Signal Transducer And Activator Of Transcription 3 mRNA Expression Research Articles

Immunological mechanism of Ermiao Powder improving inflammation in rats with collagen-induced arthritis through α7nAChR-JAK2/STAT3 pathway

This study investigated the immunological mechanisms of Ermiao powder in the treatment of rheumatoid arthritis rats through the alpha 7 nicotinic acetylcholine receptor(α7nAChR)-Janus kinases 2(JAK2)/signal transducer and activator of transcription 3(STAT3) signaling pathway. A total of 56 female Wistar rats were randomly divided into the normal group(HG, n=8), collagen-induced arthritis(CIA) model group(CM, n=8), vagotomy group(VA, n=8), sham group(SH, n=8), Ermiao Powder treatment model group(EM, n=8), Ermiao Powder treatment for vagotomy group(EV, n=8) and Ermiao Powder treatment for sham group(ES, n=8). Following the establishment of CIA models in all groups except the HG group, the rats underwent unilateral vagotomy and sham operation(only the vagus nerve was separated). Drug treatment was started 7 days after surgery and continued for 35 days. The body weight and joints of rats were recorded, the pathological changes of the spleen of rats were observed, the contents of interleukin-6(IL-6), interleukin-1β(IL-1β) and tumor necrosis factor-α(TNF-α) in serum were detected by enzyme-linked immunosorbent assay(ELISA), and the mRNA and protein expression of α7nAChR-JAK2/STAT3 pathway core genes in spleen were detected by qRT-PCR, Western blot and immunohistochemistry. RESULTS:: showed that CM group(compared with HG group) and VA group(compared with CM group and SH group) had significantly decreased body weight(P<0.05, P<0.01), increased arthritis score(P<0.05, P<0.01), swollen ankle joints with deformity, and increased and enlarged lymph nodes in the spleen. There were also notable increases in the serum levels of IL-6, IL-1β and TNF-α(P<0.05, P<0.01), and in the mRNA expressions of JAK2 and STAT3 in the spleen(P<0.05, P<0.01). The protein levels of phosphorylated JAK2(p-JAK2)/JAK2 and phospho-STAT3(p-STAT3)/STAT3 were significantly increased(P<0.05, P<0.01), and the number of JAK2, p-JAK2, STAT3 and p-STAT3 cells increased(P<0.05, P<0.01). EM group(compared with CM group) and ES group(compared with SH group) exhibited significantly increased body weight(P<0.01), decreased arthritis scores(P<0.05, P<0.01), reduced swelling of ankle joint, and decreased number and volume of lymph nodes in the spleen. Furthermore, serum levels of IL-6, IL-1β, and TNF-α decreased(P<0.05, P<0.01), the mRNA expression of JAK2 and STAT3 in spleen decreased(P<0.05, P<0.01), the protein levels of p-JAK2/JAK2 and p-STAT3/STAT3 decreased(P<0.05, P<0.01), and the number of JAK2, p-JAK2, STAT3 and p-STAT3 cells decreased(P<0.05, P<0.01), whereas the mRNA and protein expressions of α7nAChR were significantly increased(P<0.01). Compared with the VA group, there was no significant differences in weight gain and arthritis scores in the EV group. The number of lymph nodes in the spleen was not significantly reduced and the volume was still large, suggesting the inflammation was not significantly improved. The serum levels of IL-6, IL-1β and TNF-α were not significantly different, and there were no significant differences in α7nAChR, JAK2, and STAT3 mRNA expression in the spleen. The protein expression levels of p-JAK2/JAK2 and α7nAChR in spleen were lower(P<0.05, P<0.01), while p-STAT3/STAT3 protein expression was not significantly different. Besides, the two groups had no significant difference in the number of JAK2, p-JAK2, STAT3, and p-STAT3 cells. The results suggested that unilateral vagotomy promoted the increase of phosphorylated JAK2 and STAT3 expressions and exacerbated inflammation. In contrast, Ermiao Powder alleviated the inflammation in rheumatoid arthritis rats by activating the α7nAChR-mediated JAK2/STAT3 pathway through the vagus nerve, suggesting that the α7nAchR-JAK2/STAT3 pathway may be a potential target for the treatment of rheumatoid arthritis.

PEMDA-HN, an open-label, phase II, randomized controlled study of danvatirsen plus pembrolizumab compared to pembrolizumab alone in first-line recurrent and/or metastatic head and neck squamous cell carcinoma (RM HNSCC).

