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Aim: Identify hotspots and areas of high species richness for Arctic marine mammals. Location: Circumpolar Arctic. Methods: A total of 2115 biologging devices were deployed on marine mammals from 13 species in the Arctic from 2005 to 2019. Getis-Ord Gi* hotspots were calculated based on the number of individuals in grid cells for each species and for phyloge-netic groups (nine pinnipeds, three cetaceans, all species) and areas with high spe-cies richness were identified for summer (Jun-Nov), winter (Dec-May) and the entire year. Seasonal habitat differences among species’ hotspots were investigated using Principal Component Analysis. Results: Hotspots and areas with high species richness occurred within the Arctic continental-shelf seas and within the marginal ice zone, particularly in the “Arctic gateways” of the north Atlantic and Pacific oceans. Summer hotspots were generally found further north than winter hotspots, but there were exceptions to this pattern, including bowhead whales in the Greenland-Barents Seas and species with coastal distributions in Svalbard, Norway and East Greenland. Areas with high species rich-ness generally overlapped high-density hotspots. Large regional and seasonal dif-ferences in habitat features of hotspots were found among species but also within species from different regions. Gap analysis (discrepancy between hotspots and IUCN ranges) identified species and regions where more research is required. Main conclusions: This study identified important areas (and habitat types) for Arctic marine mammals using available biotelemetry data. The results herein serve as a benchmark to measure future distributional shifts. Expanded monitoring and teleme-try studies are needed on Arctic species to understand the impacts of climate change and concomitant ecosystem changes (synergistic effects of multiple stressors). While efforts should be made to fill knowledge gaps, including regional gaps and more com-plete sex and age coverage, hotspots identified herein can inform management ef-forts to mitigate the impacts of human activities and ecological changes, including creation of protected areas.

Environmental change and increasing levels of human activity are threats to marine mammals in the Arctic. Identifying marine mammal hotspots and areas of high species richness are essential to help guide management and conservation efforts. Herein, space use based on biotelemetric tracking devices deployed on 13 species (ringed seal Pusa hispida, bearded seal Erignathus barbatus, harbour seal Phoca vitulina, walrus Odobenus rosmarus, harp seal Pagophilus groenlandicus, hooded seal Cystophora cristata, polar bear Ursus maritimus, bowhead whale Balaena mysticetus, narwhal Monodon monoceros, white whale Delphinapterus leucas, blue whale Balaenoptera musculus, fin whale Balaenoptera physalus and humpback whale Megaptera novaeangliae; total = 585 individuals) in the Greenland and northern Barents Seas between 2005 and 2018 is reported. Getis-Ord Gi* hotspots were calculated for each species as well as all species combined, and areas of high species richness were identified for summer/autumn (Jun-Dec), winter/spring (Jan-May) and the entire year. The marginal ice zone (MIZ) of the Greenland Sea and northern Barents Sea, the waters surrounding the Svalbard Archipelago and a few Northeast Greenland coastal sites were identified as key marine mammal hotspots and areas of high species richness in this region. Individual hotspots identified areas important for most of the tagged animals, such as common resting, nursing, moulting and foraging areas. Location hotspots identified areas heavily used by segments of the tagged populations, including denning areas for polar bears and foraging areas. The hotspots identified herein are also important habitats for seabirds and fishes, and thus conservation and management measures targeting these regions would benefit multiple groups of Arctic animals.

espanolSe revisan los fallos de primera y segunda instancia de un juicio ejecutivo de cobro de impuesto territorial, en que Tesoreria ejecuto y emplazo al ex propietario de un bien raiz, el que fue rematado y adjudicado a un tercero, extendiendose la escritura de remate y practicandose su inscripcion conservatoria. Despues de efectuada la inscripcion, comparecio en el juicio el verdadero dueno del sitio, promoviendo incidente de nulidad de lo obrado por falta de emplazamiento, el que fundo en el hecho de haberse notificado y requerido de pago a un ex propietario. El incidente fue rechazado en ambas instancias. EnglishThe first and second instance rulings of a procedure to enforce collection of the territorial tax are reviewed, in which the Treasury collected taxes against the former owner of a real estate, which was auctioned and awarded to a third party, and in which the auction deeds were issued and registered in the Real Estate Registry in the name of the auctionee. After the registration was performed, the true owner of the real estate appeared before the court, submitting a motion of nullity of the procedure based on the lack of formal summons, founded on the fact that the summons and the request of payment were addressed against the former owner. The motion was rejected in both instances.

In the light of a recent judgement of the Constitutional Court, this comment analyzes the rule of paragraph 4 of article 171 of the Tax Code, which allows the notifi cation to and the request of payment from the debtors in the same real state to which the tax applies for purposes of the territorial tax collection procedure. In particular, it reviews the thesis according to which the executed debtor can be validly summoned in a deserted placed

This article describes the relationship between sea ice and marine mammals around Greenland, the possible effects of climate change and the importance of subsistence hunting in modern Greenland. We also summarize the biology, conservation status and utilization of the polar bear, walrus, seal and whale species of Greenland.

