Contribution of group II phospholipase A2 to arachidonic acid release and prostaglandin synthesis by mesangial cells

Interleukin 1 (IL-1) and tumour necrosis factor (TNF) have been found to increase group II phospholipase A₂ (PLA₂) synthesis and secretion by mesangial cells. In all cases 85%–90% of the enzyme is secreted from the cells and a parallel increase in prostaglandin (PGE₂) synthesis is observed. We report here that co-incubation with a monoclonal antibody that specifically binds and neutralizes rat group II PLA₂ attenuates IL-1 and TNF-stimulated PGE₂ production by 45% and 52%, respectively. CGP43182, a specific inhibitor of group II PLA₂, potently blocks IL-1- and TNF-stimulated PGE₂ synthesis in intact mesangial cells with 1C₅₀s of 1.3 and 1.0 M, respectively.     

白细胞介素-10抑制人肾系膜细胞环氧化酶-2表达及其催化的前列腺素E2释放的实验研究

目的:观察IL-10对IL-1β诱导的人系膜细胞(HMC)前列腺素E₂(PGE₂)释放及环氧化酶-2(cyclooxygenase-2,COX-2)基因和蛋白表达的影响。方法:应用放射免疫测定法检测HMC培养上清中PGE₂,应用RT-PCR和Westernblot分别检测COX-2mRNA和蛋白水平。结果:①IL-1β显著上调PGE₂释放及COX-2基因和蛋白的表达(P均＜0.01);②IL-10对基础状态下PGE₂释放及COX-2基因和蛋白表达无明显影响(P＞0.05);③IL-10可呈剂量依赖性地下调IL-1β诱导的PGE₂释放及COX-2mRNA和蛋白表达(P＜001)。结论:IL-10抑制IL-1β诱导的HMCPGE₂释放及COX-2表达,提示IL-10对HMC具有多方面抗炎作用。     

Interleukin-1β induces cytosolic PLA2 in parallel with prostaglandin E2 in rheumatoid synovial fibroblasts

We investigated the temporal relationship between the increase in enzymatic activity and protein of a high molecular weight (100 kDa), cytosolic PLA₂ (cPLA₂) in interleukin-1β (IL-1β)-treated rheumatoid synovial fibroblasts (RSF). Both of these responses increased according to a similar time-course which correlates with PGE₂ production by these cells. In contrast, 14 kDa, secreted PLA₂ (sPLA₂), which was also produced by RSF, was not affected by IL-1β treatment. These findings support that an augmentation of cPLA₂ activity, caused by an induction of cPLA₂ protein, rather than sPLA₂, is temporally associated with increased PGE₂ production in IL-1β-treated RSF.     

Anti-inflammatory Effects of Interleukin-4, Interleukin-10, and Transforming Growth Factor-β on Human Placental Cells In Vitro

PROBLEM: To determine the effects of anti-inflammatory cytokines on the production of inflammatory mediators by placental cells. METHOD OF STUDY: Cells from term human placentas were isolated and cultured in vitro in the presence of anti-inflammatory cytokines interleukin (IL)-4, IL-10, and transforming growth factor (TGF)-β. Their effects on the production of IL-8 and prostaglandin E₂ (PGE₂) were investigated under basal conditions and after stimulation with IL-1β and tumor necrosis factor (TNF)-α. RESULTS: Both IL-10 and IL-4 inhibited IL-1β- and TNF-α-induced PGE₂ production but had no significant effects on the production of PGE₂ under basal conditions. TGF-β₁ was without effect in stimulated cells, whereas under basal conditions TGF-β₁ stimulated PGE₂ production. Similar trends were seen for IL-8 production, with the exceptions that TGF-β₁ decreased the TNF-α-induced production and IL-4 decreased basal IL-8 production. CONCLUSIONS: The anti-inflammatory effects shown by IL-4, IL-10, and (to lesser extent) TGF-β may play a role in ameliorating the potentially harmful effects of pro-inflammatory mediators in the feto-placental unit.     

