BACKGROUND

Salmonella Typhimurium (STm) remain a prominent cause of bacteremia in sub-Saharan Africa. Complement-fixing antibodies to STm develop by 2 years of age. We hypothesized that STm-specific CD4⁺ T cells develop alongside this process.

METHODS

Eighty healthy Malawian children aged 0-60 months were recruited. STm-specific CD4⁺ T cells producing interferon γ, tumor necrosis factor α, and interleukin 2 were quantified using intracellular cytokine staining. Antibodies to STm were measured by serum bactericidal activity (SBA) assay, and anti-STm immunoglobulin G antibodies by enzyme-linked immunosorbent assay.

RESULTS

Between 2006 and 2011, STm bacteremias were detected in 449 children <5 years old. STm-specific CD4⁺ T cells were acquired in infancy, peaked at 14 months, and then declined. STm-specific SBA was detectable in newborns, declined in the first 8 months, and then increased to a peak at age 35 months. Acquisition of SBA correlated with acquisition of anti-STm-lipopolysaccharide (LPS) immunoglobulin G (r = 0.329 [95% confidence interval, .552-.062]; P = .01) but not anti-STm-outer membrane protein or anti-STm-flagellar protein (FliC).

CONCLUSIONS

Acquisition of STm-specific CD4⁺ T cells in early childhood is consistent with early exposure to STm or cross-reactive protein antigens priming this T-cell development. STm-specific CD4⁺ T cells seem insufficient to protect against invasive nontyphoidal Salmonella disease, but sequential acquisition of SBA to STm LPS is associated with a decline in its incidence.

OBJECTIVE

We developed a nonmyeloablative conditioning regimen for allogeneic bone marrow transplantation (BMT) followed by donor lymphocyte infusions (DLI) for treatment of chemotherapy refractory malignancies. Although the majority of patients who receive this regimen achieve lasting mixed or full allogeneic chimerism, approximately 30% show initial mixed chimerism followed by loss of the donor graft. These patients recover host hematopoiesis without significant cytopenias. To assess the role of immunologic rejection in graft loss, we compared T-cell recovery and in vitro alloresponses in six patients who lost their marrow graft to that in 16 concurrent patients with sustained donor chimerism.

METHODS

Conditioning included pretransplant cyclophosphamide (150-200 mg/kg), thymic irradiation (700 cGy), and pre- and post-transplant equine antithymocyte globulin (ATG; ATGAM). HLA-identical related donor BMT was followed by DLI at approximately day <em>35</em> in patients without graft-vs-host disease.

RESULTS

The group with transient chimerism showed significantly increased circulating host T-cell (median 416 cells/mm(3) vs 10 cells/mm(3), p<0.05) and CD8 T-cell numbers (<em>35</em>4 cells/mm(3) vs 71 cells/mm(3), p<0.05) compared to the group with stable mixed or full donor chimerism within the first 100 days post-BMT. All DLI recipients who lost chimerism following DLI had greater than 80% recipient T cells at the time of DLI, whereas those with persistent chimerism had <60% host T cells. Graft rejection was associated with the development of a sensitized anti-donor bulk cytotoxic T-lymphocyte (CTL) response in 4 of 6 evaluated patients, compared to only 1 of 10 evaluated patients with sustained chimerism (p<0.05). Additionally, 3 of 5 evaluated transient chimeras showed high anti-donor CTL precursor frequencies in limiting dilution assays, and 3 of 4 evaluated transient chimeras showed high anti-donor interleukin-2 (IL-2)-producing T-helper (T(H)) cell frequencies. High anti-donor T(H) or cytotoxic T-lymphocyte precursors were not detected in sustained chimeras.

CONCLUSIONS

These data indicate that loss of chimerism in patients receiving this nonmyeloablative regimen is due to immune-mediated rejection. This rejection appears to bemediated by recovering recipient cytolytic CD8(+) cells as well as IL-2-producing recipient T(H) cells. These data are the first to demonstrate sensitization of recipient anti-donor IL-2-producing cells in association with human marrow allograft rejection.

OBJECTIVE

To investigate a novel anticytokine therapy in patients with sepsis syndrome, and the relationship between a patient's baseline mortality risk and survival benefit.

METHODS

Data from a recent phase III, double-blind, placebo-controlled, multicenter clinical trial with patients randomized to three treatment arms: an intravenous loading dose of recombinant human interleukin-1-receptor antagonist (rhIL-1ra) or placebo, followed by a continuous infusion of rhIL-1ra (1.0 mg/kg/hr, or 2.0 mg/kg/hr), or placebo for 72 hrs.

METHODS

Sixty-three investigative centers in eight countries.

METHODS

The study population consisted of 893 patients: 302 placebo patients; 298 patients treated with 1.0 mg/kg/hr of rhIL-1ra; and 293 patients treated with 2.0 mg/kg/hr of rhIL-1ra.

