Successive treatment with cyclosporine and infliximab in steroid-refractory ulcerative colitis.

OBJECTIVES: Rescue therapy with either cyclosporine (CYS) or infliximab (IFX) is an effective option in patients with intravenous steroid-refractory attacks of ulcerative colitis (UC). In patients who fail, colectomy is usually recommended, but a second-line rescue therapy with IFX or CYS is an alternative. The aims of this study were to investigate the efficacy and tolerance of IFX and CYS as a second-line rescue therapy in steroid-refractory UC or indeterminate colitis (IC) unsuccessfully treated with CYS or IFX.METHODS: This was a retrospective survey of patients seen during the period 2000-2008 in the GETAID centers. Inclusion criteria included a delay of <1 month between CYS withdrawal (when used first) and IFX, or a delay of <2 months between IFX (when used first) and CYS, and a follow-up of at least 3 months after inclusion. Time-to-colectomy, clinical response, and occurrence of serious adverse events were analyzed.RESULTS: A total of 86 patients (median age 34 years; 49 males; 71 UC and 15 IC) were successively treated with CYS and IFX. The median (+/-s.e.) follow-up time was 22.6 (7.0) months. During the study period, 49 patients failed to respond to the second-line rescue therapy and underwent a colectomy. The probability of colectomy-free survival (+/-s.e.) was 61.3 +/- 5.3% at 3 months and 41.3 +/- 5.6% at 12 months. A case of fatal pulmonary embolism occurred at 1 day after surgery in a 45-year-old man. Also, nine infectious complications were observed during the second-line rescue therapy.CONCLUSIONS: In patients with intravenous steroid-refractory UC and who fail to respond to CYS or IFX, a second-line rescue therapy may be effective in carefully selected patients, avoiding colectomy within 2 months in two-thirds of them. The risk/benefit ratio should still be considered individually.

Molecular genetic approaches to the biology of the moss "Physcomitrella patens"

