Signal Transducer And Activator Of Transcription 3 Inhibitor Research Articles

STAT3 promotes migration and invasion of cholangiocarcinoma arising from choledochal cyst by transcriptionally inhibiting miR200c through the c-myb/MEK/ERK signaling pathway.

Signal transducer and activator of transcription 3 (STAT3) have been highlighted in cancer regulation. Its roles in Cholangiocarcinoma (CCA) arising from the choledochal cyst (CC) were unclear. Here, we attempted to elucidate the roles of STAT3 in CCA-CC and explore its mechanism. A total of 20 patients with CCA arising from CC, that underwent CC excision in the infant stage were included. The expressions of STAT3, miR200c and c-Myb in clinical samples were assessed by RT-qPCR and/or western blot. Their expression correlations in tumor tissues were evaluated by Pearson correlation analysis. Their roles in CCA cell migration and invasion were investigated by gene silence using siRNA or miRNA inhibitor mediated approach and MEK activator. The expression levels of EMT, metastasis and MEK/ERK pathway-related proteins were checked by western blot. The high expressions of STAT3 and c-Myb, and low expression of miR200c were detected in CCA samples. We defined the transcription inhibition of STAT3 in miR200c expression and the negative correlation between miR200c and c-Myb expression. Silence of STAT3 increased miR200c expression and retarded the migration and invasion of CCA cells, accompanied by decreased levels of Vimentin, N-cadherin, MMP2 and MMP9, and elevated expression of E-cadherin, resulting in inactivating MEK/ERK pathway. MiR200c inhibitor reversed the changes induced by STAT3 silence, which was restored by si-c-Myb. MEK activator significantly reversed the inactivation of the MEK/ERK pathway induced by si-STAT3+miR200c inhibitor+si-c-Myb. In summary, the silence of STAT3 suppressed metastasis and progression of CCA cells by regulating miR200c through the c-Myb mediated MEK/ERK pathway, suggesting STAT3 is the effective target for CCA arising from CC.

Src Kinase Activates Signal Transducer and Activator of Transcription 3 to Enhance Radioresistance in Triple Negative Breast Cancer Cells

Triple-negative breast cancer (TNBC) is known to be insensitive to radiation therapy, and it requires to development radiosensitizer for TNBC. We previously established a radioresistant sub-line from MDA-MB-231 cells, called 231-RR, and have demonstrated that 231-RR cells displayed high cancer stem cell (CSC) activity. And 231-RR cells were treated with dasatinib, a Src inhibitor. In the present study, we discovered that the activation of Src kinase was increased in 231-RR cells. Treatment of dasatinib, an Src inhibitor, sensitized 231-RR cells to radiotherapy, along with the increased p-γH2Axser139, indicated the enhancement of DNA damage. Dasatinib also caused the downregulation of cancer stemness factors, including c-Myc, OCT4, and the Notch intracellular domain, as well as the decrease of phosphorylation of Signal Transducer and Activator of Transcription 3 (STAT3). The treatment of C188-9, a STAT3 inhibitor, also sensitized 231-RR cells toward radiotherapy along with the increased p-γH2Axser139, but without changing the phosphorylation of Src, indicating that STAT3 is a downstream event of Src activation. In conclusion, our data suggests that the inhibitors of Src or STAT3 could function as radiosensitizers or CSC targeting agents for TNBC radiotherapy.

A new triazolyl-indolo-quinoxaline induces apoptosis in gastric cancer cells by abrogating the STAT3/5 pathway through upregulation of PTPεC.

Signal transducer and activator of transcription 3 (STAT3) and STAT5 are the transcription factors that have been studied extensively in relevance to the development of cancers in humans. Suppression of either STAT3 or STAT5-mediated signaling events has been demonstrated to be effective in inducing cytotoxicity in cancer cells. Herein, new hybrids of triazolyl-indolo-quinoxaline are synthesized and examined for their effect on the activation of STAT3 and STAT5 pathways in gastric cancer (GC) cells. Among the newly synthesized compounds, 2,3-difluoro-6-((1-(3-fluorophenyl)-1H-1,2,3-triazol-5-yl)methyl)-6H-indolo[2,3-b]quinoxaline (DTI) displayed selective cytotoxicity against GC cells over their normal counterpart. Flow cytometric analysis, annexin-V-fluorescein isothiocyanate staining, terminal deoxynucleotidyl transferase dUTP nick end labeling assay, live and dead assay, and caspase activation experiments suggested DTI as a potent inducer of apoptosis. The mechanistic approach revealed that DTI imparts cytotoxicity via downregulating the phosphorylation of STAT3Y705 and STAT5Y694/699 . DTI significantly reduced the nuclear pool of STAT3/STAT5 and reduced the DNA interaction ability of STAT3/STAT5 as evidenced by immunofluorescence and electrophoretic mobility shift assay. Further investigation revealed that inhibitory effects towards STAT proteins were mediated through the suppression of upstream kinases such as JAK1, JAK2, and Src. Treatment of GC cells with pervanadate counteracted the DTI-driven STAT3/STAT5 inhibition suggesting the involvement of tyrosine phosphatase. Upon DTI exposure, there was a significant upregulation in the mRNA and protein expression of PTPεC, which is a negative regulator of the JAK-STAT pathway. Knockdown of PTPεC suppressed the DTI-induced STATs inhibition in GC cells. Taken together, triazolyl-indolo-quinoxaline is presented as a new inhibitor of the STAT3/STAT5 pathway in GC cells.

