Focus on cutaneous and uveal melanoma specificities.

Cutaneous melanoma (CM) and uveal melanoma (UM) derive from cutaneous and uveal melanocytes that share the same embryonic origin and display the same cellular function. However, the etiopathogenesis and biological behaviors of these melanomas are very different. CM and UM display distinct landscapes of genetic alterations and show different metastatic routes and tropisms. Hence, therapeutic improvements achieved in the last few years for the treatment of CM have failed to ameliorate the clinical outcomes of patients with UM. The scope of this review is to discuss the differences in tumorigenic processes (etiologic factors and genetic alterations) and tumor biology (gene expression and signaling pathways) between CM and UM. We develop hypotheses to explain these differences, which might provide important clues for research avenues and the identification of actionable vulnerabilities suitable for the development of new therapeutic strategies for metastatic UM.

RNA interaction format: a general data format for RNA interactions.

SUMMARY: RNA molecules play crucial roles in various biological processes. They mediate their function mainly by interacting with other RNAs or proteins. At present, information about these interactions is distributed over different resources, often providing the data in simple tab-delimited formats that differ between the databases. There is no standardized data format that can capture the nature of all these different interactions in detail. AVAILABILITY AND IMPLEMENTATION: Here, we propose the RNA interaction format (RIF) for the detailed representation of RNA-RNA and RNA-Protein interactions and provide reference implementations in C/C++, Python, and JavaScript. RIF is released under licence GNU General Public License version 3 (GNU GPLv3) and is available on https://github.com/RNABioInfo/rna-interaction-format.

An in vitro method for inducing titan cells reveals novel features of yeast-to-titan switching in the human fungal pathogen Cryptococcus gattii.

Cryptococcosis is a potentially lethal fungal infection of humans caused by organisms within the Cryptococcus neoformans/gattii species complex. Whilst C. neoformans is a relatively common pathogen of immunocompromised individuals, C. gattii is capable of acting as a primary pathogen of immunocompetent individuals. Within the host, both species undergo morphogenesis to form titan cells: exceptionally large cells that are critical for disease establishment. To date, the induction, defining attributes, and underlying mechanism of titanisation have been mainly characterized in C. neoformans. Here, we report the serendipitous discovery of a simple and robust protocol for in vitro induction of titan cells in C. gattii. Using this in vitro approach, we reveal a remarkably high capacity for titanisation within C. gattii, especially in strains associated with the Pacific Northwest Outbreak, and characterise strain-specific differences within the clade. In particular, this approach demonstrates for the first time that cell size changes, DNA amplification, and budding are not always synchronous during titanisation. Interestingly, however, exhibition of these cell cycle phenotypes was correlated with genes associated with cell cycle progression including CDC11, CLN1, BUB2, and MCM6. Finally, our findings reveal exogenous p-Aminobenzoic acid to be a key inducer of titanisation in this organism. Consequently, this approach offers significant opportunities for future exploration of the underlying mechanism of titanisation in this genus.

Correction: An in vitro method for inducing titan cells reveals novel features of yeast-to-titan switching in the human fungal pathogen Cryptococcus gattii.

[This corrects the article DOI: 10.1371/journal.ppat.1010321.].

Rapid screening and detection of inter-type viral recombinants using phylo-k-mers.

MOTIVATION: Novel recombinant viruses may have important medical and evolutionary significance, as they sometimes display new traits not present in the parental strains. This is particularly concerning when the new viruses combine fragments coming from phylogenetically distinct viral types. Here, we consider the task of screening large collections of sequences for such novel recombinants. A number of methods already exist for this task. However, these methods rely on complex models and heavy computations that are not always practical for a quick scan of a large number of sequences. RESULTS: We have developed SHERPAS, a new program to detect novel recombinants and provide a first estimate of their parental composition. Our approach is based on the precomputation of a large database of 'phylogenetically-informed k-mers', an idea recently introduced in the context of phylogenetic placement in metagenomics. Our experiments show that SHERPAS is hundreds to thousands of times faster than existing software, and enables the analysis of thousands of whole genomes, or long-sequencing reads, within minutes or seconds, and with limited loss of accuracy. AVAILABILITY AND IMPLEMENTATION: The source code is freely available for download at https://github.com/phylo42/sherpas. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Generation and characterization of a tamoxifen-inducible Vsx1-CreERT2 line to target V2 interneurons in the mouse developing spinal cord.

