Interleukin-1 receptor antagonist (IL-1ra), tumor necrosis factor soluble receptors (sTNF-R) type I and II, and regulated upon activation, normal T-cell expressed and secreted (RANTES) play an important role in the modulation of primary glomerulonephritis (GN) course. The aim of the study was to assess whether pre-treatment measurements of IL-1ra, sTNF-R, and RANTES assessed conjointly may be useful as predicting factors in patients with GN. In 84 patients (45 males and 39 female) serum concentration (pg/mL) and urinary excretion (pg/mgCr) of cytokines were measured. After 12 months of therapy with steroids and cyclophosphamide the patients were divided into two subgroups: Responders (R) and Non-Responders (NR) according to the treatment results. The urinary IL-1ra, TNF-RI and RII were significantly higher in R than NR (1,732 vs 646 with P < 0.001, 13.1 vs 6.3 with P = 0.005, and 33.6 vs 14.4 with P = 0.012). The urinary RANTES excretion was increased in NR (79.6 vs 28.5; P < 0.001). The multivariable analysis showed that if conjointly assessed, only urinary IL-1ra, TNF-R I and R II, RANTES with 85% probability pointed the feature remission (R). In conclusion, the urinary excretion of IL-1ra, TNF-R I and R II, and RANTES examined conjointly are effective in predicting favorable response to immunosuppressive treatment in patients with GN.
Adult ; Cyclophosphamide/therapeutic use ; Female ; Glomerulonephritis/drug therapy/*metabolism/pathology ; Humans ; Immunosuppressive Agents/therapeutic use ; Interleukin 1 Receptor Antagonist Protein/*analysis/blood/urine ; Lymphocyte Activation ; Male ; Middle Aged ; Multivariate Analysis ; Predictive Value of Tests ; Receptors, Tumor Necrosis Factor, Type I/*analysis/blood/urine ; Receptors, Tumor Necrosis Factor, Type II/*analysis/blood/urine ; Steroids/therapeutic use ; T-Lymphocytes/immunology/metabolism

PURPOSE: Interleukin-1 (IL-1) plays a key role in inflammation and immunity and its decoy receptor, IL-1R2, has been implicated in transcriptomic and genetic studies of asthma. METHODS: Two large asthma family collections, the French-Canadian Saguenay-Lac-St-Jean (SLSJ) study and the French Epidemiological Study on the Genetics and Environment of Asthma (EGEA), were used to investigate the association of SNPs in 10 genes that modulate IL-1R2 activities with asthma, allergic asthma, and atopy. Gene-gene interactions were also tested. RESULTS: One SNP in BACE2 was associated with allergic asthma in the SLSJ study and replicated in the EGEA study before statistical correction for multiple testing. Additionally, two SNPs in the MMP2 gene were replicated in both studies prior to statistical correction and reached significance in the combined analysis. Moreover, three gene-gene interactions also survived statistical correction in the combined analyses (BACE1-IL1RAP in asthma and allergic asthma and IL1R1-IL1RAP in atopy). CONCLUSIONS: Our results highlight the relevance of genes involved in the IL-1R2 activity in the context of asthma and asthma-related traits.
Asthma ; Epidemiologic Studies ; Genetics ; Humans ; Inflammation ; Interleukin-1* ; Phenotype* ; Polymorphism, Single Nucleotide ; Receptors, Interleukin-1 Type II*

OBJECTIVETo construct pPICZalphaA-soluble interleukin-1 receptor type I (sIL-1RI) recombinant expression vector containing the gene fragment encoding the extracellular domain of sIL-1RI for its expression in Pichia pastoris.

METHODSsIL-1RI gene was amplified by RT-PCR and inserted into the yeast expression vector pPICZalphaA by digestion ligation. The recombinant plasmid pPICZalphaA-sIL1RI was transformed into E.coli Stb13, and the positive clones were analyzed by PCR and DNA sequencing. The pPICZalphaA-sIL1RI recombinant plasmid was electroporated into GS115 cells and the transformants were analyzed by PCR. After phenotype identification, the recombinant strains were induced by methanol to express the target protein, which was analyzed by Western blotting of the cell extract and supernatant.

RESULTSThe recombinant plasmid pPICZalphaA-sIL-1RI was constructed successfully, and the results of Western blotting showed that yeast induced by methanol expressed a protein of about 39 kD.

CONCLUSIONsIL-1RI protein has been successfully expressed in P.pastoris expression system, which provides the basis for further study of sIL-1RI.

Escherichia coli ; metabolism ; Gene Expression ; Genetic Vectors ; Humans ; Pichia ; metabolism ; Plasmids ; Receptors, Interleukin-1 Type I ; biosynthesis ; genetics

OBJECTIVETo isolate and characterize human rheumatoid arthritis (RA) fibroblast-like synoviocytes (FLSs).

