STAT3 Research Articles

Anti-proliferative effect of Shenqi Fuzheng injection on gastric cancer cell line SGC7901 via STAT3 pathway

Objective To study the effect of Shenqi Fuzheng injection on proliferation of gastric cancer cell line SGC7901, and to explore its mechanism associated with the STAT3 pathway. Methods The human gastric cancer cell line SGC7901 was used as the research object, the effects of different concentrations of Shenqi Fuzheng injection on the activity of SGC7901 cells were observed by MTT method, the levels of interleukin (IL)-6, IL-8 and IL-11 were detected by enzyme-linked immunosorbent assay (ELISA), and the expressions of Janus kinase 2 (JAK2), STAT3 and phosphorylated-STAT3 (p-STAT3) were detected by Western blot. Results Compared with the control group, the activity of SGC7901 cells in the treatment group decreased significantly, and the concentration of Shenqi Fuzheng injection reached 0.1 mg/ml, the inhibitory rate of SGC7901 cells reached (10.8±0.7)%, and the difference was statistically significant (P < 0.01). With the increase of Shenqi Fuzheng injection concentration, the inhibition rate of SGC7901 cells increased gradually (F = 12.319, P = 0.000). Compared with the control group, with the increase of Shenqi Fuzheng injection concentration, IL-6, IL-8 and IL-11 contents were decreased in SGC7901 cells, the differences were statistically significant (IL-6: F = 31.256, P = 0.000; IL-8: F = 16.857, P = 0.000; IL-11: F = 21.319, P = 0.000); JAK2 and p-STAT3 protein levels were significantly reduced (JAK2: F = 12.315, P = 0.000; p-STAT3: F = 16.728, P = 0.000) in the treatment groups. Conclusion Shenqi Fuzheng injection can inhibit the proliferation of SGC7901 cells, and the mechanism may be related to the inhibition of STAT3 pathway. Key words: Stomach neoplasms; STAT3 transcription factor; Signal transduction; Shenqi Fuzheng injection

Effect of continuous transversus abdominis plane block on postoperative systemic inflammatory responses of patients undergoing laparoscopic radical surgery for colorectal cancer

Objective To evaluate the effect of continuous transversus abdominis plane (TAP) block on postoperative systemic inflammatory responses of patients undergoing laparoscopic radical surgery for colorectal cancer. Methods Ninety-eight patients, aged 45-64 yr, of American Society of Anesthesiologists physical status Ⅱ or Ⅲ, weighing 50-75 kg, scheduled for elective laparoscopic radical surgery for colorectal cancer under general anesthesia, were divided into continuous patient-controlled epidural analgesia (PCEA) group (group PCEA) and continuous TAP block group (group TAPB) using a random number table, with 49 patients in each group.An epidural catheter was placed at L1, 2 interspace, a test dose of 3 ml of 2% lidocaine was given, and morphine 2 mg (in 10 ml of normal saline) was injected into the epidural space at the end of surgery in group PCEA.PCEA solution contained 0.15% ropivacaine and sufentanil 2 μg/ml (diluted to 300 ml in normal saline). PCA pump was set up with a 2 ml bolus dose, a 20 min lockout interval and background infusion at a rate of 4 ml/h.Catheterization of bilateral transversus abdominis plane was accomplished under ultrasound guidance, and 0.3% ropivacaine 20 ml was injected in group TAPB.TAP block solution contained 0.15% ropivacaine (diluted to 300 ml in normal saline). PCA pump was set up with a 6 ml bolus dose, a 30 min lockout interval and background infusion at a rate of 6 ml/h.Analgesia lasted until 48 h after surgery, and visual analogue scale score was maintained less than or equal to 3 in both groups.Parecoxib sodium 40 mg was intravenously injected as a rescue analgesic when visual analogue scale score was more than 3.Blood samples were collected from the right internal jugular vein at 24 h before operation (T0) and 48 h after operation (T1) for determination of the expression of CXCL8 mRNA in serum (by real-time polymerase chain reaction) and expression of CXCL8 and STAT3 in serum (by Western blot). The development of requirement for rescue analgesia, sensory motor dysfunction of lower extremities and nausea and vomiting after surgery was recorded. Results Compared with the baseline at T0, the expression of serum CXCL8 mRNA, CXCL8 and STAT3 was significantly up-regulated at T1 in the two groups (P 0.05). Conclusion Continuous TAP block can reduce postoperative systemic inflammatory responses of patients undergoing laparoscopic radical surgery for colorectal cancer. Key words: Nerve conduction block; Abdomen; Colon cancer; Rectal cancer; Inflammation