In the spinal cord, ventral interneurons regulate the activity of motor neurons, thereby controlling motor activities including locomotion. Interneurons arise during embryonic development from distinct progenitor domains orderly distributed along the dorso-ventral axis of the neural tube. The p2 progenitor domain generates at least five V2 interneuron populations. However, identification and characterization of all V2 populations remain currently incomplete and the mechanisms that control their development remain only partly understood. In this study, we report the generation of a Vsx1-CreERT2 BAC transgenic mouse line that drives CreERT2 recombinase expression mimicking endogenous Vsx1 expression pattern in the developing spinal cord. We showed that the Vsx1-CreERT2 transgene can mediate recombination in V2 precursors with a high efficacy and specificity. Lineage tracing demonstrated that all the V2 interneurons in the mouse developing spinal cord derive from cells expressing Vsx1. Finally, we confirmed that V2 precursors generate additional V2 populations that are not characterized yet. Thus, the Vsx1-CreERT2 line described here is a useful genetic tool for lineage tracing and for functional studies of the mouse spinal V2 interneurons.

ITGBL1 is a new immunomodulator that favors development of melanoma tumors by inhibiting natural killer cells cytotoxicity.

Resistances to immunotherapies remains a major hurdle towards a cure for melanoma in numerous patients. An increase in the mesenchymal phenotype and a loss of differentiation have been clearly associated with resistance to targeted therapies. Similar phenotypes have been more recently also linked to resistance to immune checkpoint therapies. We demonstrated here that the loss of MIcrophthalmia associated Transcription Factor (MITF), a pivotal player in melanocyte differentiation, favors the escape of melanoma cells from the immune system. We identified Integrin beta-like protein 1 (ITGBL1), a secreted protein, upregulated in anti-PD1 resistant patients and in MITFlow melanoma cells, as the key immunomodulator. ITGBL1 inhibited immune cell cytotoxicity against melanoma cells by inhibiting NK cells cytotoxicity and counteracting beneficial effects of anti-PD1 treatment, both in vitro and in vivo. Mechanistically, MITF inhibited RUNX2, an activator of ITGBL1 transcription. Interestingly, VitaminD3, an inhibitor of RUNX2, improved melanoma cells to death by immune cells. In conclusion, our data suggest that inhibition of ITGBL1 might improve melanoma response to immunotherapies.

The Gp1ba-Cre transgenic mouse: a new model to delineate platelet and leukocyte functions.

Conditional knockout (KO) mouse models are invaluable for elucidating the physiological roles of platelets. The Platelet factor 4-Cre recombinase (Pf4-Cre) transgenic mouse is the current model of choice for generating megakaryocyte/platelet-specific KO mice. Platelets and leukocytes work closely together in a wide range of disease settings, yet the specific contribution of platelets to these processes remains unclear. This is partially a result of the Pf4-Cre transgene being expressed in a variety of leukocyte populations. To overcome this issue, we developed a Gp1ba-Cre transgenic mouse strain in which Cre expression is driven by the endogenous Gp1ba locus. By crossing Gp1ba-Cre and Pf4-Cre mice to the mT/mG dual-fluorescence reporter mouse and performing a head-to-head comparison, we demonstrate more stringent megakaryocyte lineage-specific expression of the Gp1ba-Cre transgene. Broader tissue expression was observed with the Pf4-Cre transgene, leading to recombination in many hematopoietic lineages, including monocytes, macrophages, granulocytes, and dendritic and B and T cells. Direct comparison of phenotypes of Csk, Shp1, or CD148 conditional KO mice generated using either the Gp1ba-Cre or Pf4-Cre strains revealed similar platelet phenotypes. However, additional inflammatory and immunological anomalies were observed in Pf4-Cre-generated KO mice as a result of nonspecific deletion in other hematopoietic lineages. By excluding leukocyte contributions to phenotypes, the Gp1ba-Cre mouse will advance our understanding of the role of platelets in inflammation and other pathophysiological processes in which platelet-leukocyte interactions are involved.