High-content phenotypic screening has become the approach of choice for drug discovery due to its ability to extract drug-specific multi-layered data. In the field of epigenetics, such screening methods have suffered from a lack of tools sensitive to selective epigenetic perturbations. Here we describe a novel approach, Microscopic Imaging of Epigenetic Landscapes (MIEL), which captures the nuclear staining patterns of epigenetic marks and employs machine learning to accurately distinguish between such patterns. We validated the MIEL platform across multiple cells lines and using dose-response curves, to insure the fidelity and robustness of this approach for high content high throughput drug discovery. Focusing on noncytotoxic glioblastoma treatments, we demonstrated that MIEL can identify and classify epigenetically active drugs. Furthermore, we show MIEL was able to accurately rank candidate drugs by their ability to produce desired epigenetic alterations consistent with increased sensitivity to chemotherapeutic agents or with induction of glioblastoma differentiation., eLife digest Each cell contains a complete copy of a person’s genes coded in their DNA. However, for a cell to perform its specific role, it only needs a small fraction of this genetic information. The mechanisms that control which genes a cell is using fall under the umbrella of ‘epigenetics’ (meaning beyond genetics). These mechanisms involve changes in the way that DNA is organized inside the cell nucleus and changes in how accessible different parts of the genome are to various cellular components. DNA is long and fragile so, to maintain its integrity, it is wrapped around protein complexes called histones. Adding chemical modifications to histones is one of the main epigenetic mechanisms that cells use to regulate which genes are turned on and off. Several methods allow researchers to read patterns of histone modification and use this information to derive what state a cell is in and how it might behave. Improving these methods is of particular interest in drug development, where this information could reveal the effects, and side-effects, of new treatments. Unfortunately, existing techniques are costly in both time and money, and they are not well suited to analyzing epigenetic changes caused by the large numbers of compounds tested during drug development. To overcome this barrier, Farhy et al. have developed a new system called ‘Microscopic Imaging of the Epigenetic Landscape’ (MIEL for short). The system allows them to quickly analyze the epigenetic changes caused by each of a large number of different chemical compounds when they are used on cells. MIEL tags different histone modifications by staining each with a different color, and then uses automated microscopy to produce images. It then extracts information from these images using advanced image analysis tools. The changes induced by different drugs can then be compared and categorized using machine learning algorithms. To test the MIEL system, Farhy et al. grew brain cancer cells (derived from human tumors) in the lab, and treated them with compounds that target proteins involved in histone modifications. Using their newly created pipeline, Farhy et al. were able to identify the unique epigenetic changes caused by these compounds, and train the system to correctly predict which type of drug the cells had been treated with. In a different set of experiments Farhy et al. demonstrate the utility of their new pipeline in identifying drugs that induce a set of epigenetic changes associated with a reduced ability to regrow tumors. This new system could help screen thousands of compounds for their epigenetic effects, which could aid the design of new treatments for many diseases.

With the advent of automatic cell imaging and machine learning, high-content phenotypic screening has become the approach of choice for drug discovery because it can extract drug-specific multi-layered data, which could be compared to known profiles. In the field of epigenetics, such screening approaches have suffered from a lack of tools sensitive to selective epigenetic perturbations. Here we describe a novel approach, Microscopic Imaging of Epigenetic Landscapes (MIEL), which captures the nuclear staining patterns of epigenetic marks (e.g., acetylated and methylated histones) and employs machine learning to accurately distinguish between such patterns. We validated the fidelity and robustness of the MIEL platform across multiple cells lines using dose-response curves. We employed MIEL to uncover the mechanism by which bromodomain inhibitors synergize with temozolomide-mediated killing of human glioblastoma lines. To explore alternative, non-cytotoxic, glioblastoma treatment, we screen the Prestwick chemical library and documented the power of MIEL platform to identify epigenetically active drugs and accurately rank them according to their ability to produce epigenetic and transcriptional alterations consistent with the induction of glioblastoma differentiation.

Evaluation of patient’s response to chemotherapeutic drugs is often difficult and time consuming. Skin punch biopsies are easily accessible material that can be used for the evaluation of surrogate biomarkers of a patient’s response to a drug. In this study, we hypothesized that assessment of phosphorylated histone H3 in human skin punch biopsies could be used as a pharmacodynamics biomarker of patient’s response to the kinesin spindle protein inhibitor SCH2047069. To test this hypothesis, we used a human skin histoculture technique that allows culturing intact human skin in the presence of the drug. Human melanoma and skin histocultures were treated with SCH2047069, and the effect of the drug was assessed by increasing histone H3 phosphorylation using immunohistochemistry. Our results demonstrate that SCH2047069 has a significant effect on cell proliferation in human melanoma and skin histoculture and justify using human skin punch biopsies for evaluation of the pharmacodynamic changes induced by SCH2047069. Acronyms: Histone subunit H3 (H3), Kinesin spindle protein (KSP), 5-ethynyl-2’-deoxyuridine (EDU), Dimethyl sulfoxide (DMSO), Formalin-fixed paraffin embedded (FFPE).

