Suppressor Of Cytokine Signaling 3 Gene Expression Research Articles

Leptin receptor expression in blood mononuclear cells of lactating women is associated with infant body weight: Potential role as a molecular biomarker.

Molecular biomarkers of maternal leptin resistance associated with infant weight are needed. To evaluate gene expression of leptin receptor (LEPR), suppressor of cytokine signalling 3 (SOCS3) and insulin receptor in peripheral blood mononuclear cells (PBMCs) of lactating women and their relationship with infant body weight and adiposity. At day 10 postpartum, maternal gene expression in PBMCs as well as leptin and insulin concentrations in plasma and milk were assessed (n = 68). Infant weight and BMI z-scores, skinfolds and arm circumference were obtained at 10 days and/or at 3 months old. In mothers with pre-pregnancy overweight or obesity (OW/OB), LEPR expression was reduced (p = 0.013) whereas plasma and milk leptin and milk insulin concentrations were elevated. LEPR expression was positively related with infant weight z-score (Beta (95% CI): 0.40 (0.17, 0.63), p = 0.001) but not with leptin concentrations. SOCS3 expression was positively related with infant weight z-score (Beta (95% CI): 0.28 (0.04, 0.51), p = 0.024) and arm circumference (Beta (95% CI): 0.57 (0.32, 0.82), p < 0.001). Relationships remained significant after adjusting for maternal and infant confounders. LEPR and SOCS3 gene expression in PBMCs are novel maternal molecular biomarkers that reflect leptin resistance and are associated with infant body weight and adiposity.

The impact of sitagliptin in palmitic acid-induced insulin resistance in human HepG2 cells through the suppressor of cytokine signaling 3/phosphoinositide 3-kinase/protein kinase B pathway.

Patients with type 2 diabetes respond differently to sitagliptin, an oral anti-hyperglycemic medication. Patients whose blood sugar levels were effectively managed while using sitagliptin had significantly lower levels of a protein called suppressor of cytokine signaling 3 (SOCS3), according to our earlier research. In this study, we established an in vitro insulin resistance cell model for human HepG2 cells to investigate the possible mechanism of the effect of sitagliptin on glucose metabolism via the SOCS3/phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) pathway. Since insulin resistance first develops in the liver, palmitic acid was used to generate an insulin resistance cell model in human HepG2 cells, after which small interfering ribonucleic acid (siRNA)-SOCS3 and sitagliptin were used to intervene. We then examined the changes in cell viability and biochemical indices in the insulin resistance cell model. SOCS3, Akt, and glycogen synthase kinase 3beta (GSK-3β) gene expression levels were quantified using reverse transcription-polymerase chain reaction, and the protein expression levels of SOCS3, Akt, phosphorylated Akt (p-Akt), GSK-3β, and phosphorylated GSK-3β (p-GSK-3β) were quantified using Western blot. In results: the expression of the SOCS3 gene was considerably raised in both the insulin resistance model group and the insulin resistance model + siRNA-negative control group, but decreased following treatment with sitagliptin. After sitagliptin intervention, the protein expression of Akt, p-Akt, and p-GSK-3β were dramatically decreased in the model group, while SOCS3 was significantly decreased. We conclude that sitagliptin can reduce insulin resistance by downregulating SOCS3 and regulating glucose metabolism in a hypoglycemic manner.

Involvement of the IL-6 Signaling Pathway in the Anti-Anhedonic Effect of the Antidepressant Agomelatine in the Chronic Mild Stress Model of Depression.

Neuroinflammation has emerged as an important factor in the molecular underpinnings of major depressive disorder (MDD) pathophysiology and in the mechanism of action of antidepressants. Among the inflammatory mediators dysregulated in depressed patients, interleukin (IL)-6 has recently been proposed to play a crucial role. IL-6 activates a signaling pathway comprising the JAK/STAT proteins and characterized by a specific negative feedback loop exerted by the cytoplasmic protein suppressor of cytokine signalling-3 (SOCS3). On these bases, here, we explored the potential involvement of IL-6 signaling in the ability of the antidepressant drug agomelatine to normalize the anhedonic-like phenotype induced in the rat by chronic stress exposure. To this aim, adult male Wistar rats were subjected to the chronic mild stress (CMS) paradigm and chronically treated with vehicle or agomelatine. The behavioral evaluation was assessed by the sucrose consumption test, whereas molecular analyses were performed in the prefrontal cortex. We found that CMS was able to stimulate IL-6 production and signaling, including SOCS3 gene and protein expression, but the SOCS3-mediated feedback-loop inhibition failed to suppress the IL-6 cascade in stressed animals. Conversely, agomelatine treatment normalized the stress-induced decrease in sucrose consumption and restored the negative modulation of the IL-6 signaling via SOCS3 expression and activity. Our results provide additional information about the pleiotropic mechanisms that contribute to agomelatine’s therapeutic effects.

