Signal Transducer And Activator Of Transcription 1 Inhibitor Research Articles

Geniposide attenuates influenza virus-induced pneumonia by regulating inflammatory cytokines production. Evidences to elucidate the followed pathway

BackgroundInfluenza virus-induced pneumonia (IVP) is an infectious pulmonary disease characterized by exacerbated pulmonary inflammation caused by invasion of the influenza virus. IVP continues to threaten public health due to its high morbidity and mortality rates. Geniposide is one of the major bioactive constituents of G. jasminoides, which exerts antiviral and anti-inflammatory effects on influenza A virus (IAV) infection. PurposeTo investigate therapeutic effects and comprehensive mechanisms of geniposide on IAV infection and subsequent pneumonia. MethodsICR mice were infected intranasally with H1N1 (A/FM/1/47) to detect the anti-IAV activity of geniposide. Proteomics combined with function-integrated analysis were conducted to gain insight into the comprehensive mechanisms of geniposide. Subsequently, western blot was used to detect the phosphorylation of signal transducer and activator of transcription 1 (STAT1), signal transducer and activator of transcription 2 (STAT2), Interferon regulatory factor 9 (IRF9) and Janus kinase 1 (JAK1) in Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling pathway in lung tissue. Finally, RT-qPCR was used to detect the levels of interleukin 6 (IL-6), interleukin 17 (IL-17), interferon-γ (IFN-γ) and the STAT1 inhibitor (fludarabine) was used to verify the targeting between STAT1 and geniposide in RAW cells. ResultsGeniposide could significantly reduce the lung index, diminish lung pathology, decrease the virus loads and the inflammatory cytokines expression induced by IAV infection. A total of 411 differentially expressed proteins were identified among control, model, and geniposide-treated group in proteomic analysis. According to function-integrated analysis, 15 KEGG pathways were enriched and divided into 9 groups (modules), including influenza A, NOD-like receptor signaling, RIG-I-like receptor signaling, and so on. Among these modules, the most intensely interacting module pair was the NOD-like receptor signaling and influenza A, in which STAT1 and STAT2 acted as hubs with critical bridgeness role in the target network of geniposide. This indicated that geniposide may mitigate inflammation and alleviate IVP by JAK/STAT signaling pathways. Moreover, validation experiments confirmed that geniposide can significantly inhibit STAT1 and STAT2 phosphorylation as well as down-regulated expression of IL-6, IFN-γ and IL-17 in lung. Furthermore, when RAW cells were treated with the STAT1 inhibitor (fludarabine), the inhibitory effect of geniposide on IFN-γ and IL-6 was attenuated significantly. ConclusionsGeniposide can attenuate IAV-induced pneumonia by regulating inflammatory cytokines production through the JAK/STAT pathway.

TNIP3 protects against pathological cardiac hypertrophy by stabilizing STAT1

Pathological cardiac hypertrophy is one of the major risk factors of heart failure and other cardiovascular diseases. However, the mechanisms underlying pathological cardiac hypertrophy remain largely unknown. Here, we identified the first evidence that TNFAIP3 interacting protein 3 (TNIP3) was a negative regulator of pathological cardiac hypertrophy. We observed a significant upregulation of TNIP3 in mouse hearts subjected to transverse aortic constriction (TAC) surgery and in primary neonatal rat cardiomyocytes stimulated by phenylephrine (PE). In Tnip3-deficient mice, cardiac hypertrophy was aggravated after TAC surgery. Conversely, cardiac-specific Tnip3 transgenic (TG) mice showed a notable reversal of the same phenotype. Accordingly, TNIP3 alleviated PE-induced cardiomyocyte enlargement in vitro. Mechanistically, RNA-sequencing and interactome analysis were combined to identify the signal transducer and activator of transcription 1 (STAT1) as a potential target to clarify the molecular mechanism of TNIP3 in pathological cardiac hypertrophy. Via immunoprecipitation and Glutathione S-transferase assay, we found that TNIP3 could interact with STAT1 directly and suppress its degradation by suppressing K48-type ubiquitination in response to hypertrophic stimulation. Remarkably, preservation effect of TNIP3 on cardiac hypertrophy was blocked by STAT1 inhibitor Fludaradbine or STAT1 knockdown. Our study found that TNIP3 serves as a novel suppressor of pathological cardiac hypertrophy by promoting STAT1 stability, which suggests that TNIP3 could be a promising therapeutic target of pathological cardiac hypertrophy and heart failure.

