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1.
Zhongguo Xue Xi Chong Bing Fang Zhi Za Zhi ; 28(5): 541-544, 2016 Sep 05.
Artigo em Chinês | MEDLINE | ID: mdl-29469488

RESUMO

OBJECTIVE: To clone and express the basement membrane specific heparan sulfate proteoglycan core protein (H3), and to evaluate its effect in detection of human cystic echinococcosis (CE). METHODS: The H3 gene immunoscreened from the cDNA library was cloned into pGEX-4T vector. The recombinant plasmid pGEX-3X-AgB8/3 was transformed into Escherichia coli BL21 strains and induced by isopropyl-ß-D-thiogalactopyranoside (IPTG). Then the expressed recombinant protein was purified by affinity chromatography and its effect in the detection of CE was evaluated by ELISA. Meanwhile, the effects of H3 and two antigens that the research group prepared before (purified HCF and rAgB8/2) in CE detection were compared. RESULTS: The plasmid pGEX-4T-H3 was successfully constructed and H3 was successfully expressed in prokaryotic cells. The sensitivities of the recombinant H3, purified HCF and rAgB8/2 in CE detection were 84.0% (68/81), 90.1% (73/81) and 77.8% (63/81) respectively, and there was no statistical difference among them (χ2 = 4.58, P > 0.05). The cross reactions of recombinant H3 with the sera of the patients with CE, cysticercosis and schistosomiasis were 63.3% (19/30), 16.7% (5/30) and 5.0% (1/20) respectively, and the cross reaction was 0 with the sera of healthy people. The specificities of recombinant H3, purified HCF, and rAgB8/2 were 80.8% (105/130), 71.5% (93/130) and 82.3% (107/130) respectively, and there were no statistical difference among them (χ2 = 5.71, P > 0.05). CONCLUSIONS: The recombinant H3 is a potential diagnostic antigen for CE detecting.


Assuntos
Antígenos de Helmintos/imunologia , Equinococose/diagnóstico , Echinococcus granulosus/genética , Sindecana-2/imunologia , Animais , Antígenos de Helmintos/genética , Membrana Basal , Clonagem Molecular , Reações Cruzadas , Ensaio de Imunoadsorção Enzimática , Humanos , Proteínas Recombinantes/imunologia , Sindecana-2/genética
2.
Immunology ; 137(3): 214-25, 2012 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-22881146

RESUMO

T cells express the heparan sulphate proteoglycans syndecan-2 and syndecan-4. Syndecan-4 plays a T-cell inhibitory role; however, the function of syndecan-2 is unknown. In an attempt to examine this function, syndecan-2 was expressed constitutively in Jurkat T cells. Interestingly, the expression of syndecan-2 decreased the surface levels of T-cell receptor (TCR)/CD3 complex, concomitant with intracellular retention of CD3ε and partial degradation of the TCR-ζ chain. Immunofluorescence microscopy revealed that intracellular CD3ε co-located with Rab-4 endosomes. However, the intracellular pool of CD3ε did not recycle to the cell surface. The lower TCR/CD3 surface levels caused by syndecan-2 led to reduced TCR/CD3 responsiveness. We show that the cytosolic PDZ-binding domain of syndecan-2 is not necessary to elicit TCR/CD3 down-regulation. These results identify a previously unrecognized means of controlling surface TCR/CD3 expression by syndecan-2.


Assuntos
Complexo CD3/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Sindecana-2/imunologia , Regulação para Baixo , Humanos , Células Jurkat
3.
J Biol Chem ; 286(22): 19541-8, 2011 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-21482826

RESUMO

HIV-1 p17 contains C- and N-terminal sequences with positively charged residues and a consensus cluster for heparin binding. We have previously demonstrated by affinity chromatography that HIV-1 p17 binds strongly to heparin-agarose at physiological pH and to human activated CD4(+) T cells. In this study we demonstrated that the viral protein binds to heparan sulfate side chains of syndecan-2, syndecan-4, and CD44v3 purified from HeLa cells and that these heparan sulfate proteoglycans (HSPGs) co-localize with HIV-1 p17 on activated human CD4(+) T cells by confocal fluorescence analysis. Moreover, we observed a stimulatory or inhibitory activity when CD4(+) T cells were activated with mitogens together with nanomolar or micromolar concentrations of the matrix protein.


