Decreased expression of alpha-enolase inhibits the proliferation of hypoxia-induced rheumatoid arthritis fibroblasts-like synoviocytes.

OBJECTIVES: To investigate the effect of decreased alpha-enolase (ENO1) expression on rheumatoid arthritis fibroblasts-like synoviocytes (RA-FLSs) proliferation in response to hypoxia, and elucidate the possible mechanisms involved. METHODS: RA-FLSs and osteoarthritis fibroblasts-like synoviocytes (OA-FLSs) were cultured in tri-gas incubators with different oxygen concentrations (3% O2, 7% O2, and 21% O2). 3% O2 (hypoxia) and 7% O2 conditions simulated intra-articular oxygen concentrations as observed in RA and healthy individual, respectively. 21% O2 represented oxygen condition for normal cell culture. ENO1-knockdown FLSs were established using ENO1-siRNA. The expression level of ENO1 was detected using reverse transcription polymerase chain reaction or RT-PCR and Western blot. Proliferation and apoptosis of RA-FLSs and OA-FLSs were assessed using 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2 H-tetrazolium or MTS assay and flow cytometry, respectively. Western blot analysis was used to detect key proteins involved in apoptosis. RESULTS: ENO1 gene expression was remarkably upregulated, as well as its translation into protein, in RA-FLSs and OA-FLSs that were cultured in 3% O2 concentration. RA-FLSs and OA-FLSs that were cultured under hypoxic conditions hyperproliferated compared with similar cells under normaxic conditions. Neither 7% O2 nor 21% O2 condition had any significant effect on ENO1 expression. ENO1-siRNA-transfected FLSs, but not control-siRNA FLSs, showed markedly decreased proliferation. Additionally, ENO1 expression was found to promote significantly higher expression levels of the anti-apoptotic proteins Bcl-2, surviving, and cyclinB1, but inhibited the expression of cleaved caspase3. CONCLUSION: ENO1 may be crucial in the regulation of the proliferation and survival of synovial fibroblasts.

Growth differentiation factor 15, ischemia modified albumin and pregnancy-associated plasma protein A in patients with coronary artery disease.

BACKGROUND: Growth differentiation factor 15 (GDF-15), ischemia modified albumin (IMA) might aid in the early diagnosis and risk stratification of patients with coronary artery disease (CAD), while pregnancy associated plasma protein-A (PAPP-A) can serve as a useful marker of vulnerable plaques and acute coronary syndrome (ACS). We sought to determine serum levels of GDF-15, IMA, PAPP-A and evaluate their diagnostic value in different types of CAD. METHODS: We detected serum levels of GDF-15, IMA and PAPP-A in 348 patients with CAD and 205 controls. Levels of high-sensitivity C reactive protein, creatine kinase isoenzyme MB, and high-sensitivity cardiac troponin-T, which represent biomarkers of inflammation, damage or necrosis, were also evaluated. RESULTS: There were significant differences of GDF-15, IMA and PAPP-A in patients with CAD compared with the controls, where GDF-15 seemed to be associated with severity of CAD. GDF-15 demonstrated a sensitivity of 84.0% and specificity of 84.8% for diagnosis of UAP, while the negative predictive value was 87.4%. The sensitivity and specificity, negative predictive value of IMA in the diagnosis of UAP were 70.0%, 69.5%, and 70.0%, respectively. PAPP-A had the lowest sensitivity and negative predictive value compared with other markers. CONCLUSIONS: GDF-15 demonstrated a significant improvement in earlier prediction and assessment of overall patient risk of UAP comparing to IMA and PAPP-A.

Age-specific 99th percentile cutoff of high-sensitivity cardiac troponin T for early prediction of non-ST-segment elevation myocardial infarction (NSTEMI) in middle-aged patients.