La mousse haplobiontique Physcomitrella patens est utilisée comme système génétique modèle pour l'étude du développement des plantes. Cependant, l'absence d'un protocole efficace de transformation a constitué jusqu'à présent un gros désavantage méthodologique pour le développement futur de ce système expérimental. Les résultats présentés dans le premier chapitre relatent la mise au point d'un protocole de transformation basé sur la technique de transfert direct de gènes dans des protoplastes par précipitation au PEG. Un essai d'expression transitoire de gènes a été mis au point. Ce protocole a été adapté afin de permettre l'introduction in vivo d'anticorps dans des protoplastes. Le protocole modifié permet d'introduire simultanément du DNA et des IgG dans les cellules, et nous avons démontré que ces anticorps peuvent inactiver spécifiquement le produit d'un gène co-introduit (GUS), ainsi que certaines protéines impliquées dans des processus cellulaires (tubuline). Cet essai, baptisé "essai transitoire d'immuno-inactivation in vivo", devrait être directement applicable à d'autres protoplastes végétaux, et permettre l'élaboration de nouvelles stratégies dans l'étude de processus cellulaires. Le second chapitre est consacré aux expériences de transformation de la mousse avec des gènes conférant une résistance à des antibiotiques. Nos résultats démontrent que l'intégration de gènes de résistance dans le génome de P. patens est possible, mais que cet événement est rare. Il s'agit là néanmoins de la première démonstration d'une transformation génétique réussie de cet organisme. L'introduction de gènes de résistance aux antibiotiques dans les protoplastes de P. patens génère à haute fréquence des clones résistants instables. Deux classes de clones instables ont été identifiés. La caractérisation phénotypique, génétique et moléculaire de ces clones suggère fortement que les séquences transformantes sont concaténées pour former des structures de haut poids moléculaire, et que ces structures sont efficacement répliquées et maintenues dans les cellules résistantes en tant qu'éléments génétiques extrachromosomaux. Ce type de transformation nous permet d'envisager des expériences permettant l'identification des séquences génomiques impliquées dans la replication de l'ADN de mousse. Plusieurs lignées transgéniques ont été retransformées avec des plasmides portant des séquences homologues aux séquences intégrées dans le génome, mais conférant une résistance à un autre antibiotique. Les résultats présentés dans le troisième chapitre montrent que les fréquences de transformation intégrative dans les lignées transgéniques sont 10 fois plus élevées que dans la lignée sauvage, et que cette augmentation est associée à une coségrégation des gènes de résistance dans la plupart des clones testés. Ces résultats génétiques indiquent que l'intégration de séquences d'ADN étranger dans le génome de P. patens a lieu en moyenne 10 fois plus fréquemment par recombinaison homologue que par intégration aléatoire. Ce rapport homologue/aléatoire est 10000 fois supérieur aux rapports obtenus avec d'autres plantes, et fournit l'outil indispensable à la réalisation d'expériences de génétique inverse dans cet organisme à haplophase dominante. THESIS SUMMARY The moss Physcomitrella patens is used as a model genetic system to study plant development, taking advantage of the fact that the haploid gametophyte dominates in its life cycle. But further development of this model system was hampered by the lack of a protocol allowing the genetic transformation of this plant. We have developed a transformation protocol based on PEG-mediated direct gene transfer to protoplasts. Our data demonstrate that this procedure leads to the establishment of an efficient transient gene expression assay. A slightly modified protocol has been developed allowing the in vivo introduction of antibodies in moss protoplasts. Both DNA and IgGs can be loaded simultaneously, and specific antibodies can immunodeplete the product of an expression cassette (GUS) as well as proteins involved in cellular processes (tubulins). This assay, named transient in vivo immunodepletion assay, should be applicable to other plant protoplasts, and offers new approaches to study cellular processes. Transformations have been performed with bacterial plasmids carrying antibiotic resistance expression cassette. Our data demonstrate that integrative transformation occurs, but at low frequencies. This is the first demonstration of a successful genetic transformation of mosses. Resistant unstable colonies are recovered at high frequencies following transformation, and two different classes of unstable clones have been identified. Phenotypical, genetic and molecular characterisation of these clones strongly suggests that bacterial plasmids are concatenated to form high molecular arrays which are efficiently replicated and maintained as extrachromosomal elements in the resistant cells. Replicative transformation in P. patens should allow the design of experiments aimed at the identification of genomic sequences involved in moss DNA replication. Transgenic strains have been retransformed with bacterial plasmids carrying sequences homologous to the integrated transloci, but conferring resistance to another antibiotic. Our results demonstrate an order of magnitude increase of integrative transformation frequencies in transgenic strains as compared to wild-type, associated with cosegregation of the resistance genes in most of these double resistant transgenic strains. These observations provide strong genetic evidence that gene targeting occurs about ten times more often than random integration in the genome of P. patens. Such ratio of targeted to random integration is about 10 000 times higher than previous reports of gene targeting in plants, and provides the essential requirement for the development of efficient reverse genetics in the haplodiplobiontic P. patens.

Identification of opsA, a gene involved in solute stress mitigation and survival in soil, in the polycyclic aromatic hydrocarbon-degrading bacterium Novosphingobium sp. strain LH128.

The aim of this study was to identify genes involved in solute and matric stress mitigation in the polycyclic aromatic hydrocarbon (PAH)-degrading Novosphingobium sp. strain LH128. The genes were identified using plasposon mutagenesis and by selection of mutants that showed impaired growth in a medium containing 450 mM NaCl as a solute stress or 10% (wt/vol) polyethylene glycol (PEG) 6000 as a matric stress. Eleven and 14 mutants showed growth impairment when exposed to solute and matric stresses, respectively. The disrupted sequences were mapped on a draft genome sequence of strain LH128, and the corresponding gene functions were predicted. None of them were shared between solute and matric stress-impacted mutants. One NaCl-affected mutant (i.e., NA7E1) with a disruption in a gene encoding a putative outer membrane protein (OpsA) was susceptible to lower NaCl concentrations than the other mutants. The growth of NA7E1 was impacted by other ions and nonionic solutes and by sodium dodecyl sulfate (SDS), suggesting that opsA is involved in osmotic stress mitigation and/or outer membrane stability in strain LH128. NA7E1 was also the only mutant that showed reduced growth and less-efficient phenanthrene degradation in soil compared to the wild type. Moreover, the survival of NA7E1 in soil decreased significantly when the moisture content was decreased but was unaffected when soluble solutes from sandy soil were removed by washing. opsA appears to be important for the survival of strain LH128 in soil, especially in the case of reduced moisture content, probably by mitigating the effects of solute stress and retaining membrane stability.