TPS6113 Background: Signal transducer and activator of transcription 3 (STAT3) plays critical roles in promotion of an immune-suppressive tumor microenvironment and survival of tumor cells. Danvatirsen (DANVA) is a 16-nucleotide, generation 2.5 antisense oligonucleotide designed to down-regulate the expression of human STAT3 mRNA. Over 500 patients with hematologic malignancies or solid tumors have been exposed to DANVA monotherapy or in combination. A tolerable safety profile has been demonstrated for DANVA and toxicities are manageable. The SCORES study in recurrent and/or metastatic (RM) HNSCC patients, naïve to programmed cell death (ligand)1 (PD-(L)1) therapy, demonstrated that DANVA in combination with the PD-L1 inhibitory antibody durvalumab administered in the second line setting appeared to result in a higher objective response rate (ORR) (22.6% [12.3-36.2]) compared with the ORRs seen with durvalumab alone in prior studies (Cohen E. et al. Ann. Oncol., 2018). Several patients had complete responses (CR, 9.4%). The ORR was higher in patients with a PD-L1 tumor proportion score (TPS) ≥1 and ≥20, with increasing benefit noted with higher PD-L1 expression compared with historic anti-PD-(L)1 monotherapy data. The current study aims at evaluating the first-line combination of DANVA with pembrolizumab, an anti-PD-1 agent approved as first-line monotherapy for RM HNSCC in patients with PD-L1 positive disease. Methods: PEMDA-HN is a multicenter, open-label, randomized phase 2 study utilizing a Bayesian Optimal design (BOP2). Approximatively 81 RM HNSCC patients with a PD-L1 combined positive score (CPS) score ≥1 will be randomized in a 2:1 ratio stratified based on CPS<20 and CPS≥20 to receive either DANVA (3 mg/kg IV Days 1, 3, and 5 and then weekly starting on Day 8) and pembrolizumab (200 mg IV every 3 weeks) or pembrolizumab alone as a first line therapy for recurrent or metastatic disease. Eligible patients will receive study treatment until disease progression or discontinuation. The primary endpoint of the study is ORR by response evaluation criteria in solid tumors version 1.1 (RECIST 1.1) as assessed by the investigator. Key secondary objectives are safety, additional antitumor activity evaluation (CR rate, duration of response, disease control rate, progression-free and overall survival) and pharmacokinetics. Efficacy (e.g. ORR, CR rate) will be evaluated in all patients and in subsets of patients with CPS<20, CPS≥20 and CPS≥50. Exploratory endpoints include but are not limited to target engagement and biomarker evaluation. The study is being, or will be, conducted at US, South Korea and United-Kingdom study centers. Clinical trial information: NCT05814666 .

Ascorbic acid induces ferroptosis via STAT3/GPX4 signaling in oropharyngeal cancer

Ferroptosis is recognized as a new type of regulated cell death initiated by iron-dependent accumulation of lipid peroxidation. Recent studies have shown that the administration of ascorbic acid (AA) preferentially kills tumor cells by impairing iron metabolism and exerting pro-oxidant effects. Despite mounting evidence indicating the anticancer potential of AA, the underlying molecular mechanisms remain unknown. In this study, we demonstrated that AA decreased cell viability and Ki67 expression, along with its accumulation in the G0/G1 phase in FaDu and SCC-154 cell lines. Furthermore, AA exposure induced morphological changes in mitochondria associated with ferroptosis. AA-induced ferroptosis is accompanied by depletion of glutathione (GSH) and increased levels of ferrous ions (Fe2+), reactive oxygen species (ROS), and malondialdehyde (MDA). However, these ferroptotic effects were ameliorated by deferoxamine and N-acetylcysteine. Network pharmacology results showed that signal transducer and activator of transcription 3 (STAT3) is a key target of AA against oropharyngeal cancer. AA markedly downregulates the relative mRNA expression of STAT3 and glutathione peroxidase 4 (GPX4). Immunoblotting indicated that the protein levels of p-STAT3, STAT3, and GPX4 in FaDu and SCC-154 cells decreased significantly in response to AA treatment. Mechanistically, a chromatin immunoprecipitation assay confirmed that AA exposure reduced STAT3 expression in the GPX4 promoter region. Additionally, AA-induced inhibition of cell growth and ferroptosis was suppressed by STAT3 and GPX4 overexpression, respectively. In summary, AA inhibited oropharyngeal cancer cell growth in vitro by regulating STAT3/GPX4-mediated ferroptosis, which may provide a novel theoretical basis for the clinical treatment of oropharyngeal cancer with AA.

STAT3 expression in patients with fragility fractures and its effect on the biological function of osteoblasts.

