Biomedical subjects

Tong Shi

Publications and source records attributed to Tong Shi.

11 recordsLinked to original sources

PubMed2006

Neddylation of a breast cancer-associated protein recruits a class III histone deacetylase that represses NFkappaB-dependent transcription.

Neddylation has an important role in ubiquitin-mediated protein degradation through modification of cullins, which are the main substrates for NEDD8 modification. Here, we show that breast cancer-associated protein 3 (BCA3) is a NEDD8 substrate. BCA3 suppressed NFkappaB-dependent transcription through its ability to bind to p65 and the cyclin D1 promoter in a neddylation-dependent manner. Transcriptional suppression mediated by BCA3 may be attributed to the ability of neddylated BCA3 to recruit SIRT1, a class III histone deacetylase. Silencing of endogenous BCA3 in DU145 and MCF7 cells enhanced NFkappaB transcription and inhibited tumour necrosis factor (TNF)alpha-induced apoptosis. Conversely, BCA3 silencing could be reversed by over-expression of wild-type BCA3 and SENP8, a NEDD8-specific protease, but not by neddylation-deficient BCA3 or a SENP8 mutant. These results provide a crucial link between neddylation and transcriptional regulation by SIRT1, a NAD-dependent histone deacetylase that prolongs life span in yeast and worms.

Adaptor Proteins, Signal Transducing↗

PubMed2006

Alpha-4 integrins and VCAM-1, but not MAdCAM-1, are essential for recruitment of mast cell progenitors to the inflamed lung.

Normal mouse lungs lack appreciable numbers of mast cells (MCs) or MC progenitors (MCp's), yet the appearance of mature MCs in the tracheobronchial epithelial surface is a characteristic of allergic, T-cell-dependent pulmonary inflammation. We hypothesized that pulmonary inflammation would recruit MCp's to inflamed lungs and that this recruitment would be regulated by distinct adhesion pathways. Ovalbumin-sensitized and challenged mice had a greater than 28-fold increase in the number of MCp's in the lungs. In mice lacking endothelial vascular cell adhesion molecule 1 (VCAM-1) and in wild-type mice administered blocking monoclonal antibody (mAb) to VCAM-1 but not to mucosal addressin CAM-1 (MadCAM-1), recruitment of MCp's to the inflamed lung was reduced by greater than 75%. Analysis of the integrin receptors for VCAM-1 showed that in beta7 integrin-deficient mice, recruitment was reduced 73% relative to wild-type controls, and in either BALB/c or C57BL/6 mice, mAb blocking of alpha4, beta1, or beta7 integrins inhibited the recruitment of MCp's to the inflamed lung. Thus, VCAM-1 interactions with both alpha4beta1 and alpha4beta7 integrins are essential for the recruitment and expansion of the MCp populations in the lung during antigen-induced pulmonary inflammation. Furthermore, the MCp is currently unique among inflammatory cells in its partial dependence on alpha4beta7 integrins for lung recruitment.

Animals↗

PubMed2006

Anti-beta2-glycoprotein I antibodies in complex with beta2-glycoprotein I can activate platelets in a dysregulated manner via glycoprotein Ib-IX-V.

OBJECTIVE: Results of previous studies suggest that anti-beta2-glycoprotein I (anti-beta2GPI) antibodies in complex with beta2GPI activate platelets in a dysregulated manner, potentially contributing to the prothrombotic tendency associated with the antiphospholipid syndrome (APS). We undertook this study to investigate the possible contribution of the GPIb-IX-V receptor to platelet activation mediated by the anti-beta2GPI antibody-beta2GPI complex. METHODS: In vitro methods were used in the present study. The interaction between beta2GPI and the GPIbalpha subunit of the GPIb-IX-V receptor was delineated using direct binding and competitive inhibition assays. The interaction between the anti-beta2GPI antibody-beta2GPI complex and platelets was studied using a novel method in which the Fc portion of the antibody was immobilized using protein A coated onto a microtiter plate. Platelet activation was assessed by two methods; one involved measuring thromboxane B2 production and the other involved assessment of the activation of the phosphatidylinositol 3-kinase/Akt/glycogen synthase kinase 3beta intracellular signaling pathway. The contribution of the GPIbalpha receptor to platelet activation induced by the anti-beta2GPI antibody-beta2GPI complex was assessed by observing the influence of 2 anti-GPIbalpha antibodies (AK2 and SZ2) directed against distinct epitopes. RESULTS: This study showed that beta(2)GPI could bind to the GPIbalpha receptor. The anti-beta2GPI antibody-beta2GPI complex was able to activate platelets, and this effect was inhibited by anti-GPIbalpha antibody directed against epitope Leu-36-Gln-59, but not by anti-GPIbalpha antibody directed against residues Tyr-276-Glu-282. CONCLUSION: Our findings show that inappropriate platelet activation by the anti-beta2GPI antibody-beta2GPI complex via the GPIbalpha receptor may contribute to the prothrombotic tendency associated with APS.

