Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P10145 (IL-8)
 23,849 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

While CD40-CD40 ligand interactions are known to regulate B cell proliferation and differentiation, much less is known about the role this receptor plays on other cell types, especially those of nonhemopoietic origin. We report here that CD40 is expressed in normal human epidermis in situ, especially on the basal cell layer, and that it is maintained on cultured epidermal basal cells. Immunoprecipitation and SDS-PAGE analysis confirms that CD40 expressed by epidermal basal cells is immunologically related to the B cell CD40. IFN-gamma up-regulates CD40 expression on cultured keratinocytes, whereas other proinflammatory cytokines, such as IL-1 or TNF-alpha, have little effects. Using CD40-ligand-transfected L cells (CD40Lc), we demonstrated that CD40 triggering results in an enhanced secretion of both IL-8 and TNF-alpha by cultured epidermal basal cells, suggesting that CD40-CD40L interactions may play a role in amplifying the cutaneous inflammatory reactions. More importantly, we found that keratinocyte proliferation was significantly inhibited when the cells were grown on CD40Lc, as compared with CD32-transfected, or nontransfected, L cells. This inhibitory effect can be reversed substantially by pretreatment of keratinocytes with anti-CD40 mAb. In addition, inhibition of proliferation could be obtained by adding a soluble form of CD40 ligand to the keratinocyte cultures. Interestingly, inhibition of keratinocyte proliferation on CD40Lc correlates with differentiation of the cells, as assessed by morphologic analysis and increased profilaggrin content. Collectively, these results demonstrate that CD40 is expressed and functional on human epidermal basal cells and that, on these cells, CD40 ligation may be a signal for limitation of cell growth and induction of differentiation.
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PMID:CD40 ligation of human keratinocytes inhibits their proliferation and induces their differentiation. 897 85

alpha 2-Macroglobulin (alpha 2m) is a major plasma proteinase inhibitor, as well as a carrier and regulator of the function of many cytokines. IL-8 is a potent neutrophil attractant and activator, and it plays an important role in the pathogenesis of adult respiratory distress syndrome (ARDS). The concentration of both IL-8 and alpha 2m is increased in lung fluids from patients with ARDS. Therefore, interaction of IL-8 with human alpha 2m was studied. Mixtures of native and methylamine-treated alpha 2m (fast alpha 2m) with 125I-labeled IL-8 were analyzed using nonreducing gel electrophoresis. 125I-labeled IL-8 exclusively bound to fast alpha 2m, and the binding could be inhibited by unlabeled IL-8. Analysis of the IL-8-alpha 2m interaction using SDS-PAGE gels indicated that the binding was mainly noncovalent. The affinity of the binding of alpha 2m to IL-8 was measured using an equilibrium dialysis technique, and Kd was 30 nM. Bioassays revealed that fast alpha 2m did not affect IL-8-induced neutrophil degranulation or chemotaxis. However, it protected IL-8 from proteolytic degradation. In addition, IL-8 complexed to alpha 2m was detected in lung fluids from patients with ARDS. alpha 2m may therefore modulate IL-8 function in the lung.
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PMID:Studies on the interaction of IL-8 with human plasma alpha 2-macroglobulin: evidence for the presence of IL-8 complexed to alpha 2-macroglobulin in lung fluids of patients with adult respiratory distress syndrome. 902 35

Connective tissue-activating peptide III (CTAP-III) and neutrophil-activating peptide-2 (NAP-2) are both derived from a common precursor, platelet basic protein (PBP), which is stored in the alpha-granules of platelets and released upon their activation. CTAP-III is an 85-residue peptide which is converted to NAP-2 by enzymic removal of the 15 amino-terminal residues. Both peptides play a role in the early stages of wound healing and inflammation through different activities. We have cloned the cDNA for PBP and expressed constructs coding for the CTAP-III and NAP-2 polypeptides in Escherichia coli. We have purified and renatured these recombinant proteins. The integrity of the recombinant proteins has been ascertained by in vitro bioassays. CTAP-III causes 51% histamine release from the basophilic cell lin KU812 at 10(-7) M, whereas NAP-2 only causes 28% release at the same concentration. In assays on human neutrophils, NAP-2 had an EC50 of 2 x 10(-8) M in chemotaxis, an EC50 of 3 x 10(-8) M for shape change, and could displace IL-8 from neutrophils with a Kd of 7.5 x 10(-9) M. CTAP-III had no activity in these assays. The disulfide bonds have been identified by peptide mapping and sequence analysis, and are in the positions predicted by homology to interleukin-8 and platelet factor 4. Measurement of the molecular mass at physiologic concentrations by gel permeation chromatography has shown that CTAP-III forms predominantly tetramers and dimers, whereas NAP-2 is only dimetric. SDS/PAGE analysis of samples cross-linked with disuccinimidyl suberate support these topologies. We postulate a mechanism for tetramer formation based on the interaction of the amino-terminal extension in CTAP-III involving a helix-helix interaction that could stabilize the association of two CTAP-III dimers.
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PMID:Structure and bioactivity of recombinant human CTAP-III and NAP-2. 905 6