METHODSThe synovial membrane tissues were obtained from 4 RA patients, 1 chondroma patient and 1 healthy subject and FLS were isolated by means of tissue culture. The cell morphology was observed by phase-contrast microscope and the cell surface markers were detected by flow cytometry.

RESULTSThe FLSs were successfully cultured from the synovial membrane tissues with good cell homogeneity after the third passage. The FLSs of the 3rd to 7th passages were stable and proliferated actively, followed by slow proliferation and aging since the 8th passage. Flow cytometry showed that the 4th-passage FLSs from the RA patients contained 99.04% CD90(+) cells, 2.73% CD3(+) cells, 0.29% CD3(-)CD19(+) cells, 2.81% CD3(-)CD16(+)CD56(+) cells, 5.89% CD14(+) cells, and 54.17% CD55(+) cells. The presence of interleukin-1 receptor type I (IL-1RI, 158.63-/+20.32 pg/ml) and IL-1beta (4.67-/+0.82 pg/ml) were detected in the cell culture supernatant of the 4th-passage FLSs from the RA patients by enzyme-linked immunosorbent assay ELISA.

CONCLUSIONFLSs from RA patients can be effectively culture by means of tissue culture, and the cultured FLSs show high expressions of CD90, IL-1RI and IL-1beta.

Adult ; Aged ; Arthritis, Rheumatoid ; pathology ; Cell Proliferation ; Cell Separation ; Cells, Cultured ; Female ; Fibroblasts ; pathology ; Humans ; Interleukin-1beta ; metabolism ; Male ; Middle Aged ; Receptors, Interleukin-1 Type I ; metabolism ; Synovial Membrane ; cytology ; pathology ; Thy-1 Antigens ; metabolism

OBJECTIVETo investigate the association of two single-nucleotide polymorphisms (SNPs) in IL1R1 gene (rs1558641 and rs949963) with the susceptibility to asthma in children from Central China.

METHODSA case-control study was performed in the asthma group and the control group, consisting of 208 children with asthma and 223 normal children from Central China, respectively. The genotypes of two SNPs in IL1R1 gene, rs1558641 and rs949963, were identified using polymerase chain reaction-restriction fragment length polymorphism. The serum level of IL1R1 was determined by enzyme-linked immunosorbent assay.

RESULTSThere were no significant differences in genotype and allele frequencies of rs1558641 between the asthma and control groups. In terms of rs949963, the frequencies of GG genotype and alleles were significantly higher in the asthma group than in the control group (P<0.05). The asthma group had a significantly higher serum level of IL1R1 than the control group (P=0.011). Moreover, the serum level of IL1R1 was significantly higher in patients with GG genotype than in those with AA or AG genotype for rs949963 (P=0.028).

CONCLUSIONSIL1R1 SNP rs949963 is associated with the susceptibility to asthma in children from Central China and may increase the serum expression of IL1R1.

Alleles ; Asthma ; genetics ; Case-Control Studies ; Child ; Child, Preschool ; Female ; Genetic Predisposition to Disease ; Genotype ; Humans ; Infant ; Male ; Polymorphism, Single Nucleotide ; Receptors, Interleukin-1 Type I ; genetics

OBJECTIVETo investigate the effect of berberine on expressions of tumor necrosis factor alpha (TNF-alpha) and receptor type I (TNFR1) in Abeta25-35-induced inflammatory reaction in SH-SYSY cell lines.

METHODThe 5 micromol . L-1 Abeta25-35 was used to treat SH-SY5Y cells for 24 hours, in order to establish the Alzheimer's disease (AD) model. Before modeling, berberine was given for pretreatment for 2 hours. The experiment included the normal control group, the AD model group, and indometacin low dose and high dose groups. Spectrophotometry was adopted to detect the activity of LDH. Meanwhile, the level of TNF-alpha was determined by ELISA, and the expression of TNFR1 genes was detected by RT-PCR.

RESULTCompared with the normal control group, the AD cell model group showed significant increase in LDH, TNF-alpha, and TNFR1 gene and protein expressions in the culture media. After intervention with berberine, the activity of LDH and TNF-alpha reduced in cell supernatant. The intervention with berberine could down-regulate TNFR1 gene and protein expressions, particularly 1, 10 x 10(-6) mol . L-l berberine showed a more notable effect in regulating TNFR1.

CONCLUSIONBerberine has the protective effect in Abeta-induced inflammatory injury in SH-SY5Y cells. Its mechanism may be related to the expression of its anti inflammatory factor TNF-alpha and its type I receptor TNFR1. Specifically, its regulation to TNFR1 shows dose dependence.