Differential inhibitory effects of indomethacin,dexamethasone, and interferon-gamma (IFN-γ) on IL-11 production by rheumatoid synovial cells

IL-11, a member of the IL-6 type cytokines, has some biological activity related to the joint destruction in rheumatoid arthritis (RA), such as induction of osteoclast differentiation. However, its expression and regulation in rheumatoid inflamed joints has not been clarified. In the present study we examined the capacity of fresh rheumatoid synovial cells (fresh RSC) to produce IL-11, and the effect of indomethacin, dexamethasone and IFN-γ on IL-11 production. Fresh RSC obtained from eight patients with RA produced large amounts of IL-11, measured by ELISA, and showed strong expression of IL-11 mRNA, determined by Northern blotting. Indomethacin inhibited the production of IL-11 by about 55%. Prostaglandin E₂ (PGE₂) completely prevented the inhibition, suggesting that IL-11 production by fresh RSC was in part mediated by PGE₂. Dexamethasone inhibited the production of IL-11 by more than 80%. Interestingly, the inhibition was not abolished by PGE₂. IFN-γ inhibited the production of IL-11 from IL-1α-stimulated cultured rheumatoid synovial fibroblasts, although IFN-γ did not inhibit the production of IL-11 by fresh RSC. These results suggest that the production of IL-11 by rheumatoid synovia was differentially regulated by PGE₂ and IFN-γ, and that treatment with indomethacin or dexamethasone decreased the level of IL-11 at inflammatory joints in patients with RA.     

Production of tumor necrosis factor,interleukin-6 and prostaglandin E2 by LPS-stimulated rat bone marrow macrophages after thermal injury: Effect of indomethacin

The effect of thermal injury on the in vitro production of TNF, IL-6, and PGE₂ by bone marrow-derived, LPS-stimulated rat macrophages was studied. Thermal injury caused a general hyperactivity in the production of the mediators by the cells. Indomethacin, a cyclooxygenase inhibitor of PGE₂ synthesis, inhibited the production of IL-6 and PGE₂ but had no effect on the production of TNF. These results suggest that the observed low concentration of PGE₂ produced by the cells was insufficient to cause inhibition of TNF synthesis; thus, the effect of indomethacin would be undetectable. The results also suggest that indomethacin may act directly in inhibiting the production of IL-6 by the macrophages. The hyperactive effect of thermal injury on the production of inflammatory mediators by newly differentiated bone marrow derived macrophages can be important in the overall systemic response to the insult.     

Comparison of PGE2, 6-keto PGF1α and renin release from dog kidneys

Several renal cell types synthesize prostaglandin E₂ (PGE₂) and prostacyclin (PGI₂). To examine whether the release of these prostaglandins varies in proportion, prostaglandin synthesis was stimulated in anaesthetized dogs by renal arterial constriction, ureteral occlusion, intrarenal angiotensin II infusion and infusion of arachidonic acid, the precursor of PG synthesis. PGI₂ was measured as its stable hydrolysed product, 6-keto PGF_1α. The two former procedures raised PGE₂ release to 13 ± 2 pmol min^-1, 6-keto PGF_1α release to 5 ± 2 pmol min^-1 and renin release to 23 ± 5 μg AI min^-1, Angiotensin II infusion, reducing the renal blood flow by 30%, increased PGE₂ and 6-keto PGF_1α release only half as much as ureteral and renal arterial constriction, and exerted no significant effect on renin release. By increasing the infusion rate of angiotensin II up to 10 times, the renal blood flow remained unaltered in four dogs and fell to 50% of control in two dogs, but PGE₂ and 6-keto PGF_1α release did not increase further in any of the experiments. Arachidonic acid, infused at 40 and 160 μg kg^-1 min^-1, increased prostaglandin release in proportion to the infusion rate. At the highest infusion rate, PGE₂ release averaged 166± 37 pmol min^-1 and 6-keto PGF_1α release 98 ± 28 pmol min^-1. All procedures increased PGE₂ and 6-keto PGF_1α release in a fixed proportion of about 2.5:1, whereas renin release increased only during autoregulatory vasodilation.     

