Gastruloid-derived primordial germ cell-like cells develop dynamically within integrated tissues.

Primordial germ cells (PGCs) are the early embryonic precursors of gametes - sperm and egg cells. PGC-like cells (PGCLCs) can currently be derived in vitro from pluripotent cells exposed to signalling cocktails and aggregated into large embryonic bodies, but these do not recapitulate the native embryonic environment during PGC formation. Here, we show that mouse gastruloids, a three-dimensional in vitro model of gastrulation, contain a population of gastruloid-derived PGCLCs (Gld-PGCLCs) that resemble early PGCs in vivo. Importantly, the conserved organisation of mouse gastruloids leads to coordinated spatial and temporal localisation of Gld-PGCLCs relative to surrounding somatic cells, even in the absence of specific exogenous PGC-specific signalling or extra-embryonic tissues. In gastruloids, self-organised interactions between cells and tissues, including the endodermal epithelium, enables the specification and subsequent maturation of a pool of Gld-PGCLCs. As such, mouse gastruloids represent a new source of PGCLCs in vitro and, owing to their inherent co-development, serve as a novel model to study the dynamics of PGC development within integrated tissue environments.

A training programme to build cancer research capacity in low- and middle-income countries: findings from Guatemala.

PROBLEM: Guatemala is experiencing an increasing burden of cancer but lacks capacity for cancer prevention, control and research. APPROACH: In partnership with a medical school in the United States of America, a multidisciplinary Cancer Control Research Training Institute was developed at the Instituto de Cancerología (INCAN) in Guatemala City. This institute provided a year-long training programme for clinicians that focused on research methods in population health and sociocultural anthropology. The programme included didactic experiences in Guatemala and the United States as well as applied training in which participants developed research protocols responsive to Guatemala's cancer needs. LOCAL SETTING: Although INCAN is the point of referral and service for Guatemala's cancer patients, the institute's administration is also interested in increasing cancer research - with a focus on population health. INCAN is thus a resource for capacity building within the context of cancer prevention and control. RELEVANT CHANGES: Trainees increased their self-efficacy for the design and conduct of research. Value-added benefits included establishment of an annual cancer seminar and workshops in cancer pathology and qualitative analysis. INCAN has recently incorporated some of the programme's components into its residency training and established a research department. LESSONS LEARNT: A training programme for clinicians can build cancer research capacity in low- and middle-income countries. Training in population-based research methods will enable countries such as Guatemala to gather country-specific data. Once collected, such data can be used to assess the burden of cancer-related disease, guide policy for reducing it and identify priority areas for cancer prevention and treatment.

Predictive coding in musical anhedonia: A study of groove.

Groove, or the pleasurable urge to move to music, offers unique insight into the relationship between emotion and action. The predictive coding of music model posits that groove is linked to predictions of music formed over time, with stimuli of moderate complexity rated as most pleasurable and likely to engender movement. At the same time, listeners vary in the pleasure they derive from music listening: individuals with musical anhedonia report reduced pleasure during music listening despite no impairments in music perception and no general anhedonia. Little is known about musical anhedonics' subjective experience of groove. Here we examined the relationship between groove and music reward sensitivity. Participants (n = 287) heard drum-breaks that varied in perceived complexity, and rated each for pleasure and wanting to move. Musical anhedonics (n = 13) had significantly lower ratings compared to controls (n = 13) matched on music perception abilities and general anhedonia. However, both groups demonstrated the classic inverted-U relationship between ratings of pleasure & move and stimulus complexity, with ratings peaking for intermediately complex stimuli. Across our entire sample, pleasure ratings were most strongly related with music reward sensitivity for highly complex stimuli (i.e., there was an interaction between music reward sensitivity and stimulus complexity). Finally, the sensorimotor subscale of music reward was uniquely associated with move, but not pleasure, ratings above and beyond the five other dimensions of musical reward. Results highlight the multidimensional nature of reward sensitivity and suggest that pleasure and wanting to move are driven by overlapping but separable mechanisms.

Mutations in yeast Pcf11, a conserved protein essential for mRNA 3' end processing and transcription termination, elicit the Environmental Stress Response.

