Silencing Strategies Research Articles

Disinhibition of Touch-Evoked Itch in a Mouse Model of Psoriasis

Disinhibition of Touch-Evoked Itch in a Mouse Model of Psoriasis

A ditadura e os rastros da repressão no sudeste paraense: desvelando memórias sobre a Casa Azul

Entre 1972 e 1974, o sudeste do Pará, assim como parte do Maranhão e do norte do atual Tocantins, foi palco da Guerrilha do Araguaia, conflito que marcou não somente a história recente do Brasil, como deixou diversas heranças para a região onde ocorrera, incluindo-se um legado autoritário marcado pela repressão a movimentos sociais voltados para a luta pela terra no contexto da redemocratização. Em Marabá, maior município da mesorregião, fora montado durante a Guerrilha estrutura que visava combater os militantes do PCdoB em atuação na região, consistindo na formação de quartéis, bem como no uso de outros órgãos públicos no suporte as ações do Centro de Inteligência do Exército (CIE). Um desses órgãos fora o então denominado Departamento Nacional de Estradas de Rodagem (DNER), cuja sede fora apontada por camponeses e ex-soldados que por ali passaram como um centro de prisão clandestino, onde teriam ocorridos graves violações dos direitos humanos, que teriam atingido guerrilheiros e camponeses da região. Há relatos que apontam que alguns dos desaparecidos políticos no contexto da Guerrilha foram aprisionados, torturados, mortos ou vistos pela última vez no local, chamado à época de Casa Azul. Após o fim dos conflitos, o local teria sido envolvido na Operação Limpeza, como é descrita a ação onde se efetivou o apagamento dos rastros sobre as violações perpetradas durante a Guerrilha por parte de agentes das Forças Armadas. O artigo se propõe a contextualizar o processo de desvelamento de memórias sobre o local, à revelia das estratégias de silenciamento que foram criadas pelos agentes da Ditadura em relação às violações ocorridas na Guerrilha do Araguaia; objetiva, através dos rastros deixados sobre a Casa Azul, colaborar para maior compreensão sobre um dos espaços mais marcantes no que concerne as estratégias repressivas adotadas durante o regime militar no Brasil.

Targeted silencing of genes in polyploids: lessons learned from Brassica juncea-glucosinolate system.

Intron-spliced hairpin RNAi construct targeting the exonic region of BjuMYB28 driven by the native promoter is the best suited strategy for developing viable low glucosinolate lines in polyploid Brassica juncea. Targeted silencing of specific homolog(s) of a multigene family in polyploids through RNA interference (RNAi) is a challenging effort. Indian oilseed mustard (Brassica juncea), a natural allotetraploid, is expected to have 4-6 copies of every Arabidopsis gene ortholog. In the current study, we have attempted to establish the best gene silencing system suitable for BjuMYB28, a transcription factor gene, with the objective of developing low seed glucosinolate linesin B. juncea. After comparing multiple combinations of BjuMYB28 gene homologs, promoters, target regions (exon and 3' UTR) and silencing strategies (RNAi and antisense), we suggest that the intron-spliced hairpin RNAi construct targeting the specific exonic region of the BjuMYB28 gene under the control of native promoter, whose peak activity synchronises with the highest glucosinolate accumulation phase of the plant, is the best suited strategy for developing viable low glucosinolate lines in polyploid B. juncea.

A nonsense mutation in myelin protein zero causes congenital hypomyelination neuropathy through altered P0 membrane targeting and gain of abnormal function.