Therapeutic potential and protective role of GRK6 overexpression in pulmonary arterial hypertension

Abnormal proliferation of pulmonary arterial smooth muscle cells (PASMCs) is a key mechanism in the development of pulmonary arterial hypertension (PAH). Signal transducer and activator of transcription 3 (STAT3) signalling plays a critical role in modulating PASMC proliferation, and G-protein-coupled receptor kinase 6 (GRK6) regulates the STAT3 pathway. However, the mechanism underlying the relationship between GRK6 and PAH remains unclear. In this study, we aimed to investigate the role of GRK6 in PAH and determine its potential as a therapeutic target. We utilised hypoxia- and SU5416-induced PAH mouse models and a monocrotaline-induced PAH rat model to analyse the involvement of GRK6. We conducted gain- and loss-of-function experiments using mouse PASMCs. Modulation of GRK6 expression was achieved via a lentiviral vector in vitro and an adeno-associated virus serotype 1 encoding GRK6 in vivo. GRK6 was significantly downregulated in the lung tissues of PAH mice and rats, predominantly in PASMCs. Knockout of GRK6 exacerbated PAH, while both therapeutic and prophylactic overexpression of GRK6 alleviated PAH, as evidenced by a reduction in right ventricular systolic pressure, right ventricular wall to left ventricular wall plus ventricular septum ratio, pulmonary vascular media thickness, and pulmonary vascular muscularisation. Mechanistically, GRK6 overexpression attenuated hypoxia-induced PASMC proliferation and STAT3 phosphorylation. Conversely, knockdown of GRK6 promoted hypoxia-induced proliferation, which was mitigated by a STAT3 inhibitor. Our findings highlight the potential protective and beneficial roles of GRK6 in PAH; we propose a lung-targeted GRK6 gene therapy utilizing adeno-associated virus serotype 1 as a potential treatment approach for patients with PAH.

Melatonin attenuates cellular senescence and apoptosis in diabetic nephropathy by regulating STAT3 phosphorylation

AimsMelatonin is an endogenous hormone related to the regulation of biorhythm. Previous researchers have found that melatonin can ameliorate diabetic nephropathy (DN), but the mechanism remains to be elucidated. To discover the possible mechanism by which melatonin prevents DN, we investigated the potential effects of melatonin on signal transducer and activator of transcription 3 (STAT3) on the progression of cellular senescence and apoptosis. Main methodsCellular senescence, apoptosis and the underlying mechanism of melatonin were investigated both in vivo and in vitro. C57BL/6 mice were intraperitoneally injected with streptozotocin (STZ) to establish DN. For an in vitro model of DN, human renal cortex proximal epithelial tubule (HK-2) cells were exposed to high glucose conditions. Key findingsMelatonin inhibited the phosphorylation of STAT3, decreased the expression of senescence proteins p53, p21 and p16INK4A. Melatonin also downregulated the expression of apoptotic proteins, including cleaved PARP1, cleaved caspase-9 and -3. Melatonin treatment decreased the positive area of senescence-associated galactosidase (SA-β-gal) staining and the number of TUNEL-positive cells in kidneys of DN mice. In vitro, melatonin inhibited STAT3 phosphorylation and lowered cellular senescence and apoptosis markers, in a manner similar to the STAT3 inhibitor S3I-201. In addition, the inhibition effect of melatonin on cellular senescence and apoptosis in HK-2 cells was reversed by the usage of recombinant IL-6 (rIL-6), which can induce STAT3 phosphorylation. SignificanceWe, for the first time, demonstrate that melatonin inhibits STAT3 phosphorylation, which is involved in alleviating the cellular senescence and apoptosis in DN.

Autophagy inhibition mediated by trillin promotes apoptosis in hepatocellular carcinoma cells via activation of mTOR/STAT3 signaling.

