1.The effects of nuclear factor-kappa B p65 antisense oligonucleotides on expression of proinflammatory cytokines in lamina propria mononuclear cells from patients with ulcerative colitis.
Huatian GAN ; Qin OUYANG ; Youqin CHEN ; Feng LIANG
Journal of Biomedical Engineering 2003;20(2):268-272
To investigate if nuclear factor-kappa B (NF-kappa B) p65 antisense oligonucleotides might affect the expression of NF-kappa B p65 and cytokines in lamina propria mononuclear cells(LPMC) from patients with ulcerative colitis (UC). LPMC were isolated from intestinal mucosal biopsy specimens from 3 patients with UC, and cultured with or without NF-kappa B p65 antisense oligonucleotides (5'-GGAACAGTTCGTCCTATGG-3'), missense oligonucleotides (5'-GGAACAGTTCGTCTATGG-3') and dexamethasone. NF-kappa B p65 expression was determined by western blot analysis. The expression of cytokine mRNA was studied by reversal transcription-polymerase chain reaction (RT-PCR). The cytokine levels were measured by enzyme linked immunosorbent assay. The results showed that NF-kappa B p65 antisense oligonucleotides resulted in down-regulation of NF-kappa B p65 expression, blocked the expression of IL-1 beta mRNA and IL-8 mRNA, and strikingly reduced the production of IL-1 beta and IL-8, and these effects were greater than those of dexamethasone in cultured LPMC from patients with UC(P < 0.05). Therefore, the application of NF-kappa B p65 antisense oligonucleotides may serve as a novel molecular approach for the treatment of patients with UC.
Cells, Cultured
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Colitis, Ulcerative
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drug therapy
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pathology
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Cytokines
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biosynthesis
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genetics
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Humans
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Interleukin-1
;
biosynthesis
;
genetics
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Interleukin-8
;
biosynthesis
;
genetics
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Intestinal Mucosa
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cytology
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Monocytes
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drug effects
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metabolism
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NF-kappa B
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biosynthesis
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genetics
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Oligonucleotides, Antisense
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pharmacology
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RNA, Messenger
;
biosynthesis
2.Effects of exogenous IL-10 on IL-6 and ICAM-1 expression in inflammatory gingival tissue.
Pishan YANG ; Xiangmin QI ; Shaohua GE ; Min ZHAO
West China Journal of Stomatology 2002;20(5):343-345
OBJECTIVEThis study aimed at investigating effects of exogenous interleukine-10 (IL-10) on IL-6 and intercellular adhesion molecular (ICAM-1) expression in inflamed gingival tissue.
METHODSThe expression of IL-6 and ICAM-1 was examined using immunohistochemical techniques. Inflammatory gingival tissue treated with IL-10 was taken as the experimental group and the same patient's inflammatory gingival tissue without treatment of IL-10 was included into the control group.
RESULTSIL-6 expression was found mainly in monocytes, macrophages, lymphocytes, endotheliocytes and fibroblasts. The expression of ICAM-1 was found mainly in epithelial cells, monocot-macrophages, lymphocytes, endotheliocytes and fibroblasts. The immunohistochemical optical density (IOD) of the expression of IL-6 and ICAM-1 was detected by using Image-Proplus software, and the results showed that the expression in the experimental group differed significantly from that in the control group.
CONCLUSIONThe exogenous IL-10 may down-regulate IL-6 and ICAM-1 expression in inflammatory gingival tissue.
Adult ; Down-Regulation ; Female ; Gingiva ; metabolism ; Gingivitis ; metabolism ; Humans ; Intercellular Adhesion Molecule-1 ; biosynthesis ; genetics ; Interleukin-10 ; pharmacology ; Interleukin-6 ; biosynthesis ; genetics ; Male ; Middle Aged
3.Current progress in study of effect of bone morphogenetic protein on hematopoiesis.
