BACKGROUNDOsteoarthritis (OA) is the most common form of human polyarthritis. Many genetic factors have been implicated in OA. It was reported that a polymorphism in the gene of interleukin-6 (IL-6) was associated with OA of knee. The aim of this study was to determine whether functional IL-6 promoter -174G/C (rs1800795) polymorphisms confer susceptibility to knee OA.

METHODSA meta-analysis was conducted on the association between the IL-6 polymorphism and knee OA. Electronic search at PubMed, EMBASE, Weipu database, and Wanfang database was conducted to select studies. Case-control studies containing available genotype frequencies of IL-6 -174G/C were chosen, and odds ratio (OR) with 95% confidence interval (CI) was used to assess the strength of this association.

RESULTSA total of seven studies involving 6 464 subjects (knee OA 3 331 and controls 3 133) were considered in this study. The results suggested that the variant genotypes were not associated with knee OA risk in all genetic models (additive model: OR = 1.144, 95% CI 0.934-1.402, P = 0.194; recessive model: OR = 1.113, 95% CI 0.799-1.550, P = 0.526; dominant model: OR = 1.186, 95% CI 0.918-1.531, P = 0.191). A symmetric funnel plot, the Begg's test (P > 0.05), suggested that the data lacked publication bias.

CONCLUSIONSThis meta-analysis does not support the idea that rs1800795 genotype is associated with increased risk of knee OA. However, to draw comprehensive and more reliable conclusions, further prospective studies with larger numbers of participants worldwide are needed to examine the association between rs1800795 polymorphism and knee OA.

Humans ; Interleukin-6 ; genetics ; Osteoarthritis, Knee ; epidemiology ; genetics ; Polymorphism, Genetic ; genetics ; Promoter Regions, Genetic ; genetics

The immunomodulatory effect of Saposhnikoviae Radix polysaccharide(SRP) was evaluated based on the zebrafish mo-del, and its mechanism was explored by transcriptome sequencing and real-time fluorescence-based quantitative PCR(RT-qPCR). The immune-compromised model was induced by navelbine in the immunofluorescence-labeled transgenic zebrafish Tg(lyz: DsRed), and the effect of SRP on the density and distribution of macrophages in zebrafish was evaluated. The effect of SRP on the numbers of macrophages and neutrophils in wild-type AB zebrafish was detected by neutral red and Sudan black B staining. The content of NO in zebrafish was detected by DAF-FM DA fluorescence probe. The content of IL-1β and IL-6 in zebrafish was detected by ELISA. The differentially expressed genes(DEGs) of zebrafish in the blank control group, the model group, and the SRP treatment group were analyzed by transcriptome sequencing. The immune regulation mechanism was analyzed by Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)enrichment, and the expression levels of key genes were verified by RT-qPCR. The results showed that SRP could significantly increase the density of immune cells in zebrafish, increase the number of macrophages and neutrophils, and reduce the content of NO, IL-1β, and IL-6 in immune-compromised zebrafish. The results of transcriptome sequencing analysis showed that SRP could affect the expression level of immune-related genes on Toll-like receptor pathway and herpes simplex infection pathway to affect the release of downstream cytokines and interferon, thereby completing the activation process of T cells and playing a role in regulating the immune activity of the body.
Animals ; Zebrafish/genetics* ; Interleukin-6/genetics* ; Gene Expression Profiling ; Cytokines/genetics* ; Macrophages ; Transcriptome

OBJECTIVETo determine the relationship between polymorphisms in the genes encoding IL-1, IL-6, and IL-10 with primary biliary cirrhosis (PBC) in Chinese population.

METHODSWhole-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).

RESULTSThe 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.

CONCLUSIONThe 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.

Adult ; Aged ; Female ; Humans ; Interleukin-1 ; genetics ; Interleukin-10 ; genetics ; Interleukin-6 ; genetics ; Liver Cirrhosis, Biliary ; genetics ; Male ; Middle Aged ; Polymerase Chain Reaction ; methods ; Polymorphism, Restriction Fragment Length

BACKGROUNDChai Lai Prescription is a Chinese herbal compound which is used to sooth the liver, strengthen the spleen and harmonize the stomach for descending adverse Qi. We initiated the study to investigate its mechanism of treating in vitro rabbit reflux esophagitis models.