Greenland covers > 2 million km2 and, being completely surrounded by water, is the largest island in the world. The seas around Greenland cover an area from the High Arctic in the north to the Subarctic in the south and include the three Large Marine Ecosystems (LME) developed by the US National Oceanic and Atmospheric Administration (NOAA). The water masses are dominated by inflow of cold, nutrient-rich water from the Polar region through the Fram Strait in the east and the Nares Strait and the Canadian Archipelago in west, and by inflow of relatively warm and salty water from the North Atlantic. The Greenland shelf is wide in the southwest and here the combination of fronts of the cold and warm waters creates a very productive ecosystem. Only around 56,000 people live in Greenland, mainly in the central and southern part of West Greenland. There is a long tradition of fishing and hunting, and the commercial fishery is today by far the most important export industry. Two species—Greenland halibut and northern shrimp — form the basis for this industry, while several other species are utilized on a subsistence basis. Marine mammals and seabirds are also utilized for local marketing and on a subsistence basis. Other uses of the marine environment include oil exploration, but so far no discoveries have been made. Contaminant levels of, for example, persistent organic pollutants (POP) in the Greenland marine ecosystem are relatively low compared to more southern latitudes. However, relatively high concentrations are found of some contaminants in the higher trophic levels as a result of biomagnification in the food web. These compounds are of concern because, in some areas, marine mammals and seabirds constitute a significant part of the human diet. However, the threat of contaminants must be weighed against the health benefits of the consumption of traditional food. Climate change in the Arctic has been and will continue to be a main driver of future changes in the Greenland ecosystems, and these changes include increasing water temperatures, decreasing sea-ice coverage, and increasing freshwater discharge to the sea. An example of effects of such changes is the emergence of mackerel in the waters off Southeast Greenland. Another example is the retreat of species that depend on the sea-ice habitat for example, polar bears. Decreasing sea-ice coverage will influence future shipping activities and likely also oil exploration activities, which both increase the risk of causing negative impacts on the environment from increased emissions and oil accidents.

The resistance of Glioblastoma (GBM) to conventional cytotoxic drugs has prompted novel therapeutic strategies, including differentiating tumor propagating cells (TPCs) into less tumorigenic cells using small molecule inducers of TPC differentiation. However, high-throughput screening for such molecules is hampered by the lack of robust markers of GBM differentiation. To obtain a signature of differentiated TPCs, we developed “Microscopic Imaging of Epigenetic Landscapes” (MIEL), which captures patterns of nuclear staining for epigenetic marks to derive feature-fingerprints of individual cells. We confirmed MIEL’s ability to accurately distinguish multiple cell fates and identified a multiparametric epigenetic signature of differentiated TPCs. Critically, we validated epigenetic imaging-based signature using global gene expression thus providing the proof of principle for the MIEL’s ability to select and prioritize small molecules, which induce TPC differentiation.

Marine mammals are top predators in Arctic marine ecosystems and are key to ecosystem functioning. Many Arctic marine mammal species are important resources and hold special cultural significance in Arctic communities. The CBMP (Circumpolar Biodiversity Monitoring Programme) Marine Mammal Expert Network aggregated and reviewed data on the population status and trends of all 11 ice-associated marine mammal Focal Ecosystem Components (FECs) across eight Arctic Marine Areas as well as the state of current monitoring (and research) efforts for these species. Changes taking place in the physical environment in the Arctic due to global warming are affecting marine mammal behaviour, abundance, growth rates, body condition and reproduction, and impacting the resilience of marine mammal populations with concomitant effects on the people who rely on them for subsistence, economic and cultural purposes. Effective marine mammal population monitoring will need improved techniques and application at appropriate geographic scales to measure trends that can be evaluated relative to changes in climate (e.g., sea-ice cover) and human activities (e.g., hunting, shipping, mineral exploration). This presentation will summarize current marine mammal monitoring across the Arctic, the status and trends of FECs, drivers of observed trends, and knowledge and monitoring gaps.

RESUMEN: El objetivo de este trabajo fue escribir, según la literatura científica disponible más actual, los efectos que inducen el uso de dispositivos de avance mandibular, como terapia para el SAHOS, en el sistema temporomandibular de los pacientes. Se realizó una revisión de la literatura más actual (últimos 10 años; 2006-2016) a partir de la búsqueda electrónica en las bases de datos PubMed, TripData Base, Epistemonikos, The Cochrane Library y las revistas especializadas Journal of Clinical Sleep Medicine y SLEEP. Con el uso de las palabras clave: “Mandibular advancement device”, “orthodonthic appliances”, “sleep apnea syndroms”, “sleep apnea obstructive”, “Temporomandibular joints disorder”, los operadores booleanos AND y OR. Se realizó un análisis crítico de la literatura evaluando nivel de evidencia, grado de recomendación y riesgo de sesgo de cada publicación. La búsqueda en las distintas bases de datos arrojó un total de 242 documentos, de los cuales 60 fueron seleccionados por título y abstract. Luego 8 estudios fueron descartados por estar repetidos. Se aplicaron los criterios de inclusión y exclusión quedando un total de 20 artículos; se eliminaron 8 por no responder a la pregunta de investigación y se añadió 1 título mediante la búsqueda manual. Finalmente, se analizaron 13 artículos; 2 revisiones sistemáticas, 2 ensayos clínicos aleatorizados y 6 series de casos. La mayoría de los documentos incluidos concuerda en que los efectos inducidos por los DAM, sobre el complejo temporomandibular son mínimos y reversibles, sin explicitar ningún diagnóstico de TTM en particular. Sin embargo, esta evidencia viene en su mayoría de estudios recomendables, pero no concluyentes. Se necesitan más y mejores estudios para realizar un análisis y abstraer conclusiones más certeras. Estos deben ser homogéneos a la hora de clasificar TTM y definir un protocolo óptimo de avance mandibular.