Journal of Experimental Hematology 2003;11(3):312-315
This article reviews the structure, physical and chemical characteristics of bone morphogenetic protein, summarized the effects of bone morphogenetic protein on hematopoiesis in embryo and adult animal, discussed the possible mechanisms and pointed out the theoretic and practical significance on this research work.
Animals
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Bone Marrow Cells
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drug effects
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metabolism
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Bone Morphogenetic Proteins
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pharmacology
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Hematopoiesis
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drug effects
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Humans
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Interleukin-1
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biosynthesis
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Interleukin-6
;
biosynthesis
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Research
;
trends
4.Effects of baicalin on the expression of pro-MMP-1 and MMP-3 in human gingival fibroblasts and periodontal ligament cells.
Cheng-zhang LI ; Zheng-guo CAO ; Ru YANG ; Zhu-huan SHANG ; Li-jian JIN ; E F COBERT
Chinese Journal of Stomatology 2004;39(3):197-200
OBJECTIVETo investigate the influence of baicalin on the IL-1beta induced pro-MMP-1 in HGF and the effects of baicalin on MMP-3 expression in periodontal ligament cells (PDLCs).
METHODSThe amount of secreted pro-MMP-1 and MMP-3 expression was detected by ELISA and cell immunochemistry.
RESULTS(1) The amount of secreted pro-MMP-1 (3.333 +/- 0.123) microg/L increased significantly following 1 microg/L of IL-1beta, compared with control group (1.960 +/- 0.180) microg/L. Addition of baicalin to cell culture medium for 1 hour following IL-1beta decreased pro-MMP-1 secretion in a dose-dependent manner in the range of 10 approximately 1,000 microg/L. (2) 1 microg/L IL-1beta could significantly stimulate the synthesis and secretion of MMP-3 in PDLCs. (3) The baicalin could not interfere the synthesis of MMP-3, but could inhibit the release of MMP-3 from PDLCs.
CONCLUSIONSBaicalin could inhibit the secretion of pro-MMP-1 and MMP-3 expression in IL-1beta induced HGF and PDLCs, which suggests that baicalin may play an important role in preventing and treating periodontal disease.
Collagenases ; biosynthesis ; genetics ; Enzyme Precursors ; biosynthesis ; genetics ; Fibroblasts ; enzymology ; pathology ; Flavonoids ; pharmacology ; Gingiva ; enzymology ; pathology ; Humans ; Interleukin-1 ; pharmacology ; Interleukin-1beta ; Matrix Metalloproteinase 1 ; Metalloendopeptidases ; biosynthesis ; genetics ; Peptide Fragments ; pharmacology ; Periodontal Ligament ; enzymology ; pathology ; Periodontitis ; enzymology ; pathology ; Scutellaria ; chemistry
5.Construction, expression and preliminary pharmacokinetic analysis of IL-1ra mutants.
Yu-Xin WANG ; Zhi-Xin YANG ; Heng-Qi ZHU ; Xiao-Wei ZHOU ; Pei-Tang HUANG
Chinese Journal of Biotechnology 2006;22(3):472-476
Interleukin-1 receptor antagonist (IL-1ra), a member of IL-1 family, is a naturally occurring IL-1 inhibitor as "receptor antagonist", which blocks biological responses mediated by IL-1. Recombinant human IL-1ra (rhIL-1ra, Kineret) was introduced in clinical trials involving patients with RA. Between 2001 to approximately 2002, rhIL-1 ra was approved by the US Food and Drug Administration and the European Agency for the Evaluation of Medicine Procedure. Unfortunately, 10,000 to 100,000-fold excess amounts of IL-1ra are needed to relieve disease because minimal IL-1 can induce complete biological responses, and the dosage of 100 to approximately 150mg/day in a RA patient is so big that it greatly influence patients' physical, psychological and economical situation. In this study, IL-1ra mutants were established by site-specific mutagenesis to improve its stability. The sites of mutagenesis included R6 K7-AA,R93 K94-AA and K97 R98-AA. IL-1ra and its mutants were expressed in E. coli BL21 (DE3) using pTIG-Trx expressing system with the induction of IPTG. The recombinant proteins were purified by Ni2+ chelate chromatography and Sephadex G75 gel filtration chromatography. The activity of mutants is as high as IL-1ra. We characterized the pharmacokinetic profile of IL-1ra and its mutants. The third mutant's half life is 2.26 times than wt IL-1ra. The study has provided some approaches and experience for further research to improve the metabolism stability of IL-1ra.