The Pcf11 protein is an essential subunit of the large complex that cleaves and polyadenylates eukaryotic mRNA precursor. It has also been functionally linked to gene-looping, termination of RNA Polymerase II (Pol II) transcripts, and mRNA export. We have examined a poorly characterized but conserved domain (amino acids 142-225) of the Saccharomyces cerevisiae  Pcf11 and found that while it is not needed for mRNA 3' end processing or termination downstream of the poly(A) sites of protein-coding genes, its presence improves the interaction with Pol II and the use of transcription terminators near gene promoters. Analysis of genome-wide Pol II occupancy in cells with Pcf11 missing this region, as well as Pcf11 mutated in the Pol II CTD Interacting Domain, indicates that systematic changes in mRNA expression are mediated primarily at the level of transcription. Global expression analysis also shows that a general stress response, involving both activation and suppression of specific gene sets known to be regulated in response to a wide variety of stresses, is induced in the two pcf11 mutants, even though cells are grown in optimal conditions. The mutants also cause an unbalanced expression of cell wall-related genes that does not activate the Cell Wall Integrity pathway but is associated with strong caffeine sensitivity. Based on these findings, we propose that Pcf11 can modulate the expression level of specific functional groups of genes in ways that do not involve its well-characterized role in mRNA 3' end processing.

Regulation of long-range BMP gradients and embryonic polarity by propagation of local calcium-firing activity.

Many amniote vertebrate species including humans can form identical twins from a single embryo, but this only occurs rarely. It has been suggested that the primitive-streak-forming embryonic region emits signals that inhibit streak formation elsewhere but the signals involved, how they are transmitted and how they act has not been elucidated. Here we show that short tracks of calcium firing activity propagate through extraembryonic tissue via gap junctions and prevent ectopic primitive streak formation in chick embryos. Cross-regulation of calcium activity and an inhibitor of primitive streak formation (Bone Morphogenetic Protein, BMP) via NF-κB and NFAT establishes a long-range BMP gradient spanning the embryo. This mechanism explains how embryos of widely different sizes can maintain positional information that determines embryo polarity. We provide evidence for similar mechanisms in two different human embryo models and in Drosophila, suggesting an ancient evolutionary origin.

Study of secondhand smoke exposure in St. Louis City and County suggests need for comprehensive smoke-free Missouri law adoption.

This cross-sectional study provides information about secondhand smoke exposure across the St. Louis metro area and perceptions and attitudes about tobacco and health within the local hospitality industry. Results from this study support the need for passage and implementation of comprehensive smoke-free laws throughout Missouri, particularly in St. Louis City and County where efforts to pass comprehensive smoke-free laws have been unsuccessful.

Adolescent thriving: the role of sparks, relationships, and empowerment.

Although most social science research on adolescence emphasizes risks and challenges, an emergent field of study focuses on adolescent thriving. The current study extends this line of inquiry by examining the additive power of identifying and nurturing young people's "sparks," giving them "voice," and providing the relationships and opportunities that reinforce and nourish thriving. A national sample of 1,817 adolescents, all age 15 (49% female), and including 56% white, 17% Hispanic/Latino, and 17% African-American adolescents, completed an online survey that investigated their deep passions or interests (their "sparks"), the opportunities and relationships they have to support pursuing those sparks, and how empowered they feel to make civic contributions (their "voice"). Results consistently supported the hypothesis that linking one's spark with a sense of voice and supportive opportunities and relationships strengthens concurrent outcomes, particularly those reflecting prosociality, during a key developmental transition period. The three developmental strengths also predicted most outcomes to a greater degree than did demographics. However, less than 10 percent of 15-year-olds reported experiencing high levels of all three strengths. The results demonstrate the value of focusing on thriving in adolescence, both to reframe our understanding of this age group and to highlight the urgency of providing adolescents the opportunities and relationships they need to thrive.

Gastruloids: Embryonic Organoids from Mouse Embryonic Stem Cells to Study Patterning and Development in Early Mammalian Embryos.

