Role of IL-1b and TNF-a in the regulation of NGF in experimentally induced arthritis in mice

The aim of this study was to investigate the effects of IL-1β and TNF-α on NGF levels in the knee joint in experimentally induced arthritis in adult mice. Out data showed that IL-1β, but not TNF-α, induces an increase in NGF levels, while concomitant injection of both cytokines enhances the effect of IL-1β on NGF presence. Analysis of NGF levels in the knee joint of carrageenan- and IL-1β-induced inflammation after administration of antibodies against TNF-α supports this hypothesis. Our studies also showed that exogenous administration of NGF antibody reduces the enhanced level of TNF-α occurring in arthritic mice. This latter observation indirectly suggests that NGF is implicated in the upregulation of TNF-α in these animal models of joint inflammation. The functional significance of NGF or NGF antibody in inflammatory arthritis is discussed. Received: 16 March 1998 / Accepted: 2 July 1998     

Association between Serum Inflammatory Cytokines and Disease Activity in Juvenile Idiopathic Arthritis

Circulating interleukin-1β (IL-1β), IL-6, tumour necrosis factor-α (TNF-α), osteocalcin, and conventional parameters of inflammation were examined serially in 14 children with juvenile idiopathic arthritis (JIA) to determine any correlation with the disease activity. Serum IL-1β was undetectable in all JIA patients. Serum IL-6, white blood cell counts, platelet counts, erythrocyte sedimentation rate and C-reactive protein levels were significantly elevated in the active phase of JIA, whereas hemoglobin levels were significantly lower. Osteocalcin levels were decreased and TNF-α increased in active JIA status, but these differences showed no statistical significance. We concluded that inflammatory cytokines play an important role in JIA. Monitoring IL-6 in children with JIA is useful in determining disease activity and response to therapy. These findings confirm earlier reports. Received: 21 June 2000 / Accepted: 10 September 2001     

Proinflammatory cytokines,IL-1β and TNF-α, induce expression of interleukin-34 mRNA via JNK- and p44/42 MAPK-NF-κB pathway but not p38 pathway in osteoblasts

The aim of this study is to investigate the induction of interleukin-34 (IL-34) and macrophage colony-stimulating factor (M-CSF) mRNA by inflammatory cytokines and the involvement of mitogen-activated protein kinases (MAPKs) in this signaling pathway in human osteoblasts as both IL-34 and M-CSF bind to the same receptor c-FMS. Among four inflammatory cytokines [(IL-1β, IL-6, IL-17, and tumor necrosis factor-α (TNF-α)], IL-34 mRNA expression level was dramatically induced by IL-1β (17-fold) and TNF-α (74-fold). IL-1β and TNF-α activated the intracellular mitogen-activated protein kinases (MAPKs): p44/42 MAPK, p38, and c-Jun N-terminal kinase (JNK) as well as nuclear factor-κB (NF-κB) in osteoblasts. IL-1β- and TNF-α-mediated induction of IL-34 mRNA expression was decreased by JNK inhibitor. Interestingly, although treatment of MEK-1/2 inhibitor showed no reduction in the increase of IL-34 mRNA expression by cytokines, combination of MEK-1/2 inhibitor and JNK inhibitor significantly inhibited IL-1β- and TNF-α-mediated IL-34 mRNA expression level compared to those by each inhibitor alone. On the other hand, M-CSF mRNA expression level was significantly induced by both IL-1β and TNF-α by up to 7- and 11-fold, respectively. IL-1β- and TNF-α-mediated induction of M-CSF mRNA was not affected by p38, JNK, and MEK-1/2 inhibitors. However, NF-κB inhibitor completely inhibited the elevation of M-CSF mRNA expression by these cytokines. These results showed that proinflammatory cytokines, IL-1β and TNF-α, induced the expression of IL-34 mRNA via JNK and p44/42 MAPK but not p38 in human osteoblasts while p38, JNK, and p44/42 MAPK were not involved in the induction of M-CSF mRNA expression by these cytokines.     