RESULTS

An independent, sepsis-specific, log-normal regression model that predicts the risk of mortality over 28 days was applied to all patients enrolled into the rhIL-1ra sepsis study. The ability of the Predicted Risk of Mortality model to predict 28-day mortality in the placebo patients was determined and the relationship between mortality risk and efficacy of rhIL-1ra was investigated. The trial data were also analyzed using two other risk-assessment models for comparison with Predicted Risk of Mortality. A significant increase in survival time was demonstrated for all patients treated with rhIL-1ra (n = 893, p < .02 Predicted Risk of Mortality log-normal), but patients with a Predicted Risk of Mortality of < 24% derived little benefit. Retrospective examination of time-to-death data demonstrated that rhIL-1ra reduced risk of death in the first 2 days for patients with>> or = 24% Predicted Risk of Mortality (n = 580, p < .005 Predicted Risk of Mortality log-normal). This same effect was not present in patients with a Predicted Risk of Mortality of < 24% on entry into the study. The Predicted Risk of Mortality model predicted a 28-day mortality rate of 35% for placebo patients compared with 34% observed and accurately stratified patients along the full range of risks. There was a wide distribution of individual patient risks for 28-day mortality for all patients, as well as within categorical subgroups, such as shock and organ system dysfunction. Two alternate risk models were assessed and the Acute Physiology Score of Acute Physiology and Chronic Health Evaluation III also demonstrated a statistically significant survival benefit for rhIL-1ra (p = .04 Predicted Risk of Mortality log-normal) for all patients treated.

CONCLUSIONS

Using an appropriate analytic model, a statistically significant increase in survival time from rhIL-1ra was measured. A direct relationship was found between a patient's Predicted Risk of Mortality at study entry to efficacy of rhIL-1ra. Individual risk or severity assessment may be a useful tool for evaluating the clinical benefit of new therapeutic approaches to sepsis and for monitoring outcomes at the bedside.

The objectives of this study were to quantify cytokine mRNA levels and endothelial cell adhesion molecule message and protein expression in healthy wild-type and <em>interleukin</em>-10-deficient (IL-10(-/-)) mice that develop spontaneous and chronic colitis. We found that colonic message levels of IL-1, IL-6, tumor necrosis factor-alpha, interferon-gamma, lymphotoxin-beta, and transforming growth factor-beta were elevated in colitic mice 10- to <em>35</em>-fold compared with their healthy wild-type controls. In addition, colonic message levels of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and mucosal addressin cell adhesion molecule-1 (MAdCAM-1) were found to be increased 10-, 5-, and 23-fold, respectively, in colitic IL-10(-/-) mice compared with their wild-type controls. Immunoradiolabeling as well as immunohistochemistry revealed large and significant increases in vascular surface expression of colonic ICAM-1, VCAM-1, and MAdCAM-1 in the mucosa as well as the submucosa of the colons of colitic mice. These data are consistent with the hypothesis that deletion of IL-10 results in the sustained production of proinflammatory cytokines, leading to the upregulation of adhesion molecules and infiltration of mononuclear and polymorphonuclear leukocytes into the cecal and colonic interstitium.

Leptin, a 16-kD protein of the ob gene product, is produced by adipose tissue and acts on the hypothalamus to suppress neuropeptide Y secretion which reduces the appetite. It has been demonstrated that serum leptin levels in healthy subjects are correlated with body mass index(BMI). Leptin is also produced by stimulation of inflammatory cytokines such as tumor necrosis factor(TNF)-alpha and <em>interleukin</em>(IL)-1. Rheumatoid arthritis(RA) is one of the chronic inflammatory diseases in which high serum cytokine levels are noted. In this study, we measured serum leptin levels(s-leptin) by RIA kit in 20 healthy subjects (10 males and 10 females) and 49 RA patients(14 males and <em>35</em> females) and the markers for joint inflammation such as ESR and CRP. There was no difference in s-leptin between controls(male: mean +/- SD = 5.6 +/- 3.0 ng/ml; female: 7.8 +/- 4.5 ng/ml) and RA patients(male: 4.9 +/- 3.2 ng/ml; female: 9.1 +/- 5.7 ng/ml). S-leptin was correlated with BMI in both healthy subjects and RA patients. However, there was no correlation between s-leptin and the values of ESR or CRP, disease stages in RA patients. In conclusion, s-leptin in RA patients reflects BMI but not joint inflammation.

BACKGROUND

Cytokines play a key role in the pathogenesis of inflammatory diseases. An obvious question is whether patients with aggressive periodontitis, juvenile idiopathic arthritis, or rheumatoid arthritis share blood cytokine profiles distinguishing them from individuals free of disease.

METHODS

The study population consisted of Danish white adults, (<em>35</em> years of age, diagnosed with localized aggressive periodontitis (LAgP; N = 18), generalized aggressive periodontitis (GAgP; N = 27), juvenile idiopathic arthritis (JIA; N = 10), or rheumatoid arthritis (RA; N = 23) and healthy individuals with no systemic or oral diseases (control [CTRL]; N = 25). Enzyme-linked immunosorbent assays were used to determine the levels of <em>interleukin</em> (IL)-1alpha, IL-1beta, IL-1 receptor antagonist (IL-1Ra), IL-6, IL-10, tumor necrosis factor (TNF)-alpha, and lymphotoxin (LT)-alpha in peripheral blood (plasma) and unstimulated and stimulated whole blood cell cultures from the same blood collection. Autoantibodies (aAb) to IL-1alpha and IL-6 were quantitated by radioimmunoassay.

