ABSTRACT
Objective To construct an eukaryotic expression vetor of ring finger protein 6 (RNF6)and to investigate its ubiqu-itylation on insulin receptor substrate-1(IRS-1).Methods Human RNF6 coding sequence was amplified by polymerase chain reac-tion (PCR)method with human cDNA as template.The pcDNA3.1-CHA-RNF6 was constructed with routine molecular methods and transfected into hepatocarcinoma cell HepG2.Relative quantification of IRS-1 mRNA was preformed by real-time reverse tran-scription PCR.Western blot was applied to detect the expression levels of IRS-1.Results 48 h after the plasmid carring RNF6 gene transfected HepG2 cells,the mRNA level of ISR-1 gene reduced to 69% of control group.The expression of IRS-1 was markedly lower than control group (P <0.01).Conclusion The expression of IRS-1 is markedly down-regulated in HepG2 and enhancement of the ubiquitylation level of IRS-1 contribute to the disorder in insulin signal transferring.
ABSTRACT
Objective To investigate the roles of the cytokine and chemokine in protective immunity of tuberculosis(TB) patient. Methods The RANTES( CCL5) and MIP1-?( CCL3) content of the Supernatant from PBMC stimulated with IL-4、TNF-?、MIP1-?and LPS in TB patients and normal control subjects was assessed using commercially available ELISA kit. Results The RANTES secretion was increased significantly in both TNF-?plus LPS groups[ (1 731. 86?925. 60) ng/L ] and IFN-?plus LPS [(2 120.78?1 120.72)ng/L] compared with LPS group[ (1 102. 56?873. 44) ng/L] in TB patients (P 0. 05). The MIP1-?secretion in response to IL-4、TNF-?、IFN-?was unchanged significantly in PBMC of TB patients and normal subjects. Conclusion TNF-?、IFN-?and RANTES may play important roles in protective immunity of TB patient