Summary Epigenetic regulation serves as the basis for stem cell differentiation into distinct cell types, but it is unclear how global epigenetic changes are regulated during this process. Here, we tested the hypothesis that global chromatin organization affects the lineage potential of stem cells and that manipulation of chromatin dynamics influences stem cell function. Using nuclease sensitivity assays, we found a progressive decrease in chromatin digestion among pluripotent embryonic stem cells (ESCs), multipotent hematopoietic stem cells (HSCs), and mature hematopoietic cells. Quantitative high-resolution microscopy revealed that ESCs contain significantly more euchromatin than HSCs, with a further reduction in mature cells. Increased cellular maturation also led to heterochromatin localization to the nuclear periphery. Functionally, prevention of heterochromatin formation by inhibition of the histone methyltransferase G9A resulted in delayed HSC differentiation. Our results demonstrate global chromatin rearrangements during stem cell differentiation and that heterochromatin formation by H3K9 methylation regulates HSC differentiation., Graphical Abstract, Highlights • Nuclease sensitivity decreases progressively with stem cell differentiation • Distinct chromatin architecture is apparent in cells of different lineage potential • Stem cell differentiation leads to perinuclear heterochromatin localization • G9a-mediated heterochromatin formation facilitates stem cell differentiation, Forsberg and colleagues use a variety of approaches to investigate global chromatin structure and architecture in pluripotent, multipotent, and differentiated cells. They show that progressive changes in nuclease sensitivity, chromatin condensation, and heterochromatin localization correlate with the lineage potential of embryonic and adult stem cells and committed cells. Functionally, inhibition of heterochromatin formation delayed hematopoietic stem cell differentiation. These data provide new insights on the epigenetic regulation of stem cell lineage potential and differentiation.

Arctic marine mammals (AMMs) are icons of climate change, largely because of their close association with sea ice. However, neither a circumpolar assessment of AMM status nor a standardized metric of sea ice habitat change is available. We summarized available data on abundance and trend for each AMM species and recognized subpopulation. We also examined species diversity, the extent of human use, and temporal trends in sea ice habitat for 12 regions of the Arctic by calculating the dates of spring sea ice retreat and fall sea ice advance from satellite data (1979–2013). Estimates of AMM abundance varied greatly in quality, and few studies were long enough for trend analysis. Of the AMM subpopulations, 78% (61 of 78) are legally harvested for subsistence purposes. Changes in sea ice phenology have been profound. In all regions except the Bering Sea, the duration of the summer (i.e., reduced ice) period increased by 5–10 weeks and by >20 weeks in the Barents Sea between 1979 and 2013. In light of generally poor data, the importance of human use, and forecasted environmental changes in the 21st century, we recommend the following for effective AMM conservation: maintain and improve comanagement by local, federal, and international partners; recognize spatial and temporal variability in AMM subpopulation response to climate change; implement monitoring programs with clear goals; mitigate cumulative impacts of increased human activity; and recognize the limits of current protected species legislation.

Renal pelvis leiomyomas are infrequent benign tumours. These tumours are more frequent in women, usually asymptomatic and difficult to distinguish from malign kidney masses. A 27-year-old female presented with an asymptomatic renal mass discovered after abdominal ultrasound during routine check-up. Percutaneous renal biopsy was performed and reported urothelial carcinoma. After open nephroureterectomy, histopathological evaluation and immunohistochemistry were positive for Smooth Muscle Actin (SMA), Ki67

Haematopoietic stem cell (HSC) niches provide an environment essential for life-long HSC function. Intense investigation of HSC niches both feed off and drive technology development to increase our capability to assay functionally defined cells with high resolution. A major driving force behind the desire to understand the basic biology of HSC niches is the clear implications for clinical therapies. Here, with particular emphasis on cell type-specific deletion of SCL and CXCL12, we focus on unresolved issues on HSC niches, framed around some very recent advances and novel discoveries on the extrinsic regulation of HSC maintenance. We also provide ideas for possible paths forward, some of which are clearly within reach while others will require both novel tools and vision.

We present a general, entirely PCR-based strategy to construct mRNAs coding for green fluorescent protein (GFP) fusion proteins from a cDNA pool. We exemplify our approach for the chemokine receptor CXCR4. mRNA transfection of the PCR-generated fusion of CXCR4-GFP into K562 cells or primary mesenchymal stem cells (MSCs) resulted in excellent viability (90%) with more than 90% of target cells expressing easily detectable CXCR4-GFP for72 h. The fusion protein was localized in the plasma membrane and was rapidly internalized upon incubation with the CXCR4 ligand stromal cell-derived factor-1 (SDF-1). Transwell migration experiments showed significantly increased migration of CXCR4-GFP mRNA-transfected MSCs toward a gradient of SDF-1, demonstrating that mRNA-mediated chemokine receptor overexpression allows for transient initiation of chemotaxis. The presented strategy to construct a PCR-based fluorescent fusion protein can be generally applied to other genes of interest to study their function by simple overexpression and easy detection in primary cells.