Inhibition of complement C3 prevents osteoarthritis progression in guinea pigs by blocking STAT1 activation

Osteoarthritis (OA) is one of the leading causes of disability, affecting over 500 million adults worldwide. Previous studies have found that various inflammatory factors can contribute to the pathogenesis of OA, including complement factors in the synovial fluid of OA patients. However, the pathogenesis of this disease is still not known, and the only therapy of severe OA is total joint replacements. Total joint replacements are invasive, expensive, and affect quality of life. Here we show that when human articular chondrocytes are stimulated with pro-inflammatory mediator interleukin-1β (IL-1β) there is an increase in inflammatory factors including complement component 3 (C3). We also found the transcription factor, signal transducer and activator of transcription 1 (STAT1), is responsible for increased C3 expression after IL-1β stimulation in human articular chondrocytes. A specific STAT1 inhibitor, fludarabine, attenuates the hyper-expression of C3 and delays/prevents spontaneous OA in Dunkin-Hartley guinea pigs. Since fludarabine is already clinically used for chemotherapy, this study has great translational potential as a unique disease-modifying osteoarthritis drug (DMOAD) in treating primary OA.

Analysis of the role of dihydromyricetin derived from vine tea (Ampelopsis grossedentata) on multiple myeloma by activating STAT1/RIG-I axis.

Multiple myeloma (MM) is a plasma cell malignancy and remains incurable as it lacks effective curative approaches; thus, novel therapeutic strategies are desperately needed. The study aimed to explore the therapeutic role of dihydromyricetin (DHM) in MM and explore its mechanisms. Human MM and normal plasma samples, human MM cell lines, and normal plasma cells were used for in vitro experiments. Cell counting kit-8 (CCK-8), flow cytometry, and trans-well assays were performed for the assessment of cell viability, apoptosis, migration, and invasion, respectively. Quantitative real-time polymerase chain reaction (qRT-PCR) was employed to assess the mRNA expression of signal transducer and activator of transcription 1 (STAT1) and retinoic acid-inducible gene I (RIG-I). Western blotting was employed to assess E-cadherin, N-cadherin, signal transducer, STAT1, p-STAT1, and RIG-I protein expression. A tumor xenograft model was used for in vivo experiments. Here, dihydromyricetin (DHM) dose-dependently restrained viability, apoptosis, migration, and invasion, and facilitated apoptosis of U266 cells. After DHM treatment, the E-cadherin level was increased and the N-cadherin level was decreased in U266 and RPMI-8226 cells, suggesting the inhibitory effects of DHM on epithelial-mesenchymal transition (EMT) in MM. Besides, the levels of p-STAT1/STAT1 and RIG-I were down-regulated in MM. However, the STAT1 inhibitor fludarabine undid the suppressive effect of DMH on the malignant characteristics of U266 cells. Also, DHM inhibited MM tumor growth and EMT, and activated STAT1/RIG-I pathway in vivo. Collectively, this study first revealed that DHM can restrain EMT and tumor growth in MM by activating STAT1/RIG-I signaling, which provides a novel drug for the treatment of MM.

Macrophage-Derived Factors with the Potential to Contribute to Pathogenicity of HIV-1 and HIV-2: Role of CCL-2/MCP-1.

