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Dynamin-related protein 1 (Drp1) is a mitochondrial fission protein and a viable target for cardioprotection against myocardial ischaemia-reperfusion injury. Here, we reported a novel Drp1 inhibitor (DRP1i1), delivered using a cardiac-targeted nanoparticle drug delivery system, as a more effective approach for achieving acute cardioprotection. DRP1i1 was encapsulated in cubosome nanoparticles with conjugated cardiac-homing peptides (NanoDRP1i1) and the encapsulation efficiency was 99.3 ± 0.1 %. In vivo, following acute myocardial ischaemia-reperfusion injury in mice, NanoDRP1i1 significantly reduced infarct size and serine-616 phosphorylation of Drp1, and restored cardiomyocyte mitochondrial size to that of sham group. Imaging by mass spectrometry revealed higher accumulation of DRP1i1 in the heart tissue when delivered as NanoDRP1i1. In human cardiac organoids subjected to simulated ischaemia-reperfusion injury, treatment with NanoDRP1i1 at reperfusion significantly reduced cardiac cell death, contractile dysfunction, and mitochondrial superoxide levels. Following NanoDRP1i1 treatment, cardiac organoid proteomics further confirmed reprogramming of contractile dysfunction markers and enrichment of the mitochondrial protein network, cytoskeletal and metabolic regulation networks when compared to the simulated injury group. These proteins included known cardioprotective regulators identified in human organoids and in vivo murine studies following ischaemia-reperfusion injury. DRP1i1 is a promising tool compound to study Drp1-mediated mitochondrial fission and exhibits promising therapeutic potential for acute cardioprotection, especially when delivered using the cardiac-targeted cubosome nanoparticles.

Abstract Pathological reprogramming of cardiomyocyte and fibroblast proteome landscapes drive the initiation and progression of cardiac fibrosis. Although the secretome of dysfunctional cardiomyocytes is emerging as an important driver of pathological fibroblast reprogramming, our understanding of the downstream molecular players remains limited. Here, we show that cardiac fibroblast activation (αSMA+) and oxidative stress mediated by the secretome of TGFβ‐stimulated cardiomyocytes is associated with a profound reprogramming of their proteome and phosphoproteome landscape. Within the fibroblast global proteome there was a striking dysregulation of proteins implicated in extracellular matrix, protein localisation/metabolism, KEAP1‐NFE2L2 pathway, lysosomes, carbohydrate metabolism, and transcriptional regulation. Kinase substrate enrichment analysis of phosphopeptides revealed potential role of kinases (CK2, CDK2, PKC, GSK3B) during this remodelling. We verified upregulated activity of casein kinase 2 (CK2) in secretome‐treated fibroblasts, and pharmacological CK2 inhibitor TBB (4,5,6,7‐Tetrabromobenzotriazole) significantly abrogated fibroblast activation and oxidative stress. Our data provides molecular insights into cardiomyocyte to cardiac fibroblast crosstalk, and the potential role of CK2 in regulating cardiac fibroblast activation and oxidative stress.

It is challenging to directly characterize mechanical properties of soft 3D cardiac organoids with current sensors. Here the authors report an electronic skin-based all-soft organoid-sensing system which can wirelessly monitor minute force profiles of cardiac organoids in real-time in-situ.

Summary: To assess the transcriptomic profile of disease-specific cell populations, fibroblasts from patients with primary open-angle glaucoma (POAG) were reprogrammed into induced pluripotent stem cells (iPSCs) before being differentiated into retinal organoids and compared with those from healthy individuals. We performed single-cell RNA sequencing of a total of 247,520 cells and identified cluster-specific molecular signatures. Comparing the gene expression profile between cases and controls, we identified novel genetic associations for this blinding disease. Expression quantitative trait mapping identified a total of 4,443 significant loci across all cell types, 312 of which are specific to the retinal ganglion cell subpopulations, which ultimately degenerate in POAG. Transcriptome-wide association analysis identified genes at loci previously associated with POAG, and analysis, conditional on disease status, implicated 97 statistically significant retinal ganglion cell-specific expression quantitative trait loci. This work highlights the power of large-scale iPSC studies to uncover context-specific profiles for a genetically complex disease.