Prostaglandin E2 primes naive T cells for the production of anti-inflammatory cytokines

In addition to their capacity to induce pain, vasodilatation and fever, prostaglandins E (PGE) exert anti-inflammatory activities by inhibiting the release of pro-inflammatory cytokines by macrophages and T cells, and by increasing interleukin (IL)-10 production by macrophages. We here report that PGE₂, the major arachidonic acid metabolite released by antigen-presenting cells (APC), primes naive human T cells for enhanced production of anti-inflammatory cytokines and inhibition of pro-inflammatory cytokines. Unfractionated as well as CD45RO^?CD31⁺ sort-purified neonatal CD4 T cells acquire the capacity to produce a large spectrum of cytokines after priming with anti-CD3 and anti-CD28 monoclonal antibodies (mAb), in the absence of both APC and exogenous cytokines. PGE₂ primes naive T cells in a dose-dependent fashion for production of high levels of IL-4, IL-10 and IL-13, and very low levels of IL-2, interferon (IFN)-γ, tumor necrosis factor (TNF)-α, and TNF-β. PGE₂ does not significantly increase IL-4 production in priming cultures, whereas it suppresses IL-2 and IFN-γ. Addition of a neutralizing mAb to IL-4 receptor in primary cultures, supplemented or not with PGE₂, prevents the development of IL-4-producing cells but does not abolish the effects of PGE₂ on IL-10 and IL-13 as well as T helper (Th)1-associated cytokines. Addition of exogenous IL-2 in primary cultures does not alter the effects of PGE₂ on naive T cell maturation. Thus PGE₂ does not act by increasing IL-4 production in priming cultures, and its effects are partly IL-4 independent and largely IL-2 independent. Together with the recent demonstration that PGE₂ suppresses IL-12 production, our results strongly suggest that this endogenously produced molecule may play a significant role in Th subset development and that its stable analogs may be considered for the treatment of Th1-mediated inflammatory diseases.     

Cytokine-induced inhibition of bone matrix proteins is not mediated by prostaglandins

Interleukin-1 (IL-1) and tumor necrosis factor (TNF), two pleiotropic cytokines produced in inflammatory processes, inhibit bone matrix biosynthesis and stimulate prostanoid formation in osteoblasts. In the present study, the importance of prostaglandin formation in IL-1 and TNF-induced inhibition of osteocalcin and type I collagen formation has been examined. In the human osteoblastic cell line MG-63, IL-1 (10–1000 pg/ml), IL-1 (3–300 pg/ml) and TNF- (1–30 ng/ml) stimulated prostaglandin E₂ (PGE₂) formation and inhibited, 1,25(OH)₂-vitamin D3-induced osteocalcin biosynthesis as well as basal production of type I collagen. Addition of PGE₂ or increasing the endogenous formation of PGE₂ by treating the cells with arachidonic acid, bradykinin, Lys-bradykinin or des-Arg⁹-bradykinin, did not affect osteocalcin and type I collagen formation in unstimulated or 1,25(OH)₂-vitamin D3-stimulated osteoblasts. Four non-steroidal anti-inflammatory drugs, indomethacin, flurbiprofen, naproxen and meclofenamic acid, inhibited basal, IL-1 - and TNF--stimulated PGE₂ formation in the MG-63 cells without affecting IL-1 - or TNF--induced inhibition of osteocalcin and type I collagen formation. In isolated, non-transformed, human osteoblast-like cells, IL-1 and TNF- stimulated PGE₂ formation and concomitantly inhibited 1,25(OH)₂-vitamin D3-stimulated osteocalcin biosynthesis, without affecting type I collagen formation. In these cells, indomethacin and flurbiprofen abolished the effects of IL-1 and TNF- on prostaglandin formation without affecting the inhibitory effects of the cytokines on osteocalcin biosynthesis. These data show that IL-1 and TNF inhibit osteocalcin and type I collagen formation in osteoblasts independently of prostaglandin biosynthesis and that non-steroidal antiinflammatory drugs do not affect the effects of IL-1 and TNF on bone matrix biosynthesis.accepted by W. B. van den Berg     