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

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

STAT6 Promoter Polymorphism is Essential for Malaria Infection and Suppression of Parasitemia Among Infected Children

Significant morbidity and mortality among children remain the most consequential outcome of malaria infection in sub‐Saharan Africa, with host genetic variants reported to modulate infectivity and regulation of disease markers. Effector T cells are essential for a proper immune response, such responses ameliorated by regulatory T cells, which are encoded by genes implicated in cytokine cross regulation. One such gene is the signal transducer and activator of transcription 6 (STAT6), whose intronic (rs3024974) single nucleotide polymorphism (SNP), is suspected in malaria pathogenesis. To this end, regulatory SNP of the STAT6 gene was genotyped in a cohort of P. falciparum malaria infected (n=106) and uninfected, control children (n=225), ages 1–5 years, recruited from Nigeria. The genotypic distribution of rs3024974 was in Hardy‐Weinberg equilibrium, among all groups. Even though the wild type genotype (rs3024974CC) was commoner in both groups; 83.0% (infected) versus 70.7% (uninfected); the percentile frequency of rs3024974TT genotype of STAT6 was significantly higher among uninfected children compared to infected (4.9% versus 1.0% respectively). In addition, the minor allele rs3024974T was doubly prevalent among uninfected controls than infected children (17.1% versus 8.9% respectively; p=0.01). Significantly, children with the mutant genotype (rs3024974TT) were less likely to develop clinical malaria (OR 0.19; 95% CI=0.00–1.31), with the odds increased tenfold among children with wild‐type genotype (OR=2.03; 95% CI=1.10–3.86). This observation was further confirmed when examining markers of disease among infected children, with parasitemia significantly less (tenfold reduction) in those with the mutant genotype (mean parasite count: 2,986 parasites/ml of blood) versus children with wild‐type genotype (mean parasite count: 30,242 parasites/ml of blood). Despite the fact there were differences in other markers (age, weight, packed cell volume, temperature) based on rs3024974 variants, these differences were not significant. We conclude that STAT‐6 promoter polymorphism rs3024974 is a susceptibility factor for malaria infection and significant downregulation of P. falciparum parasitemia levels among infected children.Support or Funding InformationLaboratory Support and Faculty Development Award, College of Health Sciences and Technology, Rochester Institute of TechnologyThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Interleukin-13 Stimulation Reveals the Cellular and Functional Plasticity of the Airway Epithelium.

About 50% of patients with asthma exhibit chronic airway inflammation driven by the type 2 cytokines interleukin (IL)-4, IL-5, and IL-13. These patients with type 2-high asthma experience more allergic symptoms, greater airway hyperresponsiveness, and more severe mucus obstruction than patients with type 2-low asthma. Mouse models of asthma have shown that much of the airway dysfunction in these models can be generated by IL-13 stimulation of the airway epithelium alone. Both in vivo mouse model studies and in vitro studies of human mucociliary airway epithelial cultures have shown that IL-13 induces cellular remodeling of the airway epithelium through proliferation-independent transdifferentiation processes. In both humans and mice, IL-13 stimulation of the airway epithelium results in generation of hypersecretory mucin 5AC (MUC5AC)-expressing mucus cells. Whereas club cells have been shown to be the source of these mucin 5AC-positive mucus cells in mice, the origin of these mucus cells in humans is unclear. In humans, chronic IL-13 stimulation appears to result in loss of ciliated cells. Moreover, IL-13 stimulation can block ciliated cell differentiation from human basal airway epithelial cells. Coincident with IL-13 cellular remodeling are reported decreases in mucociliary transport and ciliary beat frequency. These IL-13-mediated changes in mucociliary function are accompanied by disorganization of cilia, a decrease in the ratio of mucin 5B (MUC5B) to mucin 5AC, and mucus gel tethering to the epithelial surface by mucin 5AC. These airway epithelial responses to IL-13 are mediated by multiple transcription factors, including signal transducer and activator of transcription-6 (STAT6), SAM pointed domain-containing Ets transcription factor (SPDEF), Forkhead box A2 (FOXA2), and Forkhead box J1 (FOXJ1). In addition, analysis of RNA-sequencing data derived from airway epithelial cells shows how IL-13 stimulation promotes broad changes in gene expression across the transcriptome. These results reveal the plastic nature of airway epithelial cells that enables the epithelium to undergo remodeling and functional shifts in response to IL-13 stimulation. With use of new technology, future studies should lead to greater understanding of how IL-13 and other stimuli of disease bring about airway epithelial remodeling, which may aid in the development of therapies that ameliorate airway dysfunction in asthma and other diseases.

