Melanoma differentiation associated gene-7 (mda-7) was cloned using subtraction hybridization from terminally differentiated human melanoma cells. Based on structural and functional properties, mda-7 is now recognized as interleukin-24 (<em>IL</em>-24), a new member of the expanding <em>IL</em>-10 gene family. Unique properties of mda-7/<em>IL</em>-24 include its ability to selectively induce growth suppression, apoptosis and radiosensitization in diverse human cancer cells, without causing similar effects in normal cells. The utility of mda-7/<em>IL</em>-24, administered by means of a replication-incompetent adenovirus, as a gene therapy for cancer has recently received validation in patients, highlighting an important phenomenon initially observed in pancreatic tumor cells, namely a 'potent bystander apoptosis-inducing effect' in adjacent tumor cells not initially receiving this gene product. We presently investigated the contribution of mda-7/<em>IL</em>-24 secreted by normal cells in mediating this 'bystander effect', and document that normal cells induced to produce mda-7/<em>IL</em>-24 following infection with recombinant adenoviruses expressing this cytokine secrete mda-7/<em>IL</em>-24, which modifies the anchorage-independent growth, invasiveness, survival and sensitivity to radiation of cancer cells that contain functional <em>IL</em>-<em>20</em>/<em>IL</em>-22 receptors, but not in cancer cells that lack a complete set of receptors. Moreover, the combination of secreted mda-7/<em>IL</em>-24 and radiation engenders a 'bystander antitumor effect' not only in inherently mda-7/<em>IL</em>-24 or radiation-sensitive cancer cells, but also in tumor cells overexpressing the antiapoptotic proteins bcl-2 or bcl-x(L) and displaying resistance to either treatment alone. The present studies provide definitive evidence that secreted mda-7/<em>IL</em>-24 from normal cells can induce direct antitumor and radiation-enhancing effects that are dependent on the presence of canonical receptors for this cytokine on tumor cells. Moreover, we now describe a novel means of enhancing mda-7/<em>IL</em>-24's therapeutic potential by targeting normal cells to produce and release this cancer-specific apoptosis-inducing cytokine, a strategy that could be employed as an innovative way of using this unique gene product for treating metastatic disease.

Refractory chronic graft-versus-host disease (cGVHD) is a significant complication resulting from allogeneic hematopoietic stem cell transplantation (HSCT). Mesenchymal stromal cells (MSCs) have shown promise for treating refractory cGVHD, but the favorable effects of MSCs therapy in cGVHD are complex and not fully understood. In this prospective clinical study, <em>20</em> of 23 cGVHD patients had a complete response or partial response in a 12-month follow-up study. The most marked improvements in cGVHD symptoms were observed in the skin, oral mucosa and liver. Clinical improvement was accompanied by a significantly increased number of interleukin (<em>IL</em>)-10-producing CD5+ B cells. Importantly, CD5+ B cells from cGVHD patients showed increased <em>IL</em>-10 expression after MSCs treatment, which was associated with reduced inflammatory cytokine production by T cells. Mechanistically, MSCs could promote the survival and proliferation of CD5+ regulatory B cells (Bregs), and indoleamine 2, 3-dioxygenase partially participates in the MSC-mediated effects on Breg cells. Thus, CD5+ Breg cells may have an important role in the process of MSC-induced amelioration of refractory cGVHD and may provide new clues to reveal novel mechanisms of action for MSCs.

BACKGROUND

In obese subjects, the adipose mass represents an important source of proinflammatory cytokines. We have identified a new syndrome-the normal-weight obese (NWO) syndrome-in women with normal weight and body mass index but whose fat mass is >30% of their total body weight and whose risk of developing obesity-related diseases is likely increased.

OBJECTIVE

The aim of the present study was to verify the hypothesis that NWO women are characterized by early inflammation, related to body fat mass, and that their plasma proinflammatory cytokine concentrations are greater than those of nonobese women.

METHODS

Twenty NWO, <em>20</em> preobese-obese, and <em>20</em> healthy (nonobese), age-matched white Italian women were studied. Anthropometric variables and plasma concentrations of proinflammatory cytokines and cardiovascular disease (CVD) risk factors were measured and compared between groups.

RESULTS

Plasma values and body-composition measures were significantly different between the preobese-obese and nonobese women. No significant differences in body weight, laboratory values, or CVD risk factors were found between the NWO and nonobese groups. Compared with concentrations in the NWO women, plasma concentrations of interleukin (IL)-1alpha, IL-1beta, IL-6, IL-8, and TNF-alpha were significantly lower in the nonobese group and were significantly greater in the preobese-obese group. IL-6 and TNF-alpha concentrations were related to fat mass distribution in the NWO women.

CONCLUSIONS

The proinflammatory cytokines could be regarded as significant prognostic indicators of the risk of obesity, CVD, and the metabolic syndrome in NWO women.