Absence of modulation of monokine production via endogenous cyclooxygenase or 5-lipoxygenase metabolites: MK-886 (3-[1-(4-chlorobenzyl)-3-t-butyl-thio-5-isopropylindol-2-yl]-2,2-dimethylpropanoic acid), indomethacin,or arachidonate fail to alter immunoreactive interleukin-1β, or TNF-α production by human monocytes in vitro

Human peripheral blood monocytes exposed to MK-886 (3-[1-(4-chlorobenzyl)-3-t-butyl-thio-5-isopropylindol-2-yl]-2,2-dimethylpropanoic acid) at doses which abolish formation of 5-lipoxygenase metabolites showed unaltered interleukin-1β (IL-1β) or tumor necrosis factor-α (TNF-α) levels in response to phorbol ester, concanavalin A, serum-treated zymosan, or lipopolysaccharide. Indomethacin (10 μM), alone or in combination with MK-886, also failed to modulate monokine production in response to any stimulus. Exogenous arachidonate (3–30 μM) which augmented the formation of PGE₂ and LTB₄ in the absence of stimulation, also had no effect on monokine production. LPS-induced IL-1 and TNF production occurred despite stimulation of PGE₂ synthesis. The results make a role for endogenous prostaglandins and leukotrienes in the regulation of monocyte IL-1β and TNF-α production unlikely. These data also indicate that MK-886, a novel inhibitor of 5-lipoxygenase product formation, is a potentially useful leukotriene inhibitor which does not affect monokine production.     

Pro-inflammatory cytokines and prostaglandins induce maturation of potent immunostimulatory dendritic cells under fetal calf serum-free conditions

Culture conditions for human dendritic cells (DC) have been developed by several laboratories. Most of these culture methods, however, have used conditions involving fetal calf serum (FCS) to generate DC in the presence of granulocyte-macrophage colony-stimulating factor and interleukin (IL)-4. Recently, alternative culture conditions have been described using an additional stimulation with monocyte-conditioned medium (MCM) and FCS-free media to generate DC. As MCM is a rather undefined cocktail, the yield and quality of DC generated by these cultures varies substantially. We report that a defined cocktail of tumor necrosis factor (TNF)-α, IL-1β and IL-6 equals MCM in its potency to generate DC. Addition of prostaglandin (PG)E₂ to the cytokine cocktail further enhanced the yield, maturation, migratory and immunostimulatory capacity of the DC generated. More importantly, culture conditions also influenced the outcome of the T cell response induced. DC cultured with TNF-α/IL-1/IL-6 or MCM alone induced CD4⁺ T cells that release intermediate levels of interferon (IFN)-γ and no IL-4 or IL-10. Production of IFN-γ was significantly induced by addition of PGE₂, while no effect on production of IL-4 or IL-10 was observed. Even more striking differences were observed for CD8⁺ T cells. While MCM conditions only induced IFN-γ^low, IL-4^neg cells, TNF-α/IL-1/IL-6 promoted growth of IFN-γ^intermediate, IL-4^neg CD8⁺ T cells. Addition of PGE₂ again only further polarized this pattern enhancing IFN-γ production by alloreactive CD8⁺ T cells in both cultures without inducing type 2 cytokines. Taken together, the data indicate that the defined cocktail TNF-α/IL-1/IL-6 can substitute for MCM and that addition of PGE₂ further enhances the yield and quality of DC generated. TNF-α/IL-1, IL-6 + PGE₂-cultured DC seem to be optimal for generation of IFN-γ-producing CD4/CD8⁺ T cells.     

