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Objective To investigate the expression profile variation of long non-coding RNAs (lncRNAs) in ankylosing sporidylitis (AS) and explore the role of lncRNAs in the pathogenesis of AS.Methods The peripheral blood mononuclear cells of AS patients and health controls (HC) were used to detect for differently expressed lncRNAs by microarray.The roles of lncRNAs were predicted with GO and pathway analysis.The results were verified by real time-polymerase chain reaction (PCR).Results A total of 148 lncRNAs and 134 mRNAs were detected,which had more than 2-fold differentially expressed in AS patients.Bioinformatics analysis found that GO term enrichment included protein binding,regulation of transcription,metabolism,signal transduction,et al.and might involve in toll-like receptor pathway,protein kinase,complement pathway,notch signaling pathway and so on.The expressions of three lncRNAs were estimated by real time-PCR which found that consistent with that of microarrays.Among these,D90064 was the most aberrantly expressed lncRNAs.Conclusions Several lncRNAs expression was changed significantly in AS patients in comparison with HC,which implies that those different lncRNAs may have an important role in the development and progression of AS.
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Objective This study is aimed to evaluate the association between the ABCG2 gene rs2231142 variant and gout using meta-analysis. Methods Related studies were identified by searching extensively in Chinese and foreign language databases such as Pubmed, EMBASE, Cochrane Library, CBMdisc databases and so on. The quality of included studies was assessed by using the Newcastle-Ottawa Scale (NOS). The odds ratio (OR) was calculated using a random-effects or fixed-effects model. A Q statistic was used to evaluate the heterogeneity, and Eggerˊs test and funnel plot were used to assess publication bias. Sub-group analyses on ethnicities and sex were also performed. Results A total of 10 studies, including 3 478 gout patients and 10,089 controls from 6 countries or regions, were included and identified for the current metaan-alysis. It was found that the A allele or AA genotype of the ABCG2 rs2231142 polymorphism had an increased risk for gout in the general population [A allele: OR=2.03, 95%CI (1.77, 2.34), P<0.01 and AA genotype: OR=3.01, 95%CI (2.34, 3.88), P<0.01, respectively]. Similar results were found in sub-group analyses of different gender and races. Conclusion Existing evidence indicate that rs2231142 polymorphism (the A allele and AA genotype) is associated with increased risk of gout.
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BACKGROUND:Experiments have shown that the col agen substrate has the capability of stimulating cartilage generation, but the stimulating role of different types of col agen substrates remains controversial. OBJECTIVE:To investigate the effect of type I and type II col agen on the biological characteristics of human chondrocytes cultured in vitro. METHODS:Human chondrocytes at passage were cultured onto the ordinary culture plates (ordinary plate), type I col agen-coated culture plates (type I plate), and type II col agen-coated culture plates (type II plate). cellgrowth curves were determined by MTT method after cells were cultured for 10 days. By ELISA, PCR, and 1,9-dimethyl methyleneblue technology, type I and type II col agen and glycosaminoglycan contents were quantitatively detected in cartilage cells 28 days after culture. RESULTS AND CONCLUSION:The number of cartilage cells was the highest in type II plate, which was twice of that in type I plate and five times of that in ordinary plate. Cartilage cells in type II plate secreted the least amount of type I col agen, which showed significant differences compared with the ordinary plate (P0.01). Cartilage cells in type II plate secreted the most amount of type II col agen and glycosaminoglycan, showing significant differences compared with the other two plates (P<0.01). The cartilage cells cultured in col agen plates are better than that cultured in ordinary culture plate, type II col agen culture plate is better than type I col agen culture plate in maintaining cellshape, extending the dedifferentiation pattern, and promoting celldifferentiation.