An Escherichia coli K 12 strain has been constructed for efficient expression of recombinant biologically active human IL-8 (Interleukin-8). The development of a fermentation and purification process from the laboratory scale (cells from 15 l fermentation broth) to a production scale (cells from 200 l fermentation broth) is described. Material obtained from the laboratory scale was used for initial in vitro studies and for the development of a biological assay. An upscale purification process starting from 80 l fermentation broth resulted in larger amounts of IL-8 needed for preclinical studies. This process includes a fully automated control of the initial affinity chromatography step. Finally, a production process which differed markedly from the small-scale processes was tailor-made for GMP conformity and economic considerations. It consists of a cell disruption step followed by two crossflow diafiltrations with different molecular weight cut offs and filtration rates, one cation exchange chromatography and a final dialysis step. In order to enhance the overall yield of biologically active IL-8, conditions for a resolubilisation of insoluble IL-8 present in the remaining pellet after cell disruption were worked out.
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PMID:Production of human interleukin-8 expressed in Escherichia coli: from a laboratory scale for in vitro tests via a technical scale for animal studies to a process scale for a GMP-compatible production. 918 99

IL-8 and neutrophil-activating peptide-2 (NAP-2) are two closely related C-X-C chemokines that differ in their abilities to induce chemotaxis of human polymorphonuclear leukocytes (PMN). Although two IL-8R types are expressed by PMN, only CXCR2 binds NAP-2 and IL-8 with equally high affinity. By using enriched CXCR2-transfected 293 cells, we show that high doses of IL-8 induce attenuation of chemotaxis, while equivalent doses of NAP-2 do not. Phosphorylation analysis shows that IL-8 induces higher levels of phosphorylation of the carboxyl terminus of CXCR2 than does NAP-2, suggesting that the level of phosphorylation contributes to the ability of the chemokines to attenuate the chemotactic response. To directly evaluate this difference, we analyzed the ability of receptors mutated to delete regions that highly express potential phosphorylation sites to be phosphorylated and to mediate chemotactic attenuation. We found that a carboxyl terminus-truncated mutant of CXCR2 was not phosphorylated by high doses of IL-8, as determined by in vivo phosphorylation assays and by analysis of the electrophoretic mobility of the receptors on SDS-PAGE gels. This mutated receptor had a significantly lower ability to attenuate IL-8-induced chemotaxis, indicating that the attenuation of chemotaxis is mediated by chemokine-induced receptor phosphorylation. In conclusion, the data show that the greater ability of IL-8 to induce receptor phosphorylation contributes to its more potent attenuation of chemotaxis as compared with NAP-2. This differential phosphorylation by IL-8 and NAP-2 of CXCR2 provides a basis for the divergent outcome of PMN-induced inflammation in response to these two closely related C-X-C chemokines.
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PMID:The differential ability of IL-8 and neutrophil-activating peptide-2 to induce attenuation of chemotaxis is mediated by their divergent capabilities to phosphorylate CXCR2 (IL-8 receptor B). 919 Sep 46

IL-8, a neutrophil chemotactic agent, is involved in a large number of neutrophil-driven acute and chronic inflammatory diseases. We have found that hamycin, an antifungal agent, reduces IL-8-induced migration and binding of 125I-labeled IL-8 to neutrophils by 66 and 75%, respectively. Other IL-8-induced biologic functions, such as superoxide generation, intracellular Ca2+ mobilization, and enzyme release were also reduced in hamycin-treated cells by 50 to 75%. Anti-IL-8R Ab (C-X-CR1) and IL-8 itself failed to protect the cells from the effect of hamycin. Scatchard analysis of IL-8 binding data demonstrated that while the normal cells expressed 23,000 +/- 1,704 receptors/cell (Kd = 3.5 nM), the number was reduced to 8,000 +/- 592 receptors/cell (Kd = 3.43 nM) in hamycin-treated cells. Chemical cross-linking of 125I-labeled IL-8 to its receptor followed by 10% SDS-PAGE analysis and autoradiography showed that the signals in hamycin-treated cells were considerably reduced compared with those in controls. In the immunoblot, however, the signals in control and hamycin-treated cells were almost identical. The intensity of the fluorescence emission of diphenyl hexatriene at 430 nm and membrane microviscosity measured by diphenyl hexatriene were considerably reduced in hamycin-treated cells, resulting in a reduced number of functional IL-8R, presumably by conformational change in the receptor. The study suggests that hamycin may be a potent immunomodulator of the IL-8R for alleviation of inflammatory distress.
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PMID:Hamycin inhibits IL-8-induced biologic response by modulating its receptor in human polymorphonuclear neutrophils. 936 32