Apoptosis and autophagy have been shown to act cooperatively and antagonistically in self-elimination process. On the one side, apoptosis and autophagy can act as partners to induce cell death in a coordinated or cooperative manner; on the flip side, autophagy acts as an antagonist to block apoptotic cell death by promoting cell survival. Our previous research indicated that trillin could induce apoptosis of PLC/PRF/5 cells, but the effects of trillin on autophagy as well as its functional relationship to apoptosis have not been elucidated. Here, the running study aims to investigate the function and molecular mechanism of trillin on autophagy with hepatocellular carcinoma (HCC) cells. The objective of this study is to investigate the molecular mechanism of trillin on autophagy in HCC cells. Protein levels of autophagy markers beclin1, LC3B, and p62 were detected by western blotting. 6-Hydroxyflavone and stattic were used to test the role of trillin regulation of autophagy via serine threonine kinase (AKT)/extracellular-regulated protein kinases (ERK) 1/2/mammalian target of rapamycin (mTOR)/signal transducer and activator of transcription 3 (STAT3) signaling pathway. Flow cytometry was used to detect caspase 3 activity and apoptosis in PLC/PRF/5 cells treated with trillin for 24 h with or without rapamycin, stattic, and 6-hydroxyflavone. The protein level of autophagy marker beclin1 was decreased, whilst the protein level of p62 was significantly increased by trillin treatment, indicating trillin treatment led to inhibition of autophagy in HCC cells. Trillin treatment could reduce the protein levels of p-AKT and p-ERK1/2, but enhance the protein levels of mTOR and p-mTOR, suggesting that trillin could inhibit AKT/ERK rather than mTOR. The AKT/ERK activator 6-hydroxyflavone could reverse the loss of AKT and ERK1/2 phosphorylation induced by trillin, implying that trillin impairs autophagy through activated mTOR rather than AKT/ERK. STAT3 and p-STAT3 were significantly upregulated by the trillin treatment with an increase in dose from 0 to 50 μM, suggesting that autophagy inhibition is mediated by trillin via activation of STAT3 signaling. The STAT3 inhibitor stattic significantly reversed the increased STAT3 phosphorylation at tyrosine 705 induced by trillin. The mTOR signaling inhibitor rapamycin reversed the trillin-induced mTOR phosphorylation enhancement but exerted no effects on total mTOR levels, suggesting trillin treatment led to inhibition of autophagy in HCC cells through activating mTOR/STAT3 pathway. Furthermore, caspase 3 activities and the total rate of apoptosis were increased by trillin treatment, which was reversed by rapamycin, stattic, and 6-hydroxyflavone, proving that trillin promotes apoptosis via activation of mTOR/STAT3 signaling. Trillin induced autophagy inhibition and promoted apoptosis in PLC/PRF/5 cells via the activation of mTOR/STAT3 signaling. Trillin has the potential to be a viable therapeutic option for HCC treatment.

Inhibitory effect and molecular mechanism of sinomenine on human hepatocellular carcinoma HepG2 and SK-HEP-1 cells

This study aimed to investigate the effect and molecular mechanism of sinomenine on proliferation, apoptosis, metastasis, and combination with inhibitors in human hepatocellular carcinoma HepG2 cells and SK-HEP-1 cells. The effect of sinomenine on the growth ability of HepG2 and SK-HEP-1 cells were investigated by CCK-8 assay, colony formation assay, and BeyoClick~(TM) EdU-488 staining. The effect of sinomenine on DNA damage was detected by immunofluorescence assay, and the effect of sinomenine on apoptosis of human hepatocellular carcinoma cells was clarified by Hoechst 33258 staining and CellEvent~(TM) Cystein-3/7Green ReadyProbes~(TM) reagent assay. Cell invasion assay and 3D tumor cell spheroid invasion assay were performed to investigate the effect of sinomenine on the invasion ability of human hepatocellular carcinoma cells in vitro. The effect of sinomenine on the regulation of protein expression related to the protein kinase B(Akt)/mammalian target of rapamycin(mTOR)/signal transducer and activator of transcription 3(STAT3) signaling pathway in HepG2 and SK-HEP-1 cells was examined by Western blot. Molecular docking was used to evaluate the strength of affinity of sinomenine to the target cysteinyl aspartate specific proteinase-3(caspase-3) and STAT3, and combined with CCK-8 assay to detect the changes in cell viability after combination with STAT3 inhibitor JSI-124 in combination with CCK-8 assay. The results showed that sinomenine could significantly reduce the cell viability of human hepatocellular carcinoma cells in a concentration-and time-dependent manner, significantly inhibit the clonogenic ability of human hepatocellular carcinoma cells, and weaken the invasive ability of human hepatocellular carcinoma cells in vitro. In addition, sinomenine could up-regulate the cleaved level of poly ADP-ribose polymerase(PARP), a marker of apoptosis, and down-regulate the protein levels of p-Akt, p-mTOR, and p-STAT3 in human hepatocellular carcinoma cells. Molecular docking results showed that sinomenine had good affinity with the targets caspase-3 and STAT3, and the sensitivity of sinomenine to hepatocellular carcinoma cells was diminished after STAT3 was inhibited. Therefore, sinomenine can inhibit the proliferation and invasion of human hepatocellular carcinoma cells and induce apoptosis, and the mechanism may be attributed to the activation of caspase-3 signaling and inhibition of the Akt/mTOR/STAT3 pathway. This study can provide a new reference for the in-depth research and clinical application of sinomenine and is of great significance to further promote the scientific development and utilization of sinomenine.