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Objective To analyze the infection status and variation tendency of chlamydia trachomatis (CT) ,Neisseria gonorrhoe-ae(NG) and ureaplasma urealyticum(UU) in female genital tract in northeast Sichuan province during 2005 -2012 to provide the laboratory basis for their diagnosis and treatment Methods The real-time fluorescent quantitative PCR (RT-PCR) was applied to detect the CT DNA ,NG DNA and UU DNA in 1 386 samples from female genital tract and the detection results were performed the statistical analysis .Results The total positive rate of these 3 kinds of pathogens was 62 .8% (871/1 386) .Among the simple in-fection ,UU had the highest positive rate(48 .0% ,665/1 386);the positive rates of CT and NG were only 2 .2% .In the mixed infec-tion ,the positive rate of CT + UU was highest(6 .5% ,90/1 386) ,while which of UU + NG ,CT + NG and CT+ NG+ UU was 2 .5% (35/1 386) ,0 .4% (5/1 386) and 1 .1% (15/1 386) respectively .In different age groups ,the positive rate in the age 20 years old groups were all more than 60% .The positive rate of the CT ,NG and UU pathogens in females was in continuous high level during 2005 -2012 ,and which totally showed an increasing tendency . Conclusion CT and UU are the main pathogens in female genital tract infection in this region ,and the positive rate of genital tract infection in females aged more than 20 years is higher ,the infection rate of these 3 kinds of pathogens demonstrates the increasing trend year by year ,so more attention should be paid to the detection of CT and UU in this group for guiding the clinicians to con-duct the diagnosis and treatment .
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Objective To investigate the single nucleotide polymorphisms(SNPs) rs3733591(C>T) of SLC2A9 gene in Chinese Han population, and to explore the association of this gene polymorphisms with gout susceptibility, tophi, serum uric acid levels, other clinical and laboratory data and the levels of SLC2A9 mRNA of peripheral blood mononuclear cells(PBMCs). Methods ① A total of 297 primary gout arthritis patients(GA) and 211 normal controls(NC) were enrolled into this study. The clinical and laboratory data of patients were collected. The genotypes and alleles frequencies were measured by using TaqMan ?SNP Geno-typing Assays and the possible association between gene polymorphism of SLC2A9 and gout was investigated by Chi-square test. The odds ratios(OR) and 95% confidence intervals(95%CI) were calculated. ② The lev-els of SLC2A9 mRNA on PBMCs of 86 gout patients(46 patients in remission) and controls were measured by real-time quantitative polymerase chain reaction (RT-qPCR). The nonparametric test was used to analyze the expression in different groups. Results The frequencies of genotypes and alleles of rs3733591(C>T) in gout patients were different from controls(P0.05). However, there was no significant difference in the distribution of genotypes and alleles between 30 tophaceous gout patients and 190 non-tophaceous gout patients(P>0.05). Conclusion Results of present study suggest the rs3733591(C>T) polymorphism of the SLC2A9 gene might be associated with gout development, but not with tophaceous gout. The C allele predisposes to gout, and TT genotype and T allele might protect Chinese Han population from developing gout. The rs3733591(C>T) polymorphism probably affects the susceptibility to gout by influencing the f expression of SLC2A9 mRNA susceptibility.
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Objective To measure the level of androgen receptor (AR) mRNA in peripheral blood monocytes (PBMCs) and serum testosterone level of patients with gouty arthritis (GA) and healthy controls (HC),and to explore the role of testosterone and AR in the pathogenesis of GA.Methods Chemilluminescence was used to detect the level of serum testosterone in GA [including 119 acute GA (AGA) and 60 nonacute GA (NAGA) patients] and 47 HC group.Real-time quantitative polymerase chain reaction (RT-qPCR)was used to measure AR mRNA in PBMCs from 41 GA and 35 HC.Western blotting was used to measure PBMCs AR in GA and HC for each 6 cases.One-way ANOVA,t test and Spearman's correlation were adopted for statistical analysis.Results Serum testosterone was significantly reduced in AGA and NAGA group compared to that in HC group [(6.1±1.5) ng/ml,P＜0.01,respectively],and the expression was lower in the AGA [(3.7±1.4) ng/ml] group [(4.9±2.0) ng/ml] than that in the NAGA group (P＜0.01).The level of AR mRNA and protein was much lower in the GA group than that in the HC group (P＜0.01,respectively).Negative correlations was detected between AR mRNA and uric acid in GA patients.There was negative correlation between serum testosterone and VLDL,GLU; meanwhile,positive correlation was found between serum testosterone and HDL (P＜0.05,respectively) in NAGA patients.There were no correlations between testosterone and other laboratory data.There was no correlation between AR and other laboratory data in GA patients and healthy controls (P＞0.05,respectively).Conclusion Altered expression of testosterone and its receptor may be involved in the pathogenesis of gouty inflammation.Further study will be needed to shed light on the exact role of androgen and AR in gout.