To determine the expression of signal transducer and activator of transcription 3 (STAT3) in patients with fragility fractures (FFs) and its effect on the biological function of osteoblasts. The study included 32 patients with FFs who were diagnosed and treated in the research group and 30 concurrent healthy individuals in the control group. We observed STAT3 mRNA expression in the patients with FFs and controls and altered STAT3 mRNA to detect changes in the proliferation, invasion, and apoptosis of osteoblasts. The patients with FFs presented higher serum STAT3 mRNA expression than the controls (P < 0.05). We plotted receiver operating characteristic curves based on the STAT3 mRNA expression and found that the area under the curve for STAT3 mRNA was 0.856 (P < 0.05). Transfection of STAT3 mRNA mimics resulted in increased STAT3 mRNA expression, inhibited cell proliferation as detected by an MTT assay, and increased apoptosis rate, which was determined using flow cytometry with human fetal osteoblastic cell line 1.19 cells. STAT3 mRNA expression was elevated in the serum of patients with FFs and can be used as a biomarker for the diagnosis of the disease. Regulating STAT3 mRNA can inhibit the proliferation and induce the osteoblasts apoptosis.

IL-6 Regulates the Chemosensitivity of Drug-Resistant Multiple Myeloma Cell Lines to Bortezomib through STAT3/Notch Signaling Pathway

To investigate the effect of interleukin-6 (IL-6) on the chemosensitivity of drug-resistant multiple myeloma (MM) cell lines to bortezomib (BTZ) and its mechanism. Peripheral blood samples were collected from patients with BTZ-resistant MM before and after treatment. Human MM cell lines KM3 and KM3/BTZ were cultured in vitro. ELISA was used to detect the content of IL-6 in peripheral blood of MM patients, KM3 and KM3/BTZ cells. CCK-8 assay was used to detect the drug sensitivity of KM3 and KM3 / BTZ cells to BTZ. KM3 / BTZ cells were divided into KM3/BTZ control group (normal culture for 48 h), IL-6 neutralizing antibody Anti-IL-6 group (500 ng/ml Anti-IL-6 treated for 48 h), BTZ group (300 ng/ml BTZ treated for 48 h), BTZ + Anti-IL-6 group (300 ng/ml BTZ and 500 ng/ml Anti-IL-6 treated for 48 h). The proliferation activity of KM3 / BTZ cells was detected by CCK-8 assay. The cell cycle distribution of KM3/BTZ cells was detected by flow cytometry. The apoptosis of KM3/BTZ cells was detected by Annexin V-FITC/PI double staining. The mRNA expression levels of IL-6, Notch1, signal transducer and activator of transcription 3 (STAT3) in KM3/BTZ cells were detected by real-time fluorescent quantitative PCR (qRT-PCR), and the protein expression levels of IL-6, Notch1, STAT3 in KM3/BTZ cells were detected by Western blot. The level of IL-6 in peripheral blood of patients with BTZ-resistant MM after treatment was significantly higher than that before treatment (P<0.05). The level of IL-6 in KM3/BTZ cells was significantly higher than that in KM3 cells (P<0.05). The sensitivity of KM3/BTZ cells to BTZ was significantly lower than that of KM3 cells (P<0.05), and the resistance index (RI) was 19.62. Anti-IL-6 and BTZ could inhibit the proliferation of KM3 / BTZ cells, block cell cycle, and induce apoptosis (P<0.05). Compared with single drug treatment, the combined effect of Anti-IL-6 and BTZ was more obvious on KM3/BTZ cells (P<0.05), and significantly down regulated the mRNA and protein expression of IL-6, Notch1 and STAT3 in KM3/BTZ cells (P<0.05). Antagonizing IL-6 can increase the chemosensitivity of MM cells to BTZ, and IL-6 may reduce the sensitivity of MM cells to BTZ through STAT3/Notch signaling pathway.

Therapeutic effect of Shengjiang Powder on experimental autoimmune encephalomyelitis mice through IL-6/JAK2/STAT3 signaling pathway:based on HPLC