BACKGROUND: High-sensitivity cardiac troponin T (hs-cTnT) assay is used in the diagnosis and risk assessment of patients with symptoms of myocardial infarction. This study was undertaken to establish an age-specific 99th percentile cutoff value for hs-cTnT in Chinese population, and to evaluate its potential for early prediction of non-ST-segment elevation myocardial infarction (NSTEMI) in middle-aged patients. METHODS: Troponin T levels in blood obtained from healthy Chinese adults were assayed using hs-cTnT. The distribution was plotted and 99th percentiles were determined by nonparametric statistics. Prediction performance at the conventional cutoff (14 ng/L) recommended by the Roche company was compared with the age-specific cutoff for NSTEMI in 100 middle-aged patients (40-60 years of age) with acute chest pain. RESULTS: The 99th percentile for hs-cTnT was 14 ng/L for patients ≥60 years of age and 11 ng/L for those <60. Fifty of the 100 patients were finally diagnosed with NSTEMI. The age-specific 99th percentile cutoff value of 11 ng/L identified a higher number of patients with NSTEMI than the conventional 14 ng/L cutoff (46 vs. 40 patients), although the difference was not statistically significant (P = 0.084). In addition, the sensitivity of hs-cTnT increased from 80 to 92% and the negative predictive values increased from 82.4 to 91.8%. CONCLUSION: Using 11 ng/L as a decision-making cutoff point for hs-cTnT facilitated earlier prediction of NSTEMI in middle-aged patients than the conventional 14 ng/L cutoff. Further studies are needed to confirm this finding in larger group of patients.

Proteins related to the functions of fibroblast-like synoviocytes identified by proteomic analysis.

OBJECTIVES: It is well known that the fibroblast-like synoviocytes (FLS) play a key role in pathogenesis of rheumatoid arthritis (RA). This study was performed to separate the differentially expressed proteins of FLS from the patients with RA or osteoarthritis (OA) by two-dimensional electrophoresis (2-DE), and found proteins associated with the functions of FLS by mass spectrometry (MS). METHODS: Total proteins were extracted and quantified from the primary cultured FLS from patients of RA (n=8) or OA (n=6). Proteins were separated by high-resolution 2-DE, and identified the differentially expressed proteins by MS. Western blot analyses was used to validated the expression of candidate proteins. The mRNA of these proteins was detected by semi-quantitative fluorescent PCR. RESULTS: There are 1147 protein spots from RA and 1324 protein spots from OA showed on 2-DE graphs, respectively. We have selected 84 protein spots for MS analysis, and 27 protein spots were successfully identified. We have found that protein isoaspartyl methyltransferase (PIMT) and pirin (iron-binding nuclear protein, PIR) with lower expression in RA, and thioredoxin 1(Trx-1) only expressed in RA may be associated with functions of FLS. Western Blot confirmed the expression of PIMT and pirin lower in RA, and Trx-1 expressed only in RA. The results of semi-quantitative fluorescent PCR are also consistent with 2-DE graphs. CONCLUSIONS: PIMT, pirin and Trx-1 affect the functions of FLS in some style and can be the drug targets of RA.

Improving the antibacterial property of porcine small intestinal submucosa by nano-silver supplementation: a promising biological material to address the need for contaminated defect repair.

OBJECTIVE: We hypothesize that introduction of nano-silver particles to porcine-derived small intestinal submucosa (NS-PSIS) would lead to significant enhancement in antibacterial property in repairing contaminated abdominal defect. BACKGROUND: Porcine-derived small intestinal submucosa (PSIS) is an acellular and xenogenic biological material intensively used in repairing and regenerating wounded and dysfunctional tissues. Surgical site infection (SSI) remains so far a serious problem and major challenge, particularly in contaminated tissue-deficient repairing. METHODS: Self-assembly was used to fabricate NS-PSIS. The antibacterial property was evaluated in vitro and in vivo by means of repairing full-thickness contaminated abdominal defect in rats. The native PSIS and polypropylene-oxidized regenerated cellulose were served as controls. In addition, changes in biomechanical resistance, morphology and immunohistochemistry for inflammatory reaction and neovasculation in the repaired abdominal wall were analyzed. Biosafety was investigated by pyrogen test, skin irritation test and silver measurement in vivo. RESULTS: NS-PSIS exhibited strong antibacterial activity against Staphylococcus aureus, Staphylococcus epidermidis, Escherichia coli, and Pseudomonas aeruginosa on agar diffusion, with mean diameters of inhibition zone ranging from 11.9 to 23.5 mm. There were significantly lower SSI incidence and a tendency of better abdominal wall resistance in the NS-PSIS group as compared with the PSIS or polypropylene-oxidized regenerated cellulose group after repairing contaminated abdominal defect in rats. Nano-silver modified PSIS did not change the native PSIS property in the tissue recolonization, remodeling and neovascularization. NS-PSIS was not pyrogenic or skin irritated, without silver residual in vivo after repairing contaminated abdominal defect. CONCLUSION: Nano-silver particles to PSIS lead to significant enhancements in antibacterial property in vitro and in vivo without decreasing its biomechanical resistance and biocompatibility. This study provides proof of concept for the use of nano-silver modified naturally derived PSIS as an ideal scaffold for SSI prevention in the contaminated tissue-deficient repair.