Circulating Levels of IL-1β, a Prothrombotic Cytokine,are Elevated in Unstable Angina Versus Stable Angina

Background: Multiple studies support a role for inflammation in the pathogenesis of coronary atherosclerosis and unstable cardiac syndromes. However, of the known proinflammatory cytokines, only elevated plasma levels of interleukin-6 have been linked to unstable angina. We sought to examine the plasma levels of other major proinflammatory cytokines in similar clinical settings and to determine the extent of the relationship between inflammation and unstable coronary syndromes by measuring the levels of various proinflammatory cytokines in patients with stable and unstable angina.Methods: We measured plasma levels of interleukin-1 beta (IL-1), tumor necrosis factor alpha (TNF-), and interleukin 6 (IL-6) in 97 patients: 67 with stable angina, 24 with unstable angina, and 15 healthy controls.Results: Mean levels of IL-1 were significantly higher in patients with unstable angina as compared to patients with stable angina (p = .009). Levels of IL-6 were significantly higher than control patients for both stable angina and unstable angina patients (p = .031 and .006, respectively). No significant differences were found in the levels of TNF-.Conclusions: Our results suggest that both IL-1 and IL-6 contribute to the pathogenesis of unstable angina, and that the profile of circulating plasma levels of proinflammatory cytokines differs in unstable angina from that in stable angina.Abbreviated Abstract. Multiple studies support a role for inflammation in the pathogenesis of coronary atherosclerosis and unstable cardiac syndromes. We measured plasma levels of interleukin-1 beta (IL-1), tumor necrosis factor alpha (TNF-), and interleukin 6 (IL-6) in patients with stable and unstable coronary syndromes. Levels of IL-1 and IL-6 were found to be elevated in patients with unstable coronary syndromes. No significant differences were found in the levels of TNF-. Our results suggest that both IL-1 and IL-6 contribute to the pathogenesis of unstable angina.     

Alterations of the mucosal immune system in inflammatory bowel disease

The normal intestinal immune system is under a balance in which proinflammatory and anti-inflammatory cells and molecules are carefully regulated to promote a normal host mucosal defense capability without destruction of intestinal tissue. Once this careful regulatory balance is disturbed, nonspecific stimulation and activation can lead to increased amounts of potent destructive immunologic and inflammatory molecules being produced and released. The concept of balance and regulation of normal mucosal immune and inflammatory events is indicative of how close the intestine is to developing severe inflammation. The normal intestinal mucosal immune system is constantly stimulated by lumenal contents and bacteria. The stimulatory molecules present in the intestinal lumen that activate and induce subsequent mucosal immunologic and inflammatory events include bacterial cell wall products, such as peptidoglycans and lipopolysaccharides, as well as other chemotactic and toxic bacterial products that are produced by the many different types of bacteria within the gastrointestinal tract. These highly stimulatory bacterial cell wall products are capable of activating macrophages and T lymphocytes to release potent proinflammatory cytokines, including interleukin-1 (IL-1), interleukin-6 (IL-6), and tumor necrosis factor α (TNF-α). IL-1, IL-6, and TNF-α increase the presence of human leukocyte antigen (HLA) class II antigen-presenting molecules on the surfaces of epithelial cells, endothelial cells, macrophages, and B cells, thus increasing their ability to present lumenal antigens and bacterial products. The proinflammatory cytokines IL-1 and TNF-α also increase the ability of epithelial cells, endothelial cells, macrophages, and fibroblasts to secrete potent chemotactic cytokines, such as interleukin-8 (IL-8) and monocyte chemoattractant protein-1 (MCP-1), which serve to increase the movement of macrophages and granulocytes from the circulation into the inflamed mucosa. Thus, through lumenal exposure to potent, nonspecific stimulatory bacterial products, the state of activation of the intestinal immune system and mucosal inflammatory pathways are markedly up-regulated. This raises the question of whether there is a deficiency in effective down-regulation through the absence of normally suppressive cytokines such as interleukin-10 (IL-10), transforming growth factor-β (TGF-β), interleukin-4 (IL-4), and IL-1 receptor antagonist. Normally, the turning off of the active and destructive immunologic and inflammatory events should occur following the resolution of a bacterial or viral infection that has been appropriately defended against and controlled by the mucosal immune system. In inflammatory bowel disease (IBD), however, the down-regulatory events and processes that should turn off the immunologic and inflammatory protective processes, once the pathogenic agent has been cleared, appear to be deficient or only partially effective. We may find that we ultimately are dealing with disease processes that have more than one genetic or cellular basis. The improved understanding of the immunopathophysiology of IBD will allow exploration of novel immunologic and genetic approaches, such as gene replacement therapy, administration of a suppressor cytokine or an altered cell surface antigen, the administration ofhumanized monoclonal antibodies directed against proinflammatory cytokines, or the development of newer strategies against fundamenial cell biologic mechanisms such as adhesion molecules.     