Follicular lymphoma at relapse after rituximab containing regimens: comparison of time to event intervals prior to and after (90) Y-ibritumomab-tiuxetan.

Radioimmunotherapies with Zevalin® (RIT-Z) showed encouraging results in patients with relapsed/refractory follicular lymphoma (FL), leading frequently to failure-free intervals longer than those achieved by the last previous therapy. We compared time-to-event variables obtained before and after RIT-Z in patients with relapsed FL, previously exposed to rituximab. All patients with relapsed non-transformed, non-refractory, non-rituximab-naïve FL who have been treated with RIT-Z in two different centres in Europe were included. Staging and response were assessed by contrast-enhanced CT in all patients; PET/CT was performed according to local availability. Event-free survival (EFS) and time to next treatment (TTNT) following the last previous therapy and after RIT-Z were compared. Pre-therapy characteristics were tested in univariate analyses for prediction of outcomes. A description of the patterns of relapse was also provided. Among 70 patients treated, only 16 fulfilled the inclusion criteria. They were treated with a median of 3 prior lines of chemo-immunotherapies, including a median of 2 rituximab-containing regimens; 6 patients had undergone myeloablative chemotherapy with autologous stem cell rescue (ASCT). Overall response rates were 10 (62%) CR/CRu, 3 (19%) PR and 3 (19%) PD; response rates were similar in patients with prior ASCT. After RIT-Z only few patients obtained EFS and TTNT longer than after the last previous therapy. All four patients receiving rituximab maintenance were without progression 12 months after RIT-Z. Relapses occurred in both previously and newly involved sites; a significant association was found between the number of pathologic sites involved prior to RIT-Z and subsequent TTNT. Despite the excellent response rate, the duration of response was shorter than the previous one confirming the known trend of relapses to occur earlier after subsequent treatments. Rituximab maintenance after RIT-Z showed encouraging results in terms of prolonging EFS, warranting further studies. Copyright © 2010 John Wiley &amp; Sons, Ltd.

The anti-apoptotic factor Bcl-2 can functionally substitute for the B cell survival but not for the marginal zone B cell differentiation activity of BAFF.

The TNF family ligand B cell-activating factor (BAFF, BLyS, TALL-1) is an essential factor for B cell development. BAFF binds to three receptors, BAFF-R, transmembrane activator and CAML interactor (TACI), and B cell maturation antigen (BCMA), but only BAFF-R is required for successful survival and maturation of splenic B cells. To test whether the effect of BAFF is due to the up-regulation of anti-apoptotic factors, TACI-Ig-transgenic mice, in which BAFF function is inhibited, were crossed with transgenic mice expressing FLICE-inhibitory protein (FLIP) or Bcl-2 in the B cell compartment. FLIP expression did not rescue B cells, while enforced Bcl-2 expression restored peripheral B cells and the ability to mount T-dependent antibody responses. However, many B cells retained immaturity markers and failed to express normal amounts of CD21. Marginal zone B cells were not restored and the T-independent IgG3, but not IgM, response was impaired in the TACI-IgxBcl-2 mice. These results suggest that BAFF is required not only to inhibit apoptosis of maturating B cells, but also to promote differentiation events, in particular those leading to the generation of marginal zone B cells.

A large-scale genetic validation study coupled with in vitro analyses reveal a role for vitamin Dsignaling in the pathogenesis and response to treatment of hepatitis C virus infection