We describe the acoustic behaviour of piscivorous killer whales in Norwegian and Icelandic waters. Whales were assigned to one of three activities (feeding, travelling or other), and sound recordings were made in their proximity with a single hydrophone and a digital audiotape (DAT) recorder. A quantitative analysis of the production of pulsed calls, whistles and echolocation clicks in the three activities revealed that there was a significant effect of activity on the production of these sound types. Both killer whales in Icelandic and Norwegian waters produced high rates of clicks and calls during feeding and low rates of click, calls and whistles during travelling. The differences can be used as acoustical markers and provides new possibilities for acoustic monitoring of killer whales in these areas. Based on the similarity between their prey choice, hunting strategies, phenotype and acoustic behaviour, we suggest that the killer whales in Icelandic and Norwegian waters belong to the same ecotype: Scandinavian herring-eating killer whales. Udgivelsesdato: 18 April 2007

Icelandic and Norwegian killer whales feed on herring, after debilitating them with underwater tail slaps. We analysed sound recordings of Icelandic and Norwegian killer whales engaged in feeding and other behaviour. We describe a pulsed call made by Icelandic killer whales shortly before underwater tail slaps, which had an atypical low frequency (average peak frequency: 683 ±131 Hz), long duration (3.0 ±1.1 s) and high intensity (source level 169–192 dB pp re 1 μPa @ 1 m). The low-frequency emphasis of this call was below the most sensitive hearing range of killer whales, suggesting that the call may not be optimal for interspecific communication. However, herring could easily perceive the killer whale call since the frequency content is similar to the resonant frequency of their swim bladder as well as to the most sensitive frequency band of hearing in this species. Previous studies have shown that sound may cause schooling herring to cluster. A high density of herring in a school would increas...

SUMMARYNorwegian killer whales debilitate prey by slapping their tails into herring schools. These underwater tail slaps produce a thud-like sound. It is unclear whether this sound is caused by cavitation and/or physical contact between herring and whale tail. Also the forces causing debilitation of the fish are not understood. Here we present an acoustic analysis of underwater tail slaps using a multi-channel wide (150 kHz) band recording system. Underwater tail slaps produced by Norwegian killer whales generated sounds consisting of multiple pulses with source levels of 186±5.4 dB (pp)re.1 μPa at 1 m (±1 s.d., N=4). The –3 dB and 97% energy bandwidths were 36.8±22.5 kHz and 130.5±17.5 kHz(±1 s.d., N=13), respectively, with a centre frequency of 46.1±22.3 kHz. The similarities between the acoustic properties of underwater tail slaps recorded from killer whales in Norway, and thud-like sounds recorded from killer whales in Iceland suggest that Norwegian and Icelandic killer whales use similar hunting techniques. The acoustic characteristics of sounds produced by underwater tail slaps were similar to the ones from other cavitation sound sources described in the literature, both in term of temporal and frequency features as well as in source level. We suggest that multiple factors generated by the tail slaps like particle fluctuations, turbulence, pressure changes and physical impact cause debilitation of herring.

Summary of Committee. For the complete report please refer to Sexual Medicine: Sexual Dysfunctions in Men and Women , edited by T.F. Lue, R. Basson, R. Rosen, F. Giuliano, S. Khoury, F. Montorsi, Health Publications, Paris, 2004. ABSTRACT Introduction. Ethics describe the ways in which moral life is understood. Morality comprises norms for human conduct, and addresses what is right and what is wrong. Aim. To provide a consensus‐based summary of the ethical aspects of sexual medicine. Methods. Over 200 multidisciplinary specialists from 60 countries were divided into 17 consultation committees as part of a process organized by an international consultation on sexual medicine held in Paris, June 28–July 1, 2003 in close alliance with several sexual medicine organizations. Embarking on a study on ethics in sexual medicine, 10 experts from eight countries assembled over a two‐year period to develop this consensus‐based summary. Main Outcome Measure. Although ethics are recognized as subjective, expert opinion was based on grading of evidence‐based medical literature, in addition to cultural and ethical considerations. The process also involved extensive internal committee discussion, public presentation, and debate. Results. Contemporary medical practitioners provide health care for patients from many different cultures from all around the world. Thus, it is recommended that all health professionals working in sexual medicine should above all be able to demonstrate respect, understanding, and tolerance toward the differing moral worldviews of their patients and colleagues, and the societies they represent. In sexual medicine, health professionals have an obligation to respect the autonomy of any individual that they treat, regardless of that individual's religious or socio‐cultural tradition, race, gender, or sexual orientation. Sexual rights are a necessary condition for sexual health. Sexual health requires a positive and respectful approach to sexuality and sexual relationships as well as the possibility of having pleasurable and safe sexual experiences, free of coercion, discrimination, and violence. For sexual health to be attained and maintained, the sexual rights of all persons must be respected, protected, and fulfilled. Conclusions. Additional discussion and research on ethics in sexual medicine is needed.