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Objective To detect the distribution of SLC2A9 rs10489070 polymorphism genotypes in Chinese Han population,and to explore the association of this gene polymorphism with gout susceptibility,tophi,serum uric acid levels and other clinical and laboratory data.Methods A total of 151 primary gout patients and 176.healthy controls were enrolled into this study.The genotypes and alleles frequencies were calculated by using TaqMan(R) SNP Genotyping Assays and the possible association between gene polymorphism of SLC2A9 and gout was investigated.T test,Chi-square and Fisher exact probabilities were used for statistcal analysis.Results Genotypes distribution were in Hardy-Weinberg equilibrium in gout patients and controls (P＞0.05).The frequency of CC genotype in gout patients was significantly higher than that in the controls (78.8％ vs 68.5％,P＜0.05),aand the frequency of CG genotype in gout patients was significantly lower (19.9％ vs 30.1％,P＜0.05).However,there were no statistical differences in the alleles frequencies of C and G between gout patients and controls (P＞0.05).Interestingly,there was significant difference in the distribution of genotypes between tophaceous gout patients and non-tophaceous gout patients (P＜0.05),and the frequency of CG genolype was much lower in tophaceous gout patients (0 vs 22.7％,P＜0.05).Conclusion Results of present study suggest that rs10489070 polymorphism of the SLC2A9 gene might be associated with gout development.CC genotype predisposes to gout,and CG genotype might protect Chinese Han population from gout and tophi development.
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Objective To investigate the role of adiponectin (ADP) and its receptors (ADR) in pati ents with primary gouty arthritis (GA).Methods Enzyme-linked immunosorbent assay (ELISA) was used to determine the levels of plasma ADP in 88 GA and 80 healthy controls (NC).Real time quantitative polymerase chain reaction (RT-qPCR) was employed to study the expression of ADR1 and ADR2 mRNA in peripheral blood mononuclear cells (PBMCs).The biochemical indicator of TG,HDL,LDL,VLDL,apoA1,apoB100 and uric acid (UA) were detected at the same time.T test,Spearman's correlations and regression analysis were used for statistical analysis.Results The concentration of plasma ADP was significantly lower in GA than that in NC [(6±7) μg/ml,(8±6) μg/ml,t=-3.71,P＜0.01 ],the expression of ADR1 and ADR2 mRNA in GA (ADR1:0.09±0.08,ADR2:0.0122±0.0164) was significantly increased when compared to the NC (ADR1:0.05±0.03,ADR2:0.0054±0.0024) (t=2.71,2.35,P＜0.05).In the GA patients,the level of ADP was negatively correlated with the lymphocytes count (LY) and UA (r=-0.32,-0.36,P＜0.05),and it was positively correlated with ESR and LDL level (r=0.31,0.39,P＜0.05).The expression of ADR1 mRNAwas negatively correlated with TG (r=-0.43,P＜0.05 ),but positivdy correlated with ESR and CRP level (r=0.45,0.57,P＜0.05).The expression of ADR2 mRNA was negatively correlated with glucose and UA(r=-0.50,-0.59,P＜0.05).Conclusion Altered expression of ADP and its receptors may be involved in thepathogenesis of gouty inflammation.
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objective The roles of TLRs and their signal pathway in gouty arthritis(GA)were explored.Methods TLR2 and TLR4 mRNA was measured using real-time quantitative polymerase chain reaction(RT-PCR)in PBMCs,IL-1β level was detected using ELISA in plasma,and NF-κB p65 protein level in PBMCs was measured using Western blot.Level of TLR2 mRNA,ILR4 mRNA,IL-1β,NF-κB p65protein was compared among acute GA,non-acute GA and healthy controls.Correlation between TLR2mRNA,TLR4 mRNA and serum uric acid,IL-1β level in GA patients was analyzed.One-way ANOVA was used to analyze data between multiple groups and q-test was used for two-two comparison.Spearman's analysis was applied for correlation analysis.Resuits The expression of TLR4 mRNA,NF-KB p65 protein,IL-1β arid serum uric acid level in patients with acute GA [(5.0±1.2), (7.11±0.18), (283±83)pg/ml,[585±123)μmol/L] was significantly increased compared to non-acute GA[(2.3±0.4),(0.63±0.06),(134±29)pg/ml,(493±107)μmol/Lj and healthy controls(1.1±0.6),(0.52±0.12),(97±17)pg/ml,(326±65)μmol/L](P<0.01,respectively).Significant diffefence was also observed between non-acute GA patients and healthy controls(P<0.05,respectively).The level of TLRR4 mRNA was positively correlated with uric acid and IL-1β level in GA patients(rs=0.876,0.779;P<0.05,respectively).Conclusion Innate immunity are activated by membrane-type pattern recognition receptors in primary GA.TLR4-NFκB p65-IL-1β signat transduction may participate in the inflammatory mechanisms of gout.Urate crystals in patients with gout may:be involved in the activation of TLR4 and its signal pathway.