Effect of cilostazol, a selective type-III phosphodiesterase inhibitor, on water-immersion stress-induced gastric mucosal injury in rats

Background Cilostazol, a specific type-III phosphodiesterase inhibitor, is widely used for the treatment of ischemic symptoms of peripheral vascular disease. Recent studies have reported that the mechanism of cilostazol is related to the suppression of pro-inflammatory cytokine production and improvement of local microcirculation disturbances. The pathogenesis of stress-induced gastric mucosal lesions is characterized by the activation of inflammatory cells and the production of inflammatory cytokines. The effects of cilostazol on the development of gastric mucosal lesions have not been reported. In the present study, we examined the effect of a cilostazol on water-immersion stress-induced gastric mucosal lesions. Methods Rats were subjected to water-immersion stress with or without pretreatment with a single intraperitoneal injection of the selective type-III phosphodiesterase inhibitor, cilostazol. We measured the gastric mucosal lesion and the concentrations of myeloperoxidase (MPO), interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), and cytokine-induced neutrophil chemoattractant-1 (GRO/CINC-1), as an index of neutrophil accumulation and pro-inflammatory cytokine production. Results Cilostazol ameliorated the gastric mucosal injury induced by water-immersion stress (P < 0.001). The gastric contents of MPO, TNF-α, IL-1β, and CRO/CINC-1 were all increased after water-immersion stress and were reduced to almost normal levels by cilostazol. Conclusions In this study, we demonstrated that a selective type-III phosphodiesterase inhibitor, cilostazol, inhibited stress-induced gastric inflammation and damage via suppressing the production of pro-inflammatory cytokines. Cilostazol may be useful for preventing gastric mucosal lesions.     

Nitrite/Nitrate Oxide and Cytokines Changes in Patients with Surgical Stress

Nitric oxide (NO) produced by inducible nitric oxide synthase (iNOS) is implicated in immunolog-ical and inflammatory processes. Inflammatory cytokines and endotoxin induce a large amount of NO from various cells. Surgical stress produces cytokines, which leads to systemic inflammatory response syndrome (SIRS). Continuously high production of cytokines causes a variety of complications including pneumonia, intraabdominal abscess and sepsis. We investigated the relationships between the nitrite/nitrate (NO_x) concentration and the level of cytokines in 50 patients undergoing gastroenterological surgery. We measured the levels of cytokines, including interleukin-6 (IL-6), tumor necrosis factor- (TNF-) interleukin-1 receptor antagonist (IL- 1ra), and nitrite/nitrate (NO_x) concentration in the serum and exudative fluid from the thoracic or abdominal cavity of 50 patients undergoing gastroenterological surgery in order to make clear the relationship between cytokines and NO.NO_x levels in the serum of the group with complications were higher on days 3 and 7 than in the group with no complications (P < 0.05). In the complications group, IL-6 was more elevated, and the NO_x level was synchronously elevated. In conclusion, surgical stress caused inflammatory cytokinemia. NO_x was produced during surgical stress, and when complications occurred, more NO_x was produced.     