Relationship of arterial stiffness and early mild diastolic heart failure in general middle and aged population.

AIMS: The brachial-ankle pulse wave velocity (baPWV) has been recognized as a marker that reflects arterial stiffness. We conducted an investigation to evaluate whether baPWV is independently associated with early mild diastolic heart failure (DHF) in a general middle and aged population. METHODS AND RESULTS: From 1 July 2009 until 31 August 2009, we investigated 2095 subjects for the relevant factors of heart failure in the Lujiazui Community, Shanghai. The baPWV, echocardiography, and blood sampling were performed in the morning after a 12 h overnight fast. A total of 1929 subjects had the complete data, including questionnaire, age, gender, baPWV, brain natriuretic peptide, and E/A ratio. Early mild DHF was defined as a left ventricular ejection fraction >50%, E/A ratio <0.8, and E/E' ≤ 8; finally, 482 subjects with early mild DHF and 1282 subjects with non-DHF entered into analysis. Among 1764 subjects, 31.6% of the subjects were male (average age was 58.0 ± 12.3), 35.8% of the subjects had hypertension, the average body mass index (BMI) was 24.2 ± 3.3 kg/m(2), baPWV was 1513.0 (1329.1, 1763.5) cm/s, and the baPWV was significantly correlated with the E/A ratio (r = -0.39, P < 0.01). There was a difference of the baPWV [1456.0 (1295.3, 1698.3) vs. 1670.5 (1465.6, 1910.8) cm/s] between the non-DHF group and the early mild DHF group (P < 0.01). Multiple logistic-regression analyses demonstrated that age, male gender, BMI, baPWV, posterior left ventricular wall thickness (PVWT), interventricular septal thickness (IVST), E/E' ratio, left ventricular mass index (LVMI), systolic blood pressure (≥140 mmHg), and diastolic blood pressure (≥90 mmHg) were independently correlated with early mild DHF. CONCLUSIONS: The increased arterial stiffness is associated with early mild DHF in a general middle and aged population independently of age, male gender, BMI, PVWT, IVST, E/E' ratio, LVMI, and high blood pressure. The non-invasive techniques described may allow serial measurements to be made over time to monitor baPWV changes in arteries provided the introduction of anti-arteriosclerosis therapy.

Hydrogen-rich saline ameliorates the severity of l-arginine-induced acute pancreatitis in rats.

Molecular hydrogen, which reacts with the hydroxyl radical, has been considered as a novel antioxidant. Here, we evaluated the protective effects of hydrogen-rich saline on the l-arginine (l-Arg)-induced acute pancreatitis (AP). AP was induced in Sprague-Dawley rats by giving two intraperitoneal injections of l-Arg, each at concentrations of 250mg/100g body weight, with an interval of 1h. Hydrogen-rich saline (>0.6mM, 6ml/kg) or saline (6ml/kg) was administered, respectively, via tail vein 15min after each l-Arg administration. Severity of AP was assessed by analysis of serum amylase activity, pancreatic water content and histology. Samples of pancreas were taken for measuring malondialdehyde and myeloperoxidase. Apoptosis in pancreatic acinar cell was determined with terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling technique (TUNEL). Expression of proliferating cell nuclear antigen (PCNA) and nuclear factor kappa B (NF-kappaB) were detected with immunohistochemistry. Hydrogen-rich saline treatment significantly attenuated the severity of l-Arg-induced AP by ameliorating the increased serum amylase activity, inhibiting neutrophil infiltration, lipid oxidation and pancreatic tissue edema. Moreover, hydrogen-rich saline treatment could promote acinar cell proliferation, inhibit apoptosis and NF-kappaB activation. These results indicate that hydrogen treatment has a protective effect against AP, and the effect is possibly due to its ability to inhibit oxidative stress, apoptosis, NF-kappaB activation and to promote acinar cell proliferation.

