The effect of microRNA-145 on immune-inflammatory response of foam cells by targeting CD40 / 天津医药

Objective To investigate the effect of miRNA-145 (miR-145) on immuno-inflammatory reaction of foam cells by targeting CD40. Methods Mouse macrophage cell line RAW 264.7 cells cultured in vitro were randomly divided into model group (non-transfected), miR-145 mimics group (transfected miR-145 mimics), miR-145 inhibitor group (transfected miR-145 inhibitor) and silencing CD40 sequence group (transfected siCD40). Then oxidized low density lipoprotein (ox-LDL) was used to stimulate for 24 h to establish immune inflammatory damage cell model. Quantitative real-time polymerase chain reaction (RT-qPCR) and Western blot assay were used to detect the levels of CD 40 mRNA and protein of each group. ELISA was used to detect the levels of inflammatory factors interleukin (IL)-1, IL-6 and tumor necrosis factor (TNF) -α in cell supernatant. Results Compared with model group, the levels of CD40 mRNA, CD40 protein and IL-1, IL-6, TNF-αwere all significantly decreased in miR-145 mimics group (P<0.01). After transfected with miR-145 inhibitor, the above indexes were all significantly increased than those of model group and miR-145 mimics group (P<0.01). After transfected with CD40 siRNA, the levels of CD40 mRNA, CD40 protein and IL-1, IL-6, TNF-αwere all obviously decreased compared with those of miR-145 inhibitor group (P<0.01). Conclusion MiR-145 can regulate the immune inflammatory process of foam cells through the target gene CD40, inhibit the activation of CD40/CD40L signaling pathway and inhibit inflammatory response.

Clinical significance of CD40, Ki-67 and CD34 expressions in gastric cancer and adjacent tissues / 中国医师杂志

Objective To investigate the relationship between CD40,Ki-67,and CD34 expressions and clinical significance of gastric cancer.Methods The expressions of CD40,Ki-67,and CD34 were detected by immunohistochemistry in 80 cases of gastric carcinomas and adjacent mucosas.Results The high expression of CD40 was detected in 36.3％ (29/80) cases of gastric cancer tissues,and Ki-67 was 53.8％ (43/80).The high expression of CD40 was related to lymph node metastasis,depth of invasion,and the patient's prognosis.The high expression of Ki-67 was related to histological differentiation,depth of invasion,vascular invasion,and lymph node metastasis (P ＜ 0.05).Expression of CD40 was positively related to Ki-67 and CD34 in gastric carcinomas (P ＜ 0.05).Conclusions The high expressions of CD40,Ki-67,and CD34 in gastric cancers are related to the tumor proliferation,tumor angiogenesis,and the prognosis of patients.

Association of CD40 gene polymorphisms and serum CD40 levels with the pathogenesis of systemic lupus erythematosus / 中华皮肤科杂志

Objective To explore the association of CD40 gene single nucleotide polymorphisms (SNPs) and haplotypes with the susceptibility to systemic lupus erythematosus (SLE),as well as the association of serum levels and genotypes of CD40 with the occurrence of SLE.Methods A multiplex PCR single-base extension assay (PCR-SBE) and DNA sequencing were performed to analyze 4 SNPs of the CD40 gene,including rs1883832 C/T,rs13040307 C/T,rs752118 C/T and rs3765459 G/A,in 205 patients with SLE (SLE group) and 220 healthy human controls (control group).Enzyme-linked immunosorbent assay (ELISA) was conducted to measure serum levels of CD40 in these subjects.Results Compared with the control group,the SLE group showed significantly increased serum levels of CD40 (P ＜ 0.05).There were significant differences in genotype and allele frequencies of the SNP rs1883832 C/T in the CD40 gene between the SLE group and control group (all P＜ 0.01).Relative risk analysis showed that the risk of developing SLE in rs1883832 T allele carriers was 1.517 times that in rs1883832 C allele carriers (OR =1.517,95％ CI:1.157-1.990,P=0.003).Moreover,serum levels of CD40 were significantly higher in rs1883832 T allele carriers than in rs1883832 C allele carriers (P ＜ 0.01).The risk of developing SLE was significantly increased in TCCA haplotype carriers compared with the healthy controls (OR =2.322,95％ CI:1.181-4.564,P=0.012).Conclusion The CD40 gene rs1883832 C/T polymorphism and its TCCA haplotype were both associated with the occurrence of SLE,and the rs1883832 T allele may be a gene predisposing to SLE.