Apoptotic cells trigger the ABCA1/STAT6 pathway leading to PPAR-γ expression and activation in macrophages.

The signal transducer and activator of transcription 6 (STAT6) transcription factor activates peroxisome proliferator-activated receptor gamma (PPAR-γ)-regulated gene expression in immune cells. We investigated proximal membrane signaling that was initiated in macrophages after exposure to apoptotic cells that led to enhanced PPAR-γ expression and activity, using specific siRNAs for ABCA1, STAT6, and PPAR-γ, or their antagonists. The interactions between mouse bone marrow-derived macrophages or RAW 264.7 cells and apoptotic Jurkat cells, but not viable cells, resulted in the induction of STAT6 phosphorylation as well as PPAR-γ expression and activation. Knockdown of ATP-binding cassette transporter A1 (ABCA1) after the transfection of macrophages with ABCA1-specific siRNAs reduced apoptotic cell-induced STAT6 phosphorylation as well as PPAR-γ mRNA and protein expression. ABCA1 knockdown also reduced apoptotic cell-induced liver X receptor α (LXR-α) mRNA and protein expression. Moreover, inhibition of STAT6 with specific siRNAs or the pharmacological inhibitor AS1517499AS reversed the induction of PPAR-γ, LXR-α, and ABCA1 by apoptotic Jurkat cells. PPAR-γ-specific siRNAs or the PPAR-γ antagonist GW9662 inhibited apoptotic cell-induced increases in LXR-α and ABCA1 mRNA and protein levels. Thus, these results indicate that apoptotic cells trigger the ABCA1/STAT6 pathway, leading to the activation of the PPAR-γ/LXR-α/ABCA1 pathway in macrophages.

Significance of abnormal expression of miR-21/stat3 signaling pathway in schizophrenia rats

目的 观察miR－21/stat3信号通路调控异常在精神分裂症大鼠发生中的意义。 方法 选用Wistar大鼠48只，随机数字表法分为对照组24只，模型组24只。模型组建立精神分裂症动物模型，通过Morris水迷宫实验，确定精神分裂症模型成功后，处死大鼠。采用实时荧光定量PCR(qPCR)方法观察模型组大鼠海马区脑组织中miR－21及stat3的表达情况，并与对照组比较。全部实验数据均采用SPSS 17.0统计软件包进行统计学处理。 结果 Morris水迷宫实验发现模型组大鼠找到平台的时间较对照组延迟。对照组和模型组大鼠海马区脑组织中miR－21的表达量分别为1.027和0.682，2组miR－21表达量比较差异有统计学意义(t＝7.72，P＝0.032 6)。对照组和模型组大鼠海马区脑组织中stat3的表达量分别为0.314和1.518，2组stat3表达量比较差异有统计学意义(t＝3.12，P＝0.006 1)。 结论 脑组织中海马区miR－21/stat3信号通路表达异常影响大鼠海马区神经活动，从而导致精神分裂症的发生。

Prostate cancer: Radiosensitized by STAT5 blockade.

Prostate cancer: Radiosensitized by STAT5 blockade.

Rapid environmental effects on gut nematode susceptibility in rewilded mice.