A high performance liquid chromatography(HPLC) method was established to analyze the components in Shengjiang Powder(SJP) such as emodin and curcumin and explore its therapeutic effect on experimental autoimmune encephalomyelitis(EAE) mice. To be specific, HPLC was performed to determine the content of compounds in SJP such as emodin and curcumin. A total of 72 female SPF C57 BL/6 mice were randomized into control group(equivalent volume of ultrapure water, ig), model group(equivalent volume of ultrapure water, ig), low-, medium-, and high-dose SJP groups(SJP, ig), and positive control group(prednisone acetate, ig), 12 each group. EAE was induced in mice except the control group. Administration began from the first day after immunization. The general conditions, symptom score, and body weight of the mice were recorded. On the 21 st day, mouse brain tissues were separrated. Then hematoxylin-eosin(HE) staining and Luxol Fast Blue(LFB) staining were used to detect the pathological changes of brain tissues. Immunohistochemistry(IHC) was employed to determine the myelin basic protein(MBP) level, and Western blot the expression of occludin and claudin-5, as well as the levels of interleukin-6(IL-6) and proteins in the Janus kinase 2(JAK2)/signal transducer and activator of transcription 3(STAT3) pathway and their phosphorylation levels. The mRNA expression of IL-6, JAK2, and STAT3 was detected by real-time quantitative polymerase chain reaction(qPCR). Finally, molecular docking of six main active components in SJP, including emodin and curcumin, with IL-6, JAK2 and STAT3 was performed, and the binding affinity was evaluated. The results showed that the established HPLC method demonstrated high precision, reproducibility, stability, and high recovery of samples. Compared with the model group, SJP reduced the clinical symptom score and alleviate the inflammatory infiltration of brain white matter and demyelination of EAE mice. At the same time, SJP increased the expression of occludin and claudin-5, down-regulated the mRNA expression of IL-6, JAK2, and STAT3, as well as the levels of IL-6/JAK/STAT3 proteins and the phosphorylation levels, with significant difference. Molecular docking suggested that the six active components in SJP had high binding energy with IL-6, JAK2, and STAT3 proteins. The established HPLC method is simple, accurate, and highly sensitive, which can simultaneously determine the content of emodin and curcumin in SJP. SJP may alleviate the clinical symptoms of EAE by inhibiting IL-6/JAK2/STAT3 signaling pathway, protecting the blood-brain barrier, and relieving the inflammatory response and demyelinization of brain tissue.

The Effect of STAT3 Signal Pathway Activation on Retinopathy of Prematurity.

Objective: To investigate the mechanism of activation of the signal transducer and activator of transcription 3 (STAT3) signal pathway in the process of retinopathy of prematurity (ROP).Methods: Sixty newborn Sprague-Dawley (SD) rats were randomly separated into the hyperoxia and air control groups (n = 30/in each group). The serum hepcidin level on 21 d was measured using the enzyme-linked immunosorbent assay (ELISA). The expression of HAMP and STAT3 protein in the liver was determined using reverse transcription-polymerase chain reaction (RT-PCR) and western blotting. Retinal neovasculature was evaluated by hematoxylin and eosin (HE) stain and fluorescein lectin. The retinal endothelial cells were treated with 250 μmol/L cobalt chloride for 72 h and added S3I-201. The STAT3 level was determined by western blotting.Results: The expression of STAT3 protein increased significantly after hyperoxia stimulation. The expression of HAMP mRNA in the hyperoxia group was significantly higher than that of the control group. The proliferation of retinal cells was inhibited, and the expression of STAT3 was increased. No significant difference was noted in vascular endothelial growth factor (VEGF) mRNA. The expression of STAT3 and VEGF mRNA was significantly reduced.Conclusion: The activation of the STAT3 signal pathway increased hepcidin expression, contributing to the pathogenesis of ROP. S3I-201 inhibited the expression of STAT3 and VEGF mRNA levels. This information provides potential novel therapeutic approach to the prevention and treatment of ROP.

Mechanism of oxymatrine in treatment of collagen-induced arthritis in mice

Sophorae Flavescentis Radix, the root of Sophora flavescens Ait., has been widely applied in the medical field due to its anti-inflammatory, analgesic, bacteriostatic, antiviral, antitumor, and other pharmacological effects. The present study investigated the anti-rheumatoid arthritis effect of oxymatrine(OMT), the active component of Sophorae Flavescentis Radix by observing its effect on the function of B lymphocytes in collagen-induced arthritis(CIA) mice through the Toll-like receptor 9(TLR9)/myeloid differentiation factor 88(MyD88)/signal transducer and activator of transcription 3(STAT3) pathway. The CIA model in DBA/1 J mice was induced by bovine type Ⅱ collagen and complete Freund's adjuvant(CFA). Fifteen days after the primary immunization, mice were treated with OMT for 30 days by intraperitoneal injection. Paw swelling and arthritis index(AI) score were evaluated every 3 days. Joint histopathologic changes were observed by HE staining. Magnetic-activated cell sorting(MACS) was used to isolate B lymphocytes from the spleen of CIA mice spleen. The serum expression level of interleukin(IL)-21 was examined by the enzyme-linked immunosorbent assay(ELISA). The expression of TLR9, STAT3, p-STAT3, and IL-21 in B lymphocytes was detected by Western blot. The mRNA expression of TLR9, STAT3, and IL-21 in B lymphocytes was detected by real-time fluorescence-based quantitative PCR(qRT-PCR). The results showed that OMT could significantly alleviate the paw swelling, decrease the AI score, relieve synovial inflammatory cell infiltration and hyperplasia, reduce the level of inflammatory cytokines, and inhibit the expression of TLR9, STAT3, p-STAT3, and IL-21 of B lymphocytes in CIA mice. Therefore, OMT may alleviate rheumatoid arthritis by regulating TLR9/MyD88/STAT3 pathway in B lymphocytes, providing a valuable reference for the application of OMT in the clinical treatment of rheumatoid arthritis.

