OBJECTIVE

Interleukin 10 (IL-10) is an anti-inflammatory, immunomodulatory cytokine that regulates mucosal inflammation. This study evaluated the safety, tolerance, and efficacy of recombinant human IL-10 (rhuIL-10) for mild to moderately active Crohn's disease.

METHODS

We conducted a 24-week multicenter, prospective, randomized, double-blind, placebo-controlled, and sequential-escalating-dose study. Ninety-five patients with Crohn's Disease Activity Index of 200-350, not presently undergoing corticosteroid, mesalamine, or immunosuppressive therapy, were treated with subcutaneous rhuIL-10 (1, 5, 10, or 20 microg/kg) or placebo once daily for 28 consecutive days. Patients were followed up for 20 weeks after treatment. Evaluation of safety and tolerance was the first objective, and efficacy was the second objective.

RESULTS

Adverse effects were dose-related, mild-to-moderate in severity, and reversible. Asymptomatic and reversible anemia and thrombocytopenia were observed at higher doses. No withdrawal or delayed adverse effects were evident during 20 weeks of follow-up. At the end of treatment (day 29), intent-to-treat analysis showed that 23.5% (confidence interval [CI], 6.8%-49.9%) of patients receiving 5 micro/kg rhuIL-10 experienced clinical remission and endoscopic improvement; 0% (CI, 0%-14.8%) of patients in the placebo group did. Higher doses of recombinant human IL-10 were less effective than 5 microg/kg. No rhuIL-10 serum accumulation and no antibody against IL-10 were detected after 4 weeks.

CONCLUSIONS

Subcutaneous rhuIL-10 administered daily for 28 days to patients with mild to moderately active Crohn's disease is safe, well-tolerated, and shows clinical and endoscopic improvement.

There is growing recognition that HIV-1 infection leads to an activation of the immune system that includes perturbations of cytokine expression, redistribution of lymphocyte subpopulations, cell dysfunction, and cell death. Here, we explored the relationships between HIV-1 infection and immune activation in chronically HIV-1-infected human lymph nodes. In addition to CD4 T-cell depletion, we found increased effector T-cell frequencies associated with profound up-regulation of an activation marker CD38 in naive, central memory, and effector CD4(+) and CD8(+) T cells. Likewise, Fas death receptor (CD95) was more frequently detectable on T cells from HIV-1 nodes. Dendritic cell (DC) depletion was dramatic, with plasmacytoid DCs (PDCs) 40-fold and myeloid DCs (MDCs) <em>20</em>-fold less frequent in HIV(+) nodes than in control nodes. Cytokine dysregulation was evident, with <em>IL</em>-2 and <em>IL</em>-15 as much as 2 or 3 logs greater in infected nodes than in control nodes. Thus, activated effector cells are inappropriately attracted and/or retained in lymphoid tissue in chronic HIV-1 infection. High-level cytokine expression in turn activates and retains more cells at these sites, leading to lymphadenopathy and massive bystander activation that characterizes HIV-1 infection. Strategies targeting these activation pathways may lead to new therapies.

Freshly isolated, murine epidermal Langerhans cells (LC) are weak accessory cells for primary T cell-dependent immune responses, but increase their stimulatory capacity at least <em>20</em>-fold progressively over a 3-d culture with keratinocytes. We have studied the mediators of LC maturation. LC enriched from 12-h epidermal cultures by negative selection do not survive when cultured for 60 h in standard medium. LC survive and show increased stimulatory capacity for oxidative mitogenesis and the primary MLR when 30% keratinocyte-conditioned medium is included. Of the three cytokines that are known to be produced by keratinocytes, only granulocyte/macrophage CSF (GM-CSF) maintains viability and increases stimulatory capacity. <em>IL</em>-1 alone does not keep LC alive, but further enhances the stimulatory activity when combined with GM-CSF. <em>IL</em>-3 has no effect. The increase in LC stimulatory capacity is not due to increased Ia antigen expression, which does not change between 12 and 60 h. Function is not simply due to improved viability, as GM-CSF does not enhance the function of 12-h LC when added to the short-term oxidative mitogenesis assay. By generating LC with strong stimulating activity for resting T cells, GM-CSF and <em>IL</em>-1 may be critical in the sensitization phase of T cell-mediated immunity.

OBJECTIVE

To assess the clinical response to interleukin-1 (IL-1) blockade and in vitro IL-1beta and IL-18 secretion in patients with systemic-onset juvenile idiopathic arthritis (JIA).

METHODS

Twenty-two patients with systemic-onset JIA were treated with the IL-1 receptor antagonist (IL-1Ra) anakinra. Monocytes from 18 patients and 20 healthy donors were activated by different Toll-like receptor ligands. Intracellular and secreted IL-1beta and IL-18 were analyzed by Western blotting and enzyme-linked immunosorbent assay.

RESULTS

Ten patients with systemic-onset JIA exhibited a dramatic response to anakinra and were classified as complete responders. Eleven patients had an incomplete response or no response, and 1 patient could not be classified in terms of response. Compared with patients who had an incomplete response or no response, complete responders had a lower number of active joints (P = 0.02) and an increased absolute neutrophil count (P = 0.02). In vitro IL-1beta and IL-18 secretion in response to various stimuli was not increased and was independent of treatment efficacy. Likewise, secretion of IL-1Ra by monocytes from patients with systemic-onset JIA was not impaired. An overall low level of IL-1beta secretion upon exposure to exogenous ATP was observed, unrelated to treatment responsiveness or disease activity.