Genetic and environmental factors shape host susceptibility to infection, but how and how rapidly environmental variation might alter the susceptibility of mammalian genotypes remains unknown. Here, we investigate the impacts of seminatural environments upon the nematode susceptibility profiles of inbred C57BL/6 mice. We hypothesized that natural exposure to microbes might directly (e.g., via trophic interactions) or indirectly (e.g., via microbe-induced immune responses) alter the hatching, growth, and survival of nematodes in mice housed outdoors. We found that while C57BL/6 mice are resistant to high doses of nematode (Trichuris muris) eggs under clean laboratory conditions, exposure to outdoor environments significantly increased their susceptibility to infection, as evidenced by increased worm burdens and worm biomass. Indeed, mice kept outdoors harbored as many worms as signal transducer and activator of transcription 6 (STAT6) knockout mice, which are genetically deficient in the type 2 immune response essential for clearing nematodes. Using 16S ribosomal RNA sequencing of fecal samples, we discovered enhanced microbial diversity and specific bacterial taxa predictive of nematode burden in outdoor mice. We also observed decreased type 2 and increased type 1 immune responses in lamina propria and mesenteric lymph node (MLN) cells from infected mice residing outdoors. Importantly, in our experimental design, different groups of mice received nematode eggs either before or after moving outdoors. This contrasting timing of rewilding revealed that enhanced hatching of worms was not sufficient to explain the increased worm burdens; instead, microbial enhancement and type 1 immune facilitation of worm growth and survival, as hypothesized, were also necessary to explain our results. These findings demonstrate that environment can rapidly and significantly shape gut microbial communities and mucosal responses to nematode infections, leading to variation in parasite expulsion rates among genetically similar hosts.

Ononitol monohydrate enhances PRDM16 & UCP-1 expression, mitochondrial biogenesis and insulin sensitivity via STAT6 and LTB4R in maturing adipocytes

Ononitol monohydrate (OMH), a glycoside was originally isolated from Cassia tora (Linn.). Glycosides regulate lipid metabolism but scientific validation desired. Hence, we aimed to evaluate the effect of OMH on enhancing mitochondrial potential, mitochondrial biogenesis, upregulate the expression of brown adipogenesis specific genes in maturing adipocytes. In addition, we observed the inter-relation between adipocyte and T-lymphocyte; whether, OMH treated adipocyte-condition medium stimulate T-cell chemokine linked with insulin resistance. In a dose dependent manner OMH treated to preadipocyte significantly inhibited maturation and enhanced mitochondrial biogenesis, it was confirmed by Oil red ‘O and Nile red stain without inducing cytotoxicity. The mRNA levels of adipocyte browning related genes such as, PR domain containing 16 (PRDM16), peroxisome proliferator activated receptor gamma coactivator 1 alpha (PPARγC1α) and uncoupling protein-1 (UCP-1) have been significantly upregulated. In addition, adipogenic transcription factors [such as proliferator activated receptor γ (PPARγ), CCAAT/enhancer binding protein (C/EBPα) and sterol regulatory element binding protein-1c (SREBP-1c)] and adipogenic genes were significantly down-regulated by treatment with OMH when compared to control cells. Protein expression levels of adiponectin have been increased; leptin, C/EBPα and leukotriene B4 receptor (LTB4R) were down regulated by OMH in mature adipocytes. In addition, adipocyte condition medium and OMH treated T-lymphocyte, significantly increased insulin signaling pathway related mRNAs, such as interlukin-4 (IL-4), signal transducer and activator of transcription 6 (STAT6) and decreased leukotriene B4 (LTB4). The present findings suggest that OMH increased browning factors in differentiating and maturing preadipocyte also decreased adipose tissue inflammation as well as the enhanced insulin signaling.

Genetic defect types associated with interstitial lung disease

Interstitial lung disease (ILD), also known as the diffuse parenchymal lung diseases(DPLD), are a heterogeneous group of the chronic respiratory disorders with various causes.The etiology of the ILD in infants includes the surfactant dysfunction and immunodeficiency, which are mainly caused by single gene defects or mutations.The common surfactant dysfunction is caused by the mutation of surfactant protein B gene (SFTPB), surfactant protein C gene (SFTPC), adenosine triphosphate binding cassette transport A3 (ABCA3)gene and thyroid transcription factor 1 gene, can presented as the deadly neonatal respiratory distress syndrome(RDS), and the ILD in the children.Gene mutation of the granulocyte-macrophage colony stimulating factor receptor can lead to hereditary pulmonary alveolar proteinosis(PAP), and mucins 5B gene, telomerase reverse transcriptase gene, telomerase RNA component gene were associated with adult pulmonary fibrosis.In recent years, it has been found that the mutation of some immune genes, such as STING, GATA2 and STAT5, which can lead to interstitial lung disease of children.With the development of gene techno-logy, the more etiology of gene in the interstitial lung disease are diagnosed by the new technology. Key words: Lung disease, interstitial; Gene; Surfactant

MicroRNA-31 and MicroRNA-155 Are Overexpressed in Ulcerative Colitis and Regulate IL-13 Signaling by Targeting Interleukin 13 Receptor α-1.