RESULTS

Similar patterns of slightly higher IL-10 levels in plasma were found for GAgP and RA patients and in unstimulated cultures for GAgP, RA, and JIA patients. Interestingly, unstimulated cultures also demonstrated similar patterns of higher TNF-alpha levels for these three groups of patients. Similar group patterns of periodontitis patients (LAgP and GAgP) included increased IL-1Ra levels in stimulated cultures, which also showed similar group patterns of arthritis patients (JIA and RA) with respect to higher IL-1alpha and lower LT-alpha levels. Low titers of aAb to IL-1alpha and IL-6 were found in almost all individuals.

CONCLUSIONS

Patients with aggressive periodontitis and types of arthritis presented with similar components of blood cytokine profiles distinguishing them from individuals free of disease.

OBJECTIVE

To establish an in vitro model for the investigation of destructive processes in rheumatoid arthritis (RA), to study the interaction between fibroblasts, macrophages, and chondrocytes, and to evaluate strategies to inhibit joint destruction in RA.

METHODS

Human and bovine chondrocytes cultured in sponges pretreated with native bovine embryonic extracellular matrix produced a cartilaginous matrix reflected by the incorporation of <em>35S</em> into proteoglycans. The 3-dimensional culture system was optimized for the number of chondrocytes (10(5) cells/sponge), the timing of <em>35S</em> incorporation (day 21 after chondrocyte isolation), and the medium (20% fetal calf serum). RA synovial fibroblasts (RASF; 10(5)) were added, and the matrix destruction mediated by these RASF was monitored by the release of <em>35S</em>. The system was modulated by the addition of monocytes (U937 cells), cytokines (<em>interleukin</em>-1beta [IL-1beta] and tumor necrosis factor alpha [TNFalpha]), <em>interleukin</em>-1 receptor antagonist (IL-1Ra), and monoclonal antibodies against IL-1beta and CD44.

RESULTS

RASF destroyed the bovine cartilaginous matrix within 2 weeks (days 5-12) and the human cartilaginous matrix within 3 weeks (days 10-18). Compared with the effect of RASF alone (mean +/- SD 948 +/-180 counts per minute/week), the addition of U937 cells (a monocytic cell line), IL-1beta, or TNFalpha to the incubation medium increased the destruction of human cartilaginous matrix by at least 71% up to 90% (ranging from 1,618+/-204 cpm/week to 1,802+/-307 cpm/week). IL-1Ra and anti-IL-1beta monoclonal antibodies reduced the destruction of human matrix by 45% and 35%, respectively; this was partially reversed by the addition of U937 cells. The pretreatment of RASF with anti-CD44 monoclonal antibody (an adhesion molecule and receptor for hyaluronic acid) inhibited the destruction of the cartilaginous matrix by an average of 41% over 3 weeks.

CONCLUSIONS

This model is envisioned to study distinct aspects of human destructive joint diseases under in vitro conditions and to replace and/or supplement animal experiments in basic research and drug testing. Based on the fact that proinflammatory cytokines enhance destruction whereas IL-1Ra and antibodies against IL-1beta and CD44 inhibit the process, it is concluded that anti-IL-1- and anti-CD44-directed therapies may help prevent cartilage destruction in RA.

Metastasis to the liver often occurs in patients during the natural course of pancreatic cancer. Using carcinoma cell lines established from 9 such patients, we examined phenotypes of cell lines to search for correlations with their potential to metastasize to the liver. Anti-asialo GMI-treated nude mice were used. PCI-43, -55, -24 and -6, in this order, had frequent metastases, while PCI-10, -19, -<em>35</em>, -64, and -66 did not. In vitro doubling time, surface expression of sialyl Lewis(a) (SLe(a)), VLA-4/6, LFA-I/3, CEA, E-selectin, VCAM-I, NCAM, Mac-I, HLA-ABC/ DR/DQ, ICAM-I/2, production of <em>interleukin</em>-I alpha, tumor necrosis factor-alpha, and matrix metalloproteinase, as well as susceptibility to cytotoxicity by natural killer cells, were all examined. Expression of surface SLea was significantly associated with metastasis; numbers of metastatic colonies of SLe(a)-positive and -negative cell lines were 21.6 +/- 33.9 and 6.5 +/- 14.3 (p < 0.01), respectively. Moreover, the intensity of surface SLe(a) expression of each PCI line correlated with the number of metastatic colonies in the liver. When anti-SLe(a) monoclonal antibody (MAb) was administered, the development of liver metastasis by PCI-43 cells was significantly repressed, as compared with a control MAb. Although a reverse correlation between surface ICAM-I expression and liver metastasis was noted, the species-restricted function of ICAM-I makes interpretation difficult. Collective evidence indicates that expression of SLe(a) is an important positive mediator in the hematogenous metastasis of pancreas carcinoma.