Human immunodeficiency virus type 2 (HIV-2) is known to be less pathogenic than HIV-1. However, the mechanism(s) underlying the decreased HIV-2 pathogenicity is not fully understood. Herein, we report that β-chemokine CCL2 expression was increased in HIV-1-infected human monocyte-derived macrophages (MDM) but decreased in HIV-2-infected MDM when compared to uninfected MDM. Inhibition of CCL2 expression following HIV-2 infection occurred at both protein and mRNA levels. By microarray analysis, quantitative PCR, and Western blotting, we identified that Signal Transducer and Activator of Transcription 1 (STAT1), a critical transcription factor for inducing CCL2 gene expression, was also reduced in HIV-2-infected MDM. Blockade of STAT1 in HIV-infected MDM using a STAT1 inhibitor significantly reduced the production of CCL2. In contrast, transduction of STAT1-expressing pseudo-retrovirus restored CCL2 production in HIV-2-infected MDM. These findings support the concept that CCL2 inhibition in HIV-2-infected MDM is meditated by reduction of STAT1. Furthermore, we showed that STAT1 reduction in HIV-2-infected MDM was regulated by the CUL2/RBX1 ubiquitin E3 ligase complex-dependent proteasome pathway. Knockdown of CUL2 or RBX1 restored the expression of STAT1 and CCL2 in HIV-2-infected MDM. Taken together, our findings suggest that differential regulation of the STAT1-CCL2 axis may be one of the mechanisms underlying the different pathogenicity observed for HIV-1 and HIV-2.

A chemical probe inhibitor targeting STAT1 restricts cancer stem cell traits and angiogenesis in colorectal cancer

BackgroundColorectal cancer (CRC) is a worldwide cancer with rising annual incidence. New medications for patients with CRC are still needed. Recently, fluorescent chemical probes have been developed for cancer imaging and therapy. Signal transducer and activator of transcription 1 (STAT1) has complex functions in tumorigenesis and its role in CRC still needs further investigation.MethodsRNA sequencing datasets in the NCBI GEO repository were analyzed to investigate the expression of STAT1 in patients with CRC. Xenograft mouse models, tail vein injection mouse models, and azoxymethane/dextran sodium sulfate (AOM/DSS) mouse models were generated to study the roles of STAT1 in CRC. A ligand-based high-throughput virtual screening approach combined with SWEETLEAD chemical database analysis was used to discover new STAT1 inhibitors. A newly designed and synthesized fluorescently labeled 4’,5,7-trihydroxyisoflavone (THIF) probe (BODIPY-THIF) elucidated the mechanistic actions of STAT1 and THIF in vitro and in vivo. Colonosphere formation assay and chick chorioallantoic membrane assay were used to evaluate stemness and angiogenesis, respectively.ResultsUpregulation of STAT1 was observed in patients with CRC and in mouse models of AOM/DSS-induced CRC and metastatic CRC. Knockout of STAT1 in CRC cells reduced tumor growth in vivo. We then combined a high-throughput virtual screening approach and analysis of the SWEETLEAD chemical database and found that THIF, a flavonoid abundant in soybeans, was a novel STAT1 inhibitor. THIF inhibited STAT1 phosphorylation and might bind to the STAT1 SH2 domain, leading to blockade of STAT1-STAT1 dimerization. The results of in vitro and in vivo binding studies of THIF and STAT1 were validated. The pharmacological treatment with BODIPY-THIF or ablation of STAT1 via a CRISPR/Cas9-based strategy abolished stemness and angiogenesis in CRC. Oral administration of BODIPY-THIF attenuated colitis symptoms and tumor growth in the mouse model of AOM/DSS-induced CRC.ConclusionsThis study demonstrates that STAT1 plays an oncogenic role in CRC. BODIPY-THIF is a new chemical probe inhibitor of STAT1 that reduces stemness and angiogenesis in CRC. BODIPY-THIF can be a potential tool for CRC therapy as well as cancer cell imaging.

Interleukin-35 inhibits lipopolysaccharide-induced endothelial cell activation by downregulating inflammation and apoptosis