[Association of the PADI4 gene polymorphism and HLA-DRB1 shared epitope alleles with rheumatoid arthritis].

OBJECTIVE: To investigate the association of single nucleotide polymorphisms (SNPs) of the peptidylarginine deiminase IV (PADI4) and HLA-DRB1 shared epitope (SE) alleles with rheumatoid arthritis(RA) in a Chinese population. METHODS: Four exonic SNPs of the PADI4 gene (PADI 4_89*A/G, PADI 4_90*C/T, PADI 4_92*C/G and PADI 4_104*C/T) were genotyped in 67 unrelated patients with RA and 81 healthy controls, using cDNA sequencing and T vector cloning. HLA-DRB 1*01, *04 and *10 subtypes were determined by polymerase chain reaction with sequence specific primers (PCR-SSP). RESULTS: The distributions of the 4 SNPs were different in the two groups, and increased RA susceptibility was significantly associated with the minor alleles of PADI 4_89*G (P was 0.023), PADI 4_90*T (P was 0.004), PADI 4_104*T (P was 0.003), and the haplotypes carrying the 4 minor alleles (P was 0.008). HLA-DRB1 SE alleles are composed of HLA-DRB 1*0101, *0102, *0401, *0404, *0405, *0408, *0409, *0410 and *1001. Individuals carrying the SE alleles were associated with increased RA susceptibility (P was 0.002). Individuals carrying both the SE alleles and minor alleles of the 4 SNPs were more susceptible to RA than individuals carrying neither the minor SNP alleles nor the SE alleles. CONCLUSION: The PADI4 SNPs and haplotypes are associated with RA susceptibility in Chinese. HLA-DRB1 shared epitope is also an important risky factor for RA. There may exist certain synergistic effect between the PADI4 minor alleles and the HLA-DRB1 shared epitope.

Relationship of High-Sensitivity C-Reactive Protein Concentrations and Systolic Heart Failure.

OBJECTIVE: Excessive activated proinflammatory cytokines may promote extracellular matrix alterations which induce adverse left ventricular remodeling in systolic heart failure (SHF). We sought to identify whether high-sensitivity C-reactive protein (hsCRP) levels were independently associated with SHF. METHODS: In our retrospective case-control study, 2236 subjects were included, and 260 patients had SHF. Blood sample collection, clinical laboratory tests, electrocardiogram and echocardiography examinations were performed. The questionnaires were completed by professional interviews. RESULTS: In 2236 subjects, the prevalence rate of SHF were 1.7, 1.8, 8.4 and 32.6% between hsCRP concentrations (<1 mg/L, ≥1 to <3 mg/L, ≥3 to <10 mg/L and ≥10 mg/L, respectively) (p=0.000). hsCRP concentrations (<1 mg/L, ≥1 to <3 mg/L, ≥3 to <10 mg/L and ≥10 mg/L) were associated in a linear trend with N-terminal pro-brain natriuretic peptide (NT-proBNP, p=0.000) and left ventricular ejection fraction (LVEF, p=0.000). hsCRP was also significantly related to NT-proBNP, LVEF and SHF (r=0.232, p=0.000; r=-0.358, p=0.000 and r=0.413, p=0.000, respectively). In logistic regression model, after adjusting for heart failure risk factors, compared with the low concentration of hsCRP (<1 mg/L), the high concentration of hsCRP (≥10 mg/L) was significantly independently associated with SHF (odds ratio = 10.78 [1.303 to 89.10], p=0.027). CONCLUSION: Low to high concentration of hsCRP showed a linear trend association with SHF. A high concentration of hsCRP was independently associated with SHF.