Gastruloids are embryonic organoids made from small, defined numbers of mouse embryonic stem cells (mESCs) aggregated in suspension culture, which over time form 3D structures that mimic many of the features of early mammalian development. Unlike embryoid bodies that are usually disorganized when grown over several days, gastruloids display distinct, well-organized gene expression domains demarcating the emergence of the three body axes, anteroposterior axial elongation, and implementation of collinear Hox transcriptional patterns over 5-7 days of culture. As such gastruloids represent a useful experimental system that is complementary to in vivo approaches in studying early developmental patterning mechanisms regulating the acquisition of cell fates. In this protocol, we describe the most recent method for generating gastruloids with high reproducibility, and provide a comprehensive list of possible challenges as well as steps for protocol optimization.

In vitro teratogenicity testing using a 3D, embryo-like gastruloid system.

Pharmaceuticals intended for use in patients of childbearing potential need to be tested for teratogenicity before marketing. Several pharmaceutical companies use animal-free in vitro models which allow a more rapid selection of lead compounds and contribute to 3Rs principles ('replace, reduce and refine') by streamlining the selection of promising compounds submitted to further regulatory studies in animals. Currently available in vitro models typically rely on adherent monolayer cultures or disorganized 3D structures, both of which lack the spatiotemporal and morphological context of the developing embryo. A newly developed 3D 'gastruloid' model has the potential to achieve a more reliable prediction of teratogenicity by providing a robust recapitulation of gastrulation-like events alongside morphological coordination at relatively high-throughput. In this first proof-of-concept study, we used both mouse and human gastruloids to examine a panel of seven reference compounds, with associated in vivo data and known teratogenic risk, to quantitatively assess in vitro teratogenicity. We observed several gross morphological effects, including significantly reduced elongation or decreased size of the gastruloids, upon exposure to several of the reference compounds. We also observed aberrant gene expression using fluorescent reporters, including SOX2, BRA, and SOX17, suggestive of multi-lineage differentiation defects and disrupted axial patterning. Finally, we saw that gastruloids recapitulated some of the known in vivo species-specific susceptibilities between their mouse and human counterparts. We therefore suggest that gastruloids represent a powerful tool for teratogenicity assessment by enabling relevant physiological recapitulation of early embryonic development, demonstrating their use as a novel in vitro teratogenic model system.

One-Pot Preparation of Peptide-Doped Metal-Amino Acid Framework for General Encapsulation and Targeted Delivery.

Metal-organic frameworks (MOFs), especially those made by biological molecules (bio-MOFs), have been proved to be prospective candidates for biomedical applications. However, a simple and universal bio-MOF to load different substances for precise targeting is still lacking. In this work, we propose a facile one-pot method to prepare a peptide-doped bio-MOF for general encapsulation and targeted delivery. This bio-MOF is constructed by 9-fluorenylmethyloxycarbonyl-modified histidine (Fmoc-His) as a bridging linker that coordinates with Zn2+ ions, denoted as ZFH. The Fmoc-His-Asp-Gly-Arg peptide (Fmoc-HDGR) can be easily doped into the ZFH structure with different ratios to modulate the targeting ability of ZFH-DGR. Containing both hydrophobic Fmoc and hydrophilic His moieties, this framework is compatible with encapsulating various types of payloads, including hydrophobic chemotherapeutic, hydrophilic protein, and positively/negatively charged inorganic nanoparticles. It has also been proved to be highly biocompatible and stable in circulation, exhibit the capabilities to target ανß3 integrin overexpressed on tumor cells, and trigger drug release in a low pH microenvironment at the tumor site. As a proof of concept, Doxorubicin (Dox)-loaded ZFH-DGR (ZFH-DGR/Dox) demonstrated high cell selectivity between liver hepatocellular carcinoma (HepG2) cells and normal liver (L02) cells, which express high and low ανß3 integrin, respectively. This selectivity endows ZFH-DGR/Dox precise treatment and low toxicity in Heps-bearing liver cancer mice. This work develops a de novo approach to construct a peptide-doped bio-MOF system for universal load, precise delivery, and peptide drug combination therapy in the future.