Amyloid beta-Peptides ; toxicity ; Berberine ; pharmacology ; Cell Line ; Humans ; Inflammation ; chemically induced ; metabolism ; Peptide Fragments ; toxicity ; Receptors, Interleukin-1 Type I ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism

OBJECTIVETo investigate the effects of local injections of recombinant soluble human receptors on experimental orthodontic tooth movement in the rat.

METHODS64 male Sprague-Dawley rats were observed. Starting at day 1, three groups of animals each received local injections of soluble interleukin-1 receptor II (sIL-1-R II ), soluble' tumor necrosis factor receptor I (sTNF-R I ) and their combinations. One group served as the control. The amount of tooth movement was recorded and selected tissue sections were stained with hematoxylin-eosin (HE) to observe the histological morphologic alterations of the periodontal tissues and also were stained with tartrate-resistant acid phosphatase (TRAP) histochemistry to analyze the changes of the amount and distribution of osteoclasts and odontoclasts.

RESULTSThe time-depended histomorphology changes in each receptor group were similar to those in the control group, but the resorption of alveolar bone was slighter at each time point and the surface of root appeared no or a few cement resorption signs. On day 14, the TRAP-positive cells on the surface of alveolar bone and root were reduced by approximately 50% relative to those in the control group (P<0.05). Whereas statistical tests revealed there were no significant differences among the experimental groups (P>0.05).

CONCLUSIONLocal injections of recombinant human sIL-1-R II and sTNF-R I in the orthodontic tooth of rats could reduce the amount and velocity of orthodontic tooth movement and reduce the incidence of root resorption.

Animals ; Dental Cementum ; Interleukin-1 ; Male ; Osteoclasts ; Rats ; Rats, Sprague-Dawley ; Rats, Wistar ; Receptors, Tumor Necrosis Factor ; Receptors, Tumor Necrosis Factor, Type I ; Root Resorption ; Tooth Movement Techniques ; Tooth Root

OBJECTIVETo assess the value of rheumatoid factors IgM-RF, IgA-RF, IgG-RF, interleukin-1 receptor type I (IL-1RI) and cyclin-dependent kinase 2 (CDK2) in the diagnosis of rheumatoid arthritis (RA).

METHODSIgM-RF, IgA-RF, IgG-RF, IL-1RI and CDK2 were detected in 47 patients with active RA, 47 with inactive RA, 20 with active systemic lupus erythematosus (SLE), 20 with inactive SLE, 20 with acute upper respiratory tract infection (AURTI), and 20 healthy controls using enzyme-linked immunosorbent assay (IgM-RF, IgA-RF, IgG-RF, IL-1RI) and time-resolved fluoroimmunoassay (CDK2).

RESULTSPatients with active and inactive RA showed significant differences in peripheral serum CDK2 and IgA-RF levels (P<0.05). Between active and inactive RA patients, RA patients and SLE patients, RA patients and AURTI patients, and RA patients and the control subjects, the area under curve (AUC) of the receiver operating characteristic curve (ROC) was 0.561, 0.814, 0.799, and 0.888 for IgM-RF, 0.596, 0.678, 0.729, and 0.850 for IgA-RF, 0.614, 0.718, 0.692, and 0.791 for IgG-RF, 0.646, 0.691, 0.762, and 0.835 for IL-1RI, 0.803, 0.753, 0.741, and 0.840 for CDK2, respectively.

CONCLUSIONSIgM-RF may have high value in the diagnosis and differential diagnosis of RA, and CDK2 can be useful for differential diagnosis between active and inactive RA.

Adult ; Area Under Curve ; Arthritis, Rheumatoid ; blood ; diagnosis ; Cyclin-Dependent Kinase 2 ; blood ; Female ; Humans ; Immunoglobulin A ; blood ; Immunoglobulin G ; blood ; Immunoglobulin M ; blood ; Male ; Middle Aged ; ROC Curve ; Receptors, Interleukin-1 Type I ; blood ; Rheumatoid Factor ; blood

We investigated the question of whether cholesterol catabolite can influence expression of inflammatory cytokines via Toll-like receptors (TLR) in monocytic cells. Treatment of THP-1 monocytic cells with 27-hydroxycholesterol (27OHChol) resulted in induction of gene transcription of TLR6 and elevated level of cell surface TLR6. Addition of FSL-1, a TLR6 agonist, to 27OHChol-treated cells resulted in transcription of the IL-1alpha gene and enhanced secretion of the corresponding gene product. However, cholesterol did not affect TLR6 expression, and addition of FSL-1 to cholesterol-treated cells did not induce expression of IL-1alpha . Using pharmacological inhibitors, we investigated molecular mechanisms underlying the expression of TLR6 and IL-1alpha. Treatment with Akt inhibitor IV or U0126 resulted in significantly attenuated expression of TLR6 and IL-1alpha induced by 27OHChol and 27OHChol plus FSL-1, respectively. In addition, treatment with LY294002, SB202190, or SP600125 resulted in significantly attenuated secretion of IL-1alpha . These results indicate that 27OHChol can induce inflammation by augmentation of TLR6-mediated production of IL-1alpha in monocytic cells via multiple signaling pathways.
Cholesterol ; Cytokines ; Inflammation ; Interleukin-1 ; Interleukin-1alpha* ; Toll-Like Receptors*