Effect of CD40 siRNA on inflammatory response of MRL/Lpr mice / 北京大学学报(医学版)

Objective:To observe the effect of CD40 siRNA on expression of IFN-γ,IL-17,IL-4 and anti-dsDNA antibody of systemic lupus erythematosus (SLE)animal model MRL/Lpr mice and to discuss its therapy on MRL/Lpr mice.Methods:In the study,16 female MRL/Lpr mice were randomly divided into control group (n =4),empty vector group (n =4),CD40-siRNA1 group (n =4)and CD40-siR-NA2 group (n =4).The vectors expressing siRNA against CD40 were injected by tail veil into MRL/Lpr mice,while MRL/Lpr mice in control group and empty vector group were injected with the same dose of PBS and pGFP-V-RS vector respectively.The injection was given six times and every one day.The mice were sacrificed 14 d after injection,and the spleen tissue was weighed.The pGFP-V-RS was labeled by green fluorescent protein(GFP)and the tissue sections were observed whether siRNA expressed in the spleen.The expression levels of IFN-γ,IL-17,IL-4 and anti-dsDNA antibody in the sera were detected by ELISA method on the 1st day before the first time and the 2nd,5th,8th,11th,and 14th days after last injection,and the expression levels of CD40 mRNA in spleen tissue of MRL/Lpr mice were detected by RT-PCR and the expression levels of CD40 protein in spleen tissue of MRL/Lpr mice were detected by immunohistochemistry method.Results:The expression vector of CD40-siRNA could express in the spleen of MRL/Lpr.The spleens in CD40-siRNA1 group [(78.85 ±5.61 )mg]and CD40-siRNA2 group [(80.25 ±4.07)mg]were lower than those in control [(141.88 ±7.81)mg]and empty vector group [(153.10 ±7.60)mg].The levels of IL-17,IFN-γand anti-dsDNA antibody were lower and the levels of IL-4 was higher in CD40-siRNA1 group and CD40-siRNA2 group on the 2nd,5th and 8th days after last injection than on the 1st day before the first time (P 0.05). Though the levels of anti-dsDNA antibody in CD40-siRNA1 group and CD40-siRNA2 group on the 11th day was higher than on the 8th day,there was more significance than those in control group and empty vector group (P <0.05).There was no significance between the 4 groups on the 14th day.The levels of CD40 mRNA and protein were lower in CD40-siRNA1 group and CD40-siRNA2 group than in control group and empty vector group on the 14th day after last injection (P <0.05).Conclusion:CD-40 si-RNA can reduce the concentration of IL-17,IFN-γand of anti-dsDNA antibody in serum,and at the same time,it can elevate the concentration of IL-4 and suppress CD40 mRNA and protein of spleen in MRL/Lpr.Meanwhile after suppressing CD40 mRNA and protein,it can reduce inflammatory response of the mice and the disease activity of MRL/Lpr,suggesting that CD-40 siRNA has therapy effect on SLE.

The association between the polymorphisms in the CD 40 gene and Graves′disease:a Meta analysis / 重庆医学

Objective To investigate the associations between the polymorphisms in CD40‐1C/T polymorphism(rs1883832) and Graves′disease by Meta‐analysis .Methods We searched Pubmed ,CBM ,VIP ,CNKI ,Cochrane library and Wanfang ,covering and selecting literatures according to criterions .Statistical analysis was performed by using Revman5 .2 and Stata12 .0 .Results A total of 18 eligible literatures were included(5 198 cases and 4 417 controls) ,The pooling results suggested that significant differ‐ence in allele C/T frequency and genotype frequency were observed between patients with and control subjects (P0 .05) .Conclusion This Meta analysis indicated that the polymorphisms in the CD40 gene(rs1883832) was related with Graves′disease ,but had nothing to do with family history and Graves′disease with ophthalmopathy .

The influence of CsA in CD40/CD40 ligand expression of T lymphocytes in patients with coronary artery disease / 天津医药

Objective To investigate the effect of CD40/CD40 ligand on the genesis and development of coronary artery disease (CAD),and the inhibitory effect of cyclosporine A (CsA) on CD40/CD40 ligand. Methods A total of 71 patients were divided into four groups:acute myocardial infarction group (AMI, n=19), unstable angina pectoris group (UAP, n=18), stable angina pectoris group (SAP, n=17) and normal control group (N, n=17). Flow cytometry was used to detect the expres?sion of CD40 and CD40L in peripheral blood mononuclear cells (PBMCs) of four groups. The group in which CD40 and CD40L were produced at the highest level was chosen, and a series concentrations of CsA(H1:0 mg/L, H2:0.01 mg/L, H3:0.1 mg/L, H4:1 mg/L)were used to treat the cells. Then the expressions of CD40 and CD40L were measured by flow cytome?try. Results Compared with N group,the expression of CD40 was significant higher in other groups (P0.05). The expression of CD40L was elevated and fol?lowed by different severity of CAD. There was significant difference in the expression of CD40L between groups (P<0.05) . AMI group showed the highest expression of CD40 and CD40L. After being treated with CsA, the expression of CD40 was higher in H1 group than that of H3 group and H4 group (P<0.05). The expression of CD40L was significantly higher in H1 group than that of other three groups (P < 0.05). Conclusion CD40 and CD40L may be involved in the development of CAD. Moreover, it might be restrained by CsA via regulation of CD40/CD40L.

