OBJECTIVE: To investigate the expression of cytokine signal suppressor 3 (SOCS3) in colon cancer tissue and the mechanism by which SOCS3 regulates the proliferation and invasion of colon cancer. METHODS: We collected the specimens of tumor tissues and paired adjacent tissues from 80 patients with colon cancer undergoing radical resection in our hospital between July, 2014 and May, 2017, and the expression of SOCS3 in the tissue samples was analyzed using Western blotting. We also transfected colon cancer cell line SW480 with a SOCS3-overexpressing plasmid or a small interference RNA (siRNA) for SOCS3 knockdown, and the changes in the cell proliferation and invasion capacity were evaluated using CCK-8 assay and Transwell assay, respectively. The effect of demethylation and IL-6 treatment on SOCS3 expression and the proliferation and invasion of SW480 cells were observed. RESULTS: Colon cancer tissues showed a lowered expression of SOCS3 compared with the adjacent tissues. Over-expression of SOCS3 significantly inhibited while SOCS3 knockdown obviously promoted the proliferation and invasion of SW480 cells . Demethylation treatment up-regulated SOCS3 expression and inhibited the proliferation and invasion capacity of SW480 cells; IL-6 treatment of the cells caused the reverse changes. CONCLUSIONS SOCS3 participates in the development and progression of colon cancer and serves as a potential target for colon cancer treatment. In patients with colon cancer, the low expression of SOCS3 possibly as a result of methylation may promote the proliferation and invasion of the cancer cells.
Cell Line, Tumor ; Cell Proliferation ; Colonic Neoplasms ; etiology ; pathology ; Cytokines ; Demethylation ; Disease Progression ; Humans ; Interleukin-6 ; pharmacology ; Neoplasm Invasiveness ; Neoplasm Proteins ; metabolism ; RNA, Small Interfering ; Signal Transduction ; Suppressor of Cytokine Signaling 3 Protein ; genetics ; metabolism ; Transfection

Catalpol, a major bioactive component from Rehmannia glutinosa, which has been used to treat diabetes. The present study was designed to elucidate the anti-diabetic effect and mechanism of action for catalpol in db/db mice. The db/db mice were randomly divided into six groups (10/group) according to their blood glucose levels: db/db control, metformin (positive control), and four dose levels of catalpol treatment (25, 50, 100, and 200 mg·kg), and 10 db/m mice were used as the normal control. All the groups were administered orally for 8 weeks. The levels of fasting blood glucose (FBG), random blood glucose (RBG), glucose tolerance, insulin tolerance, and glycated serum protein (GSP) and the globe gene expression in liver tissues were analyzed. Our results showed that catalpol treatment obviously reduced water intake and food intake in a dose-dependent manner. Catalpol treatment also remarkably reduce fasting blood glucose (FBG) and random blood glucose (RBG) in a dose-dependent manner. The RBG-lowering effect of catalpol was better than that of metformin. Furthermore, catalpol significantly improved glucose tolerance and insulin tolerance via increasing insulin sensitivity. Catalpol treatment significantly decreased GSP level. The comparisons of gene expression in liver tissues among normal control mice, db/db mice and catalpol treated mice (200 and 100 mg·kg) indicated that there were significant increases in the expressions of 287 genes, whichwere mainly involved in lipid metabolism, response to stress, energy metabolism, and cellular processes, and significant decreases in the expressions of 520 genes, which were mainly involved in cell growth, death, immune system, and response to stress. Four genes expressed differentially were linked to glucose metabolism or insulin signaling pathways, including Irs1 (insulin receptor substrate 1), Idh2 (isocitrate dehydrogenase 2 (NADP), mitochondrial), G6pd2 (glucose-6-phosphate dehydrogenase 2), and SOCS3 (suppressor of cytokine signaling 3). In conclusion, catalpol ecerted significant hypoglycemic effect and remarkable therapeutic effect in db/db mice via modulating various gene expressions.
Animals ; Blood Glucose ; metabolism ; Diabetes Mellitus, Experimental ; drug therapy ; genetics ; metabolism ; Disease Models, Animal ; Dose-Response Relationship, Drug ; Drugs, Chinese Herbal ; administration & dosage ; analysis ; Gene Expression ; drug effects ; Glucosephosphate Dehydrogenase ; genetics ; metabolism ; Humans ; Hypoglycemic Agents ; administration & dosage ; Insulin ; metabolism ; Insulin Receptor Substrate Proteins ; genetics ; metabolism ; Iridoid Glucosides ; administration & dosage ; analysis ; Isocitrate Dehydrogenase ; genetics ; metabolism ; Liver ; drug effects ; metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Rehmannia ; chemistry ; Suppressor of Cytokine Signaling 3 Protein ; genetics ; metabolism

OBJECTIVETo explore the mRNA expression level of Notch1 and ASB2 in bone marrow mononuclear cells of patients with P210(+) chronic myeloid leukemia and the correlation between Notch signaling pathway and ubiquitination in chronic myeloid leukemia, so as to provide the valuable information for investigating the pathogenesis of chronic myeloid leukemia and clinical treatment.