Inflammation is an essential factor contributing to sepsis-induced endothelial cell (EC) activation. Interleukin-35 (IL-35) is an anti-inflammatory/immunosuppressive cytokine that exerts protective effects on many inflammatory diseases. In this study, we investigated the effects of IL-35 on lipopolysaccharide (LPS)-induced EC activation and the potential underlying mechanism. Human umbilical vein endothelial cells (HUVECs) were incubated with LPS (1 μg/ml) for 24 h and then cocultured with different concentrations (0, 1, 10, or 100 ng/ml) of recombinant human IL-35 (rhIL-35) for 12 h. Flow cytometry analysis revealed that IL-35 inhibited LPS-induced HUVEC apoptosis in a dose-dependent manner. RT-qPCR and Western blot analyses showed significantly higher mRNA and protein levels of the adhesion molecules intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) and the inflammatory factors IL-6 and IL-8 in the LPS group than in the control group. These changes were alleviated by IL-35 treatment, suggesting that IL-35 protects ECs by downregulating inflammation. Furthermore, IL-35 induced signal transducer and activator of transcription 1 (STAT1) and STAT4 activation and promoted their interaction. Blocking STAT1 or STAT4 expression by fludarabine (STAT1 inhibitor) treatment or siRNA-STAT4-interfering fragment transfection inhibited the protective effect of IL-35 on ECs. Moreover, we observed a similar protective effect of IL-35 treatment on ECs in a mouse sepsis model induced by intraperitoneal LPS injection. This study indicated that IL-35 exerts anti-inflammatory and antiapoptotic effects on LPS-induced EC activation by activating the STAT1 and STAT4 signaling pathways.

VAV2 is required for DNA repair and implicated in cancer radiotherapy resistance

Radiotherapy remains the mainstay for treatment of various types of human cancer; however, the clinical efficacy is often limited by radioresistance, in which the underlying mechanism is largely unknown. Here, using esophageal squamous cell carcinoma (ESCC) as a model, we demonstrate that guanine nucleotide exchange factor 2 (VAV2), which is overexpressed in most human cancers, plays an important role in primary and secondary radioresistance. We have discovered for the first time that VAV2 is required for the Ku70/Ku80 complex formation and participates in non-homologous end joining repair of DNA damages caused by ionizing radiation. We show that VAV2 overexpression substantially upregulates signal transducer and activator of transcription 1 (STAT1) and the STAT1 inhibitor Fludarabine can significantly promote the sensitivity of radioresistant patient-derived ESCC xenografts in vivo in mice to radiotherapy. These results shed new light on the mechanism of cancer radioresistance, which may be important for improving clinical radiotherapy.

Dexmedetomidine Directs T Helper Cells toward Th1 Cell Differentiation via the STAT1-T-Bet Pathway

Dexmedetomidine is an α2 adrenergic receptor agonist that has been reported to modulate the polarization of CD4+ T cells. However, the underlying mechanisms by which dexmedetomidine induces T-helper 1 (Th1) cell differentiation remain poorly understood. The aim of this study was to explore the potential mechanisms through which dexmedetomidine can induce Th1 cell differentiation. Purified CD4+ T cells were stimulated with anti-CD3/anti-CD28 and then treated with dexmedetomidine. Flow cytometry analysis was adopted to measure the concentration of Th1 cells. Enzyme-linked immunosorbent assay (ELISA) and real-time quantitative polymerase chain reaction (qPCR) were performed to detect protein levels and mRNA expression, respectively, of IFN-γ and IL-4. Western blotting was used to determine the phosphorylation of signal transducer and activator of transcription 1 (STAT1) and T-bet expression. The Th1 cell subset and IFN-γ levels were elevated in the dexmedetomidine-induced CD4+ T cells. Dexmedetomidine enhanced the phosphorylation of STAT1 and the expression of T-bet in the CD4+ T cells. Atipamezole (an α2 adrenergic antagonist) and fludarabine (a STAT1 inhibitor) reversed the dexmedetomidine-induced Th1 cell differentiation. These results suggested that dexmedetomidine induced Th1 cell differentiation via the STAT1-T-bet signaling pathway.

Tregs-derived interleukin 35 attenuates endothelial proliferation through STAT1 in pulmonary hypertension.