The mediation role of SOCS3 methylation in the effect of serum testosterone on type 2 diabetes.

Previous studies reported that testosterone and DNA methylation of suppressor of cytokine signaling-3 (SOCS3) were associated with type 2 diabetes (T2D). Testosterone affects SOCS3 gene expression. Therefore, we aimed to investigate how the SOCS3 methylation mediates the relationship between testosterone and T2D among Chinese rural adults. A case-control study comprised 365 T2D patients and 651 controls was conducted. Liquid chromatography-tandem mass spectrometry and MethylTarget were used to determine the levels of serum testosterone and DNA methylation of SOCS3 gene, respectively. The odds ratio (OR) of testosterone or SOCS3 methylation for T2D was calculated using logistic regression models, and β value of testosterone for SOCS3 methylation was evaluated by linear regression models. Furthermore, through mediation analysis the mediating effect of SOCS3 methylation on the association of testosterone with T2D was estimated. After adjusting for multiple variables, the protective effect of testosterone on T2D was found in men (OR=0.61, 95% confidence interval [CI]: 0.47-0.80), and the methylation of Chr17:76356190 or Chr17:76356199 was negatively related to T2D in both men and women. Moreover, testosterone was positively associated with Chr17:76356190 methylation in men and Chr17:76356199 methylation in women (both P < .05). The mediation analysis showed that the Chr17:76356190 methylation partly mediated effect of testosterone on T2D in men was approximately 8.2%. High levels of serum testosterone in men and Chr17:76356190 and Chr17:76356199 (SOCS3) methylation were related to a lower prevalent T2D. In addition, Chr17:76356190 methylation partially mediated the effect of testosterone on T2D in Chinese rural men.

Adoptive transfer of bone marrow-derived dendritic cells (BMDCs) alleviates OVA-induced allergic airway inflammation in asthmatic mice

Airway dendritic cells (DCs) are recognized as important factors in the mechanisms of allergic inflammatory diseases. Suppressor of cytokine signaling 3 (SOCS3) is involved in regulating the functions of T cells and macrophages, but the roles of SOCS3-expressing DCs in the pathogeneses of allergic inflammatory diseases are still controversial. We compared the effects of adoptively transferred SOCS3−/− and SOCS3+/+ bone marrow-derived DCs (BMDCs) on airway inflammation in ovalbumin (OVA)-sensitized asthmatic mice. Adoptive transfer of mature DCs (lipopolysaccharide [LPS]-induced DCs, DClps) with or without SOCS3 gene expression significantly ameliorated allergic airway inflammation. SOCS3−/− DCs slightly attenuated BMDC-induced immunogenic tolerance. DClps migrated to OVA-sensitized lungs with higher efficiency than immature DCs (DCim). DClps with or without SOCS3 greatly improved lung pathology scores and alleviated airway inflammatory cell infiltration after adoptive transfer into mice; they also increased interleukin-10 (IL-10) and transforming growth factor-β (TGF-β) production and inhibited signal transducer and activator of transcription (STAT) 4 and STAT6 signaling in the lungs after OVA sensitization. In conclusion, the BMDC adoptive transfer-induced immunogenic tolerance in OVA-sensitized mice might not be due to SOCS3 gene depletion. BMDC adoptive transfer may be developed into a new approach that alleviates asthma by modulating the balance between immune tolerance and inflammation.

Expression and methylation levels of suppressor of cytokine signaling 3 in rheumatic arthritis synovial fibroblasts

BackgroundIn the present study, we aimed to understand the expression and methylation levels of the suppressor of cytokine signaling 3 (SOCS3) in rheumatoid arthritis (RA) synovial fibroblasts. MethodThe RA model was established using Freund's complete adjuvant, and then the synovial fibroblasts were isolated and cultured. Next, RNA extraction and reverse transcription were performed. The SOCS3 transcription level was detected using qPCR, and SOCS3 protein expression was detected using western blotting (WB). Lastly, methylation-specific PCR (MSP) was performed. ResultsThe RA model was successfully demonstrated. SOCS3 gene (p < .01) and protein expression levels were significantly increased in the RA rat group compared to in the wild type (WT) group. However, no significant difference was observed in the MSP products between the RA and WT groups. ConclusionThe increased expression of the SOCS3 can be correlated with the development of RA.