Protein zero (P0) is the major structural protein in peripheral myelin, and mutations in the Myelin Protein Zero (Mpz) gene produce wide-ranging hereditary neuropathy phenotypes. To gain insight in the mechanisms underlying a particularly severe form, congenital hypomyelination (CH), we targeted mouse Mpz to encode P0Q215X, a nonsense mutation associated with the disease, that we show escapes nonsense mediated decay and is expressed in CH patient nerves. The knock-in mice express low levels of the resulting truncated protein, producing a milder phenotype when compared to patients, allowing to dissect the subtle pathogenic mechanisms occurring in otherwise very compromised peripheral myelin. We find that P0Q215X does not elicit an unfolded protein response, which is a key mechanism for other pathogenic MPZ mutations, but is instead in part aberrantly trafficked to non-myelin plasma membranes and induces defects in radial sorting of axons by Schwann cells. We show that the loss of the C-terminal Tyr-Ala-Met-Leu motif is responsible for P0 mislocalization, as its addition is able to restore correct P0Q215X trafficking in vitro. Lastly, we show that P0Q215X acts through dose-dependent gain of abnormal function, as wild-type P0 is unable to rescue the hypomyelination phenotype. Collectively, these data indicate that alterations at the premyelinating stage, linked to altered targeting of P0, may be responsible for CH, and that different types of gain of abnormal function produce the diverse neuropathy phenotypes associated with MPZ, supporting future allele-specific therapeutic silencing strategies.

ADAM8 localizes to extravillous trophoblasts within the maternal-fetal interface and potentiates trophoblast cell line migration through a β1 integrin-mediated mechanism.

Does A Disintegrin And Metalloproteinase 8 (ADAM8) control extravillous trophoblast (EVT) differentiation and migration in early human placental development? ADAM8 mRNA preferentially localizes to invasive HLA-G-positive trophoblasts, associates with the acquirement of an EVT phenotype and promotes trophoblast migration through a mechanism requiring β1-integrin. Placental establishment in the first trimester of pregnancy requires the differentiation of progenitor trophoblasts into invasive EVTs that produce a diverse repertoire of proteases that facilitate matrix remodeling and activation of signaling pathways important in controlling cell migration. While multiple ADAM proteases, including ADAM8, are highly expressed by invasive trophoblasts, the role of ADAM8 in controlling EVT-related processes is unknown. First trimester placental villi and decidua (6-12 weeks' gestation), primary trophoblasts and trophoblastic cell lines (JEG3, JAR, Bewo, HTR8/SVNeo) were used to examine ADAM8 expression, localization and function. All experiments were performed on at least three independent occasions (n = 3). Placental villi and primary trophoblasts derived from IRB approved first trimester placental (n = 24) and decidual (n = 4) were used to examine ADAM8 localization and expression by in situ RNAScope hybridization, flow cytometry, quantitative PCR and immunoblot analyses. Primary trophoblasts were differentiated into EVT-like cells by plating on fibronectin and were assessed by immunofluorescence microscopy and immunoblot analysis of keratin-7, vimentin, epidermal growth factor receptor (EGFR), HLA-G and ADAM8. ADAM8 function was examined in primary EVTs and trophoblastic cell lines utilizing siRNA-directed silencing and over-expression strategies. Trophoblast migration was assessed using Transwell chambers, cell-matrix binding was tested using fibronectin-adhesion assays, and ADAM8-β1-integrin interactions were determined by immunofluorescence microscopy, co-immunoprecipitation experiments and function-promoting/inhibiting antibodies. Within first trimester placental tissues, ADAM8 preferentially localized to HLA-G+ trophoblasts residing within anchoring columns and decidua. Functional experiments in primary trophoblasts and trophoblastic cell lines show that ADAM8 promotes trophoblast migration through a mechanism independent of intrinsic protease activity. We show that ADAM8 localizes to peri-nuclear and cell-membrane actin-rich structures during cell-matrix attachment and promotes trophoblast binding to fibronectin matrix. Moreover, ADAM8 potentiates β1-integrin activation and promotes cell migration through a mechanism dependent on β1-integrin function. The primary limitation of this study was the use of in vitro experiments in examining ADAM8 function, as well as the implementation of immortalized trophoblastic cell lines. Histological localization of ADAM8 within placental and decidual tissue sections was limited to mRNA level analysis. Further, patient information corresponding to tissues obtained by elective terminations was not available. The novel non-proteolytic pro-migratory role for ADAM8 in controlling trophoblast migration revealed by this study sheds insight into the importance of ADAM8 in EVT biology and placental development. This work was supported by grants from the Natural Sciences and Engineering Research Council of Canada (NSERC-Discovery Grant) and the Canadian Institutes of Health Research (CIHR-Open Operating Grant). There are no conflicts or competing interests. NA.