Signal transducer and activator of transcription 3 as a potential therapeutic target for Graves’ orbitopathy

The roles of signal transducer and activator of transcription 3 (STAT3) in inflammation, oxidative stress, and adipogenesis during Graves’ orbitopathy (GO) are incompletely understood. Here, STAT3 expression in orbital tissues (from individuals with GO and healthy control subjects) was studied, and the role of STAT3 in GO pathogenesis was examined through small-interfering RNA (siRNA)-mediated silencing in primary orbital fibroblasts. STAT3 mRNA expression was higher in GO orbital tissues than in non-GO tissues. Treatment with proinflammatory cytokines, thyroid-stimulating hormone, or insulin-like growth factor-1 induced STAT3 mRNA in GO orbital fibroblasts, but not in non-GO cells. STAT3 silencing inhibited interleukin-1β-induced inflammatory cytokines and oxidative stress-induced haem oxygenase-1 expression. STAT3 siRNA-transfected GO orbital fibroblasts showed decreased adipocyte differentiation. STAT3 affected proinflammatory cytokine production, oxidative stress responses, and adipogenesis in an in vitro model of GO, suggesting that STAT3 mediates GO pathology, and that modulating STAT3 expression may have therapeutic potential against GO.

Novel mutations in the signal transducer and activator of transcription 3 gene are associated with sheep body weight and fatness traits.

The signal transducer and activator of transcription 3 (STAT3) gene plays a crucial role in leptin-mediated energy metabolism, upon which the growth and development of animals depend. Nevertheless, no studies have reported the effects of STAT3 gene polymorphisms on body weight and fatness modulation in sheep. This study aimed to illustrate STAT3 mRNA expression across tissues and various developmental stages of sheep and to highlight the association of STAT3 gene polymorphisms with body weight and fat-related traits in sheep, in order to identify a genetic marker that may conceivably be of value for marker-assisted selection (MAS). This study revealed that STAT3 was differentially expressed across age and sex (p < 0.05), with higher expression in the ram liver. The abundant expression of STAT3 in the liver of male sheep and increased expression in the hypothalamus and longissimus dorsi muscle from birth to six months of age may indicate the vital role of the STAT3 gene in animal growth and development. Moreover, SNP association analysis also revealed that the novel SNPs of the STAT3 gene detected in this study showed a significant association with body weight and fatness traits (p < 0.05). In conclusion, the significant genetic effects of the STAT3 gene polymorphisms on sheep growth and development revealed that STAT3 could be a marker gene for the selection of growth-related traits in sheep.

Fat mass and obesity-associated protein (FTO) mediates signal transducer and activator of transcription 3 (STAT3)-drived resistance of breast cancer to doxorubicin

ABSTRACT Excessive activation of signal transducer and activator of transcription 3 (STAT3) is implicated in breast cancer (BC) chemoresistance, but its underlying mechanism is not fully understood. There are STAT3 binding sites in fat mass and obesity-associated protein (FTO) promoter region, thus STAT3 may regulate the transcription of FTO. This study aimed to investigate the correlation between FTO and STAT3 in BC chemoresistance. Herein, FTO and STAT3 were highly expressed in doxorubicin-resistant BC (BC-DoxR) cells. CHIP assay verified the binding between STAT3 and FTO promoter in BC-DoxR cells. Dual luciferase reporter assay showed that FTO promoter activity was inhibited by S3I-201 (STAT3 inhibitor) but enhanced by epidermal growth factor (EGF, STAT3 activator) in BC-DoxR and BC cells. FTO mRNA and protein expression were suppressed by S3I-201 in BC-DoxR cells and EGF-stimulated BC cells. Notably, FTO regulated total N6-methyladenosine (m6A) levels in BC-DoxR and BC cells but could not affect STAT3 mRNA expression, indicating that FTO was not involved in the m6A modification of STAT3. However, FTO could activate STAT3 signaling in BC-DoxR and BC cells. Besides, FTO knockdown inhibited the doxorubicin resistance of BC-DoxR cells, while FTO overexpression enhanced the doxorubicin resistance and weakened the doxorubicin sensitivity of BC cells. Moreover, decreased doxorubicin resistance by STAT3 knockdown was abolished by FTO overexpression and decreased doxorubicin sensitivity by STAT3 overexpression was reversed by FTO knockdown, indicating that FTO was implicated in STAT3-mediated doxorubicin resistance and impairment of doxorubicin sensitivity of BC cells. Altogether, our findings provide a mechanism underlying BC doxorubicin resistance.