The capacity of non-pathogenic enteric bacteria to induce a pro-inflammatory response is under debate in terms of its effect on the symbiosis between the mammalian host and its commensal gut microflora. Activation of NF-kappaB and induction of interleukin-8 (<em>IL</em>-8) and CCL-<em>20</em> by the commensal Escherichia coli strain MG1655 were first studied in vitro in the human intestinal epithelial cell (IECs) lines HT29-19A and Caco-2, transfected or not with plasmids encoding dominant negative Toll-like receptor (TLR) 5 and myeloid differentiation factor-88 (MyD88) adaptor protein. The response of enterocytes in situ was then assessed using murine ileal biopsies mounted in Ussing chambers. Commensal E. coli induced NF-kappaB DNA binding, NF-kappaB transcriptional activity, CCL-<em>20</em> expression, and <em>IL</em>-8 secretion in the human IEC lines. E. coli MG1655 flagellin was necessary and sufficient to trigger this pro-inflammatory pathway via its interaction with TLR5 and the subsequent recruitment of the adaptor protein MyD88. Following epithelial cell polarization, signaling could be induced by live E. coli and flagellin on the apical side of HT29-19A. The in vivo relevance of our findings was confirmed, because immunohistochemical staining of murine ileum demonstrated expression of TLR5 in the apical part of enterocytes in situ. Furthermore, flagellin added on the mucosal side of murine ileal biopsies mounted in Ussing chambers induced a basolateral production of KC, a functional murine homolog of human <em>IL</em>-8. These findings provide strong evidence that flagellin released by flagellated commensal bacteria in the intestinal lumen can induce a pro-inflammatory response in enterocytes in vivo.

A severe burn leads to hypermetabolism and catabolism resulting in compromised function and structural changes of essential organs. The release of cytokines has been implicated in this hypermetabolic response. The severity of the hypermetabolic response following burn injury increases with age, as does the mortality rate. Due to the relationship between the hypermetabolic and inflammatory responses, we sought to compare the plasma cytokine profiles following a severe burn in adults and in children. We enrolled 25 adults and 24 children who survived a flame burn covering more than <em>20</em>% of total body surface area (TBSA). The concentrations of 22 cytokines were measured using the Linco multiplex array system (St. Charles, MO, USA). Large perturbations in the expression of pro- and anti-inflammatory cytokines were seen following thermal injury. During the first week following burn injury, IFN-gamma, <em>IL</em>-10, <em>IL</em>-17, <em>IL</em>-4, <em>IL</em>-6, and <em>IL</em>-8 were detected at significantly higher levels in adults compared with children, P < 0.05. Significant differences were measured during the second week post-burn for <em>IL</em>-1beta (higher in children) and <em>IL</em>-5 (higher in adults), P < 0.05. <em>IL</em>-18 was more abundant in children compared with adults during the third week post-burn, P < 0.05. Between post-burn d 21 and d 66, <em>IL</em>-1alpha was detected at higher concentrations in pediatric compared with adult patients, P < 0.05. Only GM-CSF expression was significantly different at all time points; it was detected at lower levels in pediatric patients, P < 0.05. Eotaxin, G-CSF, <em>IL</em>-13, <em>IL</em>-15, IP-10, MCP-1, and MIP-1alpha were detected at significantly different concentrations in adult compared with pediatric patients at multiple time points, P < 0.05. There were no differences in <em>IL</em>-12, <em>IL</em>-2, <em>IL</em>-7, or TNF levels in adult compared with pediatric burn patients at any of these time points. Following severe flame burns, the cytokine profiles in pediatric patients differ compared with those in adult patients, which may provide insight with respect to the higher morbidity rate in adults. Furthermore, the dramatic discrepancies observed in plasma cytokine detection between children and adults suggest that these two patient populations may benefit from different therapeutic interventions to achieve attenuation of the post-burn inflammatory response.

BACKGROUND

Activated platelets tether and activate myeloid leukocytes. To investigate the potential relevance of this mechanism in acute myocardial infarction (AMI), we examined cytokine induction by leukocyte-platelet adhesion and the occurrence of leukocyte-platelet conjugates in patients with AMI.

RESULTS

We obtained peripheral venous blood samples in <em>20</em> patients with AMI before and daily for 5 days after direct percutaneous transluminal coronary angioplasty (PTCA) and in <em>20</em> patients undergoing elective PTCA. Throughout the study period, CD41 immunofluorescence of leukocytes (flow cytometry) revealed increased leukocyte-platelet adhesion in patients with AMI compared with control patients (mean +/- SE of fluorescence [channels] before PTCA: 77 +/- 16 versus 35 +/- 9; P = .003). In vitro, thrombin-stimulated fixed platelets bound to neutrophils and monocytes. Within 2 hours, this resulted in increased mRNA for interleukin (<em>IL</em>),1 beta, <em>IL</em>-8, and monocyte chemoattractant protein (MCP)-1 in unfractionated leukocytes. After 4 hours, <em>IL</em>-1 beta and <em>IL</em>-8 concentration of the cell-free supernatant had increased by 268 +/- 36% and 210 +/- 7%, respectively, and cellular MCP-1 content had increased by 170 +/- 8%. Addition of activated platelets to adherent monocytes had a similar effect and was associated with nuclear factor-kappa B activation. Inhibition of binding by anti-P selectin antibodies reduced the effect of activated platelets on cytokine production.

