size, lactic dehydrogenase (LDH) release, mitochondrial ultrastructure, mitochondrial reactive oxygen species (ROS) generation rates, ATP content, protein expression of p-JAK, p-STAT3, Bcl-2 and Bax were measured. Results Compared with the I/R group, S-post significantly increased the expression of p-JAK, p-STAT3 and Bcl-2 and reduced the protein expression of Bax, which markedly decreased the myocardial infarction areas, improved the cardiac function indicators and the mitochondrial ultrastructure, decreased the mitochondrial ROS and increased the ATP content. However, the cardioprotective effects of S-post were abolished by treatment with a JAK2 selective inhibitor (p < 0.05). Conclusion This study demonstrates that the cardioprotective effects of S-post are associated with the activation of JAK2–STAT3. The mechanism may be related to an increased expression of p-JAK2 and p-STAT3 after S-post, which reduced mitochondrial ROS generation and increased mitochondrial ATP content, thereby reducing apoptosis and myocardial infarct size. PMID:28392989

  3. Qing Hua Chang Yin inhibits the LPS-induced activation of the IL-6/STAT3 signaling pathway in human intestinal Caco-2 cells.

    PubMed

    Ke, Xiao; Hu, Guanghong; Fang, Wenyi; Chen, Jintuan; Zhang, Xin; Yang, Chunbo; Peng, Jun; Chen, Youqin; Sferra, Thomas J

    2015-04-01

    Increasing evidence indicates that the pathogenesis of ulcerative colitis (UC) is highly regulated by the interleukin-6 (IL-6)/signal transducer and activator of transcription 3 (STAT3) pathway and its negative feedback regulator, suppressor of cytokine signaling 3 (SOCS3). Therefore, modulating the signaling feedback loop of IL-6/STAT3/SOCS3 may prove to be a novel therapeutic approach for the treatment of UC. Qing Hua Chang Yin (QHCY) is a traditional Chinese formulation that has long been used in clinic for the treatment of UC. We have previously reported that QHCY ameliorates acute intestinal inflammation in vivo and in vitro through the suppression of the nuclear factor-κB (NF-κB) pathway. In the present study, in order to further elucidate the mechanisms responsible for the anti-inflammatory activities of QHCY, we stimulated human intestinal Caco-2 cells with lipopolysaccharide (LPS) to create an in vitro model of an inflamed human intestinal epithelium, and evaluated the effects of QHCY on the IL-6/STAT3/SOCS3 signaling network in inflamed Caco-2 cells. The levels of IL-6 were measured by ELISA and the levels of STAT3 and SOCS3 were measured by western blot analysis. We found that QHCY significantly inhibited the LPS-induced secretion of pro-inflammatory IL-6 in the Caco-2 cells in a dose-dependent manner. Moreover, QHCY profoundly suppressed the LPS-induced phosphorylation of Janus-activated kinase 1 (JAK1), JAK2 and STAT3. Furthermore, treatment with QHCY markedly augmented the expression of SOCS3. Taken together, the findings of the present study suggest that the modulation of the IL-6/STAT3/SOCS3 signaling network may be one of the mechanisms through which QHCY exerts its anti-inflammatory effects.

  4. Peroxiredoxin 5 Protects TGF-β Induced Fibrosis by Inhibiting Stat3 Activation in Rat Kidney Interstitial Fibroblast Cells.

    PubMed

    Choi, Hoon-In; Ma, Seong Kwon; Bae, Eun Hui; Lee, JongUn; Kim, Soo Wan

    2016-01-01

    Renal fibrosis is a common final pathway of end-stage kidney disease which is induced by aberrant accumulation of myofibroblasts. This process is triggered by reactive oxygen species (ROS) and proinflammatory cytokines generated by various source of injured kidney cells. Peroxiredoxin 5 (Prdx5) is a thiol-dependent peroxidase that reduces oxidative stress by catalyzing intramolecular disulfide bonds. Along with its antioxidant effects, expression level of Prdx5 also was involved in inflammatory regulation by immune stimuli. However, the physiological effects and the underlying mechanisms of Prdx5 in renal fibrosis have not been fully characterized. Sprague-Dawley rats were subjected to unilateral ureteral obstruction (UUO) for 1 or 7 days. For the in vitro model, NRK49F cells, a rat kidney interstitial fibroblast cell lines, were treated with transforming growth factor β (TGF-β) for 0, 1, 3, or 5 days. To access the involvement of its peroxidase activity in TGF-β induced renal fibrosis, wild type Prdx5 (WT) and double mutant Prdx5 (DM), converted two active site cysteines at Cys 48 and Cys 152 residue to serine, were transiently expressed in NRK49F cells. The protein expression of Prdx5 was reduced in UUO kidneys. Upregulation of fibrotic markers, such as fibronectin and alpha-smooth muscle actin (α-SMA), declined at 5 days in time point of higher Prdx5 expression in TGF-β treated NRK49F cells. The overexpression of wild type Prdx5 by transient transfection in NRK49F cells attenuated the TGF-β induced upregulation of fibronectin and α-SMA. On the other hand, the transient transfection of double mutant Prdx5 did not prevent the activation of fibrotic markers. Overexpression of Prdx5 also suppressed the TGF-β induced upregulation of Stat3 phosphorylation, while phosphorylation of Smad 2/3 was unchanged. In conclusion, Prdx5 protects TGF-β induced fibrosis in NRK49F cells by modulating Stat3 activation in a peroxidase activity dependent manner.

  5. Peroxiredoxin 5 Protects TGF-β Induced Fibrosis by Inhibiting Stat3 Activation in Rat Kidney Interstitial Fibroblast Cells

    PubMed Central

    Choi, Hoon-In; Ma, Seong Kwon; Bae, Eun Hui; Lee, JongUn; Kim, Soo Wan

    2016-01-01

    Renal fibrosis is a common final pathway of end-stage kidney disease which is induced by aberrant accumulation of myofibroblasts. This process is triggered by reactive oxygen species (ROS) and proinflammatory cytokines generated by various source of injured kidney cells. Peroxiredoxin 5 (Prdx5) is a thiol-dependent peroxidase that reduces oxidative stress by catalyzing intramolecular disulfide bonds. Along with its antioxidant effects, expression level of Prdx5 also was involved in inflammatory regulation by immune stimuli. However, the physiological effects and the underlying mechanisms of Prdx5 in renal fibrosis have not been fully characterized. Sprague-Dawley rats were subjected to unilateral ureteral obstruction (UUO) for 1 or 7 days. For the in vitro model, NRK49F cells, a rat kidney interstitial fibroblast cell lines, were treated with transforming growth factor β (TGF-β) for 0, 1, 3, or 5 days. To access the involvement of its peroxidase activity in TGF-β induced renal fibrosis, wild type Prdx5 (WT) and double mutant Prdx5 (DM), converted two active site cysteines at Cys 48 and Cys 152 residue to serine, were transiently expressed in NRK49F cells. The protein expression of Prdx5 was reduced in UUO kidneys. Upregulation of fibrotic markers, such as fibronectin and alpha-smooth muscle actin (α-SMA), declined at 5 days in time point of higher Prdx5 expression in TGF-β treated NRK49F cells. The overexpression of wild type Prdx5 by transient transfection in NRK49F cells attenuated the TGF-β induced upregulation of fibronectin and α-SMA. On the other hand, the transient transfection of double mutant Prdx5 did not prevent the activation of fibrotic markers. Overexpression of Prdx5 also suppressed the TGF-β induced upregulation of Stat3 phosphorylation, while phosphorylation of Smad 2/3 was unchanged. In conclusion, Prdx5 protects TGF-β induced fibrosis in NRK49F cells by modulating Stat3 activation in a peroxidase activity dependent manner. PMID:26872211

  6. Granulin, a novel STAT3-interacting protein, enhances STAT3 transcriptional function and correlates with poorer prognosis in breast cancer

    PubMed Central

    Yeh, Jennifer E.; Kreimer, Simion; Walker, Sarah R.; Emori, Megan M.; Krystal, Hannah; Richardson, Andrea; Ivanov, Alexander R.; Frank, David A.

    2015-01-01

    Since the neoplastic phenotype of a cell is largely driven by aberrant gene expression patterns, increasing attention has been focused on transcription factors that regulate critical mediators of tumorigenesis such as signal transducer and activator of transcription 3 (STAT3). As proteins that interact with STAT3 may be key in addressing how STAT3 contributes to cancer pathogenesis, we took a proteomics approach to identify novel STAT3-interacting proteins. We performed mass spectrometry-based profiling of STAT3-containing complexes from breast cancer cells that have constitutively active STAT3 and are dependent on STAT3 function for survival. We identified granulin (GRN) as a novel STAT3-interacting protein that was necessary for both constitutive and maximal leukemia inhibitory factor (LIF)induced STAT3 transcriptional activity. GRN enhanced STAT3 DNA binding and also increased the time-integrated amount of LIF-induced STAT3 activation in breast cancer cells. Furthermore, silencing GRN neutralized STAT3-mediated tumorigenic phenotypes including viability, clonogenesis, and migratory capacity. In primary breast cancer samples, GRN mRNA levels were positively correlated with STAT3 gene expression signatures and with reduced patient survival. These studies identify GRN as a functionally important STAT3-interacting protein that may serve as an important prognostic biomarker and potential therapeutic target in breast cancer. PMID:26000098

  7. Early Activation of Rat Skeletal Muscle IL-6/STAT1/STAT3 Dependent Gene Expression in Resistance Exercise Linked to Hypertrophy

    PubMed Central

    Begue, Gwénaëlle; Douillard, Aymeric; Galbes, Olivier; Rossano, Bernadette; Vernus, Barbara; Candau, Robin; Py, Guillaume

    2013-01-01

    Cytokine interleukin-6 (IL-6) is an essential regulator of satellite cell-mediated hypertrophic muscle growth through the transcription factor signal transducer and activator of transcription 3 (STAT3). The importance of this pathway linked to the modulation of myogenic regulatory factors expression in rat skeletal muscle undergoing hypertrophy following resistance exercise, has not been investigated. In this study, the phosphorylation and nuclear localization of STAT3, together with IL-6/STAT3-responsive gene expression, were measured after both a single bout of resistance exercise and 10 weeks of training. Flexor Digitorum Profundus muscle samples from Wistar rats were obtained 2 and 6 hours after a single bout of resistance exercise and 72 h after the last bout of either 2, 4, or 10 weeks of resistance training. We observed an increase in IL-6 and SOCS3 mRNAs concomitant with phosphorylation of STAT1 and STAT3 after 2 and 6 hours of a single bout of exercise (p<0.05). STAT3-dependent early responsive genes such as CyclinD1 and cMyc were also upregulated whereas MyoD and Myf5 mRNAs were downregulated (p<0.05). BrdU-positive satellite cells increased at 2 and 6 hours after exercise (p<0.05). Muscle fiber hypertrophy reached up to 100% after 10 weeks of training and the mRNA expression of Myf5, c-Myc and Cyclin-D1 decreased, whereas IL-6 mRNA remained upregulated. We conclude that the IL-6/STAT1/STAT3 signaling pathway and its responsive genes after a single bout of resistance exercise are an important event regulating the SC pool and behavior involved in muscle hypertrophy after ten weeks of training in rat skeletal muscle. PMID:23451164

  8. MDM2 facilitates adipocyte differentiation through CRTC-mediated activation of STAT3

    PubMed Central

    Hallenborg, P; Siersbæk, M; Barrio-Hernandez, I; Nielsen, R; Kristiansen, K; Mandrup, S; Grøntved, L; Blagoev, B

    2016-01-01

    The ubiquitin ligase MDM2 is best known for balancing the activity of the tumor suppressor p53. We have previously shown that MDM2 is vital for adipocyte conversion through controlling Cebpd expression in a p53-independent manner. Here, we show that the proadipogenic effect of MDM2 relies on activation of the STAT family of transcription factors. Their activation was required for the cAMP-mediated induction of target genes. Interestingly, rather than influencing all cAMP-stimulated genes, inhibition of the kinases directly responsible for STAT activation, namely JAKs, or ablation of MDM2, each resulted in abolished induction of a subset of cAMP-stimulated genes, with Cebpd being among the most affected. Moreover, STATs were able to interact with the transcriptional cofactors CRTC2 and CRTC3, hitherto only reported to associate with the cAMP-responsive transcription factor CREB. Last but not least, the binding of CRTC2 to a transcriptional enhancer that interacts with the Cebpd promoter was dramatically decreased upon JAK inhibition. Our data reveal the existence of an unusual functional interplay between STATs and CREB at the onset of adipogenesis through shared CRTC cofactors. PMID:27362806

  9. Soluble egg antigens of Schistosoma japonicum induce senescence in activated hepatic stellate cells by activation of the STAT3/p53/p21 pathway

    PubMed Central

    Chen, Jinling; Pan, Jing; Wang, Jianxin; Song, Ke; Zhu, Dandan; Huang, Caiqun; Duan, Yinong

    2016-01-01

    Liver fibrosis is characterized by the activation of hepatic stellate cells (HSCs). Recent findings suggest that senescence of activated HSCs might limit the development of liver fibrosis. Based on previously observed anti-fibrotic effects of soluble egg antigens from Schistosoma japonicum in vitro, we hypothesized that SEA might play a crucial role in alleviating liver fibrosis through promoting senescence of activated HSCs. We show here that SEA inhibited expression of α-SMA and pro-collagen I and promoted senescence of activated HSCs in vitro. In addition, SEA induced an increased expression of P-p53 and p21. Knockdown of p53 inhibited the expression of p21 and failed to induce senescence of activated-HSCs. Phosphorylated STAT3 was elevated upon SEA stimulation, while loss of STAT3 decreased the level of p53 and senescence of HSCs. Results from immunoprecipitation analysis demonstrated that SOCS3 might be involved in the SEA-induced senescence in HSCs through its interaction with p53. This study demonstrates the potential capacity of SEA in restricting liver fibrosis through promoting senescence in HSCs. Furthermore, a novel STAT3-p53-p21 pathway might participate in the observed SEA-mediated senescence of HSCs. Our results suggest that SEA might carry potential therapeutic effects of restraining liver fibrosis through promoting senescence. PMID:27489164

  10. CD5 costimulation induces stable Th17 development by promoting IL-23R expression and sustained STAT3 activation.

    PubMed

    de Wit, Jelle; Souwer, Yuri; van Beelen, Astrid J; de Groot, Rosa; Muller, Femke J M; Klaasse Bos, Hanny; Jorritsma, Tineke; Kapsenberg, Martien L; de Jong, Esther C; van Ham, S Marieke

    2011-12-01

    IL-17-producing CD4(+) T helper (Th17) cells are important for immunity against extracellular pathogens and in autoimmune diseases. The factors that drive Th17 development in human remain a matter of debate. Here we show that, compared with classic CD28 costimulation, alternative costimulation via the CD5 or CD6 lymphocyte receptors forms a superior pathway for human Th17-priming. In the presence of the Th17-promoting cytokines IL-1β, IL-6, IL-23, and transforming growth factor-β (TGF-β), CD5 costimulation induces more Th17 cells that produce higher amounts of IL-17, which is preceded by prolonged activation of signal transducer and activator of transcription 3 (STAT3), a key regulator in Th17 differentiation, and enhanced levels of the IL-17-associated transcription factor retinoid-related orphan receptor-γt (ROR-γt). Strikingly, these Th17-promoting signals critically depend on CD5-induced elevation of IL-23 receptor (IL-23R) expression. The present data favor the novel concept that alternative costimulation via CD5, rather than classic costimulation via CD28, primes naive T cells for stable Th17 development through promoting the expression of IL-23R.

  11. TNF-α-inducing protein of Helicobacter pylori induces epithelial-mesenchymal transition (EMT) in gastric cancer cells through activation of IL-6/STAT3 signaling pathway.

    PubMed

    Chen, Guodong; Tang, Na; Wang, Chao; Xiao, Linqiao; Yu, Minjun; Zhao, Lanhua; Cai, Hengling; Han, Liang; Xie, Chengyuan; Zhang, Yan

    2017-03-04

    Tumor necrosis factor (TNF)-α-inducing protein (Tipα) is a newly identified carcinogenic factor secreted by Helicobacter pylori (H. pylori). Although it has been proved that Tipα is a strong inducer of epithelial-mesenchymal transition (EMT), a crucial process of migration, the exact molecular mechanism is unknown. Current evidence indicates that the oncogenic transcription factor signal transducers and activators of transcription 3 (STAT3) is inappropriately activated in multiple malignancies, including gastric cancer. In this study, we showed that Tipα significantly down-regulated the expression of EMT-related markers E-cadherin as well as up-regulated N-cadherin and vimentin in SGC7901 cells, with typical morphological changes of EMT. Tipα also promoted proliferation and migration of SGC7901 cells. Furthermore, Tipα activated interleukin-6 (IL-6)/STAT3 signaling pathway in SGC7901 cells. The effects of Tipα treatment observed was abolished when we block IL-6/STAT3 signaling pathway. Altogether, our data demonstrated that Tipα may accelerate tumor aggressiveness in gastric cancer by promoting EMT through activation of IL-6/STAT3 pathway.

  12. Salvianolic acids enhance cerebral angiogenesis and neurological recovery by activating JAK2/STAT3 signaling pathway after ischemic stroke in mice.

    PubMed

    Li, Yaoru; Zhang, Xiangjian; Cui, Lili; Chen, Rong; Zhang, Ye; Zhang, Cong; Zhu, Xingyuan; He, Tingting; Shen, Zuyuan; Dong, Lipeng; Zhao, Jingru; Wen, Ya; Zheng, Xiufen; Li, Pan

    2017-10-01

    Post-stroke angiogenesis facilitates neurovascular remodeling process and promotes neurological recovery. Proangiogenic effects of Salvianolic acids (Sals) have been reported in various ischemic disorders. However, the underlying mechanisms are still poorly understood. Previous studies of our laboratory have demonstrated that activating Janus kinase 2/signal transducer and activator of transcription 3 (JAK2/STAT3) signaling pathway is involved in the protection against cerebral ischemia/reperfusion injury. In this study, we investigated the impacts of Sals on angiogenesis and long-term neurological recovery after ischemic stroke as well as the potential mechanisms. Male mice subjected to permanent distal middle cerebral artery occlusion were administrated with Sals, 5-bromo-2'-deoxyuridine, and JAK2 inhibitor AG490 once daily from day 1 to day 14 after distal middle cerebral artery occlusion. Compared with the control group, Sals treatment significantly improved neurological recovery at day 14 and 28 after ischemic stroke. Sals enhanced post-stroke angiogenesis, pericytes and astrocytic endfeet covered ratio in the peri-infarct area. The JAK2/STAT3 signaling pathway was activated by Sals in the angiogenesis process, and inhibition of JAK2/STAT3 signaling blocked the effects of Sals on post-stroke angiogenesis and neurological recovery as well as abolished the mediation of proangiogenic factors. In summary, these data suggest that Sals administration enhances cerebral angiogenesis and promotes neurological recovery after ischemic stroke, mediated by the activation of JAK2/STAT3 signaling pathway. © 2017 International Society for Neurochemistry.

  13. Th17-type cytokines, IL-6 and TNF-α synergistically activate STAT3 and NF-kB to promote colorectal cancer cell growth.

    PubMed

    De Simone, V; Franzè, E; Ronchetti, G; Colantoni, A; Fantini, M C; Di Fusco, D; Sica, G S; Sileri, P; MacDonald, T T; Pallone, F; Monteleone, G; Stolfi, C

    2015-07-01

    Colorectal cancers (CRCs) often show a dense infiltrate of cytokine-producing immune/inflammatory cells. The exact contribution of each immune cell subset and cytokine in the activation of the intracellular pathways sustaining CRC cell growth is not understood. Herein, we isolate tumor-infiltrating leukocytes (TILs) and lamina propria mononuclear cells (LPMCs) from the tumor area and the macroscopically unaffected, adjacent, colonic mucosa of patients who underwent resection for sporadic CRC and show that the culture supernatants of TILs, but not of LPMCs, potently enhance the growth of human CRC cell lines through the activation of the oncogenic transcription factors signal transducer and activator of transcription 3 (STAT3) and nuclear factor-kappa B (NF-kB). Characterization of immune cell complexity of TILs and LPMCs reveals no differences in the percentages of T cells, natural killer T cells, natural killer (NK) cells, macrophages and B cells. However, T cells from TILs show a functional switch compared with those from LPMCs to produce large amounts of T helper type 17 (Th17)-related cytokines (that is, interleukin-17A (IL-17A), IL-17F, IL-21 and IL-22), tumor necrosis factor-α (TNF-α) and IL-6. Individual neutralization of IL-17A, IL-17F, IL-21, IL-22, TNF-α or IL-6 does not change TIL-derived supernatant-driven STAT3 and NF-kB activation, as well as their proproliferative effect in CRC cells. In contrast, simultaneous neutralization of both IL-17A and TNF-α, which abrogates NF-kB signaling, and IL-22 and IL-6, which abrogates STAT3 signaling, reduces the mitogenic effect of supernatants in CRC cells. IL-17A, IL-21, IL-22, TNF-α and IL-6 are also produced in excess in the early colonic lesions in a mouse model of sporadic CRC, associated with enhanced STAT3/NF-kB activation. Mice therapeutically given BP-1-102, an orally bioavailable compound targeting STAT3/NF-kB activation and cross-talk, exhibit reduced colon tumorigenesis and diminished expression of

  14. High density lipoprotein (HDL)-associated sphingosine 1-phosphate (S1P) inhibits macrophage apoptosis by stimulating STAT3 activity and survivin expression.

    PubMed

    Feuerborn, Renata; Becker, Susen; Potì, Francesco; Nagel, Petra; Brodde, Martin; Schmidt, Harmut; Christoffersen, Christina; Ceglarek, Uta; Burkhardt, Ralph; Nofer, Jerzy-Roch

    2017-02-01

    Macrophage apoptosis is critically involved in atherosclerosis. We here examined the effect of anti-atherogenic high density lipoprotein (HDL) and its component sphingosine-1-phosphate (S1P) on apoptosis in RAW264.7 murine macrophages. Mitochondrial or endoplasmic reticulum-dependent apoptosis was induced by exposure of macrophages to etoposide or thapsigargin/fukoidan, respectively. Cell death induced by these compounds was inhibited by S1P as inferred from reduced annexin V binding, TUNEL staining, and caspase 3, 9 and 12 activities. S1P induced expression of the inhibitor of apoptosis protein (IAP) family proteins cIAP1, cIAP2 and survivin, but only the inhibitor of survivin expression YM155 and not the cIAP1/2 blocker GDC0152 reversed the inhibitory effect of S1P on apoptosis. Moreover, S1P activated signal transducer and activator of transcription 3 (STAT3) and Janus kinase 2 (JAK2) and the stimulatory effect of S1P on survivin expression and inhibitory effects on apoptosis were attenuated by STAT3 or JAK2 inhibitors, S3I-201 or AG490, respectively. The effects of S1P on STAT3 activation, survivin expression and macrophage apoptosis were emulated by HDL, HDL lipids, and apolipoprotein (apo) M-containing HDL, but not by apoA-I or HDL deprived of S1P or apoM. In addition, JTE013 and CAY10444, S1P receptor 2 and 3 antagonists, respectively, compromised the S1P and HDL capacities to stimulate STAT3 activation and survivin expression, and to inhibit apoptosis. HDL-associated S1P inhibits macrophage apoptosis by stimulating STAT3 activity and survivin expression. The suppression of macrophage apoptosis may represent a novel mechanism utilized by HDL to exert its anti-atherogenic effects. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  15. The ETS factor ESE3/EHF represses IL-6 preventing STAT3 activation and expansion of the prostate cancer stem-like compartment

    PubMed Central

    Albino, Domenico; Civenni, Gianluca; Rossi, Simona; Mitra, Abhishek; Catapano, Carlo V.; Carbone, Giuseppina M.

    2016-01-01

    Metastatic prostate cancer represents a yet unsolved clinical problem due to the high frequency of relapse and treatment resistance. Understanding the pathways that lead to prostate cancer progression is an important task to prevent this deadly disease. The ETS transcription factor ESE3/EHF has an important role in differentiation of human prostate epithelial cells. Loss of ESE3/EHF in prostate epithelial cells determines transformation, epithelial-to-mesenchymal transition (EMT) and acquisition of stem-like properties. In this study we identify IL-6 as a direct target of ESE3/EHF that is activated in prostate epithelial cells upon loss of ESE3/EHF. ESE3/EHF and IL-6 were significantly inversely correlated in prostate tumors. Chromatin immunoprecipitation confirmed binding of ESE3/EHF to a novel ETS binding site in the IL-6 gene promoter. Inhibition of IL-6 reverted transformation and stem-like phenotype in tumorigenic ESE3/EHF knockdown prostate epithelial cell models. Conversely, IL-6 stimulation induced malignant phenotypes, stem-like behavior and STAT3 activation. Increased level of IL-6 was observed in prostatospheres compared with adherent bulk cancer cells and this was associated with stronger activation of STAT3. Human prostate tumors with IL-6 elevation and loss of ESE3/EHF were associated with STAT3 activation and displayed upregulation of genes related to cell adhesion, cancer stem-like and metastatic spread. Pharmacological inhibition of IL-6/STAT3 activation by a JAK inhibitor restrained cancer stem cell growth in vitro and inhibited self-renewal in vivo. This study identifies a novel connection between the transcription factor ESE3/EHF and the IL-6/JAK/STAT3 pathway and suggests that targeting this axis might be preferentially beneficial in tumors with loss of ESE3/EHF. PMID:27732936

  16. Characterization of phosphopeptide motifs specific for the Src homology 2 domains of signal transducer and activator of transcription 1 (STAT1) and STAT3.

    PubMed

    Wiederkehr-Adam, Michèle; Ernst, Philipp; Müller, Kurt; Bieck, Elke; Gombert, Frank O; Ottl, Johannes; Graff, Patrick; Grossmüller, Fred; Heim, Markus H

    2003-05-02

    Signal transducers and activators of transcription (STAT) 1 and STAT3 are activated by overlapping but distinct sets of cytokines. STATs are recruited to the different cytokine receptors through their Src homology (SH) 2 domains that make highly specific interactions with phosphotyrosine-docking sites on the receptors. We used a degenerate phosphopeptide library synthesized on 35-microm TentaGel beads and fluorescence-activated bead sorting to determine the sequence specificity of the peptide-binding sites of the SH2 domains of STAT1 and STAT3. The large bead library allowed not only peptide sequencing of pools of beads but also of single beads. The method was validated through surface plasmon resonance measurements of the affinities of different peptides to the STAT SH2 domains. Furthermore, when selected peptides were attached to a truncated erythropoietin receptor and stably expressed in DA3 cells, activation of STAT1 or STAT3 could be achieved by stimulation with erythropoietin. The combined analysis of pool sequencing, the individual peptide sequences, and plasmon resonance measurements allowed the definition of SH2 domain binding motifs. STAT1 preferentially binds peptides with the motif phosphotyrosine-(aspartic acid/glutamic acid)-(proline/arginine)-(arginine/proline/glutamine), whereby a negatively charged amino acid at +1 excludes a proline at +2 and vice versa. STAT3 preferentially binds peptides with the motif phosphotyrosine-(basic or hydrophobic)-(proline or basic)-glutamine. For both STAT1 and STAT3, specific high affinity phosphopeptides were identified that can be used for the design of inhibitory molecules.

  17. Bystander activity of Ad-mda7: human MDA-7 protein kills melanoma cells via an IL-20 receptor-dependent but STAT3-independent mechanism.

    PubMed

    Chada, Sunil; Mhashilkar, Abner M; Ramesh, Rajagopal; Mumm, John B; Sutton, R Bryan; Bocangel, Dora; Zheng, Mingzhong; Grimm, Elizabeth A; Ekmekcioglu, Suhendan

    2004-12-01

    The melanoma differentiation-associated gene-7 (mda-7/IL24) is a unique member of the IL-10 family of cytokines, with ubiquitous tumor cell proapoptotic activity. Transduction of tumor or normal cells with the mda-7 gene results in secretion of glycosylated MDA-7 protein. Recent data indicate that secreted MDA-7 protein functions as a pro-Th1 cytokine and as a potent antiangiogenic molecule. MDA-7 protein binds two distinct type II cytokine heterodimeric receptor complexes, IL-20R1/IL-20R2 (type 1 IL-20R) and IL-22R1/IL-20R2 (type 2 IL-20R). In this study we analyzed the activity of glycosylated secreted MDA-7 against human melanoma cells. MDA-7 protein induces phosphorylation and nuclear translocation of STAT3 in melanoma cells via both type 1 and type 2 IL-20R. MDA-7 induces dose-dependent cell death in melanoma tumor cells. MDA-7 receptor engagement results in up-regulation of BAX and subsequent apoptosis induction; this effect is mediated by STAT3-independent signaling. Additional IL-10 family members (IL-10, -19, -20, and -22) also activate STAT3; however, these ligands do not activate death pathways in melanoma. In normal cells, MDA-7 can bind to its cognate receptors and induce phosphorylation of STAT3, without cytotoxic sequelae. This study defines a tumor-selective cytotoxic bystander role for secreted MDA-7 protein and identifies a novel receptor-mediated, STAT3-independent, and PKR-independent death pathway.