Leukocytapheresis (LCAP) with a leukocyte removal filter column was administered for 45 patients with ulcerative colitis (UC). We evaluated changes in the leukocyte count and the differential percentages during LCAP. Cytokine production was assessed from each patient's peripheral mononuclear cells or monocytes. Flow cytometry was performed to assess the removal rates of activated cells and adhesion molecule positive cells by LCAP. Clinical improvement was recognized in <em>35</em> of 45 patients during intensive LCAP therapy, and it continued throughout maintenance therapy in 32 patients (71.1%). The leukocyte count was decreased to about 40% during the first 30 min, but it increased to approximately 170% at 20 min after the completion of LCAP. The concentration of tumor necrosis factor (TNF)alpha before LCAP in the effective group was higher than it was in either the ineffective group or the control group. Its level decreased to near normal range after LCAP. In the effective group, the concentrations of <em>interleukin</em> (IL)-1beta, IL-2, interferon (IFN)gamma, and IL-8 were near the normal upper limits before LCAP; however, they had decreased after LCAP. The concentration of IL-4 increased after LCAP. In the ineffective group, in contrast, the concentrations had been at or near normal before the initial LCAP treatment. Flow cytometry study revealed that LCAP could remove the activated cells and adhesion molecule positive cells more effectively. The clinical improvement and the changes observed before and after LCAP therapy suggest that LCAP is able to intervene in the causal mechanism(s) of UC.

OBJECTIVE

Recombinant human osteogenic protein 1 (OP-1) is an effective stimulator of human cartilage <em>35S</em>-proteoglycan synthesis. The present study was conducted to determine whether stimulation of human articular chondrocytes with OP-1 can help overcome <em>interleukin</em>-1beta (IL-1beta)-induced suppression of <em>35S</em>-proteoglycan synthesis.

METHODS

Human articular chondrocytes in alginate beads were maintained for 3 days in the absence (control) or presence of IL-1beta at 0.1-100 pg/ml with or without OP-1 at 50 ng/ml, in medium containing 10% fetal bovine serum (FBS). Incorporation of <em>35S</em>-sulfate into proteoglycans was quantified during the last 4 hours of culture and reported as counts per minute per microg DNA. Release of <em>interleukin</em>-1 receptor antagonist (IL-1Ra) and prostaglandin E2 into the medium was monitored by immunoassay.

RESULTS

IL-1beta at 10 pg/ml caused a 60% decrease in <em>35S</em>-proteoglycan synthesis. This could be blocked by including 500 ng/ml IL-1Ra in the medium. The presence of 50 ng/ml OP-1 in the IL-1beta-containing medium was effective in restoring <em>35S</em>-proteoglycan synthesis to the level of that found in cultures not treated with IL-1beta. The restorative effects of OP-1 and IL-1Ra were cumulative. The rate of release of prostaglandin E2 and IL-1Ra into the medium was not affected by the presence of OP-1.

CONCLUSIONS

Treatment of human articular chondrocytes with OP-1 cultured in the presence of FBS is effective in overcoming the down-regulation of proteoglycan synthesis induced by low doses of IL-1beta.

BACKGROUND

Adipocytokines are bioactive peptides that may play an important role in the regulation of glucose and lipid metabolism. In this study, we investigated the association of plasma adipocytokine concentrations with markers of triglyceride-rich lipoprotein (TRL) metabolism in men.

METHODS

Fasting adiponectin, leptin, resistin, <em>interleukin</em>-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), apolipoprotein (apo) B-48, apo C-III, and remnant-like particle (RLP)-cholesterol concentrations were measured by immunoassays and insulin resistance by homeostasis assessment (HOMA) score in 41 nondiabetic men with a body mass index of 22-<em>35</em> kg/m2. Visceral and subcutaneous adipose tissue masses (ATMs) were determined by magnetic resonance imaging and total ATM by bioelectrical impedance.

RESULTS

In univariate regression, plasma adiponectin and leptin concentrations were inversely and directly associated with plasma apoB-48, apoC-III, RLP-cholesterol, triglycerides, VLDL-apoB, and VLDL-triglycerides (P <0.05). Resistin, IL-6, and TNF-alpha were not significantly associated with any of these variables, except for a direct correction between apoC-III and IL-6 (P <0.05). In multivariate regression including HOMA, age, nonesterified fatty acids, and adipose tissue compartment, adiponectin was an independent predictor of plasma apoB-48 (beta coefficient = -0.<em>35</em>4; P = 0.048), apoC-III (beta coefficient = -0.406; P = 0.012), RLP-cholesterol (beta coefficient = -0.377; P = 0.016), and triglycerides (beta coefficient = -0.374; P = 0.013). By contrast, leptin was not an independent predictor of these TRL markers. Plasma apoB-48, apoC-III, RLP-cholesterol, and triglycerides were all significantly and positively associated with plasma insulin, HOMA, and visceral, subcutaneous, and total ATMs (P <0.05).