High-Dose TSLP Induces Co-Receptor Internalization and Signal Shutdown in Ph-like B-ALL with Overexpression of CRLF2

Background. Approximately half of all Ph-like B cell acute lymphoblastic leukemia is characterized by overexpression of CRLF2 (CRLF2 B-ALL). CRLF2 B-ALL is associated with high rates of relapse and is more prevalent in Hispanic children with Native American ancestry. CRLF2, together with the IL-7 receptor alpha (IL-7Ra), comprises a receptor complex that is activated by the cytokine, TSLP. Receptor activation by TSLP induces JAK2/STAT5 and PI3/AKT/mTOR signals that promote survival and proliferation of leukemia cells. To study the role of TSLP in CRLF2 B-ALL, we developed a patient-derived xenograft (PDX) model of CRLF2 B-ALL that allows us to vary circulating levels of human TSLP (hTSLP) in a physiologic environment. We generated PDX from patients' CRLF2 B-ALL cells and compared leukemia burden in mice with varying levels of hTSLP. CRLF2 B-ALL cells grew robustly in PDX models with hTSLP levels at or below levels present in pediatric cancer patients (~10 pg/mL). In contrast, CRLF2 B-ALL cells were essentially eliminated in PDXs with hTSLP at high physiological levels (40-140 pg/mL). These data suggest that high physiologic levels of TSLP exert a potent anti-leukemia effect in Ph-like B-ALL with overexpression of CRLF2.Objective. The objective of the proposed research was to evaluate potential mechanisms of TSLP's anti-leukemia effects.Research Design/Methods. TSLP dose response studies were performed and flow cytometry was used to evaluate the effect of TSLP on CRLF2 signaling, surface expression of TSLP receptor components (IL-7Ra and CRLF2) and expression of the Suppressor of Cytokine Signaling (SOCS) proteins in CRLF2 B-ALL cell lines (MUTZ5 and CALL-4) and CRLF2 B-ALL cells from Hispanic pediatric patients. Whole genome microarray was used to evaluate TSLP effects on SOCS gene expression in patient samples.Results. CRLF2 B-ALL cell lines cultured with TSLP showed a dose-dependent loss in the ability to induce phosphorylation of STAT5 and S6 (downstream of PI3/AKT/mTOR) following TSLP stimulation. This loss was correlated with a complete loss of surface IL-7Ra, and maintained for 24-48 hours following a pulse of high-dose, but not low-dose, hTSLP. The loss of surface CRLF2 was minimal. The loss of signaling and surface IL-7Ra could be prolonged if high-dose hTSLP levels were maintained. Similarly, preliminary studies of CRLF2 B-ALL cells from pediatric patients showed a loss of surface IL-7Ra following culture with high-dose TSLP. A potential mechanism for the effects of high-dose TSLP are the suppressor of cytokine signaling (SOCS) genes. These genes encode a family of proteins (SOCS1-7 and CISH) that regulate cytokine signaling via negative feedback through multiple mechanisms including ubiquitin-mediated cytokine receptor degradation. Whole genome microarray showed that SOCS1, SOCS2, SOCS3 and CISH mRNA are upregulated in patient CRLF2 B-ALL cells cultured with high-dose TSLP. Flow cytometry analysis showed that high-dose TSLP upregulates SOCS1 and SOCS3 proteins in CRLF2 B-ALL cell lines and in patient samples.Conclusion. These data provide evidence that TSLP exerts anti-leukemia effects by shutting down CRLF2-mediated signals and that these effects are at least partially mediated by the loss of the IL-7Ra component, and potentially through SOCS family proteins. These studies identify the human TSLP cytokine as a potential biologic therapy to treat CRLF2 B-ALL. Supported in part by 1R01CA209829 (KJP and SD), 1R43CA224723 (XM and KJP), ASH HONORS Award 2018-2019 (WBW), and Alpha Omega Alpha Carolyn L. Kuckein Student Research Fellowship 2018-2019 (WBW). DisclosuresCoats:Elf Zone, Inc.: Consultancy, Membership on an entity's Board of Directors or advisory committees. Meng:Elf Zone, Inc.: Employment. Dovat:Elf Zone, Inc.: Membership on an entity's Board of Directors or advisory committees. Payne:Elf Zone, Inc.: Consultancy, Equity Ownership, Membership on an entity's Board of Directors or advisory committees.

The novel exchange protein activated by cyclic AMP 1 (EPAC1) agonist, I942, regulates inflammatory gene expression in human umbilical vascular endothelial cells (HUVECs)