  18. Photodynamic therapy activated signaling from epidermal growth factor receptor and STAT3: Targeting survival pathways to increase PDT efficacy in ovarian and lung cancer.

    PubMed

    Edmonds, Christine; Hagan, Sarah; Gallagher-Colombo, Shannon M; Busch, Theresa M; Cengel, Keith A

    2012-12-01

    Patients with serosal (pleural or peritoneal) spread of malignancy have few definitive treatment options and consequently have a very poor prognosis. We have previously shown that photodynamic therapy (PDT) can be an effective treatment for these patients, but that the therapeutic index is relatively narrow. Here, we test the hypothesis that EGFR and STAT3 activation increase survival following PDT, and that inhibiting these pathways leads to increased PDT-mediated direct cellular cytotoxicity by examining BPD-PDT in OvCa and NSCLC cells. We found that BPD-mediated PDT stimulated EGFR tyrosine phosphorylation and nuclear translocation, and that EGFR inhibition by erlotinib resulted in reduction of PDT-mediated EGFR activation and nuclear translocation. Nuclear translocation and PDT-mediated activation of EGFR were also observed in response to BPD-mediated PDT in multiple cell lines, including OvCa, NSCLC and head and neck cancer cells, and was observed to occur in response to porfimer sodium-mediated PDT. In addition, we found that PDT stimulates nuclear translocation of STAT3 and STAT3/EGFR association and that inhibiting STAT3 signaling prior to PDT leads to increased PDT cytotoxicity. Finally, we found that inhibition of EGFR signaling leads to increased PDT cytotoxicity through a mechanism that involves increased apoptotic cell death. Taken together, these results demonstrate that PDT stimulates the nuclear accumulation of both EGFR and STAT3 and that targeting these survival pathways is a potentially promising strategy that could be adapted for clinical trials of PDT for patients with serosal spread of malignancy.

  19. Leptin-induced Growth Stimulation of Breast Cancer Cells Involves Recruitment of Histone Acetyltransferases and Mediator Complex to CYCLIN D1 Promoter via Activation of Stat3*

    PubMed Central

    Saxena, Neeraj K.; Vertino, Paula M.; Anania, Frank A.; Sharma, Dipali

    2010-01-01

    Numerous epidemiological studies documented that obesity is a risk factor for breast cancer development in postmenopausal women. Leptin, the key player in the regulation of energy balance and body weight control also acts as a growth factor on certain organs in both normal and disease state. In this study, we analyzed the role of leptin and the molecular mechanism(s) underlying its action in breast cancer cells that express both short and long isoforms of leptin receptor. Leptin increased MCF7 cell population in the S-phase of the cell cycle along with a robust increase in CYCLIN D1 expression. Also, leptin induced Stat3-phosphorylation-dependent proliferation of MCF7 cells as blocking Stat3 phosphorylation with a specific inhibitor, AG490, abolished leptin-induced proliferation. Using deletion constructs of CYCLIN D1 promoter and chromatin immunoprecipitation assay, we show that leptin induced increase in CYCLIN D1 promoter activity is mediated through binding of activated Stat3 at the Stat binding sites and changes in histone acetylation and methylation. We also show specific involvement of coactivator molecules, histone acetyltransferase SRC1, and mediator complex in leptin-mediated regulation of CYCLIN D1 promoter. Importantly, silencing of SRC1 and Med1 abolished the leptin induced increase in CYCLIN D1 expression and MCF7 cell proliferation. Intriguingly, recruitment of both SRC1 and Med1 was dependent on phosphorylated Stat3 as AG490 treatment inhibited leptin-induced recruitment of these coactivators to CYCLIN D1 promoter. Our data suggest that CYCLIN D1 may be a target gene for leptin mediated growth stimulation of breast cancer cells and molecular mechanisms involve activated Stat3-mediated recruitment of distinct coactivator complexes. PMID:17344214

  20. JAK1/STAT3 Activation through a Proinflammatory Cytokine Pathway Leads to Resistance to Molecularly Targeted Therapy in Non-Small Cell Lung Cancer.

    PubMed

    Shien, Kazuhiko; Papadimitrakopoulou, Vassiliki A; Ruder, Dennis; Behrens, Carmen; Shen, Li; Kalhor, Neda; Song, Juhee; Lee, J Jack; Wang, Jing; Tang, Ximing; Herbst, Roy S; Toyooka, Shinichi; Girard, Luc; Minna, John D; Kurie, Jonathan M; Wistuba, Ignacio I; Izzo, Julie G

    2017-10-01

    Molecularly targeted drugs have yielded significant therapeutic advances in oncogene-driven non-small cell lung cancer (NSCLC), but a majority of patients eventually develop acquired resistance. Recently, the relation between proinflammatory cytokine IL6 and resistance to targeted drugs has been reported. We investigated the functional contribution of IL6 and the other members of IL6 family proinflammatory cytokine pathway to resistance to targeted drugs in NSCLC cells. In addition, we examined the production of these cytokines by cancer cells and cancer-associated fibroblasts (CAF). We also analyzed the prognostic significance of these molecule expressions in clinical NSCLC samples. In NSCLC cells with acquired resistance to targeted drugs, we observed activation of the IL6-cytokine pathway and STAT3 along with epithelial-to-mesenchymal transition (EMT) features. In particular, IL6 family cytokine oncostatin-M (OSM) induced a switch to the EMT phenotype and protected cells from targeted drug-induced apoptosis in OSM receptors (OSMRs)/JAK1/STAT3-dependent manner. The cross-talk between NSCLC cells and CAFs also preferentially activated the OSM/STAT3 pathway via a paracrine mechanism and decreased sensitivity to targeted drugs. The selective JAK1 inhibitor filgotinib effectively suppressed STAT3 activation and OSMR expression, and cotargeting inhibition of the oncogenic pathway and JAK1 reversed resistance to targeted drugs. In the analysis of clinical samples, OSMR gene expression appeared to be associated with worse prognosis in patients with surgically resected lung adenocarcinoma. Our data suggest that the OSMRs/JAK1/STAT3 axis contributes to resistance to targeted drugs in oncogene-driven NSCLC cells, implying that this pathway could be a therapeutic target. Mol Cancer Ther; 16(10); 2234-45. ©2017 AACR. ©2017 American Association for Cancer Research.

  1. Leptin receptor expressing neurons express phosphodiesterase-3B (PDE3B) and leptin induces STAT3 activation in PDE3B neurons in the mouse hypothalamus

    PubMed Central

    Sahu, Maitrayee; Sahu, Abhiram

    2015-01-01

    Leptin signaling in the hypothalamus is critical for normal food intake and body weight regulation. Cumulative evidence suggests that besides the signal transducer and activator of transcription-3 (STAT3) pathway, several non-STAT3 pathways including the phosphodiesterase-3B (PDE3B) pathway mediate leptin signaling in the hypothalamus. We have shown that PDE3B is localized in various hypothalamic sites implicated in the regulation of energy homeostasis and that the anorectic and body weight reducing effects of leptin are mediated by the activation of PDE3B. It is still unknown if PDE3B is expressed in the long form of the leptin-receptor (ObRb)-expressing neurons in the hypothalamus and whether leptin induces STAT3 activation in PDE3B-expressing neurons. In this study, we examined co-localization of PDE3B with ObRb neurons in various hypothalamic nuclei in ObRb-GFP mice that were treated with leptin (5mg/kg, ip) for 2 hr. Results showed that most of the ObRb neurons in the arcuate nucleus (ARC, 93%), ventromedial nucleus (VMN, 94%), dorsomedial nucleus (DMN, 95%), ventral premammillary nucleus (PMv, 97%) and lateral hypothalamus (LH, 97%) co-expressed PDE3B. We next examined co-localization of p-STAT3 and PDE3B in the hypothalamus in C57BL6 mice that were treated with leptin (5mg/kg, ip) for 1 hr. The results showed that almost all p-STAT3 positive neurons in different hypothalamic nuclei including ARC, VMN, DMN, LH and PMv areas expressed PDE3B. These results suggest the possibility for a direct role for the PDE3B pathway in mediating leptin action in the hypothalamus. PMID:26297880

  2. Leptin receptor expressing neurons express phosphodiesterase-3B (PDE3B) and leptin induces STAT3 activation in PDE3B neurons in the mouse hypothalamus.

    PubMed

    Sahu, Maitrayee; Sahu, Abhiram

    2015-11-01

    Leptin signaling in the hypothalamus is critical for normal food intake and body weight regulation. Cumulative evidence suggests that besides the signal transducer and activator of transcription-3 (STAT3) pathway, several non-STAT3 pathways including the phosphodiesterase-3B (PDE3B) pathway mediate leptin signaling in the hypothalamus. We have shown that PDE3B is localized in various hypothalamic sites implicated in the regulation of energy homeostasis and that the anorectic and body weight reducing effects of leptin are mediated by the activation of PDE3B. It is still unknown if PDE3B is expressed in the long form of the leptin-receptor (ObRb)-expressing neurons in the hypothalamus and whether leptin induces STAT3 activation in PDE3B-expressing neurons. In this study, we examined co-localization of PDE3B with ObRb neurons in various hypothalamic nuclei in ObRb-GFP mice that were treated with leptin (5mg/kg, ip) for 2h. Results showed that most of the ObRb neurons in the arcuate nucleus (ARC, 93%), ventromedial nucleus (VMN, 94%), dorsomedial nucleus (DMN, 95%), ventral premammillary nucleus (PMv, 97%) and lateral hypothalamus (LH, 97%) co-expressed PDE3B. We next examined co-localization of p-STAT3 and PDE3B in the hypothalamus in C57BL6 mice that were treated with leptin (5mg/kg, ip) for 1h. The results showed that almost all p-STAT3 positive neurons in different hypothalamic nuclei including ARC, VMN, DMN, LH and PMv areas expressed PDE3B. These results suggest the possibility for a direct role for the PDE3B pathway in mediating leptin action in the hypothalamus. Copyright © 2015 Elsevier Inc. All rights reserved.

  3. p53 induces miR199a-3p to suppress SOCS7 for STAT3 activation and renal fibrosis in UUO

    PubMed Central

    Yang, Ruhao; Xu, Xuan; Li, Huiling; Chen, Jinwen; Xiang, Xudong; Dong, Zheng; Zhang, Dongshan

    2017-01-01

    The role of p53 in renal fibrosis has recently been suggested, however, its function remains controversial and the underlying mechanism is unclear. Here, we show that pharmacological and genetic blockade of p53 attenuated renal interstitial fibrosis, apoptosis, and inflammation in mice with unilateral urethral obstruction (UUO). Interestingly, p53 blockade was associated with the suppression of miR-215-5p, miR-199a-5p&3p, and STAT3. In cultured human kidney tubular epithelial cells (HK-2), TGF-β1 treatment induced fibrotic changes, including collagen I and vimentin expression, being associated with p53 accumulation, p53 Ser15 phosphorylation, and miR-199a-3p expression. Inhibition of p53 by pifithrin-α blocked STAT3 activation and the expression of miR-199a-3p, collagen I, and vimentin during TGF-β1 treatment. Over-expression of miR-199a-3p increased TGFβ1-induced collagen I and vimentin expression and restored SOCS7 expression. Furthermore, SOCS7 was identified as a target gene of miR-199a-3p, and silencing of SOCS7 promoted STAT3 activation. ChIp analyses indicated the binding of p53 to the promoter region of miR-199a-3p. Consistently, kidney biopsies from patients with IgA nephropathy and diabetic nephropathy exhibited substantial activation of p53 and STAT3, decreased expression of SOCS7, and increase in profibrotic proteins and miR-199a-3p. Together, these results demonstrate the novel p53/miR-199a-3p/SOCS7/STAT3 pathway in renal interstitial fibrosis. PMID:28240316

  4. Resistin and interleukin-6 exhibit racially-disparate expression in breast cancer patients, display molecular association and promote growth and aggressiveness of tumor cells through STAT3 activation

    PubMed Central

    Bhardwaj, Arun; Singh, Ajay P.; Tyagi, Nikhil; Marimuthu, Saravanakumar; Dyess, Donna L.; Zotto, Valeria Dal; Carter, James E.; Singh, Seema

    2015-01-01

    African-American (AA) women with breast cancer (BC) are diagnosed with more aggressive disease, have higher risk of recurrence and poorer prognosis as compared to Caucasian American (CA) women. Therefore, it is imperative to define the factors associated with such disparities to reduce the unequal burden of cancer. Emerging data suggest that inherent differences exist in the tumor microenvironment of AA and CA BC patients, however, its molecular bases and functional impact have remained poorly understood. Here, we conducted cytokine profiling in serum samples from AA and CA BC patients and identified resistin and IL-6 to be the most differentially-expressed cytokines with relative greater expression in AA patients. Resistin and IL-6 exhibited positive correlation in serum levels and treatment of BC cells with resistin led to enhanced production of IL-6. Moreover, resistin also enhanced the expression and phosphorylation of STAT3, and treatment of BC cells with IL-6-neutralizing antibody prior to resistin stimulation abolished STAT3 phosphorylation. In addition, resistin promoted growth and aggressiveness of BC cells, and these effects were mediated through STAT3 activation. Together, these findings suggest a crucial role of resistin, IL-6 and STAT3 in BC racial disparity. PMID:25868978

  5. Resistin and interleukin-6 exhibit racially-disparate expression in breast cancer patients, display molecular association and promote growth and aggressiveness of tumor cells through STAT3 activation.

    PubMed

    Deshmukh, Sachin K; Srivastava, Sanjeev K; Bhardwaj, Arun; Singh, Ajay P; Tyagi, Nikhil; Marimuthu, Saravanakumar; Dyess, Donna L; Dal Zotto, Valeria; Carter, James E; Singh, Seema

    2015-05-10

    African-American (AA) women with breast cancer (BC) are diagnosed with more aggressive disease, have higher risk of recurrence and poorer prognosis as compared to Caucasian American (CA) women. Therefore, it is imperative to define the factors associated with such disparities to reduce the unequal burden of cancer. Emerging data suggest that inherent differences exist in the tumor microenvironment of AA and CA BC patients, however, its molecular bases and functional impact have remained poorly understood. Here, we conducted cytokine profiling in serum samples from AA and CA BC patients and identified resistin and IL-6 to be the most differentially-expressed cytokines with relative greater expression in AA patients. Resistin and IL-6 exhibited positive correlation in serum levels and treatment of BC cells with resistin led to enhanced production of IL-6. Moreover, resistin also enhanced the expression and phosphorylation of STAT3, and treatment of BC cells with IL-6-neutralizing antibody prior to resistin stimulation abolished STAT3 phosphorylation. In addition, resistin promoted growth and aggressiveness of BC cells, and these effects were mediated through STAT3 activation. Together, these findings suggest a crucial role of resistin, IL-6 and STAT3 in BC racial disparity.

  6. Entinostat up-regulates the CAMP gene encoding LL-37 via activation of STAT3 and HIF-1α transcription factors

    PubMed Central

    Miraglia, Erica; Nylén, Frank; Johansson, Katarina; Arnér, Elias; Cebula, Marcus; Farmand, Susan; Ottosson, Håkan; Strömberg, Roger; Gudmundsson, Gudmundur H.; Agerberth, Birgitta; Bergman, Peter

    2016-01-01

    Bacterial resistance against classical antibiotics is a growing problem and the development of new antibiotics is limited. Thus, novel alternatives to antibiotics are warranted. Antimicrobial peptides (AMPs) are effector molecules of innate immunity that can be induced by several compounds, including vitamin D and phenyl-butyrate (PBA). Utilizing a luciferase based assay, we recently discovered that the histone deacetylase inhibitor Entinostat is a potent inducer of the CAMP gene encoding the human cathelicidin LL-37. Here we investigate a mechanism for the induction and also find that Entinostat up-regulates human β-defensin 1. Analysis of the CAMP promoter sequence revealed binding sites for the transcription factors STAT3 and HIF-1α. By using short hairpin RNA and selective inhibitors, we found that both transcription factors are involved in Entinostat-induced expression of LL-37. However, only HIF-1α was found to be recruited to the CAMP promoter, suggesting that Entinostat activates STAT3, which promotes transcription of CAMP by increasing the expression of HIF-1α. Finally, we provide in vivo relevance to our findings by showing that Entinostat-elicited LL-37 expression was impaired in macrophages from a patient with a STAT3-mutation. Combined, our findings support a role for STAT3 and HIF-1α in the regulation of LL-37 expression. PMID:27633343

  7. Interleukin-27 and interleukin-12 augment activation of distinct cord blood natural killer cells responses via STAT3 pathways.

    PubMed

    Chen, Juei-Chang; Huang, Ai-Ju; Chen, Shih-Chang; Wu, Jia-Long; Wu, Wen-Mein; Chiang, Han-Sun; Chan, Chia-Hao; Lin, Chih-Ming; Huang, Yu-Tzu

    2012-05-01

    Umbilical cord blood is rich in primitive natural killer (NK) cells, which are activated by interleukin (IL)-12. It was previously reported that a novel IL-12 family cytokine, IL-27 comprised of EBI3 and p28, was elevated in maternal serum during normal pregnancy. Thus, we compared the immune regulatory functions of IL-27 and IL-12 on mononuclear cells derived from cord blood and adult peripheral blood. After stimulation with IL-27, IL-12, and IL-27 combined with IL-12, the cytotoxicity against BJAB lymphoma cells by blood mononuclear cells was performed. Then immunofluorescence staining, reverse transcriptase-polymerase chain reaction and Western blotting were used to detect the effects of IL-27 and IL-12 in isolated NK cells. IL-27, IL-12, and IL-27 combined with IL-12 enhanced the cytotoxicity of adult peripheral blood cells and cord blood cells, but the proliferation of distinct subpopulations of cells was not evident. Similar results were also obtained with purified cord blood NK cells. Interestingly, distinct from IL-12, IL-27 could induce aggregation and morphological changes of umbilical cord blood cells. Finally, IL-27 combined with IL-12 could stimulate increased IL-27 receptor (gp130 and WSX-1) transcripts in purified cord blood NK cells. However, the activation of signal transducer and activator of transcription 3 (STAT3) in NK cells was only detected in the presence of IL-27, but not IL-12 alone. From previous results, we summarize our current understanding of the augmentation of distinct regulation of NK cells by IL-27 and IL-12. Copyright © 2012. Published by Elsevier B.V.

  8. IL-23R–activated STAT3/STAT4 is essential for Th1/Th17-mediated CNS autoimmunity

    PubMed Central

    Lee, Priscilla W.; Smith, Alan J.; Yang, Yuhong; Selhorst, Amanda J.; Liu, Yue; Racke, Michael K.; Lovett-Racke, Amy E.

    2017-01-01

    The factors that promote the differentiation of pathogenic T cells in autoimmune diseases are poorly defined. Use of genetically modified mice has provided insight into molecules necessary for the development of autoimmunity, but the sum of the data has led to contradictory observations based on what is currently known about specific molecules in specific signaling pathways. To define the minimum signals required for development of encephalitogenic T cells that cause CNS autoimmunity, myelin-specific T cells were differentiated with various cytokine cocktails, and pathogenicity was determined by transfer into mice. IL-6+IL-23 or IL-12+IL-23 generated encephalitogenic T cells and recapitulated the essential cytokine signals provided by antigen-presenting cells, and both IL-6 and IL-12 induced IL-23 receptor expression on both mouse and human naive T cells. IL-23 signaled through both STAT3 and STAT4, and disruption in STAT4 signaling impaired CNS autoimmunity independent of IL-12. These data explain why IL-12–deficient mice develop CNS autoimmunity, while STAT4-deficient mice are resistant. CD4+ memory T cells from multiple sclerosis patients had significantly higher levels of p-STAT3/p-STAT4, and p-STAT3/p-STAT4 heterodimers were observed upon IL-23 signaling, suggesting that p-STAT3/p-STAT4 induced by IL-23 signaling orchestrate the generation of pathogenic T cells in CNS autoimmunity, regardless of Th1 or Th17 phenotype. PMID:28878115

  9. IL-23R-activated STAT3/STAT4 is essential for Th1/Th17-mediated CNS autoimmunity.

    PubMed

    Lee, Priscilla W; Smith, Alan J; Yang, Yuhong; Selhorst, Amanda J; Liu, Yue; Racke, Michael K; Lovett-Racke, Amy E

    2017-09-07

    The factors that promote the differentiation of pathogenic T cells in autoimmune diseases are poorly defined. Use of genetically modified mice has provided insight into molecules necessary for the development of autoimmunity, but the sum of the data has led to contradictory observations based on what is currently known about specific molecules in specific signaling pathways. To define the minimum signals required for development of encephalitogenic T cells that cause CNS autoimmunity, myelin-specific T cells were differentiated with various cytokine cocktails, and pathogenicity was determined by transfer into mice. IL-6+IL-23 or IL-12+IL-23 generated encephalitogenic T cells and recapitulated the essential cytokine signals provided by antigen-presenting cells, and both IL-6 and IL-12 induced IL-23 receptor expression on both mouse and human naive T cells. IL-23 signaled through both STAT3 and STAT4, and disruption in STAT4 signaling impaired CNS autoimmunity independent of IL-12. These data explain why IL-12-deficient mice develop CNS autoimmunity, while STAT4-deficient mice are resistant. CD4+ memory T cells from multiple sclerosis patients had significantly higher levels of p-STAT3/p-STAT4, and p-STAT3/p-STAT4 heterodimers were observed upon IL-23 signaling, suggesting that p-STAT3/p-STAT4 induced by IL-23 signaling orchestrate the generation of pathogenic T cells in CNS autoimmunity, regardless of Th1 or Th17 phenotype.

  10. SH003 reverses drug resistance by blocking signal transducer and activator of transcription 3 (STAT3) signaling in breast cancer cells.

    PubMed

    Seo, Hye-Sook; Ku, Jin Mo; Lee, Hee-Jae; Woo, Jong-Kyu; Cheon, Chunhoo; Kim, Mia; Jang, Bo-Hyoung; Shin, Yong Cheol; Ko, Seong-Gyu

    2017-09-01

    Overcoming drug resistance is an important task for investigators and clinician to achieve successful chemotherapy in cancer patients. Drug resistance is caused by various factors, including the overexpression of P-glycoprotein (P-gp, MDR1). The development of new, useful compounds that overcome drug resistance is urgent. SH003 is extracted from the mixture of three different herbs, and its anti-cancer effect has been revealed in different cancer cell types. In the present study, we investigated whether SH003 is able to reverse drug resistance using paclitaxel-resistant breast cancer cells (MCF-7/PAC). In our experiments, SH003 significantly decreased cell growth and colony formation in MCF-7/PAC cells and parental MCF-7 cells. This growth inhibition was related to the accumulation of cells in the sub-G0/G1 apoptotic population and an increase in the number of apoptotic cells. SH003 reduced the mRNA expression of multi-drug resistance 1 (MDR1) and multi-drug resistance-associated proteins (MRPs) in MCF-7/PAC cells. SH003 also down-regulated the expression of P-gp. SH003 reversed drug efflux from MCF-7/PAC cells, resulting in rhodamine123 (Rho123) accumulation. Inhibition of drug resistance by SH003 is related to the suppression of the STAT3 signaling pathway. SH003 decreased STAT3 activation (p-STAT3) and its nuclear translocation and inhibited the secretion of VEGF and MMP-2, which are STAT3 target genes. A STAT3 inhibitor, JAK inhibitor I and an HIF-1a inhibitor decreased cell growth in MCF-7 and MCF-7/PAC cells. Taken together, these results demonstrate that SH003 can overcome drug resistance, and SH003 might be helpful for chemotherapy in cancer patients. ©2017 The Author(s).

  11. Acrp30 inhibits leptin-induced metastasis by downregulating the JAK/STAT3 pathway via AMPK activation in aggressive SPEC-2 endometrial cancer cells.

    PubMed

    Wu, Xiaomei; Yan, Qin; Zhang, Zhenbo; Du, Guiqiang; Wan, Xiaoping

    2012-05-01

    Obesity is a well-established risk factor for endometrial cancer, due in part to the adipokines generated by adipose tissue, such as adiponectin (also known as Acrp30) and leptin, which are associated with many endocrine-related cancers. Recent reports suggested that Acrp30 inhibits leptin-stimulated cell proliferation in HEC-1A and RL95-2 endometrial cancer cell lines, and that the serum leptin/Acrp30 ratio plays an important role in endometrial cancer development. We explored whether Acrp30 could reverse the leptin-induced metastasis phenotype in the SPEC-2 endometrial cancer cell line. Transcripts for Acrp30 receptors (AdipoR1 and AdipoR2) and leptin receptor (Ob-Rb) were detected by quantitative real-time RT-PCR (qRT-PCR) in six endometrial cancer cell lines. Leptin (1 µg/ml) treatment stimulated SPEC-2 cell proliferation by inducing cell cycle arrest and apoptosis, while Acrp30 (10 µg/ml) treatment inhibited the growth of SPEC-2 cells. Importantly, Acrp30 was able to inhibit leptin-induced SPEC-2 cell proliferation. Leptin promoted SPEC-2 cell invasion in a Matrigel transwell assay, while Acrp30 partly suppressed the invasion stimulated by leptin. To investigate the molecular mechanism underlying this phenomenon, we monitored the AMPK and JAK/STAT3 signaling pathways by western blotting and cell immunofluorescence. Acrp30 reduced leptin-induced STAT3 phosphorylation and nuclear translocation via activation of the MAPK pathway. AG490 (JAK/STAT3 inhibitor) reduced MMP-2 and MMP-9 protein levels in cells treated with leptin, while IL-6 (JAK/STAT3 stimulator) and Compound C (AMPK inhibitor) elevated MMP-2 and MMP-9 protein levels in cells treated with Acrp30. In conclusion, we demonstrated that Acrp30 effectively reversed the invasion stimulated by leptin, and AMPK and JAK/STAT3 pathways mediated the invasive phenotype of SPEC-2 cells.

  12. NTRK2 activation cooperates with PTEN deficiency in T-ALL through activation of both the PI3K–AKT and JAK–STAT3 pathways

    PubMed Central

    Yuzugullu, Haluk; Von, Thanh; Thorpe, Lauren M; Walker, Sarah R; Roberts, Thomas M; Frank, David A; Zhao, Jean J

    2016-01-01

    Loss of PTEN, a negative regulator of the phosphoinositide 3-kinase signaling pathway, is a frequent event in T-cell acute lymphoblastic leukemia, suggesting the importance of phosphoinositide 3-kinase activity in this disease. Indeed, hyperactivation of the phosphoinositide 3-kinase pathway is associated with the disease aggressiveness, poor prognosis and resistance to current therapies. To identify a molecular pathway capable of cooperating with PTEN deficiency to drive oncogenic transformation of leukocytes, we performed an unbiased transformation screen with a library of tyrosine kinases. We found that activation of NTRK2 is able to confer a full growth phenotype of Ba/F3 cells in an IL3-independent manner in the PTEN-null setting. NTRK2 activation cooperates with PTEN deficiency through engaging both phosphoinositide3-kinase/AKT and JAK/STAT3 pathway activation in leukocytes. Notably, pharmacological inhibition demonstrated that p110α and p110δ are the major isoforms mediating the phosphoinositide 3-kinase/AKT signaling driven by NTRK2 activation in PTEN-deficient leukemia cells. Furthermore, combined inhibition of phosphoinositide 3-kinase and STAT3 significantly suppressed proliferation of PTEN-mutant T-cell acute lymphoblastic leukemia both in culture and in mouse xenografts. Together, our data suggest that a unique conjunction of PTEN deficiency and NTRK2 activation in T-cell acute lymphoblastic leukemia, and combined pharmacologic inhibition of phosphoinositide 3-kinase and STAT3 signaling may serve as an effective and durable therapeutic strategy for T-cell acute lymphoblastic leukemia. PMID:27672444

  13. Flavanoids induce expression of the suppressor of cytokine signalling 3 (SOCS3) gene and suppress IL-6-activated signal transducer and activator of transcription 3 (STAT3) activation in vascular endothelial cells.