Glucose-6-Phosphate Isomerase (G6PI) Mediates Hypoxia-Induced Angiogenesis in Rheumatoid Arthritis.

The higher level of Glucose-6-phosphate isomerase (G6PI) has been found in both synovial tissue and synovial fluid of rheumatoid arthritis (RA) patients, while the function of G6PI in RA remains unclear. Herein we found the enrichment of G6PI in microvascular endothelial cells of synovial tissue in RA patients, where a 3% O2 hypoxia environment has been identified. In order to determine the correlation between the high G6PI level and the low oxygen concentration in RA, a hypoxia condition (~3% O2) in vitro was applied to mimic the RA environment in vivo. Hypoxia promoted cellular proliferation of rheumatoid arthritis synovial fibroblasts (RASFs), and induced cell migration and angiogenic tube formation of human dermal microvascular endothelial cells (HDMECs), which were accompanied with the increased expression of G6PI and HIF-1α. Through application of G6PI loss-of-function assays, we confirmed the requirement of G6PI expression for those hypoxia-induced phenotype in RA. In addition, we demonstrated for the first time that G6PI plays key roles in regulating VEGF secretion from RASFs to regulate the hypoxia-induced angiogenesis in RA. Taken together, we demonstrated a novel pathway regulating hypoxia-induced angiogenesis in RA mediated by G6PI.

[A study on the relativity between La protein and the stability of HBV mRNA and the expression of HBV protein].

OBJECTIVE: To study the relativity between La protein and the stability of HBV mRNA and the expression of HBV protein. METHODS: Four specific siRNAs were obtained by transcription in vitro. After transfection with the siRNAs into HepG2.2.15 cells for 3 days, the inhibitive effects of La protein were analyzed by Western blot; the content changes of HBsAg, HBeAg and HBV-DNA were detected by ECL and RT-PCR. RESULTS: In comparison to normal cells, La protein was less in the cells. There was less La protein in the cells trans-infected with siRNAs. HBsAg, the HBeAg and HBV-DNA secreted by the cells transfected with siRNA were also less than that in the normal cells. CONCLUSION: There is a correlation between La protein and HBV mRNA and the expression of HBV protein.

Genetic association of cytokines polymorphisms with autoimmune hepatitis and primary biliary cirrhosis in the Chinese.

AIM: To characterize gene polymorphism of several cytokine gene in-patients with AIH and PBC and to analyze the difference of the polymorphism distribution between Chinese patients and healthy controls. METHODS: The study population consisted of 62 patients with AIH, and 77 patients with PBC. The genetic profile of four cytokines was analyzed by restriction fragment length polymorphism after specific PCR amplification (PCR-RFLP) or sequence-specific primers PCR (SSP-PCR). The analyzed gene polymorphism included interleukin-1 (IL-1) (at position +3 953 and IL-1RN intron 2), IL-6 (at position -174), IL-10 promoter (at position -1 082, -819, and -592). The control group consisted of 160 healthy blood donors. RESULTS: The majority of Chinese people including patients and healthy controls exhibited IL-1B 1,1 genotype, and there was no significant difference in AIH, PBC patients and controls. There were highly statistically significant differences in the distribution of the IL-1RN gene polymorphism between the patients with PBC compared with controls. The frequency of IL-1RN 1,1 was significantly higher (90.9% vs 79.4%, P = 0.03) and the frequency of IL-1RN 1,2 was significantly lower in PBC patients (6.5% vs 17.5%, P = 0.01). No statistical difference was observed between AIH patients and controls. All of the 160 healthy controls and 62 cases of AIH patients exhibited IL-6-174GG genotype, and there were four cases, which expressed IL-6-174GC genotype in 77 cases of PBC patients. The frequency of IL-6-174GC was markedly significantly higher in PBC patients compared with controls (5.2% vs 0%, P = 0.004). No statistically significant difference was found in the distribution of IL-10 promoter genotype in AIH and PBC patients compared with controls. CONCLUSION: The polymorphisms of IL-1RN and IL-6 -174G/C appear to be associated with PBC in Chinese patients.