Abstract Background Silica nanoparticles (nanoSiO2) are promising systems that can deliver biologically active compounds to tissues such as the heart in a controllable manner. However, cardiac toxicity induced by nanoSiO2 has been recently related to abnormal calcium handling and energetic failure in cardiomyocytes. Moreover, the precise mechanisms underlying this energetic debacle remain unclear. In order to elucidate these mechanisms, this article explores the ex vivo heart function and mitochondria after exposure to nanoSiO2. Results The cumulative administration of nanoSiO2 reduced the mechanical performance index of the rat heart with a half-maximal inhibitory concentration (IC50) of 93 μg/mL, affecting the relaxation rate. In isolated mitochondria nanoSiO2 was found to be internalized, inhibiting oxidative phosphorylation and significantly reducing the mitochondrial membrane potential (ΔΨm). The mitochondrial permeability transition pore (mPTP) was also induced with an increasing dose of nanoSiO2 and partially recovered with, a potent blocker of the mPTP, Cyclosporine A (CsA). The activity of aconitase and thiol oxidation, in the adenine nucleotide translocase, were found to be reduced due to nanoSiO2 exposure, suggesting that nanoSiO2 induces the mPTP via thiol modification and ROS generation. In cardiac cells exposed to nanoSiO2, enhanced viability and reduction of H2O2 were observed after application of a specific mitochondrial antioxidant, MitoTEMPO. Concomitantly, CsA treatment in adult rat cardiac cells reduced the nanoSiO2-triggered cell death and recovered ATP production (from 32.4 to 65.4%). Additionally, we performed evaluation of the mitochondrial effect of nanoSiO2 in human cardiomyocytes. We observed a 40% inhibition of maximal oxygen consumption rate in mitochondria at 500 μg/mL. Under this condition we identified a remarkable diminution in the spare respiratory capacity. This data indicates that a reduction in the amount of extra ATP that can be produced by mitochondria during a sudden increase in energy demand. In human cardiomyocytes, increased LDH release and necrosis were found at increased doses of nanoSiO2, reaching 85 and 48%, respectively. Such deleterious effects were partially prevented by the application of CsA. Therefore, exposure to nanoSiO2 affects cardiac function via mitochondrial dysfunction through the opening of the mPTP. Conclusion The aforementioned effects can be partially avoided reducing ROS or retarding the opening of the mPTP. These novel strategies which resulted in cardioprotection could be considered as potential therapies to decrease the side effects of nanoSiO2 exposure.

Age-related macular degeneration (AMD) is a blinding disease characterised by dysfunction of the retinal pigmented epithelium (RPE) which culminates in disruption or loss of the neurosensory retina. Genome-wide association studies have identified >60 genetic risk factors for AMD; however, the expression profile and functional role of many of these genes remain elusive in human RPE. To facilitate functional studies of AMD-associated genes, we developed a human RPE model with integrated CRISPR interference (CRISPRi) for gene repression by generating a stable ARPE19 cell line expressing dCas9-KRAB. We performed transcriptomic analysis of the human retina to prioritise AMD-associated genes and selected TMEM97 as a candidate gene for knockdown study. Using specific sgRNAs, we showed that knockdown of TMEM97 in ARPE19 reduced reactive oxygen species (ROS) levels and exerted a protective effect against oxidative stress-induced cell death. This work provides the first functional study of TMEM97 in RPE and supports a potential role of TMEM97 in AMD pathobiology. Our study highlights the potential for using CRISPRi to study AMD genetics, and the CRISPRi RPE platform generated here provided a useful in vitro tool for functional studies of AMD-associated genes.

Extracellular vesicles (EVs) from stem cells have shown significant therapeutic potential to repair injured cardiac tissues and regulate pathological fibrosis. However, scalable generation of stem cells and derived EVs for clinical utility remains a huge technical challenge. Here, we report a rapid size-based extrusion strategy to generate EV-like membranous nanovesicles (NVs) from easily sourced human iPSCs in large quantities (yield 900× natural EVs). NVs isolated using density-gradient separation (buoyant density 1.13 g/mL) are spherical in shape and morphologically intact and readily internalised by human cardiomyocytes, primary cardiac fibroblasts, and endothelial cells. NVs captured the dynamic proteome of parental cells and include pluripotency markers (LIN28A, OCT4) and regulators of cardiac repair processes, including tissue repair (GJA1, HSP20/27/70, HMGB1), wound healing (FLNA, MYH9, ACTC1, ILK), stress response/translation initiation (eIF2S1/S2/S3/B4), hypoxia response (HMOX2, HSP90, GNB1), and extracellular matrix organization (ITGA6, MFGE8, ITGB1). Functionally, NVs significantly promoted tubule formation of endothelial cells (angiogenesis) (p < 0.05) and survival of cardiomyocytes exposed to low oxygen conditions (hypoxia) (p < 0.0001), as well as attenuated TGF-β mediated activation of cardiac fibroblasts (p < 0.0001). Quantitative proteome profiling of target cell proteome following NV treatments revealed upregulation of angiogenic proteins (MFGE8, MYH10, VDAC2) in endothelial cells and pro-survival proteins (CNN2, THBS1, IGF2R) in cardiomyocytes. In contrast, NVs attenuated TGF-β-driven extracellular matrix remodelling capacity in cardiac fibroblasts (ACTN1, COL1A1/2/4A2/12A1, ITGA1/11, THBS1). This study presents a scalable approach to generating functional NVs for cardiac repair.