CD40/CD40L system and atherosclerosis / 国际脑血管病杂志

Atherosclerosis is a chronic inflammatory disease.Atherosclerotic plaque rupture and thrombosis may result in cardio-cerebrovascular events.Inflammatory mediator CD40L widely exists in cells associated with atherosclerosis.They participate in plaque inflammatory response,release proinflammatory cytokines,degrade extracellular matrix,improve procoagulant activity,and promote the progression of atherosclerosis and plaque vulnerability.Intervening CD40/CD40L signaling system may become an effective treatment strategy to slow the progress of atherosclerosis and stabilize atherosclerotic plaques.

Correlation between CD40-1C/T gene polymorphism and serum sCD40L / 中国医师杂志

Objective To investigate the correlation between CD40-1C/T gene polymorphism and serum soluble CD40L (sCD40L) expression in cerebral infarction.Methods According to the inclusion and exclusion criteria,select the acute large artery atherosclerosis in patients with cerebral infarction as the case group,select the same period without a history of stroke examination subjects as the control group.Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technology was used to detect CD40 gene polymorphism and sequencing,double-antibody sandwich enzyme-linked immunosorbent assay (ELISA) was used to detect the expression levels of serum sCD40L.Results A total of 209 patients with large artery atherosclerotic cerebral infarction (case group) and 87 subjects without history of stroke (control group) were included.The CC genotype (31.6％) and C allele frequency (53.8％) of the case group were significantly higher than that of the control group (17.2％,39.1％) (x2 =6.94,10.69,P ＜0.01).Case group serum sCD40L expression level was significantly higher than those in the control group [(3.97 ± 1.20) vs (2.69 ±0.88)] (t =10.19,P ＜0.001).Case group serum sCD40L expression level was different among CC,CT,and TT genotypes (F =19.22,P ＜0.001),serum sCD40L level (4.55± 1.16) of CC genotype in patients was significantly higher than that of CT (3.93 ± 1.17) and TT genotypes (3.27 ± 0.90),serum sCD40L level (3.93 ± 1.17) of CT genotype in patients was significantly higher than the TT genotype (3.27 ±0.90).The serum sCD40L expression level of the control group in the CC,CT,TT has no statistically significant differences among various genotypes [(2.91 ±0.79),(2.67 ±0.89),(2.61 ±0.91),F =0.619,P =0.541).Conclusions CD40-1C/T gene polymorphism is associated with serum sCD40L expression level in cerebral infarction,serum sCD40L level of the CC genotype was significantly higher.

Correlation between CD40 -1C/T polymorphism and large artery atherosclerotic stroke / 国际脑血管病杂志

Objective To investigate the correlation between CD40 gene promoter region - 1C/T polymorphism and carotid atherosclerotic plaques and large artery atherosclerotic stroke.Methods The subjects were the patients with acute large artery atherosclerotic stroke (patient group) and the patients without history of stroke (control group).The patient group was further divided into an unstable plaque subgroup,a stable plaque subgroup,and a plaque-free subgroup.Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was used to detect CD40 - 1C/T polymorphism.Results A total of 170 patients with large artery atherosclerotic stroke (patient group) and 61 subjects without history of stroke (control group) were included.In the patient group,51 patients were in the unstable plaque subgroup,60 were in the stable plaque subgroup,and 59 were in the plaque-free subgroup.C allele frequency of the patient group was significantly higher than that of the comrol group (52.9％ vs.38.5％ ;x2 =7.466,P =0.006).In the patient group,C allele frequency of the unstable plaque subgroup (75.5％) was significantly higher than that of the stable plaque subgroup (53.3％),and both of them were significantly higher than that of the plaque-free subgroup (33.1％ ) (the stable plaque subgroup vs.the plaque-free subgroup:x2 =9.970,P =0.002; the unstable plaque subgroup vs.the stable plaque subgroups:x2 =11.680,P =0.001; the unstable plaque subgroup vs.the plaque-free subgroup:x2 =39.532,P=0.000).Multivariate logistic regressive analysis showed that hypertension (OR9.513,95％ CI 1.291 - 20.779; P =0.028),increased total cholesterol level (OR 4.235,95％ CI 1.069 -19.034; P =0.032),increased low density lipoprotein level( OR 4.201,95％ CI 1.803 - 9.672; P =0.001 )and C alleles (OR 1.759,95％ CI 1.177 - 2.738; P =0.006) are the independent risk factors of large artery atherosclerotic stroke.Conclusions CD40 - 1C/T polymorphism is associated with the risks of carotid atherosclerotic plaque formation,unstable plaque and large artery atherosclerotic stroke; C allele may be a susceptibility factor for large artery atherosclerotic stroke.