Exchange protein activated by cyclic AMP (EPAC1) suppresses multiple inflammatory actions in vascular endothelial cells (VECs), partly due to its ability to induce expression of the suppressor of cytokine signalling 3 (SOCS3) gene, the protein product of which inhibits interleukin 6 (IL6) signalling through the JAK/STAT3 pathway. Here, for the first time, we use the non-cyclic nucleotide EPAC1 agonist, I942, to determine its actions on cellular EPAC1 activity and cyclic AMP-regulated gene expression in VECs. We demonstrate that I942 promotes EPAC1 and Rap1 activation in HEK293T cells and induces SOCS3 expression and suppresses IL6-stimulated JAK/STAT3 signalling in HUVECs. SOCS3 induction by I942 in HUVECs was blocked by the EPAC1 antagonist, ESI-09, and EPAC1 siRNA, but not by the broad-spectrum protein kinase A (PKA) inhibitor, H89, indicating that I942 regulates SOCS3 gene expression through EPAC1. RNA sequencing was carried out to further identify I942-regulated genes in HUVECs. This identified 425 I942-regulated genes that were also regulated by the EPAC1-selective cyclic AMP analogue, 007, and the cyclic AMP-elevating agents, forskolin and rolipram (F/R). The majority of genes identified were suppressed by I942, 007 and F/R treatment and many were involved in the control of key vascular functions, including the gene for the cell adhesion molecule, VCAM1. I942 and 007 also inhibited IL6-induced expression of VCAM1 at the protein level and blocked VCAM1-dependent monocyte adhesion to HUVECs. Overall, I942 represents the first non-cyclic nucleotide EPAC1 agonist in cells with the ability to suppress IL6 signalling and inflammatory gene expression in VECs.

SOCS3 mRNA expression and polymorphisms as pretreatment predictor of response to HCV genotype 3a IFN-based treatment.

AimSuppressor of Cytokine Signaling 3 (SOCS3) gene belongs to SOCS family as one of the negative regulators of cytokine signaling and IFN response that function via the JAK-STAT pathway in antiviral response. SOCS3 expression and genetic polymorphism influences the pathogenesis and outcome of antiviral treatment in hepatitis C virus (HCV) infected patients. This study was designed for analysis of SOCS3 gene expression and polymorphism in Pakistani HCV patients.MethodsThis descriptive study was conducted on 250 diagnosed HCV genotype 3a infected subjects. The study population was divided into two major groups on the basis of therapeutic response i.e. sustained virological response (SVR) and non-responders/relapsers (NR). SOCS3 gene mRNA expression analysis was done by using Real time PCR technique, whereas ARMS PCR technique was used for analysis of SOCS3 gene polymorphisms i.e. 8464 A/C (rs12952093), −4874 A/G (rs4969170) and −1383 A/G, (rs4969168).ResultsGene expression analysis of SOCS3 showed that there was statistically significant increase of 2.275-fold and 3.72-fold in relative gene expression for SVR and NR as compared to normal healthy samples (p < 0.001). The distribution of rs4969168, rs4969170 and rs12952093 genotype frequencies between SVR versus NR group were not statistically significant, only the allelic frequency of rs4969170 was statistically significant (p ≤ 0.0001) with therapeutic response.ConclusionThe gene expression analysis of SOCS3 showed a clear difference in mRNA expression of SOCS3 as a possible indicator of therapeutic response rather than polymorphism of SOCS3 gene in our studied population.

JAK2, STAT3 and SOCS3 gene expression in women with and without breast cancer.

Breast cancer is a disease that arises from the accumulation of alterations in the genome of cells that make up the mammary gland. The Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) signaling pathway has been reported to participate in the development of breast cancer and is activated by adipocytokines such as leptin, which are elevated in obesity. In contrast, alterations in suppressor of cytokine signaling 3 (SOCS3) gene expression have been found in patients with decreased breast cancer metastasis. The current study sought to identify whether JAK2, STAT3 and SOCS3 gene expression is associated with body mass index (BMI) and breast cancer. This was a cross-sectional prospective study. JAK2, STAT3 and SOCS3 gene expression levels were determined using RT-qPCR from the biopsies of 26 patients with breast cancer and 43 patients with benign breast lesions. We compared the expression of these genes, relative to the housekeeping genes, ACTB and GAPDH, against BMI, clinical stage and immunohistochemistry. STAT3 gene expression was increased in breast cancer patients (p≤0.001; AUC=0.65; AUC 95% CI: 0.5-0.8), and SOCS3 expression was decreased in obese patients with benign breast lesions (p≤0.001; AUC=0.51; AUC 95% CI: 0.36-0.65). With regard to the clinical stage, there were significant differences in STAT3 gene expression between stage II and III (p≤0.011) and stage II and IV (p≤0.033) breast cancers. Among all women, there was a positive correlation between JAK2 and STAT3 expression (R=0.493, p=0.000). In addition, breast cancers that were negative for HER2 were associated with JAK2 and SOCS3 (R=0.645, p=0.003). High levels of STAT3 expression were associated with early stages of breast cancer development and patients in the control group with obesity showed higher expression of SOCS3 regarding overweight.