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Objective To investigate the expression levels of telomeric protein TPP1 and POT1mRNA in the peripheral blood mononuclear cells (PBMCs) of patients with systemic lupus erythematosus (SLE), and the relations between these gene expression levels and disease activity are explored. Methods TPP1 and POT1 genes were measured using real-time polymerase chain reaction (RT-PCR) in PBMCs. The expression levels of TPP1 and POT1 genes in PBMCs were compared between 48 SLE patients and 30 healthy indivi-duals. Results The expression levels of TPP1 and POT1 in the PBMCs of SLE patients significantly decreased compared to healthy individuals (P<0.01). The expression levels of TPP1 and POT1 were much lower in SLE patients with lupus nephritis than those in patients without lupus nephritis (P<0.01), and in LN patients, the levels of TPP1 and POT1 were negatively correlated with proteinuria. The expression levels of POT1 in active SLE patients was lower than that in inactive SLE patients (P<0.05). The expression levels of POT1 was negatively correlated with serum IgG and SLEDAI scores, but positively correlated with complement C3, but no correlation between ESR, CRP, C4,ANA and the expression levels of TPP1, POT1, SLEDAI scores. Conclusion TPP1 and POT1 genes play a key role in the etiology of SLE, and they are involved in the pathogenesis of lupus nephritis. POT1 is helpful in evaluating SLE disease activity and severity.
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Objective To study the expression level of NLRP3 (NLR family, pyrin domain containing3) inflammasome (NLRP3, ASC, caspase-1 ) mRNA in peripheral blood monocytes (PBMCs) from patients with gout arthritis (GA) and to explore the pathogenesis of GA. Methods NLRP3 inflammasome mRNA was measured using quantitative real-time PCR in PBMCs. The expression of NLRP3 inflammasome mRNA in PBMCs was compared between patients with GA (n=24) and healthy controls (n=24). β-actin was selected as the internal control. Students' t-test was used in two independent samples and Spearman's correlation was used to evaluate the relationship between mRNA level and inflammatory parameters. Results The expression of ASC mRNA in GA increased significantly when compared to healthy controls [(0.029±0.021 ) vs (0.009±0.007 ), P＜0.01 ], and the expression of NLRP3 and caspase-1 mRNA was significantly lower in patients with GA compared to healthy controls [(0.062±0.084) vs (0.133±0.106), P＜0.05; (0.025±0.014) vs (0.117±0.156), P＜0.01]. Moreover, the expression of ASC mRNA was found to correlate significantly with globulin (r=-0.547, P＜0.05) and very low density lipoprotein cholesterol (r=-0.540, P＜0.05) in GA patients,and caspase-1 was correlated to globulin(r= -0.773. P＜0.01) and very low density lipoprotein cholesterol (r=-0.465. P＜0.05). Furthermore, the ASC mRNA in GA patients was associated significantly with NLRP3 mRNA(r=-0.450, P＜0.05) and caspase-1 (r=0.604, P＜0.01 ). Conclusion Dysregulated expression of the NLRP3inflammasome is involved in the inflammatory response and plays a key role in the pathogenesis of GA.