Effects of aging and menopause on serum interleukin-6 levels and peripheral blood mononuclear cell cytokine production in healthy nonobese women

Inappropriate interleukin-6 production is thought to play a role in the development of several age-related conditions including atherosclerosis. This study aimed to determine whether aging affects circulating interleukin-6 (IL-6) levels. Healthy, nonobese women (n = 208, 44.5 ± 0.70 years, 22.4 ± 0.17 kg/m²) were categorized into four age groups (22–31, 32–41, 42–51, and 52–63 years; cross-sectional study). Cytokine levels in serum and those produced from peripheral blood mononuclear cell (PBMC) were measured. The oldest group had the highest circulating levels of IL-6 and oxidized low-density lipoprotein (ox-LDL) and higher PBMC production of IL-6, tumor necrosis factor-α (TNF-α), and interleukin-1 alpha (IL-1β). Additionally, significant interactions between age and menopause were found for serum IL-6 (P = 0.024), and TNF-α (P = 0.011) and IL-1β (P < 0.001) produced from PBMCs. Serum IL-6 levels positively correlated with age, waist–hip ratio (WHR), systolic blood pressure, circulating levels of TNF-α, IL-1β, and ox-LDL, and urinary 8-epi-prostaglandin F₂α. Multiple stepwise regression models identified the following factors for contributing to serum IL-6 levels: serum IL-1β, menopause status, WHR, and serum TNF-α in mode I (R ² = 0.302); serum IL-1β, age, serum TNF-α, and WHR (β = 0.197; P = 0.006) in model II (R ² = 0.283). Sub-analysis was performed according to menopausal status. Serum IL-6 levels were positively associated with levels of IL-6, TNF-α, and IL-1β in PBMC supernatants (unstimulated) from postmenopausal women, whereas these were negatively associated in premenopausal women. In conclusion, circulating IL-6 levels may be interactively influenced by age and menopause. Additionally, estrogen deprivation after menopause may enhance PBMC cytokine production in postmenopausal women, resulting in increased IL-6 levels which are closely related to oxidative stress.     

Systemic cytokine response after emergency and elective surgery for colorectal carcinoma

Background  Systemic cytokines (SC) are accepted mediators of host immune response. It is debated if long-term survival is influenced by emergency presentation of colorectal cancer, and the role of immunitary response is still unknown. The aim of this prospective study was to compare the SC response after emergency resection with that after elective resections of colorectal carcinoma. Materials and methods  One hundred six consecutive subjects with colorectal cancer were submitted to emergency (complete bowel obstruction; EMS, n = 50) or elective resection (ELS, n = 56) of the tumour. Sera were collected before surgery and at appropriate time points afterward and assayed for interleukin-1beta (IL-1β), tumour necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and C-reactive protein (CRP). Five-year survival was analysed according to Kaplan–Meier test. The Cox proportional hazard model was used for the multivariate analysis. Results  Pre-operative levels of IL-1β, IL-6 and CRP were statistically higher in the EMS group. Levels of TNF-α were not elevated after surgery and there was no difference between the groups. Five-year survival was significantly lower in the EMS group (p < 0.05). Conclusions  Immunitary response, as reflected by SC, was better after elective resection than after emergency resection of colorectal carcinoma and this difference may have implication in the long-term survival.     

Monocyte chemoattractant protein-1 enhances and interleukin-10 suppresses the production of inflammatory cytokines in adult rat cardiomyocytes

Objective Chemokines control the migration of leukocytes to inflamed tissue, and in particular monocyte chemoattractant protein (MCP)-1 has been implicated in the pathogenesis of several cardiovascular disorders such as chronic heart failure (CHF) and myocarditis. We hypothesised that MCP-1 may directly contribute to an inflammatory response in the cardiomyocytes, and in the present study we examined in adult rat cardiomyocytes: (i) the effect of tumour necrosis factor (TNF)a on MCP-1 production, (ii) the effect of MCP-1 on production of other inflammatory cytokines, and (iii) if the anti-inflammatory cytokine interleukin (IL)-10 could suppress any TNFα-induced MCP-1 production. Methods We used enzyme immunoassays, RNase protection assays and slot blot analysis to measure protein and mRNA levels of various cytokines in adult rat cardiomyocyte cultures. Results (i) We found a ∼6.4-fold increase of the MCP-1 level accompanied by an increase in MCP-1 mRNA accumulation in cardiomyocyte cultures after TNFα stimulation. (ii) In contrast, TNFα had no effect on IL-10 and only a modest effect on IL-1β and IL-6 levels in these cells. (iii) Importantly, MCP-1 stimulated inflammatory response in cardiomyocytes by enhancing IL-1β and IL-6 levels in these cells as found at both the protein and mRNA level. (iv) Co-stimulation with IL-10 resulted in a ∼55 % reduction in TNFα-stimulated MCP-1 levels in cardiomyocyte culture supernatants. Conclusion The present study demonstrates for the first time that MCP-1 can directly affect cardiomyocytes, and we introduce MCP-1 as a potential enhancer and IL-10 as a potential suppresser of inflammatory responses within the myocardium. Received: 10 October 2000 / Returned for 1. revision: 2 November 2000 / 1. Revision received: 5 December 2000 / Returned for 2. revision: 2 January 2001 / 2. Revision received: 5 January 2001 / Accepted: 8 January 2001     