METHODSAdult male Japanese white rabbits, weighing 1.8-2.2 kg, were divided into five groups of three each, which were: normal control group (Krebs buffer, pH7.4), esophagitis model group (Krebs buffer, pH5.8), esophagitis model proup+low-dose Chinese herbal medicine protection group (0.6 mg × ml(-1)× kg(-1)), esophagitis model group+moderate-dose Chinese herbal medicine protection group (6 mg × ml(-1)× kg(-1)), esophagitis model group+high-dose Chinese herbal medicine protection group (60 mg × ml(-1)×kg(-1)). The RT-PCR method was used to test the influence of Chai Lai Prescription on IL-1 and IL-6 in in vitro rabbit models of esophagitis. We treated the in vitro models with different doses of Chinese herbal medicine.

RESULTSEsophageal mucosa were filled with various liquids. IL-6 and IL-1β mRNA expression was increased in rabbit esophageal mucosa stimulated with acid. Chinese herbal medicine significantly reduced the levels of IL-6 and IL-1β mRNA expression in the in vitro cultured rabbit esophageal mucosa. Using Chinese herbal medicine to treat in vitro models of RE, we found that the IL-6 and IL-1β mRNA expression levels went down, near to or lower than the normal control levels, compared with the group treated with acidified buffer solution.

CONCLUSIONSChai Lai Prescription lowered the IL-1β and IL-6 cytokine mRNA levels and protected the esophageal mucosa in the in vitro models of reflux esophagitis, suggesting that the traditional Chinese herbal compound may be able to treat reflux esophagitis by inhibiting the its inflammatory mediators.

Animals ; Drugs, Chinese Herbal ; therapeutic use ; Esophagitis, Peptic ; drug therapy ; metabolism ; Interleukin-1beta ; genetics ; Interleukin-6 ; genetics ; Male ; Rabbits

OBJECTIVETo investigate the association of interleukin 6 gene (IL-6) promoter region 634C/G (rs1800796) single nucleotide polymorphism (SNP) with the genetic susceptibility to endometriosis (Ems) in south Han Chinese women.

METHODSA case-control study was performed in 432 Ems patients and 499 control women to evaluate the SNP of IL-6 634C/G by using a fluorescent quantitative PCR-based high resolution melting (HRM) method.

RESULTSThere were statistical significances in the IL-6 634C/G alleles, whether or not to carry allele G and genotype distributions between Ems patients and control women (P=0.032, 0.014 and 0.045, respectively). Allele C enhanced the risk of Ems 1.057 times while allele G reduced the risk of Ems 0.835 time. Carrying allele G reduced the risk of Ems 0.822 time, whereas not carrying allele G enhanced the risk of Ems 1.143 times. Compared with genotype CC, the risk of Ems with genotype CG reduced 0.704 time (95% CI: 0.533-0.931). There was no significant difference in whether or not carrying allele G distribution between Ems patients and control women (P=0.729).

CONCLUSIONThe present study demonstrated significant association between the SNP of IL-6 634C/G and genetic susceptibility to Ems in south Han Chinese women.

Alleles ; Case-Control Studies ; Endometriosis ; genetics ; Female ; Genetic Predisposition to Disease ; genetics ; Genotype ; Humans ; Interleukin-6 ; genetics ; Polymorphism, Single Nucleotide ; genetics

OBJECTIVETo identify the differential expressions of serum cytokines between prostate cancer (PCa) and benign prostatic hyperplasia (BPH), and provide proteomic evidence for the early diagnosis of PCa.

METHODSWe used human cytokine array to determine the profiles of the serum cytokines obtained from 6 PCa and 6 BPH patients with the PSA level within the grey scale of 4 - 10 ng/ml.

RESULTSWe identified 19 differentially expressed cytokines in the PCa patients, 16 obviously up-regulated, including IL-3, IL-6 and IL-16, and 3 markedly down-regulated, which were Fas/TNFRSF6, TRALR-3 and IGFBP-6. Most of them were involved in such cellular bioprocesses as transcription, proliferation, signal transduction, and apoptosis.

CONCLUSIONThe cytokine antibody assay permits simultaneous measurement of multiple markers in a small volume of serum, and can identify a panel of key cytokines related to the malignant biological behavior of cancer cells. And it helps to find the biomarkers for the early diagnosis, efficacy assessment and prognosis of prostate cancer.

