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Objective To investigate the regulatory effects of miR-146a on inflammation factors production in C ryptococcus neoformans treated THP-1 cells.Methods Cryptcoo ccus neoformans strains were heat killed .Fluorescence quantitative RTP-CR and ELISA were used to detect the levels of IL -1βand TNF-αin THP-1 cells treated with heat-killed Cryptococcus neoformans.In addition, the production of IL-1βand TNF-αwere analyzed before and after Dectin-1or TLR 4 blocked .THP-1 cell lines that were respectively transfected with miR-146 a mimics and inhibitors were constructed and the production of IL-1βand TNF-αin these cell lines were analyzed after interfered with Cryptococcus neoformans.Results With the interference of heat-killed Cryptococcus neoformans, the expression of miR-146a was up-regulated in THP-1 cells, but was down-regulated after Dectin-1 or TLR4 blocked.The expressions of IL-1βand TNF-αinduced by heat-killed Cryptococcus neoformans were enhanced in miR-146a mimics transfected THP-1 cells, but was inhibi-ted in inhibitors transfected THP-1 cells.Conclusion Heat-killed Cryptococcus neoformans could up-regu-late miR-146a expression in THP-1 cells via Dectin-1 and TLR.miR-146a could negatively regulate the ex-pressions of IL-1βand TNF-αinduced by Cryptococcus neoformans.
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Objective To evaluate the diagnostic performance of plasma miR-499 in acute myocardial infarction (AMI) diagnosis.Methods Diagnostic accuracy test.The suspected AMI patients,who with chest pain for more than half an hour and been admitted in the Second People's Hospital of Wuxi and First People's Hospital of chuzhou during October 2010 and July 2011,were consecutively and prospectively enrolled in the present study.Sixty apparently healthy individuals were designed as healthy control.The plasma samples of the suspected AMI patients were collected within two hours after admission.The plasma miR-499 was determined by real time polymerase chain reaction (RT-PCR).The diagnostic performance of plasma miR-499 for AMI was estimated by receiver operating characteristic (ROC) curve analysis.The association between plasma miR-499 and AMI was analyzed by multivariable logistic model.The plasma miR-499 was determined and explained in blind fashion.Results Two hundred and nine suspected AMI patients,including 131 confirmed AMI patients (46 STEMI and 85 NSTEMI) and 78 AMI free patients were enrolled in the present study.The delta cycle threshold (ΔCT) was 1.01 ± 3.34 for AMI patients,-2.76 ± 2.90 for non-AMI patients and-3.79 ± 2.21 for healthy controls.The differences had statistical significance (x2 =96.77,P ＜ 0.01).The area under curve (AUC) of plasma miR-499 was 0.80 (95％ C I:0.74-0.86),lower than that of cardiac troponin Ⅰ (AUC =0.90,95％ CI:0.86-0.94) on admission (P ＜0.01).At the optimal cut-off of 0.18,the diagnostic sensitivity and specificity were 0.69 (95％ CI:0.61-0.77) and 0.77 (95％ CI:0.66-0.86),respectively.The coefficient of correlation between plasma miR-499 and cTnI was 0.72 (P ＜0.01).The odds ratio (OR) of plasma miR-499 ＞0.18 for AMI was 2.59 (95％ CI:1.10-6.07),after adjusted cTnI.Conclusions Plasma miR-499 is a useful biomarker for AMI diagnosis.It can provide additional diagnostic information beyond cTnI.Combination utility of plasma miR-499 and cTnI may improve the diagnostic accuracy for AMI.