Interleukin-13 (IL-13) is an important Type 2 T helper (Th2) cytokine, controlling biological functions in epithelium and has been linked to asthma, atopic dermatitis and ulcerative colitis (UC). Interleukin-13 signals through IL-13 receptor α-1 (IL13RA1 (gene) and IL13Rα1 (protein)), a receptor that can be regulated by microRNAs (miRs). MicroRNAs are small non-coding single-stranded RNAs with a role in several pathologies. However, their relevance in the pathophysiology of UC, a chronic inflammatory condition of the colonic mucosa, is poorly characterised. Here, we determined the expression of IL13Rα1 in UC, its potential regulation by miRs and the subsequent effect on IL-13 signalling. Inflamed mucosa of UC patients showed decreased mRNA and protein expression of IL13RA1 when compared to healthy controls. We show that miR-31 and miR-155 are upregulated in inflamed UC mucosa and that both directly target the 3′ untranslated region of IL13RA1 mRNA. Transfection of miR-31 and miR-155 mimics reduced the expression of IL13RA1 mRNA and protein, and blocked IL-13-dependent phosphorylation of signal transducer and activator of transcription 6 (STAT6) in HT-29 cells, a gut epithelium cell line. Interleukin-13 activation of suppressor of cytokine signaling 1 (SOCS1) and eotaxin-3 (CCL26) expression was also diminished. MicroRNA-31/microRNA-155 mimics also downregulated IL13RA1 in ex vivo human inflamed UC biopsies. We propose that miR-31 and miR-155 have an important role in limiting IL-13 signalling in UC disease.

Research of the inhibitory effect and the mechanisms of plumbagin on large cell lung cancer NL9980 cell line

Objective To investigate the effects of plumbagin on human NL9980 large-cell lung cancer cells and the role of interleukin-6/signal transducers and activators of transcription 3 (IL-6/STAT3) signaling pathway in the anticancer procedure of plumbagin. Methods Different concentrations of plumbagin were administrated in NL9980 cell line. Inhibitory effects of plumbagin on NL9980 cells were analyzed using methyl thiazol tetrazolium (MTT) assay. Groups of different drug doses (5.0, 7.5, 10.0 μmol/L) and blank control (without administration of plumbagin) were set with simple random sampling method. Cells were collected after 24 h of administration. Expression of IL-6 and STAT3 and the downstream genes B-cell lymphoma/leukemia-2 (bcl-2), vascular endothelial growth factor (VEGF), and Cyclin D1 were assessed by real-time quantitative polymerase chain reaction (Real-time PCR). Enzyme linked immunosorbent assay (ELISA) was used to measure secreted IL-6 levels and western blotting to measure intracellular phospho-STAT3 levels. Results Plumbagin significantly inhibited NL9980 cells via suppressing IL-6/STAT3 signaling. Expression of all genes was inhibited by plumbagin in a dose-dependent manner (F=32.170, P=0.000). As the concentration of plumbagin increased, the expression of IL-6 mRNA decreased significantly (F=173.067, P=0.000) to 0.50±0.02, 0.44±0.1, 0.34±0.04 respectively and IL-6 protein decreased to 4.71±0.02, 4.08±0.14, 2.01±0.07 significantlly (F=1 805.177, P=0.000). Besides, the expression of STAT3 mRNA (0.86±0.08, 0.72±0.03, 0.52±0.06, F=230.918, P=0.000) and protein (0.65±0.19, 0.45±0.22, 0.31±0.11, F=17.332, P=0.000) were reduced with the increase of concentration of plumbagin. Compared with the control group, expression of downstream genes bcl-2, VEGF and Cyclin D1 mRNA were decreased significantly after administration of plumbagin (F=33.698, P=0.000; F=337.743, P=0.000; F=180.136, P=0.000), and declined gradually as drug concentration increased. Moreover, bcl-2, VEGF, and Cyclin D1 mRNA levels positively correlated with levels of IL-6 and STAT3 gene expressions. Conclusion IL-6-mediated STAT3 suppression decreased bcl-2, VEGF, and Cyclin D1 expression. Thus, plumbagin may be an effective therapy for large-cell lung cancer that acts by targeting the IL-6/STAT3 signaling pathway. Key words: Plumbagin; Interleukin-6; Signal transducers and activators of transcription 3; Large cell lung cancer

Phenotyping acute and chronic atopic dermatitis-like lesions in Stat6VT mice identifies a role for IL-33 in disease pathogenesis.