CONCLUSIONS

These data suggest that the plasma adiponectin concentration may not only link abdominal fat, insulin resistance, and dyslipidemia, but may also exert an independent role in regulating TRL metabolism.

Major controversy exists as to whether increased C-reactive protein (CRP) contributes to individual components of the metabolic syndrome or is just a secondary response to inflammatory disease processes. We measured blood pressure and metabolic phenotypes in spontaneously hypertensive rats (SHRs) in which we transgenically expressed human CRP in the liver under control of the apolipoprotein E promoter. In transgenic SHRs, serum levels of human CRP approximated the endogenous levels of CRP normally found in the rat. Systolic and diastolic blood pressures measured by telemetry were 10 to 15 mm Hg greater in transgenic SHRs expressing human CRP than in SHR controls (P<0.01). During oral glucose tolerance testing, transgenic SHRs exhibited hyperinsulinemia compared with controls (insulin area under the curve: 36±7 versus 8±2 nmol/L per 2 hours, respectively; P<0.05). Transgenic SHRs also exhibited resistance to insulin stimulated glycogenesis in skeletal muscle (174±18 versus 278±32 nmol of glucose per gram per 2 hours; P<0.05), hypertriglyceridemia (0.84±0.05 versus 0.64±0.03 mmol/L; P<0.05), reduced serum adiponectin (2.4±0.3 versus 4.3±0.6 mmol/L; P<0.05), and microalbuminuria (200±<em>35</em> versus 26±5 mg of albumin per gram of creatinine, respectively; P<0.001). Transgenic SHRs had evidence of inflammation and oxidative tissue damage with increased serum levels of <em>interleukin</em> 6 (36.4±5.2 versus 18±1.7 pg/mL; P<0.005) and increased hepatic and renal thiobarbituric acid reactive substances (1.2±0.09 versus 0.8±0.07 and 1.5±0.1 versus 1.1±0.05 nmol/L per milligram of protein, respectively; P<0.01), suggesting that oxidative stress may be mediating adverse effects of increased human CRP. These findings are consistent with the hypothesis that increased CRP is more than just a marker of inflammation and can directly promote multiple features of the metabolic syndrome.

The TH2-cytokines <em>interleukins</em>-4 and -13 severely alter gene expression of monocytic cells. We quantified the impact of <em>interleukins</em>-4 and -13 on the gene expression pattern of human peripheral blood monocytes applying a strategy that involved microarray hybridization, RT-PCR, immunohistochemistry and activity assays. After 3 days of continuous cytokine exposure the six most strongly upregulated gene products (15-lipoxygenase-1, fibronectin, monoamine oxidase-A, CD1c, CD23A, coagulation factor XIII) included four proteins with potential anti-inflammatory properties: (i) 15-lipoxygenase-1 (290-fold upregulation), (ii) fibronectin (180-fold upregulation), (iii) monoamine oxidase-A (56-fold upregulation) and (iv) coagulation factor XIII (<em>35</em>-fold upregulation). In addition, a number of other gene products, the expression of which is consistent with inflammatory resolution (annexin 1, collagen 1alpha2, laminin alpha5, TIMP3, heme oxygenase-1, CCL22, heat shock protein A8), were upregulated to a lower extent. In contrast, expression of classical pro-inflammatory gene products, such as tumor necrosis factor alpha, monocyte chemotactic protein-1, <em>interleukins</em>-1, -6, -8, -18, cyclooxygenase-2, as well as enzymes and receptors of the leukotriene cascade (5-lipoxygenase, 5-lipoxygenase activating protein, leukotriene B(4) receptor, cysteinyl leukotriene receptor 2) were significantly downregulated. These data suggest that medium-term treatment of human peripheral blood monocytes with <em>interleukins</em>-4/13 alters the gene expression pattern so that the cells might adopt a resolving phenotype.

BACKGROUND

The mechanisms of the detrimental effects of perioperative allogeneic blood transfusion are still unclear. Previous studies have suggested a higher incidence of adverse effects after the use of blood stored for prolonged time. Therefore, a possible time-dependent release of various white cell- and platelet-derived bioactive substances in stored human red cell suspensions was studied.

METHODS

Whole blood (6 units), plasma-reduced whole blood (6 units), and saline-adenine-glucose-mannitol blood (6 units) from 18 unpaid, normal blood donors were stored under standard blood bank conditions at 4 degrees C for <em>35</em> days. After refrigeration, samples were collected from all blood bags on Days 0, 2, 5, 9, 14, 21, 28, and <em>35</em> of storage. Extracellular concentrations of eosinophil cationic protein, eosinophil protein X, plasminogen activator inhibitor 1, myeloperoxidase, and <em>interleukin</em> 6 were analyzed by enzyme-linked immunosorbent assay and radioimmunoassay. The total intracellular and donor plasma levels of these substances also were analyzed at the time of blood donation.