Pirfenidone ameliorates liver steatosis by targeting the STAT3-SCD1 axis.

Previous studies reported that pirfenidone (PFD) is associated with liver disease. However, the effects of pirfenidone on energy metabolism and hepatic lipid accumulation are still poorly understood. In this study, C57BL/6J mice were randomly divided into two groups, and fed a normal chow diet (NCD) or a high-fat diet (HFD) for 16weeks. At the end of the eighth week, half of the mice fed on both diets were treated with PFD. Biochemical and lipid metabolism-related indices were analyzed. Furthermore, Hepa 1-6 cells and mouse primary hepatocytes (MPHs) were incubated with PFD with or without free fatty acid (FFA) treatment. Then, stattic (a p-STAT3 inhibitor) or Ad-shSTAT3 was used to further elucidate the effects of Signal Transducer and Activator of Transcription 3 (STAT3) signaling on PFD regulation of hepatic steatosis. PFD ameliorated obesity and hepatic lipid deposition in HFD mice by decreasing stearoyl-CoA desaturase 1 (SCD1) expression and upregulating p-STAT3 in the liver. In Hepa 1-6 cells and MPHs, PFD also down-regulated the expression of SCD1. STAT3 inhibition treatment eliminated the benefits of PFD on both SCD1 and hepatic steatosis. In summary, our data reveal that PFD may play an important role in mitigating hepatic steatosis in a STAT3-SCD1-dependent manner.

Exercise improves cardiac function and attenuates myocardial inflammation and apoptosis by regulating APJ/STAT3 in mice with stroke

Stroke can induce cardiac dysfunction without a primary cardiac disease. Exercise can promote the overall rehabilitation of stroke patients and be beneficial for all kinds of heart diseases. However, the mechanisms underlying the protective effects of exercise in stroke-induced cardiac dysfunction are poorly understood. Hence, we aimed to distinguish the different effects of acute and long-term exercise and further study the mechanism of protection against cardiomyopathy caused by stroke. Mice underwent a single acute session or long-term exercise for 30 days, followed by middle cerebral artery occlusion surgery. The expression of apoptosis-related proteins and proinflammatory factors in the heart was evaluated. Then, overexpression of apelin peptide jejunum (APJ) transfected adeno-associated virus type 9 (AAV9) and inhibition of signal transducer and activator of transcription 3 (STAT3) by Stattic were used in stroke mice or hypoxic cardiomyocytes. ML221 were used to inhibit APJ activity in exercise mouse. Thereafter, changes in apoptotic and proinflammatory factors were evaluated. The results demonstrated that chronic exercise prevented myocardial inflammation, apoptosis and cardiac dysfunction after stroke. However, acute exercise did not have similar effects. Exercise maintained the levels of APJ expression and decreased phosphorylated-STAT3 (p-STAT3) activation to protect cardiomyocytes. Moreover, APJ overexpression promoted cardiomyocyte survival and reduced p-STAT3 levels. STAT3 inhibition also reduced apoptosis and proinflammatory factors in mice hearts. Conversely, the protective effect of exercise was eliminated by APJ inhibition. This study showed that exercise can maintain APJ expression and inhibit p-STAT3, thus, conferring protection against myocardial inflammation and apoptosis induced by stroke.

Constitutive aryl hydrocarbon receptor facilitates the regenerative potential of mouse bone marrow mesenchymal stromal cells.