Aged ; Humans ; Interleukin-16 ; blood ; Interleukin-3 ; blood ; Interleukin-6 ; blood ; Male ; Middle Aged ; Prostatic Hyperplasia ; blood ; genetics ; metabolism ; Prostatic Neoplasms ; blood ; genetics ; metabolism ; Proteomics

Objective To explore the aptamer specific binding blood group antigen-binding adhesin (BabA) of Helicobacter pylori (H.pylori) for blocking of H.pylori adhering host cell. Methods H.pylori strain was cultured and its genome was extracted as templates to amplify the BabA gene by PCR with designed primers. The BabA gene obtained was cloned and constructed into prokaryotic expression plasmid, which was induced by isopropyl beta-D-galactoside (IPTG) and purified as target. The single stranded DNA (ssDNA) aptamers that specifically bind to BabA were screened by SELEX. Enzyme-linked oligonucleotide assay (ELONA) was used to detect and evaluate the characteristics of candidate aptamers. The blocking effect of ssDNA aptamers on H.pylori adhesion was subsequently verified by flow cytometry and colony counting at the cell level in vitro and in mouse model of infection, respectively. Meanwhile, the levels of cytokines, interleukin 6 (IL-6), IL-8, tumor necrosis factor α (TNF-α), IL-10 and IL-4 in the homogenate of mouse gastric mucosa cells were detected by ELISA. Results The genome of H.pylori ATCC 43504 strains was extracted and the recombinant plasmid pET32a-BabA was constructed. After induction and purification, the relative molecular mass (M_r) of the recombinant BabA protein was about 39 000. The amino acid sequence of recombinent protein was consistent with BabA protein by peptide mass fingerprint (PMF). Five candidate aptamers were selected to bind to the above recombinent BabA protein by SELEX. The aptamers A10, A30 and A42 identified the same site, while A3, A16 and the above three aptamers identified different sites respectively. The aptamer significantly blocked the adhesion of H.pylori in vitro. Animal model experiments showed that the aptamers can block the colonization of H.pylori in gastric mucosa by intragastric injection and reduce the inflammatory response. The levels of IL-4, IL-6, IL-8 and TNF-α in gastric mucosal homogenates in the model group with aptamer treatment were lower than that of model group without treatment. Conclusion Aptamers can reduce the colonization of H.pylori in gastric mucosa via binding BabA to block the adhesion between H.pylori and gastric mucosal epithelial cells.
Animals ; Mice ; Helicobacter pylori/genetics* ; Interleukin-4 ; Interleukin-6 ; Interleukin-8 ; Tumor Necrosis Factor-alpha ; Stomach ; Oligonucleotides ; Adhesins, Bacterial/genetics* ; Blood Group Antigens

OBJECTIVE: To observe the effects of moxibustion at "Mingmen" (GV 4) and "Guanyuan" (CV 4) on immune function and intestinal flora in healthy rats, thereby investigating the underlying mechanism of moxibustion on immune function. METHODS: Twenty 8-week-old SD rats were randomly divided into a young blank group and a young moxibustion group, with 10 rats in each group. Similarly, twenty 8-month-old SD rats were randomly divided into a middle-aged blank group and a middle-aged moxibustion group, with 10 rats in each group. The rats in the two moxibustion groups received moxibustion at "Mingmen" (GV 4) and "Guanyuan" (CV 4), 15 min per session, once daily, five times a week, for a total of four months. The rats in the two blank groups were fed under normal conditions. After the intervention, thymus and spleen indexes were calculated; the morphology of thymus and spleen tissues was observed using HE staining; the flow cytometry was used to detect the expression of CD and CD T lymphocytes and the CD/CD ratio was calculated; ELISA was used to measure the serum levels of tumor necrosis factor-alpha (TNF-α), interferon-gamma (IFN-γ), interleukin-6 (IL-6), interleukin-10 (IL-10), and interleukin-17 (IL-17); 16S rRNA high-throughput sequencing was used to analyze the intestinal flora. Additionally, the correlation between the relative abundance of intestinal flora and serum levels of TNF-α, IFN-γ, IL-6, IL-10 and IL-17 was analyzed. RESULTS: Compared with the young blank group, the young moxibustion group exhibited an increase in the cortical area of thymus tissue with tighter lymphocyte arrangement; compared with the middle-aged blank group, the middle-aged moxibustion group showed an increase in thymus index (P<0.05) and an increase in the cortical area of thymus tissue. There were no significant differences in spleen index between the 2 moxibustion groups and the 2 blank groups (P>0.05). There were no significant differences in the expression of CD, CD, and CD/CD ratio between the 2 moxibustion groups and the corresponding blank groups (P>0.05). Compared with the young blank group, the young moxibustion group had elevated IL-6 level (P<0.05); compared with the middle-aged blank group, the middle-aged moxibustion group had decreased IL-10 and IL-17 levels (P<0.05). Compared with the young blank group, the young moxibustion group exhibited increased Sobs index, Ace index, and Chao index (P<0.01, P<0.05), as well as increased relative abundance of Spirochaetota, Treponema, Turicibacter, Rikenellaceae_RC9_gut_group (P<0.05), and decreased relative abundance of Dubosiella (P<0.05). Compared with the middle-aged blank group, the middle-aged moxibustion group had increased relative abundance of Spirochaetota, Treponema, norank_f_Peptococcaceae (P<0.05), and decreased relative abundance of Proteobacteria, Allobaculum, and Faecalibaculum (P<0.05). Correlation analysis revealed that relative abundance of Eubacterium_xylanophilum_group and unclassified _f_Lachnospiraceae was negatively correlated with serum TNF-α level (r=-0.39, P=0.03; r=-0.24, P=0.04), while relative abundance of norank_f_norank_o_Clostridia_UCG-014 and Lactobacillus was positively correlated with serum TNF-α level (r=0.37, P=0.04; r=0.43, P=0.02). The relative abundance of Roseburia and Monoglobus was negatively correlated with serum IFN-γ level (r=-0.40, P=0.02; r=-0.44, P=0.01), while relative abundance of Lactobacillus was positively correlated with serum IL-10 level (r=0.43, P=0.02). CONCLUSION Moxibustion could improve immune function in healthy rats, and its mechanism may be related to the regulation of relative abundance of intestinal flora.
Rats ; Animals ; Moxibustion ; Rats, Sprague-Dawley ; Interleukin-10/genetics* ; Interleukin-17 ; Tumor Necrosis Factor-alpha/metabolism* ; Interleukin-6/genetics* ; Gastrointestinal Microbiome ; RNA, Ribosomal, 16S ; Interferon-gamma ; Immunity