BackgroundTo explore the source, the role and the specific mechanism of IL-35 and its downstream molecules in the development of pulmonary hypertension.Methods8–10 weeks male mice were undergoing hypoxia combined with SU5416 (HySu) to establish a pulmonary hypertension (PH) model. The phenotype of PH mice was measured by immunohistochemistry and immunofluorescence staining. The levels of two subunits (EBI3 and p35 subunits) in lung tissue were measured by real-time PCR and western blotting. EBI3 monoclonal antibody was administrated as IL-35 neutralization to offset systemic IL-35 expression. Fludarabine, an inhibitor of STAT1 (signal transducer and activator of transcription 1) was used to clarify the role of STAT1 under IL-35 treatment.ResultsAfter pulmonary hypertension, the expression of IL-35 and its two subunits (EBI3 and p35 subunits) in lung tissue were significantly increased. And the two subunits of IL-35 are highly expressed in Treg cells. Compared with the controlled PH mice, the IL-35 neutralization PH mice showed aggravated pulmonary hypertension phenotype. The specific manifestations are the increase of right ventricular systolic pressure (RVSP), the growing proportion of right heart [RV/(LV+S)], and the remodeling of pulmonary blood vessels increases. The expression of pulmonary vascular endothelium (CD31) in PH mice increased, and the proliferation ability of vascular endothelium enhanced after IL-35 was inhibited. IL-35 phosphorylates STAT1 through the receptor GP130 on pulmonary vascular endothelial cells, which in turn inhibits endothelial cell proliferation. IL-35 recombinant protein can reduce the expression of CD31 in lung tissues of PH mice. But the administration of STAT1 inhibitor made it invalid from the IL-35 effect of reversing pulmonary hypertension.ConclusionsTregs-derived IL-35 can reverse the remodeling of pulmonary blood vessels and alleviate the progression of pulmonary hypertension by reducing the proliferation of endothelial cells.

Leukocyte cell‐derived chemotaxin 2 promotes the development of nonalcoholic fatty liver disease through STAT‐1 pathway in mice

Nonalcoholic fatty liver disease (NAFLD), whose pathogenesis remains unelucidated, has become an increasingly prevalent disease globally requiring novel treatment strategies. This study aims to explore the role of leukocyte cell-derived chemotaxin 2 (LECT2), one of the known hepatokines, in the development of NAFLD. The serum LECT2 level was evaluated in patients with NAFLD and male C57BL/6 mice fed a high-fat diet (HFD) for 8weeks. Tail intravenous injection of adeno-associated virus that contained Lect2 short hairpin RNA or Lect2 overexpression plasmid was administered to mice to inhibit or increase hepatic Lect2 expression. Hepatic steatosis was evaluated by histological staining with haematoxylin and eosin and Oil Red O, and also by quantitative hepatic triglyceride measurements. RNA-seq was performed to discover the specific targets of LECT2 on NAFLD. Serum and hepatic LECT2 levels were elevated in NAFLD patients and HFD-fed mice. Inhibition of hepatic Lect2 expression alleviated HFD-induced hepatic steatosis and inflammation, whereas hepatic overexpression of Lect2 aggravated HFD-induced hepatic steatosis and inflammation. RNA-seq and bioinformatical analysis suggested that the signal transducers and activators of transcription-1 (STAT-1) pathway might play an indispensable role in the interaction between LECT2 and NAFLD. A STAT-1 inhibitor could reverse the accumulation of hepatic lipids caused by Lect2 overexpression. LECT2 expression is significantly elevated in NAFLD. LECT2 induces the occurrence and development of NAFLD through the STAT-1 pathway. LECT2 may be a potential therapeutic target for NAFLD.

Upregulation of iNOS Protects Cyclic Mechanical Stretch-Induced Cell Death in Rat Aorta Smooth Muscle Cells.

Aortic dissection and aneurysm are associated with abnormal hemodynamic loads originating from hypertension. Our previous study demonstrated that cyclic mechanical stretch (CMS, mimicked hypertension) caused the death of rat aortic smooth muscle cells (RASMCs) in a mitogen activated-protein kinases (MAPKs)-dependent manner. The current study investigated the effects of inducible nitric oxide synthase (iNOS) on CMS-induced RASMC death. cDNA microarrays for CMS-treated RASMCs showed that iNOS expression levels were increased in response to CMS. Real-time polymerase chain reaction (PCR) analysis demonstrated that this increase was p38 MAPK (p38)-dependent. NO production was also increased. This increase could be inhibited by p38 and iNOS inhibitors. Thus, CMS-induced iNOS synthesized NO. CMS-induced cell death in RASMCs was increased by the iNOS inhibitor but abrogated by the long-acting NO donor DETA-NONOate. Increased iNOS expression was confirmed in the abdominal aortic constriction mouse model. Signal transducers and activators of transcription 1 (STAT1) was activated in stretched RASMCs, and iNOS expression and NO production were inhibited by the STAT1 inhibitor nifuroxazide. Our findings suggest that RASMCs were protected by iNOS from CMS-stimulated cell death through the STAT1 and p38 signal pathways independently.