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Objective To investigate the expression of human glucocorticoid-induced tumor necrosis factor receptor(hGITR)in synovial and cartilage tissue of patients with rheumatoid arthritis(RA),and analyze their relationships with the severity of synovial inflammation in RA.Methods Expression of hGITR in synovial and cartilage biopsy material of 16 RA patients,9 osteoarthritis(OA)patients,4 contmIs were detected in situ by immunohistochemical technique and the results were analyze then.Results hGITR was strongly expressed in the vascular endothelial cells and inflammatory cells in RA and hGITR positive cells could be detected in approximately 69%of RA synovial cells.Significant differenee Was foand in hGITR expression between RA patients and control group(P<0.01).hGITR was less expressed in the cartilage than the synovial in RA.No significant difference was found in expression in the cartilage tissue expression between RA patients and the control group.Significant difference Was found in hGITR expression between synovial and cartilage in RA.Furthermore.the hGITR positive cells in synovial tissues were also found to correlate significandy with the severity of synovial inflammation in RA patients(r=0.895,P<0.01).Conclusion These results suggest that the abnormal expression of hGITR in the synovial my bean important mechanism leading to the synovial destruction found in RA.
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This study was conducted to amplify the cfpl0-esat6 fusion gene by SOE and insert into the integrating shuttle plasmid pMV361 to form the recombinant plasmid. Then another recombinant plasmid was constructed by insertinga-A g signal sequence of BCG. The two recombinant plasmids were introduced into BCG and the induced products from recombinant BCG were analyzed. In conclusion,the successful construction of rBCG expressing the fusion protein CFP10-ESAT6 will be the base of the development of novel Mycobacterium tuberclosis vaccines.
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Antigens, Bacterial , Genetics , Bacterial Proteins , Genetics , Mycobacterium bovis , Genetics , Metabolism , Mycobacterium tuberculosis , Genetics , Plasmids , Recombinant Fusion Proteins , Genetics , Allergy and Immunology , Tuberculosis VaccinesABSTRACT
This study was conducted to potentiate the expression of outer membrane protein OmpL17 of the strong virulent L. interrogans serovar Lai and investigate its immunogenicity in rabbits. The OmpL17 was cloned into prokaryotic expression vector pGEX-1lambdaT. The recombination expression plasmid pGEX-OmpL17 was transformed into E. Coli JM109. The GST fused protein GST-OmpL17 was expressed after induction by IPTG, then GST-tag was by thrombin and purified using Bulk GST purification Modules. SDS-PAGE and Western blotting analysis indicated that the molecular weight of GST-OmpL17 and OmpL17 was about 54 KDa and 28 KDa respectively. The outer membrane protein OmpL17 was subcutaneously injected into rabbits and high titre anti-OmpL17 antibody was obtained (1:4896) which could conjugate specifical with OmpL17. In conclusion, OmpL17 and specifical anti-OmpL17 antibody were obtained, which provided an experimental basis for researching pathogenic effect and immunity functions of OmpL17.
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Humans , Bacterial Outer Membrane Proteins , Genetics , Allergy and Immunology , Bacterial Proteins , Genetics , Allergy and Immunology , Cloning, Molecular , Leptospira interrogans , Genetics , Allergy and Immunology , Porins , Genetics , Allergy and Immunology , Recombinant Proteins , Genetics , Allergy and Immunology , VirulenceABSTRACT
Objective To explore new method for enhancing the efficacy of tuberculosis DNA vaccine. Methods Two recombinant plasmids were constructed, one named as pBK GM/85A encoding mouse granulocyte macrophage colony stimulating factor(GM CSF) fused to Mycobacterium tuberculosis Ag85A antigen, the other named as pBK 85A encoding Mycobacterium tuberculosis Ag85A antigen alone. Subsequently, the two plasmids were transferred into cultured COS7 cells by using cationic liposomes. The expression products were identified by Western blotting. Then, in a murine model, we compared the immunogenicity and protective immunity of the two recombinant plasmids following genetic immunization. Results All pBK GM/85A injected mice elicited higher antibody titres than that for pBK 85A injected mice. Lymphocytes obtained from the spleen of pBK GM/85A immunized mice exhibited higher lymphocyte proliferative response、IFN ? production and CTL activity than that for pBK 85A immunized mice. The protective efficacy was also higher for pBK GM/85A immunized mice than that for pBK 85A immunized mice. However, The protective efficacy for pBK GM/85A immunized mice was lower than that for BCG immunized mice. Conclusion These results showed that DNA vaccines with GM CSF/antigen fusion constructs could greatly improve the immunogenicity of DNA vaccine against Mycobacterium tuberculosis.