Correlation of Chlamydia pneumoniae infection with primary biliary cirrhosis.

AIM: To evaluate the association between Chlamydia pneumoniae (Cpn) infection and primary biliary cirrhosis (PBC). METHODS: Cpn IgG and IgM were determined by enzyme-linked immunosorbent assay (ELISA) in 41 well-established PBC patients and two race-matched control groups (post-hepatitis cirrhosis, n = 70; healthy controls, n = 57). RESULTS: The mean level and seroprevalence of Cpn IgG in PBC group and post-hepatitis cirrhosis (PHC) group were significantly higher than those in healthy controls (46.8+/-43.4 RU/mL, 49.5+/-45.2 RU/mL vs 28.3+/-32.7 RU/mL; 68.3%, 71.4%, 42.1%, respectively; P<0.05). There was a remarkably elevated seroprevalence of Cpn IgM in patients with PBC (22.0%) compared to the PHC and healthy control (HC) groups. For the PBC patients versus the HCs, the odds ratios (ORs) of the presence of Cpn IgG and IgM were 2.7 (95% CI 0.9-6.1) and 5.1 (95% CI 1.4-18.5), respectively. Though there was no correlation in the level of Cpn IgG with total IgG in sera of patients with PBC (r = -0.857, P = 0.344>0.05), Cpn IgM was related with the abnormally high concentrations of total IgM in PBC group. CONCLUSION: The results of this study do not support the hypothesis that infection with Chlamydia pneumoniae may be a triggering agent or even a causative agent in PBC, but suggest that Chlamydia pneumoniae infection probably contributes to the high level of IgM present in most patients with PBC.

[Analysis of HLA alleles polymorphism in Chinese patients with primary biliary cirrhosis].

OBJECTIVE: To investigate the frequencies of human leuckocyte antigens (HLA) -A, B and DRB1 alleles in Chinese patients with primary biliary cirrhosis (PBC) using polymerase chain reaction-based techniques, and to assess the correlation of HLA molecules with other clinical and laboratory profiles. METHODS: Genotyping of HLA-A, B, and DRB1 were performed in 65 well-characterized patients with primary biliary cirrhosis and 431 healthy controls with PCR amplification with sequence-specific primers (PCR-SSP). RESULTS: The frequency of DRB1*0701 was increased to 29.2% compared with 13.9% in the controls (PC < 0.05, OR = 2.55, 95% CI: 1.4 approximately 4.6). No association was found with HLA-DRB1*08 which had been constantly reported. The A*2 allele (53.8%) was more frequent in the PBC patient group but without a significant statistical difference. The frequencies for the other A, B and DRB1 alleles were similar between patients and healthy controls. There was no difference between patients with or without DRB1*0701 in some clinical and laboratory profiles. CONCLUSION: Susceptibility to primary biliary cirrhosis in Chinese is associated with DRB1*0701 allele and differs from people in North America, South America, North Europe and even in Japan, but the association is not restricted to any particular subgroup of patients. Valine at position 78 of HLA DRbeta1 may play an important role in the pathogenesis of primary biliary cirrhosis.

Cytotoxic T lymphocyte associated antigen-4 gene polymorphisms confer susceptibility to primary biliary cirrhosis and autoimmune hepatitis in Chinese population.

AIM: To investigate the association between Chinese patients with autoimmune hepatitis (AIH), primary biliary cirrhosis (PBC) and the polymorphisms of cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) gene promoter (-318) and exon 1 (+49). METHODS: CTLA-4 promoter (-318 T/C) and exon1 (+49A/G) polymorphisms were genotyped via restriction fragment length polymorphism methods in 62 Chinese AIH patients, 77 Chinese PBC patients and 160 healthy controls. RESULTS: We found a significant association in CTLA-4 gene exon1 49 A/G polymorphism between PBC patients and controls (P = 0.006) and the frequency of G alleles was significantly increased in comparison with controls (P = 0.0046, OR = 1.8). We also found the frequency of C alleles in promoter -318 was significantly increased in AIH patients compared with controls (P = 0.02, OR = 0.41). Although the genotype distribution of the CTLA-4 exon 1-promoter gene was not significantly different between AIH and PBC patients and controls, the occurrence of GG-CC was increased in two groups of patients (AIH: 32.3%, PBC: 37.7%, control: 22.5%). CONCLUSION: Polymorphisms of CTLA-4 gene probably confer susceptibility to AIH and PBC in Chinese population.

