Biomedical subjects

Li-huang Zhang

Publications and source records attributed to Li-huang Zhang.

4 recordsLinked to original sources

PubMed2005

[Construction of the eukaryotic expression vector containing rat transforming growth factor beta type II receptor and interferon gamma fusion genes].

OBJECTIVE: To express a bioactive fusion protein comprising rat soluble transforming growth factor beta type II receptor and interferon gamma(rsTGFbetaR II-IFN gamma) in mammalian cells. METHODS: mRNA was extracted from rat liver and the sTGFbetaR II-IFN gamma genes amplified by RT-PCR, then the two gene segments were cloned into the same pSecTag2A expression vector, and pSecTag2A/rsTGFbetaR II-IFN gamma recombinant plasmid was obtained, which was later transfected into CHO cells using liposomes. The expression of pSecTag2A/rsTGFbetaR II-IFN gamma in the supernatant was detected by ELISA and Western blotting. The bioactivities of the fusion protein were tested by sTGF betaR II-IFN gamma and IFN gamma bioassays. RESULTS: pSecTag2A/rsTGFbetaR II-IFN gamma transfectants expressed rsTGF betaR II-IFN gamma fusion protein. The purified fusion protein exhibited anti-viral activity and antagonized the proliferation-inhibitive effect of TGFbeta1 on CCL-64 cells. It inhibits the HSC activation in vitro. CONCLUSION: The pSecTag2A/rsTGFbetaR II-IFN gamma recombinant plasmid constructed in this study can express bioactive rsTGFbetaR II-IFN gamma fusion protein.

Amino Acid Sequence↗

PubMed2005

[Construction and expression of recombinant adenovirus containing human thymic stromal lymphopoietin gene].

AIM: To construct and express recombinant adenovirus bearing human thymic stromal lymphopoietin (TSLP) gene. METHODS: TSLP gene amplified from human fetal lung cells was first cloned into eukaryotic expression vector pcDNA3.1, and then subcloned into shuttle vector pShuttle. The resultant plasmid was subsequently cotransformed into E. coli BJ5183 cells with adenoviral backbone plasmid pAdEasy-1. The recombinant adenovirus plasmid containing TSLP was digested with Pac I and transfected into HEK293 cells to package recombinant adenovirus particles. The TSLP gene of the recombinant virus was detected by PCR, and its expression was analyzed by Western blot. RESULTS: Recombinant adenovirus vector bearing human TSLP gene was constructed by homologous recombination in E.coli, and recombinant adenovirus was obtained by transfecting HEK293 cells with this infectious DNA. PCR test indicated that TSLP gene was successfully integrated into the adenoviral genome, and the titer of the recombinant Ad reached 1 x 10(11) pfu/L. Meanwhile, expression of TSLP in the infected Hela cells was confirmed by Western blot. CONCLUSION: The construction of recombinant adenovirus bearing human TSLP gene and its expression mediated by this adenovirus pave a foundation for the study on the biological function of this novel cytokine.

Adenoviridae↗

PubMed2004

[Effect of Triptolide on TNFalpha-induced activation of NF-kappaB and expression of COX-2 and iNOS in human rheumatoid arthritis synovial fibroblasts].

OBJECTIVE: To explore the effects of Triptolide (TP) on TNFalpha-induced cell proliferation and expressions of cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS) and their inducing products PGE2, NO in human rheumatoid arthritis synovial fibroblasts (RASF). METHODS: Fibroblasts (RASF) were obtained from synovial tissue of patients with RA and were cultured in vitro. RASF were stimulated with TNFalpha(20 microg/L) in the presence or absence of TP(0 - 100 microg/L) for 20 h. The RASF proliferation was determined by (3)H-TdR incorporation, and the productions of PGE2 and NO in culture supernatants of RASF were detected with competitive ELISA and enzyme reduction of nitrate. Expressions of COX-2 and iNOS mRNA in RASF were analyzed by semi-quantitative RT-PCR. Expressions of COX-2 and iNOS protein were estimated by Western-blot method and cellular enzyme immunoassay in synovial fibroblasts. NF-kappaB activity in whole-cell extract of RASF was also measured by an ELISA-based method. RESULTS: TP (>20 microg/L) down-regulated markedly TNFalpha-induced COX-2 and iNOS mRNA and protein expression, and their inducing products PGE2 and NO of synovial fibroblasts. This effect was positively correlated with TP concentrations. NF-kappaB activity in TNFalpha-stimulated synovial cells was suppressed profoundly by TP treatment (IC(50) approximately 35microg/L). The activity of NF-kappaB was correlated with the levels of COX-2 and iNOS expression in TNFalpha-stimulated RASF. No change was observed in proliferation of synovial cells after treatment of TP. CONCLUSION: TP could significantly down-regulate TNFalpha-induced COX-2, iNOS expression and production of PGE2, NO in human RASF, which is associated with the suppression of NF-kappaB activity.

Arthritis, Rheumatoid↗

PubMed2004

[Construction and expression of the eukaryotic expression vector containing human soluble transforming growth factor beta receptor II].

OBJECTIVE: To construct a eukaryotic expression vector encoding the gene of extracellular region of type II transforming growth factor beta receptor (sTGFbetaR II), to express the protein in CHO cell line and to determine its biological activity. METHODS: The extracellular region (amino acids 1-159) of the human TGFbetaR II cDNA was amplified by PCR from a TGFbetaR II chimeric plasmid,and the eukaryotic expression plasmid pCDNA3.1/myc-his(-)B-sTGFbetaR II(pCDNA-sTGFbetaR II) was constructed by inserting the sTGFbetaR II cDNA into the EcoR I/Hind III-digested pCDNA. The DNA sequence was confirmed by double digestion and the pCDNA-sTGFbetaR II plasmid was transfected into the CHO cell line. The sTGFbetaR II protein was confirmed by Western blotting analysis and its biological function was determined. RESULTS: The specific protein was observed in western blotting, and the protein abrogated the growth-inhibitory effects of TGF-beta1 on mink lung epithelial cells (Mv1Lu). CONCLUSION: The eukaryotic expression plasmid pCDNA-sTGFbetaR II has been successfully constructed and the sTGFsTGFbetaR II protein with biological activity obtained.

Amino Acid Sequence↗