Effect of MiR-29b-3p Targeting STAT3 on Proliferation and Apoptosis of Acute Myeloid Leukemia Cells

To investigate the effect of miR-29b-3p on apoptosis and proliferation of acute myeloid leukemia (AML) cells by targeting signal transducer and activator of transcription 3 (STAT3). TargetScan and miRanda online databases were used to predict the binding sites of miR-29b-3p and STAT3 3'UTR. The targeting relationship between them was estimated by Dual-Luciferase reporter assay experiment. After miR-29b-3p over-expression, qPCR and Western blot were used to detect the expression of STAT3 mRNA and proteins, flow cytometry to determine the apoptosis of AML cells, and MTS to detect the changes of cell proliferation in each group. Dual-Luciferase reporter assay confirmed that STAT3 was the target gene of miR-29b-3p. After miR-29b-3p overexpression, the expression of STAT3 mRNA and protein decreased. Compared with the control groups, the proliferation of AML cells in the overexpression group decreased and the apoptosis increased (P<0.05). MiR-29b-3p can inhibit the proliferation and induce apoptosis of AML cells by down-regulating STAT3.

Curcumin Combined with Thalidomide Reduces Expression of STAT3 and Bcl-xL, Leading to Apoptosis in Acute Myeloid Leukemia Cell Lines.

IntroductionAcute myeloid leukemia (AML) is a type of blood disorder that exhibits uncontrolled growth and reduced ability to undergo apoptosis. Signal transducer and activator of transcription 3 (STAT3) is a family member of transcription factors which promotes carcinogenesis in most human cancers. This effect on AML is accomplished through deregulation of several critical genes, such as B cell lymphoma-extra-large (BCL-XL) which is anti-apoptotic protein. The aim of this study was to evaluate the effect of curcumin (CUR) and thalidomide (THAL) on apoptosis induction and also the alteration of the mRNA expression level of STAT3 and BCL-XL mRNA on AML cell line compounds.MethodsThe growth inhibitory effects of CUR and THAL and their combination were measured by MTT assay in U937 and KG-1 cell lines. The rates of apoptosis induction and cell cycle analysis were measured by concurrent staining with Annexin V and PI. The mRNA expression level of STAT3 and BCL-XL was evaluated by Real-Time PCR.ResultsCUR inhibited proliferation and induced apoptosis in both KG-1 and U937 cells and this effect increased by combination with THAL. The expression level of STAT3 and BCL-XL was significantly down-regulated in KG-1 cells after treatment by CUR and THAL and their combination.ConclusionOverall, our findings suggested that down-regulation of STAT3 and BCL-XL mRNA expression in response to CUR and THAL treatment lead to inhibition of cell growth and induction of apoptosis.

Metformin alleviates inflammatory response in non-alcoholic steatohepatitis by restraining signal transducer and activator of transcription 3-mediated autophagy inhibition in vitro and in vivo

Autophagy is an intracellular recycling and degradation process for regulating cell survival and drug resistance. Non-alcoholic steatohepatitis (NASH) is becoming a widespread disease in developing countries. However, the role of autophagy in NASH has not yet been fully elucidated. The present study determined that signal transducer and activator of transcription 3 (STAT3), in the inflammation and autophagy regulation, was the key in the progression of NASH. In NASH mouse and cell models, STAT3 mRNA and protein expressions were significantly increased, while the induction of autophagy was radically decreased. Furthermore, the effects of metformin on STAT3 expression level and NASH inflammation were investigated. The current results showed that metformin activated autophagy and decreased the mRNA expressions of inflammatory cytokines, IL-1β, IL-6, and TNF-α via inhibition of the STAT3 mRNA and protein expression. The siRNA targeting STAT3 activated autophagy and inhibited the NASH inflammatory response by reducing the mRNA expressions of the inflammatory cytokines in vivo and in vitro. The correlation between autophagy and inflammation was also explored. Autophagy induced by metformin attenuated the inflammatory response. This phenomenon of inflammation reduction was partially restored by treatment with the autophagy inhibitor 3-methylindole (3-MA). In conclusion, this study demonstrated that metformin alleviated the inflammatory response in the liver and the hepatocyte of the NASH model via STAT3-mediated autophagy induction. This mechanism provides a strategy for targeting the NASH inflammatory response.

Combined Inhibition of HDAC and EGFR Reduces Viability and Proliferation and Enhances STAT3 mRNA Expression in Glioblastoma Cells.