Parasite Infections in Multiple Sclerosis Modulate Immune Responses through a Retinoic Acid–Dependent Pathway

We recently demonstrated better outcomes in helminth-infected multiple sclerosis (MS) patients, compared with uninfected ones. The present study evaluates the role of TLR2 and retinoic acid (RA) in parasite-driven protection in MS patients. RA serum levels were significantly higher in helminth-infected MS patients than in uninfected MS subjects or healthy controls. Genes involved in RA biosynthesis and metabolism, such as Adh1 and Raldh2, as well as RA receptors and IL-10, were induced in dendritic cells (DCs) via TLR2-dependent ERK signaling. This programmed DCs to induce FOXP3(+) T regulatory cells and suppressed production of proinflammatory cytokines (IL-6, IL-12, IL-23, and TNF-α) via induction of suppressor of cytokine signaling 3 (SOCS3), an effect mediated by soluble egg Ag (SEA) obtained from Schistosoma mansoni, and by RA. SEA-activated DCs also inhibited IL-17 and IFN-γ production through autoreactive T cells. These inhibitory effects were abrogated when SOCS3 gene expression was silenced, indicating that SEA-mediated signaling inhibited production of these cytokines by T cells, through a SOCS3-dependent pathway. Overall, helminth-related immunomodulation observed in MS patients was mediated by TLR2- and RA-dependent pathways, through two different mechanisms, as follows: 1) induction of IL-10 and FOXP3(+) T regulatory cells, and 2) suppression of proinflammatory cytokine production mediated by SOCS3.

Flavanoids induce expression of the suppressor of cytokine signalling 3 (SOCS3) gene and suppress IL-6-activated signal transducer and activator of transcription 3 (STAT3) activation in vascular endothelial cells

The atherogenic cytokine IL-6 (interleukin-6) induces pro-inflammatory gene expression in VECs (vascular endothelial cells) by activating the JAK (Janus kinase)/STAT3 (signal transducer and activator of transcription 3) signalling pathway, which is normally down-regulated by the STAT3-dependent induction of the E3 ubiquitin ligase component SOCS3 (suppressor of cytokine signalling 3). Novel treatments based on the regulation of SOCS3 protein levels could therefore have value in the treatment of diseases with an inflammatory component, such as atherosclerosis. To this end we carried out a screen of 1031 existing medicinal compounds to identify inducers of SOCS3 gene expression and identified the flavanoids naringenin and flavone as effective inducers of SOCS3 protein, mRNA and promoter activity. This was in contrast with the action of traditional JAK/STAT3 inhibitors and the polyphenol resveratrol, which effectively suppress SOCS3 gene expression. Both naringenin and flavone also effectively suppressed IL-6-stimulated phosphorylation of STAT3 (Tyr705) which led to suppression of IL-6-induction of the atherogenic STAT3 target gene MCP1 (monocyte chemotactic protein-1), suggesting that their ability to induce SOCS3 gene expression is STAT3-independent. Supporting this idea was the observation that the general kinase inhibitor compound C inhibits flavone- and cAMP-dependent, but not JAK-dependent, SOCS3 induction in VECs. Indeed, the ability of flavanoids to induce SOCS3 expression requires activation of the ERK (extracellular-signal-regulated kinase)-dependent transcription factor SP3, and not STAT3. In the present paper we therefore describe novel molecular actions of flavanoids, which control SOCS3 gene induction and suppression of STAT3 signalling in VECs. These mechanisms could potentially be exploited to develop novel anti-atherogenic therapies.

Overexpression of suppressor of cytokine signaling 3 in the arcuate nucleus of juvenile Phodopus sungorus alters seasonal body weight changes

The profound seasonal cycle in body weight exhibited by the Djungarian hamster (Phodopus sungorus) is associated with the development of hypothalamic leptin resistance during long day photoperiod (LD, 16:8h light dark cycle), when body weight is elevated relative to short day photoperiod (SD, 8:16h light dark cycle). We previously have shown that this seasonal change in physiology is associated with higher levels of mRNA for the potent inhibitor of leptin signaling, suppressor of cytokine signaling-3 (SOCS3), in the arcuate nucleus (ARC) of LD hamsters relative to hamsters in SD. The alteration in SOCS3 gene expression preceded the body weight change suggesting that SOCS3 might be the molecular switch of seasonal body weight changes. To functionally characterize the role of SOCS3 in seasonal body weight regulation, we injected SOCS3 expressing recombinant adeno-associated virus type-2 (rAAV2-SOCS3) constructs into the ARC of leptin sensitive SD hamsters immediately after weaning. Hamsters that received rAAV2 expressing enhanced green fluorescent protein (rAAV2-EGFP) served as controls. ARC-directed SOCS3 overexpression led to a significant increase in body weight over a period of 12 weeks without fully restoring the LD phenotype. This increase was partially due to elevated brown and white adipose tissue mass. Gene expression of pro-opiomelanocortin was increased while thyroid hormone converting enzyme DIO3 mRNA levels were reduced in SD hamsters with SOCS3 overexpression. In conclusion, our data suggest that ARC-directed SOCS3 overexpression partially overcomes the profound seasonal body weight cycle exhibited by the hamster which is associated with altered pro-opiomelanocortin and DIO3 gene expression.