Effects of the novel immunosuppressant FTY720 in a murine rheumatoid arthritis model

We investigated the effects and mechanisms by which FTY720 (FTY) inhibits arthritis development in the SKG mouse rheumatoid arthritis (RA) model. FTY (1 mg/kg/day) administration suppressed the progression of laminarin-induced arthritis in SKG mice. FTY treatment decreased IL-6 and TNF-α expression in synovial fibroblast cells and the number of inflammatory cells overall. Bone destruction was also suppressed by treatment with FTY. The numbers of CD4⁺ and CD8⁺ T cells were significantly increased in the thymus and decreased in the spleen in FTY-treated SKG mice. FTY enhanced IL-4 production by CD4⁺ T cells stimulated by allogeneic spleen cells and inhibited prostaglandin E₂ (PGE₂) production by a TNF-α-stimulated synovial fibroblast cell line. These results indicate that FTY can inhibit arthritis in SKG mice via sequestration of autoimmune CD4⁺ T cells in the thymus, enhancement of Th2 immune responses, and inhibition of PGE₂ production by synovial cells.     

Tumour-derived prostaglandin E2 and transforming growth factor-β synergize to inhibit plasmacytoid dendritic cell-derived interferon-α

In previous studies we reported that plasmacytoid dendritic cells (PDC) infiltrating head and neck cancer tissue are functionally impaired, but the molecular basis for the functional deficiency remained unclear. Here we demonstrate that tumour-derived prostaglandin E2 (PGE₂) and transforming growth factor-β (TGF-β) increase interleukin-8 (IL-8) but synergistically inhibit interferon-α (IFN-α) and tumour necrosis factor (TNF) production of Toll-like receptor 7 (TLR7)- and Toll-like receptor 9 (TLR9)-stimulated PDC. The inhibitory effect of PGE₂ could be mimicked by the induction of cyclic AMP (cAMP) and by inhibitors of cyclooxygenase. The contribution of tumour-derived TGF-β was confirmed by the TGF-β antagonist SB-431542. Suppression of tumour-derived PGE₂ and TGF-β restored TLR-induced IFN-α production of PDC. Additionally, PGE₂- and TGF-β-treated PDC display a ‘tolerogenic’ phenotype because of a downregulation of CD40 accompanied by an upregulation of CD86. Finally, in TLR-stimulated PDC, PGE₂ and TGF-β reduce the CCR7 : CXCR4 ratio, suggesting that PDC are impaired in their ability to migrate to tumour-draining lymph nodes but are retained in stromal cell-derived factor 1 (SDF-1)-expressing tissues. Based on these data, cyclooxygenase inhibitors and TGF-β antagonists may improve TLR7- and TLR9-based tumour immunotherapy.     

Comparative Studies on the Effects of Interleukin-4 and Interleukin-13 on Cytokine and Prostaglandin E2 Production by Amnion-derived WISH Cells

PROBLEM: In hematopoietic cells, interleukin (IL)-13 shares many actions with IL-4. The effects of IL-13 in gestational tissues have yet to be reported, however. We compared the effects of IL-4 and IL-13 on the production of cytokines and prostaglandin E₂ (PGE₂) in epithelial amnion-derived WISH cells. METHOD OF STUDY: WISH cells were treated with IL-4 or IL-13 (0.08–10 ng/ml) with/without cotreatment with IL-1β (0.2 ng/ml), tumor necrosis factor-α (10 ng/ml) or epidermal growth factor (5 ng/ml). The production of IL-6, IL-8, and PGE₂ was measured by immunoassay after 16 hr. RESULTS: Both IL-4 and IL-13 inhibited PGE₂ production with indistinguishable concentration-response curves, under basal or stimulated conditions. The maximal inhibition of IL-1β-stimulated PGE₂ production (to 28% ± 10% of control) was seen at 10 ng/ml of IL-4 or IL-13. Basal IL-6 production was stimulated approximately twofold by IL-4 and IL-13, whereas IL-4 and IL-13 both inhibited cytokine-stimulated (but not basal) IL-8 production by approximately 50%. In the presence of 1 ng/ml of IL-4, IL-13 was unable to further inhibit PGE₂ production. CONCLUSIONS: The inhibition of PGE₂ and IL-8 production by IL-4 in WISH cells is mimicked by IL-13. Both cytokines, probably through binding to a common receptor complex, may share a role in suppressing inflammatory reactions within intrauterine tissues.     