Antibodies, Blocking↗

PubMed2005

SIRT3, a mitochondrial sirtuin deacetylase, regulates mitochondrial function and thermogenesis in brown adipocytes.

SIRT3 is one of the seven mammalian sirtuin homologs of the yeast Sir2 gene, which mediates the effect of caloric restriction on life span extension in yeast and Caenorhabditis elegans. Because adipose tissue is essential in energy homeostasis and also plays a role in life span determination, we decided to investigate the function of sirtuin members in fat. We report here that murine SIRT3 is expressed in brown adipose tissue and is localized on the mitochondria inner membrane. Caloric restriction activates SIRT3 expression in both white and brown adipose. Additionally, cold exposure up-regulates SIRT3 expression in brown fat, whereas elevated climate temperature reduces the expression. Enforced expression of SIRT3 in the HIB1B brown adipocytes enhances the expression of the uncoupling protein PGC-1alpha, UCP1, and a series of mitochondria-related genes. Both ADP-ribosyltransferase and deacetylase activities of SIRT3 are required for this action. Furthermore, the SIRT3 deacetylase mutant exhibits a dominant negative effect by inhibiting UCP1 expression. This inhibitive effect can be abolished by the coexpression of PGC-1alpha, indicating a major role of PGC-1alpha in the SIRT3 action. In addition, SIRT3 stimulates CREB phosphorylation, which reportedly activates PGC-1alpha promoter directly. Functionally, sustained expression of SIRT3 decreases membrane potential and reactive oxygen species production while increasing cellular respiration. Finally, SIRT3, along with genes related to mitochondrial function, is down-regulated in the brown adipose tissue of several genetically obese mice. In summary, our results demonstrate that SIRT3 activates mitochondria functions and plays an important role in adaptive thermogenesis in brown adipose.

ADP Ribose Transferases↗

PubMed2005

Intranasal CpG-oligodeoxynucleotide is a potent adjuvant of vaccine against Helicobacter pylori, and T helper 1 type response and interferon-gamma correlate with the protection.

BACKGROUND: Although a series of vaccines against Helicobacter pylori have emerged in the past 10 years, the mechanism involved in their protective effect is yet to be elucidated, and more effective vaccine adjuvants remain to be developed. In this study, CpG-oligodeoxynucleotide (CpG-ODN) was investigated as a new candidate for a H. pylori vaccine adjuvant. Furthermore, the role of T helper 1 (Th1) type response and interferon (IFN)-gamma in the protective immunity was explored. METHODS: C57BL/6 mice and IFN-gamma knockout mice were intranasally or orally immunized with H. pylori whole cell sonicate (WCS)/CpG-ODN and challenged with different doses [5 x 10(8) and 5 x 10(6) colony-forming units (CFU)] of H. pylori. The protective effect was assessed as the percentage of noninfected mice. The responsive antibodies and cytokines were analyzed using an enzyme-linked immunosorbent assay (ELISA) and flow cytometry. RESULTS: The prevention rates against H. pylori infection in mice intranasally immunized with WCS plus CpG-ODN were dramatically higher than those in sham-immunized mice (70% vs. 0%, challenged with 5 x 10(8) CFU H. pylori; 90% vs. 20%, challenged with 5 x 10(6) CFU H. pylori). Significantly higher levels of immunoglobulin G2a (IgG2a) and IFN-gamma were detected in the mice immunized with WCS/CpG than in sham-immunized controls. However, vaccination failed to effectively protect IFN-gamma knockout mice challenged with H. pylori. CONCLUSIONS: CpG-ODN given intranasally is a potent adjuvant for development of a H. pylori vaccine. Th1-type response and IFN-gamma are involved in the protection.