C2C12 myotube에서 insulin-like growth factor-I이 SOCS-3 유전자 발현에 미치는 영향

SOCS-3와 IGF-I은 근육의 분화 과정 및 근비대 기전에 있어 매우 중요한 조절자 역할을 하는 유전자 및 성장인 자이며, 최근 골격근에서 IGF-I과 SOCS-3 유전자의 상호작용에 관한 연구의 필요성이 제기되고 있다. 본 연구에서는 C2C12 myotube에서 IGF-I이 SOCS-3 유전자 발현에 미치는 영향에 대해 알아보기 위해 4일간 분화시킨 C2C12 myotube에 IGF-I을 다양한 농도(0-200 ng/ml) 및 시간(3-72 시간)에 따라 처리하였다. 그 결과 IGF-I이 SOCS-3 유전자의 단백질 발현을 시간 의존적으로 유의하게 증가시켰으며, 3 시간에서 mRNA 발현을 증가시키고, 시간이 지남에 따라 긴 시간에서는 농도 의존적으로 발현이 감소하였음을 알 수 있었다. 또한 면역형광 염색을 통해 IGF-I이 myotube에서 SOCS-3의 단백질을 발현 시켰음을 뚜렷하게 관찰 할 수 있었다. 위 결과들을 바탕으로 본 연구에서는 IGF-I의 처리가 분화된 근육 세포인 C2C12 myotube에서 SOCS-3 유전자 발현에 유의한 영향을 미쳤음을 증명하였다. 이러한 결과는 선행연구에서 보고한 운동이 SOCS-3 유전자 발현을 증가시킴에 있어서 IGF-I이 중추적인 역할을 한 것으로 생각된다. 그러나 IGF-I에 의한 SOCS-3 유전자 발현 조절 기전에 있어 관련 신호 전달체계 및 골격근 관련 유전자 발현에 미치는 영향에 관한 연구는 보다 더 이루어져야 할 것이라 사료된다. It is well known that both insulin-like growth factor-I and suppressor of cytokine signaling-3 (SOCS-3) are known to modulate various aspects of physiology in skeletal muscle cells. Furthermore, although SOCS-3 expression is related to insulin resistance in non-skeletal muscle cells and is known to interact with insulin-like growth factor-I receptor, the effect of IGF-I on SOCS-3 gene expression in skeletal muscle cells is presently unknown. C2C12 myotubes were treated with different concentrations (0-200 ng/ml) of IGF-I or for various periods of time (3-72 hr). Immunofluorescent staining image revealed that IGF-I induced SOCS-3 protein expression in a dose-dependent manner. Western blot data also showed that SOCS-3 proteins were induced by IGF-I (200 ng/ml) in C2C12 myotubes in a time-dependent manner. The level of SOCS-3 mRNA was also significantly increased after 3hr of IGF-I (10-100 ng/ml) treatment. However, the levels of SOCS-3 mRNA were significantly decreased after 24 and 48 hr of IGF-I (10-100 ng/ml) treatment compared to the control. In conclusion, SOCS-3 protein is induced by IGF-I treatment in C2C12 skeletal muscle cells and this induction is regulated pretranslationally. The modulating effect of IGF-I on SOCS-3 expression may be an important regulator of gene expression in skeletal muscle cells.

SOCS3 and IRS-1 gene expression differs between genotype 1 and genotype 2 hepatitis C virus-infected HepG2 cells

The poor response to antiviral treatment of hepatitis C virus (HCV)-infected patients with genotype 1b has been associated with a higher prevalence of metabolic syndrome. However, the molecular link between these clinical entities is not clear. The goal of this study was to clarify the role of genotype 1b and 2 in the genetic expression of suppressor of cytokine signaling 3 (SOCS3) and insulin receptor substrate 1 (IRS-1). We infected human hepatocellular carcinoma cell line (HepG2) cells with human HCV genotype 1b or 2 and measured the gene and protein expression of SOCS3 at various times. We also evaluated impairment in the insulin pathway by analysis of IRS-1 and phospho-AKT. For the control, we used HepG2 cell cultures treated with non-infectious serum. We also demonstrated the occurrence of HCV infection by the detection of both positive and negative strands in the cells and culture medium. To test infection of the HepG2 cells, we performed quantitative real-time polymerase chain reaction (qRT-PCR) of viral load at different time points. We analyzed the viral genotype in the pellet and supernatant. At each time point, we found positive and negative strands in the infected cells, while in the medium we found positive, but no negative strands. We also detected the presence of the correct genotype in the medium. Two weeks following infection when the viral load was higher, we tested genotype 1b and 2 infected cells. SOCS3 gene expression was significantly higher in genotype 1b-infected cells (median 2.56; mean 2.82+/-0.59) compared with genotype 2 (median 1.34; mean 1.46+/-0.31) (p=0.04) and control cells (median 1.09; mean 1.02+/-0.11, p=0.02). There was no difference between cells exposed to genotype 2 and control cells. Conversely, IRS-1 was significantly lower in genotype 1b-infected cells (median 15.97; mean 15.45+/-0.67) compared with genotype 2-infected cells (median 16.45; mean 16.44+/-0.01, p=0.04). Statistically significant differences were seen when comparing the pAKT/AKT ratio in genotype 1b-infected cells (0.19+/-0.034) and not genotype 1b-infected (genotype 2-infected and non-infected) cells (0.253+/-0.004, p=0.03). This inverse regulation is compatible with interactions between the molecular expression of SOCS3, IRS-1 and phospho-AKT mediated by the genotype 1b virus. Up-regulation of the SOCS3 gene might be one of the mechanisms governing non-response to therapy and expression of insulin resistance mediated via a direct mechanism at this level of genotype 1b HCV.