Assuntos
Linfócitos T CD4-Positivos/metabolismo , Antígenos HIV/metabolismo , HIV-1/metabolismo , Heparitina Sulfato/metabolismo , Receptores de Hialuronatos/metabolismo , Interleucina-2/biossíntese , Sindecana-2/metabolismo , Sindecana-4/metabolismo , Fator de Necrose Tumoral alfa/biossíntese , Produtos do Gene gag do Vírus da Imunodeficiência Humana/metabolismo , Linfócitos T CD4-Positivos/imunologia , Feminino , Antígenos HIV/genética , Antígenos HIV/imunologia , HIV-1/genética , HIV-1/imunologia , Células HeLa , Heparitina Sulfato/genética , Heparitina Sulfato/imunologia , Humanos , Receptores de Hialuronatos/genética , Receptores de Hialuronatos/imunologia , Interleucina-2/genética , Interleucina-2/imunologia , Ativação Linfocitária , Masculino , Ligação Proteica , Sindecana-2/genética , Sindecana-2/imunologia , Sindecana-4/genética , Sindecana-4/imunologia , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia , Produtos do Gene gag do Vírus da Imunodeficiência Humana/genética , Produtos do Gene gag do Vírus da Imunodeficiência Humana/imunologia
4.
Mol Immunol ; 45(10): 2905-19, 2008 May.
Artigo em Inglês | MEDLINE | ID: mdl-18342939

RESUMO

Syndecans bind to cell adhesion molecules, growth factors and cytokines, and can act as coreceptors, and in this way modulate leukocyte cell function. Here, expression of the syndecans on primary human CD4 T cells was examined. Cell stimulation dramatically increased the amount of syndecan-4, and in a lower extent that of syndecan-2. Expression of syndecan-2 and -4 show different induction kinetics. Whereas syndecan-4 expression is fast and significant, that of syndecan-2 is more delayed and short-lived decreasing its mRNA expression at day 4. Both CD45RA+ naive and CD45RA- memory CD4 T cells express syndecan-2 and -4 upon activation. When incubated with human peripheral blood lymphocytes in a mixed leukocyte reaction, anti-syndecan-4 but not anti-syndecan-2 antibodies, decreased T cell proliferation. However, cross-linking of cell-bound syndecan-2 or syndecan-4 via immobilized antibodies blocked proliferation and decreased TNF production of T cells in the presence of optimal levels of anti-CD3. These findings suggest that syndecan-2 and -4 act as inhibitors of T cell activation. We also investigated the role that MAPK signalling pathways play in control of syndecan expression in T cells. We show that production of syndecan-2 but not syndecan-4 requires signaling via p38 MAP kinase alpha/beta in T CD4 cells. As mechanisms that confer syndecan-2 expression are unknown, we analyse the chromatin hypersensitivity of syndecan-2 promoter proximal region in Jurkat T cells and endothelial cells. The analysis reveals a chromatin accessible site in the +3.5kb intronic region, concomitant with a region showing high evolutionary conservation. We isolate and analyse 5'-flanking regions of human syndecan-2 gene, by transfection assays. The +3.5kb hypersensitive site in the intronic region demonstrates basal promoter activity in Jurkat. This study provides evidence for the up-regulation of syndecan-2 and -4 in human primary CD4 T cells during in vitro activation and suggest an inhibitory role for these syndecans in CD4 T cells.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Ativação Linfocitária/imunologia , Sindecana-2/imunologia , Sindecana-4/imunologia , Complexo CD3/imunologia , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/enzimologia , Células Cultivadas , Cromatina/química , Desoxirribonuclease I/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/antagonistas & inibidores , Humanos , Memória Imunológica/efeitos dos fármacos , Íntrons/genética , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Células Jurkat , Ativação Linfocitária/efeitos dos fármacos , Teste de Cultura Mista de Linfócitos , Regiões Promotoras Genéticas/genética , Inibidores de Proteínas Quinases/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Sindecana-2/genética , Sindecana-4/genética , Regulação para Cima/efeitos dos fármacos , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores
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