METHODSBone marrow was collected from 32 patients with newly diagnosed chronic myeloid leukemia and 34 non-hematopathic and healthy individuals (control group), respectively. The mRNA expression levels of Notch1 and ASB2 were detected by real-time quantitative PCR.

RESULTSThe expression of Notch1 mRNA in CML patients were significantly different from that of healthy individuals group (t=36.3, P<0.01), which was 337.8 times of the healthy individuals. Moreover, the expression level of ASB2 mRNA in CML group was significantly higher than that in healthy individuals (t=19.4, P<0.01). The mRNA expression of Notch1 and ASB2 gene was positive correlation (r=0.504, P<0.01).

CONCLUSIONThe aberrant expression of Notch1 and Asb2 exists in patients with P210 positive CML, which may be involved in incidence and development of CML.

Bone Marrow ; metabolism ; Case-Control Studies ; Humans ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive ; genetics ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Receptor, Notch1 ; genetics ; metabolism ; Signal Transduction ; Suppressor of Cytokine Signaling Proteins ; genetics ; metabolism

Liopxin A4 (LXA4) is considered to be a crucial modulator in the inflammatory responses. In the present study, we aimed to study the effect of LXA4 on the inflammatory cytokines production induced by lipopolysaccharide (LPS) and the possible mechanism in normal human epidermal keratinocytes (NHEKs). NHEKs were isolated and cultured. The expression of toll-like receptor 4 (TLR4), LXA4 receptor (ALXR) and aryl hydrocarbon receptor (AhR) in NHEKs was detected by reverse transcription polymerase chain reaction (RT-PCR). The mRNA and protein levels of tumor necrosis factor-alpha (TNF-α) and interleukin-1β (IL-1β) were determined in NHEKs stimulated by LPS (10 μg/mL) with or without preincubation with LXA4 (100 nmol/L) for 30 min by real-time quantitative PCR (real-time qPCR) and enzyme-linked immunosorbent assay (ELISA), respectively. The expression levels of tumor necrosis factor receptor-associated factor 6 (TRAF6) and suppressors of cytokine signaling 2 (SOCS2) mRNAs and proteins, and nuclear translocation of NF-kB-p65 were measured by real-time qPCR and Western blotting, respectively. The results showed that NHEKs expressed TLR4, ALXR and AhR. LXA4 significantly inhibited the mRNA and protein expression levels of TNF-α, IL-1β and TRAF6 induced by LPS in NHEKs, and LXA4 obviously increased the expression of SOCS2 at mRNA and protein levels. The nuclear NF-kB-p65 protein expression induced by LPS was inhibited after preincubation with LXA4 in NHEKs. It was concluded that LXA4 inhibits the LPS-induced production of TNF-α and IL-1β in NHEKs by up-regulating SOCS2 and down-regulating TRAF6.
Anti-Inflammatory Agents, Non-Steroidal ; pharmacology ; Cells, Cultured ; Gene Expression Regulation ; drug effects ; Humans ; Keratinocytes ; Lipopolysaccharides ; pharmacology ; Lipoxins ; pharmacology ; NF-kappa B ; genetics ; metabolism ; Suppressor of Cytokine Signaling Proteins ; genetics ; metabolism ; TNF Receptor-Associated Factor 6 ; genetics ; metabolism ; Toll-Like Receptor 4 ; genetics ; metabolism ; Tumor Necrosis Factor-alpha ; genetics ; metabolism