Bone marrow mesenchymal stromal cells (BMSCs) are the commonly used seed cells in tissue engineering. Aryl hydrocarbon receptor (AhR) is a transcription factor involved in various cellular processes. However, the function of constitutive AhR in BMSCs remains unclear. To investigate the role of AhR in the osteogenic and macrophage-modulating potential of mouse BMSCs (mBMSCs) and the underlying mechanism. Immunochemistry and immunofluorescent staining were used to observe the expression of AhR in mouse bone marrow tissue and mBMSCs. The overexpression or knockdown of AhR was achieved by lentivirus-mediated plasmid. The osteogenic potential was observed by alkaline phosphatase and alizarin red staining. The mRNA and protein levels of osteogenic markers were detected by quantitative polymerase chain reaction (qPCR) and western blot. After coculture with different mBMSCs, the cluster of differentiation (CD) 86 and CD206 expressions levels in RAW 264.7 cells were analyzed by flow cytometry. To explore the underlying molecular mechanism, the interaction of AhR with signal transducer and activator of transcription 3 (STAT3) was observed by co-immunoprecipitation and phosphorylation of STAT3 was detected by western blot. AhR expressions in mouse bone marrow tissue and isolated mBMSCs were detected. AhR overexpression enhanced the osteogenic potential of mBMSCs while AhR knockdown suppressed it. The ratio of CD86+ RAW 264.7 cells cocultured with AhR-overexpressed mBMSCs was reduced and that of CD206+ cells was increased. AhR directly interacted with STAT3. AhR overexpression increased the phosphorylation of STAT3. After inhibition of STAT3 via stattic, the promotive effects of AhR overexpression on the osteogenic differentiation and macrophage-modulating were partially counteracted. AhR plays a beneficial role in the regenerative potential of mBMSCs partially by increasing phosphorylation of STAT3.

Synthesis and Biological Evaluation of Novel 2-Amino-1,4-Naphthoquinone Amide-Oxime Derivatives as Potent IDO1/STAT3 Dual Inhibitors with Prospective Antitumor Effects.

Indoleamine-2,3-dioxygenase 1 (IDO1) and signal transducer and activator of transcription 3 (STAT3) have emerged as significant targets in the tumor microenvironment for cancer therapy. In this study, we synthesized three novel 2-amino-1,4-naphthoquinone amide-oxime derivatives and identified them as dual inhibitors of IDO1 and STAT3. The representative compound NK3 demonstrated effective binding to IDO1 and exhibited good inhibitory activity (hIDO1 IC50 = 0.06 μM), leading to its selection for further investigation. The direct interactions between compound NK3 and IDO1 and STAT3 proteins were confirmed through surface plasmon resonance analysis. A molecular docking study of compound NK3 revealed key interactions between NK3 and IDO1, with the naphthoquinone-oxime moiety coordinating with the heme iron. In the in vitro anticancer assay, compound NK3 displayed potent antitumor activity against selected cancer cell lines and effectively suppressed nuclear translocation of STAT3. Moreover, in vivo assays conducted on CT26 tumor-bearing Balb/c mice and an athymic HepG2 xenograft model revealed that compound NK3 exhibited potent antitumor activity with low toxicity relative to 1-methyl-L-tryptophan (1-MT) and doxorubicin (DOX). Overall, these findings provided evidence that the dual inhibitors of IDO1 and STAT3 may offer a promising avenue for the development of highly effective drug candidates for cancer therapy.

A large-scale transcriptional analysis reveals herb-derived ginsenoside F2 suppressing hepatocellular carcinoma via inhibiting STAT3

BackgroundHepatocellular carcinoma (HCC) is a common type of cancer that shows great morbidity and mortality rates. However, there are limited available drugs to treat HCC. AimThe present work focused on discovering the potential anti-HCC compounds from traditional Chinese medicine (TCM) by employing high-throughput sequencing-based high-throughput screening (HTS2) together with the liver cancer pathway-associated gene signature. MethodsHTS2 assay was adopted for identifying herbs. Protein-protein interaction (PPI) network analysis and computer-aided drug design (CADD) were used to identify key targets and screen the candidate natural products of herbs. Molecular docking, network pharmacology analysis, western blotting, immunofluorescent staining, subcellular fractionation experiment, dual-luciferase reporter gene assay, surface plasmon resonance (SPR) as well as nuclear magnetic resonance (NMR) were performed to validate the ability of compound binding with key target and inhibiting its function. Moreover, cell viability, colony-forming, cell cycle assay and animal experiments were performed to examine the inhibitory effect of compound on HCC. ResultsWe examined the perturbation of 578 herb extracts on the expression of 84 genes from the liver cancer pathway, and identified the top 20 herbs significantly reverting the gene expression of this pathway. Signal transducer and activator of transcription 3 (STAT3) was identified as one of the key targets of the liver cancer pathway by PPI network analysis. Then, by analyzing compounds from top 20 herbs utilizing CADD, we found ginsenoside F2 (GF2) binds to STAT3 with high affinity, which was further validated by the results from molecular docking, SPR and NMR. Additionally, our results showed that GF2 suppresses the phosphorylation of Y705 of STAT3, inhibits its nuclear translocation, decreases its transcriptional activity and inhibits the growth of HCC in vitro and in vivo. ConclusionBased on this large-scale transcriptional study, a number of anti-HCC herbs were identified. GF2, a compound derived from TCM, was found to be a chemical basis of these herbs in treating HCC. The present work also discovered that GF2 is a new STAT3 inhibitor, which is able to suppress HCC. As such, GF2 represents a new potential anti-HCC therapeutic strategy.