CONCLUSIONS

Two subsets of systemic-onset JIA can be identified according to patient response to IL-1 blockade. The 2 subsets appear to be characterized by some distinct clinical features. In vitro secretion of IL-1beta and IL-18 by monocytes from patients with systemic-onset JIA is not increased and is independent of both treatment outcome and disease activity.

In senescent cells, a DNA damage response drives not only irreversible loss of replicative capacity but also production and secretion of reactive oxygen species (ROS) and bioactive peptides including pro-inflammatory cytokines. This makes senescent cells a potential cause of tissue functional decline in aging. To our knowledge, we show here for the first time evidence suggesting that DNA damage induces a senescence-like state in mature postmitotic neurons in vivo. About 40-80% of Purkinje neurons and <em>20</em>-40% of cortical, hippocampal and peripheral neurons in the myenteric plexus from old C57Bl/6 mice showed severe DNA damage, activated p38MAPkinase, high ROS production and oxidative damage, interleukin <em>IL</em>-6 production, heterochromatinization and senescence-associated β-galactosidase activity. Frequencies of these senescence-like neurons increased with age. Short-term caloric restriction tended to decrease frequencies of positive cells. The phenotype was aggravated in brains of late-generation TERC-/- mice with dysfunctional telomeres. It was fully rescued by loss of p21(CDKN1A) function in late-generation TERC-/-CDKN1A-/- mice, indicating p21 as the necessary signal transducer between DNA damage response and senescence-like phenotype in neurons, as in senescing fibroblasts and other proliferation-competent cells. We conclude that a senescence-like phenotype is possibly not restricted to proliferation-competent cells. Rather, dysfunctional telomeres and/or accumulated DNA damage can induce a DNA damage response leading to a phenotype in postmitotic neurons that resembles cell senescence in multiple features. Senescence-like neurons might be a source of oxidative and inflammatory stress and a contributor to brain aging.

OBJECTIVE

Patients with serious traumatic injury and major burns and an animal model of burn injury were studied to determine the effect of injury on the production of cytokines typical of the T helper-2 lymphocyte phenotype as opposed to the T helper-1 phenotype and on the production of interleukin-12.

BACKGROUND

Perturbations of natural and adoptive immunity are related to the increased susceptibility to infection manifested by seriously injured and burn patients. Earlier work has shown that impaired adoptive immunity after injury is characterized by diminished production of interleukin-2 (IL-2), a product of Th lymphocytes. Exposure of naive Th cells to certain antigens and cytokines causes conversion to either the Th-1 or the Th-2 phenotype. Th-1 cells produce IL-2 and interferon-gamma (IFN-tau) and initiate cellular immunity. Th-2 cells secrete interleukin-4 (IL-4) and interleukin-10 (IL-10) and stimulate production of certain antibodies. Conversion to the Th-1 phenotype is facilitated by IL-12, and conversion to the Th-2 phenotype is promoted by IL-4. The authors believed that serious injury might cause conversion of Th cells to the Th-2 as opposed to the Th-1 phenotype rather than generalized Th suppression.

METHODS

The authors studied circulating peripheral blood mononuclear cells (PBMC) from 16 major burn and 8 trauma patients on 32 occasions early after injury and from 13 age- and sex-matched healthy individuals for cytokine production after phytohemagglutinin stimulation. Also studied was a mouse model of 20% burn injury known to mimic the immune abnormalities seen in humans with burns. Splenocytes from burn mice, 10 to 12 per group, were studied after activation by concanavalin A or by the bacterial antigen Staphylococcus aureus Cowan strain I for cytokine production and cytokine messenger RNA expression as determined by reverse transcriptase polymerase chain reaction. Burn mice were compared with sham-burn controls and attention was focused on day 10 after burn injury, a time when IL-2 production and resistance to infection are highly suppressed. Finally, burn and sham-burn animals, 20 per group, were treated in vivo with IL-12 (25 ng daily for 5 days) and observed for mortality after septic challenge (cecal ligation and puncture [CLP]) performed on day 10 after injury.

RESULTS

Peripheral blood mononuclear cells from burn and trauma patients produced less IFN-tau, the index cytokine of Th-1 cells, than PBMCs from healthy individuals 1 to 14 days after burn injury (SE = 77.6 +/- 16 pg/mL patients vs. 141.3 +/- 35 pg/mL controls, p < 0.05). However, production of IL-4, the index cytokine of Th-2 cells, by patient PBMCs was increased (51.0 +/- 13.0 pg/mL patients vs. 26.9 +/- 2.5 controls, p < 0.05). Splenocytes from mice 10 days after burn injury, when compared with sham-burn controls, showed diminished production of IL-2 (1.04 +/- 0.91 units/mL burns vs. 5.8 +/- 0.55 units/mL controls, p < 0.05) and IFN-tau (1.05 +/- 0.7 units/mL burns vs. 12.0 +/- 8.9 units/mL controls, p < 0.05). However, burn splenocytes produced more IL-4 (2492 +/- 157.0 pg/mL burns vs. 672.0 +/- 22.7 pg/mL controls, p < 0.01) and IL-10 (695.2 +/- 20.8 pg/mL burns vs. 567.0 +/- 16.7 pg/mL controls, p < 0.05). Splenocyte production of IL-12 was also reduced after burn (0.20 +/- 0.035 units/mL) as compared with sham burn (0.46 +/- 0.08 units/mL, p < 0.05). The reduction in IL-2, IFN-tau, and IL-12 production by burn splenocytes was reflected by a tenfold decrease in expression of their respective cytokine mRNAs. In vivo IL-12 treatment of burn animals decreased mortality from CLP on day 10 after injury from 85% to 15% (sham-burn mortality after CLP, 15%, p < 0.05) and increased splenocyte IFN-tau production to supranormal levels.