Silence and resistance: Aboriginal women working within and against the archive

ABSTRACTRepresentation of Aboriginal women in colonial archives has long been linked to strategies of silencing and erasure. These representations still bear considerable weight over the ways in which Aboriginal womanhood is perceived and permitted in Australian society. This essay explores a history of Aboriginal women self-presentating both within and in direct response to colonial archives, and the modes of cultural translation which function to hold archival representations accountable to the realities of Aboriginal womanhood.

Utilizing host endogenous microRNAs to negatively regulate the replication of porcine reproductive and respiratory syndrome virus in MARC-145 cells.

MicroRNAs (miRNAs) contribute to gene regulation at the post-transcriptional level and are capable of mRNA silencing by binding to target sites exhibiting high degrees of complementarity. Therefore, cloning host miRNA-recognition sequences into the genome of RNA viruses represents a rational strategy for manipulating viral replication. Here, we performed deep sequencing to obtain small-RNA (sRNA)-expression profiles from in vitro-cultured MARC-145 cells post infection with porcine reproductive and respiratory syndrome virus (PRRSV) and chose six candidate miRNAs of different abundance (miR-21, miR-140-3p, miR-185, miR-26a, miR-505, and miR-199a) for further study. Based on the full-length cDNA clone p7USC, we constructed a number of PRRSV mutants that provided complementary base-pairing target sites for the miRNAs in 3′ untranslated regions. Our results showed that all low- and moderate- abundant miRNA-target mutants showed similar growth properties, whereas the highest-abundant miRNA-target mutant blocked both viral transcription and replication. Discontinuous mutations in high-abundant miRNA-target sites subsequently recovered viral viability and propagation. These results demonstrated the copy number of endogenous miRNAs and the extent of sRNA complementarity were key factors to silence potential mRNA expression/translation, thereby determining PRRSV viability. Interestingly, the mutant containing miR-140-target sites (v140-t) showed strong suppression of viral replication from P1 to P3 in vitro, as shown by virus titer, plaque morphology, and qRT-PCR assays. To assess genetic stability, sequencing of v140-t (P1, P3, P5 and P10) revealed spontaneous mutations preferentially located among several nucleotides near the 3′ end of the insertion region and corresponding to the “seed region” of miR-140-3p, explaining the induced viral repression and the direction of virus evolution. This approach provided a general silencing strategy for limiting PRRSV replication by endogenous miRNAs in MARC-145 cells.

Abstract 3701: Targeted Bcl2 siRNA delivery using DNA nanoparticles in cancer therapy