Differential in vitro effects of IL-4, IL-10, and IL-13 on proinflammatory cytokine production and fibroblast proliferation in rheumatoid synovium

The purpose of this study was to compare the potential of interleukin-4 (IL-4), IL-10, and IL-13 to interrupt two major inflammatory pathways in rheumatoid arthritis (RA), i.e., overexpression of proinflammatory cytokines and cytokine-mediated fibroblast growth. IL-4, IL-10, and IL-13 were all able to significantly inhibit the production of IL-1β, tumor necrosis factor-α (TNF-α), IL-6, and IL-8 by freshly isolated RA synovial tissue cells; IL-10 was most effective in terms of IL-1β and TNF-α reduction. The IL-1 receptor antagonist was enhanced by IL-4 and IL-13, but only slightly enhanced by IL-10. Spontaneous interferon-γ secretion was diminished by IL-4 and IL-10 but not by IL-13. Addition of anti-IL-10 neutralizing antibody to RA synovial tissue cells resulted in a substantial increase in IL-1β and TNF-α levels, whereas neither anti-IL-4 nor anti-IL-13 antibody had a significant effect. IL-1β-stimulated proliferation of RA synovial fibroblast cell lines was inhibited by IL-4 and IL-13, but not by IL-10; IL-4 was over tenfold more effective than IL-13. These results suggest that IL-4, IL-10, and IL-13 all have the therapeutic potential to regulate the disease activity mediated by proinflammatory cytokines in RA, but each cytokine may have different potencies. Received: 29 June 2000 / Accepted: 20 July 2000     

Modulation of IL-1β, IL-6, IL-8, TNF-α, and TGF-β secretions by alveolar macrophages under NO2 exposure

Activated alveolar macrophages (AMs) secrete interleukine (IL)1, IL-6, IL-8, tumor necrosis factor- (TNF-), and transforming growth factor- (TGF-), whose inflammatory and fibroblast-activating characteristics may play a role in the maintenance of pulmonary inflammatory processes and subsequent fibrosis. Human AMs were transferred to a gas cylinder and exposed to NO₂ in concentrations ranging from 0.1 to 0.5 ppm in synthetic air for 30 min at 37°C. AMs were fixed on a polycarbonate membrane and placed on culture medium. A culture was established, with the exposed AM (nonstimulated or stimulated with 1 g/ml lipopolysaccharide [LPS]), and the remaining cells were used to determine the cytokines. IL-1, IL-6, and IL-8 were quantified by commercial enzyme-linked immunosorbent assay kits (ELISA kits). TNF- was determined with a sandwich ELISA, using the biotin-streptavidin system. NO₂ exposure of nonstimulated AM did not result in changes in IL-1, IL-6, TNF-, and TGF- release, compared to the situation with control experiments. Exposure for 30 min to NO₂ induced a significant decrease of LPS-stimulated IL-1, IL-6, IL-8, and TNF- (p < .05). The release of TGF- was not significantly affected by NO₂ exposure. Cytotoxicity of AM was checked by trypan blue exclusion, with values ranging from 1.3 to 3.0%. NO₂ exposure of LPS-stimulated AM resulted in a functional impairment of AM after NO₂ exposure regarding IL-1, IL-6, IL-8, and TNF-. Neither the spontaneous nor the stimulated release of TGF- were influenced by NO₂.     