Changes in expression of histone deacetylases (HDACs), which epigenetically regulate chromatin structure, and mutations and amplifications of the EGFR gene, which codes for the epidermal growth factor receptor (EGFR), have been reported in glioblastoma (GBM), the most common and malignant type of brain tumor. There are likely interplays between HDACs and EGFR in promoting GBM progression, and HDAC inhibition can cooperate with EGFR blockade in reducing the growth of lung cancer cells. Here, we found that either HDAC or EGFR inhibitors dose-dependently reduced the viability of U87 and A-172 human GBM cells. In U87 cells, the combined inhibition of HDACs and EGFR was more effective than inhibiting either target alone in reducing viability and long-term proliferation. In addition, HDAC or EGFR inhibition, alone or combined, led to G0/G1 cell cycle arrest. The EGFR inhibitor alone or combined with HDAC inhibition increased mRNA expression of the signal transducer and activator of transcription 3 (STAT3), which can act either as an oncogene or a tumor suppressor in GBM. These data provide early evidence that combining HDAC and EGFR inhibition may be an effective strategy to reduce GBM growth, through a mechanism possibly involving STAT3.

MiR-506 Promotes Natural Killer Cell Cytotoxicity against Human Hepatocellular Carcinoma Cells by Targeting STAT3.

PurposeIt is well documented that natural killer (NK) cytotoxicity against hepatocellular carcinoma (HCC) cells is impaired in HCC, which might account for the failure of anti-tumor immune response. miRNAs are considered as important regulators for the development and functions of NK cells. However, the entire role of miR-506 in NK cells remains far from being addressed.Materials and MethodsThe expressions of miR-506 and signal transducer and activator of transcription 3 (STAT3) mRNA in primary NK cells from HCC patients and healthy controls were detected by quantitative real-time PCR. NK cell cytotoxicity was assessed by CFSE/7AAD cytotoxicity assay and lactate dehydrogenase assay. Luciferase reporter assay, RNA immunoprecipitation assay, and western blot were conducted to confirm the interaction between miR-506 and STAT3.ResultsmiR-506 expression was downregulated and STAT3 mRNA was upregulated in primary NK cells from HCC patients. Primary NK cells from HCC patients showed remarkably reduced cytotoxicity against SMMC7721 or HepG2 cells. NK cell cytotoxicity was positively correlated with miR-506 expression and negatively correlated with STAT3 mRNA expression. Additionally, miR-506 overexpression enhanced NK cell cytotoxicity against HCC cells, while miR-506 inhibitor showed the reverse effect. Moreover, miR-506 could suppress STAT3 expression by directly targeting 3′-untranslated regions of STAT3. A negative correlation between miR-506 and STAT3 mRNA expression in HCC patients was observed. Mechanistically, overexpressing STAT3 greatly reversed miR-506-mediated promotion of NK cell cytotoxicity against HCC cells.ConclusionmiR-506 enhanced NK cell cytotoxicity against HCC cells by targeting STAT3, suggesting that modulating miR-506 expression maybe a promising approach for enhancing NK cell-based antitumor therapies.

The changes of expressive levels of IL-17A, STAT3, and RORγt in different invasive pulmonary aspergillosis mice.

BackgroundT helper 17 (Th17) lymphocytes play an important role in Aspergillus adaptive immune response against Aspergillus fumigatus, but there is little attention focused on the different types of immunosuppressive models in which invasive pulmonary aspergillosis (IPA) develops. In addition, the expression levels of signal transducer and activator of transcription 3 (STAT3)/retinoic acid-related orphan nuclear receptor gamma (RORγt)/interleukin (IL)-17A signaling pathway, which is involved in the regulation of Th17 cells, as well as whether there are differences between two types of IPA mice models, remain unknown.Materials and methodsSix to eight weeks old female BALB/c mice were treated with cortisone acetate or cyclophosphamide to establish the immunosuppressive mice models, and then, A. fumigatus inoculum was injected to form the IPA groups and sterile saline was injected to form the control groups. Flow cytometry was performed to measure the proportion of Th17 cells in CD4+ T cells in the peripheral blood, spleen, and lung of the mice. The expression of IL-17A, RORγt, and STAT3 mRNA was detected by real-time polymerase chain reaction. Concentrations of IL-6 in the plasma and bronchoalveolar lavage fluid were measured by enzyme-linked immunosorbent assay.ResultsThe proportion of Th17 in the peripheral blood and lung tissue in neutropenic IPA mice showed a more significant increase than in non-neutropenic IPA mice (P<0.01). The IL-6 protein also showed the same trend in plasma and bronchoalveolar lavage fluid (P<0.01). Compared with the control groups, the expression of IL-17A at mRNA level in the lung was significantly increased, while RORγt/STAT3 mRNA was significantly decreased in the IPA groups (P<0.01).ConclusionThe expression of RORγt and STAT3 mRNA in the lung tissue in both groups was significantly decreased. IL-17 may play a negative role in the defense against Aspergillus through uprating IL-6.