Over-expression of SOCS-3 Gene Promotes IL-10 Production by JEG-3 Trophoblast Cells

Suppressor of cytokine signaling-3 (SOCS-3) plays an important role in negative regulation of inflammatory response. Evidence has shown that SOCS-3 and IL-10 expressions were significantly reduced in placental trophoblasts from preeclampsia. IL-10 is an anti-inflammatory cytokine. In this study, we sought to determine if enhance SOCS-3 expression could affect IL-10 production in placental trophoblasts. Placental JEG-3 cells were used. Over-expression of SOCS-3 was generated by transfection of JEG-3 cells with a green fluorescent protein (GFP) tagged SOCS-3 gene, SOCS-3/ZsGreen1, by siPORT lipid transfection. Cells transfected with ZsGreen1 vector only was used as control. Our results showed that IL-6 production was reduced in cells over-expressed with SOCS-3. Moreover, SOCS-3 transfected cells produced more IL-10 when stimulated with IL-6. The increased IL-10 production by JEG-3 cells was in a dose-dependent manner, p < 0.05. Our data suggested that enhanced SOCS-3 gene expression could promote IL-10 production by placental trophoblast cells, suggesting that SOCS-3 may play an important role in regulation of cytokine induced anti-inflammatory response in placental trophoblasts.

Influenza A Virus Inhibits Type I IFN Signaling via NF-κB-Dependent Induction of SOCS-3 Expression

The type I interferon (IFN) system is a first line of defense against viral infections. Viruses have developed various mechanisms to counteract this response. So far, the interferon antagonistic activity of influenza A viruses was mainly observed on the level of IFNβ gene induction via action of the viral non-structural protein 1 (NS1). Here we present data indicating that influenza A viruses not only suppress IFNβ gene induction but also inhibit type I IFN signaling through a mechanism involving induction of the suppressor of cytokine signaling-3 (SOCS-3) protein. Our study was based on the observation that in cells that were infected with influenza A virus and subsequently stimulated with IFNα/β, phosphorylation of the signal transducer and activator of transcription protein 1 (STAT1) was strongly reduced. This impaired STAT1 activation was not due to the action of viral proteins but rather appeared to be induced by accumulation of viral 5′ triphosphate RNA in the cell. SOCS proteins are potent endogenous inhibitors of Janus kinase (JAK)/STAT signaling. Closer examination revealed that SOCS-3 but not SOCS-1 mRNA levels increase in an RNA- and nuclear factor kappa B (NF-κB)-dependent but type I IFN-independent manner early in the viral replication cycle. This direct viral induction of SOCS-3 mRNA and protein expression appears to be relevant for suppression of the antiviral response since in SOCS-3 deficient cells a sustained phosphorylation of STAT1 correlated with elevated expression of type I IFN-dependent genes. As a consequence, progeny virus titers were reduced in SOCS-3 deficient cells or in cells were SOCS-3 expression was knocked-down by siRNA. These data provide the first evidence that influenza A viruses suppress type I IFN signaling on the level of JAK/STAT activation. The inhibitory effect is at least in part due to the induction of SOCS-3 gene expression, which results in an impaired antiviral response.

Seasonal effects of central leptin infusion on secretion of melatonin and prolactin and on SOCS-3 gene expression in ewes

