Suppressor of cytokine signaling(SOCS) 3, a novel type of cytokine signal transduction inhibitory molecules in family of SOCS, is mainly involved in Janus protein tyrosine kinase/signal transducer andantivator of transcription signaling pathway negative feedback regulation. It is involved in inflammation, oxidative stress, cell damage, and apoptosis. Meanwhile, it is closely related to atherosclerosis, obesity, glucose metabolism, insulin resistance, leptin, cancer, asthma, and rheumatic diseases. Therefore, SOCS-3 may become a therapeutic target of these diseases.
Humans ; Suppressor of Cytokine Signaling 3 Protein ; Suppressor of Cytokine Signaling Proteins

Suppressor of cytokine signaling (SOCS) is a protein family negatively regulating signal transduction pathway of a certain class of cytokines and growth factors. More than 20 members have been found in SOCS family, SOCS3 is one of them and has been studied hottest and most clearly. Recent studies demonstrated that SOCS3 abnormalities were found in patients with myeloproliferative neoplasms (MPN), suggesting that SOCS3 plays a significant role in the pathogenesis, development and metastasis in MPN. In this review, the advances of research on relationship between SOCS3 and MPN were summarized, including general profile of SOCS family; structure, function and regulation of SOCS3, relation of SOCS3 to MPN and so on.
Cytokines ; metabolism ; Humans ; Myeloproliferative Disorders ; metabolism ; Signal Transduction ; Suppressor of Cytokine Signaling 3 Protein ; Suppressor of Cytokine Signaling Proteins ; metabolism

OBJECTIVETo prepare and characterize the monoclonal antibody (mAb) against human SOCS3.

METHODSBALB/c mice were immunized with recombinant GST-SOCS3 protein, from which the spleen cells were isolated and fused with Sp2/0 cells. After several rounds of screening and cloning, the hybridoma cell strain secreting anti-SOCS3 mAb was obtained, whose specificity was evaluated using ELISA and Western blotting, and the titer, immunoglobulin subtype and affinity of the mAb were also measured.

RESULTSThe hybridoma cell strain secreting anti-SOCS3 mAb was identified to belong to IgG2a subtype. The mAb titers in cultural supernatant and acetic fluid were 1:640 and 1:25600, respectively, as determined by ELISA with affinity reaching 4.84x10(6) L/mol.

CONCLUSIONThe success in anti-SOCS3 mAb preparation provides the basis for further study of the negative regulation of cytokine signal transduction and the immunoregulation in microorganism infections.

Animals ; Antibodies, Monoclonal ; biosynthesis ; Humans ; Hybridomas ; secretion ; Mice ; Mice, Inbred BALB C ; Recombinant Fusion Proteins ; Suppressor of Cytokine Signaling 3 Protein ; Suppressor of Cytokine Signaling Proteins ; immunology

This study was purposed to investigate the effect of mutation and single nucleotide polymorphism (SNP) of suppressor of cytokine signaling (SOCS) on the typical myeloproliferative neoplasms (MPN) and its mechanism. The mutation and SNP of SOCS1, SOCS2, SOCS3 genes in 100 MPN patients were detected by RT-PCR and direct sequencing. The results showed that among 100 cases there were 21 cases with A→C polymorphism in the 63th site nucleotide of the 15 SOCS3 exon (SNP library no reported), 18 cases with A→C polymorphism in the 1779th site nucleotide of the 15 SOCS3 exon, 49 cases with A→G polymorphism in the 2249th site nucleotide of the 15 SOCS3 exon (SNP library no reported), 39 cases with T→C polymorphism in the 2366th site nucleotide of the 15 SOCS3 exon (SNP library no reported), 9 cases with T→C polymorphism in the exon of 15 SOCS2 gene (SNP library no reported). SOCS3 SNP was found in patients with significantly advanced age at diagnosis, the leukocyte count and platelet level were higher than those in patients with wild type, JAK2V617 mutations was found in 87.65% SOCS3 SNP. It is concluded that the SOCS may be an important target for anticancer therapy, the single nucleotide polymorphism of SOCS may involve to pathogenesis of MPN.
Adolescent ; Adult ; Aged ; Aged, 80 and over ; Exons ; Female ; Humans ; Male ; Middle Aged ; Mutation ; Myeloproliferative Disorders ; genetics ; Polymorphism, Single Nucleotide ; Suppressor of Cytokine Signaling 1 Protein ; Suppressor of Cytokine Signaling 3 Protein ; Suppressor of Cytokine Signaling Proteins ; genetics ; Young Adult