CONCLUSIONS

In patients with AMI, leukocyte-platelet adhesion is increased. Binding of activated platelets induces IL-1 beta, IL-8, and MCP-1 in leukocytes. Our findings suggest that leukocyte-platelet adhesion contributes to the regulation of inflammatory responses in AMI.

Interleukin-8 (<em>IL</em>-8) is a chemokine produced by a variety of cell types involved in atherogenesis and is chemotactic for neutrophils and lymphocytes. A recent study has shown that <em>IL</em>-8 is angiogenic and induces proliferation and chemotaxis of endothelial cells. The present study was undertaken to find out whether <em>IL</em>-8 is also mitogenic and chemotactic for vascular smooth muscle cells. <em>IL</em>-8 induced a concentration-dependent (0.1 to 10 nmol/L) stimulation of DNA synthesis and cell proliferation in both human and rat aortic smooth muscle cells. In addition, <em>IL</em>-8 stimulated smooth muscle cells to produce prostaglandin E2, which can inhibit <em>IL</em>-8-induced smooth muscle cell proliferation. In the presence of indomethacin (5 mumol/L), <em>IL</em>-8 (1 nmol/L) stimulated an increase in human and rat aortic smooth muscle cell number during a 3-day period of incubation by 61 +/- 16% and 59 +/- 7% (n = 4), respectively. <em>IL</em>-8 also increased DNA synthesis in human and rat aortic smooth muscle cells by 98 +/- 10% and 151 +/- 27% (n = 5), respectively. Moreover, <em>IL</em>-8 stimulated rat aortic smooth muscle cell migration by <em>20</em>-fold over the control value, with an EC50 value of 0.83 nmol/L; this chemotactic activity of <em>IL</em>-8 was also potentiated by indomethacin. Exposure of smooth muscle cells to <em>IL</em>-8 caused rapid and transient expression of the immediate-early genes c-fos and zif268 mRNA.(ABSTRACT TRUNCATED AT 250 WORDS)

We show that in vitro activation of interphotoreceptor retinoid-binding protein (IRBP)-specific T cells from C57BL/6 mice immunized with an uveitogenic IRBP peptide (IRBP(1-<em>20</em>)) under TH17-polarizing conditions is associated with increased expansion of T cells expressing the gammadelta TCR. We also show that highly purified alphabeta or gammadelta T cells from C57BL/6 mice immunized with IRBP(1-<em>20</em>) produced only small amounts of <em>IL</em>-17 after exposure to the immunizing Ag in vitro, whereas a mixture of the same T cells produced greatly increased amounts of <em>IL</em>-17. IRBP-induced T cells from IRBP-immunized TCR-delta(-/-) mice on the C57BL/6 genetic background produced significantly lower amounts of <em>IL</em>-17 than did wild-type C57BL/6 mice and had significantly decreased experimental autoimmune uveitis-inducing ability. However, reconstitution of the TCR-delta(-/-) mice before immunization with a small number of gammadelta T cells from IRBP-immunized C57BL/6 mice restored the disease-inducing capability of their IRBP-specific T cells and greatly enhanced the generation of <em>IL</em>-17(+) T cells in the recipient mice. Our study suggests that gammadelta T cells are important in the generation and activation of <em>IL</em>-17-producing autoreactive T cells and play a major role in the pathogenesis of experimental autoimmune uveitis.

OBJECTIVE

Severe alcoholic hepatitis (AH) is associated with high mortality. Tumor necrosis factor-alpha (TNFalpha) has been demonstrated to play an important role in its pathophysiology.

METHODS

Twelve patients with biopsy-confirmed AH and a Maddrey discriminant factor >32 were treated with a single infusion of the anti-TNF monoclonal antibody Infliximab at a dose of 5mg/kg body weight. Serial measurements were made for various cytokines using specific enzyme-linked immunoassays (ELISA). In four patients, liver biopsy samples were available pretreatment and on day+28 of therapy.

RESULTS

Ten of the 12 patients are alive at a median of 15 (12-<em>20</em>) months. Two patients died within 30 days from septicemia. Serum bilirubin levels, Maddrey score, neutrophil count and C-reactive protein fell significantly within the first month. There was an early, though not significant, decrease in plasma levels of proinflammatory cytokines (interleukins (<em>IL</em>)-1beta, <em>IL</em>-6, <em>IL</em>-8, interferon-gamma), whereas plasma levels of TNFalpha remained near the sensitivity limit of the assay throughout the treatment course. While TNFalpha mRNA expression in the liver did not change, expression of <em>IL</em>-8, a cytokine regulated mainly by TNFalpha, was almost absent on day+28.

CONCLUSIONS

Our data suggest that randomized controlled trials of anti-TNF antibody in severe AH are warranted.

