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Objective To investigate the expression level of Gas6 in patients with Immune Thrombocytopenia (ITP) and its clinical significance.Methods The experimental group was peripheral blood samples collected from 35 cases diagnosed with ITP in hematology department of Changhai Hospital in Shanghai from October 2013 to December 2015.Control group was peripheral blood from 35 healthy examined individuals at the same time.After separating plasm from the two group samples,the protein level of Gas6,IFN-α,IL-4,IFN-γ and IL-17 were measured by Enzyme-linked immunosorbent assay (ELISA),comparison of expressional level of the two groups was measured by t test.Pearson correlation analysis was used to decide the relation between Gas6 and cytokines such as IFN-α.Results The expression level of Gas6 in experimental and control groups was 27.28±7.56 ng/ml vs 20.51±5.39 ng/ml (t=4.314,P＜0.000 1);IFN-γ was 221.67±57.64 pg/ml vs 45.32 ±16.79 pg/ml (t=17.38,P＜0.000 1);IL-4 was 113.86±26.48 pg/ml vs 49.87±14.98 pg/ml (t=12.44,P＜0.000 1);IL-17 was 168.96±47.88 pg/ml vs 109.56±28.97 pg/ml (t=6.28,P＜0.000 1);IFN-α was 34.83±8.12 pg/ml vs 29.89 ± 5.76 pg/ml (t=2.936,P=0.004 5),all with statistical differences (P＜0.05).Pearson analysis showed that Gas6 was positively related with IL-17,IL-4,IFN γ (r=0.564,0.486,0.449,P＜0.05) and there was difference statistically,but Gas6 was not correlated with IFN-α.Conclusion Gas6 may participate in the disease formation of ITP through affection on cytokines secreted by Th cell subsets,and it was the potential therapeutic and predicted target for this disease clinically.
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Objective To test the expression level of CD69+CD4+CD25-T cells in peripheral blood from patients with autoimmune pancreatitis,and further analyze its clinical significance.Methods Peripheral blood samples from 32 patients with AIP diagnosed in hematological department,Changhai Hospital and 32 health individuals examined at the same time were collected from September 2014 to December 2016,they were classified as experimental and control groups,separately.Peripheral blood mononuclear cells (PBMCs) was acquired by density gradient centrifugation,CD69+ CD4 + CD25-T cells in PBMCs were tested by flow cytometry,and the expression level of cytokines in plasm was by ELISA.The comparison of varies between the two groups was measured by two independent samples' t test.The relationship between the two measurement data was measured by pearson correlation coefficient.Results The expression levels of CD69 + CD4 + CD25-T in experimental and control groups were 10.36％±3.68％ vs 3.99％±1.45％ (t=9.110,P＜0.0001).The expression level of TGF-β was 399.86±121.88 vs 143.87±56.22 pg/ml (t=10.79,P＜0.000 1),both with statistical significance.The levels of CD69+CD4+CD25-T in experimental was positively correlated with TGF-β (r=0.653,P＜0.001) and negatively with IL-4,IFN-γ,IL-2 (r=-0.442,-0.567,-0.351,P＜0.05) and there was statistical significance.Conclusion CD69+CD4 +CD25-T cells might involve the immunopathology of AIP and could be the potential biomarker for clinical diagnosis and therapy.