Down regulation of peroxisome proliferator-activated receptorγ expression by inflammatory cytokines and its reversal by thiazolidinediones

Aims/hypothesis. Previous studies show that inflammatory cytokines play a part in the development of insulin resistance. Thiazolidinediones were developed as insulin-sensitizing drugs and are ligands for the peroxisome proliferator-activated receptorγ (PPARγ). We hypothesized that the anti-diabetic mechanism of thiazolidinediones depends on the quantity of PPARγ in the insulin resistant state in which inflammatory cytokines play a part. Methods. We isolated rat PPARγ1 and γ2 cDNAs and examined effects of various cytokines and thiazolidinediones on PPARγ mRNA expression in rat mature adipocytes. Results. Various inflammatory cytokines, such as tumour necrosis factor-α (TNF-α), interleukin-1α (IL-1α), IL-1β, IL-6 and leukaemia inhibitory factor decreased PPARγ mRNA expression. In addition, hydrogen peroxide, lysophosphatidylcholine or phorbol 12-myristate 13-acetate also decreased the expression of PPARγ. The suppression of PPARγ mRNA expression caused by 10 nmol/l of TNF-α was reversed 60 % and 55 % by treatment with 10^–4 mol/l of troglitazone and 10^–4 mol/l of pioglitazone, respectively. The suppression of glucose transporter 4 mRNA expression caused by TNF-α was also reversed by thiazolidinediones. Associated with the change of PPARγ mRNA expression, troglitazone improved glucose uptake suppressed by TNF-α. Conclusion/interpretation. Our study suggests that inflammatory cytokines could be factors that regulate PPARγ expression for possible modulation of insulin resistance. In addition, we speculate that the regulation of PPARγ mRNA expression may contribute to the anti-diabetic mechanism of thiazolidinediones. [Diabetologia (1999) 42: 702–710] Received: 11 August 1998 and in final revised form: 12 February 1999     

Cytokine inhibitors: soluble tumor necrosis factor receptor 1 and interleukin-1 receptor antagonist in Behçet’s disease

Serum levels of proinflammatory cytokines, interleukin-1 beta (IL-1), tumor necrosis factor alpha, (TNF-), and their inhibitors, IL-1 receptor antagonist (IL-1ra) and soluble TNF receptor 1 (sTNFR1), were determined by enzyme-linked immunosorbent assay in 104 patients with Behçets disease (65 active, 39 inactive) and 40 healthy controls. The levels of IL-1 and IL-1ra were significantly higher in both active and inactive patients than in control subjects (P<0.01 and P< 0.01, respectively). The concentrations of TNF- and sTNFR1 were found to be higher in active patients than in controls (P< 0.01 and P< 0.001, respectively). There were no significant differences in the serum levels of these cytokines and their inhibitors between active and inactive patients. Significant increases in mean C-reactive protein level and erythrocyte sedimentation rate were found in patients with active vs inactive disease (P< 0.001 and P< 0.05, respectively). C-reactive protein values correlated with erythrocyte sedimentation rate but not with cytokines or their inhibitors. Our conclusion is that elevated serum TNF- and sTNFR1 seem to be important inflammatory mediators in Behçets disease. The statistically significant increase in these levels may arise from the severity of inflammation in the tissue or organ involved.     

Differential effect of IL-1β and TNF-α on the production of IL-6, IL-8 and PGE2 in fibroblast-like synoviocytes and THP-1 macrophages

Inflammation in the joint of rheumatoid arthritis is a complex immune reaction facilitated by various factors, such as cytokines, cells and hypoxia. Thus, we evaluated their relative capacity to produce proinflammatory mediators in response to IL-1β, TNF-α or IL-17 under hypoxia or normoxia in fibroblast-like synoviocytes (FLSs) and macrophages. The level of IL-6 expression was strongly increased in both FLSs and THP-1 macrophages in response to IL-1β and TNF-α, but the level by TNF-α was less than that by IL-1β. In contrast, the expression of IL-8 in both cell types was strongly stimulated by both IL-1β and TNF-α. In FLSs, PGE₂ production increased only in response to IL-1β; and no effect was observed in THP-1 cells and TNF-α-stimulated FLSs. In addition, the production by IL-17 was extremely low when compared with those induced by IL-1β or TNF-α in FLSs and THP-1 cells. Hypoxia (2% O₂) decreased IL-1β-stimulated production of PGE₂, even though it increased the expression of mRNA and protein of COX-2. These results suggest that IL-1β and TNF-α differentially regulate gene expression in FLSs and macrophages under hypoxia or normoxia.     