Background and Aims: Vitamin D is an important modulatorof numerous cellular processes. Some of us recently observedan association of the 1a-hydroxylase promoter polymorphismCYP27B1-1260 rs10877012 with sustained virologic response (SVR)in a relatively small number of German patients with chronichepatitis C. In the present study, we aimed to validate thisassociation in a large and well characterized patient cohort, theSwiss Hepatitis C Cohort Study (SCCS). In addition, we examinedthe effect of vitamin D on the hepatitis C virus (HCV) life cyclein vitro.Methods: CYP27B1-1260 rs10877012 and IL28B rs12979860 singlenucleotide polymorphisms (SNPs) were genotyped in 1049 patientswith chronic hepatitis C from the SCCS, of whom 698 were treatedwith pegylated interferon-a (PEG-IFN-a) and ribavirin. In addition,112 patients with spontaneous clearance of HCV were examined.SNPs were correlated with variables reflecting the natural courseand treatment outcome of chronic hepatitis C. The effect of1,25-(OH)2D3 (calcitriol) on HCV replication and viral particleproduction was investigated in vitro using human hepatoma celllines (Huh-7.5) harbouring subgenomic replicons and cell culturederivedHCV.Results: The CYP27B1-1260 rs10877012 genotype was notassociated with SVR in patients with the good-response IL28Brs1279860 CC genotype. However, in patients with poor-responseIL28B rs1279860 genotype CT and TT, CYP27B1-1260 rs10877012was a significant independent predictor of SVR (15% difference inSVR between rs10877012 genotype AA vs. CC, p = 0.030, OR = 1.495,95% CI = 1.038-2.152). The CYPB27-1260 rs10877012 genotype wasneither associated with spontaneous clearance of HCV, nor withliver fibrosis progression rate, inflammatory activity of chronichepatitis C, or HCV viral load. Physiological doses of 1,25-(OH)2D3did not significantly affect HCVRNA replication or infectiousparticle production in vitro.Conclusions: The results of this large-scale genetic validationstudy reveal a role of vitamin D metabolism in the responseto treatment in chronic hepatitis C, but 1,25-(OH)2D3 does notexhibit a significant direct inhibitory antiviral effect. Thus, theability of vitamin D to modulate immunity against HCV shouldbe investigated.

La caverna de Josep Palau i Fabre (1952-1969): teatre versus al·legoria filosòfica per a representar i impedir el final tràgic i fosc del lluminós art de pensar.

What is the use of performing the myth of the cave from book VII of the Republic by Plato? Josep Palau i Fabre, considers that, in Plato's dialogues, the speakers are mere instruments at the service of his dialectical goal. The aim of this article is to show how, by turning the myth into a tragedy and also by relying on Heraclitus's conflict or war of opposites, the playwright succeeds in favoring a sort of thought which is not one-sided or univocal. On the contrary, in Palau i Fabre's La Caverna, the tragic hero, that is, the released prisoner transformed by the light of Reality and finally killed by his "cavemates" -after having been imprisoned again and having tried to rescue them from their ignorance or shadows-, still leaves to them his powerful experience of the agonistikos thought, which might bear fruit in their life to come.

La caverna de Josep Palau i Fabre (1952-1969): teatro versus alegoría filosófica para representar e impedir el final trágico y sombrío del luminoso arte de pensar.

What is the use of performing the myth of the cave from book VII of the Republic by Plato? Josep Palau i Fabre, considers that, in Plato's dialogues, the speakers are mere instruments at the service of his dialectical goal. The aim of this article is to show how, by turning the myth into a tragedy and also by relying on Heraclitus's conflict or war of opposites, the playwright succeeds in favoring a sort of thought which is not one-sided or univocal. On the contrary, in Palau i Fabre's La Caverna, the tragic hero, that is, the released prisoner transformed by the light of Reality and finally killed by his "cavemates" -after having been imprisoned again and having tried to rescue them from their ignorance or shadows-, still leaves to them his powerful experience of the agonistikos thought, which might bear fruit in their life to come.

In vivo activation of PPAR target genes by RXR homodimers.

The ability of a retinoid X receptor (RXR) to heterodimerize with many nuclear receptors, including LXR, PPAR, NGF1B and RAR, underscores its pivotal role within the nuclear receptor superfamily. Among these heterodimers, PPAR:RXR is considered an important signalling mediator of both PPAR ligands, such as fatty acids, and 9-cis retinoic acid (9-cis RA), an RXR ligand. In contrast, the existence of an RXR/9-cis RA signalling pathway independent of PPAR or any other dimerization partner remains disputed. Using in vivo chromatin immunoprecipitation, we now show that RXR homodimers can selectively bind to functional PPREs and induce transactivation. At the molecular level, this pathway requires stabilization of the homodimer-DNA complexes through ligand-dependent interaction with the coactivator SRC1 or TIF2. This pathway operates both in the absence and in the presence of PPAR, as assessed in cells carrying inactivating mutations in PPAR genes and in wild-type cells. In addition, this signalling pathway via PPREs is fully functional and can rescue the severe hypothermia phenotype observed in fasted PPARalpha-/- mice. These observations have important pharmacological implications for the development of new rexinoid-based treatments.