Effects of topical corticosteroids on inflammatory mediator–induced eicosanoid release by human airway epithelial cells

Background: Airway epithelial cells are among the first cells to come in contact with aerosolized corticosteroids. However, the relative potencies and time course of action of the several commonly used aerosolized corticosteroids on eicosanoid production by airway epithelial cells are unknown. Objectives: This study compared the effects of fluticasone, budesonide, and triamcinolone on eicosanoid output by human airway epithelial cells in vitro. We also determined the spectrum of eicosanoids affected and the mechanism for corticosteroid action. Methods: Cultured BEAS-2B airway epithelial cells (a transformed cell line) were exposed to corticosteroids (1 nmol/L to 1 μmol/L) for 2 to 48 hours and then assayed for basal- and bradykinin (BK)-stimulated eicosanoid output. The eicosanoid profile was identified by HPLC in tritiated arachidonic acid prelabelled cells, and PGE₂, the major eicosanoid product, was quantitated by RIA. The effect of corticosteroids on the immunoreactivity of key proteins involved in eicosanoid metabolism (ie, cyclooxygenase [COX], phospholipase A2 [PLA₂], and Clara cell protein, a PLA₂ inhibitor) was determined by Western blotting. Results: Eicosanoid output was largely confined to prostaglandins with values of 5 ± 2 and 82 ± 35 ng PGE₂/10⁶ cells for basal- and BK stimulation, respectively (n = 8). All 3 corticosteroids inhibited basal- and BK-induced PGE₂ output in a dose- and time-dependent manner. Fluticasone and budesonide completely eliminated PGE₂ output in nanomolar concentrations in contrast to triamcinolone, which required micromolar concentration. The rank order of potency was: fluticasone = budesonide > triamcinolone. The time course of action for PGE₂ inhibition also differed, with budesonide acting more slowly than the other 2 corticosteroids (P = .04). All 3 corticosteroids markedly reduced COX2 with little effect on COX1, cPLA₂ (Type IV), or iPLA₂ (Type VI) immunoreactivity or their relative distribution in cytosol versus membrane fractions. Clara cell protein immunoreactivity was undetectable in control and corticosteroid-treated cell lysates. Conclusion: These results show that in a human airway epithelial cell line, the 3 inhaled corticosteroids commonly used to treat asthma differ in onsets of action as inhibitors of prostaglandin synthesis and vary considerably in potency. All 3 corticosteroids act mechanistically in similar fashion by inhibiting COX2 synthesis. (J Allergy Clin Immunol 1999;103:1081-91.)     

The kallikrein-kininogen-kinin system in chronic inflammation

We examined bradykinin's effects on macrophages and fibroblasts, two cell types important in the pathogenesis of chronic inflammation. Bradykinin stimulated release of proteins of 18 kDa from macrophages. These proteins caused increased thymocyte proliferation (interleukin 1, IL-1) and completely inhibited lipoprotein lipase (tumor necrosis factor, TNF). When fibroblasts were incubated with bradykinin, PGE₂ synthesis was stimulated. Pretreatment with IL-1 or TNF dramatically amplified bradykinin-stimulated PGE₂ synthesis. Thus, bradykinin is involved in a positive feedback loop in which bradykinin activates macrophages to release potent inflammatory cytokines; these in turn amplify responsiveness of bradykinin target tissues.     

Correlation of leukocyte interleukin-1 production with the stimulation of prostaglandin and tissue factor synthesis by human umbilical vein endothelial cells