Tobacco talk: reflections on corporate power and the legal framing of consumption.

This article examines how North Carolina tobacco farmers think about the moral ambiguities of tobacco business. Drawing on ethnographic research with tobacco farmers and archival research on the tobacco industry, I specify the core psychological defense mechanisms that tobacco companies have crafted for people associated with the industry. I also document local social, cultural, and economic factors in rural North Carolina that underpin ongoing rural dependence on tobacco despite the negativity that surrounds tobacco and structural adjustments. This article contributes to our knowledge about tobacco farmers and tobacco farming communities, which is important for tobacco-control strategies. I reflect on ethical and economic paradoxes related to the rise of corporate social responsibility in the tobacco industry, where an official legal framing of consumption, focused on informed adult consumer autonomy and health education, is promoted to undermine more robust public health prevention efforts.

Pluripotent stem cell models of early mammalian development.

Pluripotent stem cells derived from the early mammalian embryo offer a convenient model system for studying cell fate decisions in embryogenesis. The last 10 years have seen a boom in the popularity of two-dimensional micropatterns and three-dimensional stem cell culture systems as a way to recreate the architecture and interactions of particular cell populations during development. These methods enable the controlled exploration of cellular organization and patterning during development, using cell lines instead of embryos. They have established a new class of in vitro model system for pre-implantation and peri-implantation embryogenesis, ranging from models of the blastocyst stage, through gastrulation and toward early organogenesis. This review aims to set these systems in context and to highlight the strengths and suitability of each approach in modelling early mammalian development.

Conspicuous angiotropism of malignant melanoma involving the brain: implications for extravascular migratory metastasis.

Angiotropism is the presence of tumor cells closely apposed to the abluminal surfaces of blood and lymphatic vessels without intravasation. Previous studies have strongly suggested that angiotropism in melanoma could be a marker for extravascular migratory metastasis, the migration of tumor cells along the external surfaces of vessels. We describe for the first time a patient with malignant melanoma of the brain most likely metastatic, which was floridly angiotropic as evidenced by extensive spread of melanoma cells along the external surfaces of brain microvessels. The location of this angiotropic melanoma in the brain, together with the analogies between extravascular migratory metastasis and the neoplastic glial invasion of the nervous system, reinforces the hypothesis of extravascular migration of melanoma cells as a means of tumor spread, particularly along the abluminal surfaces of vessels, in the brain and in other organs.

Racial/ethnic differences in bystander CPR in Los Angeles, California.

BACKGROUND: Bystander CPR (BCPR) has been demonstrated to improve rates of return of spontaneous circulation, survival to hospital admission, and quality of life in survivors. While previous studies have shown that African Americans are less likely to receive BCPR than Caucasians even after adjusting for variables such as socioeconomic status, BCPR rates in Latinos have not been reported. OBJECTIVE: To describe BCPR rates in an urban African American and Latino population as compared to Caucasians. METHODS: A retrospective analysis of the Cardiac Arrest Resuscitation Evaluation in Los Angeles (CARE-LA) database combined with the California Death Statistical Master File (CDSMF). The combined database included location, race/ethnicity/ethnic background, witnessed status, socioeconomic status, and other variables that have previously been associated with differing rates of BCPR. RESULTS: There were 814 individuals included in the final study group (53% Caucasian, 28% African American, 19% Latino). African Americans and Latinos were younger than the Caucasians, had more events in the home and had a bystander CPR rate of 13% compared to 24% for the Caucasians (OR=0.47 (95%CI: 0.30-0.74) for African Americans and OR=0.48 (95%CI:0.28-0.80) for the Latinos). Bystander CPR was found to be an independent predictor of survival to hospital discharge and, after adjustment, Latino ethnicity was associated with lower rates of bystander CPR (OR 0.45 (95%CI:0.22-0.92)). CONCLUSION: After adjusting for other variables, Latinos in Los Angeles receive bystander CPR at approximately half the rate of Caucasians.

Spiritual development: a missing priority in youth development.

