Inhibition of Secretion of Interleukin (IL)-12/IL-23 Family Cytokines by 4-Trifluoromethyl-celecoxib Is Coupled to Degradation via the Endoplasmic Reticulum Stress Protein HERP

Interleukin-12 (IL-12), p80, and IL-23 are structurally related cytokines sharing a p40 subunit. We have recently demonstrated that celecoxib and its COX-2-independent analogue 4-trifluoromethyl-celecoxib (TFM-C) inhibit secretion but not transcription of IL-12 (p35/p40) and p80 (p40/p40). This is associated with a mechanism involving altered cytokine-chaperone interaction in the endoplasmic reticulum (ER). In the present study, we found that celecoxib and TFM-C also block secretion of IL-23 (p40/p19 heterodimers). Given the putative ER-centric mode of these compounds, we performed a comprehensive RTPCR analysis of 23 ER-resident chaperones/foldases and associated co-factors. This revealed that TFM-C induced 1.5-3-fold transcriptional up-regulation of calreticulin, GRP78, GRP94, GRP170, ERp72, ERp57, ERdj4, and ERp29. However, more significantly, a 7-fold up-regulation of homocysteine-inducible ER protein (HERP) was observed. HERP is part of a high molecular mass protein complex involved in ER-associated protein degradation (ERAD). Using co-immunoprecipitation assays, we show that TFM-C induces protein interaction of p80 and IL-23 with HERP. Both HERP siRNA knockdown and HERP overexpression coupled to cycloheximide chase assays revealed that HERP is necessary for degradation of intracellularly retained p80 by TFM-C. Thus, our data suggest that targeting cytokine folding in the ER by small molecule drugs could be therapeutically exploited to alleviate in appropriate inflammation in autoimmune conditions.

New York City: 12 December, 1980

This article compares two documentary treatments of the Central Park vigil for John Lennon in 1980: Happy Birthday to John (Jonas Mekas, 1995, 16mm, 18 min.), and Dix minutes de silence pour John Lennon/Ten Minutes Silence for John Lennon (Raymond Depardon, 1980, 16mm, 10 min.). Mekas and Depardon might seem an improbable combination but as the article demonstrates there are affinities, if not direct points of convergence, in outlook and documentary method: both sensibilities have been shaped by migrant experiences, and much of their work, for all its formal and structural differences, is preoccupied with experiences of exile and displacement, rootedness and the meaning of home, the country and the city (and in Mekas’s case, the country in the city). Mekas and Depardon are also Europeans who have developed an intimate social and artistic relationship with New York City; both are concerned with the place of autobiography in their work, using captions, inter-titles, diary entries, photographs, and 1st person commentary to complicate relations between the imaginary and the documentary. In addition to discussing the significance of these preoccupations, and differences in the manner in which both filmmakers witness the apotheosis of Lennon as cultural martyr (and natural New Yorker), the article also examines the phenomenon of public mourning, and how it often displaces its ostensible subject: associatively, in the case of Mekas; incidentally, in the case of Depardon; and intentionally, in the case of the mass media, and popular culture.

Participation of 8-12 year old children with cerebral palsy: a cross-sectional European Survey

Objectives
To evaluate how involvement in life situations (participation) in children with cerebral palsy varies with type and severity of impairment and to investigate geographical variation in participation.

Design
Cross sectional study. Trained interviewers visited parents of children with cerebral palsy; multilevel multivariable regression related participation to impairments, pain, and sociodemographic characteristics.

Setting
Eight European regions with population registers of children with cerebral palsy; one further region recruited children from multiple sources.

Participants
1174 children aged 8-12 with cerebral palsy randomly selected from the population registers, 743 (63%) joined in the study; the further region recruited 75 children.

Main outcome measure
Children’s participation assessed by the Life-H questionnaire covering 10 main areas of daily life. Scoring ignored adaptations or assistance required for participation.

Results
Children with pain and those with more severely impaired walking, fine motor skills, communication, and intellectual abilities had lower participation across most domains. Type of cerebral palsy and problems with feeding and vision were associated with lower participation for specific domains, but the sociodemographic factors examined were not. Impairment and pain accounted for up to a sixth of the variation in participation. Participation on all domains varied substantially between regions: children in east Denmark had consistently higher participation than children in other regions. For most participation domains, about a third of the unexplained variation could be ascribed to variation between regions and about two thirds to variation between individuals.

Conclusions
Participation in children with cerebral palsy should be assessed in clinical practice to guide intervention and assess its effect. Pain should be carefully assessed. Some European countries facilitate participation better than others, implying some countries could make better provision. Legislation and regulation should be directed to ensuring this happens.