RESULTS

Eosinophil cationic protein, eosinophil protein X, and myeloperoxidase increased 10- to 25-fold (p < 0.05) in a time-dependent manner in whole blood, plasma-reduced whole blood, and saline-adenine-glucose-mannitol blood during storage for <em>35</em> days. Plasminogen activator inhibitor 1 increased threefold to sixfold (p < 0.05) in whole blood and plasma-reduced whole blood, but not in saline-adenine-glucose-mannitol blood. Interleukin 6 was not detected in either plasma or samples obtained from the blood bags.

CONCLUSIONS

Stored whole blood, plasma-reduced whole blood, and saline-adenine-glucose-mannitol blood may release white cell- and platelet-derived bioactive substances in a time-dependent manner, which may be related to the detrimental effects of perioperative blood transfusions. Therefore, prestorage white cell reduction should be considered for further improvement of red cell suspensions.

The impact of human recombinant beta-<em>interleukin</em>-1 (IL-1) on adrenocortical stimulation was investigated. This study asked three questions: Does IL-1 increase the corticosterone levels of rat serum? Is there a direct effect on the adrenal cortex? What is the mechanism of this effect? The intraperitoneal injection of IL-1 (70 micrograms) in anesthetized male Fisher rats resulted in elevated corticosterone levels at 30 minutes and reached a maximum at 180 minutes (94 +/- 12 versus 34 +/- 4 micrograms/dl, p less than 0.01). Next, the adrenal glands from separate animals were perfused in situ. Corticosterone secretion was significantly increased (p less than 0.01) 90 minutes after a single arterial bolus of <em>35</em> micrograms of IL-1. The response to IL-1 was dose dependent, beginning at 3.5 micrograms and reaching a maximum at <em>35</em> micrograms. The addition of indomethacin (3 mumol/L) completely abolished the stimulatory effect of IL-1. This study demonstrates that IL-1 increases rat serum corticosterone levels, IL-1 directly stimulates the adrenal cortex, and the stimulation may be mediated through prostaglandin synthesis. This is the first evidence that IL-1 has a direct stimulatory effect on the adrenal cortex.

<em>Interleukin</em>-<em>35</em> (IL-<em>35</em>), an IL-12 cytokine family member, mediates the immune inhibitory function of regulatory T cells (Treg). We assayed the presence of IL-<em>35</em> in paraffin-embedded human pancreas cancer (PCAN) and unexpectedly found IL-<em>35</em> was expressed mainly by epithelial derived PCAN cells, but not by Treg. We further examined the expression and effect of exogenous IL-<em>35</em> in human PCAN cell lines and found IL-<em>35</em> promoted growth and inhibited apoptosis in PCAN cell lines. IL-<em>35</em> induced proliferation correlated with an increase in cyclin B, cyclin D, cdk2, and cdk4 and a decrease in p27 expression, while inhibition of apoptosis was associated with an increase in Bcl-2 and a decrease in TRAILR1. We conclude IL-<em>35</em> is produced by PCAN in vivo and promotes PCAN cell line growth in vitro. These results might indicate an important new role for IL-<em>35</em> as an autocrine growth factor in PCAN growth.

Exposure to ambient fine particulate matter (PM2.5) increases risks for cardiovascular disorders (CVD). However, the mechanisms and components responsible for the effects are poorly understood. Based on our previous murine exposure studies, a translational pilot study was conducted in female residents of Jinchang and Zhangye, China, to test the hypothesis that specific chemical component of PM2.5 is responsible for PM2.5 associated CVD. Daily ambient and personal exposures to PM2.5 and <em>35</em> elements were measured in the two cities. A total of 60 healthy nonsmoking adult women residents were recruited for measurements of inflammation biomarkers. In addition, circulating endothelial progenitor cells (CEPCs) were also measured in 20 subjects. The ambient levels of PM2.5 were comparable between Jinchang and Zhangye (47.4 and 54.5 µg/m(3), respectively). However, the levels of nickel, copper, arsenic, and selenium in Jinchang were 82, 26, 12, and 6 fold higher than Zhangye, respectively. The levels of C-reactive protein (3.44 ± 3.46 vs. 1.55 ± 1.13), <em>interleukin</em>-6 (1.65 ± 1.17 vs. 1.09 ± 0.60), and vascular endothelial growth factor (117.6 ± 217.0 vs. 22.7 ± 21.3) were significantly higher in Jinchang. Furthermore, all phenotypes of CEPCs were significantly lower in subjects recruited from Jinchang than those from Zhangye. These results suggest that specific metals may be important components responsible for PM2.5-induced cardiovascular effects and that the reduced capacity of endothelial repair may play a critical role.

OBJECTIVE

This study aims to investigate expression of <em>interleukin</em> 12 (IL-12) family cytokines IL-12, IL-23, IL-27 and IL-<em>35</em> in systemic lupus erythematosus (SLE) patients and the effect of glucocorticoid (GC) treatment on their expression.

METHODS

Plasma concentration of IL-12, IL-23, IL-27, IL-<em>35</em>, IL-6 and anti-double-stranded DNA (dsDNA) antibodies in 30 newly diagnosed severe SLE patients and 30 matched healthy subjects was measured by enzyme-linked immunosorbent assay. The correlation between the levels of IL-12 family cytokines and the levels of IL-6 or anti-dsDNA antibodies was analyzed by Spearman rank correlation.