CONCLUSIONS

Serious injury induced diminished production of IL-1 2 and a shift to the Th-2 phenotype with increased production of IL-4 and IL-10, cytokines known to inhibit Th-1 function. The ability of exogenous IL-12 to restore Th-1 cytokine production and resistance to infection suggests a therapeutic role for IL-12 in the immune dysfunction seen after major injury.

OBJECTIVE

To update response duration and survival data for patients with metastatic renal cell carcinoma treated with high-dose interleukin (IL)-2.

METHODS

Two hundred fifty-five assessable patients were entered onto seven phase II clinical trials. Recombinant IL-2 600,000 or 720,000 IU/kg was administered by 15-minute intravenous infusion every 8 hours for up to 14 consecutive doses over 5 days as clinically tolerated with maximal support. A second, identical cycle of treatment was scheduled following 5 to 9 days of rest, and courses could be repeated every 6 to 12 weeks in stable or responding patients. All data were updated as of December 1998, with report forms completed by the clinical investigators. These data had last been updated as part of the Food and Drug Administration reporting requirements in 1996.

RESULTS

Objective responses previously have been reported in 37 of 255 patients (15%) with 17 complete responses (7%) and 20 partial responses (8%). These data remain unchanged from previous reports. Median response duration for all objective responders remains unchanged at 54 months, but the range now extends from 3 to>> 131 months. Median duration for all complete responses has not yet been reached, but was at least 80 months (range, 7->> 131 mo) at the time of this analysis. Median duration for all partial responses remains 20 months (range, 3->> 126 mo). Median survival time for all 255 patients remains 16.3 months, with 10% to 20% of patients estimated to be alive 5 to 10 years after treatment with high-dose IL-2.

CONCLUSIONS

With prolonged follow-up, treatment with high-dose recombinant IL-2 remains extremely effective for a subset of patients with metastatic renal cell carcinoma.

BACKGROUND

Hypereosinophilic syndrome (HES) is a heterogeneous group of rare disorders defined by persistent blood eosinophilia>> or =1.5 x 10(9)/L, absence of a secondary cause, and evidence of eosinophil-associated pathology. With the exception of a recent multicenter trial of mepolizumab (anti-IL-5 mAb), published therapeutic experience has been restricted to case reports and small case series.

OBJECTIVE

The purpose of the study was to collect and summarize baseline demographic, clinical, and laboratory characteristics in a large, diverse cohort of patients with HES and to review responses to treatment with conventional and novel therapies.

METHODS

Clinical and laboratory data from 188 patients with HES, seen between January 2001 and December 2006 at 11 institutions in the United States and Europe, were collected retrospectively by chart review.

RESULTS

Eighteen of 161 patients (11%) tested were Fip1-like 1-platelet-derived growth factor receptor alpha (FIP1L1-PDGFRA) mutation-positive, and 29 of 168 patients tested (17%) had a demonstrable aberrant or clonal T-cell population. Corticosteroid monotherapy induced complete or partial responses at 1 month in 85% (120/141) of patients with most remaining on maintenance doses (median, 10 mg prednisone equivalent daily for 2 months to 20 years). Hydroxyurea and IFN-alpha (used in 64 and 46 patients, respectively) were also effective, but their use was limited by toxicity. Imatinib (used in 68 patients) was more effective in patients with the FIP1L1-PDGFRA mutation (88%) than in those without (23%; P < .001).

CONCLUSIONS

This study, the largest clinical analysis of patients with HES to date, not only provides useful information for clinicians but also should stimulate prospective trials to optimize treatment of HES.

<em>IL</em>-10-related cytokines include <em>IL</em>-<em>20</em> and <em>IL</em>-22, which induce, respectively, keratinocyte proliferation and acute phase production by hepatocytes, as well as <em>IL</em>-19, melanoma differentiation-associated gene 7, and AK155, three cytokines for which no activity nor receptor complex has been described thus far. Here, we show that mda-7 and <em>IL</em>-19 bind to the previously described <em>IL</em>-<em>20</em>R complex, composed by cytokine receptor family 2-8/<em>IL</em>-<em>20</em>Ralpha and DIRS1/<em>IL</em>-<em>20</em>Rbeta (type I <em>IL</em>-<em>20</em>R). In addition, mda-7 and <em>IL</em>-<em>20</em>, but not <em>IL</em>-19, bind to another receptor complex, composed by <em>IL</em>-22R and DIRS1/<em>IL</em><em>20</em>Rbeta (type II <em>IL</em>-<em>20</em>R). In both cases, binding of the ligands results in STAT3 phosphorylation and activation of a minimal promoter including STAT-binding sites. Taken together, these results demonstrate that: 1) <em>IL</em>-<em>20</em> induces STAT activation through <em>IL</em>-<em>20</em>R complexes of two types; 2) mda-7 and <em>IL</em>-<em>20</em> redundantly signal through both complexes; and 3) <em>IL</em>-19 signals only through the type I <em>IL</em>-<em>20</em>R complex.