Abstract Background: Short interfering RNA (siRNA), highly capable of knockdown of specific genes, has emerged as a promising molecular therapeutic tool in targeted cancer treatment. Nonetheless, only limited success has been achieved in the systematic administration of siRNA. Therefore, the lack of a robust and versatile siRNA delivery system is a critical issue in translating this therapeutic tool for cancer treatment. Recent development in DNA nanotechnology has made programmable DNA nanoparticles (DNPs) a potent drug delivery platform. Rationally designed DNPs have emerged as facile delivery vehicles since their physicochemical properties can be precisely controlled compared to other delivery systems. This study focuses on the development of tumor targeted novel DNP-based siRNA delivery system to knockdown Bcl2 gene, as a targeted cancer therapeutic. Methods: Structural DNA technology was applied to design a library of DNPs with different sizes and shapes. Flow cytometry, confocal imaging, and transmission electron microscopy (TEM) were utilized to study the cellular internalization of DNPs. The efficacy of Bcl2 knockdown by DNP-siBcl2 and the resulting influence on cell growth and progression were assessed in cancer cells (in vitro) and in mice bearing corresponding tumor xenografts (in vivo). Results: The correlation between DNPs design (size, shape, tumor-targeting) and internalization efficiency in different cancer cell lines remains elusive. We investigated the cellular uptake of different sizes and shapes of DNPs with or without aptamer (nucleolin targeting molecule) in multiple human cancer cell lines. The cellular uptake efficiency of DNPs was influenced by targeting ligand, size, shape, and cell line. We observed distinct stages of the internalization process of a gold nanoparticle (AuNP)-tagged DNP, using high-resolution TEM. This study provides detailed understanding of cellular uptake and intracellular trafficking of DNPs in cancer cells. Anti-cancer therapeutic potential of siBcl2 delivered by DNPs were studied in mice bearing DMS53 tumor xenografts. Significant tumor growth inhibition was observed in mice treated with DNPs-siBcl2 (5 mg/kg or 10 mg/kg, IV) relative to vehicle alone and naked siBcl2. No toxicity was observed by pathologic examination of lung, liver, kidney, heart, brain, or spleen. Conclusions: Our study offers new insights for future optimization of DNP-based drug delivery systems for cancer therapy. Our novel aptamer conjugated DNP formulations demonstrated substantial cellular internalization. Targeting Bcl2 and its downstream signaling intermediaries reduced cellular growth. We validated the strategy of silencing of Bcl2 using DNPs in order to inhibit cancer progression in vivo. We believe that the DNPs and methodologies developed in this project will be applicable to knockdown of Bcl2 and other gene targets and may be an excellent platform to apply in future anti-cancer therapy. Citation Format: Mohammad Aminur Rahman, Pengfei Wang, Dongsheng Wang, Sreenivas Nannapaneni, Selwyn J. Hurwitz, Zhuo G. Chen, Yonggang Ke, Dong M. Shin. Targeted Bcl2 siRNA delivery using DNA nanoparticles in cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3701.

Narratives and the romantic genre in IR: dominant and marginalized stories of Arab Rebellion in Libya

The article shows how the rebellion against Gaddafi in Libya in 2011 was romanticized in the British newspaper media and among the political elite. Combining insights from literary studies and employing a method of narrative analysis which focuses on the elements of setting, characterization and emplotment, it illustrates the process of narrative romanticization by emphasizing story elements which constitute the rebellion in an emotional setting in which the rebel is characterized as a young and brave underdog fighting against a brutal and oppressive regime for an ideal such as democracy, freedom and a better future. While romantic narratives were dominant in the discourse on Libya at the time, other less positive narratives which for example emphasize human right violations by rebels were marginalized through a strategy of silencing, denial, ridicule and justification. While the dominance of romantic narratives of rebellion aided the legitimation of British military intervention, the marginalization of negative counter-narratives contributed to the ignorance of extremism and set a bad precedent for the role of human rights in post-conflict Libya.

CRISPR/Cas13a targeting of RNA virus in plants.

This approach is quite promising to control plant viral diseases and create synthetic networks to better understand the structure/function relationship in RNA and proteins. Plant viruses are obligate intracellular parasites which causes enormous losses in crop yield worldwide. These viruses replicate into infected cells by highjacking host cellular machinery. Over the last two decades, diverse approaches such as conventional breeding, transgenic approach and gene silencing strategies have been used to control RNA viruses, but escaped due to high rate of mutation. Recently, a novel CRISPR enzyme, called Cas13a, has been used engineered to confer RNA viruses resistance in plants. Here, we summarize the recent breakthrough of CRISPR/Cas13a and its applications in RNA biology.

Experimental Analysis of Mimivirus Translation Initiation Factor 4a Reveals Its Importance in Viral Protein Translation during Infection of Acanthamoeba polyphaga.