The Stat6VT mouse model of atopic dermatitis (AD) is induced by T-cell-specific expression of a constitutively active form of the protein signal transducer and activator of transcription 6 (STAT6). Although AD-like lesions are known to develop in Stat6VT mice, this study was designed to determine if these mice develop acute and chronic phases of disease similar to humans. To address this, AD-like lesions from Stat6VT mice were harvested at two different timepoints relative to their onset. Lesions harvested within 1week after development were defined as acute lesions, and those present for 1month or more were defined as chronic lesions. Acute and chronic AD-like lesions from Stat6VT mice exhibited histologic findings and cytokine expression patterns similar to acute and chronic AD lesions in humans. Further analysis revealed increased levels of interleukin (IL)-33 transcripts in AD-like lesions compared to Stat6VT nonlesional and wild-type skin controls. Immunofluorescence also revealed increased numbers of IL-33+ keratinocytes in Stat6VT lesional skin and localized IL-33+ keratinocytes to a keratin 5+ subset. Furthermore, AD-like disease was more severe in IL-33-deficient Stat6VT mice compared to IL-33-sufficient Stat6VT mice. These studies suggest that Stat6VT mice can serve as a model of acute and chronic AD and that IL-33 may attenuate inflammation in this system.

Effects of knockdown of Stat5 on invasion and epithelial mesenchymal transition of thyroid carcinoma TT cells

Objective To investigate the effects of knockdown of signal transducer and activator of transcription 5 (Stat5) on invasion and epithelial mesenchymal transition (EMT) of thyroid carcinoma TT cells. Methods Thyroid carcinoma TT cells were cultured and divided into Stat5-small interfering RNA (siRNA) group and NC-siRNA group, transfected with Stat5 siRNA and negative control NC-siRNA respectively. After transfection, the numbers of invasive cells were determined by Transwell model and the expressions of matrix metalloproteinase (MMP) family genes, EMT marker genes, angiogenesis genes in the cells were determined by fluorescent quantitative PCR. Results The numbers of invasive cells in Stat5-siRNA group were significantly lower than those in NC-siRNA group (42.39±5.82 vs. 64.41±8.49, t=-4.784, P=0.001; 51.23±6.38 vs. 78.54±9.52, t=-5.329, P<0.001; 60.35±8.35 vs. 98.42±11.25, t=-6.076, P<0.001) after the transfection of siRNA for 12 hours, 18 hours and 24 hours. Twenty-four hours after transfection of siRNA, the mRNA expressions of MMP1, MMP2, MMP9, MMP13, N-cadherin, Vimentin, vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), angiogenin (Ang)-1 and Ang-2 in Stat5-siRNA group were significantly lower than those in NC-siRNA group (0.42±0.07 vs. 1.03±0.15, t=-8.240, P<0.001; 0.35±0.06 vs. 1.01±0.13, t=-10.307, P<0.001; 0.29±0.05 vs. 1.05±0.18, t=-9.097, P<0.001; 0.54±0.08 vs. 0.98±0.12, t=-6.822, P<0.001; 0.38±0.06 vs. 1.04±0.18, t=-7.778, P<0.001; 0.29±0.04 vs. 1.06±0.12, t=-12.612, P<0.001; 0.36±0.07 vs. 1.06±0.14, t=-10.000, P<0.001; 0.43±0.08 vs. 0.97±0.12, t=-8.372, P<0.001; 0.25±0.03 vs. 1.03±0.12, t=-14.100, P<0.001; 0.19±0.03 vs. 0.99±0.13, t=-13.408, P<0.001). The mRNA expression of E-cadherin was significantly higher than that in NC-siRNA group (2.88±0.42 vs. 0.98±0.12, t=9.726, P<0.001). Conclusion Knockdown of Stat5 inhibits the invasion and EMT of thyroid carcinoma TT cells. Key words: Thyroid neoplasms; Neoplasm invasiveness; Signal transducer and activator of transcription 5; Epithelial mesenchymal transition

Genetic polymorphisms in HLA-DP and STAT4 are associated with IgA nephropathy in a Southwest Chinese population.