[Study on the relationship of CTLA-4 -318, +49 polymorphisms with autoimmune hepatitis and primary biliary cirrhosis in a Chinese population].

OBJECTIVE: To investigate the association between Chinese patients with autoimmune hepatitis (AIH), primary biliary cirrhosis (PBC) and the polymorphisms of cytotoxic T lymphocyte -associated antigen-4 (CTLA-4) gene promoter (-318) and exon 1 (+49). METHODS: The CTLA-4 promoter (-318 T/C) and exon 1 (+49A/G) polymorphisms were genotyped via restriction fragment length polymorphism methods in 62 Chinese AIH patients, 77 Chinese PBC patients and 160 healthy controls. RESULTS: There was no difference in the distribution of CTLA-4 promoter -318 T/C polymorphisms between AIH patients and controls, but the C allele frequency was significantly increased in patients with AIH, compared to controls (P=0.02, OR=2.43). The distribution of CTLA-4 gene exon 1 49 A/G genotypes exhibited significant difference between PBC patients and controls (P=0.006), and the frequency of G allele showed a significant increase in PBC group as compared with controls (P=0.0046, OR=1.8). Although the genotype distribution of the CTLA-4 exon 1-promoter gene displayed no significant difference between AIH and PBC patients and controls, the occurrence of GG-CC was increased in the patients of the two groups (AIH: 32.3%, PBC: 37.7%; control: 22.5%). CONCLUSION: The above findings suggest that the polymorphisms of CTLA-4 gene probably confer susceptibility to AIH and PBC in the Chinese population.

[Genetic association of vitamin D receptor polymorphisms with primary biliary cirrhosis and autoimmune liver diseases on Chinese].

OBJECTIVE: To study the association between the polymorphisms of vitamin D receptor (VDR) gene and autoimmune liver diseases and (AIH) and primary biliary cirrhosis (PBC) in Chinese. METHODS: PCR-RELP was used to investigate the polymorphisms in exon 2, and exon 7 to exon 9 of VDR among 49 AIH patients, 58 PBC patients, and 160 healthy controls, all Chinese. VDR polymorphisms were assessed by FokI, BsmI, ApaI, and TaqI endonucleases restriction fragment length polymorphism analysis after specific polymerase chain reaction (PCR) amplification. RESULTS: The distribution of VDR gene polymorphism in Chinese was similar to those in Koreans and Japanese, and different from those in the Germans and Spaniards. The percentage of ff phenotype carriers was significantly higher in the AIH patients than in the healthy controls (34.7% vs. 48.1%, chi(2) = 5.47, P = 0.019) and the percentage of Ff phenotype carriers was lower in the AIH patients than in the healthy controls (34.7% vs. 48.1%, P = 0.057). The percentage of Bb phenotype carriers was significantly lower in the PBC patients than in the healthy controls (5.2% vs. 17.5%, P = 0.021) and. the percentage of bb phenotype carriers was significantly higher in the PBC patients than in the healthy controls (94.8% vs. 80.6%, chi(2) = 6.52, P = 0.01). We also detected a significant association of the BsmI polymorphisms in PBC patients in comparison with controls (P = 0.01). Furthermore we found the difference in the FokI, BsmI, ApaI, and TaqI genotype distribution between Chinese health controls and Caucasian health controls. CONCLUSION: There is a significant association between FokI polymorphism and AIH and a significant association between the BsmI polymorphisms and PBC in Chinese.

[Prediction and identification of autoepitopes of PDC-E2 specific CD8+ CTL in primary biliary cirrhosis patients].