Addressing the spiritual development of young people has the potential to strengthen youth work and its outcomes. Spiritual development matters because it is an intrinsic part of being human and because young people themselves view it as important. This article reviews the research that points to positive impacts of spiritual development for youth and notes that in an increasingly pluralistic society, everyone needs to build skills for negotiating religious and spiritual diversity. The authors propose that spiritual development involves, in part, the dynamic interplay of three dimensions: belonging and connecting, awareness and awakening, and a way of living. Three initial challenges and opportunities are emerging: empowering youth to explore core developmental issues, motivation and focus, and multisector engagement.

Mitochondria: promising organelle targets for cancer diagnosis and treatment.

Mitochondria, the energy supply factories for cell-life activities, play important roles in controlling epigenetics, differentiation and initiation, and the execution of apoptosis. These functions of the mitochondria contribute to cell adaptation to challenging microenvironment conditions. In past decades, mitochondrial malfunction has been revealed to be closely related to the occurrence and development of a variety of human disorders, including cancer and multiple neurodegenerative diseases. The disturbance of the mitochondrial genome (mtDNA) or mitochondrial vital functions, e.g., the production of adenosine triphosphate (ATP) and the generation of reactive oxygen species (ROS), can potentially be involved in disease pathogenesis. Recent research has shown that the precise monitoring of mitochondrial environments can provide potential directions for cancer diagnosis. Furthermore, mitochondrial-targeted cancer treatment exhibits unparalleled superiority for enhanced tumor therapy. Therefore, in this review, we focus on mitochondrial-based cancer diagnosis via monitoring mitochondrial respiration or mitophagy. Current approaches using mitochondrial-based cancer treatments, including targeting mitochondrial ATP, mitochondrial membrane permeability, and mitochondrial ROS levels and mtDNA, are also summarized. This review will provide insights into mitochondrial-mediated tumor monitoring and mitochondrial-based therapy.

Programmed death ligand 1 (PD-L1) expression influences the immune-tolerogenic microenvironment in antiretroviral therapy-refractory Kaposi's sarcoma: A pilot study.

Upregulation of programmed death ligand 1 (PD-L1) is a mechanism of immune escape utilized by a variety of tumors. PD-L1 expression in tumor cells or in the surrounding infiltrate correlates with clinical responsiveness to novel therapies targeting the PD-1/PD-L1 immune checkpoint. In the context of HIV-1 infection, Kaposi's sarcoma (KS) is largely responsive to restoration of immunity following combination antiretroviral therapy (cART), but there is a subset that is not. We hypothesized that this subset of cART-refractory KS may utilize the PD-L1 pathway of immune escape. We found that PD-L1 expressing KS had a denser CD8+ T cell (p = 0.03) and PD-L1 positive macrophage peritumoral infiltrate (p = 0.04) to suggest the involvement of PD-L1 in shaping an immune-tolerogenic microenvironment in cART-refractory KS. The presence of PD-L1 expression in association with immune-infiltrating cells provides rationale for the clinical development PD-1/PD-L1-targeted checkpoint inhibitors in cART-refractory KS.

Experience With Prehospital Damage Control Capability in Modern Conflict: Results From Surgical Resuscitation Team Use.

BACKGROUND: Early resuscitation and damage control surgery (DCS) are critical components of modern combat casualty care. Early and effective DCS capabilities can be delivered in a variety of settings through the use of a mobile surgical resuscitation team (SRT). METHODS: Twelve years of after-action reports from SRTs were reviewed. Demographics, interventions, and outcomes were analyzed. RESULTS: Data from 190 casualties (185 human, five canine) were reviewed. Among human casualties, 12 had no signs of life at intercept and did not survive. Of the remaining 173 human casualties, 96.0% were male and 90.8% sustained penetrating injuries. Interventions by the SRT included intravascular access (50.9%) and advanced airway establishment (29.5%). Resuscitation included whole blood (3.5%), packed red blood cells (20.8%), and thawed plasma (11.0%). Surgery was provided for 63 of the 173 human casualties (36.4%), including damage control laparotomy (23.8%) and arterial injury shunting or repair (19.0%). SRTs were effectively used to augment an existing medical treatment facility (70.5%), to facilitate casualty transport (13.3%), as an independent surgical entity at a forward ground structure (9.2%), and in mobile response directly to the point of injury (6.9%). Overall survival was 97.1%. CONCLUSION: An SRT provides a unique DCS capability that can be successfully used in a variety of flexible roles.