IgA nephropathy (IgAN) is the most common chronic glomerular disease worldwide. Genetic factors are thought to be crucial in the pathogenesis of IgAN. However, few data are available on the relationship between human leucocyte antigen (HLA) and signal transducer and activator of transcription 4 (STAT4) polymorphisms and IgAN susceptibility in the Chinese population. Therefore, we examined HLA-DP/DQ and STAT4 polymorphisms (rs3077, rs9277535, rs7453920 and rs7574865) in a total of 630 subjects including 140 IgAN and 490 healthy controls in Chinese. There were significant associations between IgAN patients and healthy controls in the allele frequency of rs3077, rs9277535 and rs7574865. In addition, the genotypes of rs3077, rs9277535 and rs7574865 were also significantly associated with IgAN under recessive models. Moreover, the haplotypes block AAG, AGG, GAG and GGA in the HLA gene significantly correlated with the risk of IgAN. This is the first study demonstrating the significant associations of SNP rs3077, rs9277535 and rs7574865 and the haplotypes in the HLA gene with the risk of IgAN in a Southwest Chinese population. This research provides a new insight into the significant relationship between HLA-DP and STAT4 polymorphisms and the susceptibility to IgAN.

A STAT6 Inhibitor AS1517499 Reduces Preventive Effects of Apoptotic Cell Instillation on Bleomycin-Induced Lung Fibrosis by Suppressing PPARγ

Background/Aims: The signal transducer and activator of transcription 6 (STAT6) transcription factor mediates PPARγ-regulated gene expression in macrophages. However, it remains largely unknown how proximal membrane signaling events initiated by apoptotic cell recognition upregulate PPARγ expression and activate the lung homeostatic program. Methods: The STAT6 inhibitor AS1517499 was used to determine the role of STAT6 in mediating PPARγ activity, anti-inflammatory effects, and anti-fibrotic effects induced by apoptotic cell instillation after bleomycin treatment into C57BL/6 mice. Bronchoalveolar lavage fluid, alveolar macrophages and lungs were harvested at days 2, 7, and 14 and then analyzed by real-time PCR, immunoblotting, ELISA, immunocytochemistry and immunohistochemistry assays. Results: Our data demonstrate that apoptotic cell instillation after bleomycin results in prolonged enhancement of STAT6 phosphorylation in alveolar macrophages and lung. Co-administration of the STAT6 inhibitor, AS1517499, reversed the enhanced PPARγ expression and activity induced by apoptotic cell instillation after bleomycin treatment. By reducing the expression of PPARγ target genes, including CD36, macrophage mannose receptor, and arginase 1, AS1517499 inhibited efferocytosis and restored pro-inflammatory cytokine expression, neutrophil recruitment, protein levels, hydroxyproline content, and expression of fibrosis markers, including type 1 collagen α2, fibronectin, and α-smooth muscle actin. STAT6 inhibition reversed the expression profile of hepatocyte growth factor and interleukin-10. Conclusion: These results indicate that prolonged STAT6 activation following one-time apoptotic cell instillation facilitates continuous PPARγ activation, resulting in the resolution of bleomycin-induced lung inflammation and fibrosis.

Association of STAT6 rs324011 Gene Polymorphism with Susceptibility of Atopic Bronchial Asthma in Egyptian Children

Background: Bronchial asthma is a common disease with multiple determinants that include genetic variation. Signal transducer and activator of transcription 6 (STAT6) is a central molecule in the signal transduction pathway used by interleukin-4 (IL-4) in immunoglobulin E (IgE) isotype switching. Study Objective was to investigate whether single nucleotide polymorphism (SNP) in STAT6 is associated with atopic asthma susceptibility in Egyptian children and to correlate the effect of STAT6 gene polymorphism on IL-4 and total IgE levels. Methods and Findings: the study was conducted on 60 children with atopic asthma in addition, 30 healthy children as controls. The enzyme-linked immunosorbent assay technique (ELISA) was used to measure serum IL-4 and total IgE levels. Genotyping pattern for the STAT6 rs324011 polymorphism was done using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique. Discussion: Our data revealed that rs324011 TT genotype was significantly associated with increased susceptibility of developing bronchial asthma (OR=5.55, 95% CI=1.43-21.44) in comparison to rs324011 CC and CT genotypes. Meanwhile, in bronchial asthma group, serum IL-4 and total IgE levels were higher in patients with CT and TT genotypes compared to those with CC genotype. Conclusion: The T allele variant of STAT6 rs324011 polymorphism may be associated with increased susceptibility of the development of atopic bronchial asthma in Egyptian children; however, further studies are needed to verify these preliminary results.