OBJECTIVE

The aims of this study were to determine whether IL-17(+) T cells were present in CD4 and CD8 interphotoreceptor retinoid-binding protein (IRBP)-specific T cells and to determine the role of antigen-specific and nonspecific IL-17(+) T cells in the pathogenesis of experimental autoimmune uveitis (EAU).

METHODS

B6 mice were immunized with uveitogenic peptide IRBP1-20. In vivo-primed T cells were separated and stimulated with the immunizing peptide. Intracellular expression of IFN-gamma and IL-17 by the T cells was assessed, and the pathogenic activity of the activated T cells was determined.

RESULTS

A subset of autoreactive IRBP-specific CD8 T cells expressed IL-17. IRBP-specific T cells preferentially expressed IL-17 when expanded by IL-23, whereas IFN-gamma-expressing cells were dominant when the T cells were cultured with IL-2. Importantly, both expanded T-cell populations were uveitogenic. In addition, IL-23 promoted the expansion of antigen-specific and non-antigen-specific IL-17(+) T cells, whereas TGF-beta and IL-6 acted only on non-antigen-specific IL-17(+) T cells. Only the antigen-specific IL-17(+) T cells were uveitogenic. The activation of autoreactive IL-17(+) T cells was markedly increased in vivo by the mycobacterial component of CFA and pertussis toxin (PTX) and in vitro by the ligation of Toll-like receptors.

CONCLUSIONS

IL-17(+) T cells can be readily detected among activated autoreactive and bystander T cells and may play a major role in the pathogenesis of EAU.

OBJECTIVE

Hepatocyte apoptosis, the hallmark of non-alcoholic steatohepatitis (NASH) contributes to liver injury and fibrosis. Although, both the intrinsic and extrinsic apoptotic pathways are involved in the pathogenesis of NASH, the final common step of apoptosis is executed by a family of cysteine-proteases termed caspases. Thus, our aim was to ascertain if administration of Emricasan, a pan-caspase inhibitor, ameliorates liver injury and fibrosis in a murine model of NASH.

METHODS

C57/BL6J-mice were fed regular chow or high fat diet (HFD) for <em>20</em> weeks. All mice were treated with vehicle or Emricasan.

RESULTS

Mice fed a HFD diet demonstrate a five-fold increase in hepatocyte apoptosis by the TUNEL assay and a 1.5-fold and 1.3-fold increase in caspase-3 and-8 activities respectively; this increase in apoptosis was substantially attenuated in mice fed a HFD treated with Emricasan (HFD-Em). Likewise, liver injury and inflammation were reduced in mice fed HFD-Em as compare to HFD by measuring serum aspartate aminotransferase and alanine aminotransferase levels, NAS histological score and IL 1-β, TNF-α, monocyte chemoattractant protein (MCP-1) and C-X-C chemokine ligand-2 (CXCL2) quantitative reverse-transcription polymerase chain reaction (qPCR). These differences could not be attributed to differences in hepatic steatosis as liver triglycerides content were similar in both HFD groups. Hepatic fibrosis was reduced by Emricasan in HFD animals by decreasing αSMA (a marker for hepatic stellate cell activation), fibrosis score, Sirius red staining, hydroxyproline liver content and profibrogenic cytokines by qPCR.

CONCLUSIONS

In conclusion, these data demonstrate that in a murine model of NASH, liver injury and fibrosis are suppressed by inhibiting hepatocytes apoptosis and suggests that Emricasan may be an attractive antifibrotic therapy in NASH.

To delineate the functional significance of <em>IL</em>-17 Receptor (<em>IL</em>-17RA) and characterize the <em>IL</em>-17 producing T cell (Th17) subpopulation in psoriatic arthritis (PsA). Mononuclear cells from blood and synovial fluid (SF) were obtained from PsA (n=<em>20</em>), rheumatoid arthritis (RA, n=<em>20</em>) and osteoarthritis (OA, n=<em>20</em>) patients. Synoviocytes (FLS) were isolated from the synovium of RA (n=5), PsA (n=5) and OA (n=5) patients. <em>IL</em>-17RA expression in FLS was identified by western blotting (WB) and flowcytometry. T lymphocytes derived from the SF of these patients were studied to identify and phenotype the Th17 cells. The functional significance of <em>IL</em>-17RA was determined by evaluating its regulatory role on the production of proinflammatory cytokines and endopeptidase. <em>IL</em>-17RA expression was found to be significantly higher in FLS of RA (15.7%±4.9) and PsA (4.5%±0.9) in comparison to OA (1.14%±0.9). Western blot analyses showed that the relative intensity (RI) of <em>IL</em>-17RA protein was higher in RA and PsA compared to OA (Fisher exact, P<0.01). A significant enrichment of <em>IL</em>-17-producing CD4+ T cells (7.9%±2.8) was observed in the SF of PsA patients compared to that of OA patients (P<.001). Compared to OA-FLS, recombinant <em>IL</em>-17 induced higher levels of <em>IL</em>-6, <em>IL</em>-8, and MMP-3 production in PsA-FLS. Blockage of <em>IL</em>-17RA with an anti-<em>IL</em>-17RA antibody inhibited the production of <em>IL</em>-6, <em>IL</em>-8, and MMP-3. This is the first report to demonstrate the functional significance of <em>IL</em>-17RA in PsA. Results of this study support the hypothesis that <em>IL</em>-17RA blocking antibodies have the potential to be a therapeutic option for psoriatic arthritis.