    PubMed

    Wiejak, Jolanta; Dunlop, Julia; Mackay, Simon P; Yarwood, Stephen J

    2013-09-01

    The atherogenic cytokine IL-6 (interleukin-6) induces pro-inflammatory gene expression in VECs (vascular endothelial cells) by activating the JAK (Janus kinase)/STAT3 (signal transducer and activator of transcription 3) signalling pathway, which is normally down-regulated by the STAT3-dependent induction of the E3 ubiquitin ligase component SOCS3 (suppressor of cytokine signalling 3). Novel treatments based on the regulation of SOCS3 protein levels could therefore have value in the treatment of diseases with an inflammatory component, such as atherosclerosis. To this end we carried out a screen of 1031 existing medicinal compounds to identify inducers of SOCS3 gene expression and identified the flavanoids naringenin and flavone as effective inducers of SOCS3 protein, mRNA and promoter activity. This was in contrast with the action of traditional JAK/STAT3 inhibitors and the polyphenol resveratrol, which effectively suppress SOCS3 gene expression. Both naringenin and flavone also effectively suppressed IL-6-stimulated phosphorylation of STAT3 (Tyr⁷⁰⁵) which led to suppression of IL-6-induction of the atherogenic STAT3 target gene MCP1 (monocyte chemotactic protein-1), suggesting that their ability to induce SOCS3 gene expression is STAT3-independent. Supporting this idea was the observation that the general kinase inhibitor compound C inhibits flavone- and cAMP-dependent, but not JAK-dependent, SOCS3 induction in VECs. Indeed, the ability of flavanoids to induce SOCS3 expression requires activation of the ERK (extracellular-signal-regulated kinase)-dependent transcription factor SP3, and not STAT3. In the present paper we therefore describe novel molecular actions of flavanoids, which control SOCS3 gene induction and suppression of STAT3 signalling in VECs. These mechanisms could potentially be exploited to develop novel anti-atherogenic therapies.

  14. Serum amyloid A3 exacerbates cancer by enhancing the suppressive capacity of myeloid-derived suppressor cells via TLR2-dependent STAT3 activation.

    PubMed

    Lee, Jung-Mi; Kim, Eun-Kyung; Seo, Hyungseok; Jeon, Insu; Chae, Min-Ji; Park, Young-Jun; Song, Boyeong; Kim, Yun-Sun; Kim, Yeon-Jeong; Ko, Hyun-Jeong; Kang, Chang-Yuil

    2014-06-01

    Myeloid-derived suppressor cells (MDSCs), which suppress diverse innate and adaptive immune responses and thereby provide an evasion mechanism for tumors, are emerging as a key population linking inflammation to cancer. Although many inflammatory factors that induce MDSCs in the tumor microenvironment are known, the crucial components and the underlying mechanisms remain elusive. In this study, we proposed a novel mechanism by which serum amyloid A3 (SAA3), a well-known inflammatory factor, connects MDSCs with cancer progression. We found that SAA3 expression in BALB/c mice increased in monocytic MDSCs (Mo MDSCs) with tumor growth. The induction of SAA3 by apo-SAA treatment in Mo MDSCs enhanced their survival and suppressive activity, while it inhibited GM-CSF-induced differentiation. Endogenous SAA3 itself contributed to the increase in the survival and suppressive activity of Mo MDSCs. We demonstrated that SAA3 induced TLR2 signaling, in turn increasing the autocrine secretion of TNF-α, that led to STAT3 activation. In addition, activated STAT3 enhanced the suppressive activity of Mo MDSCs. Furthermore, SAA3 induction in Mo MDSCs contributed to accelerating tumor progression in vivo. Collectively, these data suggest a novel mechanism by which Mo MDSCs mediate inflammation through SAA3-TLR2 signaling and thus exacerbate cancer progression by a STAT3-dependent mechanism. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Clinical relevance of circulating anti-ENA and anti-dsDNA secreting cells from SLE patients and their dependence on STAT-3 activation.

    PubMed

    de la Varga Martínez, Raquel; Rodríguez-Bayona, Beatriz; Añez, Gustavo A; Medina Varo, Fermín; Pérez Venegas, José J; Brieva, José A; Rodríguez, Carmen

    2017-07-01

    Disturbances of plasma cell homeostasis and auto-antibody production are hallmarks of systemic lupus erythematosus. The aim of this study was to explore the presence of circulating anti-ENA and anti-dsDNA antibody-secreting cells, to determine their dependence on plasma cell-niche cytokines and to analyze their clinical value. The study was performed in SLE patients with serum anti-ENA and/or anti-dsDNA antibodies (n = 57). Enriched B-cell fractions and sorted antibody-secreting cells (CD19(low) CD38(high) ) were obtained from blood. dsDNA- and ENA-specific antibody-secreting cells were identified as cells capable of active auto-antibody production in culture. The addition of a combination of IL-6, IL-21, BAFF, APRIL, and CXCL12 to the cultures significantly augmented auto-antibody production and antibody-secreting cell proliferation, whereas it diminished apoptosis. The effect on auto-antibody production was dependent on STAT-3 activation as it was abrogated in the presence of the JAK/STAT-3 pathway inhibitors ruxolitinib and stattic. Among patients with serum anti-dsDNA antibodies, the detection of circulating anti-dsDNA-antibody-secreting cells was associated with higher disease activity markers. In conclusion, auto-antibody production in response to plasma cell-niche cytokines that are usually at high levels in SLE patients is dependent on JAK/STAT-3 activation. Thus, patients with circulating anti-dsDNA antibody-secreting cells and active disease could potentially benefit from therapies targeting the JAK/STAT3 pathway. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  16. Lumbar puncture-administered resveratrol inhibits STAT3 activation, enhancing autophagy and apoptosis in orthotopic rat glioblastomas

    PubMed Central

    Xue, Song; Xiao-Hong, Shu; Lin, Sha; Jie, Bian; Li-Li, Wang; Jia-Yao, Gu; Shun, Shi; Pei-Nan, Li; Mo-Li, Wu; Qian, Wang; Xiao-Yan, Chen; Qing-You, Kong; Peng, Zhang; Hong, Li; Jia, Liu

    2016-01-01

    Trans-resveratrol suppresses glioblastoma growth in vitro, but its effects on intracranial glioblastomas remain untested. Resveratrol crosses the blood–brain barrier, and lumbar puncture (LP) greatly increases its bioavailability in rat brains; therefore, we investigated the effectiveness of LP-administered resveratrol on orthotopic rat glioblastomas. Twenty-four tumor-bearing rats were separated into two groups: Group 1 receiving 100 μl saline containing 0.3% DMSO and Group 2 receiving 100 μl resveratrol (300 μM). Treatments started 3 days after transplantation in 2-day intervals until death. Intracranial drug availabilities, tumor sizes, average life spans and the impacts on STAT3 signaling, apoptosis and autophagy rates were evaluated. MRI imaging revealed that average tumor size in the LP group (495.8 ± 22.3 mm2) was smaller than the control groups (810.3 ± 56.4 mm2; P<0.05). The mean survival time in the LP group (22.2 ± 2.1 d) was longer than control animals (16.0 ± 1.8 d; P<0.05). LP resveratrol-treated glioblastomas showed less Cyclin D1 staining, enhanced autophagy with up-regulated LC3 and Beclin1 expression, and widely distributed apoptotic foci around tumor capillaries with suppressed STAT3 expression and nuclear translocation. In conclusion, LP-delivered resveratrol efficiently inhibited orthotopic rat glioblastoma growth by inactivating STAT3 signaling and enhancing autophagy and apoptosis. PMID:27716625

  17. STAT3-dependent VEGF production from keratinocytes abrogates dendritic cell activation and migration by arsenic: a plausible regional mechanism of immunosuppression in arsenical cancers.

    PubMed

    Hong, Chien-Hui; Lee, Chih-Hung; Chen, Gwo-Shing; Chang, Kee-Lung; Yu, Hsin-Su

    2015-02-05

    Arsenic remains an important environmental hazard that causes several human cancers. Arsenic-induced Bowen's disease (As-BD), a skin carcinoma in situ, is the most common arsenical cancer. While great strides have been made in our understanding of arsenic carcinogenesis, how host immunity contributes to this process remains unknown. Patients with As-BD have an impaired contact hypersensitivity response. Although impaired T cell activation has been well-documented in arsenical cancers, how dendritic cell (DC), the key cell regulating innate immunity, regulates the immune response in arsenical cancers remains unclear. Using myeloid derived DC (MDDC) from patients with As-BD and normal controls as well as bone marrow derived DC (BMDC) from mice fed with or without arsenic, we measured the migration of DC. As-BD patients showed an impaired CCL21-mediated MDDC migration in vitro. Arsenic-fed mice had defective DC migration toward popliteal lymph nodes when injected with allogenic BMDCs via foot pad. Using skin from As-BD and normal controls, we found an increased expression of STAT3, a transcriptional factor contributing to impaired DC activation. Arsenic induced STAT3 activation and the production of VEGF in keratinocytes. The increase in VEGF was blocked by inhibiting STAT3 with RNA interference or pharmaceutically with JSI-124. While VEGF by itself minimally induced the expression of CD86 and MHC-II in MDDC, arsenic induced-MDDC activation was abolished by VEGF pretreatment. We concluded that the STAT3-VEGF axis in keratinocytes inhibits DC migration in the microenvironment of As-BD, indicating that cellular interactions play an important role in regulating the disease course of arsenical cancers. Copyright © 2015. Published by Elsevier Ireland Ltd.

  18. Salinomycin Promotes Anoikis and Decreases the CD44+/CD24- Stem-Like Population via Inhibition of STAT3 Activation in MDA-MB-231 Cells.

    PubMed

    An, Hyunsook; Kim, Ji Young; Oh, Eunhye; Lee, Nahyun; Cho, Youngkwan; Seo, Jae Hong

    2015-01-01

    Triple-negative breast cancer (TNBC) is an aggressive tumor subtype with an enriched CD44+/CD24- stem-like population. Salinomycin is an antibiotic that has been shown to target cancer stem cells (CSC); however, the mechanisms of action involved have not been well characterized. The objective of the present study was to investigate the effect of salinomycin on cell death, migration, and invasion, as well as CSC-like properties in MDA-MB-231 breast cancer cells. Salinomycin significantly induced anoikis-sensitivity, accompanied by caspase-3 and caspase-8 activation and PARP cleavage, during anchorage-independent growth. Salinomycin treatment also caused a marked suppression of cell migration and invasion with concomitant downregulation of MMP-9 and MMP-2 mRNA levels. Notably, salinomycin inhibited the formation of mammospheres and effectively reduced the CD44+/CD24- stem-like population during anchorage-independent growth. These observations were associated with the inhibition of STAT3 phosphorylation (Tyr705). Furthermore, interleukin-6 (IL-6)-induced STAT3 activation was strongly suppressed by salinomycin challenge. These findings support the notion that salinomycin may be potentially efficacious for targeting breast cancer stem-like cells through the inhibition of STAT3 activation.

  19. Anti-angiogenic activity of thienopyridine derivative LCB03-0110 by targeting VEGFR-2 and JAK/STAT3 Signalling.

    PubMed

    Kim, Byung-Hak; Lee, Yoonji; Yoo, Hyun; Cui, Minghua; Lee, Sungwoon; Kim, Sun Young; Cho, Jong Un; Lee, Hyangsook; Yang, Beom-Seok; Kwon, Young-Guen; Choi, Sun; Kim, Tae-Yoon

    2015-07-01

    Vascular endothelial growth factor receptor-2 (VEGFR-2) and Janus kinase (JAK)/signal transducer and activator of transcription 3 (STAT3) signalling are important for tumor angiogenesis and metastasis. In this study, we identified (3-(2-(3-(morpholinomethyl)phenyl)thieno[3,2-b]pyridin-7-ylamino)phenol (LCB03-0110) as a potent angiogenesis inhibitor. LCB03-0110 inhibited VEGFR-2 and JAK/STAT3 signalling in primary cultured human endothelial cells and cancer cells. An in vitro kinase assay and molecular modelling revealed that LCB03-0110 inhibited VEGFR-2, c-SRC and TIE-2 kinase activity via preferential binding at the ATP-binding site of their kinases. LCB03-0110 successfully occupied the hydrophobic pocket of VEGFR-2, c-SRC and TIE-2. LCB03-0110 also inhibited hypoxia-induced HIF/STAT3 and EGF- or angiopoietin-induced signalling cascades. In addition, LCB03-0110 inhibited VEGF-induced proliferation, viability, migration and capillary-like tube formation. LCB03-0110 also suppressed the sprouting of endothelial cells in the rat aorta and the formation of new blood vessels in the mouse Matrigel plug assay, but also suppressed pulmonary metastasis and tumor xenograft in mice. Our results suggest that LCB03-0110 is a potential candidate small molecule for blocking angiogenesis mediated by aberrant activation of VEGFR-2 and JAK/STAT3 signalling. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  20. Cell confluency-induced Stat3 activation regulates NHE3 expression by recruiting Sp1 and Sp3 to the proximal NHE3 promoter region during epithelial dome formation.

    PubMed

    Su, Hsiao-Wen; Wang, Shainn-Wei; Ghishan, Fayez K; Kiela, Pawel R; Tang, Ming-Jer

    2009-01-01

    Activation of signal transducer and activator of transcription-3 (Stat3) during cell confluency is related to its regulatory roles in cell growth arrest- or survival-related physiological or developmental processes. We previously demonstrated that this signaling event triggers epithelial dome formation by transcriptional augmentation of sodium hydrogen exchanger-3 (NHE3) expression. However, the detailed molecular mechanism remained unclear. By using serial deletions, site-directed mutagenesis, and EMSA analysis, we now demonstrate Stat3 binding to an atypical Stat3-response element in the rat proximal NHE3 promoter, located adjacent to a cluster of Sp cis-elements (SpA/B/C), within -77/-36 nt of the gene. SpB (-58/-55 nt) site was more effective than SpA (-72/-69 nt) site for cooperative binding of Sp1/Sp3. Increasing cell density had no effect on Sp1/Sp3 expression but resulted in their increased binding to the SpA/B/C probe along with Stat3 and concurrently with enhanced nuclear pTyr705-Stat3 level. Immunoprecipitation performed with the nuclear extracts demonstrated physical interaction of Stat3 and Sp1/Sp3 triggered by cell confluency. Stat3 inhibition by overexpression of dominant-negative Stat3-D mutant in MDCK cells or by small interfering RNA-mediated knockdown in Caco-2 cells resulted in inhibition of the cell density-induced NHE3 expression, Sp1/Sp3 binding, and NHE3 promoter activity and in decreased dome formation. Thus, during confluency, ligand-independent Stat3 activation leads to its interaction with Sp1/Sp3, their recruitment to the SpA/B/C cluster in a Stat3 DNA-binding domain-dependent fashion, increased transcription, and expression of NHE3, to coordinate cell density-mediated epithelial dome formation.

  1. HIV-1 gp120 influences the expression of microRNAs in human monocyte-derived dendritic cells via STAT3 activation.

    PubMed

    Masotti, Andrea; Donninelli, Gloria; Da Sacco, Letizia; Varano, Barbara; Del Cornò, Manuela; Gessani, Sandra

    2015-06-27

    MicroRNAs (miRs) are an abundant class of small non-coding RNAs (~22 nt) that reprogram gene expression by targeting mRNA degradation and translational disruption. An emerging concept implicates miR coupling with transcription factors in myeloid cell development and function, thus contributing to host defense and inflammation. The important role that these molecules play in the pathogenesis of HIV-1 is only now emerging. We provide evidence that exposure of monocyte-derived dendritic cells (MDDCs) to recombinant HIV-1 R5 gp120, but not to CCR5 natural ligand CCL4, influences the expression of a panel of miRs (i.e., miR-21, miR-155 and miR-181b) regulated by STAT3 and potentially targeting genes belonging to the STAT3 signaling pathway. The blockage of gp120-induced STAT3 activation impairs gp120 capacity to modulate the expression level of above mentioned miRs. Predictive analysis of miR putative targets emphasizes that these miRs share common target genes. Furthermore, gene ontology and pathway enrichment analysis outline that these genes mainly belong to biological processes related to regulation of transcription, in a complex network of interactions involving pathways relevant to HIV-DC interaction. Overall, these results point to gp120-triggered modulation of miR expression via STAT3 activation as a novel molecular mechanism exploited by HIV-1 to affect DC biology and thus modulate the immune response through complex regulatory loops involving, at the same time, miRs and transcription factors.

  2. Autocrine activity of BDNF induced by the STAT3 signaling pathway causes prolonged TrkB activation and promotes human non-small-cell lung cancer proliferation

    PubMed Central

    Chen, Bo; Liang, Yan; He, Zheng; An, Yunhe; Zhao, Weihong; Wu, Jianqing

    2016-01-01

    Brain-derived neurotrophic factor (BDNF) is a member of the neurotrophin superfamily, which has been implicated in the pathophysiology of the nervous system. Recently, several studies have suggested that BDNF and/or its receptor, tropomyosin related kinase B (TrkB), are involved in tumor growth and metastasis in several cancers, including prostate cancer, neuroblastoma, pancreatic ductal carcinoma, hepatocellular carcinoma, and lung cancer. Despite the increasing emphasis on BDNF/TrkB signaling in human tumors, how it participates in primary tumors has not yet been determined. Additionally, little is known about the molecular mechanisms that elicit signaling downstream of TrkB in the progression of non-small-cell lung cancer (NSCLC). In this study, we report the significant expression of BDNF in NSCLC samples and show that BDNF stimulation increases the synthesis of BDNF itself through activation of STAT3 in lung cancer cells. The release of BDNF can in turn activate TrkB signaling. The activation of both TrkB and STAT3 contribute to downstream signaling and promote human non-small-cell lung cancer proliferation. PMID:27456333

  3. Human IL6 stimulates bovine satellite cell proliferation through a Signal transducer and activator of transcription 3 (STAT3)-dependent mechanism.

    PubMed

    Brandt, A M; Kania, J M; Reinholt, B M; Johnson, S E

    2017-08-31

    Bovine satellite cell (bSC) myogenesis and skeletal muscle hypertrophy occur through the orchestrated actions of multiple autocrine and paracrine growth factors. Intimate to the bSC niche is IL6, a dual-purpose cytokine with proinflammatory and mitogenic properties. The objective of the experiment was to examine the effects of IL6 on proliferation and differentiation of bSC in vitro. Treatment of primary bSC cultures with recombinant bovine IL6 (bIL6) failed to alter myogenesis owing to the absence of intracellular signal transduction. The cytokine was able to stimulate phosphorylation of signal transducer and activator of transcription 3 tyrosine 705 (STAT3(Y705)) in Madin-Darby bovine kidney (MDBK) epithelial cells, thus demonstrating bioactivity. Media supplemented with recombinant human IL6 (hIL6) caused phosphorylation of STAT3(Y705) in bSC and increased (P < 0.05) proliferation. Inclusion of a STAT3 inhibitor in the media blunted phosphorylation of the STAT3(Y705) and suppressed (P < 0.05) hIL6-mediated bSC proliferation. Morphologic and biochemical measures of bSC differentiation remained unchanged (P > 0.05) following treatment for 48 h with hIL6. These results support a role for hIL6 as a bSC mitogen in vitro. The inability of bIL6 to initiate an intracellular signal in bSC requires further investigation. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Induction of Sca-1 via activation of STAT3 system in the duct cells of the mouse submandibular gland by ligation of the main excretory duct.

    PubMed

    Purwanti, Nunuk; Karabasil, Mileva Ratko; Matsuo, Shinsuke; Chen, Gang; Javkhlan, Purevjav; Azlina, Ahmad; Hasegawa, Takahiro; Yao, Chenjuan; Akamatsu, Tetsuya; Hosoi, Kazuo

    2011-11-01

    To examine the very initial step that takes place immediately after tissue injury and is linked to tissue regeneration, we employed the submandibular gland (SMG), which was injured by ligation of its main excretory duct (MED). Ligation of the MED of the SMG in mice induced the expression of Sca-1, a protein marker of hematopoietic stem cells. In the normal gland, a low level of Sca-1 was expressed, which was localized predominantly in the excretory duct cells. At 1 day after ligation, Sca-1 expression increased prominently in almost all of cells in the duct system, but not in the acinar cells. The level of Sca-1 mRNA had begun to increase at 6 h after ligation and continuously rose thereafter until it reached a plateau, which occurred ∼12 h after ligation. STAT3 phosphorylated at its tyrosine-705 (p-STAT3) in the ligated gland increased immediately after ligation, and it was localized in the nuclei of all duct cells. The results of an EMSA revealed the specific binding of a nuclear extract to the sequence of the γ-interferon activation site (GAS) present in the Sca-1 promoter and confirmed that such binding increased after ligation. Thus the present study suggests that STAT3, having been phosphorylated following MED ligation, was transferred to the nucleus, where it bound to the GAS element in the promoter of Sca-1 gene, resulting in promotion of Sca-1 gene expression. Actual prevention of STAT3 phosphorylation reduced the ligation-induced Sca-1 elevation.

  5. Oncogenic transformation of human lung bronchial epithelial cells induced by arsenic involves ROS-dependent activation of STAT3-miR-21-PDCD4 mechanism

    PubMed Central

    Pratheeshkumar, Poyil; Son, Young-Ok; Divya, Sasidharan Padmaja; Wang, Lei; Zhang, Zhuo; Shi, Xianglin

    2016-01-01

    Arsenic is a well-documented human carcinogen. The present study explored the role of the onco-miR, miR-21 and its target protein, programmed cell death 4 (PDCD4) in arsenic induced malignant cell transformation and tumorigenesis. Our results showed that treatment of human bronchial epithelial (BEAS-2B) cells with arsenic induces ROS through p47phox, one of the NOX subunits that is the key source of arsenic-induced ROS. Arsenic exposure induced an upregulation of miR-21 expression associated with inhibition of PDCD4, and caused malignant cell transformation and tumorigenesis of BEAS-2B cells. Indispensably, STAT3 transcriptional activation by IL-6 is crucial for the arsenic induced miR-21 increase. Upregulated miR-21 levels and suppressed PDCD4 expression was also observed in xenograft tumors generated with chronic arsenic exposed BEAS-2B cells. Stable shut down of miR-21, p47phox or STAT3 and overexpression of PDCD4 or catalase in BEAS-2B cells markedly inhibited the arsenic induced malignant transformation and tumorigenesis. Similarly, silencing of miR-21 or STAT3 and forced expression of PDCD4 in arsenic transformed cells (AsT) also inhibited cell proliferation and tumorigenesis. Furthermore, arsenic suppressed the downstream protein E-cadherin expression and induced β-catenin/TCF-dependent transcription of uPAR and c-Myc. These results indicate that the ROS-STAT3-miR-21-PDCD4 signaling axis plays an important role in arsenic -induced carcinogenesis. PMID:27876813

  6. XPO1 (CRM1) inhibition represses STAT3 activation to drive a survivin-dependent oncogenic switch in triple-negative breast cancer.

    PubMed

    Cheng, Yan; Holloway, Michael P; Nguyen, Kevin; McCauley, Dilara; Landesman, Yosef; Kauffman, Michael G; Shacham, Sharon; Altura, Rachel A

    2014-03-01

    Inhibition of XPO1 (CRM1)-mediated nuclear export of multiple tumor suppressor proteins has been proposed as a novel cancer therapeutic strategy to turn off oncogenic signals and enhance tumor suppression. Survivin is a multifunctional protein with oncogenic properties when expressed in the cytoplasm that requires the XPO1-RanGTP complex for its nuclear export. We investigated the antitumor mechanisms of the drug-like selective inhibitors of nuclear export (SINE) XPO1 antagonists KPT-185, KPT-251 KPT-276, and KPT-330 in estrogen receptor-positive and triple-negative breast cancer (TNBC) cell lines and xenograft models of human breast tumors. KPT compounds significantly inhibited breast cancer cell growth and induced tumor cell death, both in vitro and in vivo. These drugs initially promoted survivin accumulation within tumor cell nuclei. However, their major in vitro effect was to decrease survivin cytoplasmic protein levels, correlating with the onset of apoptosis. XPO1 inhibition repressed Survivin transcription by inhibiting CREB-binding protein-mediated STAT3 acetylation, and blocking STAT3 binding to the Survivin promoter. In addition, caspase-3 was activated to cleave survivin, rendering it unavailable to bind X-linked inhibitor of apoptosis protein and block the caspase cascade. Collectively, these data demonstrate that XPO1 inhibition by SINE compounds represses STAT3 transactivation to block the selective oncogenic properties of survivin and supports their clinical use in TNBC.

  7. Activation of IL-11/STAT3 pathway in preconditioned human skeletal myoblasts blocks apoptotic cascade under oxidant stress.

    PubMed

    Idris, Niagara Muhammad; Ashraf, Muhammad; Ahmed, Rafeeq P H; Shujia, Jiang; Haider, Khawaja H

    2012-01-01

    To determine whether our novel approach of diazoxide-induced stem cell preconditioning might be extrapolated to human skeletal myoblasts to support their survival under lethal oxidant stress. Using an in vitro model of H(2)O(2) treatment of human skeletal myoblasts, we report the ability of diazoxide-preconditioned human skeletal myoblasts to express cytokines and growth factors, which act in an autocrine and paracrine fashion to promote their own survival. Preconditioning of skeletal myoblasts was cytoprotective and significantly reduced their apoptotic index (p < 0.05). IL-11 gene and protein expression was significantly increased in preconditioned skeletal myoblasts. Transfection of skeletal myoblasts with IL-11-specific siRNA incurred their death under oxidant stress. The cytoprotective effect of diazoxide preconditioning was blocked by Erk1/2 inhibitor PD98059 (20-100 µM), which abrogated STAT-3 phosphorylation, thus confirming a possible involvement of Erk1/2/STAT3 signaling downstream of IL-11 in cell survival. We also investigated the time course of subcellular changes and signaling pathway of skeletal myoblasts apoptosis under oxidant stress before and after preconditioning. Apoptosis was induced in skeletal myoblasts with 100-500 µM H(2)O(2) for time points ranging from 1 to 24 h. Release of lactate dehydrogenase, disruption of the mitochondrial membrane potential and cytochrome-c translocation into cytoplasm were the earliest signs of apoptosis. Total Akt protein remained unchanged whereas marked reduction in pAkt was observed in the native skeletal myoblasts. Terminal dUTP nick end-labeling and annexin-V positivity were significantly increased after 4 h. Ultra-structure studies showed condensed chromatin, shriveled nuclei and swollen mitochondria. These data suggest that skeletal myoblasts undergo apoptosis under oxidant stress in a time-dependent manner and preconditioning of skeletal myoblasts significantly prevented their apoptosis via IL-11/STAT3

  8. Non-canonical Stat3 signaling in cancer.

    PubMed

    Srivastava, Jaya; DiGiovanni, John

    2016-12-01

    Stat3 is a member of the signal transducers and activators of transcription family and is a known regulator of essential biologic processes including angiogenesis, apoptosis, cell cycle progression, and cell migration. Canonical Stat3-mediated signaling involves tyrosine phosphorylation on specific residues that leads to homodimerization and translocation to the nucleus. For many years it was presumed that most, if not all, of the functions of Stat3, both normal and aberrant, were due to the canonical cytokine and growth factor signaling mechanisms. Recent studies suggest that Stat3 functions through alternate non-canonical pathways to bring about some of these biological functions both in normal cells as well as during cancer development and progression. A number of studies have now shown that Stat3 has a function in mitochondria and that unphosphorylated Stat3 (uStat3) can also function as a transcription factor broadening the potential mechanisms involved in Stat3 action. In this review article, we discuss these two main non-canonical functions of Stat3 and their potential roles in oncogenesis. Given the many facets of Stat3 signaling, additional comprehensive investigations are required to fully understand the role of non-canonical Stat3 signaling in cancer and whether these pathways can be targeted for cancer prevention and treatment. © 2015 Wiley Periodicals, Inc. © 2015 Wiley Periodicals, Inc.

