New Therapeutic Strategies for Systemic Sclerosis—a Critical Analysis of the Literature

Systemic sclerosis (SSc) is a multi-system disease characterized by skin fibrosis and visceral disease. Therapy is organ and pathogenesis targeted. In this review, we describe novel strategies in the treatment of SSc. Utilizing the MEDLINE and the COCHRANE REGISTRY, we identified open trials, controlled trials, for treatment of SSc from 1999 to April 2005. We used the terms scleroderma, systemic sclerosis, Raynaud's phenomenon, pulmonary hypertension, methotrexate, cyclosporin, tacrolimus, relaxin, low-dose penicillamine, IVIg, calcium channel blockers, losartan, prazocin, iloprost, N-acetylcysteine, bosentan, cyclophosphamide, lung transplantation, ACE inhibitors, anti-thymocyte globulin, and stem cell transplantation. Anecdotal reports were omitted

Maternal gestational vitamin D supplementation and offspring bone health (MAVIDOS): a multicentre, double-blind, randomised placebo-controlled trial.

BACKGROUND: Maternal vitamin D status has been associated with bone mass of offspring in many, but not all, observational studies. However, maternal vitamin D repletion during pregnancy has not yet been proven to improve offspring bone mass in a randomised controlled trial. We aimed to assess whether neonates born to mothers supplemented with vitamin D during pregnancy have greater whole-body bone mineral content (BMC) at birth than those of mothers who had not received supplementation. METHODS: The Maternal Vitamin D Osteoporosis Study (MAVIDOS) was a multicentre, double-blind, randomised, placebo-controlled trial that recruited pregnant women from three study sites in the UK (Southampton, Oxford, and Sheffield). Eligible participants were older than 18 years, with a singleton pregnancy, gestation of less than 17 weeks, and a serum 25-hydroxyvitamin D (25[OH]D) concentration of 25-100 nmol/L at 10-17 weeks' gestation. P'articipants were randomly assigned (1:1), in randomly permuted blocks of ten, to either cholecalciferol 1000 IU/day or matched placebo, taken orally, from 14 weeks' gestation (or as soon as possible before 17 weeks' gestation if recruited later) until delivery. Participants and the research team were masked to treatment allocation. The primary outcome was neonatal whole-body BMC, assessed within 2 weeks of birth by dual-energy x-ray absorptiometry (DXA), analysed in all randomly assigned neonates who had a usable DXA scan. Safety outcomes were assessed in all randomly assigned participants. This trial is registered with the International Standard Randomised Controlled Trial registry, ISRCTN 82927713, and the European Clinical Trials Database, EudraCT 2007-001716-23. FINDINGS: Between Oct 10, 2008, and Feb 11, 2014, we randomly assigned 569 pregnant women to placebo and 565 to cholecalciferol 1000 IU/day. 370 (65%) neonates in the placebo group and 367 (65%) neonates in the cholecalciferol group had a usable DXA scan and were analysed for the primary endpoint. Neonatal whole-body BMC of infants born to mothers assigned to cholecalciferol 1000 IU/day did not significantly differ from that of infants born to mothers assigned to placebo (61·6 g [95% CI 60·3-62·8] vs 60·5 g [59·3-61·7], respectively; p=0·21). We noted no significant differences in safety outcomes, apart from a greater proportion of women in the placebo group with severe post-partum haemorrhage than those in the cholecalciferol group (96 [17%] of 569 mothers in the placebo group vs 65 [12%] of 565 mothers in the cholecalciferol group; p=0·01). No adverse events were deemed to be treatment related. INTERPRETATION: Supplementation of women with cholecalciferol 1000 IU/day during pregnancy did not lead to increased offspring whole-body BMC compared with placebo, but did show that 1000 IU of cholecalciferol daily is sufficient to ensure that most pregnant women are vitamin D replete, and it is safe. These findings support current approaches to vitamin D supplementation in pregnancy. Results of the ongoing MAVIDOS childhood follow-up study are awaited. FUNDING: Arthritis Research UK, Medical Research Council, Bupa Foundation, and National Institute for Health Research