Rank signaling links the development of invariant γδ T cell progenitors and Aire(+) medullary epithelium.

The thymic medulla provides a specialized microenvironment for the negative selection of T cells, with the presence of autoimmune regulator (Aire)-expressing medullary thymic epithelial cells (mTECs) during the embryonic-neonatal period being both necessary and sufficient to establish long-lasting tolerance. Here we showed that emergence of the first cohorts of Aire(+) mTECs at this key developmental stage, prior to αß T cell repertoire selection, was jointly directed by Rankl(+) lymphoid tissue inducer cells and invariant Vγ5(+) dendritic epidermal T cell (DETC) progenitors that are the first thymocytes to express the products of gene rearrangement. In turn, generation of Aire(+) mTECs then fostered Skint-1-dependent, but Aire-independent, DETC progenitor maturation and the emergence of an invariant DETC repertoire. Hence, our data attributed a functional importance to the temporal development of Vγ5(+) γδ T cells during thymus medulla formation for αß T cell tolerance induction and demonstrated a Rank-mediated reciprocal link between DETC and Aire(+) mTEC maturation.

Adipose tissue-derived stem cells protect the primordial follicle pool from both direct follicle death and abnormal activation after ovarian tissue transplantation.

PURPOSE: To investigate whether adipose tissue-derived stem cells (ASCs) protect the primordial follicle pool, not only by decreasing direct follicle loss but also by modulating follicle activation pathways. METHODS: Twenty nude mice were grafted with frozen-thawed human ovarian tissue from 5 patients. Ten mice underwent standard ovarian tissue transplantation (OT group), while the remaining ten were transplanted with ASCs and ovarian tissue (2-step/ASCs+OT group). Ovarian grafts were retrieved on days 3 (n = 5) and 10 (n = 5). Analyses included histology (follicle count and classification), immunohistochemistry (c-caspase-3 for apoptosis and LC3B for autophagy), and immunofluorescence (FOXO1 for PI3K/Akt activation and YAP for Hippo pathway disruption). Subcellular localization was determined in primordial follicles on high-resolution images using structured illumination microscopy. RESULTS: The ASCs+OT group showed significantly higher follicle density than the OT group (p = 0.01). Significantly increased follicle atresia (p < 0.001) and apoptosis (p = 0.001) were observed only in the OT group. In primordial follicles, there was a significant shift in FOXO1 to a cytoplasmic localization in the OT group on days 3 (p = 0.01) and 10 (p = 0.03), indicating follicle activation, while the ASCs+OT group and non-grafted controls maintained nuclear localization, indicating quiescence. Hippo pathway disruption was encountered in primordial follicles irrespective of transplantation, with nuclear YAP localized in their granulosa cells. CONCLUSION: We demonstrate that ASCs exert positive effects on the ovarian reserve, not only by protecting primordial follicles from direct death but also by maintaining their quiescence through modulation of the PI3K/Akt pathway.

Accurate and live peroxisome biogenesis evaluation achieved by lentiviral expression of a green fluorescent protein fused to a peroxisome targeting signal 1.

Peroxisomes are ubiquitous organelles formed by peroxisome biogenesis (PB). During PB, peroxisomal matrix proteins harboring a peroxisome targeting signal (PTS) are imported inside peroxisomes by peroxins, encoded by PEX genes. Genetic alterations in PEX genes lead to a spectrum of incurable diseases called Zellweger spectrum disorders (ZSD). In vitro drug screening is part of the quest for a cure in ZSD by restoring PB in ZSD cell models. In vitro PB evaluation is commonly achieved by immunofluorescent staining or transient peroxisome fluorescent reporter expression. Both techniques have several drawbacks (cost, time-consuming technique, etc.) which we overcame by developing a third-generation lentiviral transfer plasmid expressing an enhanced green fluorescent protein fused to PTS1 (eGFP-PTS1). By eGFP-PTS1 lentiviral transduction, we quantified PB and peroxisome motility in ZSD and control mouse and human fibroblasts. We confirmed the stable eGFP-PTS1 expression along cell passages. eGFP signal analysis distinguished ZSD from control eGFP-PTS1-transduced cells. Live eGFP-PTS1 transduced cells imaging quantified peroxisomes motility. In conclusion, we developed a lentiviral transfer plasmid allowing stable eGFP-PTS1 expression to study PB (deposited on Addgene: #133282). This tool meets the needs for in vitro PB evaluation and ZSD drug discovery.