BACKGROUND

Atopic dermatitis (AD) is a chronic and relapsing inflammatory skin disease characterized by the predominant infiltration of TH2-type cells in lesional skin. Thymus and activation-regulated chemokine (TARC/CCL17) is a chemokine that attracts CC chemokine receptor 4-positive (CCR4+) or CCR8+ cells.

OBJECTIVE

The purpose of this study was to investigate the participation of TARC in AD.

METHODS

We measured serum TARC levels in 40 patients with AD, <em>20</em> healthy control subjects, and <em>20</em> patients with psoriasis. We also examined disease activity by using SCORAD score; serum soluble E-selectin, soluble <em>IL</em>-2 receptor, IgE, and GM-CSF levels; and eosinophil numbers in peripheral blood, as well as correlations between TARC levels and these factors. The positivity of CCR4 of CD4+CD45RO+ cells in PBMCs was examined by using FACS analysis. Immunohistochemical staining of TARC and GM-CSF was performed in the lesional skin of patients with AD.

RESULTS

The serum TARC levels of patients with AD were significantly higher than those of healthy control subjects and patients with psoriasis. The serum TARC levels significantly correlated with eosinophil number (r = 0.61), SCORAD score (r = 0.60), and serum soluble E-selectin levels (r = 0.58) and weakly correlated with serum soluble IL-2 receptor levels (r = 0.34) in patients with AD. The TARC levels of patients with AD decreased after the treatment in accordance with the improvement of clinical symptoms. The CCR4 positivity of CD4+CD45RO+ cells in PBMCs of patients with AD was also higher than that of healthy control subjects. Immunohistochemical staining revealed that TARC was positive in keratinocytes in the epidermis and in vascular endothelial cells, T cells, and dendritic cells in the dermis.

CONCLUSIONS

Serum TARC levels are associated with disease activity of AD, and TARC may play an important role in the pathogenesis of AD.

BACKGROUND

Severe pulmonary arterial hypertension (SPH) is a frequently lethal condition characterized by pulmonary vascular remodeling and right heart strain or failure. SPH is also often associated with autoimmune and collagen vascular disorders.

OBJECTIVE

To study the effects of T cells on the development of experimental SPH.

METHODS

Athymic nude rats lacking T cells were treated with a single subcutaneous injection of vascular endothelial growth factor (VEGF) receptor blocker SU5416 (<em>20</em> mg/kg) to induce pulmonary vascular endothelial cell apoptosis. Immunohistochemical analysis and <em>IL</em>-4 levels of the lung tissue were performed. Cell death and proliferation were assessed by Western blot and immunohistochemistry.

RESULTS

In contrast to SU5416-treated euthymic rats that develop SPH only in combination with chronic hypoxia, athymic nude rats developed SPH and vascular remodeling (similar to clinical SPH) at normoxic conditions as demonstrated by measurements of pulmonary artery pressure and right ventricle hypertrophy. Pulmonary arterioles became occluded with proliferating endothelial cells and were surrounded by mast cells, B cells, and macrophages. IL-4, proliferating cell nuclear antigen, and collagen type I levels were markedly increased in SU5416-treated athymic rat lungs. Antibody deposition was noted along the vascular endothelium in rats with SPH. Finally, protection from SPH was conferred by immune challenge with spleen cells from euthymic nude rats.

CONCLUSIONS

These studies demonstrate the importance of a complete, intact immune system in protecting against pulmonary angioproliferation in this new model of SPH as well as the importance of intact VEGF receptor signaling for lung endothelial cell homeostasis.

OBJECTIVE

To evaluate ixekizumab, an anti-interleukin-17A (anti-IL-17A) monoclonal antibody, in 2 populations of rheumatoid arthritis (RA) patients: biologics-naive patients and patients with an inadequate response to tumor necrosis factor (TNF) inhibitors.

METHODS

In this phase II, randomized, double-blind study, placebo or ixekizumab was administered subcutaneously to 260 biologics-naive patients and 188 patients with an inadequate response to TNF inhibitors at weeks 0, 1, 2, 4, 6, 8, and 10 with concomitant disease-modifying antirheumatic drugs. The primary objective was to determine the dose-response relationship of ixekizumab as measured by the proportion of biologics-naive patients meeting the American College of Rheumatology 20% improvement criteria (ACR20) at week 12.