RESULTS

Significantly higher levels of plasma IL-12, IL-23, IL-27, IL-<em>35</em>, IL-6 and anti-dsDNA antibodies were observed in SLE patients compared with healthy controls (p < 0.05), and after prednisone treatment, the serum levels of IL-12 family cytokines decreased significantly. Moreover, serum levels of IL-12, IL-23, IL-27 and IL-<em>35</em> were correlated with serum levels of IL-6 and anti-dsDNA antibodies in pre-treatment as well as post-treatment SLE patients.

CONCLUSIONS

SLE patients have increased plasma levels of IL-12 family cytokines and GCs can downregulate the expression of them in SLE patients. Therefore, members of the IL-12 family may be involved in the pathophysiological process of SLE.

BACKGROUND

Saposhnikovia divaricata (SD), called "Fangfeng" in China, is commonly used in clinical compound prescription for treatment of rheumatoid arthritis (RA), but its actions on RA have not been clarified. The present study aims to determine the anti-inflammatory activity of SD chromone extract (SCE), the major bioactive component of SD, on collagen-induced arthritis (CIA) rats, and elucidate its underlying mechanisms with regards to its molecular basis of action on human fibroblast-like synoviocytes derived from RA patients (HFLS-RA).

METHODS

CIA model on rats was constructed by injection of bovine type II collagen. Rats were pre-treated with different dosages of SCE from 3 days before till <em>35</em> days after model building. The progression of CIA was evaluated by macroscopic scoring, X-ray observation and hematoxylin and eosin (HE) staining of paws. HFLS-RA were pre-treated with different concentrations of SCE prior to stimulation with 10 ng/ml of tumor necrosis factor (TNF) α. By radioimmunoassay (RIA) and enzyme-linked immunosorbent assay (ELISA), levels of <em>interleukin</em> (IL)-1β, IL-6, TNFα and prostaglandin E2 (PGE2) were quantified respectively. Nuclear factor (NF-κB) p65 expression and DNA-binding activity were tested by immunohistochemisty and electrophoretic mobility shift assay (EMSA) respectively. Phosphorylation of extracellular signal-regulated kinase (ERK), Jun N-terminal kinase (JNK) and p38 MAPKs were examined by immunohistochemisty staining and western blot analysis.

RESULTS

Histological examination and radiological observation demonstrated that SCE significantly reduced the inflammatory responses in the joints of CIA rats. SCE inhibited the production of TNFα, IL-1β, and IL-6 in the joint tissues and sera. The level of PGE2 in sera was also decreased by SCE. Moreover, SCE treatment in vivo was able to reduce protein level of NF-κB, the transcriptional factor closely related to the inflammatory process, in articular synovium and cartilage of CIA rats. In addition, SCE inhibited p-ERK, p-JNK and p-p38 expression, which were considered to be involved in the phosphorylation of transcription factor NF-κB and the transcription of pro-inflammatory factors. Further, SCE inhibited NF-κB DNA binding activity and attenuated the phosphorylation of ERK, JNK and p38 MAPKs, in a concentration-dependent manner in cultured HFLS-RA.

CONCLUSIONS

These results highlight the anti-arthritic potential of SCE, and provide further evidence of the involvement of the NF-κB and MARKs inhibition in the effects of SCE.

Chondroitin sulfate E proteoglycan was extracted in the presence of protease inhibitors from 6 X 10(9) mouse bone marrow-derived, <em>interleukin</em> 3-dependent mast cells, of which 3 X 10(7) had been biosynthetically labeled with [<em>35S</em>]sulfate or [3H]glycine. Chondroitin sulfate E proteoglycan was purified to apparent homogeneity by density-gradient centrifugation, differential molecular weight dialysis, DEAE-52 ion exchange chromatography, and Sepharose CL-4B gel filtration chromatography. Chondroitin sulfate E proteoglycan, radiolabeled with [3H]glycine or [<em>35S</em>]sulfate, filtered as a single peak of radioactivity on Sepharose CL-4B with a Kav of 0.41. When purified [3H]glycine-labeled proteoglycan was digested with chondroitinase ABC and subjected to gel filtration, all of the radioactivity was shifted to a lower molecular weight. As assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis the Mr of the peptide core obtained by chondroitinase ABC treatment was approximately 10,000. The purified proteoglycan was resistant to degradation by collagenase, clostripain, trypsin, chymotrypsin, elastase, chymopapain, V8 protease, proteinase K, and Pronase, as assessed by gel filtration chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Analysis of the core peptide of the intact proteoglycan revealed that glycine, serine, and glutamic acid/glutamine accounted for 70% of the total amino acids and were present in a molar ratio of 4.3/1.6/1.0. When analyzed for neutral hexose content by gas-liquid chromatography, the proteoglycan contained approximately 2% of its weight as mannose, fucose, galactose, and other sugars, indicating that oligosaccharides were linked to the peptide core. The mouse bone marrow-derived mast cell chondroitin sulfate E proteoglycan, like the rat serosal mast cell heparin proteoglycan, is markedly protease resistant, has highly sulfated glycosaminoglycans, and contains a peptide core that is rich in serine and glycine. These characteristics of the mast cell class of intracellular proteoglycans may contribute to their function in stimulus-induced granule secretion as well as in mediator storage, including retention of cationic neutral proteases.