Genetic susceptibility to aspergillosis in allogeneic stem-cell transplantation

Invasive aspergillosis (IA) is a major threat to positive outcomes for allogeneic stem-cell transplantation (allo-SCT) patients. Despite presenting similar degrees of immunosuppression, not all individuals at-risk ultimately develop infection. Therefore, the traditional view of neutropenia as a key risk factor for aspergillosis needs to be accommodated within new conceptual advances on host immunity and its relationship to infection. Polymorphisms in innate immune genes, such as those encoding TLRs, cytokines and cytokine receptors, have recently been associated with susceptibility to IA in allo-SCT recipients. This suggests that understanding host-pathogen interactions at the level of host genetic susceptibility will allow the formulation of new targeted and patient-tailored antifungal therapeutics, including improved donor screening.Fundação para a Ciência e a Tecnologia (FCT) - SFRH/BD/65962/2009, SFRH/BPD/46292/2008Specific Targeted Research Projects MANASP (LSHE-CT-2006), contract number 037899 (FP6), Italian Project PRIN2007KLCKP8_004

Performance of the CMS Cathode Strip Chambers with Cosmic Rays

The Cathode Strip Chambers (CSCs) constitute the primary muon tracking device in the CMS endcaps. Their performance has been evaluated using data taken during a cosmic ray run in fall 2008. Measured noise levels are low, with the number of noisy channels well below 1%. Coordinate resolution was measured for all types of chambers, and fall in the range 47 microns to 243 microns. The efficiencies for local charged track triggers, for hit and for segments reconstruction were measured, and are above 99%. The timing resolution per layer is approximately 5 ns

MED12 Alterations in Both Human Benign and Malignant Uterine Soft Tissue Tumors

The relationship between benign uterine leiomyomas and their malignant counterparts, i.e. leiomyosarcomas and smooth muscle tumors of uncertain malignant potential (STUMP), is still poorly understood. The idea that a leiomyosarcoma could derive from a leiomyoma is still controversial. Recently MED12 mutations have been reported in uterine leiomyomas. In this study we asked whether such mutations could also be involved in leiomyosarcomas and STUMP oncogenesis. For this purpose we examined 33 uterine mesenchymal tumors by sequencing the hot-spot mutation region of MED12. We determined that MED12 is altered in 66.6% of typical leiomyomas as previously reported but also in 11% of STUMP and 20% of leiomyosarcomas. The mutated allele is predominantly expressed in leiomyomas and STUMP. Interestingly all classical leiomyomas exhibit MED12 protein expression while 40% of atypical leiomyomas, 50% of STUMP and 80% of leiomyosarcomas (among them the two mutated ones) do not express MED12. All these tumors without protein expression exhibit complex genomic profiles. No mutations and no expression loss were identified in an additional series of 38 non-uterine leiomyosarcomas. MED12 mutations are not exclusive to leiomyomas but seem to be specific to uterine malignancies. A previous study has suggested that MED12 mutations in leiomyomas could lead to Wnt/β-catenin pathway activation however our immunohistochemistry results show that there is no association between MED12 status and β-catenin nuclear/cytoplasmic localization. Collectively, our results show that subgroups of benign and malignant tumors share a common genetics. We propose here that MED12 alterations could be implicated in the development of smooth muscle tumor and that its expression could be inhibited in malignant tumors

Clinical and molecular characterization of a cohort of patients with novel nucleotide alterations of the Dystrophin gene detected by direct sequencing