  9. Erythropoietin promotes regeneration of adult CNS neurons via Jak2/Stat3 and PI3K/AKT pathway activation.

    PubMed

    Kretz, Alexandra; Happold, Caroline J; Marticke, Julia K; Isenmann, Stefan

    2005-08-01

    The cytokine hormone erythropoietin (EPO) has proved neuroprotective in CNS injury, and clinical trials for ischemic stroke are ongoing. The capability of EPO to restore postmitotic CNS architecture and function by fibre regeneration has not been examined. Here, we compared in vitro outgrowth capacity of adult retinal ganglion cells (RGCs) following optic nerve (ON) lesion in the presence and absence of EPO. Immediate EPO conditioning in vivo, or delayed EPO treatment of cultures with 10--10,000 IU rhEPO significantly increased numbers (2.66-fold) and length (8.31-fold) of newly generated neurites, without evoking rheological complications. EPO induced Stat3 phosphorylation in RGCs, and inhibition of Jak2/Stat3 abolished EPO-induced growth. EPO-facilitated neuritogenesis was paralleled by upregulation of Bcl-X(L), a Bcl-2 homologue capable of promoting RGC regeneration. The PI3K/Akt pathway was also involved in antiapoptotic and regeneration-enhancing EPO actions. In conclusion, EPO treatment may offer a unique dual-function strategy for neuroprotection and regeneration.

  10. PSM/SH2B1 splice variants: critical role in src catalytic activation and the resulting STAT3s-mediated mitogenic response.

    PubMed

    Zhang, Manchao; Deng, Youping; Riedel, Heimo

    2008-05-01

    A role of PSM/SH2B1 had been shown in mitogenesis and extending to phenotypic cell transformation, however, the underlying molecular mechanism remained to be established. Here, four alternative PSM splice variants and individual functional protein domains were compared for their role in the regulation of Src activity. We found that elevated cellular levels of PSM variants resulted in phenotypic cell transformation and potentiated cell proliferation and survival in response to serum withdrawal. PSM variant activity presented a consistent signature pattern for any tested response of highest activity observed for gamma, followed by delta, alpha, and beta with decreasing activity. PSM-potentiated cell proliferation was sensitive to Src inhibitor herbimycin and PSM and Src were found in the same immune complex. PSM variants were substrates of the Src Tyr kinase and potentiated Src catalytic activity by increasing the V(max) and decreasing the K(m) for ATP with the signature pattern of variant activity. Dominant-negative PSM peptide mimetics including the SH2 or PH domains inhibited Src catalytic activity as well as Src-mediated phenotypic cell transformation. Activation of major Src substrate STAT3 was similarly potentiated by the PSM variants in a Src-dependent fashion or inhibited by PSM domain-specific peptide mimetics. Expression of a dominant-negative STAT3 mutant blocked PSM variant-mediated phenotypic cell transformation. Our results implicate an essential role of the PSM variants in the activation of the Src kinase and the resulting mitogenic response--extending to phenotypic cell transformation and involving the established Src substrate STAT3.

  11. Cloning of a new type II cytokine receptor activating signal transducer and activator of transcription (STAT)1, STAT2 and STAT3.

    PubMed Central

    Dumoutier, Laure; Lejeune, Diane; Hor, Simon; Fickenscher, Helmut; Renauld, Jean-Christophe

    2003-01-01

    In the present paper, we report the identification of a new gene encoding a transmembrane protein of 520 amino acids, showing 22% amino acid identity with the extracellular domain of the interleukin (IL)-20 receptor. This gene, termed likely interleukin or cytokine receptor-2 ( LICR2 ), is located on chromosome 1, at 25 kb from the IL22R (IL-22 receptor) gene, and is constitutively expressed in most tissues. A chimaeric receptor, consisting of the extracellular domain of the IL-10 receptor alpha chain and the intracellular domain of LICR2, activated signal transducer and activator of transcription (STAT)1, STAT2, STAT3 and STAT5 upon IL-10 stimulation, in a Janus kinase 1-dependent manner. In contrast, none of the IL-10-related cytokines described so far could activate LICR2-transfected cells, suggesting that LICR2 is a signalling receptor for a new cytokine of the IL-10 family. PMID:12521379

  12. The prenylflavonoid Icaritin enhances osteoblast proliferation and function by signal transducer and activator of transcription factor 3 (STAT-3) regulation of C-X-C chemokine receptor type 4 (CXCR4) expression.

    PubMed

    Lim, Rzl; Li, L; Chew, N; Yong, E L

    2017-08-31

    In this study, we examined the effects of a natural prenylflavonoid Icaritin (ICT), on human osteoblast proliferation and osteogenic function. We observed that ICT dose-dependently enhanced osteoblast proliferation by ~15% over a 7day period. This increase in cell proliferation was associated with corresponding increases in osteoblast functions as measured by ALP secretion, intracellular calcium ions influx and calcium deposition. These anabolic effects were associated with a 4-fold increase in CXCR4 mRNA and protein expression. Silencing of CXCR4 protein expression using small interfering RNA reversed ICT-induced increase in cell proliferation, ALP activity and calcium deposition. Interestingly, we observed that ICT dose-dependently increased STAT-3 phosphorylation; and this resulted in increased binding of phosphorylated STAT-3 to the promoter region of the CXCR4 gene, to increase CXCR4 protein expression. Furthermore, we found that inhibition of STAT-3 phosphorylation resulted in a decrease in CXCR4 protein expression; whilst increasing phosphorylation of STAT-3 using a constitutive active STAT-3 vector significantly increased CXCR4 levels. Moreover, the chemical inhibition of STAT-3 phosphorylation annulled our previously observed ICT-induced increases of osteoblast proliferation and function. Finally, in a rat model of estrogen-deficient osteoporosis, ICT restored both osteoblasts numbers and CXCR4 expression. Taken together, both cellular and animal models support the novel findings that ICT; through the phosphorylation of STAT-3, up-regulated CXCR4, to increase osteoblast proliferation and function. Copyright © 2017. Published by Elsevier Inc.

  13. Alternative Roles of STAT3 and MAPK Signaling Pathways in the MMPs Activation and Progression of Lung Injury Induced by Cigarette Smoke Exposure in ACE2 Knockout Mice

    PubMed Central

    Hung, Yi-Han; Hsieh, Wen-Yeh; Hsieh, Jih-Sheng; Liu, Fon-Chang; Tsai, Chin-Hung; Lu, Li-Che; Huang, Chen-Yi; Wu, Chien-Liang; Lin, Chih-Sheng

    2016-01-01

    Inflammation-mediated abnormalities in the renin-angiotensin system (RAS) and expression of matrix metalloproteinases (MMPs) are implicated in the pathogenesis of lung injury. Angiotensin converting enzyme II (ACE2), an angiotensin converting enzyme (ACE) homologue that displays antagonist effects on ACE/angiotensin II (Ang II) axis, could also play a protective role against lung diseases. However, the relationship between ACE2 and MMPs activation in lung injury is still largely unclear. The purpose of this study is to investigate whether MMPs activity could be affected by ACE2 and which ACE2 derived signaling pathways could be also involved via using a mouse model with lung injury induced by cigarette smoke (CS) exposure for 1 to 3 weeks. Wild-type (WT; C57BL/6) and ACE2 KO mice (ACE2-/-) were utilized to study CS-induced lung injury. Increases in the resting respiratory rate (RRR), pulmonary immunokines, leukocyte infiltration and bronchial hyperplasia were observed in the CS-exposed mice. Compared to WT mice, more serious physiopathological changes were found in ACE2-/- mice in the first week of CS exposure. CS exposure increased pulmonary ACE and ACE2 activities in WT mice, and significantly increased ACE in ACE2-/- mice. Furthermore, the activity of pulmonary MMPs was decreased in CS-exposed WT mice, whereas this activity was increased in ACE2-/- mice. CS exposure increased the pulmonary p-p38, p-JNK and p-ERK1/2 level in all mice. In ACE2-/- mice, a significant increase p-STAT3 signaling was detected; however, no effect was observed on the p-STAT3 level in WT mice. Our results support the hypothesis that ACE2 deficiency influences MMPs activation and STAT3 phosphorylation signaling to promote more pulmonary inflammation in the development of lung injury. PMID:27019629

  14. Alternative Roles of STAT3 and MAPK Signaling Pathways in the MMPs Activation and Progression of Lung Injury Induced by Cigarette Smoke Exposure in ACE2 Knockout Mice.

    PubMed

    Hung, Yi-Han; Hsieh, Wen-Yeh; Hsieh, Jih-Sheng; Liu, Fon-Chang; Tsai, Chin-Hung; Lu, Li-Che; Huang, Chen-Yi; Wu, Chien-Liang; Lin, Chih-Sheng

    2016-01-01

    Inflammation-mediated abnormalities in the renin-angiotensin system (RAS) and expression of matrix metalloproteinases (MMPs) are implicated in the pathogenesis of lung injury. Angiotensin converting enzyme II (ACE2), an angiotensin converting enzyme (ACE) homologue that displays antagonist effects on ACE/angiotensin II (Ang II) axis, could also play a protective role against lung diseases. However, the relationship between ACE2 and MMPs activation in lung injury is still largely unclear. The purpose of this study is to investigate whether MMPs activity could be affected by ACE2 and which ACE2 derived signaling pathways could be also involved via using a mouse model with lung injury induced by cigarette smoke (CS) exposure for 1 to 3 weeks. Wild-type (WT; C57BL/6) and ACE2 KO mice (ACE2(-/-)) were utilized to study CS-induced lung injury. Increases in the resting respiratory rate (RRR), pulmonary immunokines, leukocyte infiltration and bronchial hyperplasia were observed in the CS-exposed mice. Compared to WT mice, more serious physiopathological changes were found in ACE2(-/-) mice in the first week of CS exposure. CS exposure increased pulmonary ACE and ACE2 activities in WT mice, and significantly increased ACE in ACE2(-/-) mice. Furthermore, the activity of pulmonary MMPs was decreased in CS-exposed WT mice, whereas this activity was increased in ACE2(-/-) mice. CS exposure increased the pulmonary p-p38, p-JNK and p-ERK1/2 level in all mice. In ACE2(-/-) mice, a significant increase p-STAT3 signaling was detected; however, no effect was observed on the p-STAT3 level in WT mice. Our results support the hypothesis that ACE2 deficiency influences MMPs activation and STAT3 phosphorylation signaling to promote more pulmonary inflammation in the development of lung injury.

  15. Role of STAT3 in Cancer Metastasis and Translational Advances

    PubMed Central

    Patil, Prachi; Gude, Rajiv P.

    2013-01-01

    Signal transducer and activator of transcription 3 (STAT3) is a latent cytoplasmic transcription factor, originally discovered as a transducer of signal from cell surface receptors to the nucleus. It is activated by tyrosine phosphorylation at position 705 leading to its dimerization, nuclear translocation, DNA binding, and activation of gene transcription. Under normal physiological conditions, STAT3 activation is tightly regulated. However, compelling evidence suggests that STAT3 is constitutively activated in many cancers and plays a pivotal role in tumor growth and metastasis. It regulates cellular proliferation, invasion, migration, and angiogenesis that are critical for cancer metastasis. In this paper, we first describe the mechanism of STAT3 regulation followed by how STAT3 is involved in cancer metastasis, then we summarize the various small molecule inhibitors that inhibit STAT3 signaling. PMID:24199193

  16. Intermolecular disulfide bond influences unphosphorylated STAT3 dimerization and function.

    PubMed

    Butturini, Elena; Gotte, Giovanni; Dell'Orco, Daniele; Chiavegato, Giulia; Marino, Valerio; Canetti, Diana; Cozzolino, Flora; Monti, Maria; Pucci, Piero; Mariotto, Sofia

    2016-10-01

    Signal transducer and activator of transcription 3 (STAT3) is a transcription factor activated by the phosphorylation of tyrosine 705 in response to many cytokines and growth factors. Recently, the roles for unphosphorylated STAT3 (U-STAT3) have been described in response to cytokine stimulation, in cancers, and in the maintenance of heterochromatin stability. It has been reported that U-STAT3 dimerizes, shuttles between the cytoplasm and nucleus, and binds to DNA, thereby driving genes transcription. Although many reports describe the active role of U-STAT3 in oncogenesis in addition to phosphorylated STAT3, the U-STAT3 functional pathway remains elusive.In this report, we describe the molecular mechanism of U-STAT3 dimerization, and we identify the presence of two intermolecular disulfide bridges between Cys367 and Cys542 and Cys418 and Cys426, respectively. Recently, we reported that the same cysteines contribute to the redox regulation of STAT3 signaling pathway both in vitro and in vivo The presence of these disulfides is here demonstrated to largely contribute to the structure and the stability of U-STAT3 dimer as the dimeric form rapidly dissociates upon reduction in the S-S bonds. In particular, the Cys367-Cys542 disulfide bridge is shown to be critical for U-STAT3 DNA-binding activity. Mutation of the two Cys residues completely abolishes the DNA-binding capability of U-STAT3. Spectroscopic investigations confirm that the noncovalent interactions are sufficient for proper folding and dimer formation, but that the interchain disulfide bonds are crucial to preserve the functional dimer. Finally, we propose a reaction scheme of U-STAT3 dimerization with a first common step followed by stabilization through the formation of interchain disulfide bonds. © 2016 The Author(s); published by Portland Press Limited on behalf of the Biochemical Society.

  17. AICAR induces astroglial differentiation of neural stem cells via activating the JAK/STAT3 pathway independently of AMP-activated protein kinase.

    PubMed

    Zang, Yi; Yu, Li-Fang; Pang, Tao; Fang, Lei-Ping; Feng, Xu; Wen, Tie-Qiao; Nan, Fa-Jun; Feng, Lin-Yin; Li, Jia

    2008-03-07

    Neural stem cell differentiation and the determination of lineage decision between neuronal and glial fates have important implications in the study of developmental, pathological, and regenerative processes. Although small molecule chemicals with the ability to control neural stem cell fate are considered extremely useful tools in this field, few were reported. AICAR is an adenosine analog and extensively used to activate AMP-activated protein kinase (AMPK), a metabolic "fuel gauge" of the biological system. In the present study, we found an unrecognized astrogliogenic activity of AICAR on not only immortalized neural stem cell line C17.2 (C17.2-NSC), but also primary neural stem cells (NSCs) derived from post-natal (P0) rat hippocampus (P0-NSC) and embryonic day 14 (E14) rat embryonic cortex (E14-NSC). However, another AMPK activator, Metformin, did not alter either the C17.2-NSC or E14-NSC undifferentiated state although both Metformin and AICAR can activate the AMPK pathway in NSC. Furthermore, overexpression of dominant-negative mutants of AMPK in C17.2-NSC was unable to block the gliogenic effects of AICAR. We also found AICAR could activate the Janus kinase (JAK) STAT3 pathway in both C17.2-NSC and E14-NSC but Metformin fails. JAK inhibitor I abolished the gliogenic effects of AICAR. Taken together, these results suggest that the astroglial differentiation effect of AICAR on neural stem cells was acting independently of AMPK and that the JAK-STAT3 pathway is essential for the gliogenic effect of AICAR.

  18. STAT1 and STAT3 in tumorigenesis

    PubMed Central

    Avalle, Lidia; Pensa, Sara; Regis, Gabriella; Novelli, Francesco; Poli, Valeria

    2012-01-01

    The transcription factors STAT1 and STAT3 appear to play opposite roles in tumorigenesis. While STAT3 promotes cell survival/proliferation, motility and immune tolerance and is considered as an oncogene, STAT1 mostly triggers anti-proliferative and pro-apoptotic responses while enhancing anti-tumor immunity. Despite being activated downstream of common cytokine and growth factor receptors, their activation is reciprocally regulated and perturbation in their balanced expression or phosphorylation levels may re-direct cytokine/growth factor signals from proliferative to apoptotic, or from inflammatory to anti-inflammatory. Here we review the functional canonical and non-canonical effects of STAT1 and STAT3 activation in tumorigenesis and their potential cross-regulation mechanisms. PMID:24058752

  19. Downregulated expression of microRNA-124 in pediatric intestinal failure patients modulates macrophages activation by inhibiting STAT3 and AChE

    PubMed Central

    Xiao, Yong-Tao; Wang, Jun; Lu, Wei; Cao, Yi; Cai, Wei

    2016-01-01

    Intestinal inflammation plays a critical role in the pathogenesis of intestinal failure (IF). The macrophages are essential to maintain the intestinal homeostasis. However, the underlying mechanisms of intestinal macrophages activation remain poorly understood. Since microRNAs (miRNAs) have pivotal roles in regulation of immune responses, here we aimed to investigate the role of miR-124 in the activation of intestinal macrophages. In this study, we showed that the intestinal macrophages increased in pediatric IF patients and resulted in the induction of interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α). The miRNA fluorescence in situ hybridization analysis showed that the expression of miR-124 significantly reduced in intestinal macrophages in IF patients. Overexpression of miR-124 was sufficient to inhibit intestinal macrophages activation by attenuating production of IL-6 and TNF-α. Further studies showed that miR-124 could directly target the 3′-untranslated region of both signal transducer and activator of transcription 3 (STAT3) and acetylcholinesterase (AChE) mRNAs, and suppress their protein expressions. The AChE potentially negates the cholinergic anti-inflammatory signal by hydrolyzing the acetylcholine. We here showed that intestinal macrophages increasingly expressed the AChE and STAT3 in IF patients when compared with controls. The inhibitors against to STAT3 and AChE significantly suppressed the lipopolysaccharides-induced IL-6 and TNF-α production in macrophages. Taken together, these findings highlight an important role for miR-124 in the regulation of intestinal macrophages activation, and suggest a potential application of miR-124 in pediatric IF treatment regarding as suppressing intestinal inflammation. PMID:27977009

  20. Induction of neurite outgrowth by interleukin-6 is accompanied by activation of Stat3 signaling pathway in a variant PC12 cell (E2) line.

    PubMed

    Wu, Y Y; Bradshaw, R A

    1996-05-31

    PC12-E2 cells, a stable variant subcloned from native cell populations, produce neurites in a rapid, transcription-independent manner upon exposure to nerve growth factor (NGF) or basic fibroblast growth factor (bFGF). They also give a similar morphological response to interleukin-6 (IL-6), which is, however, transcription-dependent and with a slower onset, a phenomenon basically not observed in native PC12 cells. The response profile of PC12-E2 cells to NGF and bFGF is similar to that observed for native PC12 cells pre-exposed (primed) to NGF, and such cells also respond to IL-6 in a fashion indistinguishable from PC12-E2 cells. Mechanistically, NGF and bFGF induce a sustained phosphorylation and activation of ERK1 and ERK2 in both cells, while IL-6 produces only a transient and weak tyrosine phosphorylation. However, it does stimulate a prolonged and biphasic tyrosine phosphorylation and nuclear translocation of Stat3 (signal transducers and activators of transcription 3; at least 24 h) and, to a lesser extent, Stat1. Gel shift and supershift analyses confirm that IL-6 predominantly activates Stat3 (and some Stat1) and stimulates sis-inducible element binding activity. Other members of the same cytokine subfamily, including ciliary neurotrophic factor and leukemia inhibitory factor, also cause a transient initial phase of tyrosine phosphorylation and activation of Stat1 and Stat3 (up to 1 h) but fail to stimulate a second phase of response and do not produce significant neurites. These results suggest that sustained signaling of either STAT or ERK pathways in PC12-E2 cells leads to induction of neuronal differentiation. However, only the latter is effective in native PC12 cells as the activation of Stat3 and Stat1 in native PC12 cells by IL-6 fails to induce neuronal differentiation. Thus, the response of PC12-E2 cells to IL-6 suggests the constitutive expression of a required factor(s) for differentiation, that is induced in native PC12 cells by NGF or b

  1. Chemokine CCL2-CCR2 Signaling Induces Neuronal Cell Death via STAT3 Activation and IL-1β Production after Status Epilepticus.

    PubMed

    Tian, Dai-Shi; Peng, Jiyun; Murugan, Madhuvika; Feng, Li-Jie; Liu, Jun-Li; Eyo, Ukpong B; Zhou, Li-Jun; Mogilevsky, Rochelle; Wang, Wei; Wu, Long-Jun

    2017-08-16

    Elevated levels of chemokine C-C motif ligand 2 (CCL2) and its receptor CCR2 have been reported in patients with temporal lobe epilepsy and in experimental seizures. However, the functional significance and molecular mechanism underlying CCL2-CCR2 signaling in epileptic brain remains largely unknown. In this study, we found that the upregulated CCL2 was mainly expressed in hippocampal neurons and activated microglia from mice 1 d after kainic acid (KA)-induced seizures. Taking advantage of CX3CR1(GFP/+):CCR2(RFP/+) double-transgenic mice, we demonstrated that CCL2-CCR2 signaling has a role in resident microglial activation and blood-derived monocyte infiltration. Moreover, seizure-induced degeneration of neurons in the hippocampal CA3 region was attenuated in mice lacking CCL2 or CCR2. We further showed that CCR2 activation induced STAT3 (signal transducer and activator of transcription 3) phosphorylation and IL-1β production, which are critical for promoting neuronal cell death after status epilepticus. Consistently, pharmacological inhibition of STAT3 by WP1066 reduced seizure-induced IL-1β production and subsequent neuronal death. Two weeks after KA-induced seizures, CCR2 deficiency not only reduced neuronal loss, but also attenuated seizure-induced behavioral impairments, including anxiety, memory decline, and recurrent seizure severity. Together, we demonstrated that CCL2-CCR2 signaling contributes to neurodegeneration via STAT3 activation and IL-1β production after status epilepticus, providing potential therapeutic targets for the treatment of epilepsy.SIGNIFICANCE STATEMENT Epilepsy is a global concern and epileptic seizures occur in many neurological conditions. Neuroinflammation associated with microglial activation and monocyte infiltration are characteristic of epileptic brains. However, molecular mechanisms underlying neuroinflammation in neuronal death following epilepsy remain to be elucidated. Here we demonstrate that CCL2-CCR2 signaling is

  2. Activation of the FGFR-STAT3 pathway in breast cancer cells induces a hyaluronan-rich microenvironment that licenses tumor formation

    PubMed Central

    Bohrer, Laura R.; Chuntova, Pavlina; Bade, Lindsey K.; Beadnell, Thomas C.; Leon, Ronald P.; Brady, Nicholas J.; Ryu, Yungil; Goldberg, Jodi E.; Schmechel, Stephen C.; Koopmeiners, Joseph S.; McCarthy, James B.; Schwertfeger, Kathryn L.

    2014-01-01

    Aberrant activation of fibroblast growth factor receptors (FGFRs) contributes to breast cancer growth, progression and therapeutic resistance. Due to the complex nature of the FGF/FGFR axis, and the numerous effects of FGFR activation on tumor cells and the surrounding microenvironment, the specific mechanisms through which aberrant FGFR activity contributes to breast cancer are not completely understood. We show here that FGFR activation induces accumulation of hyaluronan (HA) within the extracellular matrix (ECM) and that blocking HA synthesis decreases proliferation, migration and therapeutic resistance. Furthermore, FGFR-mediated HA accumulation requires activation of the signal transducer and activator of transcription 3 (STAT3) pathway, which regulates expression of hyaluronan synthase 2 (HAS2) and subsequent HA synthesis. Using a novel in vivo model of FGFR-dependent tumor growth, we demonstrate that STAT3 inhibition decreases both FGFR-driven tumor growth and HA levels within the tumor. Finally, our results suggest that combinatorial therapies inhibiting both FGFR activity and HA synthesis is more effective than targeting either pathway alone and may be a relevant therapeutic approach for breast cancers associated with high levels of FGFR activity. In conclusion, these studies indicate a novel targetable mechanism through which FGFR activation in breast cancer cells induces a pro-tumorigenic microenvironment. PMID:24197137

  3. Anti-myeloma activity of a multi targeted kinase inhibitor, AT9283, via potent Aurora Kinase and STAT3 inhibition either alone or in combination with lenalidomide

    PubMed Central

    Santo, Loredana; Hideshima, Teru; Cirstea, Diana; Bandi, Madhavi; Nelson, Erik A.; Gorgun, Gullu; Rodig, Scott; Vallet, Sonia; Pozzi, Samantha; Patel, Kishan; Unitt, Christine; Squires, Matt; Hu, Yiguo; Chauhan, Dharminder; Mahindra, Anuj; Munshi, Nikhil C.; Anderson, Kenneth C.; Raje, Noopur

    2014-01-01

    Purpose Aurora Kinases, whose expression is linked to genetic instability and cellular proliferation, are under investigation as novel therapeutic targets in multiple myeloma (MM). Here, we investigated the preclinical activity of a small molecule–multi-targeted kinase inhibitor, AT9283, with potent activity against Aurora kinase A (AURKA), Aurora kinase B (AURKB) and Janus Kinase 2/3. Experimental design We evaluated the in vitro anti myeloma activity of AT9283 alone and in combination with lenalidomide and the in vivo efficacy by using a Xenograft mouse model of human MM. Results Our data demonstrated AT9283 induced cell growth inhibition and apoptosis in MM. Studying the apoptosis mechanism of AT9283 in MM, we observed features consistent with both AURKA and AURKB inhibition, e.g increase of cells with polyploid DNA content, decrease in phospho-Histone H3, and decrease of phospho-Aurora A. Importantly, AT9283 also inhibited STAT3 tyrosine phosphorylation in MM cells. Genetic depletion of STAT3, AURKA or AURKB showed growth inhibition of MM cells, suggesting a role of AT9283-induced inhibition of these molecules in the underlying mechanism of MM cell death. In vivo studies demonstrated decreased MM cell growth and prolonged survival in AT9283-treated mice compared to controls. Importantly, combination studies of AT9283 with lenalidomide showed significant synergistic cytotoxicity in MM cells, even in the presence of bone marrow stromal cells (BMSCs). Enhanced cytotoxicity was associated with increased inhibition of pSTAT3 and pERK. Conclusions Demonstration of in vitro and in vivo anti-MM activity of AT9283 provides the rationale for the clinical evaluation of AT9283 as monotherapy and in combination in patients with MM. PMID:21430070

  4. Selective JAK/STAT3 signalling regulates transcription of colony stimulating factor-2 and -3 in Concanavalin-A-activated mesenchymal stromal cells.

    PubMed

    Zgheib, Alain; Pelletier-Bonnier, Émilie; Levros, Louis-Charles; Annabi, Borhane

    2013-08-01

    Human bone marrow-derived mesenchymal stromal cells (MSCs) express Toll-like receptors (TLRs) and produce cytokines and chemokines, all of which contribute to these cells' immunomodulatory and proangiogenic properties. Among the secreted cytokines, colony-stimulating factors (CSFs) regulate angiogenesis through activation of endothelial cell proliferation and migration. Since MSC are recruited within hypoxic tumors where they signal paracrine-regulated angiogenesis, the aim of this study was to evaluate which CSF members are expressed and are inducible in activated MSC. Furthermore, we investigated the JAK/STAT signal transducing pathway that may impact on CSF transcription. MSC were activated with Concanavalin-A (ConA), a TLR-2/6 agonist as well as a membrane type-1 matrix metalloproteinase (MT1-MMP) inducer, and we found increased transcription of granulocyte macrophage-CSF (GM-CSF, CSF-2), granulocyte CSF (G-CSF, CSF-3), and MT1-MMP. Gene silencing of either STAT3 or MT1-MMP prevented ConA-induced phosphorylation of STAT3, and reversed ConA effects on CSF-2 and CSF-3. Treatment with the Janus Kinase (JAK)2 inhibitor AG490 antagonized the ConA induction of MT1-MMP and CSF-2, while the pan-JAK inhibitor Tofacitinib reversed ConA-induced CSF-2 and -3 gene expression. Silencing of JAK2 prevented the ConA-mediated increase of CSF-2, while silencing of JAK1, JAK3 and TYK2 prevented the increase in CSF-3. Given that combined TLR-activation and locally-produced CSF-2 and CSF-3 could regulate immunomodulation and neovascularization, pharmacological targeting of TLR-2/6-induced MT1-MMP/JAK/STAT3 signalling pathway may prevent MSC contribution to tumor development. Copyright © 2013 Elsevier Ltd. All rights reserved.

  5. Activation of the FGFR-STAT3 pathway in breast cancer cells induces a hyaluronan-rich microenvironment that licenses tumor formation.