The role of adaptive immunity in obesity-associated adipose tissue (AT) inflammation and insulin resistance (IR) is controversial. We employed flow cytometry and quantitative PCR to assess T-cell recruitment and activation in epididymal AT (eAT) of C57BL/6 mice during 4-22 weeks of a high-fat diet (HFD (60% energy)). By week 6, eAT mass and stromal vascular cell (SVC) number increased threefold in mice fed HFD, coincident with onset of IR. We observed no increase in the proportion of CD3(+) SVCs or in gene expression of CD3, interferon-γ (IFN-γ), or regulated upon activation, normal T-cell expressed and secreted (RANTES) during the first 16 weeks of HFD. In contrast, CD11c(+) macrophages (MΦ) were enriched sixfold by week 8 (P < 0.01). SVC enrichment for T cells (predominantly CD4(+) and CD8(+)) and elevated IFN-γ and RANTES gene expression were detected by <em>20</em>-22 weeks of HFD (P < 0.01), coincident with the resolution of eAT remodeling. HFD-induced T-cell priming earlier in the obesity time course is suggested by (i) elevated (fivefold) interleukin-12 (<em>IL</em>-12)p40 gene expression in eAT by week 12 (P ≤ 0.01) and (ii) greater IFN-γ secretion from phorbol myristate acetate (PMA)/ionophore-stimulated eAT explants at week 6 (onefold, P = 0.08) and week 12 (fivefold, P < 0.001). In conclusion, T-cell enrichment and IFN-γ gene induction occur subsequent to AT macrophage (ATMΦ) recruitment, onset of IR and resolution of eAT remodeling. However, enhanced priming for IFN-γ production suggests the contribution of CD4(+) and/or CD8(+) effectors to cell-mediated immune responses promoting HFD-induced AT inflammation and IR.

Little is known of the function of the T cells in the inflammatory infiltrate in Helicobacter pylori associated gastritis. This study thus measured T cell in vivo activation by enumerating the frequency of interferon gamma (IFN gamma) and interleukin 4 (<em>IL</em> 4) secreting cells isolated from the gastric antral mucosa in patients with or without gastritis and in H pylori positive and negative gastritis. Fifty four samples were examined for cytokine secretion. Four antral biopsy specimens from each patient (n = 51) were taken during diagnostic endoscopy. One was used to estimate histological gastritis and the presence of H pylori, and three of the samples were used to isolate T cells by enzymatic digestion. IFN gamma and <em>IL</em> 4 secreting cells were enumerated by ELISPOT. Thirty four samples had gastritis and 79% of those were H pylori positive. None of the samples from non-inflamed mucosa had H pylori. The numbers of IFN gamma secreting cells per 10(5) T cells were higher in gastritis than in normal mucosa (145 v <em>20</em> IFN gamma spots, p < 0.01), and higher in H pylori negative than H pylori positive gastritis (371 v 110 IFN gamma spots, p < 0.05). The frequencies of <em>IL</em> 4 secreting cells did not differ between gastritis and non-inflamed mucosa. In conclusion, there is an increase in IFN gamma secreting cells but not in <em>IL</em> 4 secreting cells in H pylori positive and negative gastritis. It is not known if this TH1 type reaction has a pathogenetic or protective role.

Necrotizing enterocolitis (NEC) is a devastating neonatal intestinal inflammatory disease, occurring primarily in premature infants, causing significant morbidity and mortality. The pathogenesis of NEC is associated with an excessive inflammatory <em>IL</em>-8 response. In this study, we hypothesized that this excessive inflammatory response is related to an immature expression of innate immune response genes. To address this hypothesis, intestinal RNA expression analysis of innate immune response genes was performed after laser capture microdissection of resected ileal epithelium from fetuses, NEC patients and children and confirmed in ex vivo human intestinal xenografts. Changes in mRNA levels of toll-like receptors (TLR)-2 and -4, their signaling molecules and transcription factors (MyD88, TRAF-6 and NFκB1) and negative regulators (SIGIRR, IRAK-M, A-<em>20</em> and TOLLIP) and the effector <em>IL</em>-8 were characterized by qRT-PCR. The expression of TLR2, TLR4, MyD88, TRAF-6, NFκB1 and <em>IL</em>-8 mRNA was increased while SIGIRR, IRAK-M, A-<em>20</em> and TOLLIP mRNA were decreased in fetal vs. mature human enterocytes and further altered in NEC enterocytes. Similar changes in mRNA expression were observed in immature, but not mature, human intestinal xenografts. Confirmation of gene expression was also validated with selective protein measurements and with suggested evidence that immature TRL4 enterocyte surface expression was internalized in mature enterocytes. Cortisone, an intestinal maturation factor, treatment corrected the mRNA differences only in the immature intestinal xenograft. Using specific siRNA to attenuate expression of primary fetal enterocyte cultures, both TOLLIP and A-<em>20</em> were confirmed to be important when knocked down by exhibiting the same excessive inflammatory response seen in the NEC intestine. We conclude that the excessive inflammatory response of the immature intestine, a hallmark of NEC, is due to a developmental immaturity in innate immune response genes.

Systemic onset juvenile idiopathic arthritis (SoJIA) represents up to <em>20</em>% of juvenile idiopathic arthritis. We recently reported that interleukin (<em>IL</em>) 1 is an important mediator of this disease and that <em>IL</em>-1 blockade induces clinical remission. However, lack of specificity of the initial systemic manifestations leads to delays in diagnosis and initiation of therapy. To develop a specific diagnostic test, we analyzed leukocyte gene expression profiles of 44 pediatric SoJIA patients, 94 pediatric patients with acute viral and bacterial infections, 38 pediatric patients with systemic lupus erythematosus (SLE), 6 patients with PAPA syndrome, and 39 healthy children. Statistical group comparison and class prediction identified genes differentially expressed in SoJIA patients compared with healthy children. These genes, however, were also changed in patients with acute infections and SLE. An analysis of significance across all diagnostic groups identified 88 SoJIA-specific genes, 12 of which accurately classified an independent set of SoJIA patients with systemic disease. Transcripts that changed significantly in patients undergoing <em>IL</em>-1 blockade were also identified. Thus, leukocyte transcriptional signatures can be used to distinguish SoJIA from other febrile illnesses and to assess response to therapy. Availability of early diagnostic markers may allow prompt initiation of therapy and prevention of disabilities.