Animals
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Escherichia coli
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genetics
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metabolism
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Female
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Humans
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Interleukin 1 Receptor Antagonist Protein
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biosynthesis
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genetics
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pharmacokinetics
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Mutagenesis, Site-Directed
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methods
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Mutant Proteins
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biosynthesis
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pharmacokinetics
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Recombinant Proteins
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biosynthesis
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genetics
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pharmacokinetics
6.Interleukin-1beta expression and phospholipase A(2) activation after intestinal ischemia/reperfusion injury.
Guang-Tao YAN ; Xiu-Hua HAO ; Hui XUE ; Lu-Huan WANG ; Ying-Li LI ; Li-Ping SHI
Acta Physiologica Sinica 2002;54(1):28-32
The experiments were carried out to explore the interactions between IL-1 beta gene expression, protein level and phospholipase A(2) PLA(2) inhibition after intestinal ischemia/reperfusion injury. Using a rat intestinal ischemia/reperfusion injury model, after collecting the serum, lung lavage, abdomen cavity lavage and important organ tissue samples from control, injury and PLA(2) inhibitor treated groups, IL-1 beta level was measured by radioimmunoassay, and the mRNA expression of IL-1 beta and type II PLA (2)was determined by RT-PCR. After 6 h of injury, the IL-1 beta level in serum was significantly higher than that in the control group; an increase in IL-1 beta was also observed in abdomen cavity lavage 1 or 3 h after injury. IL-1 beta was significantly increased in liver tissue after injury, but was not changed obviously in the lung, kidney and intestinal tissues. IL-1 beta in the lung lavage was significantly higher than that of control group. The mRNA expression of IL-1 beta in lung tissue was increased after injury, but type II PLA(2) mRNA expression was decreased. There were different changes in IL-1 beta level and gene expression after treatment with PLA(2) inhibitor chloroquine, cyclo-oxidase inhibitor indomethacin, or PAF receptor antagonist SR27417 respectively after injury. All these results indicate that after intestinal ischemia/reperfusion injury, the IL-1 beta level and mRNA gene expression are significantly increased, however, the relationship among IL-1 beta, PLA(2) activation and its metabolite release remains to be further elucidated.
Animals
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Female
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Gene Expression
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Interleukin-1
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biosynthesis
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metabolism
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Intestines
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blood supply
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Ischemia
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metabolism
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Male
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Phospholipases A
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biosynthesis
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metabolism
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RNA, Messenger
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biosynthesis
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Rats
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Rats, Wistar
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Reperfusion Injury
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metabolism
7.The expression of TNF-alpha and IL-1 beta in lung tissues following liver ischemia-reperfusion injury in rats.
Hong-mei WANG ; Fei TANG ; Jian-long ZHANG ; Wen-an BIAN
Chinese Journal of Hepatology 2005;13(5):383-384
Animals
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Interleukin-1
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biosynthesis
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genetics
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Liver
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blood supply
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Lung
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metabolism
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Male
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RNA, Messenger
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biosynthesis
;
genetics
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Rats
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Rats, Wistar
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Reperfusion Injury
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metabolism
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Tumor Necrosis Factor-alpha
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biosynthesis
;
genetics
8.Effect of heart benefiting recipe in controlling IL-1beta, IL-6 and APPmRNA expression in brain of beta-amyloid protein induced rat model of dementia.