    PubMed

    Bohrer, Laura R; Chuntova, Pavlina; Bade, Lindsey K; Beadnell, Thomas C; Leon, Ronald P; Brady, Nicholas J; Ryu, Yungil; Goldberg, Jodi E; Schmechel, Stephen C; Koopmeiners, Joseph S; McCarthy, James B; Schwertfeger, Kathryn L

    2014-01-01

    Aberrant activation of fibroblast growth factor receptors (FGFR) contributes to breast cancer growth, progression, and therapeutic resistance. Because of the complex nature of the FGF/FGFR axis, and the numerous effects of FGFR activation on tumor cells and the surrounding microenvironment, the specific mechanisms through which aberrant FGFR activity contributes to breast cancer are not completely understood. We show here that FGFR activation induces accumulation of hyaluronan within the extracellular matrix and that blocking hyaluronan synthesis decreases proliferation, migration, and therapeutic resistance. Furthermore, FGFR-mediated hyaluronan accumulation requires activation of the STAT3 pathway, which regulates expression of hyaluronan synthase 2 (HAS2) and subsequent hyaluronan synthesis. Using a novel in vivo model of FGFR-dependent tumor growth, we demonstrate that STAT3 inhibition decreases both FGFR-driven tumor growth and hyaluronan levels within the tumor. Finally, our results suggest that combinatorial therapies inhibiting both FGFR activity and hyaluronan synthesis is more effective than targeting either pathway alone and may be a relevant therapeutic approach for breast cancers associated with high levels of FGFR activity. In conclusion, these studies indicate a novel targetable mechanism through which FGFR activation in breast cancer cells induces a protumorigenic microenvironment.

  6. Arecoline induces HA22T/VGH hepatoma cells to undergo anoikis - involvement of STAT3 and RhoA activation

    PubMed Central

    2010-01-01

    Background Our previous study showed that, in basal cell carcinoma cells, arecoline reduces levels of the tumor cell survival factor interleukin-6 (IL-6), increases levels of tumor suppressor factor p53, and elicits cell cycle arrest, followed by apoptosis. In preliminarily studies, we observed that arecoline induces detachment of the human-derived hepatoma cell line HA22T/VGH from the extracellular matrix. In the present study, we explored the fate of the detached HA22T/VGH cells and investigated the underlying mechanism. Methods HA22T/VGH cells or primary cultured rat hepatocytes were treated with arecoline, then changes in morphology, viability, apoptosis, and the expression of surface β1-integrin, apoptosis-related proteins, and IL-6 were examined. Furthermore, activation of the signal transducer and activator of transcription 3 (STAT3) pathway and the RhoA/Rock signaling pathway, including p190RhoGAP and Src homology-2 domain-containing phosphatase SHP2, was examined. Results A low concentration of arecoline (≤ 100 μg/ml) caused cytoskeletal changes in HA22T/VGH cells, but not hepatocytes, and this was accompanied by decreased β1-integrin expression and followed by apoptosis, indicating that HA22T/VGH cells undergo anoikis after arecoline treatment. IL-6 expression and phosphorylation of STAT3, which provides protection against anoikis, were inhibited and levels of downstream signaling proteins, including Bcl-XL and Bcl-2, were decreased, while Bax expression, mitochondrial cytochrome c release, and caspase-3 activity were increased. In addition, phosphorylation/activation of p190RhoGAP, a RhoA inhibitor, and of its upstream regulator, SHP2, was inhibited by arecoline treatment, while Rho/Rock activation was increased. Addition of the RhoA inhibitor attenuated the effects of arecoline. Conclusions This study demonstrated that arecoline induces anoikis of HA22T/VGH cells involving inhibition of STAT3 and increased RhoA/Rock activation and that the STAT3

  7. Arecoline induces HA22T/VGH hepatoma cells to undergo anoikis - involvement of STAT3 and RhoA activation.

    PubMed

    Cheng, Hsiao-Ling; Su, Shu-Jem; Huang, Li-Wen; Hsieh, Bau-Shan; Hu, Yu-Chen; Hung, Thu-Ching; Chang, Kee-Lung

    2010-05-28

    Our previous study showed that, in basal cell carcinoma cells, arecoline reduces levels of the tumor cell survival factor interleukin-6 (IL-6), increases levels of tumor suppressor factor p53, and elicits cell cycle arrest, followed by apoptosis. In preliminarily studies, we observed that arecoline induces detachment of the human-derived hepatoma cell line HA22T/VGH from the extracellular matrix. In the present study, we explored the fate of the detached HA22T/VGH cells and investigated the underlying mechanism. HA22T/VGH cells or primary cultured rat hepatocytes were treated with arecoline, then changes in morphology, viability, apoptosis, and the expression of surface beta1-integrin, apoptosis-related proteins, and IL-6 were examined. Furthermore, activation of the signal transducer and activator of transcription 3 (STAT3) pathway and the RhoA/Rock signaling pathway, including p190RhoGAP and Src homology-2 domain-containing phosphatase SHP2, was examined. A low concentration of arecoline (STAT3, which provides protection against anoikis, were inhibited and levels of downstream signaling proteins, including Bcl-XL and Bcl-2, were decreased, while Bax expression, mitochondrial cytochrome c release, and caspase-3 activity were increased. In addition, phosphorylation/activation of p190RhoGAP, a RhoA inhibitor, and of its upstream regulator, SHP2, was inhibited by arecoline treatment, while Rho/Rock activation was increased. Addition of the RhoA inhibitor attenuated the effects of arecoline. This study demonstrated that arecoline induces anoikis of HA22T/VGH cells involving inhibition of STAT3 and increased RhoA/Rock activation and that the STAT3 and RhoA/Rock signaling pathways

  8. Protective effect of Fenofibrate in renal ischemia reperfusion injury: Involved in suppressing kinase 2 (JAK2)/transcription 3 (STAT3)/p53 signaling activation.

    PubMed

    Lv, J; Wang, X; Liu, S Y; Liang, P F; Feng, M; Zhang, L L; Xu, A P

    2015-12-01

    Renal ischemia reperfusion (I/R) injury is a common reason of acute kidney injury. Apoptosis play an important role in the IRI. Fenofibrate, one of agonist Peroxisome proliferator-activated receptor-alpha (PPARα) has the effect of anti-apoptosis. This study was to explore the effect of Fenofibrate on renal ischemia reperfusion injury and its mechanism. IRI was induced by bilateral renal ischemia for 45 min followed by reperfusion for 24h. Eighteen male C57BL/6 mice were randomly divided into three groups: Sham group (Sham), IRI group (IRI), I/R-Fenofibrate group (FEN). Fenofibrate was injected at 45 min before renal ischemia. Renal histology, function, and the expression of Bax, Bcl-2, Bcl-xl p21, p53, Caspase3, CytC, p-JAK2, p-STAT3 and p-PPAR-α were assessed. Fenofibrate precondition can significantly alleviate the renal dysfunction, the pathological change, up-regulate the expression of p-PPAR-α, Bcl-2, Bcl-xl, Caspase3 and down-regulate the expression of p-JAK2, p-STAT3, p53, p21, CytC and Bax induced by renal IR injury. Fenofibrate precondition can protect mice against IRI by suppressing p53-mediating apoptosis which was associated with inhibiting JAK2/STAT3 signaling activation though further activating PPAR-α. Our findings suggest that Fenofibrate could be useful for preventing IR-induced renal injury. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

  9. Antitumor activity of IL-32β through the activation of lymphocytes, and the inactivation of NF-κB and STAT3 signals

    PubMed Central

    Yun, H-M; Oh, J H; Shim, J-H; Ban, J O; Park, K-R; Kim, J-H; Lee, D H; Kang, J-W; Park, Y H; Yu, D; Kim, Y; Han, S B; Yoon, D-Y; Hong, J T

    2013-01-01

    Cytokine and activation of lymphocytes are critical for tumor growth. We investigated whether interleukin (IL)-32β overexpression changes other cytokine levels and activates cytotoxic lymphocyte, and thus modify tumor growth. Herein, IL-32β inhibited B16 melanoma growth in IL-32β-overexpressing transgenic mice (IL-32β mice), and downregulated the expressions of anti-apoptotic proteins (bcl-2, IAP, and XIAP) and cell growth regulatory proteins (Ki-67 antigen (Ki-67) and proliferating cell nuclear antigen (PCNA)), but upregulated the expressions of pro-apoptotic proteins (bax, cleaved caspase-3, and cleaved caspase-9). IL-32β also inhibited colon and prostate tumor growth in athymic nude mice inoculated with IL-32β-transfected SW620 colon or PC3 prostate cancer cells. The forced expression of IL-32β also inhibited cell growth in cultured colon and prostate cancer cells, and these inhibitory effects were abolished by IL-32 small interfering RNA (siRNA). IL-10 levels were elevated, but IL-1β, IL-6, and tumor necrosis factor-alpha (TNF-α) levels were reduced in the tumor tissues and spleens of IL-32β mice, and athymic nude mice. The number of cytotoxic T (CD8+) and natural killer (NK) cells in tumor tissues, spleen, and blood was significantly elevated in IL-32β mice and athymic nude mice inoculated with IL-32β-transfected cancer cells. Constituted activated NF-κB and STAT3 levels were reduced in the tumor tissues of IL-32β mice and athymic nude mice, as well as in IL-32β-transfected cultured cancer cells. These findings suggest that IL-32β inhibits tumor growth by increasing cytotoxic lymphocyte numbers, and by inactivating the NF-κB and STAT3 pathways through changing of cytokine levels in tumor tissues. PMID:23703385

  10. HAVCR/KIM-1 activates the IL-6/STAT-3 pathway in clear cell renal cell carcinoma and determines tumor progression and patient outcome.

    PubMed

    Cuadros, Thaïs; Trilla, Enric; Sarró, Eduard; Vilà, Maya R; Vilardell, Jordi; de Torres, Inés; Salcedo, Mayte; López-Hellin, Joan; Sánchez, Alex; Ramón y Cajal, Santiago; Itarte, Emilio; Morote, Juan; Meseguer, Anna

    2014-03-01

    Renal cell carcinoma (RCC), the third most prevalent urological cancer, claims more than 100,000 lives/year worldwide. The clear cell variant (ccRCC) is the most common and aggressive subtype of this disease. While commonly asymptomatic, more than 30% of ccRCC are diagnosed when already metastatic, resulting in a 95% mortality rate. Notably, nearly one-third of organ-confined cancers treated by nephrectomy develop metastasis during follow-up care. At present, diagnostic and prognostic biomarkers to screen, diagnose, and monitor renal cancers are clearly needed. The gene encoding the cell surface molecule HAVCR1/KIM-1 is a suggested susceptibility gene for ccRCC and ectodomain shedding of this molecule may be a predictive biomarker of tumor progression. Microarray analysis of 769-P ccRCC-derived cells where HAVCR/KIM-1 levels have been upregulated or silenced revealed relevant HAVCR/KIM-1-related targets, some of which were further analyzed in a cohort of 98 ccRCC patients with 100 month follow-up. We found that HAVCR/KIM-1 activates the IL-6/STAT-3/HIF-1A axis in ccRCC-derived cell lines, which depends on HAVCR/KIM-1 shedding. Moreover, we found that pSTAT-3 S727 levels represented an independent prognostic factor for ccRCC patients. Our results suggest that HAVCR/KIM-1 upregulation in tumors might represent a novel mechanism to activate tumor growth and angiogenesis and that pSTAT-3 S727 is an independent prognostic factor for ccRCC.

  11. The ROS-mediated activation of IL-6/STAT3 signaling pathway is involved in the 27-hydroxycholesterol-induced cellular senescence in nerve cells.

    PubMed

    Liu, Jiao; Liu, Yun; Chen, Juan; Hu, Chunyan; Teng, Mengying; Jiao, Kailin; Shen, Zhaoxia; Zhu, Dongmei; Yue, Jia; Li, Zhong; Li, Yuan

    2017-07-22

    The oxysterol 27-hydroxycholesterol (27HC) is a selective estrogen receptor modulator (SERMs), which like endogenous estrogen 17β-estradiol (E2) induces the proliferation of ER-positive breast cancer cells in vitro. Interestingly, the observation that 27HC induces adverse effects in neural system, distinguishing it from E2. It has been suggested that high levels of circulating cholesterol increase the entry of 27HC into the brain, which may induce learning and memory impairment. Based on this evidence, 27HC may be associated with neurodegenerative processes and interrupted cholesterol homeostasis in the brain. However, the biological events that participate in this process remain largely elusive. In the present study, we demonstrated that 27HC induced apparent cellular senescence in nerve cells. Senescence-associated β-galactosidase (SA-β-Gal) assay revealed that 27HC induced senescence in both BV2 cells and PC12 cells. Furthermore, we demonstrated that 27HC promoted the accumulation of cellular reactive oxygen species (ROS) in nerve cells and subsequently activation of IL-6/STAT3 signaling pathway. Notably, treatment with the ROS scavenger N-acetylcysteine (NAC) markedly blocked 27HC-induced ROS production and activation of IL-6/STAT3 signaling pathway. Either blocking the generation of ROS or inhibition of IL-6/STAT3 both attenuated 27HC-induced cellular senescence. In sum, these findings not only suggested a mechanism whereby 27HC induced cellular senescence in nerve cells, but also helped to recognize the 27HC as a novel harmful factor in neurodegenerative diseases. Copyright © 2017. Published by Elsevier Ltd.

  12. Reduced TCR-dependent activation through citrullination of a T-cell epitope enhances Th17 development by disruption of the STAT3/5 balance.

    PubMed

    Tibbitt, Christopher; Falconer, Jane; Stoop, Jeroen; van Eden, Willem; Robinson, John H; Hilkens, Catharien M U

    2016-07-01

    Citrullination is a post-translational modification of arginine that commonly occurs in inflammatory tissues. Because T-cell receptor (TCR) signal quantity and quality can regulate T-cell differentiation, citrullination within a T-cell epitope has potential implications for T-cell effector function. Here, we investigated how citrullination of an immunedominant T-cell epitope affected Th17 development. Murine naïve CD4(+) T cells with a transgenic TCR recognising p89-103 of the G1 domain of aggrecan (agg) were co-cultured with syngeneic bone marrow-derived dendritic cells (BMDC) presenting the native or citrullinated peptides. In the presence of pro-Th17 cytokines, the peptide citrullinated on residue 93 (R93Cit) significantly enhanced Th17 development whilst impairing the Th2 response, compared to the native peptide. T cells responding to R93Cit produced less IL-2, expressed lower levels of the IL-2 receptor subunit CD25, and showed reduced STAT5 phosphorylation, whilst STAT3 activation was unaltered. IL-2 blockade in native p89-103-primed T cells enhanced the phosphorylated STAT3/STAT5 ratio, and concomitantly enhanced Th17 development. Our data illustrate how a post-translational modification of a TCR contact point may promote Th17 development by altering the balance between STAT5 and STAT3 activation in responding T cells, and provide new insight into how protein citrullination may influence effector Th-cell development in inflammatory disorders. © 2016 The Authors. European Journal of Immunology published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Prostaglandin E1 reduces the keratinocyte toxicity of sorafenib by maintaining signal transducer and activator of transcription 3 (STAT3) activity and enhancing the cAMP response element binding protein (CREB) activity.

    PubMed

    Shichiri, Hiroaki; Yamamoto, Kazuhiro; Tokura, Maya; Ishida, Takahiro; Uda, Atsushi; Bito, Toshinori; Nishigori, Chikako; Nakagawa, Tsutomu; Hirano, Takeshi; Yano, Ikuko; Hirai, Midori

    2017-04-01

    Hand-foot skin reaction (HFSR) is a common side effect of multiple tyrosine kinase inhibitors (mTKIs). HFSR can necessitate dose reductions or interruption of therapy owing to its negative effect on the quality of life. Therefore, effective use of mTKIs requires measures to prevent HFSR. We evaluated the effect of prostaglandin E1 (PGE1) on HFSR, because PGE1 is already used to treat bed sores and skin ulcers and has established angiogenic and antiproliferative effects in keratinocytes. We found that the pathogenesis of sorafenib-induced HFSR is characterized by a decrease in levels of a phosphorylated signal transducer and activator of transcription 3 (STAT3). We investigated the effect of PGE1 on the sorafenib-mediated reduction in phosphorylated STAT3 levels in HaCaT human epidermal keratinocytes. In cells treated with sorafenib, phosphorylated STAT3 levels decreased in a concentration-dependent manner, and this effect was blocked in cells treated with sorafenib and PGE1. Furthermore, the expression of phosphorylated STAT3, the antiapoptotic proteins myeloid cell leukemia-1 (Mcl-1) and survivin decreased in cells pretreated with an inhibitor of cAMP response element binding protein (CREB). Cell viability increased in cells treated with sorafenib and PGE1 compared with that in cells treated with sorafenib alone, and these effects were not observed in STAT3 knockdown HaCaT cells. Collectively, these findings indicate that PGE1 blocks the inhibitory effects of sorafenib on cell growth by maintaining the activity of STAT3 and enhancing the CREB activity. Therefore, PGE1 might represent an effective treatment for the prevention of sorafenib-induced HFSR. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. AZD9150, a Next-Generation Antisense Oligonucleotide Inhibitor of STAT3 with Early Evidence of Clinical Activity in Lymphoma and Lung Cancer

    PubMed Central

    Hong, David; Kurzrock, Razelle; Kim, Youngsoo; Woessner, Richard; Younes, Anas; Nemunaitis, John; Fowler, Nathan; Zhou, Tianyuan; Schmidt, Joanna; Jo, Minji; Lee, Samantha J.; Yamashita, Mason; Hughes, Steven G.; Fayad, Luis; Piha-Paul, Sarina; Nadella, Murali VP; Mohseni, Morvarid; Lawson, Deborah; Reimer, Corinne; Blakey, David C.; Xiao, Xiaokun; Hsu, Jeff; Revenko, Alexey; Monia, Brett P.; MacLeod, A. Robert

    2017-01-01

    Next-generation sequencing technologies have greatly expanded our understanding of cancer genetics. Antisense technology is an attractive platform with the potential to translate these advances into improved cancer therapeutics, because antisense oligonucleotide (ASO) inhibitors can be designed on the basis of gene sequence information alone. Recent human clinical data have demonstrated the potent activity of systemically administered ASOs targeted to genes expressed in the liver. Here, we describe the preclinical activity and initial clinical evaluation of a class of ASOs containing constrained ethyl modifications for targeting the gene encoding the transcription factor STAT3, a notoriously difficult protein to inhibit therapeutically. Systemic delivery of the unformulated ASO, AZD9150, decreased STAT3 expression in a broad range of preclinical cancer models and showed antitumor activity in lymphoma and lung cancer models. AZD9150 preclinical activity translated into single-agent antitumor activity in patients with highly treatment-refractory lymphoma and non-small cell lung cancer in a phase I dose escalation study. PMID:26582900

  15. Mechanisms of Unphosphorylated STAT3 Transcription Factor Binding to DNA*

    PubMed Central

    Timofeeva, Olga A.; Chasovskikh, Sergey; Lonskaya, Irina; Tarasova, Nadya I.; Khavrutskii, Lyuba; Tarasov, Sergey G.; Zhang, Xueping; Korostyshevskiy, Valeriy R.; Cheema, Amrita; Zhang, Lihua; Dakshanamurthy, Sivanesan; Brown, Milton L.; Dritschilo, Anatoly

    2012-01-01

    Phosphorylation of signal transducer and activator of transcription 3 (STAT3) on a single tyrosine residue in response to growth factors, cytokines, interferons, and oncogenes activates its dimerization, translocation to the nucleus, binding to the interferon γ (gamma)-activated sequence (GAS) DNA-binding site and activation of transcription of target genes. STAT3 is constitutively phosphorylated in various cancers and drives gene expression from GAS-containing promoters to promote tumorigenesis. Recently, roles for unphosphorylated STAT3 (U-STAT3) have been described in response to cytokine stimulation, in cancers, and in maintenance of heterochromatin stability. However, the mechanisms underlying U-STAT3 binding to DNA has not been fully investigated. Here, we explore STAT3-DNA interactions by atomic force microscopy (AFM) imaging. We observed that U-STAT3 molecules bind to the GAS DNA-binding site as dimers and monomers. In addition, we observed that U-STAT3 binds to AT-rich DNA sequence sites and recognizes specific DNA structures, such as 4-way junctions and DNA nodes, within negatively supercoiled plasmid DNA. These structures are important for chromatin organization and our data suggest a role for U-STAT3 as a chromatin/genome organizer. Unexpectedly, we found that a C-terminal truncated 67.5-kDa STAT3 isoform recognizes single-stranded spacers within cruciform structures that also have a role in chromatin organization and gene expression. This isoform appears to be abundant in the nuclei of cancer cells and, therefore, may have a role in regulation of gene expression. Taken together, our data highlight novel mechanisms by which U-STAT3 binds to DNA and supports U-STAT3 function as a transcriptional activator and a chromatin/genomic organizer. PMID:22378781

  16. Targeting STAT3 in cancer: how successful are we?

    PubMed Central

    Yue, Peibin; Turkson, James

    2008-01-01

    Background Aberrant activation of the signal transducer and activator of transcription (STAT)3 occurs in many human tumors. Moreover, studies utilizing genetic and pharmacological approaches to modulate constitutive STAT3 activity have provided compelling evidence for the critical role of aberrant STAT3 activity in malignant transformation and tumor progression, and thereby validated STAT3 as a novel cancer drug target. Objective This review is intended to be a full coverage of the efforts to develop direct STAT3 inhibitors and will provide a discussion on the inhibitory modalities developed to date. Methods Review of the literature focused on the modalities and mechanisms that directly target and inhibit the STAT protein or its functions. Results/conclusion While a variety of STAT3 inhibitors have been identified that induce antitumor cell effects in vitro and in vivo, the landscape remains murky. With a few exceptions, most of the STAT3 inhibitors reported to date have not undergone an in vivo efficacy, pharmacology or toxicity testing. Also, there is no evidence, per the published literature of an impending clinical development for the few agents that were reported to exhibit in vivo efficacy. Overall, there is the need for a reassessment of the ongoing strategies to target STAT3 intended not only for refinement, but also for incorporating some new technologies to strengthen our efforts and ensure the success – sooner, rather than later – of identifying suitable anti-STAT3 agents for development into clinically useful anticancer therapeutics. PMID:19053881

  17. Cannabinoid receptor CB1 regulates STAT3 activity and its expression dictates the responsiveness to SR141716 treatment in human glioma patients' cells

    PubMed Central

    Ciaglia, Elena; Torelli, Giovanni; Pisanti, Simona; Picardi, Paola; D'Alessandro, Alba; Laezza, Chiara; Malfitano, Anna Maria; Fiore, Donatella; Zottola, Antonio Christian Pagano; Proto, Maria Chiara; Catapano, Giuseppe; Gazzerro, Patrizia; Bifulco, Maurizio

    2015-01-01

    Herein we show that a majority of human brain tumor samples and cell lines over-expressed cannabinoid receptor CB1 as compared to normal human astrocytes (NHA), while uniformly expressed low levels of CB2. This finding prompted us to investigate the therapeutic exploitation of CB1 inactivation by SR141716 treatment, with regard to its direct and indirect cell-mediated effects against gliomas. Functional studies, using U251MG glioma cells and primary tumor cell lines derived from glioma patients expressing different levels of CB1, highlighted SR141716 efficacy in inducing apoptosis via G1 phase stasis and block of TGF-β1 secretion through a mechanism that involves STAT3 inhibition. According to the multivariate role of STAT3 in the immune escape too, interestingly SR141716 lead also to the functional and selective expression of MICA/B on the surface of responsive malignant glioma cells, but not on NHA. This makes SR141716 treated-glioma cells potent targets for allogeneic NK cell-mediated recognition through a NKG2D restricted mechanism, thus priming them for NK cell antitumor reactivity. These results indicate that CB1 and STAT3 participate in a new oncogenic network in the complex biology of glioma and their expression levels in patients dictate the efficacy of the CB1 antagonist SR141716 in multimodal glioma destruction. SIGNIFICANCE CB1 is implicated in the regulation of cellular processes linked to survival, proliferation, invasion and angiogenesis in several physio-pathological conditions. We shed light on previously unrecognized molecular mechanism of CB1-mediated modulation of human glioma progression and provide the first and original demonstration of CB1-STAT3 axis as a new target and predictor biomarkers of the benefit from specific therapies. Indeed CB1 antagonism capable of tumoral cell division' control while making the glioma immunovisible and engaging the immune system to fight it may represent a hopeful alternative to other established

  18. Inhibition of STAT3 by Anticancer Drug Bendamustine

    PubMed Central

    Iwamoto, Kazunori; Uehara, Yutaka; Inoue, Yukie; Taguchi, Kyoko; Muraoka, Daisuke; Ogo, Naohisa; Matsuno, Kenji; Asai, Akira

    2017-01-01

    Bendamustine (BENDA), which bears the bis(2-chloroethyl)amino moiety, is an alkylating agent that stops the growth of cancer cells by binding to DNA and interfering with its replication. However, the mechanism of action underlying its excellent clinical efficacy remains unclear. In this work, we report that BENDA inhibits signal transducer and activator of transcription 3 (STAT3). In an AlphaScreen-based biochemical assay using recombinant human STAT3, binding of STAT3–Src homology 2 (SH2) to the phosphotyrosine (pTyr, pY) peptide was inhibited by BENDA but not by the inactive metabolite dihydroxy bendamustine (HP2). When a single point mutation of C550A or C712A was introduced into recombinant human STAT3, its sensitivity to BENDA was substantially reduced, suggesting that these cysteine residues are important for BENDA to inhibit STAT3. Furthermore, BENDA suppressed the function of cellular STAT3 as a transcriptional activator in a human breast cancer cell line, MDA-MB-468, with constitutively activated STAT3. A competitive pull-down assay using biotinylated BENDA (Bio-BENDA) revealed that BENDA bound tightly to cellular STAT3, presumably through covalent bonds. Therefore, our results suggest that the anticancer effects of BENDA may be associated, at least in part, with its inhibitory effect on the SH2 domain of STAT3. PMID:28125678

  19. CD147 promotes Src-dependent activation of Rac1 signaling through STAT3/DOCK8 during the motility of hepatocellular carcinoma cells

    PubMed Central

    Zhao, Pu; Zhang, Yang; Fu, Zhi-Guang; Chen, Zhi-Nan; Jiang, Jian-Li

    2015-01-01

    Metastasis is considered a dynamic process in tumor development that is related to abnormal migration and invasion. Tumor cells can move as individual cells in two interconvertible modes: mesenchymal-type and amoeboid. Previously, we reported that the interaction between CD147 and Annexin II can inhibit the amoeboid movement in hepatocellular carcinoma (HCC) cells. However, the mechanism of CD147 involved in mesenchymal movement is still unclear. Notably, our results show overexpression of CD147 led to mesenchymal-type movement in HCC cells. Evidence indicated that the mesenchymal-type cell movement induced by CD147 was Src dependent, as observed by confocal microscopy and Rac1 activity assay. The phosphorylation of Src (pY416-Src) can be up-regulated by CD147, and this regulation is mediated by focal adhesion kinase (FAK). Next, we identified DOCK8 as a GEF for Rac1, a key molecule driving mesenchymal-type movement. We also found that Src promotes STAT3 phosphorylation and STAT3 facilitates DOCK8 transcription, thus enhancing DOCK8 expression and Rac1 activation. This study provides a novel mechanism of CD147 regulating mesenchymal-type movement in HCC cells. PMID:25428919

  20. The mitochondrial-derived peptide humanin activates the ERK1/2, AKT, and STAT3 signaling pathways and has age-dependent signaling differences in the hippocampus

    PubMed Central

    Kim, Su-Jeong; Guerrero, Noel; Wassef, Gabriella; Xiao, Jialin; Mehta, Hemal H.; Cohen, Pinchas; Yen, Kelvin

    2016-01-01

    Humanin is a small secreted peptide that is encoded in the mitochondrial genome. Humanin and its analogues have a protective role in multiple age-related diseases including type 2 diabetes and Alzheimer's disease, through cytoprotective and neuroprotective effects both in vitro and in vivo. However, the humanin-mediated signaling pathways are not well understood. In this paper, we demonstrate that humanin acts through the GP130/IL6ST receptor complex to activate AKT, ERK1/2, and STAT3 signaling pathways. Humanin treatment increases phosphorylation in AKT, ERK 1/2, and STAT3 where PI3K, MEK, and JAK are involved in the activation of those three signaling pathways, respectively. Furthermore, old mice, but not young mice, injected with humanin showed an increase in phosphorylation in AKT and ERK1/2 in the hippocampus. These findings uncover a key signaling pathway of humanin that is important for humanin's function and also demonstrates an age-specific in vivo effect in a region of the brain that is critical for memory formation in an age-dependent manner. PMID:27384491

  1. CD147 promotes Src-dependent activation of Rac1 signaling through STAT3/DOCK8 during the motility of hepatocellular carcinoma cells.