Mice islet allograft tolerance induced by blockade of OX40/OX40L and CD40/CD154 costimulation pathway / 中华器官移植杂志

Objective To investigate the effects of blockade of OX40/OX40L costimulation pathway on mice islet allograft tolerance in CD40/CD154 costimulation pathway blockade mice.Methods C57BL/6 mice were induced into diabetes mellitus as recipients,and were transplanted with DBA/2 mice islets.The recipients were divided into four groups,(1) treated with IgG as controls,(2) anti-OX40L mAb,(3) anti-CD154,(4) combined treatment of anti-OX40L mAb and anti CD154mAb.The mean survival time (MST) of islet allograft was observed.The expression of OX40 in activated T cells of CD154 deficient mice was detected.Effector T cells were obtained from the spleen of CD154 deficient mice cultured with or without anti-OX40L mAb for 3 days.The proliferation of T cells was assayed.Results The MST in the control group,anti-OX40L mAb group,anti-CD154 mAb group and anti OX40L mAb + anti-CD154 mAb group was 19,22,48,and ＞150 days respectively (P ＜0.05).The OX40 expression was readily induced in the 66％ activated T effector cells.CD154 deficient T effector cells proliferation was inhibited by the addition of anti-OX40L mAb in the culture in a dose-dependent fashion.Conclusion The blockade of OX40/OX40L costimulation pathway can promote islet allograft tolerance in CD40/CD154 costimulation pathway blockade mice by inhibiting the proliferation of T cells.

Increased p190RhoGEF expression in activated B cells correlates with the induction of the plasma cell differentiation

Previously, we demonstrated that the p190 Rho guanine nucleotide exchange factor (p190RhoGEF) was induced following CD40 stimulation of B cells. In this study, we examined whether p190RhoGEF and a downstream effector molecule RhoA are required for B cell differentiation. Expression of p190RhoGEF positively correlated with the expression of surface markers and transcriptional regulators that are characteristic of mature B cells with plasma cell (PC) phenotypes. Moreover, either the overexpression of p190RhoGEF or the expression of a constitutively active RhoA drove cellular differentiation toward PC phenotypes. B cell maturation was abrogated in cells that overexpressed p190RhoGEF and a dominant-negative form of RhoA simultaneously. CD40-mediated maturation events were also abrogated in cells that overexpressed either dominant-negative p190RhoGEF or RhoA. Together, these data provide evidence that p190RhoGEF signaling through RhoA in CD40-activated B cells drives the induction of the PC differentiation.

Association between plasma levels of soluble leukocyte differentiation antigens CD40/CD40 ligand and kidney damage in preeclamptic patients / 中华妇产科杂志