ORMDL proteins are a conserved new family of endoplasmic reticulum membrane proteins

Background: Annotations of completely sequenced genomes reveal that nearly half of the genes identified are of unknown function, and that some belong to uncharacterized gene families. To help resolve such issues, information can be obtained from the comparative analysis of homologous genes in model organisms. Results: While characterizing genes from the retinitis pigmentosa locus RP26 at 2q31-q33, we have identified a new gene, ORMDL1, that belongs to a novel gene family comprising three genes in humans (ORMDL1, ORMDL2 and ORMDL3), and homologs in yeast, microsporidia, plants, Drosophila, urochordates and vertebrates. The human genes are expressed ubiquitously in adult and fetal tissues. The Drosophila ORMDL homolog is also expressed throughout embryonic and larval stages, particularly in ectodermally derived tissues. The ORMDL genes encode transmembrane proteins anchored in the endoplasmic reticulum (ER). Double knockout of the two Saccharomyces cerevisiae homologs leads to decreased growth rate and greater sensitivity to tunicamycin and dithiothreitol. Yeast mutants can be rescued by human ORMDL homologs. Conclusions: From protein sequence comparisons we have defined a novel gene family, not previously recognized because of the absence of a characterized functional signature. The sequence conservation of this family from yeast to vertebrates, the maintenance of duplicate copies in different lineages, the ubiquitous pattern of expression in human and Drosophila, the partial functional redundancy of the yeast homologs and phenotypic rescue by the human homologs, strongly support functional conservation. Subcellular localization and the response of yeast mutants to specific agents point to the involvement of ORMDL in protein folding in the ER.

Stratégies thérapeutiques dans les myopathies inflammatoires [Therapeutic strategies in idiopatic inflammatory myopathies]

Idiopathic inflammatory myopathies, such as polymyositis and dermatomyositis, share common clinical features such as progressive, symmetrical muscle weakness prevailing in the lower limbs, associated sometimes with muscle pains. High CK and typical biopsy insure the diagnosis. Possible causes for secondary myopathies and associated diseases should be actively investigated. The search for autoantibodies helps to better classify inflammatory myopathies and to better define the prognosis of the myopathy. Glucocorticoids are the cornerstone of the early phase therapy. Glucocorticoid-sparing agents, such as azathioprine and methotrexate, are second line agents but can be readily prescribed. In case of therapeutic resistance, a rescue treatment (ciclosporine, immunoglobulins, rituximab, cyclophosphamide) could be considered.

The Role of Interleukin-2 in Memory CD8 Cell Differentiation

The current literature on the role of interleukin (IL)-2 in memory CD8(+) T-cell differentiation indicates a significant contribution of IL-2 during primary and also secondary expansion of CD8(+) T cells. IL-2 seems to be responsible for optimal expansion and generation of effector functions following primary antigenic challenge. As the magnitude of T-cell expansion determines the numbers of memory CD8(+) T cells surviving after pathogen elimination, these events influence memory cell generation. Moreover, during the contraction phase of an immune response where most antigen-specific CD8(+) T cells disappear by apoptosis, IL-2 signals are able to rescue CD8(+) T cells from cell death and provide a durable increase in memory CD8(+) T-cell counts. At the memory stage, CD8(+) T-cell frequencies can be boosted by administration of exogenous IL-2. Significantly, only CD8(+) T cells that have received IL-2 signals during initial priming are able to mediate efficient secondary expansion following renewed antigenic challenge. Thus, IL-2 signals during different phases of an immune response are key in optimizing CD8(+) T-cell functions, thereby affecting both primary and secondary responses of these T cells.