T cell activation is initiated by signal 1 through an antigen–specific TCR recognition and engagement, then a second co-stimulation signal is needed. This co-stimulation is provided by the interaction of IMR receptors and ligands expressed on the T cells and APC or tumor cells. These receptor-ligand interactions can either stimulate or inhibit T cell activation and they are the main targets for the new therapeutics developed in the oncology field. The establishment of Ag-specific human T cell clones is essential to study IMR biology and even more to test different biologics targeting those molecules. In our laboratory, we developed primary-cell based assays to test such biologics. In those assays allo-Ag-specific human CD4+ T cell clones are activated by HLA-DR-positive JY cells, a human B cell line, to proliferate and to produce IFNγ. To analyze the efficacy of PD-1/Lag3 and PD-1/TIGIT biologics, we generated by lentivirus transduction, different JY cell line clones expressing PD-L1, CD155 or PD-L1/CD155. The JY clones isolated after infection showed a stable expression of the IMRs and they modulated activation of the CD4+ T cell clones.

Haematopoietic stem cells (HSCs) self-renew for life, thereby making them one of the few blood cells that truly age(1,2). Paradoxically, although HSCs numerically expand with age, their functional activity declines over time, resulting in degraded blood production and impaired engraftment following transplantation(2). While many drivers of HSC ageing have been proposed(2-5), the reason why HSC function degrades with age remains unknown. Here we show that cycling old HSCs in mice have heightened levels of replication stress associated with cell cycle defects and chromosome gaps or breaks, which are due to decreased expression of mini-chromosome maintenance (MCM) helicase components and altered dynamics of DNA replication forks. Nonetheless, old HSCs survive replication unless confronted with a strong replication challenge, such as transplantation. Moreover, once old HSCs re-establish quiescence, residual replication stress on ribosomal DNA (rDNA) genes leads to the formation of nucleolar-associated gamma H2AX signals, which persist owing to ineffective H2AX dephosphorylation by mislocalized PP4c phosphatase rather than ongoing DNA damage. Persistent nucleolar gamma H2AX also acts as a histone modification marking the transcriptional silencing of rDNA genes and decreased ribosome biogenesis in quiescent old HSCs. Our results identify replication stress as a potent driver of functional decline in old HSCs, and highlight the MCM DNA helicase as a potential molecular target for rejuvenation therapies.

Blockade of the checkpoint inhibitor PD-1 has demonstrated remarkable success in the clinic for the treatment of a growing list of different cancers. However, several tumor types are resistant to anti-PD-1 monotherapy. This observation has spurred numerous combination studies to reveal what additional therapeutic interventions may complement anti-PD1 blockade. Recently it has been shown that immunogenic cell death (ICD), induced by radiation and/or chemotherapy, improves T cell responses against different tumor types. ICD is characterized by damage-associated molecular patterns (DAMPs) including surface expression of calreticulin, and release of ATP and HMGB1. Recognition of DAMPs triggers dendritic cell maturation and functions that are critical for tumor antigen-specific T cell activation. Thus therapies that evoke ICD may further augment anti-tumor immunity elicited by anti-PD-1. In mouse syngeneic tumor models, we observed combinatorial anti-tumor activity of anti-PD1 and the cyclin-dependent kinase inhibitor, dinaciclib. We hypothesized that dinaciclib potentiates the effects of anti-PD-1 by eliciting ICD. Indeed, tumor cells treated with dinaciclib express the hallmarks of ICD including HMGB1 and ATP release and surface expression of calreticulin. Dinaciclib treatment also increases tumor cell phagocytosis and induces dendritic cell maturation. Furthermore, mice immunized with dinaciclib-treated tumor cells are resistant to subsequent tumor challenge. Tumors from mice receiving anti-PD1 and dinaciclib have increased T cell infiltration and dendritic cell activation, indicating the overall quality of the immune response generated may be improved with the combo. Taken together, these findings suggest a potential mechanism for the observed synergy between dinaciclib and anti-PD1. Dinaciclib induces immunogenic cell death, converting the tumor cell into an endogenous vaccine and thereby boosting the effects of anti-PD-1. Citation Format: Dewan Mohammed Sakib Hossain, Fernando Ugarte, Anandi Sawant, Mingmei Cai, Venkataraman Sriram, Elaine Pinheiro, Svetlana Sadekova, Alissa Chackerian. Dinaciclib induces immunogenic cell death and enhances anti-PD-1 mediated tumor suppression. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 562.

Introduction Disturbances in testicular integrity have the potential to cause severe concerns about masculinity, body image, and sexual function. Testicular volume replacement surgery with prosthesis has been known about since 1941, although esthetic procedures for testicular augmentation have not been satisfactory. Aim To describe an unprecedented testicular augmentation surgical technique that is specially focused on preserving testicular function and providing a favorable esthetic outcome. Methods We present a case of a 45‐year‐old man with body dysmorphic disorder. Surgical treatment using a new technique by placing a chin implant on the testicle was offered and accepted. Results Good symmetry between both testicles was achieved. No complications were reported. One month after the procedure, the patient started a satisfactory sex life. Functional monitoring was normal during the year after the procedure. Conclusion This technique is unprecedented in literature, it represents a safe and effective therapeutic alternative; nevertheless, more experience in performing this procedure is needed. Ugarte y Romano F and Gonzalez Serrano A. Testicular augmentation using chin implants. J Sex Med 2013;10:2582–2585.