Objective To investigate the variance levels of plasma soluble leukocyte differentiation antigens CD40 (sCD40) and soluble CD40 ligand (sCD40L) in preeclamptic patients with renal damage and its relationship. Methods A total of 63 pregnant women attended the Department of Obstetrics, Affiliated Hospital of Qingdao University Medical College between August 2008 and June 2010. In the present study included 28 pregnant women with mild preeclampsia and 35 patients with severe preeclampsia. Thirty matched normotensive pregnant women were enrolled in the study as the control group. Expression of sCD40 and sCD40L were determined by ELISA. At the same time, the blood routine, C reaction protein ( CRP),urine routine, 24 hours urine protein excretion, and serum uric acid (UA), creatinine (Cr), blood urea nitrogen (BUN) were measured. The correlation analysis was performed between the sCD40/sCD40L and the blood biochemical indexes in 3 groups. Results ( 1 ) The median levels of CRP in severe preeclampsia (10. 8 mg/L)and mild preeclampsia group(7. I mg/L)are significantly higher than that of control group (3. 3 mg/L,P ＜ 0. 05 ); The level of CRP in severe preeclampsia group was also higher than that of mild preeclampsia group ( P ＜ 0. 05 ). The median gestational age at delivery in severe preeclampsia ( 32. 5 weeks)was significantly less than that of mild preeclampsia group ( 37. 2 weeks) and normal group ( 38. 6 weeks,P ＜ 0. 05). However no significant differences were observed between mild preeclampsia group and normal group ( P ＞0. 05 ). The platelet count in severe preeclampsia ( 132 × 109/L) was significantly less than those of mild preeclampsia group (212 × 109/L) and normal group ( 216 × 109/L, P ＜ 0. 01 ), but no significant differences were observed in blood platelet amount between mild preeclampsia group and normal group ( P ＞0. 05 ). There was no significant difference in hemoglobin level and white blood cell in three groups ( P ＞0. 05). (2) The sCD40 plasma concentration in severe, mild preeclampsia and normal group was 133.6,126. 5 and 90. 7 ng/L, respectively. The sCD40 L plasma concentrations were 12. 5, 10. 4 and 4. 4 ng/L respectively in the 3 groups. 24 hours urinary protein quantitative was 4. 5 g/d,0. 8 g/d and 0 in the 3 groups respectively. And the UA level was 486 μ mol/L,289 μmol/L and 162 μmol/L. In the above three groups,the monitoring indicators were significantly higher in women with severe preeclampsia group compared with mild preeclampsia and control groups (P ＜ 0. 01 ), and there were also higher in mild preeclampsia group than that in control groups ( P ＜ 0. 01 ). The level of plasma Cr ( 89 μmol/L) and BUN ( 5. 32 mmol/L) in severe preeclampsia group were higher than those of mild preeclampsia group (66 μmol/L and 4. 49mmol/L) and control group ( 57 μmol/L and 3.32 mmol/L, P ＜ 0. 05 ). There was no significant difference between mild preeclampsia group and normal group (P ＞ 0. 05 ). (3) The correlation analysis indicated that the level of sCD40 has a positive correlation with 24 hours urinary protein quantitative( r = 0. 434, P ＜ 0. 05 ),also significant positive correlation( r =0. 536,0. 528 ,P ＜ 0. 01 ) between the level of sCD40 and UA or CRP in women with preeclampsia. There was no significant correlation between the level of sCD40 and systolic blood pressure, diastolic blood pressure, delivery gestational age, Cr, BUN, and platelet count(r =0. 135,0. 183, -0. 133,0. 190,0. 167, -0. 221 ,all P ＞0. 05 ). There were positive correlation between the level of sCD40L and 24 hours urine protein excretion, either UA or CRP( r =0. 591,0. 445,0. 539 ,all P ＜0. 01 ). No significant correlation was found between sCD40 L and systolic blood pressure, diastolic blood pressure,delivery gestational age, Cr, BUN, and platelet count( r =0. 178,0. 212, -0. 292,0. 144,0. 135, -0. 273,all P ＞0. 05). There was significant positive correlation between plasma sCD40 and sCD40L ( r =0. 707 ,P ＜0. 01 ). There was no relationship between the level of sCD40, sCD40L and the blood biochemical indexes in normotensive pregnant women ( P ＞ 0. 05 ). Conclusions The plasma concentrations of sCD40 and sCD40 L are significantly higher in pregnant women with preeclampsia compared with the control, which may be involved in the development of preeclampsia and contribute to the kidney damage. The variance levels of sCD40 and sCD40L may be also related to the severity of preeclampsia.

Influence of CTLA4Ig and CD40Ig gene local co-transfection on the survival of xenogenic renal allografts / 中华器官移植杂志

Objective To observe the efficacy of CTLA4Ig and CD40Ig gene local cotransfection on the survival of renal allografts.Methods The kidneys of guinea pig were transfected with PcDNA 3.1+-CTLA4Ig and/or PcDNA 3.1+ -CD40Ig gene by Lipo2000,and the transfected kidneys were transplanted to SD rats.The recipients were divided into group 1 (transfected with PcDNA 3.1 +,mock group),group 2 (transfected with PcDNA 3.1 + -CD40Ig,CD40Ig group),group 3 (transfected with PcDNA3.1+ -CTLA4Ig,CTLA4Ig group) and group 4 (co-transfected with PcDNA 3.1+-CD40Ig and CTLA4Ig,CTLA4Ig+ CD40Ig group).The effects of CTLA4Ig and CD40Ig transfection were determined by Western blotting.The serum creatinine (Scr) in the recipients,the pathological changes of the allografts and the survival of renal allografts were observed.Results Significantly prolonged allografts survival time was observed in group 2 (40.7 ± 10.9 days),group 3 (49.3 ± 9.5 days) and group 4 (75.7 ± 8.0 days) as compared with group 1 (6.8 ± 1.9days),especially allografts got the longest survival time in group 4 (75.7± 8.0 days).The serum creatinine level was reduced in group 4 at 30th day as compared with that in group 2 and group 3.The lymphocytes infiltrating rate in the grafts was lowest in group 4 after transplantation.Conclusion Local co-transfection of CTLA4Ig and CD40Ig genes can prolong the survival time of renal allografts significantly.