Adjuvants, Immunologic↗

PubMed2004

Domain V of beta2-glycoprotein I binds factor XI/XIa and is cleaved at Lys317-Thr318.

The fifth domain (DV) of beta2-glycoprotein I (beta2GPI) is important for binding a number of ligands including phospholipids and factor XI (FXI). Beta2GPI is proteolytically cleaved in DV by plasmin but not by thrombin, VIIa, tissue plasminogen activator, or uPA. Following proteolytic cleavage of DV by plasmin, beta2GPI retains binding to FXI but not to phospholipids. Native beta2GPI, but not cleaved beta2GPI, inhibits activation of FXI by thrombin and factor XIIa, attenuating a positive feedback mechanism for additional thrombin generation. In this report, we have defined the FXI/FXIa binding site on beta2GPI using site-directed mutagenesis. We show that the positively charged residues Lys284, Lys286, and Lys287 in DV are essential for the interaction of beta2GPI with FXI/FXIa. We also demonstrate that FXIa proteolytically cleaves beta2GPI at Lys317-Thr318 in DV. Thus, FXIa cleavage of beta2GPI in vivo during thrombus formation may accelerate FXI activation by decreasing the inhibitory effect of beta2GPI.

Binding Sites↗

PubMed2004

Effect of beta2-glycoprotein I null mutation on reproductive outcome and antiphospholipid antibody-mediated pregnancy pathology in mice.

beta(2)-Glycoprotein I (beta(2)GPI) is a principal target antigen for antiphospholipid antibodies associated with recurrent pregnancy loss and fetal growth restriction in women. The significance of disrupted beta(2)GPI activity in contributing to pregnancy pathology in antiphospholipid syndrome (APS) is not clear. In this study the physiological requirement for functional beta(2)GPI in pregnancy was investigated by evaluating reproductive outcomes in beta(2)GPI null mutant (beta(2)GPI-/-) mice. beta(2)GPI-/- mice were fertile and carried viable fetuses to term. However, there was an 18% reduction in the number of viable implantation sites in beta(2)GPI-/- mice and reduced fetal weight and fetal:placental weight ratio in late gestation, suggesting compromised placental function. Placental architecture was altered in beta(2)GPI-/- implantation sites with a 24% increase in the junctional zone: labyrinthine ratio, but placentae showed no evidence of increased thrombosis in the absence of beta(2)GPI. The effect of beta(2)GPI genotype on pregnancy success after passive transfer of human and mouse antibodies reactive with beta(2)GPI was also explored. Two of five anti-beta(2)GPI antibodies induced pregnancy loss in beta(2)GPI+/+ mice but beta(2)GPI-/- mice were refractory to antibody-induced pregnancy failure. We conclude that functional beta(2)GPI is not essential for successful pregnancy in mice, but optimal placental development and fetal growth require this molecule. Together these data are consistent with pathogenic mechanisms in antiphospholipid syndrome involving both neutralization of beta(2)GPI function and beta(2)GPI-immunoglobulin complex formation.

Animals↗

PubMed2004

Beta 2-Glycoprotein I binds factor XI and inhibits its activation by thrombin and factor XIIa: loss of inhibition by clipped beta 2-glycoprotein I.

Activation of factor XI (FXI) by thrombin in vivo plays a role in coagulation by providing an important positive feedback mechanism for additional thrombin generation. FXI is activated in vitro by thrombin, or FXIIa in the presence of dextran sulfate. In this report, we investigated the effect of beta(2)-glycoprotein I (beta(2)GPI) on the activation of FXI. beta(2)GPI bound FXI in vitro and inhibited its activation to FXIa by thrombin and FXIIa. The affinity of the interaction between beta(2)GPI and FXI was equivalent to the interaction between FXI and high molecular weight kininogen. Inhibition of FXI activation occurred with lower concentrations of beta(2)GPI than found in human plasma. Proteolytic clipping of beta(2)GPI by plasmin abolished its inhibition of FXI activation. The results suggest a mechanism of regulation whereby physiological concentrations of beta(2)GPI may attenuate thrombin generation in vivo by inhibition of FXI activation. Plasmin cleavage of beta(2)GPI provides a negative feedback that counteracts its inhibition of FXI activation.