Discovery, Optimization, and Evaluation of Novel N-(Benzimidazol-5-yl)-1,3,4-thiadiazol-2-amine Analogues as Potent STAT3 Inhibitors for Cancer Treatment.

Signal transducer and activator of transcription 3 (STAT3) is an attractive target for cancer therapy. However, identifying potent and selective STAT3 small-molecule inhibitors with drug-like properties remains challenging. Based on a scaffold combination strategy, compounds with a novel N-(benzimidazol-5-yl)-1,3,4-thiadiazol-2-amine scaffold were designed and their inhibition of the interleukin-6 (IL-6)/JAK/STAT3 pathway was tested in HEK-Blue IL-6 reporter cells. After optimization of lead compound 12, compound 40 was identified as a selective STAT3 inhibitor that directly binds the SH2 domain to inhibit STAT3 phosphorylation, translocation, and downstream gene transcription. Compound 40 exhibited antiproliferative activities against STAT3-overactivated DU145 (IC50 value = 2.97 μM) and MDA-MB-231 (IC50 value = 3.26 μM) cancer cells and induced cell cycle arrest and apoptosis. In the DU145 xenograft model, compound 40 showed in vivo antitumor efficacy following intraperitoneal administration, with a tumor growth inhibition rate of 65.3% at 50 mg/kg, indicating promise for further development.

Platinum(IV) Complexes as Inhibitors of STAT3 and Regulators of the Tumor Microenvironment To Control Breast Cancer.

Interplay between breast cancer (BC) cells and the tumor microenvironment (TME) influences the outcome of cancer treatment. Aberrant activation of signal transducer and activator of transcription 3 (STAT3) promotes the interaction and causes immunosuppression and drug resistance. Platinum(IV) complexes SPP and DPP bearing pterostilbene-derived axial ligand(s) were synthesized to inhibit the JAK2-STAT3 pathway in BC cells and regulate the TME. These complexes exerted remarkable antiproliferative activity against the triple-negative BC cells, suppressed the expression of phosphorylated STAT3 and STAT3-related cyclooxygenase-2 and IL-6, and activated caspase-3 and cleaved poly ADP-ribose polymerase, preventing the repair of DNA lesions and inducing apoptosis. Furthermore, DPP promoted the maturation and antigen presentation of dendritic cells, repressed the proliferation and differentiation of myeloid-derived suppressor cells and regulatory T cells, and facilitated the expansion of T cells. As a consequence, DPP showed excellent anticancer activity against BC with almost no general toxicity in vivo as a potential chemoimmunotherapeutic agent.

CLCF1 signaling restrains thermogenesis and disrupts metabolic homeostasis by inhibiting mitochondrial biogenesis in brown adipocytes

Great progress has been made in identifying positive regulators that activate adipocyte thermogenesis, but negative regulatory signaling of thermogenesis remains poorly understood. Here, we found that cardiotrophin-like cytokine factor 1 (CLCF1) signaling led to loss of brown fat identity, which impaired thermogenic capacity. CLCF1 levels decreased during thermogenic stimulation but were considerably increased in obesity. Adipocyte-specific CLCF1 transgenic (CLCF1-ATG) mice showed impaired energy expenditure and severe cold intolerance. Elevated CLCF1 triggered whitening of brown adipose tissue by suppressing mitochondrial biogenesis. Mechanistically, CLCF1 bound and activated ciliary neurotrophic factor receptor (CNTFR) and augmented signal transducer and activator of transcription 3 (STAT3) signaling. STAT3 transcriptionally inhibited both peroxisome proliferator-activated receptor-γ coactivator (PGC) 1α and 1β, which thereafter restrained mitochondrial biogenesis in adipocytes. Inhibition of CNTFR or STAT3 could diminish the inhibitory effects of CLCF1 on mitochondrial biogenesis and thermogenesis. As a result, CLCF1-TG mice were predisposed to develop metabolic dysfunction even without external metabolic stress. Our findings revealed a brake signal on nonshivering thermogenesis and suggested that targeting this pathway could be used to restore brown fat activity and systemic metabolic homeostasis in obesity.