Human leukocyte suspensions (neutrophils 80–85%, monocyte 15–20%) were incubated alone or with cultured human umbilical vein endothelial cells. Leukocytes were either directly added to the endothelial cell cultures or separated from them by a 0.4 micron insert filter. Supernatants or cell lysates were obtained at 0.5, 1, 2, and 4 hours of incubation. Supernatants were assayed for the prostacyclin (PGI₂) metabolite 6-keto prostaglandin F₁ and prostaglandin E₂ (PGE₂) by radioimmunoassay and for interleukin-1 (IL-1) by the thymocyte co-mitogen assay. Cell lysates were analyzed for cell-associated procoagulant activity (PCA). Co-incubation of endothelial cells with leukocytes stimulated the synthesis of PGI₂, PGE₂, and PCA. These biochemical changes correlated partially with the release of IL-1 beta. The results suggest that IL-1 released in monocyte/neutrophil co-cultures can produce prothrombotic (increased PCA expression) and inflammatory changes (increased synthesis of vasodilatory and permeability enhancing PGI₂ and PGE₂) in endothelial cells. Neutrophils may represent a source of the released IL-1 and/or may act to stimulate monocyte release of this cytokine and thus play an important role in vascular pathology by a mechanism unrelated to their more direct cytotoxic activity.     

Echinocystic Acid Inhibits IL-1β-Induced COX-2 and iNOS Expression in Human Osteoarthritis Chondrocytes

Echinocystic acid (EA), a pentacyclic triterpene isolated from the fruits of Gleditsia sinensis Lam, displays a range of pharmacological activities including anti-inflammatory and antioxidant effects. However, the effect of EA on IL-1β-stimulated osteoarthritis chondrocyte has not been reported. The purpose of this study was to assess the effects of EA on IL-1β-stimulated human osteoarthritis chondrocyte. Chondrocytes were stimulated with IL-1β in the absence or presence of EA. NO and PGE₂ production were measured by Griess reagent and ELISA. The expression of COX-2, iNOS, nuclear factor-κB (NF-κB), inhibitory kappa B (IκBα), c-Jun N-terminal kinase (JNK), p38, and extracellular signal-regulated kinase (ERK) were detected by Western blot analysis. The results showed that EA suppressed IL-1β-induced collagenase-3 (MMP-13), NO, and PGE₂ production in a dose-dependent manner. IL-1β up-regulated the expression of COX-2 and iNOS, and the increase was inhibited by EA. Furthermore, IL-1β-induced NF-κB and mitogen-activated protein kinase (MAPK) activation were inhibited by EA. In conclusion, EA effectively attenuated IL-1β-induced inflammatory response in osteoarthritis chondrocyte which suggesting that EA may be a potential agent in the treatment of osteoarthritis.     

Febrile response induced by cecal ligation and puncture (CLP) in rats: involvement of prostaglandin E<Subscript>2</Subscript> and cytokines

The purpose of the present study was to better understand the events involved in the febrile response induced by cecal ligation and puncture (CLP), a complex infectious process. To this end, we conducted in vivo experiments in rats examining (1) fever development, (2) bacterial number in the infection focus and in blood, (3) peripheral and hypothalamic synthesis of cytokines, (4) hypothalamic and cerebrospinal fluid (CSF) synthesis of prostaglandin E₂ (PGE₂), (5) the effect of anti-IL-6 antibody on fever, and (6) the effect of celecoxib on fever and hypothalamic synthesis of PGE₂ after CLP induction. We found that CLP promotes fever and animal death depending on the number of punctures. The peak of CLP-induced fever overlapped with the maximal increase in the number of bacteria in the infectious focus and blood, which occurred at 6 and 12 h. The peak of the febrile response also coincided with increased amounts of interleukin (IL)-1β, IL-6 and IL-10 in the peritoneal exudate and serum; IL-6 in the hypothalamus and PGE₂ in the CSF and predominantly in the hypothalamus. Moreover, intracerebroventricularly injected anti-IL-6 antibody reduced the febrile response while celecoxib reduced the fever and PGE₂ amount in the hypothalamus induced by CLP. Tumor necrosis factor (TNF)-α peaked at 3 h at all sites studied. Conversely, IL-10 concentration decreased in the hypothalamus. These findings show that the peak of CLP-induced fever is accompanied by an increase of bacteria in peritoneal fluid (local infection) and blood; local synthesis of pyrogenic (IL-1β, IL-6) and antipyretic (IL-10) cytokines and central production of IL-6 and PGE₂, suggesting that these last are the central mediators of this response.