The role of <em>interleukin</em> (IL)-13, a Th2 cytokine sharing many of the features of IL-4, has not previously been examined in patients with visceral leishmaniasis (VL). We examined sera from Iranian patients with VL caused by Leishmania infantum. Serum IL-13 was detected in 50% (22/44) of patients with active primary disease. In comparison, IL-10 was detected in 79.5% (<em>35</em>/44), interferon gamma (IFN gamma) in 38.5% (17/44), and IL-4 in only 5% (2/44) of these patients. With few exceptions all 3 cytokines were undetectable after clinical recovery following antimony therapy. Five of 7 patients (71%) who failed antimony therapy and had relapsing disease had similar levels of IL-10 to patients with active primary disease. However, with only 1 exception, IL-13, IFN gamma and IL-4 were not detected in such patients. These data suggest that relapsing disease may result from defective cellular immunity, unrelated to immunosuppression mediated by IL-10.

Fangchinoline and tetrandrine are the major alkaloids from Stephania tetrandrae S. Moore which has been used traditionally for the treatment of inflammatory diseases in oriental countries including Korea. Both fangchinoline and tetrandrine showed anti-inflammatory effects on mouse ear edema induced by croton oil. In addition, the effects of fangchinoline and tetrandrine on cyclooxygenase, murine <em>interleukin</em>-5 (mIL-5) and human <em>interleukin</em>-6 (hIL-6) were examined in vitro to investigate the anti-inflammatory action mechanisms. One hundred micromolar of fangchinoline showed <em>35</em>% of inhibition on cyclooxygenase, but the same concentration of tetrandrine did not show any inhibition. On the other hand, 12.5 microM of tetrandrine exhibited 95% of inhibition on mIL-5 activity, while fangchinoline did not show any effects. However, 4 microM of fangchinoline and 6 microM of tetrandrine showed 63 and 86% of inhibitions on hIL-6 activity, respectively. These results suggest that biochemical mechanisms of fangchinoline and tetrandrine on anti-inflammation are significantly different even though they are similar in chemical structure.

OBJECTIVE

To search for genetic association between microsatellite marker loci and sibling pairs with nodal osteoarthritis (NOA).

METHODS

Using the affected sibling pair method of analysis, genomic DNA from 66 sib pairs with NOA was analysed for association with highly polymorphic microsatellite marker loci. The microsatellite markers were amplified using polymerase chain reaction and typed on polyacrylamide gels.

RESULTS

A significant association (p < 0.05) was identified between NOA and two loci on the short arm of chromosome 2 (2q 23-<em>35</em>). Candidate genes for osteoarthritis in this region include: fibronectin, a glycoprotein present in the extracellular matrix of normal cartilage; the alpha 2 chain of collagen type V, a major constituent of bone; and the <em>interleukin</em>-8 receptor, important in the regulation of neutrophil activation and chemotaxis.

CONCLUSIONS

The chromosomal region 2q 23-<em>35</em> requires further detailed study in NOA. Confirmation of these findings in large independent data sets and further analysis of candidate genes in this region will be important in unravelling the molecular basis for this common disease.

We investigated the pattern of down-regulation of cytokine production in endotoxin (lipopolysaccharide [LPS]) tolerance. A 4-day treatment with LPS (<em>35</em> micrograms per mouse) was followed by a challenge on day 6 with one more injection of LPS. Circulating tumor necrosis factor (TNF) and <em>interleukin</em>-6 (IL-6) could not be induced >> 99% inhibition) by LPS in LPS-tolerant mice; colony-stimulating factor (CSF) was also down-regulated by more than 95%, whereas interferon (IFN) and IL-1 syntheses were only partially inhibited. To study the mechanism of cytokine down-regulation in tolerance, we attempted to reverse the tolerant state by pretreatment with phorbol 12-myristate 13-acetate (PMA) (4 micrograms per mouse) 10 min before the LPS challenge. PMA completely restored IL-6 production and partially that of CSF. PMA had no effect on IFN production and inhibited the induction of IL-1. TNF production was also not restored by PMA. To investigate the role of endogenously produced cytokines in the development of LPS tolerance, we administered IL-6, TNF, or IL-1 alpha, using the same treatment schedule as that for LPS. Whereas IL-6 had no effect, IL-1 alpha or TNF induced partial tolerance to LPS in terms of inhibition of LPS-stimulated TNF and IL-6 production. However, a full LPS-tolerant state could not be induced by administration of recombinant cytokines, suggesting the existence of additional mechanisms, such as a loss of LPS receptors or changes in release of soluble binding proteins.