Abnormal levels of microRNA (miR)-155, which regulate inflammation and immune responses, have been demonstrated in the colonic mucosa of patients with inflammatory bowel diseases (IBD), although its role in disease pathophysiology is unknown. We investigated the role of miR-155 in the acquisition and maintenance of an activated phenotype by intestinal myofibroblasts (IMF), a key cell population contributing to mucosal damage in IBD. IMF were isolated from colonic biopsies of healthy controls, ulcerative colitis (UC) and Crohn's disease (CD) patients. MiR-155 in IMF was quantified by quantitative reverse transcription-PCR in basal condition and following exposure to TNF-alpha, interleukin (IL)-1beta, lipopolysaccharide (LPS) or TGF-beta1. The effects of miR-155 mimic or inhibitor transfection on cytokine release and suppressor of cytokine signaling 1 (SOCS1) expression were assessed by enzyme-linked immunosorbent assay and western blot, respectively. Regulation of the target gene SOCS1 expression by miR-155 was assessed using luciferase reporter construct. We found that miR-155 was significantly upregulated in UC as compared with control- and CD-derived IMF. Moreover, TNF-alpha and LPS, but not TGF-beta1 and IL-1beta, significantly increased miR-155 expression in IMF. Ectopic expression of miR-155 in control IMF augmented cytokines release, whereas it downregulated SOCS1 expression. MiR-155 knockdown in UC-IMF reduced cytokine production and enhanced SOCS1 expression. Luciferase reporter assay demonstrated that miR-155 directly targets SOCS1. Moreover, silencing of SOCS1 in control IMF significantly increased IL-6 and IL-8 release. In all, our data suggest that inflammatory mediators induce miR-155 expression in IMF of patients with UC. By downregulating the expression of SOCS1, miR-155 wires IMF inflammatory phenotype.
Adult ; Aged ; Cells, Cultured ; Colitis, Ulcerative/*genetics/immunology/*pathology ; Cytokines/immunology ; Female ; *Gene Expression Regulation ; Humans ; Intestinal Mucosa/immunology/metabolism/pathology ; Male ; MicroRNAs/*genetics ; Middle Aged ; Myofibroblasts/immunology/metabolism/*pathology ; Suppressor of Cytokine Signaling Proteins/*genetics ; Tumor Necrosis Factor-alpha/immunology ; Up-Regulation ; Young Adult

Abnormal levels of microRNA (miR)-155, which regulate inflammation and immune responses, have been demonstrated in the colonic mucosa of patients with inflammatory bowel diseases (IBD), although its role in disease pathophysiology is unknown. We investigated the role of miR-155 in the acquisition and maintenance of an activated phenotype by intestinal myofibroblasts (IMF), a key cell population contributing to mucosal damage in IBD. IMF were isolated from colonic biopsies of healthy controls, ulcerative colitis (UC) and Crohn's disease (CD) patients. MiR-155 in IMF was quantified by quantitative reverse transcription-PCR in basal condition and following exposure to TNF-alpha, interleukin (IL)-1beta, lipopolysaccharide (LPS) or TGF-beta1. The effects of miR-155 mimic or inhibitor transfection on cytokine release and suppressor of cytokine signaling 1 (SOCS1) expression were assessed by enzyme-linked immunosorbent assay and western blot, respectively. Regulation of the target gene SOCS1 expression by miR-155 was assessed using luciferase reporter construct. We found that miR-155 was significantly upregulated in UC as compared with control- and CD-derived IMF. Moreover, TNF-alpha and LPS, but not TGF-beta1 and IL-1beta, significantly increased miR-155 expression in IMF. Ectopic expression of miR-155 in control IMF augmented cytokines release, whereas it downregulated SOCS1 expression. MiR-155 knockdown in UC-IMF reduced cytokine production and enhanced SOCS1 expression. Luciferase reporter assay demonstrated that miR-155 directly targets SOCS1. Moreover, silencing of SOCS1 in control IMF significantly increased IL-6 and IL-8 release. In all, our data suggest that inflammatory mediators induce miR-155 expression in IMF of patients with UC. By downregulating the expression of SOCS1, miR-155 wires IMF inflammatory phenotype.
Adult ; Aged ; Cells, Cultured ; Colitis, Ulcerative/*genetics/immunology/*pathology ; Cytokines/immunology ; Female ; *Gene Expression Regulation ; Humans ; Intestinal Mucosa/immunology/metabolism/pathology ; Male ; MicroRNAs/*genetics ; Middle Aged ; Myofibroblasts/immunology/metabolism/*pathology ; Suppressor of Cytokine Signaling Proteins/*genetics ; Tumor Necrosis Factor-alpha/immunology ; Up-Regulation ; Young Adult