Ligation of CD40 receptor in human B lymphocytes triggers the 5-lipoxygenase pathway to produce reactive oxygen species and activate p38 MAPK

Previously, we reported that CD40-induced production of reactive oxygen species (ROS) by NADPH oxidase requires the TNF receptor-associated factor (TRAF) 3, as well as the activities of phosphatidylinositol 3-kinase (PI3K) and Rac1. Here we investigated the possible mechanisms of the production of ROS after CD40 ligation in B cells. We describe an alternative ROS production pathway that is triggered by CD40 ligation, involves 5-lipoxygenase (5-LO), and results in activation of p38 MAPK. Our studies in Raji human B lymphomas revealed that CD40-induced ROS production by 5-LO also requires the activities of PI3K and Rac1. In contrast to the NADPH oxidase pathway, however, TRAF molecules are not required for the CD40-induced ROS production by 5-LO. The association of CD40 with 5-LO is dependent on CD40 ligation in Raji B cells, and co-immunoprecipitation experiments using epitope-tagged proteins transiently expressed in human embryonic kidney 293T cells revealed the role of the regulatory subunit of PI3K, p85, in this association. Collectively, these data suggest a separate pathway for the CD40-induced ROS production in B cells and demonstrate that this pathway requires 5-LO via direct association of p85 with both CD40 and 5-LO.

CD40/CD40L system and atherosclerosis and cerebral infarction / 国际脑血管病杂志

Cerebral artery atherosclerosis is the main reason leading to cerebral infarction. CD40/CD40L overexpression will stimulate the immune and inflammatory responses,leading to local inflammatory cell infiltration within the atherosclerotic plaque, triggering plaque rupture, and thus causing cerebral infarction. Studies have suggested that that CD40L is associated with the severity of cerebral infarction, Therefore, clinical detection of CD40L can be used as an indicator for identifying the severity of cerebral infarction to guide clinical treatment.

Effects of Tongxinluo capsule on endodermis function and the expression of CD40, CD40L in artherosclerosis rabbits / 中国基层医药

Objective To investigate the therapeutic effects of Tongxinluo and its influence on the expression of CD40 and CD40L in artherosclerosis rabbits. Methods 60 male rabbits were divided into three groups randomly,20 of each group: normal control group, model group and Tongxinluo group. The rabbits in model group and Tongxinluogroup were used to prepare the atherosclerosis model induced by high-cholesterol diet. The levels of ET-1, NOwere measurde. Before and 12 weeks after durg treatment,serum total cholesterol ,plaque areas and ratios of intima/media thickness were detected. The expression of CD40 and CD40L mRNA were determinated by quantitive RT-PCR. Results Serum total cholesterol level of Tongxinluo group was decreased compared with model group ( P < 0. 05 ), and they were both much higher than those of normal control group ( P < 0. 01 ); as compared with the model group, aortic plaque areas and ratios of intima/media thickness in Tongxinluo group reduced significantly [ (0.56 ± 0. 07) vs ( 1.16±0.08),P<0.01;(36.88±2.38)% vs (76.58 ±2.86) %,P <0. 0l] ;The expression of CD40 and CD40L mRNA also decreased in Tongxinluo group with statistic significance[ (0.798 ± 0.115 )、 (0. 592 ± 0. 132) vs (0.686±0. 132) 、(0.498 ±0. 108) ,P <0. 01 ) ]. Conclusion Tongxinluo had the ability to anti-artherosclerosis and its possible mechanism was down-regulate the expression of CD40 and CD40L.

Rosiglitazone down-regulates lipopolysaccharide-induced expression of CD40 and intercellular adhesion molecule 1 in rat peritoneal mesothelial cells through a NF-κB dependent mechanism / 中华肾脏病杂志