Ischemia-reperfusion induces death receptor-independent necroptosis via calpain-STAT3 activation in a lung transplant setting.

Ischemia-reperfusion (I/R)-induced lung injury undermines lung transplantation (LTx) outcomes by predisposing lung grafts to primary graft dysfunction (PGD). Necrosis is a feature of I/R lung injury. However, regulated necrosis (RN) with specific signaling pathways has not been explored in an LTx setting. In this study, we investigated the role of RN in I/R-induced lung injury. To study I/R-induced cell death, we simulated an LTx procedure using our cell culture model with human lung epithelial (BEAS-2B) cells. After 18 h of cold ischemic time (CIT) followed by reperfusion, caspase-independent cell death, mitochondrial reactive oxygen species production, and mitochondrial membrane permeability were significantly increased. N-acetyl-Leu-Leu-norleucinal (ALLN) (calpain inhibitor) or necrostatin-1 (Nec-1) [receptor interacting serine/threonine kinase 1 (RIPK1) inhibitor] reduced these changes. ALLN altered RIPK1/RIPK3 expression and mixed lineage kinase domain-like (MLKL) phosphorylation, whereas Nec-1 did not change calpain/calpastatin expression. Furthermore, signal transducer and activator of transcription 3 (STAT3) was demonstrated to be downstream of calpain and regulate RIPK3 expression and MLKL phosphorylation during I/R. This calpain-STAT3-RIPK axis induces endoplasmic reticulum stress and mitochondrial calcium dysregulation. LTx patients' samples demonstrate that RIPK1, MLKL, and STAT3 mRNA expression increased from CIT to reperfusion. Moreover, the expressions of the key proteins are higher in PGD samples than in non-PGD samples. Cell death associated with prolonged lung preservation is mediated by the calpain-STAT3-RIPK axis. Inhibition of RIPK and/or calpain pathways could be an effective therapy in LTx.

Aberrant phosphorylation of signal transducer and activator of transcription 3 protein of the CD4+ T cells in patients with primary immune thrombocytopenia.

: Primary immune thrombocytopenia (ITP) is an autoimmune disorder characterized by decreased platelet count of which dysfunctional cellular immunity in its pathogenesis. Signal transducer and activator of transcription 3 (STAT3) is critical for the differentiation of T cells. The present study was aimed to investigate the STAT3 protein phosphorylation of CD4 T cells in ITP patients. Fourteen patients of newly diagnosed ITP with complete remission (R group) and other 15 newly diagnosed ITP patients with nonresponse (N group) after corticosteroids therapy were included. Sixteen healthy human volunteers were served as normal controls (C group). Blood samples were collected before and after the therapy. Serum levels of IL-6 were measured by ELISA. The phosphorylation of STAT3 protein (pSTAT3) and the percentage of Th17 cells of the CD4 T cells were analyzed by flow cytometry. The STAT3 mRNA expression was examined by Real-time PCR. The level of IL-6 in the R group was higher than that in the C group (P = 0.02) whereas no difference was found between groups of N and C. The basal level of pSTAT3 in the R group was significantly higher when compared with N group (P = 0.002). The percentage of Th17 cells of the CD4 T cells in the ITP patients was numerically higher than that in the controls (P = 0.03). Our results indicate that ITP patient with higher basal level of pSTAT3 might have more favorite response to the corticosteroid therapy, which warrants further investigation on the prognostic role of pSTAT3 in ITP.

IL-10/STAT3 is reduced in childhood obesity with hypertriglyceridemia and is related to triglyceride level in diet-induced obese rats

BackgroundThe prevalence of childhood obesity and obesity-related metabolic disorder such as dyslipidemia has sharply increased in the past few decades. Chronic low-grade inflammation is associated with the development of comorbidities and poor prognosis in obesity. This study aims to evaluate interleukin-10 (IL-10) in childhood obesity with hypertriglyceridemia.MethodWe evaluated IL-10 and signal transducer and activator of transcription 3 (STAT3) mRNA expression in adipose tissue (AT) as well as serum IL-10 in 62 children of 3 groups and in high-fat diet (HFD) induced obese rat. Expression of IL-10 and STAT3 protein in AT of diet-induced obese rats were examined over feed period.ResultsAdipose IL-10 and STAT3 mRNA expression and serum IL-10 reduced in obese children with hypertriglyceridemia and in HFD obese rats. The protein expression of IL-10 and STAT3 decreased in AT of obese rats compared with the control rats at end time. Expression of IL-10 mRNA was negatively correlated to TG and LDL-C levels, and positively correlated to HDL-C, adiponectin and serum IL-10 levels.ConclusionsIL-10 expression and its downstream JAK-STAT pathway are down-regulated in obese children with hypertriglyceridemia and in HFD obese rats.