Hormonal regulation of pro-inflammatory and lipid peroxidation processes in liver of old ovariectomized female rats

There is now a large body of evidence suggesting that the decline in ovarian function with menopause is associated with spontaneous increases in pro-inflammatory cytokines. On the other hand, oxidative stress has been implicated in the pathogenesis of several alterations due to menopause, and can arise through the increased production of lipid peroxides (LPO) and/or a deficiency of antioxidant defense. The aim of the present study was to investigate the effect of aging and ovariectomy on various physiological parameters related to inflammation and oxidative stress in livers obtained from old female rats and the influence of chronic exogenous administration of estrogens, phytoestrogens and growth hormone on these. Thirty-six female Wistar rats of 22 months of age were used in the present study. Twelve of them remained intact, and the other 24 had been ovariectomized at 12 months of age. Intact animals were divided into two groups and treated for 10 weeks with GH or saline, and ovariectomized animals were divided into four groups and treated for the same time with GH, estrogens, phytoestrogens or saline. A group of 2 month old intact female rats was used as young control. Protein expression of iNOS, HO-1, IL-6, TNFα, and IL-1β were determined by Western blot analysis. The levels of NO _x , LPO, TNFα, IL-1β, IL-6 and IL-10 were determined in different fractions of the liver. Levels of LPO in the liver homogenates as well as iNOS protein expression and NO _x levels were increased in old rats as compared to young animals; this effect was more evident in ovariectomized animals. Pro-inflammatory cytokines TNF-α, IL-1β and IL-6 were significantly increased and anti-inflammatory IL-10 decreased during ageing and after ovariectomy. Aging also significantly increased expression of HO-1 protein and ovariectomized rats showed an additional increase. Hormonal administration to the ovariectomized groups decreased NO _x , LPO levels and pro-inflammatory cytokines as compared with untreated rats. Significant rise in IL-10 and reductions in the iNOS, IL-6, TNFα and IL-1β proteins expression were also found. Oxidative stress and inflammation induced during aging in the liver are more marked in castrated than in intact old females. Administration of the different hormonal replacement therapies was able to inhibit the induction of pro-inflammatory cytokines and iNOS, decreased the levels of oxidative stress markers and had therapeutic potential in the prevention of liver injury.     

Comparison of serum IL-1β, sIL-2R,IL-6, and TNF-α levels with disease activity parameters in ankylosing spondylitis

Ankylosing spondylitis (AS) is a chronic, inflammatory, rheumatological disease affecting primarily the sacroiliac joint and vertebral column, with an etiology that remains obscure. Cytokines are soluble proteins that have specific roles in inflammatory response, arranging the interaction between cells of the immune system both in natural and specific immune reactions. This study was planned to evaluate the relation between the level of cytokines and the clinical and laboratory findings of patients with AS compared to healthy subjects. In this study, we demonstrated increased serum levels of soluble interleukin-2 receptor (sIL-2R), Interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α) in patients with AS compared with healthy subjects. Only IL-1β levels were not increased in AS patients. We found a correlation between C-reactive protein and IL-6 levels and between erythrocyte sedimentation rate and sIL-2R, IL-6 and TNF-α levels. Only the sIL-2R level was correlated with Bath AS Metrology Index and Bath AS Functional Index. We suggest that sIL-2R, IL-6, and TNF-α may have a role in the pathogenesis of AS and that their serum levels can be used as disease activity parameters and tools for diagnosis.     