<p>Abstract</p> <p>Background</p> <p>Duchenne and Becker Muscular dystrophies (DMD/BMD) are allelic disorders caused by mutations in the dystrophin gene, which encodes a sarcolemmal protein responsible for muscle integrity. Deletions and duplications account for approximately 75% of mutations in DMD and 85% in BMD. The implementation of techniques allowing complete gene sequencing has focused attention on small point mutations and other mechanisms underlying complex rearrangements.</p> <p>Methods</p> <p>We selected 47 patients (41 families; 35 DMD, 6 BMD) without deletions and duplications in <it>DMD </it>gene (excluded by multiplex ligation-dependent probe amplification and multiplex polymerase chain reaction analysis). This cohort was investigated by systematic direct sequence analysis to study sequence variation. We focused our attention on rare mutational events which were further studied through transcript analysis.</p> <p>Results</p> <p>We identified 40 different nucleotide alterations in DMD gene and their clinical correlates; altogether, 16 mutations were novel. DMD probands carried 9 microinsertions/microdeletions, 19 nonsense mutations, and 7 splice-site mutations. BMD patients carried 2 nonsense mutations, 2 splice-site mutations, 1 missense substitution, and 1 single base insertion. The most frequent stop codon was TGA (n = 10 patients), followed by TAG (n = 7) and TAA (n = 4). We also analyzed the molecular mechanisms of five rare mutational events. They are two frame-shifting mutations in the <it>DMD </it>gene 3'end in BMD and three novel splicing defects: IVS42: c.6118-3C>A, which causes a leaky splice-site; c.9560A>G, which determines a cryptic splice-site activation and c.9564-426 T>G, which creates pseudoexon retention within IVS65.</p> <p>Conclusion</p> <p>The analysis of our patients' sample, carrying point mutations or complex rearrangements in <it>DMD </it>gene, contributes to the knowledge on phenotypic correlations in dystrophinopatic patients and can provide a better understanding of pre-mRNA maturation defects and dystrophin functional domains. These data can have a prognostic relevance and can be useful in directing new therapeutic approaches, which rely on a precise definition of the genetic defects as well as their molecular consequences.</p

Studies of a Ring-Cleaving Dioxygenase Illuminate the Role of Cholesterol Metabolism in the Pathogenesis of Mycobacterium tuberculosis

Mycobacterium tuberculosis, the etiological agent of TB, possesses a cholesterol catabolic pathway implicated in pathogenesis. This pathway includes an iron-dependent extradiol dioxygenase, HsaC, that cleaves catechols. Immuno-compromised mice infected with a ΔhsaC mutant of M. tuberculosis H37Rv survived 50% longer than mice infected with the wild-type strain. In guinea pigs, the mutant disseminated more slowly to the spleen, persisted less successfully in the lung, and caused little pathology. These data establish that, while cholesterol metabolism by M. tuberculosis appears to be most important during the chronic stage of infection, it begins much earlier and may contribute to the pathogen's dissemination within the host. Purified HsaC efficiently cleaved the catecholic cholesterol metabolite, DHSA (3,4-dihydroxy-9,10-seconandrost-1,3,5(10)-triene-9,17-dione; kcat/Km = 14.4±0.5 µM−1 s−1), and was inactivated by a halogenated substrate analogue (partition coefficient<50). Remarkably, cholesterol caused loss of viability in the ΔhsaC mutant, consistent with catechol toxicity. Structures of HsaC:DHSA binary complexes at 2.1 Å revealed two catechol-binding modes: bidentate binding to the active site iron, as has been reported in similar enzymes, and, unexpectedly, monodentate binding. The position of the bicyclo-alkanone moiety of DHSA was very similar in the two binding modes, suggesting that this interaction is a determinant in the initial substrate-binding event. These data provide insights into the binding of catechols by extradiol dioxygenases and facilitate inhibitor design