    PubMed

    Wang, Shi-Jie; Cui, Hong-Yong; Liu, Yan-Mei; Zhao, Pu; Zhang, Yang; Fu, Zhi-Guang; Chen, Zhi-Nan; Jiang, Jian-Li

    2015-01-01

    Metastasis is considered a dynamic process in tumor development that is related to abnormal migration and invasion. Tumor cells can move as individual cells in two interconvertible modes: mesenchymal-type and amoeboid. Previously, we reported that the interaction between CD147 and Annexin II can inhibit the amoeboid movement in hepatocellular carcinoma (HCC) cells. However, the mechanism of CD147 involved in mesenchymal movement is still unclear. Notably, our results show overexpression of CD147 led to mesenchymal-type movement in HCC cells. Evidence indicated that the mesenchymal-type cell movement induced by CD147 was Src dependent, as observed by confocal microscopy and Rac1 activity assay. The phosphorylation of Src (pY416-Src) can be up-regulated by CD147, and this regulation is mediated by focal adhesion kinase (FAK). Next, we identified DOCK8 as a GEF for Rac1, a key molecule driving mesenchymal-type movement. We also found that Src promotes STAT3 phosphorylation and STAT3 facilitates DOCK8 transcription, thus enhancing DOCK8 expression and Rac1 activation. This study provides a novel mechanism of CD147 regulating mesenchymal-type movement in HCC cells.

  2. Elevated level of Interleukin-35 in colorectal cancer induces conversion of T cells into iTr35 by activating STAT1/STAT3

    PubMed Central

    Xie, Guohua; Zhou, Yunlan; Yue, Chaoyan; Yuan, Xiangliang; Zheng, Yingxia; Wang, Weiwei; Deng, Lin; Shen, Lisong

    2016-01-01

    IL-35 is a novel heterodimeric and inhibitory cytokine, composed of interleukin-12 subunit alpha (P35) and Epstein-Barr virus -induced gene 3 (EBI3). IL-35 has been reported to be produced by a range of cell types, especially regulatory T cells, and to exert immunosuppressive effects via the STATx signaling pathway. In this study, we demonstrated that IL-35 expression was elevated in both serum and tumors in patients with colorectal cancer. IL-35 mainly expressed in CD4+ T cells in human colorectal cancer tumors and adjacent tissues. Increased IL-35 expression in tumor-adjacent tissues was significantly associated with tumor metastasis. IL-35 inhibited the proliferation of CD4+CD25− T effector cells in vitro in a dose-dependent manner, and its suppression was partially reversed by applying IL-35-neutralizing antibodies. IL-35 treatment activated the phosphorylation of both STAT1 and STAT3 in human CD4+ T cells. Meanwhile, IL-35 induced a positive feedback loop to promote its own production. We observed that Tregs obtained from colorectal cancer patients were capable of inducing more IL-35 production. In addition, EBI3 promoter-driven luciferase activity was higher than that of the mock plasmid after IL-35stimulation. Thus, our study indicates that the high level of IL-35 in colorectal cancer promotes the production of IL-35 via STAT1 and STAT3, which suppresses T cell proliferation and may participate in tumor immunotolerance. PMID:27682874

  3. Pyranocoumarins isolated from Peucedanum praeruptorum Dunn suppress lipopolysaccharide-induced inflammatory response in murine macrophages through inhibition of NF-κB and STAT3 activation.

    PubMed

    Yu, Peng-Jiu; Jin, Hong; Zhang, Jun-Yan; Wang, Guang-Fa; Li, Jing-Rong; Zhu, Zheng-Guang; Tian, Yuan-Xin; Wu, Shao-Yu; Xu, Wei; Zhang, Jia-Jie; Wu, Shu-Guang

    2012-06-01

    Praeruptorin C, D, and E (PC, PD, and PE) are three pyranocoumarins isolated from the dried root of Peucedanum praeruptorum Dunn of Umbelliferae. In the present study, we investigated the anti-inflammatory effect of these compounds in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophage cells. Pyranocoumarins significantly inhibited LPS-induced production of nitric oxide, interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α). The mRNA and protein expressions of inducible nitric oxide synthase, IL-6, and TNF-α were also suppressed by these compounds. Both PD and PE exhibited greater anti-inflammatory activities than PC. Further study showed that pyranocoumarins suppressed the cytoplasmic loss of inhibitor κB-α protein and inhibited the translocation of NF-κB from cytoplasm to nucleus. In addition, pyranocoumarins suppressed LPS-induced STAT3 tyrosine phosphorylation. Taken together, the results suggest that pyranocoumarins may exert anti-inflammatory effects in LPS-stimulated RAW 264.7 macrophages through the inhibition of NF-κB and STAT3 activation.

  4. The mitochondrial-derived peptide humanin activates the ERK1/2, AKT, and STAT3 signaling pathways and has age-dependent signaling differences in the hippocampus.

    PubMed

    Kim, Su-Jeong; Guerrero, Noel; Wassef, Gabriella; Xiao, Jialin; Mehta, Hemal H; Cohen, Pinchas; Yen, Kelvin

    2016-07-26

    Humanin is a small secreted peptide that is encoded in the mitochondrial genome. Humanin and its analogues have a protective role in multiple age-related diseases including type 2 diabetes and Alzheimer's disease, through cytoprotective and neuroprotective effects both in vitro and in vivo. However, the humanin-mediated signaling pathways are not well understood. In this paper, we demonstrate that humanin acts through the GP130/IL6ST receptor complex to activate AKT, ERK1/2, and STAT3 signaling pathways. Humanin treatment increases phosphorylation in AKT, ERK 1/2, and STAT3 where PI3K, MEK, and JAK are involved in the activation of those three signaling pathways, respectively. Furthermore, old mice, but not young mice, injected with humanin showed an increase in phosphorylation in AKT and ERK1/2 in the hippocampus. These findings uncover a key signaling pathway of humanin that is important for humanin's function and also demonstrates an age-specific in vivo effect in a region of the brain that is critical for memory formation in an age-dependent manner.

  5. Porcine Reproductive and Respiratory Syndrome Virus Antagonizes JAK/STAT3 Signaling via nsp5, Which Induces STAT3 Degradation.

    PubMed

    Yang, Liping; Wang, Rong; Ma, Zexu; Xiao, Yueqiang; Nan, Yuchen; Wang, Yu; Lin, Shaoli; Zhang, Yan-Jin

    2017-02-01

    Signal transducer and activator of transcription 3 (STAT3) is a pleiotropic signaling mediator of many cytokines, including interleukin-6 (IL-6) and IL-10. STAT3 is known to play critical roles in cell growth, proliferation, differentiation, immunity and inflammatory responses. The objective of this study was to determine the effect of porcine reproductive and respiratory syndrome virus (PRRSV) infection on the STAT3 signaling since PRRSV induces a weak protective immune response in host animals. We report here that PRRSV infection of MARC-145 cells and primary porcine pulmonary alveolar macrophages led to significant reduction of STAT3 protein level. Several strains of both PRRSV type 1 and type 2 led to a similar reduction of STAT3 protein level but had a minimal effect on its transcripts. The PRRSV-mediated STAT3 reduction was in a dose-dependent manner as the STAT3 level decreased, along with incremental amounts of PRRSV inocula. Further study showed that nonstructural protein 5 (nsp5) of PRRSV induced the STAT3 degradation by increasing its polyubiquitination level and shortening its half-life from 24 h to ∼3.5 h. The C-terminal domain of nsp5 was shown to be required for the STAT3 degradation. Moreover, the STAT3 signaling in the cells transfected with nsp5 plasmid was significantly inhibited. These results indicate that PRRSV antagonizes the STAT3 signaling by accelerating STAT3 degradation via the ubiquitin-proteasomal pathway. This study provides insight into the PRRSV interference with the JAK/STAT3 signaling, leading to perturbation of the host innate and adaptive immune responses. The typical features of immune responses in PRRSV-infected pigs are delayed onset and low levels of virus neutralizing antibodies, as well as weak cell-mediated immunity. Lymphocyte development and differentiation rely on cytokines, many of which signal through the JAK/STAT signaling pathway to exert their biological effects. Here, we discovered that PRRSV antagonizes the

  6. Study of the UV Light Conversion of Feruloyl Amides from Portulaca oleracea and Their Inhibitory Effect on IL-6-Induced STAT3 Activation.

    PubMed

    Hwang, Joo Tae; Kim, Yesol; Jang, Hyun-Jae; Oh, Hyun-Mee; Lim, Chi-Hwan; Lee, Seung Woong; Rho, Mun-Chual

    2016-06-30

    Two new feruloyl amides, N-cis-hibiscusamide (5) and (7'S)-N-cis-feruloylnormetanephrine (9), and eight known feruloyl amides were isolated from Portulaca oleracea L. and the geometric conversion of the ten isolated feruloyl amides by UV light was verified. The structures of the feruloyl amides were determined based on spectroscopic data and comparison with literature data. The NMR data revealed that the structures of the isolated compounds showed cis/trans-isomerization under normal laboratory light conditions. Therefore, cis and trans-isomers of feruloyl amides were evaluated for their convertibility and stability by UV light of a wavelength of 254 nm. After 96 h of UV light exposure, 23.2%-35.0% of the cis and trans-isomers were converted to trans-isomers. Long-term stability tests did not show any significant changes. Among all compounds and conversion mixtures collected, compound 6 exhibited the strongest inhibition of IL-6-induced STAT3 activation in Hep3B cells, with an IC50 value of 0.2 μM. This study is the first verification of the conversion rates and an equilibrium ratio of feruloyl amides. These results indicate that this natural material might provide useful information for the treatment of various diseases involving IL-6 and STAT3.

  7. Roscovitine attenuates intimal hyperplasia via inhibiting NF-κB and STAT3 activation induced by TNF-α in vascular smooth muscle cells.

    PubMed

    He, Ming; Wang, Chao; Sun, Jia-Huan; Liu, Yu; Wang, Hong; Zhao, Jing-Shan; Li, Yun-Feng; Chang, Hong; Hou, Jian-Ming; Song, Jun-Na; Li, Ai-Ying; Ji, En-Sheng

    2017-08-01

    Roscovitine is a selective CDK inhibitor originally designed as anti-cancer agent, which has also been shown to inhibit proliferation in vascular smooth muscle cells (VSMCs). However, its effect on vascular remodeling and its mechanism of action remain unknown. In our study, we created a new intimal hyperplasia model in male Sprague-Dawley rats by trypsin digestion method, which cause to vascular injury as well as the model of rat carotid balloon angioplasty. Roscovitine administration led to a significant reduction in neointimal formation and VSMCs proliferation after injury in rats. Western blot analysis revealed that, in response to vascular injury, TNF-α stimulation induced p65 and STAT3 phosphorylation and promoted translocation of these molecules into the nucleus. p65 can physically associate with STAT3 and bind to TNF-α-regulated target promoters, such as MCP-1 and ICAM-1, to initiate gene transcription. Roscovitine can interrupt activation of NF-κB and reduce expression of TNF-α-induced proinflammatory gene, thus inhibiting intimal hyperplasia. These findings provide a novel mechanism to explain the roscovitine-mediated inhibition of intimal hyperplasia induced by proinflammatory pathways. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Discovery of Potent Anticancer Agent HJC0416, an Orally Bioavailable Small Molecule Inhibitor of Signal Transducer and Activator of Transcription 3 (STAT3)

    PubMed Central

    Chen, Haijun; Yang, Zhengduo; Ding, Chunyong; Xiong, Ailian; Wild, Christopher; Wang, Lili; Ye, Na; Cai, Guoshuai; Flores, Rudolfo M.; Ding, Ye; Shen, Qiang; Zhou, Jia

    2014-01-01

    In a continuing effort to develop orally bioavailable small-molecule STAT3 inhibitors as potential therapeutic agents for human cancer, a series of novel diversified analogues based on our identified lead compound HJC0149 (1) (5-chloro-N-(1,1-dioxo-1H-1λ6-benzo[b]thiophen-6-yl)-2-hydroxybenzamide, Eur. J. Med. Chem. 2013, 62, 498–507) have been rationally designed, synthesized, and pharmacologically evaluated. Molecular docking studies and biological characterization supported our earlier findings that the O-alkylamino-tethered side chain on the hydroxyl group is an effective and essential structural determinant for improving biological activities and druglike properties of these molecules. Compounds with such modifications exhibited potent antiproliferative effects against breast and pancreatic cancer cell lines with IC50 values from low micromolar to nanomolar range. Among them, the newly discovered STAT3 inhibitor 12 (HJC0416) displayed an intriguing anticancer profile both in vitro and in vivo (i.p. & p.o.). More importantly, HJC0416 is an orally bioavailable anticancer agent as a promising candidate for further development. PMID:24904966

  9. The ROS-mediated activation of STAT-3/VEGF signaling is involved in the 27-hydroxycholesterol-induced angiogenesis in human breast cancer cells.

    PubMed

    Zhu, Dongmei; Shen, Zhaoxia; Liu, Jiao; Chen, Juan; Liu, Yun; Hu, Chunyan; Li, Zhong; Li, Yuan

    2016-12-15

    Breast cancer (BC) is the leading cause of cancer-related mortality among females worldwide, and angiogenesis plays a crucial role in BC progression. 27-Hydroxycholesterol (27HC) is an endogenous selective estrogen receptor modulator, which promotes the growth and metastasis of BC. Here, we further found that, 27HC improved the angiogenic ability of BC in a VEGF-dependent manner. For the molecular mechanisms, on one hand, as an estrogen-like factor, 27HC enhanced the expression of VEGF by the classical ERα/VEGF signaling in ER-positive BC cells; on the other hand, in both ER-positive and ER-negative BC cells, 27HC enhanced the generation of ROS, which in turn activated the STAT-3/VEGF signaling in an ER independent manner. Either blocking the generation of ROS or knockdown of STAT-3 attenuated the 27HC-induced autocrine of VEGF and angiogenesis. These findings not only suggested a mechanism whereby 27HC enhanced the angiogenesis, but also helped to recognize the 27HC as a novel potential harmful factor in BC, especially in the menopause patients.

  10. STAT3 in Cancer—Friend or Foe?

    PubMed Central

    Zhang, Hai-Feng; Lai, Raymond

    2014-01-01

    The roles and significance of STAT3 in cancer biology have been extensively studied for more than a decade. Mounting evidence has shown that constitutive activation of STAT3 is a frequent biochemical aberrancy in cancer cells, and this abnormality directly contributes to tumorigenesis and shapes many malignant phenotypes in cancer cells. Nevertheless, results from more recent experimental and clinicopathologic studies have suggested that STAT3 also can exert tumor suppressor effects under specific conditions. Importantly, some of these studies have demonstrated that STAT3 can function either as an oncoprotein or a tumor suppressor in the same cell type, depending on the specific genetic background or presence/absence of specific coexisting biochemical defects. Thus, in the context of cancer biology, STAT3 can be a friend or foe. In the first half of this review, we will highlight the “evil” features of STAT3 by summarizing its oncogenic functions and mechanisms. The differences between the canonical and non-canonical pathway will be highlighted. In the second half, we will summarize the evidence supporting that STAT3 can function as a tumor suppressor. To explain how STAT3 may mediate its tumor suppressor effects, we will discuss several possible mechanisms, one of which is linked to the role of STAT3β, one of the two STAT3 splicing isoforms. Taken together, it is clear that the roles of STAT3 in cancer are multi-faceted and far more complicated than one appreciated previously. The new knowledge has provided us with new approaches and strategies when we evaluate STAT3 as a prognostic biomarker or therapeutic target. PMID:24995504

  11. Imatinib Reverses Doxorubicin Resistance by Affecting Activation of STAT3-Dependent NF-κB and HSP27/p38/AKT Pathways and by Inhibiting ABCB1

    PubMed Central

    Sims, Jonathan T.; Ganguly, Sourik S.; Bennett, Holly; Friend, J. Woodrow; Tepe, Jessica; Plattner, Rina

    2013-01-01

    Despite advances in cancer detection and prevention, a diagnosis of metastatic disease remains a death sentence due to the fact that many cancers are either resistant to chemotherapy (conventional or targeted) or develop resistance during treatment, and residual chemoresistant cells are highly metastatic. Metastatic cancer cells resist the effects of chemotherapeutic agents by upregulating drug transporters, which efflux the drugs, and by activating proliferation and survival signaling pathways. Previously, we found that c-Abl and Arg non-receptor tyrosine kinases are activated in breast cancer, melanoma, and glioblastoma cells, and promote cancer progression. In this report, we demonstrate that the c-Abl/Arg inhibitor, imatinib (imatinib mesylate, STI571, Gleevec), reverses intrinsic and acquired resistance to the anthracycline, doxorubicin, by inducing G2/M arrest and promoting apoptosis in cancer cells expressing highly active c-Abl and Arg. Significantly, imatinib prevents intrinsic resistance by promoting doxorubicin-mediated NF-κB/p65 nuclear localization and repression of NF-κB targets in a STAT3-dependent manner, and by preventing activation of a novel STAT3/HSP27/p38/Akt survival pathway. In contrast, imatinib prevents acquired resistance by inhibiting upregulation of the ABC drug transporter, ABCB1, directly inhibiting ABCB1 function, and abrogating survival signaling. Thus, imatinib inhibits multiple novel chemoresistance pathways, which indicates that it may be effective in reversing intrinsic and acquired resistance in cancers containing highly active c-Abl and Arg, a critical step in effectively treating metastatic disease. Furthermore, since imatinib converts a master survival regulator, NF-κB, from a pro-survival into a pro-apoptotic factor, our data suggest that NF-κB inhibitors may be ineffective in sensitizing tumors containing activated c-Abl/Arg to anthracyclines, and instead might antagonize anthracycline-induced apoptosis. PMID:23383209

  12. Targeting cancer initiating cells by promoting cell differentiation and restoring chemosensitivity via dual inactivation of STAT3 and Src activity using an active component of Antrodia cinnamomea mycelia

    PubMed Central

    Chan, Ik-On; Chen, Chin-Chu; Sheu, Sen-Je; Lin, Ting-wei; Chou, Shiu-Huey; Liu, Chung-Ji; Lee, Te-Chang; Lo, Jeng-Fan

    2016-01-01

    Cancer initiating cells (CICs) represent a subpopulation of cancer cells, which are responsible for tumor growth and resistance to chemotherapy. Herein, we first used a cell-based aldehyde dehydrogenase (ALDH) activity assay to identify that YMGKI-2 (also named as Ergone), an active component purified from Antrodia cinnamomea Mycelia extract (ACME), effectively abrogated the ALDH activity and abolished the CICs in head and neck squamous cell carcinoma cells (HNSCCs). Consequently, YMGKI-2 treatment suppressed self-renewal ability and expression of stemness signature genes (Oct-4 and Nanog) of sphere cells with enriched CICs. Moreover, YMGKI-2 treated sphere cells displayed reduction of CICs properties and promotion of cell differentiation, but not significant cytotoxicity. YMGKI-2 treatment also attenuated the tumorigenicity of HNSCC cells in vivo. Mechanistically, treatment of YMGKI-2 resulted in inactivation of STAT3 and Src. Lastly, combinatorial treatments with YMGKI-2 and standard chemotherapeutic drugs (cisplatin or Fluorouracil) restored the chemosensivity on sphere cells and cisplatin-resistant HNSCC cells. Together, we demonstrate that YMGKI-2 treatment effectively induces differentiation and reduces tumorigenicity of CICs. Further, combined treatment of YMGKI-2 and conventional chemotherapy can overcome chemoresistance. These results suggest that YMGKI-2 treatment may be used to improve future clinical responses in head and neck cancer treatment through targeting CICs. PMID:27682875

  13. p-21 activated kinase 4 (PAK4) maintains stem cell-like phenotypes in pancreatic cancer cells through activation of STAT3 signaling

    PubMed Central

    Tyagi, Nikhil; Marimuthu, Saravanakumar; Bhardwaj, Arun; Deshmukh, Sachin K.; Srivastava, Sanjeev K.; Singh, Ajay P.; McClellan, Steven; Carter, James E.; Singh, Seema

    2015-01-01

    Pancreatic cancer (PC) remains a highly lethal malignancy due to its unusual chemoresistance and high aggressiveness. A subpopulation of pancreatic tumor cells, known as cancer stem cells (CSCs), is considered responsible not only for tumor-maintenance, but also for its widespread metastasis and therapeutic failure. Here we investigated the role of p-21 activated kinase 4 (PAK4) in driving PC stemness properties. Our data demonstrate that triple-positive (CD24+/CD44+/EpCAM+) subpopulation of pancreatic CSCs exhibits greater level of PAK4 as compared to triple-negative (CD24−/CD44−/EpCAM−) cells. Moreover, PAK4 silencing in PC cells leads to diminished fraction of CD24, CD44, and EpCAM positive cells. Furthermore, we show that PAK4-silenced PC cells exhibit decreased sphere-forming ability and increased chemo-sensitivity to gemcitabine toxicity. PAK4 expression is also associated with enhanced levels of stemness-associated transcription factors (Oct4/Nanog/Sox2 and KLF4). Furthermore, our data show decreased nuclear accumulation and transcriptional activity of STAT3 in PAK4-silenced PC cells and restitution of its activity leads to restoration of stem cell phenotypes. Together, our findings deliver first experimental evidence for the involvement of PAK4 in PC stemness and support its clinical utility as a novel therapeutic target in PC. PMID:26546043

  14. Targeting cancer initiating cells by promoting cell differentiation and restoring chemosensitivity via dual inactivation of STAT3 and src activity using an active component of antrodia cinnamomea mycelia.

    PubMed

    Chang, Ching-Wen; Chen, Yu-Syuan; Chen, Chien-Chih; Chan, Ik-On; Chen, Chin-Chu; Sheu, Sen-Je; Lin, Ting-Wei; Chou, Shiu-Huey; Liu, Chung-Ji; Lee, Te-Chang; Lo, Jeng-Fan

    2016-11-08

    Cancer initiating cells (CICs) represent a subpopulation of cancer cells, which are responsible for tumor growth and resistance to chemotherapy. Herein, we first used a cell-based aldehyde dehydrogenase (ALDH) activity assay to identify that YMGKI-2 (also named as Ergone), an active component purified from Antrodia cinnamomea Mycelia extract (ACME), effectively abrogated the ALDH activity and abolished the CICs in head and neck squamous cell carcinoma cells (HNSCCs). Consequently, YMGKI-2 treatment suppressed self-renewal ability and expression of stemness signature genes (Oct-4 and Nanog) of sphere cells with enriched CICs. Moreover, YMGKI-2 treated sphere cells displayed reduction of CICs properties and promotion of cell differentiation, but not significant cytotoxicity. YMGKI-2 treatment also attenuated the tumorigenicity of HNSCC cells in vivo. Mechanistically, treatment of YMGKI-2 resulted in inactivation of STAT3 and Src. Lastly, combinatorial treatments with YMGKI-2 and standard chemotherapeutic drugs (cisplatin or Fluorouracil) restored the chemosensivity on sphere cells and cisplatin-resistant HNSCC cells. Together, we demonstrate that YMGKI-2 treatment effectively induces differentiation and reduces tumorigenicity of CICs. Further, combined treatment of YMGKI-2 and conventional chemotherapy can overcome chemoresistance. These results suggest that YMGKI-2 treatment may be used to improve future clinical responses in head and neck cancer treatment through targeting CICs.

  15. Activation of tumor suppressor LKB1 by honokiol abrogates cancer stem-like phenotype in breast cancer via inhibition of oncogenic Stat3.

    PubMed

    Sengupta, S; Nagalingam, A; Muniraj, N; Bonner, M Y; Mistriotis, P; Afthinos, A; Kuppusamy, P; Lanoue, D; Cho, S; Korangath, P; Shriver, M; Begum, A; Merino, V F; Huang, C-Y; Arbiser, J L; Matsui, W; Győrffy, B; Konstantopoulos, K; Sukumar, S; Marignani, P A; Saxena, N K; Sharma, D

    2017-06-05

    Tumor suppressor and upstream master kinase Liver kinase B1 (LKB1) plays a significant role in suppressing cancer growth and metastatic progression. We show that low-LKB1 expression significantly correlates with poor survival outcome in breast cancer. In line with this observation, loss-of-LKB1 rendered breast cancer cells highly migratory and invasive, attaining cancer stem cell-like phenotype. Accordingly, LKB1-null breast cancer cells exhibited an increased ability to form mammospheres and elevated expression of pluripotency-factors (Oct4, Nanog and Sox2), properties also observed in spontaneous tumors in Lkb1(-/-) mice. Conversely, LKB1-overexpression in LKB1-null cells abrogated invasion, migration and mammosphere-formation. Honokiol (HNK), a bioactive molecule from Magnolia grandiflora increased LKB1 expression, inhibited individual cell-motility and abrogated the stem-like phenotype of breast cancer cells by reducing the formation of mammosphere, expression of pluripotency-factors and aldehyde dehydrogenase activity. LKB1, and its substrate, AMP-dependent protein kinase (AMPK) are important for HNK-mediated inhibition of pluripotency factors since LKB1-silencing and AMPK-inhibition abrogated, while LKB1-overexpression and AMPK-activation potentiated HNK's effects. Mechanistic studies showed that HNK inhibited Stat3-phosphorylation/activation in an LKB1-dependent manner, preventing its recruitment to canonical binding-sites in the promoters of Nanog, Oct4 and Sox2. Thus, inhibition of the coactivation-function of Stat3 resulted in suppression of expression of pluripotency factors. Further, we showed that HNK inhibited breast tumorigenesis in mice in an LKB1-dependent manner. Molecular analyses of HNK-treated xenografts corroborated our in vitro mechanistic findings. Collectively, these results present the first in vitro and in vivo evidence to support crosstalk between LKB1, Stat3 and pluripotency factors in breast cancer and effective anticancer modulation

  16. Expression of IL-24, an activator of the JAK1/STAT3/SOCS3 cascade, is enhanced in inflammatory bowel disease.

    PubMed

    Andoh, Akira; Shioya, Makoto; Nishida, Atsushi; Bamba, Shigeki; Tsujikawa, Tomoyuki; Kim-Mitsuyama, Shokei; Fujiyama, Yoshihide

    2009-07-01

    IL-24 is a member of the IL-10 family of cytokines. In this study, we investigated IL-24 expression in the inflamed mucosa of patients with inflammatory bowel disease (IBD), and characterized the molecular mechanisms responsible for IL-24 expression in human colonic subepithelial myofibroblasts (SEMFs). IL-24 expression in the IBD mucosa was evaluated by immunohistochemical methods. IL-24 mRNA and protein expression was determined by real-time PCR and ELISA, respectively. AP-1 and C/EBP DNA-binding activity and IL-24 promoter activity were assessed by EMSA analysis and a reporter gene assay, respectively. IL-24 mRNA expression was significantly elevated in active lesions from patients who have ulcerative colitis and Crohn's disease. Colonic SEMFs were identified as a major source of IL-24 in the mucosa. IL-1beta, but not IL-17A, TNF-alpha, or IFN-gamma, significantly enhanced IL-24 mRNA and protein expression in isolated colonic SEMFs. The IL-1beta-induced IL-24 mRNA expression was mediated by the activation of the transcription factors, AP-1 and C/EBP-beta. Induction of IL-24 mRNA stabilization was also involved in the effects of IL-1beta. IL-24 induced JAK1/STAT-3 phosphorylation and SOCS3 expression in HT-29 colonic epithelial cells. IL-24 did not modulate the proliferation of HT-29 cells, but significantly increased the mRNA expression of membrane-bound mucins (MUC1, MUC3, and MUC4). IL-24 derived from colonic SEMFs acts on colonic epithelial cells to elicit JAK1/STAT-3 activation and the expression of SOCS3 and mucins, supporting their suppressive effects on mucosal inflammation in IBD.

  17. Constitutive Phosphorylation of STAT3 by the CK2-BLNK-CD5 Complex.

    PubMed

    Rozovski, Uri; Harris, David M; Li, Ping; Liu, Zhiming; Jain, Preetesh; Veletic, Ivo; Ferrajoli, Alessandra; Burger, Jan; O'Brien, Susan; Bose, Prithviraj; Thompson, Philip; Jain, Nitin; Wierda, William; Keating, Michael J; Estrov, Zeev

    2017-01-27

    In chronic lymphocytic leukemia (CLL), STAT3 is constitutively phosphorylated on serine 727 and plays a role in the pathobiology of CLL. However, what induces constitutive phosphorylation of STAT3 is currently unknown. Mass spectrometry was used to identify casein kinase 2 (CK2), a serine/threonine kinase that co-immunoprecipitated with serine phosphorylated STAT3 (pSTAT3). Furthermore, activated CK2 incubated with recombinant STAT3 induced phosphorylation of STAT3 on serine 727. Although STAT3 and CK2 are present in normal B- and T-cells, STAT3 is not constitutively phosphorylated in these cells. Further study found that CD5 and BLNK co-expressed in CLL, but not in normal B- or T-cells, are required for STAT3 phosphorylation. To elucidate the relationship of CD5 and BLNK to CK2 and STAT3, STAT3 was immunoprecipitated from CLL cells and CK2, CD5, and BLNK were detected in the immunoprecipitate. Conversely, STAT3, CD5, and BLNK were in the immunoprecipitate of CLL cells immunoprecipitated with CK2 antibodies. Furthermore, siRNA knockdown of CD5 or BLNK, or treatment with CD5-neutralizing antibodies significantly reduced the levels of serine pSTAT3 in CLL cells. Finally, confocal microscopy determined that CD5 is cell membrane bound and fractionation studies revealed that the CK2/CD5/BLNK/STAT3 complex remains in the cytoplasm, whereas serine pSTAT3 is shuttled to the nucleus.