Stattic ameliorates the cecal ligation and puncture-induced cardiac injury in septic mice via IL-6-gp130-STAT3 signaling pathway

AimSepsis-induced cardiac dysfunction is the leading cause of higher morbidity and mortality with poor prognosis in septic patients. Our recent previous investigation provides evidence of the hallmarks of signal transducer and activator of transcription3 (STAT3) activation in sepsis and targeting of STAT3 with Stattic, a small-molecule inhibitor of STAT3, has beneficial effects in various septic tissues. We investigated the possible cardioprotective effects of Stattic on cardiac inflammation and dysfunction in mice with cecal ligation and puncture (CLP)-induced sepsis. Main methodsA polymicrobial sepsis model was induced by CLP in mice and Stattic (25 mg/kg) was intraperitoneally given at one and twelve hours after CLP operation. The cecum was exposed in sham-control mice without CLP. After 18 h of surgery, electrocardiogram (ECG) for anaesthized mice was registered followed by collecting of samples of blood and tissues for bimolecular and histopathological assessments. Myeloperoxidase, a marker of neutrophil infiltration, was assessed immunohistochemically. Key findingsCLP profoundly impaired cardiac functions as evidenced by ECG changes in septic mice as well as elevation of cardiac enzymes, and inflammatory markers with myocardial histopathological and immunohistochemical alterations. While, Stattic markedly reversed the CLP-induced cardiac abnormalities and restored the cardiac function by its anti-inflammatory activities. SignificanceStattic treatment had potential beneficial effects against sepsis-induced cardiac inflammation, dysfunction and damage. Its cardioprotective effects were possibly attributed to its anti-inflammatory activities by targeting STAT3 and downregulation of IL-6 and gp130. Our investigations suggest that Stattic could be a promising target for management of cardiac sepsis and inflammation-related cardiac damage.

Anthelmintic nitazoxanide protects against experimental pulmonary fibrosis.

Nitazoxanide is a therapeutic anthelmintic drug. Our previous studies found that nitazoxanide and its metabolite tizoxanide activated adenosine 5'-monophosphate-activated protein kinase (AMPK) and inhibited signal transducer and activator of transcription 3 (STAT3) signals. As AMPK activation and/or STAT3 inhibition are targets for treating pulmonary fibrosis, we hypothesized that nitazoxanide would be effective in experimental pulmonary fibrosis. The mitochondrial oxygen consumption rate of cells was measured by using the high-resolution respirometry system Oxygraph-2K. The mitochondrial membrane potential of cells was evaluated by tetramethyl rhodamine methyl ester (TMRM) staining. The target protein levels were measured by using western blotting. The mice pulmonary fibrosis model was established through intratracheal instillation of bleomycin. The examination of the lung tissues changes were carried out using haematoxylin and eosin (H&E), and Masson staining. Nitazoxanide and tizoxanide activated AMPK and inhibited STAT3 signalling in human lung fibroblast cells (MRC-5 cells). Nitazoxanide and tizoxanide inhibited transforming growth factor-β1 (TGF-β1)-induced proliferation and migration of MRC-5 cells, collagen-I and α-smooth muscle cell actin (α-SMA) expression, and collagen-I secretion from MRC-5 cells. Nitazoxanide and tizoxanide inhibited epithelial-mesenchymal transition (EMT) and inhibited TGF-β1-induced Smad2/3 activation in mouse lung epithelial cells (MLE-12 cells). Oral administration of nitazoxanide reduced the bleomycin-induced mice pulmonary fibrosis and, in the established bleomycin-induced mice, pulmonary fibrosis. Delayed nitazoxanide treatment attenuated the fibrosis progression. Nitazoxanide improves the bleomycin-induced pulmonary fibrosis in mice, suggesting a potential application of nitazoxanide for pulmonary fibrosis treatment in the clinic.

Long non-coding RNA MM2P suppresses M1-polarized macrophages-mediated excessive inflammation to prevent sodium taurocholate-induced acute pancreatitis by blocking SHP2-mediated STAT3 dephosphorylation.