Objective To investigate the effect and mechanism by which PPARγ ligand, rosiglitasone, regulates the expression of CD40 and intercellular adhesion molecule 1 (ICAM-1) in the rat peritoneal mesothelial cells (RPMCs) induced by lipopolysaccharide (LPS). Methods RPMCs were harvested from Sprague-Dawley rat peritoneal cavity and maintained under defined in vitro conditions. The cells were randomly divided into groups as follows: medium, LPS (5 mg/L), LPS (5 mg/L)+BAY11-7085(5 μmol/L, NF-κB inhibitor), rosiglitazone (10 μmol/L or 20 μmol/L, peroxisome proliferator-activated receptor γ activator), LPS (5 mg/L)+rosiglitazone (10 μmol/L)+GW9662 (3 μmol/L, peroxisome proliferator-aetivatcd receptor γ antagonist), and LPS (5 mg/L)+vehicle (DMSO 0.2 ml/L). The expressions of CD40 and ICAM-1 RNA in RPMCs were examined by RT-PCR after 3 hour treatment, and the protein expressions of CD40, ICAM-1, p-NF-κB p65 and p-IκBα were examined by Western blot or immunofluorescence after 24 hour treatment. Results Following treatment with LPS, both the expressions of CD40 and ICAM-1 protein in RPMCs were up-regulated significantly (P<0.05), and the phosphoralation of p65 was increased greatly (1.10±0.17 vs 0.55±0.06, P<0.05). BAY11-7085 (5 μmol/L) significantly decreased the protein expression of p-p65 (0.22±0.11 vs 1.10±0.17, P<0.01), CD40 (0.34±0.02 vs 0.50±0.06, P<0.05) and ICAM-1 (0.35±0.16 vs 0.74±0.03, P<0.05). Pretreated with rosiglitazone for 3 h then added with LPS for 1 h, the levels of p-p65, CD40 and ICAM-1 in RPMCs were significantly decreased compared with those of LPS group (0.77±0.08 vs 0.90±0.10, P<O.01; 0.79±0.16 vs 0.99±0.06, P<0.05; 0.83±0.20 vs 1.22±0.13, P<0.05). However, for the group pretreated with rosiglitazone(10 μmol/L) and GW9662(3 μmol/L) for 3 h then added with LPS, the levels of p-p65 in RPMCs did not change significantly compared with those of rosiglitazone-pretreated group. The expressions of CD40 and ICAM-1 in RPMCs were significantly increased compared with those of rosiglitazone-pretreated group (0.95±0.19 vs 0.79±0.16, and 1.04±0.24 vs 0.83±0.20, P<0.05). Conclusion Rosiglitazone can decrease LPS-induced expression of CD40 and ICAM-1 in RPMCs by inhibition of NF-κB activation, which suggests that rosiglitazone may mediate its antiinflammatory effect through a NF-κB dependent mechanism.

The roles of CD40-CD40L in carotid atherosclerotic plaque stability and their correlation with matrix metalloproteinase 9 / 中华神经科杂志

Objective To investigate the expression of CD40,CD40L and MMP9 in carotid atherosclerotic plaques and evaluate their roles in carotid atherosclerotic plaque stability.Methods Carotid atherosclerotic plaques were isolated in carotid eversion endarterectomy (GEE) in 37 patients with high-grade stenosis (>70%) including 20 stroke (A group) and 17 non-stroke patients (B group).The control group included samples of normal carotid artery from 11 normal individuals,The RNA expression levels of CD40,CD40 L and MMP9 in all A,B and control groups were quantitatively detected by real-time quantification polymerase chain reaction (PCR) and the protein expression levels were detected by Western blotting analysis.The expression and distribution of CD40,CD40L and MMP9 in carotid atherosclerotic plaques were detected by immunohistochemistry staining.Then correlations between CD40-CD40L and MMP9 were statistically analyzed.Results The relative CD40 mRNA level in high-grade stenosis of A group,B group and normal control were 2.41±0.43,1.03±0.38 and 0.31±0.12,respectively,and MMP9 mRNA 6.88±1.57,1.90±0.44 and 0.39±0.12,respectively.The levels of CD40 and MMP9 mRNA in A group were significantly higher than those in B group (P=0.000),the levels of CD40 and MMP9 in B group were significantly higher than those in controls (P=0.000).There was a linear correlation between CD40 and MMP9 mRNA (r=0.929,P=0.000).However,there were no significantly difference in mRNA levels of CD40L between carotid atherosclerosis and controls.The protein expression levels of CD40,CD40L and MMP9 in A group were significantly higher than those in B group (FCD40=104.100,P=0.000;FCD40L=129.932,P=0.000;FMMP9=13.565,P=0.021) and B group higher than normal controls (FCD40=115.848,P = 0.000;FCD40L= 30.482,P=0.005;FMMP9=35.557,P=0.004).The areas of positive staining of CD40,CD40L and MMP9 in immunochemistry study in A group were significantly higher than those in B group and B group was significantly higher than controls.There were linear correlations between positive staining areas Of CD40 and CD40L,CD40 and MMP9,CD40L and MMP9 (r=0.963,0.959,0.929,P=0.000).Expressions of CD40,CD40L and MMP9 were significantly higher in the shoulder areas of the atherosclerotic plaques than in other areas.Conclusions The CD40-CD40L has an important role in the formation of carotid atherosclerosis and plaque instability,probably by up-regulating MMP9.The expression of CD40L may be regulated by post-transcriptional modification to exert biological effects.