Recently, suppressor of cytokine signaling-3 (SOCS3) has been shown to be an inducible endogenous negative regulator of Janus kinase/signal transducers and activators of transcription (JAK/STAT) pathway which is relevant in inflammatory response, while its functions in acute liver failure and HBV-induced acute-on-chronic liver failure (HBV-ACLF) have not been fully elucidated. In this study, we explored the role of SOCS3 in the development of mouse hepatitis virus strain 3 (MHV-3)-induced acute liver failure and its expression in liver and peripheral blood mononuclear cells (PBMCs) of patients with HBV-ACLF. Inflammation-related gene expression was detected by real-time PCR, immunohistochemistry and Western blotting. The correlation between SOCS3 level and liver injury was studied. Our results showed that the SOCS3 expression was significantly elevated in both the liver tissue and PBMCs from patients with HBV-ACLF compared to mild chronic hepatitis B (CHB). Moreover, a time course study showed that SOCS3 level was increased remarkably in the liver of BALB/cJ mice at 72 h post-infection. Pro-inflammatory cytokines, interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α, were also increased significantly at 72 h post-infection. There was a close correlation between hepatic SOCS3 level and IL-6, and the severity of liver injury defined by alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, respectively. These data suggested that SOCS3 may play a pivotal role in the pathogenesis of MHV-3-induced acute liver failure and HBV-ACLF.
Adult ; Alanine Transaminase ; blood ; Animals ; Aspartate Aminotransferases ; blood ; Blotting, Western ; End Stage Liver Disease ; genetics ; pathology ; virology ; Female ; Gene Expression ; Hepatitis, Viral, Animal ; genetics ; pathology ; virology ; Host-Pathogen Interactions ; Humans ; Interleukin-1beta ; genetics ; metabolism ; Interleukin-6 ; genetics ; metabolism ; Leukocytes, Mononuclear ; metabolism ; virology ; Liver Failure, Acute ; genetics ; pathology ; virology ; Male ; Mice, Inbred BALB C ; Middle Aged ; Murine hepatitis virus ; physiology ; Reverse Transcriptase Polymerase Chain Reaction ; Severity of Illness Index ; Suppressor of Cytokine Signaling 3 Protein ; Suppressor of Cytokine Signaling Proteins ; blood ; genetics ; metabolism ; Tumor Necrosis Factor-alpha ; genetics ; metabolism ; Young Adult

BACKGROUNDSome hepatitis B extracellular antigen (HBeAg)-positive chronic hepatitis B (CHB) patients in their immune active phase can clear the virus spontaneously and enter into an inactive hepatitis B virus (HBV) carrier state, indicating a benign prognosis. In this study, the association between cytokine-inducible SRC homology 2 domain protein (CISH) gene polymorphisms at -292 (rs414171) and the spontaneous clearance of HBV in HBeAg-positive CHB patients in immune the active phase was investigated.

METHODSSeventy HBeAg-positive CHB patients in the immune active phase were followed up for 76 weeks without antiviral therapy. The alanine transaminase, aspartate transaminase, HBV DNA, HBeAg and hepatitis B extracellular antibody levels were tested regularly. At week 76, 27 patients were classified into group A (HBV DNA level below 2 104 IU/ml and the value of HBeAg declined below 10% of the baseline at week 76), and 43 patients were classified into group B (HBV DNA level higher than 2×10(4) IU/ml or the value of HBeAg did not decline substantially at week 76). CISH (rs414171) polymorphisms were also tested using the iPLEX system.

RESULTSThe HBV DNA levels at week 12 were significantly greater in group B compared with group A (group A: (6.87±1.40) log10IU/ml; group B: (7.61±1.38) log10IU/ml, P = 0.034) and the HBeAg values were greater in group B at week 28 compared with group A (P = 0.001). The differences in HBV DNA and HBeAg values increased between the groups over time. Sixteen patients in group A and 11 in group B were genotype AA. Those with genotype AT or TT included 11 in group A and 31 in group B (AA vs. AT and TT, odds ratio 4.10 (95% confidence interval: 1.462-11.491), P = 0.006).

CONCLUSIONCISH gene polymorphisms at -292 (rs414171) are associated with HBV clearance in HBeAg-positive CHB patients in the immune active phase, and AA is a favorable genotype for this effect.

Adolescent ; Adult ; Aged ; Female ; Genotype ; Hepatitis B e Antigens ; metabolism ; Hepatitis B virus ; genetics ; immunology ; Hepatitis B, Chronic ; immunology ; Humans ; Male ; Middle Aged ; Polymorphism, Genetic ; genetics ; Suppressor of Cytokine Signaling Proteins ; genetics ; Young Adult

BACKGROUNDIdiopathic pulmonary fibrosis (IPF) is a progressive diffuse parenchymal disease with a poor prognosis. A variety of cytokines and chemokines are involved in its pathophysiology. The aim of this study was to evaluate the clinical features in IPF patients with the expression of suppressor of cytokine signaling 1 (SOCS-1), which acts as a negative regulator of cytokine signaling.