T-cell-based immunotherapies can be effective in the treatment of large vascularized tumors, but they rely on adoptive transfer of substantial numbers ( approximately <em>20</em> million) of tumor-specific T cells administered together with vaccination and high-dose interleukin (<em>IL</em>)-2. In this study, we report that approximately 10,000 T cells gene-engineered to express a single-chain <em>IL</em>-12 molecule can be therapeutically effective against established tumors in the absence of exogenous <em>IL</em>-2 and vaccine. Although <em>IL</em>-12-engineered cells did not perist long-term in hosts, they exhibited enhanced functionality and were detected in higher numbers intratumorally along with increased numbers of endogenous natural killer and CD8(+) T cells just before regression. Importantly, transferred T cells isolated from tumors stably overproduced supraphysiologic amounts of <em>IL</em>-12, and the therapeutic effect of <em>IL</em>-12 produced within the tumor microenvironment could not be mimicked with high doses of exogenously provided <em>IL</em>-12. Furthermore, antitumor effects could be recapitulated by engineering wild-type open-repertoire splenocytes to express both the single-chain <em>IL</em>-12 and a recombinant tumor-specific T-cell receptor (TCR), but only when individual cells expressed both the TCR and <em>IL</em>-12, indicating that arrested migration of T cells at the tumor site was required for their activities. Successful tumor eradication was dependent on a lymphodepleting preconditioning regimen that reduced the number of intratumoral CD4(+) Foxp3(+) T regulatory cells. Our findings reveal an approach to genetically modify T cells to reduce the cell number needed, eliminate the need for vaccines or systemic <em>IL</em>-2, and improve immunotherapy efficacy based on adoptive transfer of gene-engineered T cells.

<em>IL</em>-21, a newly described cytokine belonging to the <em>IL</em>-2 gamma-chain receptor cytokine family (that includes <em>IL</em>-2, <em>IL</em>-7, and <em>IL</em>-15), has been described as an important regulator of the cellular immune response. In this study, the role of <em>IL</em>-21 in the generation of a human Ag-specific CD8+ T cell response is characterized by tracking a rare, but measurable population of self-Ag-specific T cells in vitro. Autologous dendritic cells pulsed with the melanoma antigen recognized T cells 1 self-peptide were used to stimulate CD8+ T cells from HLA-A2+ healthy donors and melanoma patients. We demonstrate that exposure to <em>IL</em>-21 increased the total number of MART-1-specific CD8+ T cells that could be elicited by>><em>20</em>-fold and, at the clonal level, enriched for a population of high-affinity CD8+ T cells with a peptide dose requirement more than 1 log(10)-fold less than their untreated counterparts. Phenotypic analysis of T cells from <em>IL</em>-21-treated cultures revealed a unique population of CD45RO+ CD28(high) CD8+ T cells, a phenotype that was stable for at least 4 wk after <em>IL</em>-21 exposure. These CD28(high) CD8+ T cells produced <em>IL</em>-2 upon Ag stimulation and represent potential helper-independent CTLs. Our studies demonstrate a significant role for <em>IL</em>-21 in the primary Ag-specific human CTL response and support the use of <em>IL</em>-21 in the ex vivo generation of potent Ag-specific CTLs for adoptive therapy or as an adjuvant cytokine during in vivo immunization against tumor Ags.

As the applications of industrial nanoparticles are being developed, the concerns on the environmental health are increasing. Cytotoxicities of titanium dioxide nanoparticles of different concentrations (5, 10, <em>20</em> and 40 microg/ml) were evaluated in this study using a cultured human bronchial epithelial cell line, BEAS-2B. Exposure of the cultured cells to nanoparticles led to cell death, reactive oxygen species (ROS) increase, reduced glutathione (GSH) decrease, and the induction of oxidative stress-related genes such as heme oxygenase-1, thioredoxin reductase, glutathione-S-transferase, catalase, and a hypoxia inducible gene. The ROS increase by titanium dioxide nanoparticles triggered the activation of cytosolic caspase-3 and chromatin condensation, which means that titanium dioxide nanoparticles exert cytotoxicity by an apoptotic process. Furthermore, the expressions of inflammation-related genes such as interleukin-1 (<em>IL</em>-1), interleukin-6 (<em>IL</em>-6), interleukin-8 (<em>IL</em>-8), TNF-a, and C-X-C motif ligand 2 (CXCL2) were also elevated. The induction of <em>IL</em>-8 by titanium dioxide nanoparticles was inhibited by the pre-treatment with SB<em>20</em>3580 and PD98059, which means that the <em>IL</em>-8 was induced through p38 mitogen-activated protein kinase (MAPK) pathway and/or extracellular signal (ERK) pathway. Uptake of the nanoparticles into the cultured cells was observed and titanium dioxide nanoparticles seemed to penetrate into the cytoplasm and locate in the peri-region of the nucleus as aggregated particles, which may induce direct interactions between the particles and cellular molecules, to cause adverse biological responses.

BACKGROUND

An anti-inflammatory cytokine profile on whole-blood stimulation in vitro is associated with fatal outcome of meningococcal disease. We investigated whether an anti-inflammatory cytokine profile in the circulation is associated with adverse outcome in other infectious diseases.