Killer whales (Orcinus orca) feeding on herring (Clupea harengus) were observed both from the surface and underwater. We refer to one of the feeding techniques used by killer whales as the carousel method, whereby whales cooperatively herded herring into a tight ball close to the surface. During herding and feeding, whales swam around and under a school of herring, performing much lobtailing and porpoising. When the herring were gathered into a tight ball whales often swam with the white underside of their body towards the fish and emitted large bubbles close to the surface. While feeding, whales spent more time circling around the ball of fish than eating. Ball formation is a known defence mechanism used by schooling fish, and the effort by killer whales appeared to be directed towards keeping the ball very dense and close to the surface. The whales stunned their prey by slapping the edge of the school with the underside of their flukes and then ate the stunned fish one by one. The tail slaps created a loud banging sound which could have been either a by-product or an aid to stunning the prey.

Knowledge about harbour porpoise and bottlenose dolphin occurrence in Cardigan Bay Special Area of Conservation (SAC), Wales, is limited to daylight hours during summer, when conditions are suitable for traditional visual surveys. T-PODs are autonomous instruments programmed to log time-cues of species-specific echolocation signals for long periods of time. Here we investigated bottlenose dolphin and harbour porpoise habitat use and partitioning by deploying ten calibrated T-PODs in Cardigan Bay SAC for one year. The T-PODs detected both species all year round with a peak of detections in April–October for dolphins and in October–March for porpoise, revealing a previously unknown importance of the place to harbour porpoise during winter. Though the two species are sympatric, simultaneous detections of both species were rare and indication of temporal habitat partitioning between the two species in some parts of the SAC was observed. The one location where simultaneous detections were not as rare was close to the stretch of shoreline where stranding of porpoises killed by dolphins are most common, suggesting that the observed spatiotemporal overlap leads to inter-specific interactions, in some cases fatal for the porpoise.

Objective Internal tandem duplication (ITD) mutations of the FLT3 receptor are associated with a high incidence of relapse in acute myeloid leukemia (AML). Expression of the CXCR4 receptor in FLT3-ITD−positive AML is correlated with poor outcome, and inhibition of CXCR4 was shown to sensitize AML blasts toward chemotherapy. The aim of this study was to evaluate the impact of FLT3-ITD on cell proliferation and CXCR4-dependent migration in human hematopoietic progenitor cells and to investigate their response to CXCR4 inhibition. Materials and Methods We used primary blasts from patients with FLT3-ITD or FLT3 wild-type AML. In addition, human CD34 + hematopoietic progenitor cells were transduced to >70% with retroviral vectors containing human FLT3-ITD. Results We found that FLT3-ITD transgene overexpressing human hematopoietic progenitor cells show strongly reduced migration toward stromal-derived factor−1 in vitro and display significantly reduced bone marrow homing in nonobese diabetic severe combined immunodeficient mice. Cocultivation of FLT3-ITD−positive AML blasts or hematopoietic progenitor cells on bone marrow stromal cells resulted in a strong proliferation advantage and increased early cobblestone area−forming cells compared to FLT3−wild-type AML blasts. Addition of the CXCR4 inhibitor AMD3100 to the coculture significantly reduced both cobblestone area−forming cells and proliferation of FLT3-ITD−positive cells, but did not affect FLT3−wild-type cells—highlighting the critical interaction between CXCR4 and FLT3-ITD. Conclusion CXCR4 inhibition to decrease cell proliferation and to control the leukemic burden may provide a novel therapeutic strategy in patients with advanced FLT3-ITD−positive AML.

Objective The Notch signaling pathway has been shown to play a role in bone marrow−derived stromal cell differentiation, however, the precise outcome of Notch activation remains controversial. The aim of this study was to evaluate the effect of Notch signaling in primary human bone marrow−derived stromal cells (hBMSCs). Materials and Methods hBMSCs were transduced to >90% with lentiviral vectors containing either human notch1 intracellular domain ( NICD ), jagged1 , or dominant negative mastermind1 . Cells were exposed to adipogenic and osteogenic differentiation stimuli and differentiation was quantified by oil red or alizarin red staining, alkaline phosphatase liver/bone/kidney (ALPL) activity and expression of adipogenic or osteogenic marker genes. Results NICD and jagged1 transgene−expressing hBMSCs demonstrated enhanced mineralization, nodule formation, and ALPL activity in osteogenic differentiation media. These findings correlated with increased gene expression of bone morphogenetic protein 2 and ALPL . In contrast, NICD or jagged1 transgene expression strongly inhibited adipocyte formation and reduced peroxisome proliferator-activated receptor-γ , fatty acid binding protein 4 , and adiponectin precursor gene expression. Co-overexpression of dominant negative mastermind1 and NICD or jagged1 led to a partial rescue of the differentiation phenotypes. In addition, high endogenous jagged1 expression levels were observed in hBMSCs samples with strong ALPL activity compared to a group of samples with low ALPL activity. Conclusion In summary, our data suggest that induction of Notch signaling enhances the osteogenic differentiation of hBMSCs while inhibiting the adipogenic fate.