Mesophases in Nearly 2D Room-Temperature Ionic Liquids

Computer simulations of (i) a [C(12)mim][Tf2N] film of nanometric thickness squeezed at kbar pressure by a piecewise parabolic confining potential reveal a mesoscopic in-plane density and composition modulation reminiscent of mesophases seen in 3D samples of the same room-temperature ionic liquid (RTIL). Near 2D confinement, enforced by a high normal load, as well as relatively long aliphatic chains are strictly required for the mesophase formation, as confirmed by computations for two related systems made of (ii) the same [C(12)mim][Tf2N] adsorbed at a neutral solid surface and (iii) a shorter-chain RTIL ([C(4)mim][Tf2N]) trapped in the potential well of part i. No in-plane modulation is seen for ii and iii. In case ii, the optimal arrangement of charge and neutral tails is achieved by layering parallel to the surface, while, in case iii, weaker dispersion and packing interactions are unable to bring aliphatic tails together into mesoscopic islands, against overwhelming entropy and Coulomb forces. The onset of in-plane mesophases could greatly affect the properties of long-chain RTILs used as lubricants.

Molecular and clinical features of the myeloproliferative neoplasm associated with JAK2 exon 12 mutations

Although approximately 95% of patients with polycythemia vera (PV) harbor the V617F mutation in JAK2 exon 14, several mutations in exon 12 have been described in the remaining patients. We conducted a European collaborative study to define the molecular and clinical features of patients harboring these mutations. Overall, 106 PVs were recruited and 17 different mutations identified. Irrespective of the mutation, two-thirds of patients had isolated erythrocytosis, whereas the remaining subjects had erythrocytosis plus leukocytosis and/or thrombocytosis. Compared with JAK2 (V617F)-positive PV patients, those with exon 12 mutations had significantly higher hemoglobin level and lower platelet and leukocyte counts at diagnosis but similar incidences of thrombosis, myelofibrosis, leukemia, and death. In a multivariable analysis, age more than 60 years and prior thrombosis predicted thrombosis. These findings suggest that, despite the phenotypical difference, the outcome of JAK2 exon 12 mutations-positive PV is similar to that of JAK2 (V617F)positive PV. (Blood. 2011; 117(10):2813-2816)

Interactions of 12-lipoxygenase with phospholipase A(2) isoforms following platelet activation through the glycoprotein VI collagen receptor

Recent studies implicate the collagen receptor, glycoprotein VI (GPVI) in activation of platelet 12-lipoxygenase (p12-LOX). Herein, we show that GPVI-stimulated 12-hydro(peroxy)eicosatetraenoic acid (H(P)ETE) synthesis is inhibited by palmityl trifluromethyl ketone or oleyloxyethyl phosphocholine, but not bromoenol lactone, implicating secretory and cytosolic, but not calcium-independent phospholipase A(2) (PLA(2)) isoforms. Also, following GPVI activation, 12-LOX co-immunoprecipitates with both cytosolic and secretory PLA(2), (sPLA(2)). Finally, venoms containing sPLA(2) acutely activate p12-LOX in a dose-dependent manner. This study shows that platelet 12-H(P)ETE generation utilizes arachidonate substrate from both c- and sPLA(2) and that 12-LOX functionally associates with both PLA(2) isoforms. (C) 2004 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Platelet 12-lipoxygenase activation via glycoprotein VI - Involvement of multiple signaling pathways in agonist control of H(P)ETE synthesis

Lipoxygenases (LOX) contribute to vascular disease and inflammation through generation of bioactive lipids, including 12-hydro(pero)xyeicosatetraenoic acid (12-H(P)ETE). The physiological mechanisms that acutely control LOX product generation in mammalian cells are uncharacterized. Human platelets that contain a 12-LOX isoform (p12-LOX) were used to define pathways that activate H( P) ETE synthesis in the vasculature. Collagen and collagen-related peptide (CRP) (1 to 10 mug/mL) acutely induced platelet 12-H(P)ETE synthesis. This implicated the collagen receptor glycoprotein VI ( GPVI), which signals via the immunoreceptor-based activatory motif (ITAM)-containing FcRgamma chain. Conversely, thrombin only activated at high concentrations (> 0.2 U/mL), whereas U46619 and ADP alone were ineffective. Collagen or CRP-stimulated 12-H( P) ETE generation was inhibited by staurosporine, PP2, wortmannin, BAPTA/AM, EGTA, and L-655238, implicating src-tyrosine kinases, PI3-kinase, Ca2+ mobilization, and p12-LOX translocation. In contrast, protein kinase C (PKC) inhibition potentiated 12-H( P) ETE generation. Finally, activation of the immunoreceptor tyrosine-based inhibitory motif (ITIM)-containing platelet endothelial cell adhesion molecule (PECAM-1) inhibited p12-LOX product generation. This study characterizes a receptor-dependent pathway for 12-H(P) ETE synthesis via the collagen receptor GPVI, which is negatively regulated by PECAM-1 and PKC, and demonstrates a novel link between immune receptor signaling and lipid mediator generation in the vasculature.