METHODSIPF patients (n = 20) and healthy controls (n = 16) were included in this study. The expression of SOCS-1 was analyzed in peripheral blood mononuclear cells (PBMC) of subjects using RT-PCR. Interleukin 4 (IL-4), transforming growth factor β1 (TGF-β1) and type I collagen expression were also analyzed in each individual using enzyme-linked immunosorbent assay (ELISA). The clinical characteristics of IPF patients were delineated. These results were analyzed by SPSS13.0 statistics software.

RESULTSSOCS-1 mRNA expression was significantly decreased in the PBMC of IPF patients compared with healthy controls; serum levels of IL-4 and TGF-β1 were higher in IPF patients. The patients with lower expression of SOCS-1 developed lower percentage of forced vital capacity (FVC%) and DLCO/VA. A patients' SOCS-1 mRNA level was negatively correlated with serum levels of IL-4, and negatively correlated with their high-resolution computed tomography (HRCT) scores.

CONCLUSIONSSOCS-1 mRNA can be detected in PBMC, and it is down-regulated in IPF patients. The expression of SOCS-1 is associated with the severity of IPF patients' symptoms, so it might be the predictor of disease severity. SOCS-1 might play an important role in IPF by reducing the expression of the T helper type 2 (Th2) cell-related cytokine IL-4.

Adult ; Aged ; Aged, 80 and over ; Enzyme-Linked Immunosorbent Assay ; Female ; Humans ; Idiopathic Pulmonary Fibrosis ; metabolism ; Interleukin-4 ; metabolism ; Leukocytes, Mononuclear ; metabolism ; Male ; Middle Aged ; Suppressor of Cytokine Signaling 1 Protein ; Suppressor of Cytokine Signaling Proteins ; genetics ; metabolism ; Transforming Growth Factor beta1 ; metabolism

The microRNA (miRNA) regulation mechanisms associated with atherosclerosis are largely undocumented. Specific selection and efficient validation of miRNA regulation pathways involved in atherosclerosis development may be better assessed by contemporary microarray platforms applying cross-verification methodology. A screening platform was established using both miRNA and genomic microarrays. Microarray analysis was then simultaneously performed on pooled atherosclerotic aortic tissues from 10 Apolipoprotein E (apoE) knockout mice (apoE-/-) and 10 healthy C57BL/6 (B6) mice. Differentiated miRNAs were screened and cross-verified against an mRNA screen database to explore integrative mRNA-miRNA regulation. Gene set enrichment analysis was conducted to describe the potential pathways regulated by these mRNA-miRNA interactions. High-throughput data analysis of miRNA and genomic microarrays of knockout and healthy control mice revealed 75 differentially expressed miRNAs in apoE-/- mice at a threshold value of 2. The six miRNAs with the greatest differentiation expression were confirmed by real-time quantitative reverse-transcription PCR (qRT-PCR) in atherosclerotic tissues. Significantly enriched pathways, such as the type 2 diabetes mellitus pathway, were observed by a gene-set enrichment analysis. The enriched molecular pathways were confirmed through qRT-PCR evaluation by observing the presence of suppressor of cytokine signaling 3 (SOCS3) and SOCS3-related miRNAs, miR-30a, miR-30e and miR-19b. Cross-verified high-throughput microarrays are optimally accurate and effective screening methods for miRNA regulation profiles associated with atherosclerosis. The identified SOCS3 pathway is a potentially valuable target for future development of targeted miRNA therapies to control atherosclerosis development and progression.
Animals ; Aorta/metabolism/pathology ; Apolipoproteins E/*deficiency/metabolism ; Atherosclerosis/genetics/pathology ; Down-Regulation/genetics ; Gene Expression Profiling ; *Gene Expression Regulation ; Gene Regulatory Networks/genetics ; Immunohistochemistry ; Male ; Mice ; Mice, Inbred C57BL ; Mice, Knockout ; MicroRNAs/*genetics/metabolism ; Models, Biological ; *Oligonucleotide Array Sequence Analysis ; RNA, Messenger/genetics/metabolism ; Reproducibility of Results ; Reverse Transcriptase Polymerase Chain Reaction ; Signal Transduction/*genetics ; Suppressor of Cytokine Signaling Proteins/genetics/metabolism ; Up-Regulation/genetics