RESULTS

Using a logistic regression model defined a priori, a statistically significant dose-response relationship as measured by ACR20 response rates at week 12 was detected in biologics-naive patients (P = 0.031). For patients with an inadequate response to TNF inhibitors, ACR20 responses at week 12 were significantly better with ixekizumab than placebo (P < 0.05). Decreases in the Disease Activity Score in 28 joints using the C-reactive protein level (DAS28-CRP), Clinical Disease Activity Index (CDAI), and CRP level from baseline were observed at week 12 in the ixekizumab groups in both populations (P < 0.05 versus placebo). Onset of action was rapid in some dose groups in both populations, with improvements in the ACR20, DAS28-CRP, CRP levels, and CDAI observed by day 3 (P < 0.05). Adverse events occurred with similar frequencies overall in the ixekizumab and placebo groups. Infections were more frequent with ixekizumab than placebo (biologics-naive 25% versus 19%; inadequate responders to TNF inhibitors 27% versus 25%). No mycobacterial or invasive fungal infections were reported.

CONCLUSIONS

Ixekizumab improved RA signs and symptoms in RA patients who were either naive to biologics treatment or had an inadequate response to TNF inhibitors. The safety profile was similar to that of other biologic agents, with no unexpected safety concerns.

Proinflammatory cytokines have been shown to influence the expression and function of the glucocorticoid receptor (GR). Specifically, several studies have found that cytokines induce a decrease in GR function, as evidenced by reduced sensitivity to glucocorticoid effects on functional end points. To investigate the potential mechanism(s) involved, we examined the impact of the proinflammatory cytokine, interleukin-1alpha (<em>IL</em>-1alpha), on 1) GR translocation from cytoplasm to nucleus using GR immunostaining, 2) cytosolic radioligand GR binding, and 3) GR-mediated gene transcription in L929 cells stably transfected with the mouse mammary tumor virus-cholamphenicol acetyltransferase reporter gene. L929 cells were treated with <em>IL</em>-1alpha (100 and 1000 U/ml) for 24 h in the presence or absence of dexamethasone (Dex; 10 nM to 1 microM). <em>IL</em>-1alpha inhibited Dex-induced GR translocation and alone induced GR up-regulation. Pretreatment with <em>IL</em>-1alpha followed by Dex treatment for 1.5 h led to about <em>20</em>% inhibition of Dex-induced GR-mediated gene transcription, whereas coincubation of <em>IL</em>-1alpha plus Dex for 24 h inhibited Dex-induced GR-mediated gene activity up to 42%. The latter effect was reversed by the <em>IL</em>-1 receptor antagonist. These results suggest that cytokines produced during an inflammatory response may induce GR resistance in relevant cell types by direct effects on the GR, thereby providing an additional pathway by which the immune system can influence the hypothalamic-pituitary-adrenal axis.

BACKGROUND

Atherosclerosis is an inflammatory disease in which monocytes and macrophages have been suggested to play an essential role. The underlying signaling mechanisms are unknown thus far. We hypothesized that the human isoform of Toll-like receptor (hTLR)-4 is involved in monocyte activation of patients with accelerated forms of atherosclerosis.

RESULTS

Expression of hTLR4 on circulating monocytes from 30 controls, <em>20</em> patients with stable angina (SA), 40 patients with unstable angina (UA), and 28 patients with acute myocardial infarction (AMI) was compared with the use of flow-cytometry and reverse transcription-polymerase chain reaction. Regulation of interleukin (<em>IL</em>)-12 and B7-1 as downstream events of TLR4 activation was analyzed after lipopolysaccharide stimulation of monocytes. TLR4-transfected Chinese hamster ovary (CHO) cells were used to identify potential hTLR4 ligands in the serum of patients with UA or AMI. Circulating hTLR4+/CD14+ monocytes were approximately 2.5-fold increased above controls and patients with SA in the UA and AMI groups (P<0.0001). This was paralleled by enhanced transcript levels of TLR4 and Myd88 in patients with UA and AMI (P<0.0001) and increased expression of <em>IL</em>-12 (UA 35.5+/-7.8, AMI 31.8+/-7.7 versus SA 2.2+/-0.5, controls 2.1+/-0.3 pg/mL; P<0.0002) and B7-1 (UA 27.3+/-14.4, AMI 22.6+/-11.1 versus SA 3.4+/-2.5, controls 2.4+/-2.3%; P<0.0001). Compared with serum from patients with UA and AMI, challenging TLR4-transfected CHO cells with serum from SA patients yielded only a weak response (P<0.0001). Coincubation with anti-heat shock protein 60 inhibited CHO cell activation.

CONCLUSIONS

UA and AMI are associated with enhanced expression and signaling events downstream of hTLR4 in circulating monocytes. These observations suggest hTLR4 activation as a signaling mechanism in immune-mediated progression of atherosclerosis.