Hip fractures and characteristics of living area: a fine-scale spatial analysis in France.

We investigated the association between hip fracture incidence and living area characteristics in France. The spatial distribution of hip fracture incidence was heterogeneous and there was a significant relationship between social deprivation, urbanization, health access, and hip fracture risk. INTRODUCTION: Several studies have shown great disparities in spatial repartition of hip fractures (HF). The aim of the study was to analyze the association between HF incidence and characteristics of the living area. METHODS: All patients aged 50 or older, living in France, who were hospitalized for HF between 2012 and 2014 were included, using the French national hospital discharge database. Standardized incidence ratio (SIR) was calculated for each spatial unit and adjusted on age and sex. An ecological regression was performed to analyze the association between HF standardized incidence and ecological variables. We adjusted the model for neighborhood spatial structure. We used three variables to characterize the living areas: a deprivation index (French-EDI); healthcare access (French standardized index); land use (percentage of artificialized surfaces). RESULTS: A total of 236,328 HF were recorded in the French hospital national database, leading to an annual HF incidence of 333/100,000. The spatial analysis revealed geographical variations of HF incidence with SIR varying from 0.67 (0.52; 0.85) to 1.45 (1.23; 1.70). There was a significant association between HF incidence rates and (1) French-EDI (trend p = 0.0023); (2) general practitioner and nurse accessibility (trend p = 0.0232 and p = 0.0129, respectively); (3) percentage of artificialized surfaces (p < 0.0001). CONCLUSION: The characteristics of the living area are associated with significant differences in the risk of hip fracture of older people.

OSF-Builder: A New Tool for Constructing and Representing Evolutionary Histories Involving Introgression.

Introgression is an evolutionary process which provides an important source of innovation for evolution. Although various methods have been used to detect introgression, very few methods are currently available for constructing evolutionary histories involving introgression. In this article, we propose a new method for constructing such evolutionary histories whose starting point is a species forest (consisting of a collection of lineage trees, usually arising as a collection of clades or monophyletic groups in a species tree), and a gene tree for a specific allele of interest, or allele tree for short. Our method is based on representing introgression in terms of a certain "overlay" of the allele tree over the lineage trees, called an overlaid species forest (OSF). OSFs are similar to phylogenetic networks although a key difference is that they typically have multiple roots because each monophyletic group in the species tree has a different point of origin. Employing a new model for introgression, we derive an efficient algorithm for building OSFs called OSF-Builder that is guaranteed to return an optimal OSF in the sense that the number of potential introgression events is minimized. As well as using simulations to assess the performance of OSF-Builder, we illustrate its use on a butterfly data set in which introgression has been previously inferred. The OSF-Builder software is available for download from https://www.uea.ac.uk/computing/software/OSF-Builder.

α1AMP-Activated Protein Kinase Protects against Lipopolysaccharide-Induced Endothelial Barrier Disruption via Junctional Reinforcement and Activation of the p38 MAPK/HSP27 Pathway.

Vascular hyperpermeability is a determinant factor in the pathophysiology of sepsis. While, AMP-activated protein kinase (AMPK) is known to play a role in maintaining endothelial barrier function in this condition. Therefore, we investigated the underlying molecular mechanisms of this protective effect. α1AMPK expression and/or activity was modulated in human dermal microvascular endothelial cells using either α1AMPK-targeting small interfering RNA or the direct pharmacological AMPK activator 991, prior to lipopolysaccharide (LPS) treatment. Western blotting was used to analyze the expression and/or phosphorylation of proteins that compose cellular junctions (zonula occludens-1 (ZO-1), vascular endothelial cadherin (VE-Cad), connexin 43 (Cx43)) or that regulate actin cytoskeleton (p38 MAPK; heat shock protein 27 (HSP27)). Functional endothelial permeability was assessed by in vitro Transwell assays, and quantification of cellular junctions in the plasma membrane was assessed by immunofluorescence. Actin cytoskeleton remodeling was evaluated through actin fluorescent staining. We consequently demonstrate that α1AMPK deficiency is associated with reduced expression of CX43, ZO-1, and VE-Cad, and that the drastic loss of CX43 is likely responsible for the subsequent decreased expression and localization of ZO-1 and VE-Cad in the plasma membrane. Moreover, α1AMPK activation by 991 protects against LPS-induced endothelial barrier disruption by reinforcing cortical actin cytoskeleton. This is due to a mechanism that involves the phosphorylation of p38 MAPK and HSP27, which is nonetheless independent of the small GTPase Rac1. This results in a drastic decrease of LPS-induced hyperpermeability. We conclude that α1AMPK activators that are suitable for clinical use may provide a specific therapeutic intervention that limits sepsis-induced vascular leakage.