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Objective To test the expression of MIF in peripheral blood mononuclear cells (PBMCs)from patients with Cry-tococcal Meningitis and further discuss its clinical significance.Methods Peripheral blood from 42 patients with Crytococcal Meningitis diagnosed in Changhai Hospital,Shanghai and 42 healthy individuals examined at the same time was collected from August,2012 to November,2015.PBMCs were separated by density gradient centrifugation method,mRNA relative expression of MIF in PBMCs was measured by PCR,the level of MIF,IL-17,IL-1β,TNF-α,IFN-γand IL-4 in plasma was tested by Enzyme-linked immunosorbent assay (ELISA).The comparison for expression level of cytokines between the two groups was by two-independent samples t test.Pearson correlation coefficient was used to measure the relation between MIF and other cytokines.Results The protein levels of MIF in experimental and controlled groups were 34.17±7.88 ng/ml vs 10.89±2.76 ng/ml(t=18.07,P<0.0001),while relative expression of RNA was 2.87±0.94 vs 1.95±0.89(t=4.606,P<0.0001),and there was statistical significance (P<0.005).Pearson correlation analysis showed that MIF was positively related with IL-1β,IL-17 (r=0.467,0.401,P<0.01),with statisticaldifference.Conclusion MIF may involve in the im-mune regulation for Crytococcal Meningitis by affecting the secretion and function of cytokines as IL-1β,IL-17,and it was potential target and monitored biomarker for this disease.
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Objective To explore the change of microRNA let-7b in heat acclimatization/heat stroke rat models and its relation with HSP70,IL-6,TNF-α,TGF-3 and IL-12,and analyze its clinical significance.Methods 60 male rats with almost the same anal temperature,weight,and weeks' age were selected from Laboratory Animal Center in the Second Military Medical University.They were randomly divided into control,heat acclimatization,heat stroke groups averagely.Heat acclimatization/heat stroke rat models were built in hot climate simulated animal tank,blood was collected from cordis apex and serum was separated.Real-time fluorescent quantitative polymerase chain reaction (qRT-PCR) was used to test microRNA let-7b in serum,enzyme-linked immunosorbent reaction (ELISA) was to measure the protein level of HSP70,IL-6,TNF-α,TGF-β and IL-12 in serum.Kruskal-Wallis H test was used to analyze quantitative data in the three groups,Spearman rank correlation coefficient was used to reveal relation between two variables in heat stroke group.Results M values of miRNA let-7b in control,heat acclimatization,heat stroke groups were 0.99,1.04 and 1.93 separately,Q values were 0.30,0.25 and 0.44 (x2 =38.95,P＜0.001),separately,with statistical significance.The results of pairwise comparison showed no statistical difference in control and heat acclimatization (P＞0.05),but there were differences in control and heat stroke,heat acclimatization and heat stroke groups statistically (P＜0.05).Spearman correlation analysis showed that in heat stroke group,let-7b was positively related with HSP70,TNF-α and IL-6 (rs =0.579,0.498 and 0.609,P＜0.05) with statistical significance.Conclusion miR let-7b might involve in the pathology of heat stroke,it provided a potential biomarker for monitoring patients in high temperature and humidity clinically.
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Objective To analyze the affection and clinical significance of CD26/DPP4 on CD4+T cells and its cytokines in patients withCrytococcalMeningitis.Methods Peripheral blood was collected from 36 patients diagnosed withCrytococcal Meningitis in Changhai Hospital and Changzheng Hospital,Shanghai from August,2011 to December,2015,meanwhile 36 health controls’was also acquired.Peripheral blood mononuclear cell (PBMC)was separated by density gradient centrifuga-tion,CD26+CD4+T and CD26-CD4+T cell groups were classified by Flow Cytometry,the expression level of cytokines was tested by reverse transcriptase-polymerase chain reaction (RT-PCR).The correlation between DPP4 activity,CD26+CD4+T (%)and APACHE II score,IL-17,TNF-α,IL-4,IFN-γwas measured by Pearson coefficient.Results CD26+CD4+T(%)between experimental and control groups was 13.35±3.83 vs 8.39±2.14 (t=6.78,P<0.000 1).DPP4 activity was 50.89±17.21 mU/ml vs 73.83±20.24 mU/ml (t=5.18,P<0.000 1),with statistically significant differences.In ex-perimental groups,CD26+CD4+T (%)was positively related with APACHE II score,IL-17,TNF-α(r=0.431,0.564, 0.688,P=0.003 8,0.001,0.004 6).DPP4 activity was negatively interrelated with APACHE II score,IL-17,TNF-α,IFN-γ(r=-0.544,-0.489,-0.678,-0.734;P<0.001).Conclusion CD26/DPP4 may be involved in the pathogenesis of Crytococcal Meningitis through regulation of Th subgroups,and it was the potential therapeutic target and the predicted marker of the disease.