ICRC Factsheet, June 2010

Three Classes of Animals: 1. Pets are animals living with owners for purposes of love, affection, and company. 2. Emotional Support Animals provide some therapeutic benefit to person with mental or psychiatric disability, requiring no specific training. The mere presence of this animal mitigate the effects of the emotional or mental disability. 3. Service Animals are any animal individually trained to do work or perform tasks for the benefit of an individual with a physical, intellectual, and mental disability—IE guiding individuals with impaired vision, providing protection or rescue work, pulling a wheel chair, or fetching dropped items.

Early use of TIPS in patients with cirrhosis and variceal bleeding.

Background Patients with cirrhosis in ChildPugh class C or those in class B who have persistent bleeding at endoscopy are at high risk for treatment failure and a poor prognosis, even if they have undergone rescue treatment with a transjugular intrahepatic porto - systemic shunt (TIPS). This study evaluated the earlier use of TIPS in such patients. Methods We randomly assigned, within 24 hours after admission, a total of 63 patients with cirrhosis and acute variceal bleeding who had been treated with vasoactive drugs plus endoscopic therapy to treatment with a polytetrafluoroethylene-covered stent within 72 hours after randomization (early-TIPS group, 32 patients) or continuation of vasoactive-drug therapy, followed after 3 to 5 days by treatment with propranolol or nadolol and long-term endoscopic band ligation (EBL), with insertion of a TIPS if needed as rescue therapy (pharmacotherapyEBL group, 31 patients). Results During a median follow-up of 16 months, rebleeding or failure to control bleeding occurred in 14 patients in the pharmacotherapyEBL group as compared with 1 patient in the early-TIPS group (P=0.001). The 1-year actuarial probability of remaining free of this composite end point was 50% in the pharmacotherapyEBL group versus 97% in the early-TIPS group (P<0.001). Sixteen patients died (12 in the pharmacotherapyEBL group and 4 in the early-TIPS group, P=0.01). The 1-year actuarial survival was 61% in the pharmacotherapyEBL group versus 86% in the early-TIPS group (P<0.001). Seven patients in the pharmacotherapyEBL group received TIPS as rescue therapy, but four died. The number of days in the intensive care unit and the percentage of time in the hospital during follow-up were significantly higher in the pharmacotherapyEBL group than in the early-TIPS group. No significant diferences were observed between the two treatment groups with respect to serious adverse events. Conclusions In these patients with cirrhosis who were hospitalized for acute variceal bleeding and at high risk for treatment failure, the early use of TIPS was associated with signif icant reductions in treatment failure and in mortality. (Current Controlled Trials number, ISRCTN58150114.)

Nadolol plus isosorbide mononitrate alone or associated with band ligation in the prevention of recurrent bleeding: A multicenter randomized controlled trial.

Background and aims: Previous clinical trials suggest that adding non-selective beta-blockers improves the efficacy of endoscopic band ligation (EBL) in the prevention of recurrent bleeding, but no study has evaluated whether EBL improves the efficacy of beta-blockers + isosorbide-5-mononitrate. The present study was aimed at evaluating this issue in a multicentre randomised controlled trial (RCT) and to correlate changes in hepatic venous pressure gradient (HVPG) during treatment with clinical outcomes. Methods: 158 patients with cirrhosis, admitted because of variceal bleeding, were randomised to receive nadolol+isosorbide-5-mononitrate alone (Drug: n=78) or combined with EBL (Drug+EBL; n=80). HVPG measurements were performed at randomisation and after 4¿6 weeks on medical therapy. Results: Median follow-up was 15 months. One-year probability of recurrent bleeding was similar in both groups (33% vs 26%: p=0.3). There were no significant differences in survival or need of rescue shunts. Overall adverse events or those requiring hospital admission were significantly more frequent in the Drug+EBL group. Recurrent bleeding was significantly more frequent in HVPG non-responders than in responders (HVPG reduction ¿20% or ¿12 mm Hg). Among non-responders recurrent bleeding was similar in patients treated with Drugs or Drugs+EBL. Conclusions: Adding EBL to pharmacological treatment did not reduce recurrent bleeding, the need for rescue therapy, or mortality, and was associated with more adverse events. Furthermore, associating EBL to drug therapy did not reduce the high rebleeding risk of HVPG non-responders.