OBJECTIVEKawasaki disease (KD) is an acute febrile, multi-system endangeitis, which is mainly found in early childhood. Its etiology is still unknown. A great deal of clinical evidence and epidemiologic data suggest that KD is correlated with an acute immune dysfunction caused by infection. Many evidences in the past suggested that over-expression of proinflammatory cytokines, co-stimulatory molecules and chemokines, which were observed in KD, may contribute to the pathologic lesion of vascular endothelial cells. But the causative factors are still unknown. Toll-like receptor is a type I trans-membrane protein which could recognize ligands of pathogenic microbes, induce interferon beta (IFN-beta) and promote gene transcription of proinflammatory cytokines, co-stimulatory molecules and chemokines. This study was designed to investigate the role of MyD88-independent signal transduction of Toll-like receptor 4 in immunological pathogenesis of KD.

METHODSThirty-two children with KD and 16 age-matched healthy children were studied. Reverse-transcription PCR (RT-PCR) and real-time PCR were used to evaluate the mRNA levels of Toll-like receptor 4 and the molecules such as Toll-IL-1-receptor domain containing adaptor inducing IFN-beta (TRIF), TRIF-related adaptor molecule (TRAM), TANK-binding kinase 1 (TBK-1), IFN-beta, interferon-gamma-inducible protein 10 (IP-10), regulated on activation normal T cells expressed and secreted (RANTES), inducible nitric oxide synthase (iNOS) and suppressor of cytokine signaling 1 (SOCS-1) in monocytes/macrophages (MC), which participate in MyD88-independent signal transduction of toll-like receptors. Expression of costimulatory molecules such as CD40 in MC was analyzed by flow cytometry. Methylation-specific PCR was performed to analyze the methylation status of cytosine-phosphate-guanine (CpG) motif in SOCS-1 gene.

RESULTS(1) Compared with healthy controls, transcription levels of the molecules such as TLR4, TRIF, TRAM, TBK-1 and IFN-beta, were significantly up-regulated during acute phase of KD (P < 0.05), and down-regulated after treatment with intravenous immunoglobulin therapy. (2) Expression of iNOS and chemokines such as IP10 and RANTES in MC during acute phase of KD was remarkably elevated (P < 0.05), and down-regulated to some extents after treatment with intravenous immunoglobulin therapy. (3) Expression of costimulatory molecule CD40 in MC increased significantly during acute phase of KD [(6.19 +/- 2.25)% vs. (2.00 +/- 1.37)%, P < 0.05], while the protein levels of CD40 in KD-coronary artery lesion (CAL)(+) group was found to be significantly higher than that of KD-CAL-group [KD-CAL, (9.63 +/- 2.96)% vs. (4.12 +/- 1.91)%, P < 0.05]. (4) Expression levels of SOCS-1 mRNA were significantly up-regulated during acute phase of KD [(4.31 +/- 0.83) x 10(-3) vs. (1.09 +/- 0.23) x 10(-3), P < 0.05], and the levels of SOCS-1 gene in KD-CAL(+) group was found to be significantly lower than that of KD-CAL(-) group [(5.73 +/- 1.04) x 10(-3) vs (1.94 +/- 0.46) x 10(-3), P < 0.05]. (5) The CpG island of SOCS-1 DNA in KD patients was remarkably demethylated [(26.9 +/- 8.6)% vs (5.9 +/- 1.4)%, P < 0.05], and demethylation levels of SOCS-1 in KD-CAL(-) group were higher than that in KD-CAL+ group [(35.1 +/- 10.3)% vs. (13.2 +/- 3.7)%, P < 0.05].

CONCLUSIONAberrant activation of MyD88-independent pathways of Toll-like receptor 4 may be one of the factors causing disturbed immunological function in KD.

Child ; Humans ; Interleukin-1 ; metabolism ; Macrophages ; drug effects ; pathology ; Nitric Oxide Synthase Type II ; metabolism ; Pyrimidinones ; pharmacology ; Signal Transduction ; drug effects ; physiology ; Thiazoles ; pharmacology ; Toll-Like Receptor 4 ; metabolism ; Toll-Like Receptors ; deficiency ; drug effects ; metabolism