ABSTRACT
Objective To scan protein expression profile of immune complexes (ICs) derived from the synovial tissue of the patients with rheumatoid arthritis (RA) based on liquid chromatography-tandem mass spectrometry (LC-MS).Methods The samples of synovial fluid were obtained from knee joints of the patients with RA and osteoarthritis (OA) used as control during therapeutic arthrocentesis in knee jiont at the Department of Orthopedics of Jinling Hospital,School of Medicine,Nanjing University.The protein expression profile of ICs was identified by enrichment strategy based on immunoprecipitation and LC-MS analysis.The value of fraction of total (FOT) was used to estimate protein abundance and screen the up-and down-regulated proteins.The function enrichment,interaction network and signal pathway of differential proteins were analyzed using softwares David and String.Results A total of 511 and 526 protein spots in ICs of RA and OA patients were identified respectively.Among them,170 proteins existed only in RA group.45 and 85 proteins in RA group were statistically up-and down-expressed compared with controls.Conclusion HSP90AA1,HSP70,HLAG,Thioredoxin,Annexin A2 and vitronectin may be involved in the pathogenesis of RA through different paths and possible to become promising diagnostic indicators or new therapeutic targets for RA.
ABSTRACT
Objective The inhibitor of differentiation 3 (Id3) is an important transcriptional regulation factor, which participates in tumorigenesis, cell proliferation, and cell apoptosis.β-catenin, as a central molecule of the Wnt signaling pathway, is critical for tumor development.This study aimed to evaluate the expressions of these two molecules and the regulatory effect of Id3 on β-catenin in different tumor cells.Methods Total RNA was extracted using the Trizol Reagent.The relative mRNA expression levels of Id3 and β-catenin in tumor cells were detected by quantitative real-timePCR(qRT-PCR).The recombinant eukaryotic expression vector pEGFP/Id3 with the human Id3 gene was transfected into A549, A549/ DDP and SW-480 cells using the non-liposome-mediated method.The protein expressions of Id3 and β-catenin were determined by Western blot.Results The expression of Id3 was significantly lower in the colorectal cancer cell lines SW-480 and HT-29 than in A549 and other tumor cells (P0.05).Western blot showed the same results.Conclusion The expression levels of Id3 and β-catenin vary in different tumor cell lines.Anabnormally high level of β-catenin is an important risk factor for colorectal cancer, and the down-regulatedexpression of β-catenin after eogenous transfection of Id3 may provide some new ideas for target gene therapies of colorectal cancer.
ABSTRACT
Objective Autoantibodies and corresponding immune complexes involved directly in the development of rheuma -toid arthritis ( RA) .The effect of anti-mutated citrulline vimentin antibodies contained immune complex ( AMCV-IC) on the function of fibroblast-like synoviocytes ( FLSs) is still unclear .The aim of the study was to investigate the effect of AMCV-IC in serum of RA pa-tients on the proliferation of RA-FLSs. Methods Serum samples of RA , SLE ( systemic lupus erythematosus ) and healthy people were collected from outpatients and hospitalized patients in Nanjing General Hospital of Nanjing Military Region from March 2012 to March 2013 , including 20 RA patients ( 10 patients with serum AM-CV+and 10 patients with serum AMCV -) , 10 SLE patients and 10 healthy people .Serum AMCV of SLE patients and healthy people was negative .Serum IC was extracted by affinity chromatography with im-mobilized protein G and was divided into AMCV +-IC group,AMCV--IC group,SLE-IC group, and control group.The levels of AMCV were detected by ELISA .We cultivated RA patients'arthral FLSs in vitro .FLSs were stimulated with AMCV +-IC,AMCV--IC, SLE-IC and CON-IC (50,100,200,and 400μg/mL),respectively.And the effect of IC on RA-FLSs was detected by CCK-8. Results Compared with AMCV --IC group (1.37 ±0.86, 1.31 ±0.10),SLE-IC group (1.36 ±0.06, 1.38 ±0.03) and CON-IC group (1.08 ±0.02, 1.04 ±0.06),the A values of AMCV +-IC group (1.53 ±0.11, 1.58 ±0.05) were obviously higher after stimulating RA-FLSs with 50 and 100μg/mL IC for 24 h (P<0.05);Similarly, compared with AMCV --IC group (1.32 ±0.04, 1.38 ±0.07), SLE-IC group (1.31 ±0.10, 1.29 ±0.01) and CON-IC group (1.13 ±0.06, 1.03 ±0.09),the A values of AMCV +-IC group (1.53 ±0.16, 1.57 ±0.06) were obviously higher after stimulating RA-FLSs with 200 and 400μg/mL IC for 24 h (P<0.05). Meanwhile , the A values of AMCV --IC group and SLE-IC group were obviously higher than CON-IC group under different concentra-tions of IC(P<0.05). Conclusion AMCV+-IC can stimulate the proliferation of RA-FLSs more effectively compared with AM-CV--IC, SLE-IC and CON-IC under different concentrations of IC and involve in the pathogenesis of RA .