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Objective To investigate the IL‐17A/F and IL‐22 ,the Th17 related cytokine ,in the serum of cryptococcal meningitis patients and its clinical implications .Methods We collected peripheral blood from 22 cryptococcal meningitis patients and 20 matched healthy controls from February 2011 to February 2013 .Then the peripheral blood mononuclear cells(PBMC)were seperat‐ed with isodensity centrifugation .The RT‐PCR and ELISA were used to detect the mRNA and protein lever of Th17 related cyto‐kine such as IL‐17A ,IL‐17F and IL‐22 .In addition ,the relationship between these cytokines and cerebrospinal fluid characteristics were analyzed .Results The serum concentration of IL‐17A and IL‐22 were much higher than that of healthy controls (P<0 .01) . Their concentration were even higher in patients with poor prognosis factors ,including higher cerebrospinal fluid protein level and cryptococcal antigen titer ,lower cerebrospinal fluid glucose concentration and increased cerebrospinal fluid open pressure .Conclusion IL‐17A and IL‐22 were involved in the pathogenesis of cryptococcal meningitis and may be a potential prognosis biomarker for cryptococcal meningitis.
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Objective To investigate whether the hyper-reactivity of monocytes from the patients with primary biliary cirrhosis (PBC) to LPS and Poly I ∶ C was associated with miR-146a.Methods Peripheral blood mononuclear cells from PBC patients and healthy controls were stimulated with 10 μg/ml of LPS or Poly I ∶ C.The levels of IL-1 β,IL-6,TNF-α and IFN-α in the culture medium were detected by ELISA.The relative expressions of miR-146a,miR-21,let-7e,miR-155 and miR-125b were detected by RT-PCR.The regulatory role of miR-146a on the production of inflammatory factors in LPS or Poly I ∶ C stimulated THP-1 cells was studied by gain-and-loss function assay.Results The up-regulation of miR146a,which was induced by both LPS or Poly I ∶ C,was impaired in the monocytes from PBC patients.The production of LPS or Poly I ∶ C induced inflammatory factor,could be enhanced by miR-146a down-regulation.Conclusion The hyper-reactivity of monocytes from PBC patients to LPS and Poly I ∶ C was associated with miR-146a.
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Objective To investigate the association between IL-22 and the pathogenesis of coronary artery atherosclerosis(AS).Methods The relative expression of IL-22 mRNA in PBMC from 30 AS patients and 8 patients without any signs of coronary artery stenosis was detected by RT-PCR.Serum IL-22 levels of 22 patients without any signs of coronary artery stenosis and 79 AS patients were detected by ELISA.CRL-1730 cells(human umbilical vein endothelial cells) were stimulated with oxidized low density lipoprotein (ox-LDL) at different dosage for 24 h,and the expression of IL-22R1 was detected by flowcytometry.The proliferation ability of CRL-1730 cells treated with IL-22(20 ng/ml) and/or ox-LDL(100 μg/ml)was measured by MTS assay,and the expression of basic fibroblast growth factor(bFGF) was detected by RTPCR and ELISA.Results Decreased IL-22 expression in PBMC and serum was observed as worsen of AS.The expression of IL-22R1 in ox-LDL treated CRL-1730 cells was increased in dose dependent manner.OxLDL decreased proliferation ability,as well as bFGF expression and releasing,of CRL-1730 cells.This effect of ox-LDL was partially rescued by IL-22.Conclusion IL-22 may have anti-atherosclerosis effect.This effect may be mediated by regulating bFGF expression and endothelial cells proliferation ability in the presence of IL-22.
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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.