  18. Mutations in the signal transducer and activator of transcription 3 (STAT3) and diagnostic guidelines for the Hyper-IgE Syndrome

    PubMed Central

    Woellner, Cristina; Gertz, E. Michael; Schäffer, Alejandro A.; Lagos, Macarena; Perro, Mario; Glocker, Erik-Oliver; Pietrogrande, Maria C.; Cossu, Fausto; Franco, José L.; Matamoros, Nuria; Pietrucha, Barbara; Heropolitańska-Pliszka, Edyta; Yeganeh, Mehdi; Moin, Mostafa; Español, Teresa; Ehl, Stephan; Gennery, Andrew R.; Abinun, Mario; Bręborowicz, Anna; Niehues, Tim; Kilic, Sara Sebnem; Junker, Anne; Turvey, Stuart E.; Plebani, Alessandro; Sánchez, Berta; Garty, Ben-Zion; Pignata, Claudio; Cancrini, Caterina; Litzman, Jiri; Sanal, Özden; Baumann, Ulrich; Bacchetta, Rosa; Hsu, Amy P.; Davis, Joie N.; Hammarström, Lennart; Davies, E. Graham; Eren, Efrem; Arkwright, Peter D.; Moilanen, Jukka S.; Viemann, Dorothee; Khan, Sujoy; Maródi, László; Cant, Andrew J.; Freeman, Alexandra F.; Puck, Jennifer M.; Holland, Steven M.; Grimbacher, Bodo

    2010-01-01

    Background The hyper-IgE syndrome (HIES) is a primary immunodeficiency characterized by infections of the lung and skin, elevated serum IgE, and involvement of the soft and tissues. Recently, HIES has been associated with heterozygous dominant-negative mutations in STAT3 and severe reductions of Th17 cells. Objective To determine whether there is a correlation between the genotype and phenotype of HIES patients and to establish diagnostic criteria to distinguish between STAT3 mutated and STAT3 wild-type patients. Methods We collected clinical data, determined Th17 cell numbers, and sequenced STAT3 100 patients with a strong clinical suspicion of HIES and serum IgE >1000 IU/mL. explored diagnostic criteria by using a machine-learning approach to identify which features best predict a STAT3 mutation. Results In 64 patients we identified 31 different STAT3 mutations, 18 of which are novel. These included mutations at splice sites and outside the previously implicated DNA-binding and SH2 domains. A combination of five clinical features predicted STAT3 mutations with 85% accuracy. Th17 cells were profoundly reduced in patients harboring STAT3 mutations, while 10 out of 13 patients without mutations had low (<1%) Th17 cells but were distinct markedly reduced IFN-γ producing CD4+ T cells. Conclusions We propose the following diagnostic guidelines for STAT3-deficient HIES: Possible: IgE >1000 IU/mL plus a weighted score of clinical features >30 based on recurrent pneumonia, newborn rash, pathologic bone fractures, characteristic face, and high palate. Probable: Above plus lack of Th17 cells or a family history for definitive HIES. Definitive: Above plus a dominant-negative heterozygous mutation in STAT3. PMID:20159255

  19. Genetic Interactions of STAT3 and Anticancer Drug Development

    PubMed Central

    Fang, Bingliang

    2014-01-01

    Signal transducer and activator of transcription 3 (STAT3) plays critical roles in tumorigenesis and malignant evolution and has been intensively studied as a therapeutic target for cancer. A number of STAT3 inhibitors have been evaluated for their antitumor activity in vitro and in vivo in experimental tumor models and several approved therapeutic agents have been reported to function as STAT3 inhibitors. Nevertheless, most STAT3 inhibitors have yet to be translated to clinical evaluation for cancer treatment, presumably because of pharmacokinetic, efficacy, and safety issues. In fact, a major cause of failure of anticancer drug development is lack of efficacy. Genetic interactions among various cancer-related pathways often provide redundant input from parallel and/or cooperative pathways that drives and maintains survival environments for cancer cells, leading to low efficacy of single-target agents. Exploiting genetic interactions of STAT3 with other cancer-related pathways may provide molecular insight into mechanisms of cancer resistance to pathway-targeted therapies and strategies for development of more effective anticancer agents and treatment regimens. This review focuses on functional regulation of STAT3 activity; possible interactions of the STAT3, RAS, epidermal growth factor receptor, and reduction-oxidation pathways; and molecular mechanisms that modulate therapeutic efficacies of STAT3 inhibitors. PMID:24662938

  20. Stevia and stevioside protect against cisplatin nephrotoxicity through inhibition of ERK1/2, STAT3, and NF-κB activation.

    PubMed

    Potočnjak, Iva; Broznić, Dalibor; Kindl, Marija; Kropek, Matija; Vladimir-Knežević, Sanda; Domitrović, Robert

    2017-09-01

    We investigated the effect of natural sweetener Stevia rebaudiana and its constituent stevioside in cisplatin (CP)-induced kidney injury. Male BALB/cN mice were orally administered 10, 20, and 50 mg/kg body weight of Stevia rebaudiana ethanol extract (SE) or stevioside 50 mg/kg, 48 h after intraperitoneal administration of CP (13 mg/kg). Two days later, CP treatment resulted in histopathological changes showing kidney injury. Increased expression of 4-hydroxynonenal (4-HNE), 3-nitrotyrosine (3-NT), and heme oxygenase-1 (HO-1) in mice kidneys suggested oxidative stress. CP treatment also increased renal expression of nuclear factor-kappaB (NF-κB) p65 subunit and phosphorylated inhibitor of NF-κB (IκBα), as well as expression of pro-inflammatory cytokine tumor necrosis factor-alpha (TNF-α). Induction of apoptosis and inhibition of the cell cycle in kidneys was evidenced by increased expression of p53, Bax, caspase-9, and p21, proteolytic cleavage of poly (ADP-ribose) polymerase (PARP), with concomitant suppression of Bcl-2 and cyclin D1 expression. The number of apoptotic cells in kidneys was also assessed. CP administration resulted in activation of extracellular signal-regulated kinases 1 and 2 (ERK1/2) and signal transducer and activator of transcription 3 (STAT3). Both SE and stevioside attenuated CP nephrotoxicity by suppressing oxidative stress, inflammation, and apoptosis through mechanism involving ERK1/2, STAT3, and NF-κB suppression. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. Inorganic Phosphate Prevents Erk1/2 and Stat3 Activation and Improves Sensitivity to Doxorubicin of MDA-MB-231 Breast Cancer Cells.

    PubMed

    Sapio, Luigi; Sorvillo, Luca; Illiano, Michela; Chiosi, Emilio; Spina, Annamaria; Naviglio, Silvio

    2015-09-01

    Due to its expression profile, triple-negative breast cancer (TNBC) is refractory to the most effective targeted therapies available for breast cancer treatment. Thus, cytotoxic chemotherapy represents the mainstay of treatment for early and metastatic TNBC. Therefore, it would be greatly beneficial to develop therapeutic approaches that cause TNBC cells to increase their sensitivity to cytotoxic drugs. Inorganic phosphate (Pi) is emerging as an important signaling molecule in many cell types. Interestingly, it has been shown that Pi greatly enhances the sensitivity of human osteosarcoma cell line (U2OS) to doxorubicin. We investigated the effects of Pi on the sensitivity of TNBC cells to doxorubicin and the underlying molecular mechanisms, carrying out flow cytometry-based assays of cell-cycle progression and cell death, MTT assays, direct cell number counting and immunoblotting experiments. We report that Pi inhibits the proliferation of triple-negative MDA-MB-231 breast cancer cells mainly by slowing down cell cycle progression. Interestingly, we found that Pi strongly increases doxorubicin-induced cytotoxicity in MDA-MB-231 cells by apoptosis induction, as revealed by a marked increase of sub-G1 population, Bcl-2 downregulation, caspase-3 activation and PARP cleavage. Remarkably, Pi/doxorubicin combination-induced cytotoxicity was dynamically accompanied by profound changes in Erk1/2 and Stat3 protein and phosphorylation levels. Altogether, our data enforce the evidence of Pi acting as a signaling molecule in MDA-MB-231 cells, capable of inhibiting Erk and Stat3 pathways and inducing sensitization to doxorubicin of TNBC cells, and suggest that targeting Pi levels at local sites might represent the rationale for developing effective and inexpensive strategies for improving triple-negative breast cancer therapy.

  2. Role of STAT3 pathway in genitourinary tumors

    PubMed Central

    Santoni, Matteo; Conti, Alessandro; Piva, Francesco; Massari, Francesco; Ciccarese, Chiara; Burattini, Luciano; Cheng, Liang; Lopez-Beltran, Antonio; Scarpelli, Marina; Santini, Daniele; Tortora, Giampaolo; Cascinu, Stefano; Montironi, Rodolfo

    2015-01-01

    The STAT3 is often dysregulated in genitourinary tumors. In prostate cancer, STAT3 activation correlates with Gleason score and pathological stage and modulates cancer stem cells and epithelial–mesenchymal transition. In addition, STAT3 promotes the progression from carcinoma in situ to invasive bladder cancer and modulates renal cell carcinoma angiogenesis by increasing the expression of HIF1α and VEGF. STAT3 is also involved in the response to tyrosine kinase inhibitors sunitinib and axitinib, in patients with metastatic renal cell carcinoma, and to second-generation androgen receptor inhibitor enzalutamide in patients with advanced prostate cancer. In this review, we describe the role of STAT3 in genitourinary tumors, thus describing its potential for future therapeutic strategies. PMID:28031890

  3. Design, Synthesis and in vitro Characterization of Novel Hybrid Peptidomimetic Inhibitors of STAT3 Protein

    PubMed Central

    Shahani, Vijay M.; Yue, Peibin; Fletcher, Steven; Sharmeen, Sumaiya; Sukhai, Mahadeo A.; Luu, Diana P.; Zhang, Xiaolei; Sun, Hong; Zhao, Wei; Schimmer, Aaron D.; Turkson, James; Gunning, Patrick T.

    2011-01-01

    Aberrant activation of oncogenic signal transducer and activator of transcription 3 (STAT3) protein signaling pathways has been extensively implicated in human cancers. Given STAT3’s prominent dysregulatory role in malignant transformation and tumorigenesis, there has been a significant effort to discover STAT3-specific inhibitors as chemical probes for defining the aberrant STAT3-mediated molecular events that support the malignant phenotype. To identify novel, STAT3-selective inhibitors suitable for interrogating STAT3 signaling in tumor cells, we explored the design of hybrid molecules by conjugating a known STAT3 inhibitory peptidomimetic, ISS610 to the high-affinity STAT3-binding peptide motif derived from the ILR/gp-130. Several hybrid molecules were examined in in vitro biophysical and biochemical studies for inhibitory potency against STAT3. Lead inhibitor 14aa was shown to strongly bind to STAT3 (KD = 900 nM), disrupt STAT3:phosphopeptide complexes (Ki = 5 μM) and suppress STAT3 activity in in vitro DNA-binding activity/ electrophoretic mobility shift assay (EMSA). Moreover, lead STAT3 inhibitor 14aa induced a time-dependent inhibition of constitutive STAT3 activation in v-Src transformed mouse fibroblasts (NIH3T3/v-Src), with 80 % suppression of constitutively-active STAT3 at six hours following treatment of NIH3T3/v-Src. However, STAT3 activity recovered at 24 hours after treatment of cells, suggesting potential degradation of the compound. Results further showed a suppression of aberrant STAT3 activity in NIH3T3/v-Src by the treatment with compound 14aa-OH, which is the non-pTyr version of compound 14aa. The effect of compounds 14aa and 14aa-OH are accompanied by a moderate loss of cell viability. PMID:21216604

  4. Human Cytomegalovirus IE1 Protein Disrupts Interleukin-6 Signaling by Sequestering STAT3 in the Nucleus

    PubMed Central

    Reitsma, Justin M.; Sato, Hiromi; Nevels, Michael

    2013-01-01

    In the canonical STAT3 signaling pathway, binding of agonist to receptors activates Janus kinases that phosphorylate cytoplasmic STAT3 at tyrosine 705 (Y705). Phosphorylated STAT3 dimers accumulate in the nucleus and drive the expression of genes involved in inflammation, angiogenesis, invasion, and proliferation. Here, we demonstrate that human cytomegalovirus (HCMV) infection rapidly promotes nuclear localization of STAT3 in the absence of robust phosphorylation at Y705. Furthermore, infection disrupts interleukin-6 (IL-6)-induced phosphorylation of STAT3 and expression of a subset of IL-6-induced STAT3-regulated genes, including SOCS3. We show that the HCMV 72-kDa immediate-early 1 (IE1) protein associates with STAT3 and is necessary to localize STAT3 to the nucleus during infection. Furthermore, expression of IE1 is sufficient to disrupt IL-6-induced phosphorylation of STAT3, binding of STAT3 to the SOCS3 promoter, and SOCS3 gene expression. Finally, inhibition of STAT3 nuclear localization or STAT3 expression during infection is linked to diminished HCMV genome replication. Viral gene expression is also disrupted, with the greatest impact seen following viral DNA synthesis. Our study identifies IE1 as a new regulator of STAT3 intracellular localization and IL-6 signaling and points to an unanticipated role of STAT3 in HCMV infection. PMID:23903834

  5. The JAK2 Inhibitor, AZD1480, Potently Blocks Stat3 Signaling and Oncogenesis in Solid Tumors

    PubMed Central

    Hedvat, Michael; Huszar, Dennis; Herrmann, Andreas; Gozgit, Joseph M.; Schroeder, Anne; Sheehy, Adam; Buettner, Ralf; Proia, David; Kowolik, Claudia M.; Xin, Hong; Armstrong, Brian; Bebernitz, Geraldine; Weng, Shaobu; Wang, Lin; Ye, Minwei; McEachern, Kristen; Chen, Huawei; Morosini, Deborah; Bell, Kirsten; Alimzhanov, Marat; Ioannidis, Stephanos; McCoon, Patricia; Cao, Zhu A.; Yu, Hua; Jove, Richard; Zinda, Michael

    2009-01-01

    Summary Persistent activation of Stat3 is oncogenic and is prevalent in a wide variety of human cancers. Chronic cytokine stimulation is associated with Stat3 activation in some tumors, implicating cytokine receptor-associated Jak family kinases. Using Jak2 inhibitors, we demonstrate a central role of Jaks in modulating basal and cytokine-induced Stat3 activation in human solid tumor cell lines. Inhibition of Jak2 activity is associated with abrogation of Stat3 nuclear translocation and tumorigenesis. The Jak2 inhibitor, AZD1480, suppresses the growth of human solid tumor xenografts harboring persistent Stat3 activity. We demonstrate the essential role of Stat3 downstream of Jaks by inhibition of tumor growth using shRNA targeting Stat3. Our data support a key role of Jak kinase activity in Stat3-dependent tumorigenesis. PMID:19962667

  6. Suppression by Δ(9)-tetrahydrocannabinol of the primary immunoglobulin M response by human peripheral blood B cells is associated with impaired STAT3 activation.

    PubMed

    Ngaotepprutaram, Thitirat; Kaplan, Barbara L F; Carney, Stephen; Crawford, Robert; Kaminski, Norbert E

    2013-08-09

    This study was undertaken to gain insights into the mechanism for Δ(9)-tetrahydrocannabinol (Δ(9)-THC)-mediated suppression of primary immunoglobulin M (IgM) responses in humans. An in vitro activation model, which employs cell surface-expressed CD40 ligand (CD40L) and recombinant cytokines (interleukin (IL)-2, -6, and -10), was used to differentiate human peripheral blood (HPB) naïve B cells into IgM secreting cells. Pretreatment with Δ(9)-THC significantly decreased the number of IgM secreting cells as determined by ELISPOT. The attenuation of IgM secretion by Δ(9)-THC involved, at least in part, the impairment of plasma cell differentiation as evidenced by suppression of immunoglobulin joining chain (IgJ) mRNA expression. The analysis at each of two different stages critically involved in plasma cell differentiation indicates that Δ(9)-THC impaired both the primary activation stage and proliferation of B cells. Interestingly, Δ(9)-THC selectively suppressed the surface expression of CD80, but not other measured B-cell activation markers (CD69, CD86, and ICAM1). Furthermore, pretreatment with Δ(9)-THC was accompanied by a robust decrease of STAT3 phosphorylation, whereas the phosphorylation of the p65 NFκB subunit was not affected. Collectively, these data provide new insights into the mechanisms for impaired B cell function by Δ(9)-THC.

  7. STAT3 upregulation in pituitary somatotroph adenomas induces growth hormone hypersecretion.

    PubMed

    Zhou, Cuiqi; Jiao, Yonghui; Wang, Renzhi; Ren, Song-Guang; Wawrowsky, Kolja; Melmed, Shlomo

    2015-04-01

    Pituitary somatotroph adenomas result in dysregulated growth hormone (GH) hypersecretion and acromegaly; however, regulatory mechanisms that promote GH hypersecretion remain elusive. Here, we provide evidence that STAT3 directly induces somatotroph tumor cell GH. Evaluation of pituitary tumors revealed that STAT3 expression was enhanced in human GH-secreting adenomas compared with that in nonsecreting pituitary tumors. Moreover, STAT3 and GH expression were concordant in a somatotroph adenoma tissue array. Promoter and expression analysis in a GH-secreting rat cell line (GH3) revealed that STAT3 specifically binds the Gh promoter and induces transcription. Stable expression of STAT3 in GH3 cells induced expression of endogenous GH, and expression of a constitutively active STAT3 further enhanced GH production. Conversely, expression of dominant-negative STAT3 abrogated GH expression. In primary human somatotroph adenoma-derived cell cultures, STAT3 suppression with the specific inhibitor S3I-201 attenuated GH transcription and reduced GH secretion in the majority of derivative cultures. In addition, S3I-201 attenuated somatotroph tumor growth and GH secretion in a rat xenograft model. GH induced STAT3 phosphorylation and nuclear translocation, indicating a positive feedback loop between STAT3 and GH in somatotroph tumor cells. Together, these results indicate that adenoma GH hypersecretion is the result of STAT3-dependent GH induction, which in turn promotes STAT3 expression, and suggest STAT3 as a potential therapeutic target for pituitary somatotroph adenomas.

  8. The antihistamines clemastine and desloratadine inhibit STAT3 and c-Myc activities and induce apoptosis in cutaneous T-cell lymphoma cell lines.

    PubMed

    Döbbeling, Udo; Waeckerle-Men, Ying; Zabel, Franziska; Graf, Nicole; Kündig, Thomas M; Johansen, Pål

    2013-02-01

    Mycosis fungoides and its leukaemic counterpart Sézary syndrome are the most frequent cutaneous T-cell lymphomas (CTCL), and there is no cure for these diseases. We evaluated the effect of clinically approved antihistamines on the growth of CTCL cell lines. CTCL cell lines as well as blood lymphocytes from patients with Sézary syndrome were cultured with antihistamines, and the cell were analysed for proliferation, apoptosis and expression of programmed death molecules and transcription factors. The two antihistamines clemastine and desloratadine, currently used for symptom alleviation in allergy, induced potent reduction of the activities of the constitutively active transcription factors c-Myc, STAT3, STAT5a and STAT5b in mycosis fungoides and Sézary syndrome cell lines. This inhibition was followed by apoptosis and cell death, especially in the Sézary syndrome-derived cell line Hut78 that also showed increased expression of the programmed death-1 (PD-1) after clemastine treatment. In lymphocytes isolated from Sézary syndrome patients, the CD4-positive fraction underwent apoptosis after clemastine treatment, while CD4-negative lymphocytes were little affected. Because both c-Myc and STAT transcription factors are highly expressed in proliferating tumours, their inhibition by clemastine, desloratadine and other inhibitors could complement established chemotherapies not only for cutaneous T-cell lymphomas but perhaps also other cancers.

  9. Idiopathic pancreatitis in a patient with a STAT3 mutation

    PubMed Central

    Peppers, Brian; Frith, John; Tcheurekdjian, Haig; Hostoffer, Robert

    2016-01-01

    Background: Hyperimmunoglobulin E syndrome (HIES) is a rare primary immunodeficiency characterized by recurrent skin infections with abscesses, recurrent pneumonias with pneumatoceles, and immunoglobulin E levels of >10 times the upper limit of normal. Case: The patient described herein had a classic case of signal transducer and activator of transcription 3 (STAT3) deficiency associated with HIES diagnosed several years before this particular presentation. He demonstrated extraimmune manifestations of the disease as well, including characteristic facies and a history of skeletal fractures. In addition, the patient had several distinct episodes of idiopathic pancreatitis for which a full gastrointestinal workup had been performed. STAT3 mutation was confirmed by genotyping at the time of diagnosis of HIES. Conclusions: STAT3, a mammalian protein that regulates cell growth, survival, and differentiation, has been linked to human pancreatic carcinogenesis as well as the above-mentioned immune deficiency. Mouse studies demonstrated that genetic ablation of STAT3 exacerbates the course of acute pancreatitis, whereas normal pancreatic STAT3 seems to have a protective effect against necrotizing pancreatitis. An association between STAT3 mutations and pancreatitis has not yet been revealed in humans. Here we describe a case of acute pancreatitis that presented in a patient with STAT3 mutation. PMID:27103560

  10. Hepatocyte-specific Smad7 deletion accelerates DEN-induced HCC via activation of STAT3 signaling in mice

    PubMed Central

    Feng, T; Dzieran, J; Yuan, X; Dropmann, A; Maass, T; Teufel, A; Marhenke, S; Gaiser, T; Rückert, F; Kleiter, I; Kanzler, S; Ebert, M P; Vogel, A; ten Dijke, P; Dooley, S; Meindl-Beinker, N M

    2017-01-01

    TGF-β signaling in liver cells has variant roles in the dynamics of liver diseases, including hepatocellular carcinoma (HCC). We previously found a correlation of high levels of the important endogenous negative TGF-β signaling regulator SMAD7 with better clinical outcome in HCC patients. However, the underlying tumor-suppressive molecular mechanisms are still unclear. Here, we show that conditional (TTR-Cre) hepatocyte-specific SMAD7 knockout (KO) mice develop more tumors than wild-type and corresponding SMAD7 transgenic mice 9 months after diethylnitrosamine (DEN) challenge, verifying SMAD7 as a tumor suppressor in HCC. In line with our findings in patients, Smad7 levels in both tumor tissue as well as surrounding tissue show a significant inverse correlation with tumor numbers. SMAD7 KO mice presented with increased pSMAD2/3 levels and decreased apoptosis in the tumor tissue. Higher tumor incidence was accompanied by reduced P21 and upregulated c-MYC expression in the tumors. Activation of signal transducer and activator of transcription factor 3 signaling was found in Smad7-deficient mouse tumors and in patients with low tumoral SMAD7 expression as compared with surrounding tissue. Together, our results provide new mechanistic insights into the tumor-suppressive functions of SMAD7 in hepatocarcinogenesis. PMID:28134936

  11. IL-22+CD4+ T cells promote colorectal cancer stemness via STAT3 transcription factor activation and induction of the methyltransferase DOT1L

    PubMed Central

    Peng, Dongjun; Zhao, Lili; Zhao, Ende; Vatan, Linda; Szeliga, Wojciech; Dou, Yali; Owens, Scott; Zgodzinski, Witold; Majewski, Marek; Wallner, Grzegorz; Fang, Jingyuan; Huang, Emina; Zou, Weiping

    2014-01-01

    Little is known about how the immune system impacts human colorectal cancer invasiveness and stemness. Here we detected interleukin-22 (IL-22) in patient colorectal cancer tissues that was produced predominantly by CD4+ T cells. In a mouse model, migration of these cells into the colon cancer microenvironment required the chemokine receptor CCR6 and its ligand CCL20. IL-22 acted on cancer cells to promote activation of the transcription factor STAT3 and expression of the histone 3 lysine 79 (H3K79) methytransferase DOT1L. The DOT1L complex induced the core stem cell genes NANOG, SOX2 and Pou5F1, resulting in increased cancer stemness and tumorigenic potential. Furthermore, high DOT1L expression and H3K79me2 in colorectal cancer tissues was a predictor of poor patient survival. Thus, IL-22+ cells promote colon cancer stemness via regulation of stemness genes which negatively affects patient outcome. Efforts to target this network might be a strategy in treating colorectal cancer patients. PMID:24816405

  12. Dominant negative stat3 mutant inhibits interleukin-6-induced Jak-STAT signal transduction.

    PubMed

    Kaptein, A; Paillard, V; Saunders, M

    1996-03-15

    Interleukin-6 (IL-6) induces tyrosine phosphorylation and activation of the latent transcription factor Stat3 in HepG2 cells. Mutation of Stat3 tyrosine 705 to phenylalanine (Y705F) inhibits IL-6-induced tyrosine phosphorylation of this Stat3 mutant in transfected HepG2 cells. In cotransfections of HepG2 cells, the Stat3 mutant Y705F causes a reduction of the tyrosine phosphorylation of wild type Stat3-FLAG. Moreover, Y705F inhibits the action of endogenous Stat3 in cotransfected cells, reducing IL-6 induction of a Stat3-responsive reporter construct. Y705F therefore acts as a dominant negative mutation of Stat3.

  13. 20(S)-25-methoxyl-dammarane-3β, 12β, 20-triol negatively regulates activation of STAT3 and ERK pathways and exhibits anti-cancer effects in HepG2 cells.

    PubMed

    Ai, Hui-Han; Zhou, Zi-Long; Sun, Lu-Guo; Yang, Mei-Ting; Li, Wei; Yu, Chun-Lei; Song, Zhen-Bo; Huang, Yan-Xin; Wu, Yin; Liu, Lei; Yang, Xiao-Guang; Zhao, Yu-Qing; Bao, Yong-Li; Li, Yu-Xin

    2017-09-01

    The pro-inflammatory cytokine interleukin 6 (IL-6), via activating its downstream JAK/STAT3 and Ras/ERK signaling pathways, is involved in cell growth, proliferation and anti-apoptotic activities in various malignancies. To screen inhibitors of IL-6 signaling, we constructed a STAT3 and ERK dual-pathway responsive luciferase reporter vector (Co.RE). Among several candidates, the natural compound 20(S)-25-methoxyl-dammarane-3β, 12β, 20-triol (25-OCH3-PPD, GS25) was identified to clearly inhibit the luciferase activity of Co.RE. GS25 was confirmed to indeed inhibit activation of both STAT3 and ERK pathways and expression of downstream target genes of IL-6, and to predominantly decrease the viability of HepG2 cells via induction of cell cycle arrest and apoptosis. Interestingly, GS25 showed preferential inhibition of HepG2 cell viability relative to normal liver L02 cells. Further investigation showed that GS25 could not induce apoptosis and block activation of STAT3 and ERK pathways in L02 cells as efficiently as in HepG2 cells, which may result in differential effects of GS25 on malignant and normal liver cells. In addition, GS25 was found to potently suppress the expression of endogenous STAT3 at a higher concentration and dramatically induce p38 phosphorylation in HepG2 cells, which could mediate its anti-cancer effects. Finally, we demonstrated that GS25 also inhibited tumor growth in HepG2 xenograft mice. Taken together, these findings indicate that GS25 elicits its anti-cancer effects on HepG2 cells through multiple mechanisms and has the potential to be used as an inhibitor of IL-6 signaling. Thus, GS25 may be developed as a treatment for hepatocarcinoma with low toxicity on normal liver tissues as well as other inflammation-associated diseases.

  14. Induction of caspase-dependent extrinsic apoptosis by apigenin through inhibition of signal transducer and activator of transcription 3 (STAT3) signalling in HER2-overexpressing BT-474 breast cancer cells.