Energy-Chirp Compensation in a Laser Wakefield Accelerator.

The energy spread in laser wakefield accelerators is primarily limited by the energy chirp introduced during the injection and acceleration processes. Here, we propose the use of longitudinal density tailoring to reduce the beam chirp at the end of the accelerator. Experimental data sustained by quasi-3D particle-in-cell simulations show that broadband electron beams can be converted to quasimonoenergetic beams of ≤10% energy spread while maintaining a high charge of more than 120 pC. In the linear and quasilinear regimes of wakefield acceleration, the method could provide even lower, subpercent level, energy spread.

Ulipristal acetate for the management of large uterine fibroids associated with heavy bleeding: a review.

Ulipristal acetate (UPA), a selective progesterone receptor modulator (SPRM), offers new therapeutic options for the clinical management of large uterine fibroids associated with heavy menstrual bleeding or with other moderate or severe symptoms (bulk symptoms, pelvic pain, decreased quality of life). SPRM are synthetic compounds that exert an agonist or antagonist effect on target tissues by their binding to progesterone receptors. UPA reduces fibroid size, controls bleeding in a high percentage of women and significantly improves quality of life. The present review aims to provide insights into UPA indications and its mechanism of action.

Gene expression changes in uterine myomas in response to ulipristal acetate treatment.

RESEARCH QUESTION: Does ulipristal acetate (UPA) modify the expression of genes related to apoptosis or the extracellular matrix in uterine myomas and are any modifications associated with a clinical response? DESIGN: Targeted analysis of 176 apoptosis- or extracellular-matrix-related genes was conducted using polymerase chain reaction (PCR) arrays. Relevant results were validated by quantitative PCR. Four groups were established: responsive short-term (one course, n = 9), responsive long-term (two to four courses, n = 9), non-responsive (n = 9), and the control group who was not given any hormone therapy (n = 9). The clinical response was monitored by medical imagery and considered significant when volume reduction was greater than 25%. RESULTS: Compared with untreated myomas, significant changes in expression of four genes were found in UPA-treated myomas. Gene expression of integrin subunit beta 4 was repressed by UPA treatment (fold change [FC] = -12.50, P < 0.001, q < 0.001), tenascin-C expression was downregulated in UPA-responsive patients (FC = -2.50, P = 0.010, q = 0.090), survivin was repressed in short-term UPA-responsive tumours (FC = -7.69, P < 0.001, q = 0.010), and catenin delta 2 gene expression was upregulated in non-responsive myomas (FC = +7.36, P < 0.001, q = 0.010). CONCLUSION: This characterization provides the first molecular distinction between myomas responsive or non-responsive to UPA treatment.

Progesterone Receptor Isoforms, Nuclear Corepressor-1 and Steroid Receptor Coactivator-1 and B-Cell Lymphoma 2 and Akt and Akt Phosphorylation Status in Uterine Myomas after Ulipristal Acetate Treatment: A Systematic Immunohistochemical Evaluation.