OBJECTIVE: To identify autoepitopes of E2 subunit of pyruvate dehydrogenase complex (PDC-E2) specific CD8+ CTL in primary biliary cirrhosis (PBC) patients. METHODS: An online database SYFPEITHI was applied to predict HLA-A*0201 restricted epitopes which located in PDC-E2 30-50 aa and 150-190 aa where B-cell epitopes clustered with CD4+ T-cell epitopes. T2 cell line reconstitution and stabilization assay, induction of specific CTL lines from peripheral blood mononuclear cells (PBMCs) of patients with PBC and cytotoxicity of peptides-induced CTL were performed to screen the epitopes from those candidates. RESULTS: Five potential epitopes were predicted by database. Of the 5 candidates, two peptides 159-167 aa and 165-174 aa, with highly binding activity to HLA-A*0201 molecules, could stimulate PBMCs from most HLA-A*0201 positive PBC patients to proliferate and peptide-induced CTL lines showed specific cytotoxicity. CONCLUSION: Peptides of KLSEGDLLA (159-167 aa) and LLAEIETDKA (165-174 aa) in the inner lipoyl domain of PDC-E2 are HLA-A*0201 restricted CD8+ CTL immunodominant epitopes in PBC.

[Genetic association between interleukins gene polymorphisms with primary biliary cirrhosis in Chinese population].

OBJECTIVE: To determine the relationship between polymorphisms in the genes encoding IL-1, IL-6, and IL-10 with primary biliary cirrhosis (PBC) in Chinese population. METHODS: Whole-blood samples were taken from 77 patients with PBC and 160 healthy controls. DNA was extracted and the polymorphisms at positions IL-1 +3953, IL-1RN intron 2, IL-6 -174, and IL-10 -1082, -819, and -592 were determined by using sequence-specific polymerase chain reaction (SSP) or polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). RESULTS: The frequency of IL-1RN1,1 allele in PBC group was significantly higher than in control group (90.9% vs 79.4%, P=0.026), and the frequency of IL-1RN1,2 in PBC group was significantly lower than in control group (6.5% vs 18.8%, P=0.013). There was no significant difference in the frequence of IL-1RN*2 allele between PBC group and control group (P=0.06). Of the 77 patients with PBC, 4 patients were IL-6 -174GC, 73 were IL-6 174GG. All the 160 health controls are IL-6 -174GG (P=0.0036). The frequence of IL-6 -174C allele in PBC group was significantly higher than that in control group (P=0.0038). No significant differences of polymorphisms for IL-1 +3953 and IL-10 (-1082, -819 and -592) were found between PBC group and control group. CONCLUSION: The polymorphisms of IL-1RN and IL-6 -174G/C appear to be associated with PBC, and the polymorphisms of IL-1 +3953 and IL-10 promoter gene are not associated with PBC in a Chinese population.

[Genetic association of tumor necrosis factor (TNF)-alpha polymorphisms with primary biliary cirrhosis and autoimmune liver diseases in a Chinese population].

OBJECTIVE: Autoimmune hepatitis (AIH) and primary biliary cirrhosis (PBC) are two autoimmune diseases of unknown etiology. Genetic factors appear to be involved in the pathogenesis of both diseases. Tumor necrosis factor (TNF)-alpha is one of the proinflammatory cytokines and immunomodulators, and is implicated in the pathogenesis of AIH and PBC. In this study, we studied the association between Chinese patients with AIH, PBC and the polymorphisms in promoter-region polymorphisms of the TNF-alpha gene at position -308 and -238. METHODS: We have investigated four candidate gene loci in 49 patients with AIH, 58 patients with PBC, and 160 healthy controls. The polymorphisms were assessed by the PCR specifically for the single-nucleotide polymorphisms. RESULTS: We found the difference in the TNF-alpha gene at position -308 genotype distributions between Chinese health controls and Caucasian health controls. Although the percent of TNF-alpha*2 was decrease on PBC patient group (10.34% vs. 16.88%), there was no significant difference between PBC patients and health control in the Chinese. There were also no significant differences between AIH and health control on the TNF-alpha gene at position -308 and -238. CONCLUSION: Our findings suggest that the TNF-alpha promoter-region polymorphisms distribution is different between differe of ethnic groups; there are no genetic links of the TNF-alpha promoter-region polymorphisms to AIH and PBC in Chinese.