The ability of polymorphonuclear leukocytes (PMNs) to modulate endothelial cell (EC) activation was investigated. Adding PMNs to cultured HUVECs resulted in a release of IL-6 (888 +/- 71 pg/ml, a 35-fold increase over release by the two cell types alone) and IL-8 (45.2 +/- 14.5 ng/ml, a 6.4-fold over PMN release alone and a 173-fold increase over EC release alone). In contrast, the release of TNF-alpha, IL-1beta, and platelet-derived growth factor was not affected by the EC-PMN coculture. Neutralizing mAbs to ICAM-1 or beta2 integrins or a physical segregation of PMNs and ECs did not reduce EC stimulation. In contrast, cell-free supernatants of PMNs recapitulated EC activation with an 18-fold up-regulation of EC IL-6 mRNA. The filtration of PMN supernatant or PMN pretreatment with metabolic antagonists or membrane cross-linking agents all suppressed EC activation. By flow cytometry, PMNs released in the supernatant, heterogeneous membrane-derived microparticles containing discrete proteins of 28 to 250 kDa as resolved by SDS-PAGE. PMN microparticle formation was enhanced by inflammatory stimuli, including formyl peptide and phorbol ester, and was time-dependent, reaching a plateau after a 1-h incubation from stimulation. Purified PMN microparticles induced EC IL-6 release in a reaction that was quantitatively indistinguishable from that observed with unfractionated PMN supernatant and unaffected by a neutralizing Ab to soluble IL-6R. These findings demonstrate that membrane microparticles released from stimulated PMNs are competent inflammatory mediators to produce EC activation and cytokine gene induction.
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PMID:Endothelial cell activation by leukocyte microparticles. 978 Feb 16

We investigated whether intercellular adhesion molecule-1 (ICAM-1) transduces outside-in signals for the production of chemokines IL-8 and RANTES in endothelial cells. Cross-linking of ICAM-1 induced IL-8 and RANTES mRNA expressions and increased their protein synthesis and secretions in endothelial cells. Furthermore, ICAM-1 cross-linking activated 44- and 42-kDa mitogen-activated protein (MAP) kinases (ERK1 and ERK2) in endothelial cells, as indicated by the electrophoretic mobility shift of MAP kinases on SDS-polyacrylamide gels. Finally, the specific MEK inhibitor PD98059 inhibited ICAM-1-induced IL-8 and RANTES production in endothelial cells. Taken together, these results indicate that stimulation of ICAM-1 induces IL-8 and RANTES production through the activation of 44- and 42-kDa MAP kinases in endothelial cells, suggesting that ICAM-1-induced chemokine production in endothelial cells would further attract and activate leukocytes to induce intense inflammation.
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PMID:Cross-linking of intercellular adhesion molecule-1 induces interleukin-8 and RANTES production through the activation of MAP kinases in human vascular endothelial cells. 978 8

The native GroEL-like protein was purified from Campylobacter rectus, a putative periodontal pathogen, by affinity chromatography on ATP-agarose followed by high performance liquid chromatography on Superose 6. The purified 64-kDa protein (denatured form of GroEL-like protein) was immunoreactive by SDS-PAGE and Western immunoblotting with the monoclonal antibody directed against heat shock protein 60 of human origin. The native GroEL-like protein stimulated both interleukin-6 (IL-6) and IL-8 secretion by a confluent monolayer of human gingival fibroblast in their culture supernatant. During the 22-h incubation, the amounts of IL-6 and IL-8 were increased by 5.4- and 3.5-fold, respectively. These data suggested that the GroEL-like protein might be considered to be a virulence factor of C. rectus in periodontal disease.
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PMID:The GroEL-like protein from Campylobacter rectus: immunological characterization and interleukin-6 and -8 induction in human gingival fibroblast. 978 45

The main goal of the present study was to investigate the response of the human skin equivalent Apligraf in vitro to the application of irritant substances and its predictivity as a screening tool for cumulative skin irritant potential in humans. Vaseline, calcipotriol, trans-retinoic acid, and sodium lauryl sulfate were applied to Apligraf in vitro for 24 h. Cell viability (lactate dehydrogenase leakage), release and mRNA expression of the proinflammatory cytokines IL-1alpha and IL-8, and morphological changes were assessed. The same products were applied to 30 healthy volunteers in a double-blind, randomized, vehicle-controlled within-subject study. The skin reactions after repeated 24-h applications over 3 weeks under Finn chamber patches were monitored by visual scoring and biophysical methods (trans-epidermal water loss, chromametry, and blood flow). Sodium lauryl sulfate was cytotoxic to Apligraf, and increased the release and expression of cytokines at low (0.2%, 0. 4%), but not at high (0.8%, 1%) concentrations. It induced severe irritancy in vivo. Trans-retinoic acid increased the expression and release of cytokines with no detectable cytotoxicity and showed moderate irritancy in humans. Although calcipotriol did neither affect cell viability nor the production of cytokines, it induced morphological signs of irritation and was mildly irritant for healthy volunteers. Vaseline was innocuous in vivo and induced no changes in Apligraf. In conclusion, the cumulative skin irritation potential of the tested products could be predicted with Apligraf in a sensitive and specific manner, by monitoring cytotoxicity, proinflammatory cytokines, and morphological changes.
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PMID:Use of human skin equivalent Apligraf for in vitro assessment of cumulative skin irritation potential of topical products. 1073 42


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