Downregulation of STAT1 induces the differentiation of neural stem cells through JNK pathway

Neural stem cells (NSCs) generated neurons and glial cells. Thus, it is a preferable candidate to the cell replacement-based therapy against neural disorders. The signaling pathways that regulate differentiation of NSCs are widely studied. In the current study, we used in vitro culture system to elucidate the role of signal transducer and activator of transcription 1 (STAT1) in NSCs’ differentiation. Downregulation of STAT1 inhibited the proliferation of NSCs. Meanwhile, we also found STAT1 regulation could control the differentiation of NSCs. More neurons and glia cells were generated from NSCs with STAT1 silencing. This process was mediated by the JNK/STAT1 signaling. STAT1 inhibitor promoted differentiation of NSCs. After transplantation, we observed more neurons generated from NSCs with shRNA-STAT1 treatment. Collectively, this work showed an efficient way to regulate neuronal differentiation of NSCs through regulating the STAT1 expression. This is likely to provide source and theoretical support to cell replacement based theory.

Activation of the STAT1 Pathway Accelerates Periodontitis in Nos3-/- Mice

Early studies on the etiology and pathogenesis of hypertension have shown that it has a considerable association with inflammation and the immune response as well as periodontitis. Clinical studies have also shown that hypertension can promote the periodontal tissue destruction caused by periodontitis. However, the underlying mechanisms remain unclear. This study aimed to explore the possible mechanisms of how hypertension aggravates periodontitis. Treatment with or without the signal transducer and activator of transcription 1 (STAT1) inhibitor fludarabine was performed in an endothelial nitric oxide synthase gene knockout-related (Nos3-/-) mouse model with the hypertension phenotype of periodontitis induced by bacteria. Micro–computed tomography, immunohistochemistry, Western blot, quantitative reverse transcription polymerase chain reaction, immunofluorescence, and ELISA were performed. We demonstrated that Nos3-/--related hypertension increases bone resorption and periodontal destruction in periodontitis lesion areas, which can be inhibited by the STAT1 inhibitor. Experimental data also showed that Nos3-/- significantly increased macrophage infiltration and proinflammatory cytokine expression in the periodontitis lesion area, which is dependent on the angiotensin II–induced STAT1 pathway. Inhibition of STAT1 in vivo can decrease the expression of proinflammatory cytokines and macrophage infiltration. Furthermore, data in this study showed that Nos3-/--related hypertension further downregulated the STAT3 anti-inflammatory function and its downstream chemokine expression in a STAT1-dependent manner. By applying RAW 264.7 and L929 cell lines and monocytes isolated from Nos3-/- mice, we confirmed that activation of the STAT1 pathway inhibits STAT3 and its downstream pathway and promotes inflammatory cytokine expression in vitro. Collectively, our current study demonstrated that STAT1 plays an indispensable role in the Nos3-/--related hypertension with aggravation of periodontitis, suggesting that STAT1 may be a key target for the treatment of periodontitis with hypertension.

Role of STAT1 and Oxidative Stress in Gentamicin-Induced Hair Cell Death in Organ of Corti.