Systemic Stimulation of TLR2 Impairs Neonatal Mouse Brain Development

Background: Inflammation is associated with perinatal brain injury but the underlying mechanisms are not completely characterized. Stimulation of Toll-like receptors (TLRs) through specific agonists induces inflammatory responses that trigger both innate and adaptive immune responses. The impact of engagement of TLR2 signaling pathways on the neonatal brain is still unclear. The aim of this study was to investigate the potential effect of a TLR2 agonist on neonatal brain development. Methodology/Principal Findings: Mice were injected intraperitoneally (i.p.) once a day from postnatal day (PND) 3 to PND11 with endotoxin-free saline, a TLR2 agonist Pam$_{3}$CSK$_{4}$ (5 mg/kg) or Lipopolysaccharide (LPS, 0.3 mg/kg). Pups were sacrificed at PND12 or PND53 and brain, spleen and liver were collected and weighed. Brain sections were stained for brain injury markers. Long-term effects on memory function were assessed using the Trace Fear Conditioning test at PND50. After 9 days of Pam$_{3}$CSK$_{4}$ administration, we found a decreased volume of cerebral gray matter, white matter in the forebrain and cerebellar molecular layer that was accompanied by an increase in spleen and liver weight at PND12. Such effects were not observed in Pam$_{3}$CSK$_{4}$-treated TLR 2-deficient mice. Pam$_{3}$CSK$_{4}$-treated mice also displayed decreased hippocampus neuronal density, and increased cerebral microglia density, while there was no effect on caspase-3 or general cell proliferation at PND12. Significantly elevated levels of IL-1β, IL-6, KC, and MCP-1 were detected after the first Pam$_{3}$CSK$_{4}$ injection in brain homogenates of PND3 mice. Pam$_{3}$CSK$_{4}$administration did not affect long-term memory function nor the volume of gray or white matter. Conclusions/Significance: Repeated systemic exposure to the TLR2 agonist Pam$_{3}$CSK$_{4}$ can have a short-term negative impact on the neonatal mouse brain

Native human adipose stromal cells: localization, morphology and phenotype

International audienceObjectives:Beside having roles in energy homeostasis and endocrine modulation, adipose tissue (AT) is now considered a promising source of mesenchymal stromal cells (adipose-derived stromal cells or ASCs) for regenerative medicine. Despite numerous studies on cultured ASCs, native human ASCs are rarely investigated. Indeed, the phenotype of ASCs in their native state, their localization within AT and comparison with bone marrow-derived mesenchymal stromal cells (BM-MSCs) has been poorly investigated.Design:To address these issues, the stroma vascular fraction (SVF) of human AT was extracted and native cell subtypes were isolated by immunoselection to study their clonogenic potential in culture. Immunohistology on samples of human AT in combination with reconstruction of confocal sections were performed in order to localize ASCs.Results:Compared with BM-MNCs, all native ASCs were found in the CD34(+) cell fraction of the AT-SVF. Native ASCs expressed classical mesenchymal markers described for BM-MSCs. Interestingly, CD34 expression decreased during ASC cell culture and was negatively correlated with cell proliferation rate. Immunohistological analysis revealed that native ASCs exhibited specific morphological features with protrusions. They were found scattered in AT stroma and did not express in vivo pericytic markers such as NG2, CD140b or alpha-smooth muscle actin, which appeared during the culture process. Finally, ASCs spontaneous commitment to adipocytic lineage was enhanced in AT from obese humans.Conclusions:The use of complementary methodological approaches to study native human ASCs revealed their immunophenotype, their specific morphology, their location within AT and their stemness. Furthermore, our data strongly suggest that human ASCs participate in adipogenesis during AT development.International Journal of Obesity advance online publication, 25 January 2011; doi:10.1038/ijo.2010.269

The dynamic cilium in human diseases

Cilia are specialized organelles protruding from the cell surface of almost all mammalian cells. They consist of a basal body, composed of two centrioles, and a protruding body, named the axoneme. Although the basic structure of all cilia is the same, numerous differences emerge in different cell types, suggesting diverse functions. In recent years many studies have elucidated the function of 9+0 primary cilia. The primary cilium acts as an antenna for the cell, and several important pathways such as Hedgehog, Wnt and planar cell polarity (PCP) are transduced through it. Many studies on animal models have revealed that during embryogenesis the primary cilium has an essential role in defining the correct patterning of the body. Cilia are composed of hundreds of proteins and the impairment or dysfunction of one protein alone can cause complete loss of cilia or the formation of abnormal cilia. Mutations in ciliary proteins cause ciliopathies which can affect many organs at different levels of severity and are characterized by a wide spectrum of phenotypes. Ciliary proteins can be mutated in more than one ciliopathy, suggesting an interaction between proteins. To date, little is known about the role of primary cilia in adult life and it is tempting to speculate about their role in the maintenance of adult organs. The state of the art in primary cilia studies reveals a very intricate role. Analysis of cilia-related pathways and of the different clinical phenotypes of ciliopathies helps to shed light on the function of these sophisticated organelles. The aim of this review is to evaluate the recent advances in cilia function and the molecular mechanisms at the basis of their activity