The Acanthamoeba polyphaga mimivirus is the first giant virus ever described, with a 1.2-Mb genome which encodes 979 proteins, including central components of the translation apparatus. One of these proteins, R458, was predicted to initiate translation, although its specific role remains unknown. We silenced the R458 gene using small interfering RNA (siRNA) and compared levels of viral fitness and protein expression in silenced versus wild-type mimivirus. Silencing decreased the growth rate, but viral particle production at the end of the viral cycle was unaffected. A comparative proteomic approach using two-dimensional difference-in-gel electrophoresis (2D-DIGE) revealed deregulation of the expression of 32 proteins in silenced mimivirus, which were defined as up- or downregulated. Besides revealing proteins with unknown functions, silencing R458 also revealed deregulation in proteins associated with viral particle structures, transcriptional machinery, oxidative pathways, modification of proteins/lipids, and DNA topology/repair. Most of these proteins belong to genes transcribed at the end of the viral cycle. Overall, our data suggest that the R458 protein regulates the expression of mimivirus proteins and, thus, that mimivirus translational proteins may not be strictly redundant in relation to those from the amoeba host. As is the case for eukaryotic initiation factor 4a (eIF4a), the R458 protein is the prototypical member of the ATP-dependent DEAD box RNA helicase mechanism. We suggest that the R458 protein is required to unwind the secondary structures at the 5' ends of mRNAs and to bind the mRNA to the ribosome, making it possible to scan for the start codon. These data are the first experimental evidence of mimivirus translation-related genes, predicted to initiate protein biosynthesis.IMPORTANCE The presence in the genome of a mimivirus of genes coding for many translational processes, with the exception of ribosome constituents, has been the subject of debate since its discovery in 2003. In this work, we focused on the R458 mimivirus gene, predicted to initiate protein biosynthesis. After silencing was performed, we observed that it has no major effect on mimivirus multiplication but that it affects protein expression and fitness. This suggests that it is effectively used by mimivirus during its developmental cycle. Until large-scale genetic manipulation of giant viruses becomes possible, the silencing strategy used here on mimivirus translation-related factors will open the way to understanding the functions of these translational genes.

Genome-wide profiling of sRNAs in the Verticillium dahliae-infected Arabidopsis roots

ABSTRACTSmall RNAs (sRNAs, including small interfering RNAs [siRNAs] and micro RNAs [miRNAs]) are key mediators of RNA silencing (or RNA interference), which play important roles in plant development and response to biotic and abiotic stimulation. Verticillium wilt is a plant vascular disease caused by the soil-borne fungal pathogens, such as Verticillium dahliae. We previously reported that V. dahliae infection increased two plant endogenous miRNAs that were exported to fungal cell to silence virulence genes. To investigate plant sRNAs in genome-wide response to V. dahliae infection, in this study, we constructed two sRNA libraries from Arabidopsis roots with and without V. dahliae infection, respectively. In total, 31 conserved miRNAs were found to be differentially expressed during the early stage of infection with V. dahliae using sRNA sequencing. Among these, the expression levels of miR160, miR164, miR166, miR167, miR390 and miR156h were confirmed by northern blot. Reverse transcription quantitative real time polymerase chain reaction results showed that the induction of miRNAs (miR160, miR164, miR166 and miR167) upon V. dahliae infection downregulated the expression of their targeted genes (ARF10, NAC1, PHV and ARF6), respectively. In addition, we identified specific phased siRNAs generated from distinct regions of two libraries. Profiling of these miRNAs and sRNAs lay the foundation for further understanding and utilising the host-induced gene silencing strategy to control plant vascular pathogens.

“Disgusting Details Which Are Best Forgotten”: Disclosures of Child Sexual Abuse in Twentieth-Century Britain

For those who by the end of the twentieth century came to be termed “survivors” of child sexual abuse, different genres and forms have been available to narrate and evaluate that abuse. This article explores the reception and practical results of such disclosures: the unpredictable effects of telling, and the strategies of containment, silencing, or disbelief that greeted disclosures. I make note of the ethical challenges of writing the history of child sexual abuse and conclude that twenty-first-century observers have been too ready to perceive much of the previous century as a period of profound silence in relation to child sexual abuse. At the same time, historical and sociological accounts have also been too ready to claim the final third of the twentieth century as a period of compulsive disclosure and fluency in constructing sexual selves. The history of child sexual abuse reveals significant barriers to disclosure in the 1970s and 1980s, despite new visibility of child sexual abuse in the media and through feminist sexual politics. Attention to such obstacles suggests the need to rethink narratives of “permissive” sexual change to acknowledge more fully the ongoing inequities and hierarchies in sexual candor and voice.