The t(4;14)(p16.3;q32), associated with 10-<em>20</em>% of cases of multiple myeloma (MM), deregulates the expression of MMSET and FGFR3. To assess the potential of FGFR3 as a drug target, we evaluated the effects of selective inhibitors on MM and control cell lines. SU5402 and PD173074 specifically inhibited the growth of the two t(4;14)-positive MM lines, KMS-11 and OPM-2. Importantly, inhibition was still observed in the presence of <em>IL</em>-6, a growth factor known to play an important role in MM. Both compounds induced a dose-dependent reduction in cell viability and an increase in apoptosis, accompanied by a decrease in extracellular signal-related kinase phosphorylation. In contrast, no inhibition was seen with either compound against t(4;14)-negative cell lines or NCI-H929, a t(4;14)-positive, FGFR3-negative MM cell line. FGFR3 is thus a plausible candidate for targeted therapy in a subset of MM patients.

OBJECTIVE

To determine the role of vision and visual attention factors in automobile crash involvement.

METHODS

Drivers aged 65 to 84 years were identified during the baseline interview (1993-1995) of the Salisbury Eye Evaluation (SEE) Study. Crash involvement through December 1997 was determined from Maryland State motor vehicle records. Vision tests at baseline included distance acuity at normal and low luminance, contrast sensitivity, glare sensitivity, stereoacuity, and visual fields. Visual attention was evaluated with the Useful Field of View Test (UFOV; Visual Awareness, Chicago, IL). Survival analysis was used to determine the relative risk of a crash as a function of demographic variables, miles driven, vision, and visual attention.

RESULTS

One hundred twenty (6.7%) of the 1801 drivers were involved in a crash during the observation interval. Glare sensitivity and binocular field loss were significant predictors of crash involvement (P < 0.05). For those with moderate or better vision (<3 letters for glare sensitivity and <20 points missed for binocular visual fields) increased glare sensitivity or reduced visual fields were, paradoxically, associated with a reduction in crash risk, whereas for those with poorer levels of vision, increased glare sensitivity or reduced visual fields were associated with increased crash risk. Worse UFOV score was associated with increased crash risk.

CONCLUSIONS

Glare sensitivity, visual field loss, and UFOV were significant predictors of crash involvement. Acuity, contrast sensitivity, and stereoacuity were not associated with crashes. These results suggest that current vision screening for drivers' licensure, based primarily on visual acuity, may miss important aspects of visual impairment.

Patients with head and neck squamous cell carcinoma (HNSCC) have profound immune defects. These defects are associated with a poor prognosis and are mediated, in part, by immune inhibitory CD34(+) progenitor cells, whose numbers are increased in the peripheral blood of HNSCC patients. Immune inhibitory CD34(+) cells are also present within HNSCC tumors. A phase IB clinical trial was conducted with HNSCC patients to determine if treatment with the differentiation-inducer 25-hydroxyvitamin D(3) could diminish CD34(+) cell levels and improve a panel of immune parameters. Here we present the results of treatment with orally administered escalating doses (<em>20</em>, 40, 60 microg) of 25-hydroxyvitamin D(3), with an emphasis on the six patients who received the maximum dosage of 60 microg per day. Peripheral blood was collected at 0, 1, 2, 4, and 6 weeks, and assessed for markers of immune activity. Although no clinical responses were observed, results of this pilot study demonstrated that treatment of HNSCC patients with 25-hydroxyvitamin D(3 )reduces the number of immune suppressive CD34(+) cells, increases HLA-DR expression, increases plasma <em>IL</em>-12 and IFN-gamma levels, and improves T-cell blastogenesis. In contrast, 25-hydroxyvitamin D(3) treatment did not modulate plasma <em>IL</em>-1beta, <em>IL</em>-2, <em>IL</em>-4, <em>IL</em>-6, <em>IL</em>-10, GM-CSF, or TGF-beta levels.

Asthma and atopy are related conditions that may share similar genetic susceptibility. Linkage studies have identified a region on chromosome 5q that contains biologic candidates for both asthma and atopy phenotypes, including several proinflammatory cytokines. Interleukin (<em>IL</em>)-13, one of the candidate genes in the region, is directly involved in the regulation of immunoglobulin E and has been associated with both asthma and atopy. We sought to identify new polymorphisms in the <em>IL</em>-13 gene, and evaluated the involvement of a subset of these variants in asthma and atopy in a case-control study using probands and spouses from a Dutch asthma family study. <em>IL</em>-13 was sequenced in <em>20</em> probands and <em>20</em> unaffected spouses, and 10 polymorphisms were identified, four novel and six previously reported. Three single nucleotide (nt) polymorphisms (SNPs) were detected in the 5'-promoter region, two in intron 1, and five in exon 4. Only one of the exon 4 SNPs resulted in an amino-acid change (Arg130Gln). We analyzed three SNPs in <em>IL</em>-13 in an extended group of 184 probands and their spouses: one in the promoter region (-1111), the Arg130Gln (nt position 4257), and a 3' untranslated region SNP (nt position 4738). The most significant associations were observed to asthma (P = 0.005), bronchial hyperresponsiveness (P = 0.003), and skin-test responsiveness (P = 0.03) with the -1111 promoter. These results provide evidence that variation in the <em>IL</em>-13 gene is involved in the pathogenesis of asthma and atopy. Further investigation is required to determine which specific alleles or combination of alleles contribute to these phenotypes, and the possible downstream effects of the resulting change in <em>IL</em>-13 levels or activity.