The study was aimed to investigate the effect of IL-2 on the acquired immune response of naive T cells, and to explore the influence of IL-2 on suppressor of cytokine signaling 3 (SOCS-3) expression of naive T cells, as well as to elucidate the role of SOCS-3 on antigen specific immune response in vitro. Naive DO11.10 T cells were pre-stimulated with IL-2 (50 U/ml), and stimulated with OVA(323 - 329) antigen after removing IL-2, then the proliferation of naive DO11.10 T cells was detected. Naive DO11.10 T cells were stimulated with IL-2 (50 U/ml), and SOCS-3 expression was detected by real-time PCR. Naive DO11.10 T cells were stimulated with OVA(323 - 329) antigen, and SOCS-3 expression was detected by means of (3)H-TdR. The results showed that after IL-2 pre-stimulation, the proliferation of naive DO11.10 T cells decreased significantly when stimulated with OVA(323 - 329) antigen; SOCS-3 expression of naive DO11.10 T cells was up-regulated significantly after IL-2 stimulation, the up-regulation began obviously at 4 hours and reached peak at 6 hours. SOCS-3 expression on naive DO11.10 T cells was down-regulated markedly after OVA(323 - 329) antigen stimulation, the expression level of SOCS-3 was lowest on day 2 and returned to normal on day 4 after stimulation. It is concluded that the antigen specific immune response of naive DO11.10 T cells is inhibited after pre-stimulation with IL-2, which may be mediated by SOCS-3.
Animals ; Gene Expression ; Humans ; Interleukin-2 ; immunology ; pharmacology ; Mice ; Suppressor of Cytokine Signaling 3 Protein ; Suppressor of Cytokine Signaling Proteins ; immunology ; metabolism ; T-Lymphocytes ; immunology

OBJECTIVETo study the mechanism underlying the effect of the SOCS3 rs4969170 A/G alleles on the occurrence of insulin resistance (IR) in patients with chronic hepatitis C.

METHODSThe promoter region of the SOCS3 gene was amplified by PCR,and luciferase expression vectors were constructed and transfected into HepG2,Huh7 cell lines.The relative luciferase activity of each expression vector was assessed by the dual luciferase reporter gene assay system.Western blotting was used to detect SOCS3 protein expression in PBMCs from groups of patients with the rs4969170 AA and AG genotypes.The state of IR in eight patients was evaluated by determining their HOMA-IR.

RESULTSThe pGL3-A, PGL3-G and pGL3-control vectors showed significantly different luciferase expression in the HepG2 cells (0.121 00 ± 0.022 07,0.027 00+/-0.012 49 and 0.043 33 ± 0.005 51; F =48.068, P=0.001) and in the Huh7 cell lines (0.164 70 ± 0.007 10,0.027 33 ± 0.017 04 and 0.033 67 ± 0.014 98; F =115.137, P=0.001). The expression of SOCS3 protein was significantly higher in the rs4969170 AA genotype group than in the AG genotype group (1.22 ± 0.40 vs. 0.30 ± 0.19; t =4.149, P=0.006).The IR index of patients with the rs4969170 AA genotype and the AG genotype was 4.11 ± 2.62 and 1.47 ± 1.01 respectively.There were three patients with IR in the rs4969170 AA genotype group and one in the rs4969170 AG group. There was no statistically significant difference between the two genotype groups (t=1.881, P=0.109).

CONCLUSIONSThe SOCS3 rs4969170 A haplotype may enhance transcriptional activity of the gene promoter to regulate gene expression, thereby increasing intracellular SOCS3 protein level and ultimately interfering with insulin signaling and causing IR in patients with chronic hepatitis C.

Cell Line, Tumor ; Genes, Reporter ; Genotype ; Haplotypes ; Hepatitis C, Chronic ; Humans ; Insulin Resistance ; Luciferases ; Polymorphism, Single Nucleotide ; Suppressor of Cytokine Signaling 3 Protein ; Suppressor of Cytokine Signaling Proteins

Suppressor of cytokine signaling (SOCS) is a new family of proteins produced in cells. It may play an important role in classic negative feedback loop to regulate cytokine signal transduction. SOCS-1 was observed and confirmed firstly. Expression of SOCS-1 can inhibit cytokine signal transduction of some cytokines, such as IL-6, LIF, OSM, INF-gamma, GH, and so on, many immune responses are regulated by them in vivo. Abnormal expression of SOCS-1 is closely related to some human diseases. It plays an important role in the development of leukemia, rheumatoid arthritis, liver cirrhosis and liver cancer. In this review, the advances of research on the relationship between SOCS-1 and cytokine, and its correlation with some diseases were summarized.
DNA Methylation ; Humans ; Interleukin-6 ; biosynthesis ; Intracellular Signaling Peptides and Proteins ; Leukemia ; etiology ; genetics ; Leukemia Inhibitory Factor ; biosynthesis ; Suppressor of Cytokine Signaling 1 Protein ; Suppressor of Cytokine Signaling Proteins ; biosynthesis ; genetics ; physiology