    PubMed

    Seo, Hye-Sook; Jo, Jae Kyung; Ku, Jin Mo; Choi, Han-Seok; Choi, Youn Kyung; Woo, Jong-Kyu; Kim, Hyo In; Kang, Soo-Yeon; Lee, Kang Min; Nam, Koong Won; Park, Namkyu; Jang, Bo-Hyoung; Shin, Yong Cheol; Ko, Seong-Gyu

    2015-10-23

    Phytoestrogen intake is known to be beneficial to decrease breast cancer incidence and progression. But its molecular mechanisms of action are still unknown. The present study aimed to examine the effect of apigenin on proliferation and apoptosis in HER2-expressing breast cancer cells. In our experiments, apigenin inhibited the proliferation of BT-474 cells in a dose- and time-dependent manner. Apigenin also inhibited clonogenic survival (anchorage-dependent and -independent) of BT-474 cells in a dose-dependent manner. These growth inhibitions were accompanied with an increase in sub-G0/G1 apoptotic populations. Apigenin-induced extrinsic a caspase-dependent apoptosis up-regulating the levels of cleaved caspase-8 and cleaved caspase-3, and inducing the cleavage of poly (ADP-ribose) polymerase (PARP). Whereas, apigenin did not induce apoptosis via intrinsic mitochondrial apoptosis pathway since this compound did not decrease mitochondrial membrane potential without affecting the levels of B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (BAX). Apigenin reduced the expression of phospho-JAK1, phospho-JAK2 and phospho-STAT3 and decreased signal transducer and activator of transcription 3 (STAT3) dependent luciferase reporter gene activity in BT-474 cells. Apigenin inhibited CoCl2-induced VEGF secretion and decreased the nuclear translocation of STAT3. Our study indicates that apigenin induces apoptosis through inhibition of STAT3 signalling and could serve as a useful compound to prevent or treat HER2-overexpressing breast cancer.

  15. Differences in antiproliferative effect of STAT3 inhibition in HCC cells with versus without HBV expression

    SciTech Connect

    Hong, Yun; Zhou, Lin; Xie, Haiyang; Wang, Weilin; Zheng, Shusen

    2015-06-05

    Chronic infection with hepatitis B virus (HBV) plays an important role in the etiology of hepatocellular carcinoma (HCC). Signal transducer and activator of transcription 3 (STAT3) inactivation could inhibit the tumor growth of HCC. In this study, differential antiproliferative effect of STAT3 inhibition was observed with HBV-related HCC cells being more resistant than non-HBV-related HCC cells. Resistance of HBV-related HCC cells to STAT3 inhibition was positively correlated to the expression of HBV. Enhanced ERK activation after STAT3 blockade was detected in HBV-related HCC cells but not in non-HBV-related HCC cells. Combined ERK and STAT3 inhibition eliminates the discrepancy between the two types of HCC cells. Moderate reduced HBV expression was found after STAT3 inhibition. These findings disclose a discrepancy in cellular response to STAT3 inhibition between non-HBV-related and HBV-related HCC cells and underscore the complexity of antiproliferative effect of STAT3 inactivation in HBV-related HCC cells. - Highlights: • HBV endows HCC cells with resistance to STAT3 inactivation on proliferation. • Abnormal ERK activation after STAT3 inhibition in HBV-related HCC cells. • Combined ERK and STAT3 inhibition eliminates the discrepancy. • STAT3 inhibition moderately reduces HBV expression.

  16. STAT3 phosphorylation in injured axons before sensory and motor neuron nuclei: potential role for STAT3 as a retrograde signaling transcription factor.

    PubMed

    Lee, Nancy; Neitzel, Karen L; Devlin, Brenda K; MacLennan, A John

    2004-07-05

    STAT3 is a latent transcription factor that is activated by plasma membrane growth factor receptor complexes. Conditional gene disruption data indicate that it contributes to the survival of cranial motor neurons after peripheral nerve lesion. In agreement, levels of activated STAT3 (Tyr705-phosphorylated STAT3) have been shown to increase in the nuclei of adult cranial motor neurons during their regeneration after the same injury. The data presented here demonstrate that STAT3 is similarly but not identically affected in sciatic motor neurons after sciatic nerve injury. In addition, we find that sensory neuron nuclei also display an analogous increase in activated STAT3, thereby supporting a role for STAT3 in the survival and regeneration of these cells. Most interesting, the present data indicate that peripheral nerve lesion leads to a very rapid activation of STAT3 in axons at the lesion site. This response increases during the first 24 hours after injury and extends back to the motor and sensory neurons such that phospho-STAT3-immunoreactive axons are first detected in the dorsal root ganglia and ventral spinal cord at the same postlesion time intervals at which the activated STAT3 is first detected in the neuronal nuclei. Together these data raise the possibility that axonal STAT3, activated at the injury site, acts as a retrograde signaling transcription factor, which promotes the survival and regeneration of both sensory and motor neurons.

  17. Pancreatic STAT3 protects mice against caerulein-induced pancreatitis via PAP1 induction.

    PubMed

    Shigekawa, Minoru; Hikita, Hayato; Kodama, Takahiro; Shimizu, Satoshi; Li, Wei; Uemura, Akio; Miyagi, Takuya; Hosui, Atsushi; Kanto, Tatsuya; Hiramatsu, Naoki; Tatsumi, Tomohide; Takeda, Kiyoshi; Akira, Shizuo; Takehara, Tetsuo

    2012-12-01

    The signal transducer and activator of transcription 3 (STAT3) is a transcription factor that controls expressions of several genes involved in cell survival, proliferation and differentiation, and tissue inflammation. However, the significance of pancreatic STAT3 in acute pancreatitis remains unclear. We generated conditional STAT3 knockout (stat3(Δ/Δ)) mice by crossing stat3(flox/flox) mice with Pdx1-promoter Cre transgenic mice. Caerulein administration activated pancreatic STAT3 and induced acute pancreatitis as early as 3 hours in wild-type mice, and full recovery from the induced pancreatic injury was observed within 7 days. The levels of serum amylase and lipase and histologic scores of pancreatic necrosis and inflammatory cell infiltration were significantly higher at 3 hours in stat3(Δ/Δ) mice than in stat3(flox/flox) mice. Pancreatic recovery after pancreatitis was significantly delayed in stat3(Δ/Δ) mice compared with stat3(flox/flox) mice. Although stat3(flox/flox) mice had marked production in the pancreas of pancreatitis-associated protein 1 (PAP1), a serum acute phase protein, this induction was completely abrogated in stat3(Δ/Δ) mice. Enforced production of PAP1 by a hydrodynamic procedure in the liver significantly suppressed pancreatic necrosis and inflammation and also promoted pancreatic regeneration and recovery in stat3(Δ/Δ) mice to levels similar to those observed in stat3(flox/flox) mice. In conclusion, pancreatic STAT3 is indispensable for PAP1 production, and this STAT3/PAP1 pathway plays a protective role in caerulein-induced pancreatitis.

  18. ENMD-1198, a novel tubulin-binding agent reduces HIF-1alpha and STAT3 activity in human hepatocellular carcinoma(HCC) cells, and inhibits growth and vascularization in vivo.

    PubMed

    Moser, Christian; Lang, Sven A; Mori, Akira; Hellerbrand, Claus; Schlitt, Hans J; Geissler, Edward K; Fogler, William E; Stoeltzing, Oliver

    2008-07-23

    Hepatocellular carcinoma (HCC) represents a highly vascularized tumor entity and the process of angiogenesis is essential for the growth of HCC. Importantly, the pro-angiogenic transcription factors HIF-1alpha and STAT3 have been implicated in HCC progression, thus representing interesting targets for molecular targeted therapy. We hypothesized that therapeutic inhibition of HIF-1alpha could be achieved by using a novel tubulin-binding agent (ENMD-1198). ENMD-1198 is an analog of 2-methoxyestradiol (2ME2) with antiproliferative and antiangiogenic activity. The human HCC cell lines HUH-7 and HepG2 were used for experiments. Effects of ENMD-1198 on constitutive and inducible (hypoxia, growth factors) activation of signaling cascades, including HIF-1alpha and STAT3, were investigated by Western blotting. Changes in VEGF expression were determined by real-time PCR. Effects of ENMD-1198 on cancer cell migration and invasion were evaluated in in vitro-assays. The growth-inhibitory effects of ENMD-1198 (200 mg/kg/day) were determined in a subcutaneous tumor model (HUH-7). ENMD-1198 inhibited the phosphorylation of MAPK/Erk, PI-3K/Akt and FAK. Moreover, activation of HIF-1alpha and STAT3 was dramatically reduced by ENMD-1198, which resulted in lower VEGF mRNA expression (P < 0.05). In addition, tumor cell migratory and invasive properties were significantly inhibited (P < 0.05, for both). In vivo, treatment with ENMD-1198 led to a significant reduction in tumor growth, tumor vascularization, and numbers of proliferating tumor cells (P < 0.05 for all). The novel microtubule destabilizing agent ENMD-1198 is suitable for inhibiting HIF-1alpha and STAT3 in human HCC cells and leads to reduced tumor growth and vascularization in vivo. Hence, inhibition of HIF-1alpha and STAT3 could prove valuable for therapy of hepatocellular carcinoma.

  19. ENMD-1198, a novel tubulin-binding agent reduces HIF-1alpha and STAT3 activity in human hepatocellular carcinoma(HCC) cells, and inhibits growth and vascularization in vivo

    PubMed Central

    Moser, Christian; Lang, Sven A; Mori, Akira; Hellerbrand, Claus; Schlitt, Hans J; Geissler, Edward K; Fogler, William E; Stoeltzing, Oliver

    2008-01-01

    Background Hepatocellular carcinoma (HCC) represents a highly vascularized tumor entity and the process of angiogenesis is essential for the growth of HCC. Importantly, the pro-angiogenic transcription factors HIF-1α and STAT3 have been implicated in HCC progression, thus representing interesting targets for molecular targeted therapy. We hypothesized that therapeutic inhibition of HIF-1α could be achieved by using a novel tubulin-binding agent (ENMD-1198). ENMD-1198 is an analog of 2-methoxyestradiol (2ME2) with antiproliferative and antiangiogenic activity. Methods The human HCC cell lines HUH-7 and HepG2 were used for experiments. Effects of ENMD-1198 on constitutive and inducible (hypoxia, growth factors) activation of signaling cascades, including HIF-1α and STAT3, were investigated by Western blotting. Changes in VEGF expression were determined by real-time PCR. Effects of ENMD-1198 on cancer cell migration and invasion were evaluated in in vitro-assays. The growth-inhibitory effects of ENMD-1198 (200 mg/kg/day) were determined in a subcutaneous tumor model (HUH-7). Results ENMD-1198 inhibited the phosphorylation of MAPK/Erk, PI-3K/Akt and FAK. Moreover, activation of HIF-1α and STAT3 was dramatically reduced by ENMD-1198, which resulted in lower VEGF mRNA expression (P < 0.05). In addition, tumor cell migratory and invasive properties were significantly inhibited (P < 0.05, for both). In vivo, treatment with ENMD-1198 led to a significant reduction in tumor growth, tumor vascularization, and numbers of proliferating tumor cells (P < 0.05 for all). Conclusion The novel microtubule destabilizing agent ENMD-1198 is suitable for inhibiting HIF-1α and STAT3 in human HCC cells and leads to reduced tumor growth and vascularization in vivo. Hence, inhibition of HIF-1α and STAT3 could prove valuable for therapy of hepatocellular carcinoma. PMID:18651980

  20. STAT3 as a target for inducing apoptosis in solid and hematological tumors

    PubMed Central

    Siddiquee, Khandaker Al Zaid; Turkson, James

    2008-01-01

    Studies in the past few years have provided compelling evidence for the critical role of aberrant Signal Transducer and Activator of Transcription 3 (STAT3) in malignant transformation and tumorigenesis. Thus, it is now generally accepted that STAT3 is one of the critical players in human cancer formation and represents a valid target for novel anticancer drug design. This review focuses on aberrant STAT3 and its role in promoting tumor cell survival and supporting the malignant phenotype. A brief evaluation of the current strategies targeting STAT3 for the development of novel anticancer agents against human tumors harboring constitutively active STAT3 will also be presented. PMID:18227858

  1. Le Carbone, a charcoal supplement, modulates DSS-induced acute colitis in mice through activation of AMPKα and downregulation of STAT3 and caspase 3 dependent apoptotic pathways.

    PubMed

    Afrin, Mst Rejina; Arumugam, Somasundaram; Rahman, Md Azizur; Karuppagounder, Vengadeshprabhu; Sreedhar, Remya; Harima, Meilei; Suzuki, Hiroshi; Nakamura, Takashi; Miyashita, Shizuka; Suzuki, Kenji; Ueno, Kazuyuki; Watanabe, Kenichi

    2017-02-01

    Le Carbone (LC) is a charcoal supplement, which contains a large amount of dietary fibers. Several studies suggested that charcoal supplement may be beneficial for stomach disorders, diarrhea, gas and indigestion. But no studies address whether LC intake would suppress inflammation, cell proliferation or disease progression in colitis. In the present study, the effect of LC on experimental colitis induced by dextran sulfate sodium (DSS) in mice and its possible mechanism of action were examined. A study was designed for 8days, using C57BL/6 female mice that were administered with 3% DSS in drinking water for 7days followed by another 1day consumption of normal water with or without treatment. LC suspension was administered daily for 7days via oral gavage using 5mg/mouse in treatment group and normal group was supplied with drinking water. LC suspension significantly attenuated the loss of body weight and shortening of colon length induced by DSS. The disease activity index, histopathologic changes were significantly reduced by LC treatment. The inflammatory mediators TNFα, IL-1β, p-STAT3 and p-NF-κB induced in the colon by DSS were markedly suppressed by LC. The increased activation of AMPKα in the colon was also detected in LC group. Furthermore, the apoptotic marker protein cleaved caspase 3 was down-regulated and anti-apoptotic proteins Bcl2 and Bcl-xL were significantly up-regulated by LC treatment. Taken together, our results demonstrate the ability of LC to inhibit inflammation, apoptosis and give some evidence for its potential use as adjuvant treatment of inflammatory bowel disease.

  2. Myricitrin attenuates endothelial cell apoptosis to prevent atherosclerosis: An insight into PI3K/Akt activation and STAT3 signaling pathways.

    PubMed

    Qin, Meng; Luo, Yun; Meng, Xiang-bao; Wang, Min; Wang, Hong-wei; Song, Shi-yu; Ye, Jing-xue; Pan, Rui-le; Yao, Fan; Wu, Ping; Sun, Gui-bo; Sun, Xiao-bo

    2015-07-01

    Blood vessel endothelial dysfunction induced by oxidized low-density lipoprotein (ox-LDL) has been implicated in the pathogenesis of atherosclerosis and vasculopathy. The ox-LDL-elicited reactive oxygen species (ROS) release has been assumed to serve a critical function in endothelial damage. Myricitrin (from Myrica cerifera) is a natural antioxidant that has strong anti-oxidative, anti-inflammatory, and anti-nociceptive activities. However, the protective effect of myricitrin on ROS-induced endothelial cell injury and its related molecular mechanisms have never been investigated. This study demonstrates that myricitrin can inhibit ox-LDL-induced endothelial apoptosis and prevent plaque formation at an early stage in an atherosclerotic mouse model. The administration of myricitrin in vivo decreases the thickness of the vascular wall in the aortic arch of ApoE-/- mice. In vitro study shows that ox-LDL-induced human umbilical vein endothelial cell apoptosis can be reduced upon receiving myricitrin pre-treatment. Treatment with myricitrin significantly attenuated ox-LDL-induced endothelial cell apoptosis by inhibiting LOX-1 expression and by increasing the activation of the STAT3 and PI3K/Akt/eNOS signaling pathways. At the same time, our result demonstrates that myricitrin treatment optimizes the balance of pro/anti-apoptosis proteins, including Bax, Bad, XIAP, cIAP-2, and survivin. Our study suggests that myricitrin treatment can effectively protect cells from ox-LDL-induced endothelial cell apoptosis, which results in reduced atherosclerotic plaque formation. This result indicates that myricitrin can be used as a drug candidate for the treatment of cardiovascular diseases. Copyright © 2015. Published by Elsevier Inc.

  3. High mobility group box 1 induces the activation of the Janus kinase 2 and signal transducer and activator of transcription 3 (JAK2/STAT3) signaling pathway in pancreatic acinar cells in rats, while AG490 and rapamycin inhibit their activation

    PubMed Central

    Wang, Guoliang; Zhang, Jingchao; Dui, Danhua; Ren, Haoyuan; Liu, Jin

    2016-01-01

    The pathogenesis of severe acute pancreatitis (SAP) remains unclear. The Janus kinase and signal transducer and activator of transcription (JAK/STAT) pathway is important for various cytokines and growth factors. This study investigated the effect of the late inflammatory factor high mobility group box 1 (HMGB1) on the activation of JAK2/STAT3 in pancreatic acinar cells and the inhibitory effects of AG490 (a JAK2 inhibitor) and rapamycin (a STAT3 inhibitor) on this pathway. Rat pancreatic acinar cells were randomly divided into the control, HMGB1, AG490, and rapamycin groups. The mRNA levels of JAK2 and STAT3 at 10, 30, 60, and 120 minutes were detected using reverse transcription polymerase chain reaction (RT-PCR). The protein levels of JAK2 and STAT3 at 60 and 120 minutes were observed using Western blotting. Compared with the control group, the HMGB1 group exhibited significantly increased levels of JAK2 mRNA at each time point; STAT3 mRNA at 30, 60, and 120 minutes; and JAK2 and STAT3 proteins at 60 and 120 minutes (p < 0.01). Compared with the HMGB1 group, the AG490 and rapamycin groups both exhibited significantly decreased levels of JAK2 mRNA at each time point (p < 0.05); STAT3 mRNA at 30, 60, and 120 minutes (p < 0.01); and JAK2 and STAT3 proteins at 60 and 120 minutes (p < 0.01). HMGB1 induces the activation of the JAK2/STAT3 signaling pathway in rat pancreatic acinar cells, and this activation can be inhibited by AG490 and rapamycin. The results of this study may provide new insights for the treatment of SAP. PMID:27754827

  4. Cross-talk between KLF4 and STAT3 regulates axon regeneration

    NASA Astrophysics Data System (ADS)

    Qin, Song; Zou, Yuhua; Zhang, Chun-Li

    2013-10-01

    Cytokine-induced activation of signal transducer and activator of transcription 3 (STAT3) promotes the regrowth of damaged axons in the adult central nervous system (CNS). Here we show that KLF4 physically interacts with STAT3 upon cytokine-induced phosphorylation of tyrosine 705 (Y705) on STAT3. This interaction suppresses STAT3-dependent gene expression by blocking its DNA-binding activity. The deletion of KLF4 in vivo induces axon regeneration of adult retinal ganglion cells (RGCs) via Janus kinase (JAK)-STAT3 signalling. This regeneration can be greatly enhanced by exogenous cytokine treatment, or removal of an endogenous JAK-STAT3 pathway inhibitor called suppressor of cytokine signalling 3 (SOCS3). These findings reveal an unexpected cross-talk between KLF4 and activated STAT3 in the regulation of axon regeneration that might have therapeutic implications in promoting repair of injured adult CNS.

  5. Cannabinoid receptor CB1 regulates STAT3 activity and its expression dictates the responsiveness to SR141716 treatment in human glioma patients' cells.

    PubMed

    Ciaglia, Elena; Torelli, Giovanni; Pisanti, Simona; Picardi, Paola; D'Alessandro, Alba; Laezza, Chiara; Malfitano, Anna Maria; Fiore, Donatella; Pagano Zottola, Antonio Christian; Proto, Maria Chiara; Catapano, Giuseppe; Gazzerro, Patrizia; Bifulco, Maurizio

    2015-06-20

    Herein we show that a majority of human brain tumor samples and cell lines over-expressed cannabinoid receptor CB1 as compared to normal human astrocytes (NHA), while uniformly expressed low levels of CB2. This finding prompted us to investigate the therapeutic exploitation of CB1 inactivation by SR141716 treatment, with regard to its direct and indirect cell-mediated effects against gliomas. Functional studies, using U251MG glioma cells and primary tumor cell lines derived from glioma patients expressing different levels of CB1, highlighted SR141716 efficacy in inducing apoptosis via G1 phase stasis and block of TGF-β1 secretion through a mechanism that involves STAT3 inhibition. According to the multivariate role of STAT3 in the immune escape too, interestingly SR141716 lead also to the functional and selective expression of MICA/B on the surface of responsive malignant glioma cells, but not on NHA. This makes SR141716 treated-glioma cells potent targets for allogeneic NK cell-mediated recognition through a NKG2D restricted mechanism, thus priming them for NK cell antitumor reactivity. These results indicate that CB1 and STAT3 participate in a new oncogenic network in the complex biology of glioma and their expression levels in patients dictate the efficacy of the CB1 antagonist SR141716 in multimodal glioma destruction.

  6. Identification and functional evaluation of two STAT3 variants in grass carp: Implication for the existence of specific alternative splicing of STAT3 gene in teleost.

    PubMed

    Du, Linyong; Zhou, Hong; Qin, Lei; Wei, He; Zhang, Anying; Yang, Kun; Wang, Xinyan

    2017-11-01

    A STAT family member, STAT3, becomes activated as a DNA binding protein in response to cytokines and growth factors. In teleost, STAT3 cDNA has been cloned and identified in a few species, but only a single STAT3 transcript is revealed in these studies. In the present study, two variants of STAT3 gene generated by alternative splicing were isolated from grass carp and nominated as STAT3α1 and STAT3α2 based on the homology with their mammalian orthologs. In particular, the homologs of STAT3α1/2 were also found in various fish species, including zebrafish, takifugu, tilapia, medaka and goldfish. Intriguingly, sequence alignment and genomic structure analysis revealed that fish STAT3α1/2 are generated through similar alternative splicing events, implying the potential physiological significance of generating STAT3 variants in fish. Grass carp STAT3α1/2 (gcSTAT3α1/2) were ubiquitously expressed although the transcript levels of STAT3α2 were markedly higher than STAT3α1 in all examined tissues. In vivo and in vitro studies showed that the expression patterns of these two variants were similar under the stimulation of immune stimuli. To reveal the role of gcSTAT3α1/2 in fish immunity, their phosphorylation and involvement in IL-17A/F1 mRNA expression were demonstrated in grass carp peripheral blood lymphocytes upon LPS or PHA challenge, providing evidence for the functional conservation of STAT3 signaling in fish. These findings also raise a question of whether both gcSTAT3α1/2 participate in transcriptional regulation in fish. Actually, our results showed that both of them had the ability to translocate into the nucleus upon activation, and to amplify IL-10 signaling, indicating the existence of STAT3 isoforms with functional redundancy in teleost. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Suppression of autophagy augments the radiosensitizing effects of STAT3 inhibition on human glioma cells

    SciTech Connect

    Yuan, Xiaopeng; Du, Jie; Hua, Song; Zhang, Haowen; Gu, Cheng; Wang, Jie; Yang, Lei; Huang, Jianfeng; Yu, Jiahua Liu, Fenju

    2015-01-15

    Radiotherapy is an essential component of the standard therapy for newly diagnosed glioblastoma. To increase the radiosensitivity of glioma cells is a feasible solution to improve the therapeutic effects. It has been suggested that inhibition of signal transducer and activator of transcription 3 (STAT3) can radiosensitize glioma cells, probably via the activation of mitochondrial apoptotic pathway. In this study, human malignant glioma cells, U251 and A172, were treated with an STAT3 inhibitor, WP1066, or a short hairpin RNA plasmid targeting STAT3 to suppress the activation of STAT3 signaling. The radiosensitizing effects of STAT3 inhibition were confirmed in glioma cells. Intriguingly, combination of ionizing radiation exposure and STAT3 inhibition triggered a pronounced increase of autophagy flux. To explore the role of autophagy, glioma cells were treated with 3-methyladenine or siRNA for autophagy-related gene 5, and it was demonstrated that inhibition of autophagy further strengthened the radiosensitizing effects of STAT3 inhibition. Accordingly, more apoptotic cells were induced by the dual inhibition of autophagy and STAT3 signaling. In conclusion, our data revealed a protective role of autophagy in the radiosensitizing effects of STAT3 inhibition, and inhibition of both autophagy and STAT3 might be a potential therapeutic strategy to increase the radiosensitivity of glioma cells. - Highlights: • Inactivation of STAT3 signaling radiosensitizes malignant glioma cells. • STAT3 inhibition triggers a significant increase of autophagy flux induced by ionizing radiation in glioma cells. • Suppression of autophagy further strengthens the radiosensitizing effects of STAT3 inhibition in glioma cells. • Dual inhibition of autophagy and STAT3 induce massive apoptotic cells upon exposure to ionizing radiation.

  8. Loss of STAT3 in Lymphoma Relaxes NK Cell-Mediated Tumor Surveillance.

    PubMed

    Putz, Eva Maria; Hoelzl, Maria Agnes; Baeck, Julia; Bago-Horvath, Zsuzsanna; Schuster, Christian; Reichholf, Brian; Kern, Daniela; Aberger, Fritz; Sexl, Veronika; Hoelbl-Kovacic, Andrea

    2014-01-27

    The transcription factors and proto-oncogenes STAT3 and STAT5 are highly activated in hematological malignancies and represent promising therapeutic targets. Whereas the importance of STAT5 as tumor promoter is beyond doubt, the role of STAT3 in hematological cancers is less well understood. Both, enforced as well as attenuated expression of STAT3 were reported in hematopoietic malignancies. Recent evidence implicates STAT3 as key player for tumor immune surveillance as it both mediates the production of and response to inflammatory cytokines. Here we investigated the effects of STAT3 deletion in a BCR/ABL-induced lymphoma model, which is tightly controlled by natural killer (NK) cells in vivo. Upon STAT3 deletion tumor growth is significantly enhanced when compared to STAT3-expressing controls. The increased tumor size upon loss of STAT3 was accompanied by reduced NK cell infiltration and decreased levels of the cytokine IFN-γ and the chemokine RANTES. Upon transplantation into NK cell-deficient mice differences in lymphoma size were abolished indicating that STAT3 expression in the tumor cells controls NK cell-dependent tumor surveillance. Our findings indicate that STAT3 inhibition in lymphoma patients will impair NK cell-mediated tumor surveillance, which needs to be taken into account when testing STAT3 inhibitors in preclinical or clinical trials.

  9. Stat3 orchestrates interaction between endothelial and tumor cells and inhibition of Stat3 suppresses brain metastasis of breast cancer cells

    PubMed Central

    Chou, Ping-Chieh; Zhou, Aidong; Yang, Phillip; Conrad, Charles A.; Aldape, Kenneth D.; Priebe, Waldemar; Patterson, Cam; Sawaya, Raymond; Xie, Keping; Huang, Suyun

    2015-01-01

    Brain metastasis is a major cause of morbidity and mortality in patients with breast cancer. Our previous studies indicated that Stat3 plays an important role in brain metastasis. Here, we present evidence that Stat3 functions at the level of the microenvironment of brain metastases. Stat3 controlled constitutive and inducible VEGFR2 expression in tumor-associated brain endothelial cells. Furthermore, inhibition of Stat3 by WP1066 decreased the incidence of brain metastases and increased survival in a preclinical model of breast cancer brain metastasis. WP1066 inhibited Stat3 activation in tumor-associated endothelial cells, reducing their infiltration and angiogenesis. WP1066 also inhibited breast cancer cell invasion. Our results indicate that WP1066 can inhibit tumor angiogenesis and brain metastasis mediated by Stat3 in endothelial and tumor cells. PMID:25881542

  10. Stat3 orchestrates interaction between endothelial and tumor cells and inhibition of Stat3 suppresses brain metastasis of breast cancer cells.

    PubMed

    Lee, Hsueh-Te; Xue, Jianfei; Chou, Ping-Chieh; Zhou, Aidong; Yang, Phillip; Conrad, Charles A; Aldape, Kenneth D; Priebe, Waldemar; Patterson, Cam; Sawaya, Raymond; Xie, Keping; Huang, Suyun

    2015-04-30

    Brain metastasis is a major cause of morbidity and mortality in patients with breast cancer. Our previous studies indicated that Stat3 plays an important role in brain metastasis. Here, we present evidence that Stat3 functions at the level of the microenvironment of brain metastases. Stat3 controlled constitutive and inducible VEGFR2 expression in tumor-associated brain endothelial cells. Furthermore, inhibition of Stat3 by WP1066 decreased the incidence of brain metastases and increased survival in a preclinical model of breast cancer brain metastasis. WP1066 inhibited Stat3 activation in tumor-associated endothelial cells, reducing their