OBJECTIVE: To investigate whether ulipristal acetate (UPA) treatment modifies the expression of progesterone receptor (PR), its nuclear cofactors steroid receptor coactivator-1 (SRC1) and nuclear corepressor-1 (NCoR1), prosurvival factor B-cell lymphoma 2 (Bcl-2), and Akt in uterine myomas. PATIENTS: Prospective study of 59 women with symptomatic myomas undergoing myomectomy. Forty-two patients were treated preoperatively with UPA; the remaining 17 were not and they served as controls. METHOD: Tissue microarrays were obtained from surgical specimens and immunohistochemistry was performed. Blinded quantification of expression of PR (PR-A vs. PR-B), coactivator SRC1 and corepressor NCoR1, and prosurvival factor Bcl-2, and Akt and evaluation of Akt phosphorylation levels. RESULTS: Compared with the control group, UPA does not alter PR protein levels or expression patterns in myomas, and the PR-A/PR-B ratio was similar, as well as cytoplasmic or nuclear expression of cofactors SRC1 and NCoR1. Bcl-2 was heterogeneously expressed throughout the samples and no significant modification in expression was evidenced. No significant difference was found in Akt expression and phosphorylation between treated and untreated myomas. CONCLUSION: This study did not find any significant change in the expression of the studied factors in myomas after UPA exposure. In conclusion, various theories on myomas cells proposed on the basis of in vitro studies are not supported in vivo.

Navigating the highlights of phase III trials: a watchful eye on evidence-based radiotherapy.

BACKGROUND: Phase III randomized controlled trials (RCTs) are the cornerstone of evidence-based oncology. However, there is no exhaustive review describing the radiotherapy RTCs characteristics. The objective of the present study was to describe features of all phase III RCTs including at least a radiation therapy. METHODS AND MATERIALS: Requests were performed in the Medline database (via PubMed). The latest update was performed in April 2016, using the following MESH terms: 'clinical trials: phase III as topic', 'radiotherapy', 'brachytherapy', as keywords. RESULTS: A total of 454 phase III RCTs were identified. Studies were mainly based on open (92.1%) multicenter (77.5%) designs, analyzed in intend to treat (67.6%), aiming at proving superiority (91.6%) through overall survival assessment (46.5%). Most frequently studied malignancies were head and neck (21.8%), lung (14.3%) and prostate cancers (9.9%). Patients were mainly recruited with a locally advanced disease (73.7%). Median age was 59 years old. Out of 977 treatment arms, 889 arms experienced radiotherapy, mainly using 3D-conformal radiotherapy (288 arms, 32.4%). Intensity-modulated techniques were tested in 12 arms (1.3%). The intervention was a non-cytotoxic agent addition in 89 studies (19.6%), a radiation dose/fractionation modification in 74 studies (16.3%), a modification of chemotherapy regimen in 63 studies (13.9%), a chemotherapy addition in 63 studies (13.9%) and a radiotherapy addition in 53 trials (11.7%). With a median follow-up of 50 months, acute all-grade and grade 3-5 toxicities were reported in 49.6% and 69.4% of studies, respectively. Radiotherapy technique, follow-up and late toxicities were reported in 60.1%, 74%, and 31.1% of studies, respectively. CONCLUSION: Phase III randomized controlled trials featured severe limitations, since a third did not report radiotherapy technique, follow-up or late toxicities. The fast-paced technological evolution creates a discrepancy between literature and radiotherapy techniques performed in daily-routine, suggesting that phase III methodology needs to be reinvented.

The K+ channel TASK1 modulates ß-adrenergic response in brown adipose tissue through the mineralocorticoid receptor pathway.

Brown adipose tissue (BAT) is essential for adaptive thermogenesis and dissipation of caloric excess through the activity of uncoupling protein (UCP)-1. BAT in humans is of great interest for the treatment of obesity and related diseases. In this study, the expression of Twik-related acid-sensitive K(+) channel (TASK)-1 [a pH-sensitive potassium channel encoded by the potassium channel, 2-pore domain, subfamily K, member 3 (Kcnk3) gene] correlated highly with Ucp1 expression in obese and cold-exposed mice. In addition, Task1-null mice, compared with their controls, became overweight, mainly because of an increase in white adipose tissue mass and BAT whitening. Task1(-/-)-mouse-derived brown adipocytes, compared with wild-type mouse-derived brown adipocytes, displayed an impaired ß3-adrenergic receptor response that was characterized by a decrease in oxygen consumption, Ucp1 expression, and lipolysis. This phenotype was thought to be caused by an exacerbation of mineralocorticoid receptor (MR) signaling, given that it was mimicked by corticoids and reversed by an MR inhibitor. We concluded that the K(+) channel TASK1 controls the thermogenic activity in brown adipocytes through modulation of ß-adrenergic receptor signaling.