METHODS

We enrolled 464 consecutive patients (272 men, 192 women) who presented to hospital with fever >> or = 38.2 degrees C). On admission we measured plasma interleukin 10 (IL-10) and tumour necrosis factor alpha (TNF alpha), and collected clinical and microbiological data on the febrile illness, then followed up all patients for clinical outcome.

RESULTS

In at least 399 of the 464 patients fever was caused by infection. 33 patients died after a median hospital stay of 11 days (interquartile range 3-20). Concentrations of IL-10 were significantly higher in non-survivors (median 169 pg/mL [IQR 83-530]) than in survivors (median 88 pg/mL [42-235], p=0.042). When dichotomised around the median, the mortality risk was two times higher in patients who had high concentrations of IL-10 than in those with low concentrations (relative risk 2.39 [95% CI 1.07-5.33]), in patients with low and high concentrations of TNF alpha. In the 406 patients without haemodynamic deterioration in the first 24 h, IL-10 was higher and TNF alpha lower in patients who died than in those who survived. The ratio of IL-10 to TNF alpha was higher in non-survivors (median 6.9 [3.0-21.0]) than in survivors (median 3.9 [2.0-7.0], p=0.040). This ratio was highest in patients who died without underlying disease (median 21.5 [5.0-25.0]). Age, sex, and duration of fever before admission did not explain the differences in IL-10 and TNF alpha.

CONCLUSIONS

An anti-inflammatory cytokine profile of a high ratio of IL-10 to TNF alpha is associated with fatal outcome in febrile patients with community-acquired infection. Our findings caution against a widespread use of proinflammatory cytokine inhibition in patients with sepsis.

Exogenously added <em>IL</em>-10 rapidly inhibited Staphylococcus aureus- or LPS-induced cytokine mRNA expression in human PBMCs and monocytes, with a maximal effect observed when <em>IL</em>-10 was added from <em>20</em> h before until 1 h after the addition of the inducers. Nuclear run-on assays revealed that the inhibition of <em>IL</em>-12 p40, <em>IL</em>-12 p35, and TNF-alpha was at the gene transcriptional level and that the addition of <em>IL</em>-10 to S. aureus- or LPS-treated PBMCs did not affect mRNA stability. The inhibitory activity of <em>IL</em>-10 was abrogated by cycloheximide (CHX), suggesting the involvement of a newly synthesized protein(s). The addition of CHX at 2 h before S. aureus or LPS also inhibited the accumulation of <em>IL</em>-12 p40 mRNA, but did not inhibit <em>IL</em>-12 p35 and TNF-alpha mRNA. This finding suggests that p40 transcription is regulated through a de novo synthesized protein factor(s), whereas the addition of CHX at 2 h after S. aureus activation caused superinduction of the <em>IL</em>-12 p40, <em>IL</em>-12 p35, and TNF-alpha genes. These results indicate that in human monocytes, the mechanism(s) of <em>IL</em>-10 suppression of both <em>IL</em>-12 p40 and <em>IL</em>-12 p35 genes is primarily seen at the transcriptional level, and that the induction of the <em>IL</em>-12 p40 and p35 genes have different requirements for de novo protein synthesis.

Synthetic oligodeoxynucleotides containing CpG dinucleotides (CpG-ODN) mimic the immunostimulatory qualities of bacterial DNA. We asked whether immunostimulation by CpG-ODN predisposes for a commitment toward a Th1 vs a Th2 response in Leishmania major infection, a model for a lethal Th2-driven disease, in BALB/c mice. CpG-ODN induced Th1 effector T cells in vitro and conveyed protective immunity to disease-prone BALB/c mice in vivo. Conversion to a Th1-driven resistant phenotype was associated with <em>IL</em>-12 production and maintained the expression of <em>IL</em>-12R beta2-chains. Most strikingly, CpG-ODN were even curative when given as late as <em>20</em> days after lethal L. major infection, indicating that CpG-ODN revert an established Th2 response. These findings imply an important role of bacterial DNA and CpG-ODN in the instruction of adaptive immune responses. They also point to the therapeutic potential of CpG-ODN in redirecting curative Th1 responses in Th2-driven disorders.

BACKGROUND

IL-17 signaling has been implicated in development and persistence of asthma. Cytokine-targeted strategies blocking IL-17 receptor signaling may be beneficial in asthma treatment.

OBJECTIVE

To determine efficacy and safety of brodalumab, a human anti-IL-17 receptor A monoclonal antibody, in subjects with inadequately controlled moderate to severe asthma taking regular inhaled corticosteroids.

METHODS

Three hundred two subjects were randomized to brodalumab (140, 210, or 280 mg) or placebo. Primary endpoint was change in Asthma Control Questionnaire (ACQ) score from baseline to Week 12. Secondary endpoints included FEV1, symptom scores, and symptom-free days. Prespecified subgroup analyses were conducted to identify potential responsive subpopulations. Analyses included randomized subjects receiving one or more doses of investigational product using last-observation-carried-forward imputation.

RESULTS

Demographics and baseline characteristics were generally balanced among groups (n = 302; n = 226 brodalumab). For the overall study population, no treatment differences were observed. Nine prespecified subgroups were examined without corrections for multiple testing. In only the high-reversibility subgroup (post-bronchodilator FEV1 improvement ≥ 20%; n = 112) was an ACQ change with nominal significance noted; ACQ responses were nominally significant in the 210-mg group (estimated treatment difference, 0.53) but not significant in the higher 280-mg group (estimated treatment difference, 0.38). Adverse events, generally balanced among groups, were most commonly asthma, upper respiratory tract infection, and injection site reaction.