Oxidative stress plays a critical role in gentamicin-induced hair cell death. Previous work has implicated the cytoplasmic transcription factor signal transducer and activator of transcription 1 (STAT1) as a potential mediator of drug-induced ototoxicity, but role in aminoglycosides is largely unknown. This study investigated aminoglycosides-induced cell death, exploring contributions of reactive oxygen species and STAT1 pathway in injury and protection. Neonatal murine organ of Corti explants from 2 to 3 day postnatal pups (n = 96) were treated with gentamicin at (4 μM, 50 μM) for 4 to 72 hours, with/without protectants. Effects on STAT1 pathway and gentamicin-induced hair cell death were measured with 50 μM Epigallocatechin gallate (EGCG, a STAT1 inhibitor) and all-trans retinoic acid (atRA, a STAT1 activator). Hair cell morphology was evaluated and hair cell loss was quantified with cytocochleograms. Mitochondrial membrane potential was assayed and superoxide generation and suppression was measured with dihydroethidium (DHE) staining. Co-administration of 50 μM EGCG conferred protection from 4 μM gentamicin toxicity (p < 0.001), whereas atRA potentiated gentamicin-induced hair cell death (p < 0.001). On immunohistochemistry, STAT1 phosphorylation at theserine 727 (Ser) residues was increased at 72 hours with 4 μM gentamicin. With administration of 50 μM gentamicin, there was activation of STAT1 Tyr at 4 hours and STAT1 Ser at 16 hours. Gentamicin dissipated mitochondrial membrane potentials, and EGCG attenuated gentamicin-induced oxidative stress at 72 hours. EGCG protected outer hair cells from gentamicin toxicity in a cochlear explant model, with the underlying mechanism involving both reactive oxygen species (ROS) suppression and STAT1 inhibition.

Dimerization of EGFR and HER2 induces breast cancer cell motility through STAT1-dependent ACTA2 induction.

The dimerization of EGFR and HER2 is associated with poor prognosis such as induction of tumor growth and cell invasion compared to when EGFR remains as a homodimer. However, the mechanism for events after dimerization in breast cancer models is not clear. We found that expressions of alpha-smooth muscle actin (ACTA2) and signal transducer and activator of transcription 1 (STAT1) significantly increased with transient or stable overexpression of HER2 in EGFR-positive breast cancer cells. ACTA2 and STAT1 expression was also increased in HER2-positive breast cancer patients. In contrast, ACTA2 expression was decreased by HER2 siRNA. Next, we investigated the co-relation between STAT1 and ACTA2 expression. Basal ACTA2 expression was significantly decreased by treatment with the STAT1 inhibitor fludarabine or the JAK2 inhibitor AG490. In contrast, ACTA2 expression was increased by STAT1 overexpression. Levels of ACTA2, STAT1, and HER2 were increased and relapse free survival was decreased in high-risk breast cancer patients. We also investigated the effect of ACTA2 on cell motility, which was suppressed by ACTA2 shRNA overexpression in MDA-MB231 HER2 and 4T1 mammary carcinoma cells. The number of lung metastatic nodules was significantly decreased in ACTA2 knockdown mice. Taken together, these results demonstrated that induction of ACTA2 by EGFR and HER2 dimerization was regulated through a JAK2/STAT1 signaling pathway, and aberrant ACTA2 expression accelerated the invasiveness and metastasis of breast cancer cells.

STAT1 inhibitor alleviates spinal cord injury by decreasing apoptosis.

Spinal cord injury (SCI) is typically caused by trauma or disease, and it severely affects patients' motor function. The relationship between signal transducers and activators of transcription-1 (STAT1) and neuronal death after cerebral focal ischemia has been comprehensively studied, but its role in SCI remains largely unknown. This study investigated the protective effect of an STAT1 inhibitor on SCI. Thirty SD rats were SCI-induced and were then randomly divided into two groups (N = 15 each), either receiving STAT1 or the STAT1 inhibitor S1491 by intraperitoneal injection. The motor dysfunction of the rats was evaluated by behavioral scores, followed by the examination of SCI by hematoxylin and eosin staining. Apoptosis was also detected by Western blot and terminal deoxynucleotidyl transferase-mediated dUTP nick-end-labeling (TUNEL) assay. The motor functions of rats receiving STAT1 did not score as well as the STAT1 inhibitor group (P < 0.01). Further assays showed remarkable improvements in pathological damage to spinal code tissue in STAT1 inhibitor-treated rats, along with lower Bax and higher Bcl-2 expression. The STAT1 inhibitor also suppressed the occurrence of TUNEL-positive cells compared to the STAT1-treated group. In summary, we suggest that the STAT1 inhibitor alleviates SCI by decreasing apoptosis.

AMPK Suppresses Vascular Inflammation In Vivo by Inhibiting Signal Transducer and Activator of Transcription-1.