Efficacy and Safety of Tranexamic Acid in Prehospital Traumatic Hemorrhagic Shock: Outcomes of the Cal-PAT Study.

INTRODUCTION: The California Prehospital Antifibrinolytic Therapy (Cal-PAT) study seeks to assess the safety and impact on patient mortality of tranexamic acid (TXA) administration in cases of trauma-induced hemorrhagic shock. The current study further aimed to assess the feasibility of prehospital TXA administration by paramedics within the framework of North American emergency medicine standards and protocols. METHODS: This is an ongoing multi-centered, prospective, observational cohort study with a retrospective chart-review comparison. Trauma patients identified in the prehospital setting with signs of hemorrhagic shock by first responders were administered one gram of TXA followed by an optional second one-gram dose upon arrival to the hospital, if the patient still met inclusion criteria. Patients administered TXA make up the prehospital intervention group. Control group patients met the same inclusion criteria as TXA candidates and were matched with the prehospital intervention patients based on mechanism of injury, injury severity score, and age. The primary outcomes were mortality, measured at 24 hours, 48 hours, and 28 days. Secondary outcomes measured included the total blood products transfused and any known adverse events associated with TXA administration. RESULTS: We included 128 patients in the prehospital intervention group and 125 in the control group. Although not statistically significant, the prehospital intervention group trended toward a lower 24-hour mortality rate (3.9% vs 7.2% for intervention and control, respectively, p=0.25), 48-hour mortality rate (6.3% vs 7.2% for intervention and control, respectively, p=0.76), and 28-day mortality rate (6.3% vs 10.4% for intervention and control, respectively, p=0.23). There was no significant difference observed in known adverse events associated with TXA administration in the prehospital intervention group and control group. A reduction in total blood product usage was observed following the administration of TXA (control: 6.95 units; intervention: 4.09 units; p=0.01). CONCLUSION: Preliminary evidence from the Cal-PAT study suggests that TXA administration may be safe in the prehospital setting with no significant change in adverse events observed and an associated decreased use of blood products in cases of trauma-induced hemorrhagic shock. Given the current sample size, a statistically significant decrease in mortality was not observed. Additionally, this study demonstrates that it may be feasible for paramedics to identify and safely administer TXA in the prehospital setting.

Empiric acyclovir is infrequently initiated in the emergency department to patients ultimately diagnosed with encephalitis.

STUDY OBJECTIVE: We evaluate the frequency of empiric acyclovir administration to patients in the emergency department (ED) who are ultimately diagnosed with encephalitis. METHODS: We conducted an explicit retrospective medical record review of consecutive patients discharged with a final diagnosis of herpes simplex encephalitis or viral encephalitis not otherwise specified for the period 1993 to 2003. The frequency of ED administration of empiric acyclovir was measured for patients who met the inclusion criteria of fever, neuropsychiatric abnormality, and cerebrospinal fluid pleocytosis with a negative Gram's stain result in the ED. RESULTS: Of the 90 patients reviewed, 24 (27%) met the inclusion criteria of fever, neuropsychiatric abnormality, and cerebrospinal fluid pleocytosis with a negative Gram's stain result in the ED. Of these 24 patients, 7 (29%) received empiric acyclovir in the ED, 6 (86%) patients after cerebrospinal fluid results were available, with a median time to administration of 1.5 hours (95% confidence interval [CI] 0 to 3.1 hours). The remaining 17 (71%) patients did not receive acyclovir in the ED, with median times of 16 hours (95% CI 7.5 to 44 hours) before initiation of acyclovir in inpatient settings. CONCLUSION: The majority of patients in our institution who were ultimately diagnosed with encephalitis did not receive empiric acyclovir in the ED, despite clinical presentations consistent with encephalitis.