“Disgusting Details Which Are Best Forgotten”: Disclosures of Child Sexual Abuse in Twentieth-Century Britain

AbstractFor those who by the end of the twentieth century came to be termed “survivors” of child sexual abuse, different genres and forms have been available to narrate and evaluate that abuse. This article explores the reception and practical results of such disclosures: the unpredictable effects of telling, and the strategies of containment, silencing, or disbelief that greeted disclosures. I make note of the ethical challenges of writing the history of child sexual abuse and conclude that twenty-first-century observers have been too ready to perceive much of the previous century as a period of profound silence in relation to child sexual abuse. At the same time, historical and sociological accounts have also been too ready to claim the final third of the twentieth century as a period of compulsive disclosure and fluency in constructing sexual selves. The history of child sexual abuse reveals significant barriers to disclosure in the 1970s and 1980s, despite new visibility of child sexual abuse in the media and through feminist sexual politics. Attention to such obstacles suggests the need to rethink narratives of “permissive” sexual change to acknowledge more fully the ongoing inequities and hierarchies in sexual candor and voice.

GPER mediates the angiocrine actions induced by IGF1 through the HIF-1\u03b1/VEGF pathway in the breast tumor microenvironment

BackgroundThe G protein estrogen receptor GPER/GPR30 mediates estrogen action in breast cancer cells as well as in breast cancer-associated fibroblasts (CAFs), which are key components of microenvironment driving tumor progression. GPER is a transcriptional target of hypoxia inducible factor 1 alpha (HIF-1α) and activates VEGF expression and angiogenesis in hypoxic breast tumor microenvironment. Furthermore, IGF1/IGF1R signaling, which has angiogenic effects, has been shown to activate GPER in breast cancer cells.MethodsWe analyzed gene expression data from published studies representing almost 5000 breast cancer patients to investigate whether GPER and IGF1 signaling establish an angiocrine gene signature in breast cancer patients. Next, we used GPER-positive but estrogen receptor (ER)-negative primary CAF cells derived from patient breast tumours and SKBR3 breast cancer cells to investigate the role of GPER in the regulation of VEGF expression and angiogenesis triggered by IGF1. We performed gene expression and promoter studies, western blotting and immunofluorescence analysis, gene silencing strategies and endothelial tube formation assays to evaluate the involvement of the HIF-1α/GPER/VEGF signaling in the biological responses to IGF1.ResultsWe first determined that GPER is co-expressed with IGF1R and with the vessel marker CD34 in human breast tumors (n = 4972). Next, we determined that IGF1/IGF1R signaling engages the ERK1/2 and AKT transduction pathways to induce the expression of HIF-1α and its targets GPER and VEGF. We found that a functional cooperation between HIF-1α and GPER is essential for the transcriptional activation of VEGF induced by IGF1. Finally, using conditioned medium from CAFs and SKBR3 cells stimulated with IGF1, we established that HIF-1α and GPER are both required for VEGF-induced human vascular endothelial cell tube formation.ConclusionsThese findings shed new light on the essential role played by GPER in IGF1/IGF1R signaling that induces breast tumor angiogenesis. Targeting the multifaceted interactions between cancer cells and tumor microenvironment involving both GPCRs and growth factor receptors has potential in future combination anticancer therapies.

RNAi-Mediated Specific Gene Silencing as a Tool for the Discovery of New Drug Targets in Giardia lamblia; Evaluation Using the NADH Oxidase Gene.

The microaerophilic protozoan Giardia lamblia is the agent causing giardiasis, an intestinal parasitosis of worldwide distribution. Different pharmacotherapies have been employed against giardiasis; however, side effects in the host and reports of drug resistant strains generate the need to develop new strategies that identify novel biological targets for drug design. To support this requirement, we have designed and evaluated a vector containing a cassette for the synthesis of double-stranded RNA (dsRNA), which can silence expression of a target gene through the RNA interference (RNAi) pathway. Small silencing RNAs were detected and quantified in transformants expressing dsRNA by a stem-loop RT-qPCR approach. The results showed that, in transformants expressing dsRNA of 100–200 base pairs, the level of NADHox mRNA was reduced by around 30%, concomitant with a decrease in enzyme activity and a reduction in the number of trophozoites with respect to the wild type strain, indicating that NADHox is indeed an important enzyme for Giardia viability. These results suggest that it is possible to induce the G. lamblia RNAi machinery for attenuating the expression of genes encoding proteins of interest. We propose that our silencing strategy can be used to identify new potential drug targets, knocking down genes encoding different structural proteins and enzymes from a wide variety of metabolic pathways.