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Objective To investigate the effect of miR-146a on the proliferation and interleukin (IL)-2 production of T helper cells from primary biliary cirrhosis (PBC) patients.Methods MiR-146a in the peripheral blood mononuclear cells (PBMC),monocytes,T helper cells,cytotoxic T cells and B cells from 20 confirmede PBC patients and age/sex matched healthy controls were detected by quantitative PCR.By gainand-loss of function,the miR-146a's effect on anti-CD3/anti-CD28 activated T helper's proliferation and IL-2 production ability were measured by CCK-8 approach and enzyme linked immunosorbent assays (ELISA),respectively.Statistical analysis were carried out by t-test.Results PBMCs (0.46±0.20 vs 1.00±0.26; t=7.47,P＜0.01),T helpers (0.33±0.13 vs 1.00±0.14; t=6.15,P＜0.01) and monocytes (0.56±0.11 vs 1.00±0.11; t=4.97,P＜0.05),but not B cells (0.91±0.06 vs 1.00±0.14; t=0.97,P＞0.05) and cytotoxic T cells (0.98±0.15vs 1.00±0.12; t=0.22; P＞0.05) from PBC patients had lower miR-146a expression level than that of healthy controls.Inducible up expression of miR-146a was observed in PBC patients'T helpers stimulated with antiCD3/anti-CD28 (1.00±0.18 vs 9.12±2.05; t=8.81; P＜0.01).The activated T helpers from PBC patients had higher proliferative ability [PBC:0.35±0.06 (A); healthy controls:0.26±0.04 (A); t=2.83; P＜0.05] and increased IL-2 production [PBC: (685.60±109.19 pg/ml)]; Healthy controls: [(512.20±72.26) pg/ml; t=2.96; P＜0.05 ] than those of healthy controls.For activated T helpers,the proliferation ability,as well as IL-2 production,was enhanced by miR-146a.Conclusion MiR-146a can down regulate the proliferation and IL-2 releasing of activated T helpers.The reduced miR-146a expression enhances IL-2 production and promotes proliferation of T helper of PBC patients,thus,may be involved in the pathogenesis of PBC.
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Objective To investigate the increased expression of microRNA-146a(miR-146a) in peripheral blood mononuclear cells (PBMC) of patients with chronic immune thrombocytopenic purpura (ITP) and its clinical significance. Methods Twenty-eight patients with chronic ITP and 28 healthy controls matched with age and gender were enrolled in this study. Fluorescent quantitative PCR reaction was used to detect the relative expression of miR-146a in their PBMC. The serum concentration of TNF-α, IL-2,IL-1 β and IFN-γ were measured by ELISA. CCK-8 method was used to detect the proliferation ability of PBMC , which transfected with miR-146a mimics or inhibitor and then stimulated with platelet . Results The relative expression of miR-146a in ITP patients was higher than that of healthy controls. The increased expression of miR-146a was negatively correlated with the serum TNF-α, IL-2 and IFN-γ. The PBMC transfected with miR-146a mimics had reduced expression of IL-2 and proliferation when stimulated with platelet.In contrast, the opposite effect was observed with the miR-146a inhibitors transfection. Conclusion MiR146a was involved in the pathogenesis of chronic ITP by controlling IL-2 production and PBMC proliferation.Thus, it may be a potential therapy target for chronic ITP.
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Objective To detect the expression of IL-27 in PBC (primary biliary cirrhosis)patients and the possible involvement of IL-27 signal pathway in PBC. Methods The gene transcription and protein expression levels of IL-27 in patients with PBC, chronic hepatitis B(CHB) group and healthy controls(HC) were measured by real-time PCR, ELISA, flow cytometry and immunohistochemistry. AST,ALP, ALT, TBIL, GGT were determined and their correlation with IL-27 was also analyzed. Results IL-27 was significantly elevated in patients with PBC and IL-27 is present in the liver tissues of patients with PBC. Expression of IL-27 on CD4+T cells was increased in patients with PBC(72.40% ±6.22% ) and CHB(59.40% ± 7.03%) compared with HC(1.70%±0.55%,P＜0.01). Expression of IL-27 protein was increased in patients with PBC [( 126.25 ± 36.00 ) pg/ml] compared with CHB [( 51.81 ± 23.30 )pg/ml, P ＜ 0. 01] and HC[(34.19 ± 9.70) pg/ml, P ＜ 0.01], and it was positively correlated with GGT( r = 0.554, P＜0.01) and TBIL (r = 0.559,P＜0.01), but no correlation with ALT, AST, ALP.Conclusion These facts indicated the key role of IL-27 in the immune inflammatory reaction in patients with PBC.
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OBJECTIVE To develop a suitable and easy method for detecting AmpC ?-lactamases.METHODS The function with different cloxacillin concentrations in cloxacillin-potentiated disc diffusion test was compared and estimated with FOX three-dimensional extract test to show the best concentration of cloxacillin.RESULTS Cefoxitin three-dimesional test indicated that 48 strains were detected out to produce AmpC ?-lactamases among 130 clinical isolates of Gram-negative bacilli strains.when the discs contained 200 ?g cloxacillin the CAZ,CTX,CRO and ATM alone could separately detect 87.5%,87.5%,87.5% and 33.3% highly producing AmpC ?-lactamases strains.When the combination of CTX with CRO,almost 95.8% could be detected out.CONCLUSIONS With the combination of CTX and CRO,the cloxacillin-potentiated disc diffusion test can specifically detect AmpC ?-lactamases and easy to be used in clinical laboratories.