Hui ZHOU ; Wei-kang ZHAO ; Guo-qin JIN
Chinese Journal of Integrated Traditional and Western Medicine 2004;24(6):529-533
OBJECTIVETo investigate the neuro-immune regulatory mechanism of Heart Benefiting recipe (HBR), an effective recipe for treatment of Alzheimer's disease (AD).
METHODSUsing immunohistochemical and RT-PCR methods, the neuro-immunological pathological changes in the AD rat model induced by beta-amyloid protein (A beta1-40) via lateral cerebral ventricle injection, including mainly the glial fibrillary acidic protein expression and inflammatory cytokines IL-1beta, IL-6mRNA and beta-amyloid protein precursor (APPmRNA) gene expression were studied. And the effects of HBR on these parameters were observed.
RESULTSDeposition of A beta in cerebral tissue could induce activation of stellate glial cells and abnormal increased levels of inflammatory cytokines (IL-1beta and IL-6mRNA), also the elevation of APPmRNA level. HBR could effectively control the above-mentioned pathological changes.
CONCLUSIONHBR could effectively control the inflammation and the A beta immune cascade reaction in brain of AD patients, it is one of the important therapeutic mechanisms of the recipe.
Alzheimer Disease ; chemically induced ; metabolism ; Amyloid beta-Peptides ; Amyloid beta-Protein Precursor ; biosynthesis ; genetics ; Animals ; Brain ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Interleukin-1 ; biosynthesis ; genetics ; Interleukin-6 ; biosynthesis ; genetics ; Male ; Neuroprotective Agents ; pharmacology ; RNA, Messenger ; biosynthesis ; genetics ; Random Allocation ; Rats
9.Interleukin-1 beta, -6 and interferon-gamma productions in patients undergoing continuous ambulatory peritoneal dialysis.
Kyu Hun CHOI ; In Hee LEE ; Sug Kyun SHIN ; Hyun Jin NOH ; Shin Wook KANG ; Ho Young LEE ; Dae Suk HAN
Yonsei Medical Journal 1998;39(3):240-246
In order to investigate cytokine productions in patients undergoing continuous ambulatory peritoneal dialysis (CAPD), we studied the production of interleukin (IL)-1 beta, -6 and interferon (IFN)-gamma by cultured peripheral blood mononuclear cells (PBMC) in peritonitis-free CAPD patients. The correlation of cytokine production with plasma parathyroid hormone (PTH) and albumin levels was also evaluated. While the release of IL-1 beta was not markedly different from controls release of IL-6 from 24-hour cultured PBMCs was significantly greater than that of controls, (Mean +/- S.D., IL-6: 2186.8 +/- 1217.9 pg/ml, vs 1516.3 +/- 767.9, P <0.05). The addition of lipopolysaccharide (LPS, 10 micrograms/ml, significantly stimulated IL-1 beta and -6 production of PBMCs in CAPD patients and controls, compared to an unstimulated condition. The LPS-induced IL-1 beta production was also not markedly different from controls, whereas LPS-induced IL-6 production was significantly higher than controls (IL-6: 13,220.7 +/- 7177.4 vs 7411.4 +/- 1236.9, P <0.05). However, the percentage increases of IL-6 production stimulated with LPS in CAPD patients were not significantly different from controls (p > 0.05). No difference of baseline IFN-gamma was detected between CAPD patients controls, but phytohemagglutinin (PHA, 10 micrograms/ml)-stimulated IFN-gamma release was significantly higher in CAPD patients than controls (2425.9 +/- 1565.0 pg/ml vs 1364.0 +/- 755.1, P <0.05). There was no significant correlation between PTH and, IL-1 beta, serum albumin level and LPS-stimulated IL-6 production (r = 0.54, P <0.05). In conclusion, CAPD seems to partly induce activation of PBMCs with an enhanced release of IL-6 and IFN-gamma, and CAPD patients with higher serum albumin levels tend to show higher IL-6 production in immune response.