Eighteen Caucasian (white, Middle East and Asian) children diagnosed by paediatric rheumatologists in the UK and France as having systemic juvenile idiopathic arthritis (sJIA) were enrolled in this open label, single dose trial. All patients had evidence of continued symptoms and disease activity for at least three months while receiving >0.2 mg/kg/day of prednisolone, or its equivalent, prior to recruitment. Twelve patients also received methotrexate (< or =<em>20</em> mg/m2/week). The patients were divided into three groups receiving 2, 4 or 8 mg/kg of MRA (tocilizumab) by intravenous infusion. No evidence of dose-limiting toxicity was observed and there were no dose-limiting safety issues. MRA appeared to be dramatically effective, with clinical and laboratory responses observed by 48 h post infusion, and these improvements continued well after serum MRA was undetectable. Eleven patients achieved the JIA definition of improvement (at least 3 of 6 core set criteria with a 30% improvement and no more than one worsened by 30%) and eight achieved>> or =50% improvement. There were no observable differences with age. Clinical improvement in these children was observed for up to eight weeks, supporting the hypothesis that <em>IL</em>-6 is a key cytokine in the upregulation of genes crucial in the inflammation processes of sJIA, and the possibility of sequestration of MRA in the extra-vascular compartment needs to be considered.

BACKGROUND

The objective of this study was to investigate the effects of tumor necrosis factor (TNF)-α inhibitors on circulating T helper-type 17 (Th17) cells and Th17-related cytokines in patients with rheumatoid arthritis (RA).

METHODS

The frequencies of circulating Th17 cells and serum levels of Th17-related cytokines were determined using flow cytometry analysis and ELISA, respectively, in 48 RA patients both before (baseline) and six months after anti-TNF-α therapy. Therapeutic response was evaluated using European League Against Rheumatism (EULAR) response criteria.

RESULTS

Significantly higher baseline frequencies of circulating Th17 cells and serum levels of interleukin (<em>IL</em>)-6, <em>IL</em>-17, <em>IL</em>-21, <em>IL</em>-23 and TNF-α were observed in active RA patients than in 12 healthy controls (all P < 0.001). After anti-TNF-α therapy, 36 patients (75%) were EULAR responders (<em>20</em> good responders and 16 moderate responders) and 12 (25.0%) were non-responders. The mean levels of circulating Th17 cells and <em>IL</em>-17 significantly decreased (1.13% vs. 0.79%; 43.1 pg/ml vs. 27.8 pg/ml; respectively, both P < 0.001) in parallel with clinical remission in responders. Levels of <em>IL</em>-6, <em>IL</em>-21, <em>IL</em>-23 and TNF-α were significantly decreased after anti-TNF-α therapy in responders. In contrast, the mean levels of circulating Th17 cells and <em>IL</em>-17 significantly increased after anti-TNF-α therapy (2.94% vs. 4.23%; 92.1 pg/ml vs. 148.6 pg/ml; respectively, both P < 0.05) in non-responders. Logistic regression analysis identified a high baseline level of <em>IL</em>-17 as a significant predictor of poor therapeutic response.

CONCLUSIONS

The beneficial effect of anti-TNF-α therapy might involve a decrease in Th17-related cytokines in responders, whereas rising levels of circulating Th17-cells and IL-17 were observed in patients with an inadequate response to anti-TNF-α therapy.

Identification of the components of protective immunity are crucial for the development of effective prophylactic and therapeutic vaccine strategies. Analysis of HIV-specific responses in exposed but uninfected individuals might thus provide a unique resource to elucidate the components and correlates of protective immunity to HIV. In the present study we analyzed HIV-specific cytotoxic and helper T lymphocyte responses in health care workers (HCW) exposed to body fluids from HIV-positive individuals. HCW exposed to blood from HIV-negative individuals as well as healthy donors served as controls. Cytotoxic T lymphocyte (CTL) responses to HIV envelope (env) peptides were detected in 7/<em>20</em> (35%) HCW exposed to HIV-positive blood and in none of the <em>20</em> health care workers exposed to uninfected blood or the seven healthy blood donors studied. HIV-specific CTL responses were detected only after in vitro stimulation, and were MHC class I restricted. No MHC class I restriction elements were uniformly identified among the different responders. 21/28 (75%) HCW exposed to contaminated blood responded to env as measured by <em>IL</em>-2 production to the peptides, in contrast to only 9/38 (24%) HCW exposed to HIV seronegative blood and 3/35 (9%) healthy blood donors. All the HIV exposed individuals were seronegative on repeated ELISA tests, and no evidence of infection was obtained by PCR analysis. These findings indicate that a single exposure to HIV can induce CTL immunity to HIV antigens, in the absence of other evidence of infection.