Recent studies have demonstrated photoperiodic changes in leptin sensitivity of seasonal mammals. Herein, we examined the interaction of season (long days (LD) versus short days (SD)) and recombinant ovine leptin (roleptin) on secretion of melatonin and prolactin (PRL) and on mRNA expression of suppressor of cytokine signaling-3 (SOCS-3) in the medial basal hypothalamus (MBH) in sheep. Twenty-four Polish Longwool ewes, surgically fitted with third ventricle (IIIV) cannulas, were utilized in a replicated switchback design involving 12 ewes per season. Within-season and replicate ewes were assigned randomly to one of three treatments (four ewes/treatment) and infused centrally three times at 0, 1 and 2 h beginning at sunset. Treatments were 1) control, Ringer-Locke buffer; 2) L1, roleptin, 0.5 microg/kg BW; and 3) L2, roleptin, 1.0 microg/kg BW. Jugular blood samples were collected at 15-min intervals beginning immediately before the start of infusions and continued for 6 h. At the end of blood sampling, a washout period of at least 3 days elapsed before ewes were re-randomized and treated with one of the treatments described above (four ewes/treatment). Ewes were then killed and brains were collected for MBH processing. Leptin treatments increased (P<0.001) circulating leptin concentrations compared with controls during both seasons in a dose-dependent manner. Overall, mean plasma concentrations of melatonin were greater (P<0.001) during LD than SD. However, leptin treatments increased melatonin concentrations during SD in a dose-dependent manner and decreased it during LD. Similarly, plasma concentrations of PRL were greater (P<0.001) during LD than SD. However, unlike changes in melatonin, circulating PRL decreased (P<0.001) in response to leptin during LD. Semi-quantitative PCR revealed that leptin increased (P<0.001) SOCS-3 expression in the MBH region during LD in a dose-dependent manner. Data provide evidence that secretion of photoperiodic hormones such as melatonin and PRL are inversely regulated by leptin during SD and LD. However, the increase in expression of SOCS-3 in the MBH during LD compared with SD fails to fully explain these effects.

Different Transcription Profiles of &amp;lt;italic&amp;gt;SOCS-3&amp;lt;/italic&amp;gt;, &amp;lt;italic&amp;gt;ob&amp;lt;/italic&amp;gt; and &amp;lt;italic&amp;gt;IGF-I&amp;lt;/italic&amp;gt; Genes and their Possible Correlations in Obese and Lean Pigs

Pig breeds have significant differences in fat deposition and muscle development ability. However, the molecular mechanism behind these differences is still unknown. In this study, the expression patterns of three candidate genes, suppressor of cytokine signaling 3 (SOCS-3), obesity (ob) and insulin-like growth factor I (IGF-I), which are involved in adipose metabolism or muscle development, were analyzed. Total RNA was extracted from dorsal subcutaneous adipose tissue and longissimus of 8-month-old Bamei and Largewhite pigs. Semiquantitative reverse transcription-polymerase chain reaction was used to determine the expression levels of the SOCS-3 and ob genes in adipose tissue, and SOCS-3 and IGF-I genes in muscle tissue. The results showed that in adipose tissue the expression level of SOCS-3 was significantly higher in Bamei (obese) pigs than that in Largewhite (lean) pigs (P<0.01). However, in muscle tissue it was significantly lower in Bamei than that in Largewhite pigs (P<0.01). Furthermore, the expression of SOCS-3 was positively correlated to that of ob in adipose tissue and that of IGF-I in muscle tissue. These findings suggest that the difference in SOCS-3 gene expression levels in adipose and muscle tissues, the relationship between SOCS-3 and ob in adipose tissue, and that between SOCS-3 and IGF-I in muscle tissue, might contribute to the different fat deposition and muscle development ability between obese and lean pigs.

1141635071 Suppressor of Cytokine Signalling 3 (SOCS3) controls trophoblast invasion and proliferation

Introduction: Signal Transducer and Activator of Transcription 3 (STAT3) is an intracellular mediator of trophoblast invasion. In other cells, Suppressor Of Cytokine Signalling 3 (SOCS3) is known to be a major antagonist of STAT3 activation. For auto‐regulation and to avoid excessive signals, SOCS3 gene expression is partly induced by STAT3.Methods: The human choriocarcinoma cell line JEG‐3 was used for all experiments. STAT3 expression and STAT3 tyrosine (705) phosphorylation were analyzed by polyacrylamid gele electrophoresis followed by Western blotting. Interleukin‐6 (IL‐6) or Leukemia Inhibitory Factor (LIF) were used for stimulation. SOCS3 expression was silenced by RNA interference. For this approach in a first step, small interfering RNA (siRNA) was used. Subsequently, Jeg‐3 cells were transfected with genes for the respective, self‐designed small hairpin RNA (shRNA), neomycin resistance and green fluorescent protein. Application of neomycin and cell sorting by green fluorescence were used for elimination of non‐transfected cells. Non‐genomic siRNA and shRNA were respectively used for controls. Colorimetric proliferation assays were performed.Results: In IL‐6 stimulated cells, SOCS3 knockdown leads to increased STAT3 tyrosine phosphorylation. After LIF stimulation, proliferation in SOCS3 knockdown cells is significantly higher than in controls. STAT3 expression is not affected by SOCS3 knockdown.Conclusion: SOCS3 plays a major role in intracellular regulation of trophoblast proliferation and invasiveness. The balance of STAT3 and SOCS3 may tune the behavior of invading trophoblast cells.