CONCLUSIONS

Inhibition of IL-17 receptor A did not produce a treatment effect in subjects with asthma. The results of the high-reversibility subgroup analysis are of uncertain significance, requiring further study of brodalumab in this asthma subpopulation. Clinical trial registered with www.clinicaltrials.gov (NCT01199289).

B-1a cells are distinguished from conventional B cells (B2) by their developmental origin, their surface marker expression and their functions. They were originally identified as a B cell subset of fetal origin that expresses the pan-T cell surface glycoprotein, CD5. B-1a cells also differ from B2 by the expression levels of several surface markers, including IgM, IgD, CD43 and B2<em>20</em> [R. Berland, H.H. Wortis, Origins and functions of B-1 cells with notes on the role of CD5. Ann Rev Immunol, <em>20</em> (<em>20</em>02) 253-300.]. The majority of B-1a cells are located in peritoneal and pleural cavities. Compared to B2 cells, B-1a are long-lived, non-circulating, with reduced BCR diversity and affinity [A.B. Kantor, C.E. Merrill, L.A. Herzenberg, J.L. Hillson, An unbiased analysis of V-H-D-J(H) sequences from B-1a, B-1b, and conventional B cells. J Immunol, 158 (1997) 1175-1186.]. B-1a cells are largely responsible for the production of circulating IgM referred to as natural antibodies. These low affinity antibodies are polyreactive and constitute as such a first line of defense against bacterial pathogens [M.C. Carroll, A.P. Prodeus, Linkages of innate and adaptive immunity. Curr Opin Immunol, 10 (1998) 36-40.]. This polyreactivity also results into the recognition of autoantigens, which serves in the clearance of apoptosis products. The weak autoreactivity of the B-1a cells has been postulated to play a role in autoimmune pathogenesis. In addition, other characteristics, such as the production of high level of <em>IL</em>-10 [A. O'Garra, R. Chang, N. Go, R. Hastings, G. Haughton, M. Howard, et al. Ly-1 B (B-1) cells are the main source of B cell-derived interleukin 10. Eur J Immunol, 22 (1992) 711-717.] and enhanced antigen presentation capacities [C. Mohan, L. Morel, P. Yang, E.K. Wakeland, Accumulation of splenic B1a cells with potent antigen-presenting capability in NZM2410 lupus-prone mice. Arthritis and Rheumatism, 41 (1998) 1652-1662.], have implicated B-1a cells in autoimmunity. This review will discuss the current understandings of their role in autoimmune diseases with focus on lupus.

OBJECTIVE

Proinflammatory cytokines such as tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta have been implicated in the pathogenesis of myocardial dysfunction in ischemia-reperfusion injury, sepsis, chronic heart failure, viral myocarditis, and cardiac allograft rejection. Although circulating TNF-alpha and IL-1beta are both often elevated in septic shock, it remains unknown whether TNF-alpha or IL-1beta are the factors induced during sepsis that directly depress human myocardial function, and if so, whether the combination synergistically depresses myocardial function. Furthermore, the mechanism(s) by which these cytokines induce human myocardial depression remain unknown. We hypothesized the following: a) TNF-alpha and IL-1beta directly depress human myocardial function; b) together, TNF-alpha and IL-1beta act synergistically to depress human myocardial function; and c) inhibition of ceramidase or nitric oxide synthase attenuates myocardial depression induced by TNF-alpha or IL-1beta by limiting proximal cytokine signaling or production of myocardial nitric oxide (NO).

METHODS

Prospective, randomized, controlled study.

METHODS

Experimental laboratory in a university hospital.

METHODS

Freshly obtained human myocardial trabeculae.

METHODS

Human atrial trabeculae were obtained at the time of cardiac surgery, suspended in organ baths, and field simulated at 1 Hz, and the developed force was recorded. After a 90-min equilibration, TNF-alpha (1.25, 12.5, 125, or 250 pg/mL for 20 mins), IL-1beta (6.25, 12.5, 50, or 200 pg/mL for 20 mins), or TNF-alpha (1.25 pg/mL) plus IL-1beta (6.25 pg/mL) were added to the bath, and function was measured for the subsequent 100 mins after the 20-min exposure. To assess the roles of the sphingomyelin and NO pathways in TNF-alpha and IL-1beta cross-signaling, the ceramidase inhibitor N-oleoyl ethanolamine (1 microM) or the NO synthase inhibitor N(G)-monomethyl-L-arginine (10 microM) was added before TNF-alpha (125 pg/mL) or IL-1beta (50 pg/mL).

RESULTS

TNF-alpha and IL-1beta each depressed human myocardial function in a dose-dependent fashion (maximally depressing to 16.2 + 1.9% baseline developed force for TNF-alpha and 25.7 + 6.3% baseline developed force for IL-1beta), affecting systolic relatively more than diastolic performance (each p < .05). However, when combined, TNF-alpha and IL-1beta at concentrations that did not individually result in depression (p>> .05 vs. control) resulted in contractile depression (p < .05 vs. control). Inhibition of myocardial sphingosine or NO release abolished the myocardial depressive effects of either TNF-alpha or IL-1beta.

CONCLUSIONS

TNF-alpha and IL-1beta separately and synergistically depress human myocardial function. Sphingosine likely participates in the TNF-alpha and IL-1beta signal leading to human myocardial functional depression. Therapeutic strategies to reduce production or signaling of either TNF-alpha or IL-1beta may limit myocardial dysfunction in sepsis.