Antiphospholipid Syndrome↗

PubMed2003

[The role of Th1-type response and gamma-interferon in the protection induced by Helicobacter pylori vaccine].

OBJECTIVE: To determine whether CpG-oligodeoxynucleotide (CpG-ODN) can be an adjuvant for the development of a Helicobacter pylori (H. pylori) vaccine, and explore the role of Th1-type response and gamma-interferon (gamma-IFN) in protection induced by vaccine. METHODS: Mice were intranasally or orally immunized with H. pylori whole cell sonicate (WCS) and CpG-ODN and challenged with 5 x 10(6) or 5 x 10(8) organisms. The mice were killed 2 and 8 weeks later respectively, the sera, saliva, gastric juice were collected to measure the concentrations of IgG, IgG1, IgG2a and IgA by ELISA, their spleen cells were isolated for analysis of intracellular gamma-IFN and interleukin-4 (IL-4) by flow cytometry. RESULTS: The prevention rate against H. pylori infection was 70% in mice intranasally immunized with WCS plus CpG-ODN and challenged with 5 x 10(8) CFU H. pylori, and 90% in those challenged with 5 x 10(6) CFU H. pylori. Significantly higher levels of IgG1 and IFN-gamma were found in the mice immunized with WCS/CpG than those in sham-immunized controls (P < 0.05). Spontaneous elimination of H. pylori infection occurred in two of ten sham-immunized mice, whose IgG1 and IFN-gamma levels were higher than those of other sham-immunized ones. CONCLUSION: CpG-Oligodeoxynucleotide is a potent adjuvant of vaccine against H. pylori, and Th1-type response and gamma-IFN are highly associated with the protective effect.

Animals↗

PubMed2003

[The role of nuclear factor kappa B in secretion of interleukin-8 by gastric cancer cell line SGC 7901 induced by Helicobacter pyiori].

OBJECTIVE: To investigate the role of nuclear factor kappaB (NF-kappaB) in the gastric inflammation induced by Helicobacter pylori (H. pylori). METHODS: SGC-7901 was transfected with IkappaB (NF-kappaB inhibitor gene) by electroporation, (beta)lacZ activity assay was used to examine transfected efficacy. Expression of IkappaB was assessed by Western-blot. Different concentration of live and heat-killed Hp (ATCC 43504) and supernatant of liquid culture were cocultured with SGC7901-IkappaB and its negative control SGC7901- neo. Activation of intracellular NF-kappaB was examined by electrophoretic mobility shift analyses (EMSA) and luciferase report gene assay at different time point. IL-8 levels were measured by ELISA at different time point. RESULTS: IL-8 release was evident 4 hours after infection of SGC-7901-neo with H. pylori, and this effect was dose-time dependent. SGC-7901-IkappaB in which NF-kappaB has not been activated could not secret IL-8 after infection with H. pylori. CONCLUSION: Secretion of IL-8 by gastric epithelial cell upon H pylori infection is dependent on activation of NF-kappaB.

Cell Line, Tumor↗

PubMed2002

[Relation between Beta-2-glycoprotein I and hepatitis B virus surface antigen].

OBJECTIVE: To clarify the binding character between Beta-2-glycoprotein I (Beta-2-GPI) and HBsAg. METHODS: Beta-2-GPI was purified from human plasma and labelled with biotin. Solid phase enzyme linked absorbance assay was used to investigate its binding with HBsAg. RESULTS: Biotinylated Beta-2-GPI was found to bind HBsAg and the reaction could be inhibited by excess unlabelled Beta-2-GPI. CONCLUSIONS: Beta-2-GPI may play a role in hepatitis B virus infection.

Binding Sites↗