Interação entre as vias de sinalização CD40/CD40L e os PPARs / Interections between CD40/CD40L and PPARs signaling pathways

O receptor CD40 e seu ligante CD40L possuem um papel importante na interface entre a resposta imune inata e a adaptativa. Disfunções desta via de sinalização são descritas em doenças de origem inflamatória e autoimunes. Em Lúpus eritematoso sistêmico (LES) foi descrito um aumento nos níveis séricos de CD40L solúvel, que participa na produção de autoanticorpos. Receptores ativados por proliferadores de peroxisomos (PPARs) são fatores de transcrição que inicialmente foram descritos como envolvidos apenas no metabolismo lipídico, mas que atualmente são também descritos como atuantes no controle da resposta imune. Com isso, nosso objetivo é determinar se a ativação dos PPARs modula o processo inflamatório através da interação com CD40/CD40L in vitro ou in vivo. Células de linhagem monocítica humana THP-1 foram tratadas por 24 horas com forbol-éster (PMA, 40 nM) e posteriormente estimuladas com CD40L recombinante (rhCD40L, 1 g/ml) por diferentes períodos. Transcritos de mRNA foram analisados por real time PCR e os resultados expressos como razão da expressão do gene housekeeping GAPDH. As células THP-1 apresentam um aumento na expressão de PPAR e após 16 e 2 horas de estímulo com rhCD40L, respectivamente. Estas células também foram estimuladas com LPS (10 g/ml) e LPS+rhCD40L para sabermos se a resposta obtida anteriormente era específica ao estímulo com rhCD40L. O resultado mostra que há uma diminuição na expressão de PPAR e após o estimulo com LPS ou LPS+rhCD40L, indicando que nessas condições a modulação da expressão de PPARs é especifica para a via de sinalização CD40/CD40L. Foi medida também a expressão de CD36, que é descrito na literatura como um indicador da atividade de PPARs. O resultado mostra que o estímulo com CD40L promove um aumento de CD36, o que indica indiretamente que o PPAR estava ativo neste modelo experimental. Para mostrar a interação direta destas duas vias de sinalização, silenciamos o gene de PPAR por siRNA e posteriormente anlisamos...

The membrane receptor CD40 and its ligand CD40L play an important role in the interface between innate and acquired immunity. Dysfunction of this signaling pathway was described in inflammatory and autoimmune diseases. In systemic lupus erythematosus (SLE), increased serum levels of soluble CD40L have been detected, where it plays a significant role in the generation of auto-antibodies. Peroxisome proliferator activator receptors (PPARs) are transcription factors originally described in lipid metabolism. More recently, they were also characterized as inflammatory modulators. Therefore, our objective was to determine whether the activation of PPARs may modulate the inflammatory process through interaction with the CD40/CD40L signaling pathway in vitro and in vivo. Macrophages derived from the human monocytic cell line THP-1 by 24h-treatment with PMA (40 nM) were stimulated with human recombinant CD40L (rhCD40L, 1 g/ml) for different periods. Messenger RNA (mRNA) transcripts for PPAR , and were determined by real time PCR and expressed as a ratio of the housekeeping gene GAPDH transcripts. THP-1 cells express a basal level of PPAR and gene transcription, which is increased 16 and 2 hours after exposure to rhCD40L, respectively. We also stimulated the THP-1 cells with LPS (10 g/ml) and LPS+rhCD40L to see if the increase of PPAR was a response specific to the rhCD40L stimuli. The data show that there is a decrease in PPAR and genes expression upon LPS or LPS+rhCD40L stimulation, indicating that in these times (2 and 16 hours) the response is specific for the CD40/CD40L signaling pathway. Increased expression of CD36 is known as an indicator of PPARs activity. We measured CD36 and saw an increase of this receptor after rhCD40L stimulus, indicating indirectly that PPARs were active in this experimental model. To prove the direct interaction between CD40/CD40L and PPAR , we silenced the PPAR gene by siRNA and analyzed the expression of CD80, which is known...

The expression of CD40 and CD40L on the surface of peripheral blood mononuclear cells in asthmatic rats and the effect of antiCD40L McAb on Th1 and Th2 cytokines / 中国医师杂志

Objective To investigate the expression of CD40 and CD40L on the surface of peripheral blood mononuclear cells(PBMCs)in asthmatic rats and the effect of anti-CD40L McAb on cytokines of it.Methods Flow cytometry and RT-PCR were used to detect the expression of CD40 and CD40L of PBMCs ih asthmatic rats.After the PBMCs Was treated with anti.CIMOL McAb.ELISA was used to detect the levels of IL-4 and IFN-γin the supematants of cultured cells.Results Compared with the normal control group.the expression of CD40 and CD40L of PBMCs in asthImatic rats increased(P＜0.05).Compared with the untreated group,the level of IL-4 and the ratio of IL4/IFN-γ decreased after the PBMCs were treated with anti-CD40L McAb(P＜0.05).Conclusion The expression of CD40 and CD40L on the surface of PBMCs in asthmatic rats Was unregulated.Anti-CD40L Mcab Can decrease the level of IL-4 and the ratio of IL_4/IFN-γ.