Activation of AMPK suppresses inflammation, but the underlying mechanisms remain poorly understood. This study was designed to characterize the molecular mechanisms by which AMPK suppresses vascular inflammation. In cultured human aortic smooth muscle cells, pharmacologic or genetic activation of AMPK inhibited the signal transducer and activator of transcription-1 (STAT1), while inhibition of AMPK had opposite effects. Deletion of AMPKα1 or AMPKα2 resulted in activation of STAT1 and in increases in proinflammatory mediators, both of which were attenuated by administration of STAT1 small interfering RNA or fludarabine, a selective STAT1 inhibitor. Moreover, AMPK activation attenuated the proinflammatory actions induced by STAT1 activators such as interferon-γ and angiotensin II (AngII). Mechanistically, we found that AMPK activation increased, whereas AMPK inhibition decreased, the levels of mitogen-activated protein kinase phosphatase-1 (MKP-1), an inducible nuclear phosphatase, by regulating proteasome-dependent degradation of MKP-1. Gene silencing of MKP-1 increased STAT1 phosphorylation and prevented 5-aminoimidazole-4-carboxyamide ribonucleoside–reduced STAT1 phosphorylation. Finally, we found that infusion of AngII caused a more severe inflammatory response in AMPKα2 knockout mouse aortas, all of which were suppressed by chronic administration of fludarabine. We conclude that AMPK activation suppresses STAT1 signaling and inhibits vascular inflammation through the upregulation of MKP-1.

Gomisin A Enhances Tumor Necrosis Factor-α-Induced G1 Cell Cycle Arrest &lt;i&gt;via&lt;/i&gt; Signal Transducer and Activator of Transcription 1-Mediated Phosphorylation of Retinoblastoma Protein

Gomisin A, a dibenzocyclooctadiene lignan isolated from the fruit of Schisandra chinensis, has been reported as an anti-cancer substance. In this study, we investigated the effects of gomisin A on cancer cell proliferation and cell cycle arrest in HeLa cells. Gomisin A significantly inhibited cell proliferation in a dose-dependent manner after 72 h treatment, especially in the presence of tumor necrosis factor-α (TNF-α), due to cell cycle arrest in the G1 phase with the downregulation of cyclin D1 expression and Retinoblastoma (RB) phosphorylation. In addition, gomisin A in combination with TNF-α strongly suppressed the expression of signal transducer and activator of transcription 1 (STAT1). Inhibition of STAT1 pathways by a small-interfering RNA against STAT1 and AG490 Janus kinase (JAK) kinase inhibitor AG490 reduced the cyclin D1 expression and RB phosphorylation, indicating that JAK-mediated STAT1 activation is involved in gomisin A-induced G1 cell cycle arrest.

STAT1 promotes radioresistance of CD44+/CD24−/low cells in breast cancer

Breast cancer-initiating cells are a relatively radioresistant subpopulation of breast cancer cells. However, the mechanism of this radioresistance is still unclear. This study aimed to investigate the effect of radiation on the levels of signal transducer and activator of transcription 1 (STAT1) in mammospheres of cancer-initiating cells and monolayer cultures of MCF-7 cells. We isolated CD44(+)/CD24(-/low) cancer-initiating cells from MCF-7 cells and propagated them as mammospheres. Next we used realtime quantitative reverse-transcriptase polymerase chain reaction to examine the mRNA level of STAT1 in mammospheres of breast cancer-initiating cells and monolayer cultures of MCF-7 cells. The apoptosis rate and surviving fraction using clonogenic assays was observed after treating the cells with a STAT1 inhibitor. After irradiation, the STAT1 level in the mammospheres was higher than that in the monolayer cultures. STAT1 inhibitor treatment did not cause significant changes in the apoptosis rate and surviving fraction in the MCF-7 monolayer cultures. However, the inhibitor treatment caused significant differences in the apoptosis rate and surviving fraction in mammospheres. Our study provides the first evidence that STAT1 signaling contributes to radioresistance in breast cancer-initiating cells and reveals STAT1 as a promising target to reduce radioresistance and enhance the efficacy of radiotherapy for breast cancer.