M1 macrophage-mediated excessive inflammatory response plays a key role in the onset and progression of acute pancreatitis (AP), and this study aimed to investigate the role and underlying mechanisms by which the macrophage polarization-related long noncoding RNA (lncRNA) MM2P participated in the regulation of AP progression. By performing quantitative reverse-transcription PCR (qRT-PCR) assay, lncRNA MM2P was found to bedownregulated in both sodium taurocholate-induced AP model mice tissues and lipopolysaccharide (LPS)-stimulated RAW264.7 cells, and gain-of-function experiments confirmed that overexpression of lncRNA MM2P counteracted inflammatory responses, reduced macrophage infiltration and facilitated M1-to-M2 transformation of macrophages to ameliorate AP development in vitro and in vivo. Further mechanical experiments revealed that lncRNA MM2P inhibited Src homology 2 containing protein tyrosine phosphatase 2 (SHP2)-mediated signal transducer and activator of transcription 3 (STAT3) dephosphorylation to activate the STAT3 signaling, and silencing of SHP2 suppressed M1 type skewing in LPS-induced RAW264.7 cells. Interestingly, our rescuing experiments verified that lncRNA MM2P-induced suppressing effects on M1-polarization of LPS-treated RAW264.7 cells were abrogated by co-treating cells with STAT3 inhibitor stattic. Collectively, our data for the first time revealed that lncRNA MM2P suppressed M1-polarized macrophages to attenuate the progression of sodium taurocholate-induced AP, and lncRNA MM2P might be an ideal biomarker for AP diagnosisand treatment.

Protein inhibitor of activated STAT3 (PIAS3) attenuates psoriasis and associated inflammation.

Psoriasis is a common chronic inflammatory multisystem disease accompanied by hyperproliferation and inflammation of epidermal keratinocytes. Signal transducer and activator of transcription 3 (STAT3) is constitutively activated and plays an important role in epidermal keratinocytes of human psoriatic skin lesions. In this study, we investigated the effects of an endogenous STAT3 inhibitor, a protein inhibitor of activated STAT3 (PIAS3), on the proliferation and inflammation of psoriatic cells. The expression of PIAS3 in psoriatic tissues and healthy skin was analyzed using the Gene Expression Omnibus database and clinical samples. The human immortalized epidermal (HaCaT) cells were used to establish an in vitro psoriasis-like cell model. 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-thethrazolium (MTS) assay was used to detect cell proliferation. Flow cytometry was used to determine apoptosis levels. Real-time PCR, western blotting, and enzyme-linked immunosorbentassay (ELISA)were used to detect the expression levels of related factors. Furthermore, a mouse model of imiquimod (IMQ)-induced psoriatic dermatitis was established to verify the in vitro experimental results. The results showed that the mRNA and protein expression levels of PIAS3 were lower in psoriatic lesions than in normal tissues. PIAS3 inhibited the proliferation and promoted apoptosis of M5-induced HaCaT cells. Simultaneously, the mRNA and protein expression levels of tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), interleukin-8 (IL-8), and keratin 17 (K17) were significantly decreased and that of p53 was increased, thereby inhibiting the inflammatory response and promoting apoptosis. PIAS3 inhibited the transcription activity of STAT3 and noncanonical nuclear factor-kappaB (NF-κB). Furthermore, PIAS3 attenuated IMQ-induced psoriasis-like inflammation in mice. Our findings suggest that PIAS3 plays an important role in psoriasis by regulating the STAT3/NF-κB signaling pathway and p53. The lack of PIAS3 may represent a novel mechanism underlying the pathogenesis of psoriasis.

Targeting crosstalk of STAT3 between tumor-associated M2 macrophages and Tregs in colorectal cancer

ABSTRACT A comprehensive analysis of the molecular mechanism underlying colorectal tumor evaluated the development of colorectal cancer (CRC) and proposed targeting small molecular inhibitors. Nonetheless, the adoptive resistance of these therapies remains a challenge with respect to achieving an effective clinical response. Thus, identifying the molecular mechanisms guiding CRC growth is essential. The results of The Cancer Genome Atlas (TCGA) dataset analysis demonstrated a critical role of signal transducer and activator of transcription 3 (STAT3) pathway in tumor immune suppression via modulation of the recruitment of Treg cells and M2 type tumor-associated macrophages. The in vivo experiments elucidate that targeting STAT3 pathways markedly reduce the proportions of TAMs and Tregs by inhibiting tumor progression. These findings revealed crosstalk between Treg cells and M2 macrophages, proving a potential therapeutic strategy for CRC therapy. Combinatorial treatment with STAT3 inhibitor and programmed death 1 (PD-1) antibody therapy effectively prevents CRC tumor growth in a mouse model with high anti-tumor immunity. In summary, targeting STAT3 disrupts the interaction between Treg cells and M2 macrophages and improves the anti-tumor response in CRC, thereby offering a promising strategy to treat patients with CRC.