Adult
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Cells, Cultured
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Female
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Human
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Interferon Type II/biosynthesis*
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Interleukin-1/biosynthesis*
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Interleukin-6/biosynthesis*
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Male
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Middle Age
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Monocytes/metabolism
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Parathyroid Hormones/blood
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Peritoneal Dialysis, Continuous Ambulatory*
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Serum Albumin/analysis
10.Characterization of a small molecule inhibitor of tumor necrosis factor-alpha production.
Gao-yun YANG ; Zhi-qiang XIE ; Ge QIAN ; Wen-ying CUI ; Jun-yin ZHAO ; Jian-zhong ZHANG ; Shi LIAN
Chinese Medical Journal 2010;123(20):2883-2887
BACKGROUNDNumerous studies have shown that reducing the level of tumor necrosis factor-alpha (TNFα) through the use of anti-TNF antibodies or soluble TNF receptor is a safe and efficacious treatment to inflammatory diseases such as rheumatoid arthritis. Therefore, novel approaches to achieve this outcome are desired. The aim of this study was to investigate the characterization of a small molecule inhibitor, Y316, which blocks TNF mRNA upregulation and TNF production by lipopolysaccharides (LPS) stimulated monocytes.
METHODSPeripheral blood mononuclear cells (PBMC) from healthy volunteers were plated in 24-well plates and stimulated with LPS (1 µg/ml), phorbol-12-myristate-13-acetate (PMA) (100 ng/ml), zymosan (10 µg/ml) and Tsst (100 ng/ml). Supernatants were collected after 4-hour culture at 37°C, and quantitative determination of TNFα, interleukin-1β (IL-1β), IL-6, IL-8, IL-10 and IL-2 production in the supernatants was performed by colorimetric enzyme-linked immunosorbent assay (ELISA). Total RNA of PBMC was isolated and cytokine mRNA quantitation was performed by using a RNA level measuring kit (R & D Systems). PBMC were pretreated with Y316 (10 µmol/L, 1 µmol/L, 0.1 µmol/L, 0.01 µol/L and 0.001 µmol/L) or dimethyl sulfoxide at 37°C for 10 minutes, and then stimulated with LPS or PMA, protein concentrations of p44.42, IKBα, P38 and Jun NH2-terminal kinase were determined by Western blotting. Cyclic adenosine-3',5'-monophosphate (cAMP) of PBMC was measured by enzyme immunoassay kit (Amersham Pharmacia Biotech).
RESULTSY316 blocked TNF production and inhibited the upregulation of TNF mRNA levels in response to LPS, and also prevented the production of IL-1 and IL-6. In contrast, Y316 augmented the production of IL-10 in LPS-stimulated monocytes. Y316 failed to prevent the production of IL-2 and TNF in antigen-stimulated T cells, suggesting that its effects may be cell-type specific. Y316 prevented the phosphorylation and activation of the MAPK, ERK, and therefore appeared to mediate its effects on TNF by acting at an early point in the signaling cascade induced in response to LPS. There was no effect of Y316 on cAMP levels either alone or in the presence of LPS.
CONCLUSIONSY316 appears to be a small molecule inhibiting TNF production, which may act via a novel mechanism. Identification of the target of Y316 may lead to the development of alternative strategies for achieving selective cytokine inhibition.
Anti-Inflammatory Agents ; pharmacology ; Extracellular Signal-Regulated MAP Kinases ; metabolism ; Humans ; Interleukin-1 ; antagonists & inhibitors ; biosynthesis ; Interleukin-6 ; antagonists & inhibitors ; biosynthesis ; Lipopolysaccharides ; pharmacology ; Monocytes ; drug effects ; immunology ; Phosphorylation ; Tumor Necrosis Factor-alpha ; antagonists & inhibitors ; biosynthesis