There are two distinct subtypes of multiple sclerosis in Asians, opticospinal (OS-multiple sclerosis) and conventional (C-multiple sclerosis). In OS-multiple sclerosis, selective and severe involvement of the optic nerves and spinal cord is characteristic, though its mechanisms are unknown. The present study aimed to find out possible differences in the cytokine/chemokine profiles in CSF between OS-multiple sclerosis and C-multiple sclerosis and to delineate the relationships between these profiles and neuroimaging and pathological features. Sixteen cytokines/chemokines, namely interleukin (<em>IL</em>)-1beta, <em>IL</em>-2, <em>IL</em>-4, <em>IL</em>-5, <em>IL</em>-6, <em>IL</em>-7, <em>IL</em>-8, <em>IL</em>-10, <em>IL</em>-12 (p70), <em>IL</em>-13, <em>IL</em>-17, interferon (IFN)-gamma, tumour necrosis factor (TNF)-alpha, granulocyte colony-stimulating factor (G-CSF), monocyte chemoattractant protein-1 (MCP-1) and macrophage inflammatory protein-1beta (MIP-1beta), were measured simultaneously in CSF supernatants from 40 patients with relapsing-remitting multiple sclerosis (<em>20</em> OS-multiple sclerosis and <em>20</em> C-multiple sclerosis) at relapse and 19 control patients with spinocerebellar degeneration (SCD), together with intracellular production of IFN-gamma and <em>IL</em>-4 in CSF CD4+ T cells. In CSF supernatants relative to controls, <em>IL</em>-17, MIP-1beta, <em>IL</em>-1beta and <em>IL</em>-13 were only significantly increased in OS-multiple sclerosis patients, while TNF-alpha was only significantly increased in C-multiple sclerosis patients, using a cut-off level of 1 pg/ml. <em>IL</em>-8 was significantly elevated in both OS-multiple sclerosis and C-multiple sclerosis patients. MCP-1 was significantly decreased in both OS-multiple sclerosis and C-multiple sclerosis patients, while <em>IL</em>-7 was only significantly decreased in C-multiple sclerosis patients. <em>IL</em>-17, <em>IL</em>-8 and <em>IL</em>-5 were significantly higher in OS-multiple sclerosis patients than in C-multiple sclerosis patients. The increases in <em>IL</em>-17 and <em>IL</em>-8 in OS-multiple sclerosis were still significant even after exclusion of the patients undergoing various immunomodulatory therapies. Assays of intracellular cytokine production revealed that both the IFN-gamma+<em>IL</em>-4- T-cell percentage and intracellular IFN-gamma/<em>IL</em>-4 ratio in CSF cells were significantly greater in C-multiple sclerosis patients than in controls. Contrarily, OS-multiple sclerosis patients showed not only a significantly greater percentage of IFN-gamma+<em>IL</em>-4- T cells than controls but also a significantly higher percentage of IFN-gamma-<em>IL</em>-4+ T cells than C-multiple sclerosis patients. Among the cytokines elevated in multiple sclerosis, only <em>IL</em>-8 showed a significant positive correlation with the Expanded Disability Status Scale of Kurtzke score. Both the length of the spinal cord lesions on MRI and the CSF/serum albumin ratio had a significant positive correlation with <em>IL</em>-8 and <em>IL</em>-17 in multiple sclerosis, in which the spinal cord lesions were significantly longer in OS-multiple sclerosis than in C-multiple sclerosis. Three of six spinal cord specimens from autopsied OS-multiple sclerosis cases demonstrated numerous myeloperoxidase-positive neutrophils infiltrating necrotic lesions. These findings strongly suggest that in OS-multiple sclerosis, in addition to the Th1 cell upregulation seen in C-multiple sclerosis, intrathecal activation of the <em>IL</em>-17/<em>IL</em>-8 axis inducing heavy neutrophil infiltration contributes to extensive spinal cord lesion formation.

BACKGROUND

There are no treatments currently available for peanut allergy. Sublingual immunotherapy (SLIT) is a novel approach to the treatment of peanut allergy.

OBJECTIVE

We sought to investigate the safety, clinical effectiveness, and immunologic changes with SLIT in children with peanut allergy.

METHODS

In this double-blind, placebo-controlled study subjects underwent 6 months of dose escalation and 6 months of maintenance dosing followed by a double-blind, placebo-controlled food challenge.

RESULTS

Eighteen children aged 1 to 11 years completed 12 months of dosing and the food challenge. Dosing side effects were primarily oropharyngeal and uncommonly required treatment. During the double-blind, placebo-controlled food challenge, the treatment group safely ingested <em>20</em> times more peanut protein than the placebo group (median, 1,710 vs 85 mg; P = .011). Mechanistic studies demonstrated a decrease in skin prick test wheal size (P = .0<em>20</em>) and decreased basophil responsiveness after stimulation with 10(-2) μg/mL (P = .009) and 10(-3) μg/mL (P = .009) of peanut. Peanut-specific IgE levels increased over the initial 4 months (P = .002) and then steadily decreased over the remaining 8 months (P = .003), whereas peanut-specific IgG4 levels increased during the 12 months (P = .014). Lastly, <em>IL</em>-5 levels decreased after 12 months (P = .015). No statistically significant changes were found in <em>IL</em>-13 levels, the percentage of regulatory T cells, or <em>IL</em>-10 and IFN-γ production.

CONCLUSIONS

Peanut SLIT is able to safely induce clinical desensitization in children with peanut allergy, with evidence of immunologic changes suggesting a significant change in the allergic response. Further study is required to determine whether continued peanut SLIT is able to induce long-term immune tolerance.