The CXC chemokine receptor 4 (CXCR4) and its ligand stromal derived factor 1 (SDF-1) regulate egress and homing of hematopoietic stem cells. Activation of sphingosine-1-phosphate (S1P) receptors (S1P(1-5)) modulates chemokine-induced migration of lymphocytes and hematopoietic stem cells. To analyze the influence of S1P(1) on SDF-1-dependent chemotaxis and trafficking, we overexpressed S1P(1) in CD34+ mobilized peripheral blood progenitor cells (PBPCs). Using a gamma-retroviral vector, transgene overexpression was achieved in more than 90% of target cells. S1P(1) transgene positive PBPCs showed enhanced chemotaxis towards S1P. S1P(1) overexpression resulted in reduced CXCR4 surface expression levels and strong inhibition of SDF-1-dependent ERK1/2 phosphorylation and Ca(2+) flux. Furthermore, SDF-1-dependent migration of S1P(1) overexpressing PBPCs or Jurkat cells was reduced up to 10-fold. Sublethally irradiated NOD/SCID mice were transplanted with 6-day cultured PBPCs overexpressing either S1P(1)-IRES-GFP or GFP alone. Screening for GFP positive human cells in the mouse bone marrow 20h after transplantation revealed an eightfold reduction in bone marrow homing of S1P(1) transgene expressing cells. Our data suggest that S1P(1) acts as an inhibitor of CXCR4-dependent migration of hematopoietic cells to sites of SDF-1 production.

DNA-Vector transfection of eukaryotic cells needs transport of the DNA to the nucleus for efficient transcription and subsequent translation. In suspension cells most DNA-transfection methods are either inefficient (e.g. lipofection) or stressful for the cells (electroporation or nucleofection), which leads to high cell losses during the transfection process. We report the use of in vitro transcribed, capped and polyadenylated mRNA for the transfection of leukemic cell lines in comparison to DNA delivery. Templates for in vitro transcription were generated by PCR. The forward primer included the T7-promoter and the reverse primer a fragment of the beta-globin 3'UTR to increase the in vivo half life of the mRNA. Electroporation of GFP-mRNA into Jurkat, K562 or Kg1a lead to a GFP overexpression in 95 % of the transfected cells, with survival rates of 80 to 90%. Electroporation with pEGFP-N1 using the same pulse conditions resulted in GFP-overexpression in 62% of Jurkat or 9% of K562, with survival rates of 50% and 40 %, respectively. Our results suggest that electroporation of DNA is more toxic than mRNA transfection. GFP-expression in mRNA transfected cells was visible two hours after the transfection and peaked at about 12h. In contrast to DNA-transfection, mRNA mediated GFP expression levels were uniform in transfected cells. To investigate whether mRNA transfection results in functional transgene expression, mRNA coding the chemokine receptor CXCR4 was generated and electroporated into CXCR4 negative K562 cells. Flow cytometric analyses showed CXCR4 overexpression in more than 90% of transfected cells. Calcium flux measurements displayed a robust increase of intracellular free calcium in CXCR4-mRNA transfected K562 upon binding of the CXCR4 ligand SDF-1, while no calcium flux was visible in GFP-mRNA transfected or untransfected K562. Electroporation of GFP-mRNA into CD34+ hematopoietic progenitor cells or mesenchymal stem cells resulted in GFP overexpression in > 50% of transfected cells, respectively. Experiments are under way to further improve the mRNA transfection for primary cells. mRNA transfection is a highly efficient means for transient overexpression of transgenes in various cell lines and primary cells.

The objective of the present study was to determine the efficacy and safety of a fixed dose of vardenafil in the treatment of patients with erectile dysfunction (ED).This was an open label, prospective and multicentric trial. After a 4-week wash out period, all patients received 20 mg of vardenafil given on demand for 12 weeks. Primary efficacy variables were the erectile function domain of the International Index of Erectile Function (IIEF), answers to questions 2 and 3 of the Sexual Encounter Profile (SEP) and the Global Assessment Question (GAQ). All adverse events were recorded and reported.229 patients were screened. 177 received at least one dose of vardenafil and were included in the safety analysis. Mean age was 54.4 years old. Etiology of ED was organic or mixed in 77% of the patients. Erectile function domain of the IIEF changed from a basal mean score of 14.8 to 25.5 at the end of the study. 80.5% of the patients reported erections of rigidity and duration enough for satisfactory sexual intercourse and 93.3% improved their erections at the end of the study. Adverse events were mild to moderate and the most common were headache, dyspepsia, rhinitis and facial flushing. The drop out rate due to adverse events was 1.7%.This multicenter study confirms the high efficacy of this new phosphodiesterase type 5 inhibitor, vardenafil. There was a low rate of discontinuations due to adverse events and a favorable safety profile. The results of this study are similar to the results of other studies conducted in other parts of the world.