OBJECTIVETo establish the activation of NF-kappaB in middle ear cholesteatoma, discuss the relationship of NF-kappaB and the gene expression of IL-6 and explore the pathogenesis of middle ear cholesteatoma.

METHODSTen cases of cholesteatoma and 6 cases of normal external meatal skin were obtained from middle ear surgery. The NF-kappaB DNA binding activity and the mRNA level of IL-6 in these two kinds of tissues were detected by electrophoretic motility shift assay (EMSA) and Reverse transcription polymerase chain reaction (RT-PCR) respectively. The relationship of NF-kappaB DNA binding activity and the mRNA level of IL-6 were analyzed.

RESULTSThe NF-kappaB DNA binding activities of cholesteatoma [(15.9 +/- 8.2)%] were higher than those in normal skin [(1.36 +/- 0.94)%, t = 3.502, P < 0.05]. The expression of IL-6 mRNA was increased significantly in patients with cholesteatoma, as compared with that in the control specimens (t = 2.166, P < 0.05) and had a significant positive correlation with NF-kappaB DNA binding activity (r = 0.752, P < 0.05).

CONCLUSIONSThe IL-6 mRNA expression in cholesteatoma is closely related with the activity of NF-kappaB. It is tempting to speculate that NF-kappaB play a key role in the activation of cytokine in cholesteatoma.

Cholesteatoma, Middle Ear ; genetics ; metabolism ; Gene Expression Regulation ; Humans ; Interleukin-6 ; genetics ; metabolism ; NF-kappa B ; metabolism ; RNA, Messenger ; genetics

Based on site-specific transposition of an expression cassette into a baculovirus shuttle vector (Bacmid) which propagated in Escherichia coli, the Bac-to-Bac System provides a rapid and efficient method to generate recombinant baculoviruses and is widely used for high level expression of heterologous proteins. And the efficiency of recombinant baculovirus infecting cells plays an important role on the protein expression. In this study, we introduced an EGFP expression cassette driven by polyhedrin promoter into the p74 locus of Bacmid by homologous recombination. The target Bacmid-egfp was then transformed into E. coli DH10B containing the transposition helper plasmid to gain a new transposition receipt strain E. coli DH10Bac-egfp. Because of the intact attTn7 sites and lacZ', target gene cloned in a pFastBac vector can be transposed into the Bacmid-egfp shutter vector to construct recombinant baculovirus, which would allow the tracing of the target protein expression and the recombinant Bacmid transfection or recombinant baculoviral infection under fluorescence microscopes. Recombinant virus Bac-egfp-DsRed was constructed by transposing DsRed into the Bacmid-egfp in E. coliDHl0Bac-egfp, and the Sf9 cells infected with the recombinant virus expressed DsRed and EGFP efficiently. Another protein IL-6 fused with 6 x his tag was expressed and purified sucessfully from Sf9 cells infected with recombinant virus Bac-egfp-6 x his-IL6 constructed by the improved Bac-to-Bac system.
Animals ; Baculoviridae ; genetics ; Green Fluorescent Proteins ; genetics ; Interleukin-6 ; biosynthesis ; genetics ; Plasmids ; Polymerase Chain Reaction ; Recombinant Proteins ; biosynthesis ; Spodoptera