Bevacizumab-mediated tumor vasculature remodelling improves tumor infiltration and antitumor efficacy of GD2-CAR T cells in a human neuroblastoma preclinical model

ABSTRACTGD2-redirected chimeric antigen receptor (CAR) T lymphocytes represent a promising therapeutic option for immunotherapy of neuroblastoma (NB). However, despite the encouraging therapeutic effects observed in some hematological malignancies, clinical results of CAR T cell immunotherapy in solid tumors are still modest.Tumor driven neo-angiogenesis supports an immunosuppressive microenvironment that influences treatment responses and is amenable to targeting with antiangiogenic drugs. The latter agents promote lymphocyte tumor infiltration by transiently reprogramming tumor vasculature, and may represent a valid combinatorial approach with CAR T cell immunotherapy.In light of these considerations, we investigated the anti-NB activity of GD2-CAR T cells combined with bevacizumab (BEV) in an orthotopic xenograft model of human NB. Two weeks after tumor implantation, mice received BEV or GD2-CAR T cells or both by single intravenous administration. GD2-CAR T cells exerted a significant anti-NB activity only in combination with BEV, even at the lowest concentration tested, which per se did not inhibit tumor growth. When combined with BEV, GD2-CAR T cells massively infiltrated tumor mass where they produced interferon-γ (IFN-γ), which, in turn, induced expression of CXCL10 by NB cells. IFN-γ, and possibly other cytokines, upregulated NB cell expression of PD-L1, while tumor infiltrating GD2-CAR T cells expressed PD-1. Thus, the PD-1/PD-L1 axis can limit the anti-tumor efficacy of the GD2-CAR T cell/BEV association.This study provides a strong rationale for testing the combination of GD2-CAR T cells with BEV in a clinical trial enrolling NB patients. PD-L1 silencing or blocking strategies may further enhance the efficacy of such combination.

Familial amyloid polyneuropathy.

Transthyretin familial amyloid polyneuropathy is the most disabling hereditary polyneuropathy of adult onset because of a point mutation of transthyretin gene. This review updates our knowledge about natural history of the disease, phenotypes, diagnosis tools for small and large fibers involvement, expert's consensus for both symptomatic and asymptomatic follow-up, and treatment's research. Access to TTR gene sequencing permit diagnosis and first reports of the disease in nonendemic countries (EU countries, United States, China, India). Most studies showed a more severe natural history of the neuropathy in nonendemic countries. First European consensus for management has been established. New long-term results allow selection of best candidates for liver transplantation based on phenotype and cardiac involvement. Multimodal evaluation of small fiber neuropathy and resonance magnetic neurography are under development. New results are available for long-term effect of tafamidis in late-onset patients. TTR gene silencing drugs are subject to phase 3 clinical trials. New methods for the evaluation of the disease are being developed. The TTR gene silencing strategy will be available by the end of 2017.

Corrigendum: The Generation of Mouse and Human Huntington Disease iPS Cells Suitable for In vitro Studies on Huntingtin Function.

[This corrects the article on p. 253 in vol. 10, PMID: 28848389.].

The mediation of childbirth: ‘Joyful’ birthing and strategies of silencing on a Facebook discussion group

The mediation of parenting has recently occupied sociologists and media, communication and cultural studies scholars alike. This article locates itself within this developing strand of research, as it explores discourses of intensive motherhood on a Facebook discussion group that provides support and advice for a specific approach to and philosophy of childbirth. Presenting findings from an analysis of main posts and comments made on them, I tease out the brighter and darker sides of the performance of motherhood in anticipation of birth on social media - reading these against discussions about the self-managing, intensive mother who is responsible for making the very best decisions for her child.