Meningeal solitary fibrous tumor/hemangiopericytoma: Emphasizing on STAT 6 immunohistochemistry with a review of literature.

The 2016 central nervous system (CNS) World Health Organisation (WHO) Update has merged the entities of meningeal solitary fibrous tumor (SFT) and hemangiopericytoma (HPC) into a single entity based on the presence of the nerve growth factor 1A (NGFI-A) binding protein 2 (NAB2)- signal transducer and activator of transcription 6 (STAT6) gene fusion in these tumors. Immunohistochemical (IHC) staining with STAT6 results in a strong nuclear positivity confirming the diagnosis. Meningeal SFT/HPCs are currently histologically graded according to a three-tiered system. Grade I (SFT phenotype) is benign, whereas grades II and III (HPC phenotype) are malignant and require radiotherapy in addition to gross total resection. The objectives were to review the cases diagnosed as meningeal SFT or HPC between 2010 and 2017 and classify them into SFT (grade I) or HPC (grades II and III) phenotypes; to confirm the diagnosis by performing STAT6 immunohistochemistry; and to observe and record the histological features in detail and correlate the tumor grades with their behavior. The published literature on the subject was also reviewed. A total of 21 cases diagnosed between 2010 and 2017 as meningeal SFT or HPC were included in the study. All cases were reviewed by the authors and were categorized and graded according to histologic phenotype and mitotic count. STAT6 immunohistochemistry was performed in all the cases. The epidemiological data and histologic findings in each case were recorded in detail. The follow-up of patients was obtained. Fifteen patients were males and six were females. The mean age was 43.5 years. The mean tumor size was 6.8 cm. The tumor specimens in 20 out of 21 cases corresponded to the HPC phenotype, of which 6 were in grade II while 14 were in grade III. Thus, over 95% cases had malignant lesions. The tumor in all the 21 cases recruited for the study showed immunohistochemical positivity for SAT6, while CD34 was positive in all the 18 tumor in which it was performed. The follow-up was available in 14 of the patients. Recurrence occurred in six patients who had either a grade II or a grade III tumor and three patients died (including one patient with a grade III tumor. This patient died a month after initial resection although there was no evidence of recurrence). Radiotherapy was given to only 4 out of 14 patients in whom follow-up was available. These rare tumors need to be accurately diagnosed and optimally treated (gross total resection and radiotherapy) to improve the prognosis.

Toll-Interacting Protein, Tollip, Inhibits IL-13-Mediated Pulmonary Eosinophilic Inflammation in Mice

Toll-interacting protein (Tollip) is a key negative regulator of innate immunity by preventing excessive proinflammatory responses. Tollip genetic variation has been associated with airflow limitation in asthma subjects and Tollip expression. Whether Tollip regulates lung inflammation in a type 2 cytokine milieu (e.g., IL-13) is unclear. Our goal was to determine the in vivo role of Tollip in IL-13-mediated lung eosinophilic inflammation and the underlying mechanisms. Tollip-knockout (KO) and wild-type (WT) mice were inoculated intranasally with recombinant mouse IL-13 protein to examine lung inflammation. To determine how Tollip regulates inflammation, alveolar macrophages and bone marrow-derived macrophages from Tollip KO and WT mice were cultured with or without IL-13 and/or IL-33. IL-13-treated Tollip KO mice significantly increased lung eosinophilic inflammation and eotaxin-2 (CCL24) levels compared with the WT mice. IL-13- treated Tollip KO (vs. WT) macrophages, in the absence and particularly in the presence of IL-33, increased expression of the IL-33 receptor ST2L and CCL24, which was in part dependent on enhanced activation of interleukin (IL)-1 receptor-associated kinase 1 (IRAK1) and signal transducer and activator of transcription 6 (STAT6). Our results suggest that Tollip downregulates IL-13-mediated pulmonary eosinophilia in part through inhibiting the activity of the ST2L/IL-33/IRAK1 axis and STAT6.