Comparison of anti-rheumatic effects of local RNAi-based therapy in collagen induced arthritis rats using various cytokine genes as molecular targets

RNA interference (RNAi) provides a powerful means of sequence-specific gene silencing. Several studies show that RNAi may provide promising strategies to treat human diseases by suppressing disease responsible genes in vivo. In locomotor diseases, the progression of collagen-induced arthritis (CIA) is suppressed by tumor necrosis factor-α (TNF-α)-specific small interfering RNA (siRNA) delivered into the joint. The aim of this study, is to compare the effects of intraarticularly administered siRNAs targeting TNF-α, interleukin-1β (IL-1β), interleukin-6 (IL-6) and receptor activator of NF-κB ligand (RANKL) on CIA in rats. We confirmed that the silencing effects of siRNA duplexes specific for rat TNF-α, IL-1β, IL-6 and RANKL in vitro. Each siRNA was also delivered into the knee joint of CIA rats by the in vivo electroporation method 7, 10, 13 and 16 days after immunization with collagen. Local delivery of TNF-α or IL-1β-specific siRNA ameliorated CIA in rats effectively at the gross morphological, radiographical and histological evaluations. Our results suggested that TNF-α and IL-1β were the cytokines to be targeted in the joint for the treatment of rheumatoid arthritis. The in vivo siRNA transfection method may be useful for selection of target molecules to be silenced for treatment of joint diseases.     

Serum levels of tumor necrosis factor alpha,interleukin-1 alpha and beta in healthy elderly subjects

To determine the age-related changes in monokine secretion, the serum concentrations of tumor necrosis factor alpha (TNF), interleukin-1 alpha (IL-1α) and interleukin-1 beta (IL-1β) were measured in 31 young healthy subjects (21–35 years old) and 19 healthy elderly subjects (65–83 years old). There were no differences between the two groups with respect to serum concentrations of these cytokines. We also measured in a subgroup of young (n=8–11) and elderly (n=8–10) subjects the in vitro secretion of TNF and IL-1β in response to adding to the medium lipopolysaccharide (LPS, 2 ng/ml), interferon gamma (50 U/ml) or advanced glycosylation endproduct of bovine serum albumin (250 ug/ml). Only LPS-stimulated IL-1β production was significantly reduced in elderly subjects (456.7 ± 229.9 vs 1118.8 ± 494.8 pg/ml). It is concluded that with the possible exception of LPS-stimulated IL-1β production, monokine secretion measured in this study is well preserved in elderly subjects.     

Regulation of expression of cytokines and growth factors in osteoarthritic cartilage explants

Osteoarthritis (OA) is a degenerative joint disease characterised by the breakdown of the extracellular matrix of chondrocytes in the affected joints. Cytokines and growth factors which are known to play a role in the synthesis and degradation of cartilage matrix have been shown to be upregulated in osteoarthritic cartilage. This upregulation resulted in two different phenotypes, overexpressing either TNF-α and IL-6 or IL-1β, TGFβ₁, IL-4 and IL-10. To investigate the hierarchy among growth factors and cytokines involved in cartilage metabolism, we analysed osteoarthritic cartilage explants for their responses to human recombinant (rh) cytokines and growth factors. The cytokine expression patterns of the explants before and after in vitro culture were compared by immunohistological staining of cartilage sections. We found a coordinate expression of TNF-α and IL-6 on the one hand, and of IL-1β, TGFβ₁, IL-4 and IL-10 on the other. Although TNF-α and IL-6 stimulated each other’s expression, they downregulated TGF β₁, IL-4 and IL-10 or IL-1β, TGF β₁ and IFNg, respectively. IL-1β upregulated the expression of TGF β₁, IL-4 and IL-10, and jointly these four cytokines and growth factors downregulated IL-6. Both of the expression patterns described for OA cartilage can be explained by these regulatory mechanisms. Interestingly, no cytokine efficiently downregulated TNF-α, and even though IL-1β is upregulated in one of the OA phenotypes, none of the growth factors and cytokines tested – except for IL-1β itself – seemed capable of mediating this upregulation. This unresponsiveness to cytokine stimulation might hint at a genetic cause for the elevated expression in the respective phenotypes. Received: 15 September 2000 / Accepted: 17 April 2001