In order to construct recombinant adenovirus vector expressing Suppressor of cytokine signaling 3 (SOCS3) and obtain infectious adenoviral particles, SOCS3 gene was amplified from plasmid pcDNA3-SOCS3 and subcloned into the adenovirus shuttle plasmid pAdTrack-CMV. After sequence confirmation, the recombinant shuttle plasmid pAdTrack-CMV-SOCS3 was linearized by Pme I, and then transformed into BJ5183 competent cell, the recombinant plasmid pAd-SOCS3 was obtained by homologous recombination between pAdTrack-CMV-SOCS3 and the adenoviral backbone plasmid pAdEasy-1 in BJ5183. The pAd-SOCS3 was linearized by Pac I and transfected into HEK293 cells via liposome. The recombinant adenovirus was packaged and amplified in HEK293 cells. After purifying, virus titer was determined by tissue culture infectious dose 50 (TCID50). Using the recombinant adenoviruses to infect porcine primary adipocytes, the expression of green fluorescent protein (GFP) was observed by fluorescent microscopy, and SOCS3 gene was identified by RT-PCR and Western blotting. Restriction enzyme and PCR analysis demonstrated that the recombinant adenovirus vector was constructed correctly, and the virus titer reached 1.2x10(9) PFU/mL. The result of RT-PCR and Western blotting showed that SOCS3 mRNA and protein expression was remarkably increased in porcine primary adipocytes infected with recombinant adenovirus. In conclusion, this study successfully constructed the recombinant adenovirus containing SOCS3 gene, and can be helpful for further research on the function of SOCS3.
Adenoviridae ; genetics ; metabolism ; Adipocytes ; metabolism ; Animals ; Cloning, Molecular ; Green Fluorescent Proteins ; biosynthesis ; genetics ; HEK293 Cells ; Humans ; RNA, Messenger ; biosynthesis ; genetics ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; Suppressor of Cytokine Signaling 3 Protein ; Suppressor of Cytokine Signaling Proteins ; biosynthesis ; genetics ; Swine ; Transfection

Recombinant human granulocyte-colony-stimulating factor (rhG-CSF) can widely regulate human immunologic response. In the protocol of peripheral blood stem/progenitor cell mobilization, rhG-CSF can change the numbers and functions of T cells. Then the results can impact the incidence of graft-versus-host disease after allogeneic peripheral blood stem/progenitor cell transplantation. The regulation of rhG-CSF on T cell is an indirect action which is based on the direct action to monocytes and dendritic cells. The numerous IL-10 secreted by monocytes plays a key role in cytokines production, proliferative response and cytotoxicity of T cells. Endogenous IL-10 can induce high expression of SOCS3 and the SOCS3 is very important for regulating the signal transduction of the activities of T cells. In this review influences of rhG-CSF on T-cells in mobilization process and related mechanisms were elaborated with emphasis.
Blood Donors ; Granulocyte Colony-Stimulating Factor ; pharmacology ; therapeutic use ; Hematopoietic Stem Cell Mobilization ; methods ; Humans ; Interleukin-10 ; biosynthesis ; Recombinant Proteins ; Suppressor of Cytokine Signaling 3 Protein ; Suppressor of Cytokine Signaling Proteins ; biosynthesis ; T-Lymphocytes ; cytology ; drug effects ; metabolism

OBJECTIVETo investigate the effect of DNA methyhransferase l (DNMT1) gene silencing on methylation of suppressor of cytokine signaling (SOCS-1) in multiple myeloma RPMI 8226 cells.

METHODSRecombinant plasmid pshRNA-DNMTl was transfected into multiple myeloma RPMI 8226 cells by lipofectamine 2000. RT-PCR and Western blot were used to detect the mRNA and protein expression of DNMTl in RPMI 8226 cells respectively before and after transfection. Methylation-specific polymerase chain reaction was used to detect the methylation level change of SOCS-1 gene in RPMI8226 cells transfected.

RESULTSDNMTl targeted short hairpin RNA(shRNA) was successfully inserted into the plasmid vector pshRNA. RT-PCR results showed that the relative mRNA expression level of DNMTI gene in RPMI 8226 cells transfected with pshRNA was 0.176±0.004 which was significantly lower than that in cells transfected by empty vector (0.956±0.033, P<0.01). Western blot analysis showed that the relative expression level of DNMT1 protein of RPMI 8226 cells transfected by pshRNA was 0.065±0.014, which was significantly lower than that in transfected cells by empty vector(0.415±0.027) (P<0.05). These results indicated that the recombinant plasmid pshRNA could effectively knock down the expression of DNMT1 gene in RPMI 8226 cells. Methylation analysis showed that the methylation level of SOCS-1 gene was obviously reduced after transfection.

CONCLUSIONDNMT1 gene in RPMI 8226 cell can be silenced by shRNA. DNMT1 gene silencing can significantly induce SOCS-1 gene hypomethylation, which indicates that DNMT1 may play an important role in the process of SOCS-1 hypermethylation.

Cell Line, Tumor ; DNA Methylation ; Gene Silencing ; Genetic Vectors ; Humans ; Multiple Myeloma ; RNA, Messenger ; RNA, Small Interfering ; Repressor Proteins ; Suppressor of Cytokine Signaling 1 Protein ; Suppressor of Cytokine Signaling Proteins ; Transfection