Metabolism has been shown to alter cell fate in human pluripotent stem cells (hPSC). However, current understanding is almost exclusively based on work performed at 20% oxygen (air), with very few studies reporting on hPSC at physiological oxygen (5%). In this study, we integrated metabolic, transcriptomic, and epigenetic data to elucidate the impact of oxygen on hPSC. Using 13C-glucose labeling, we show that 5% oxygen increased the intracellular levels of glycolytic intermediates, glycogen, and the antioxidant response in hPSC. In contrast, 20% oxygen increased metabolite flux through the TCA cycle, activity of mitochondria, and ATP production. Acetylation of H3K9 and H3K27 was elevated at 5% oxygen while H3K27 trimethylation was decreased, conforming to a more open chromatin structure. RNA-seq analysis of 5% oxygen hPSC also indicated increases in glycolysis, lysine demethylases, and glucose-derived carbon metabolism, while increased methyltransferase and cell cycle activity was indicated at 20% oxygen. Our findings show that oxygen drives metabolite flux and specifies carbon fate in hPSC and, although the mechanism remains to be elucidated, oxygen was shown to alter methyltransferase and demethylase activity and the global epigenetic landscape.

Human induced pluripotent stem cells (iPSCs) can be differentiated in vitro into bona fide cardiomyocytes for disease modelling and personalized medicine. Mitochondrial morphology and metabolism change dramatically as iPSCs differentiate into mesodermal cardiac lineages. Inhibiting mitochondrial fission has been shown to promote cardiac differentiation of iPSCs. However, the effect of hydrazone M1, a small molecule that promotes mitochondrial fusion, on cardiac mesodermal commitment of human iPSCs is unknown. Here, we demonstrate that treatment with M1 promoted mitochondrial fusion in human iPSCs. Treatment of iPSCs with M1 during embryoid body formation significantly increased the percentage of beating embryoid bodies and expression of cardiac-specific genes. The pro-fusion and pro-cardiogenic effects of M1 were not associated with changes in expression of the α and β subunits of adenosine triphosphate (ATP) synthase. Our findings demonstrate for the first time that hydrazone M1 is capable of promoting cardiac differentiation of human iPSCs, highlighting the important role of mitochondrial dynamics in cardiac mesoderm lineage specification and cardiac development. M1 and other mitochondrial fusion promoters emerge as promising molecular targets to generate lineages of the heart from human iPSCs for patient-specific regenerative medicine.

Metabolism is central to embryonic stem cell (ESC) pluripotency and differentiation, with distinct profiles apparent under different nutrient milieu, and conditions that maintain alternate cell states. The significance of altered nutrient availability, particularly oxygen, and metabolic pathway activity has been highlighted by extensive studies of their impact on preimplantation embryo development, physiology, and viability. ESC similarly modulate their metabolism in response to altered metabolite levels, with changes in nutrient availability shown to have a lasting impact on derived cell identity through the regulation of the epigenetic landscape. Further, the preferential use of glucose and anaplerotic glutamine metabolism serves to not only support cell growth and proliferation but also minimise reactive oxygen species production. However, the perinuclear localisation of spherical, electron-poor mitochondria in ESC is proposed to sustain ESC nuclear-mitochondrial crosstalk and a mitochondrial-H2O2 presence, to facilitate signalling to support self-renewal through the stabilisation of HIFα, a process that may be favoured under physiological oxygen. The environment in which a cell is grown is therefore a critical regulator and determinant of cell fate, with metabolism, and particularly mitochondria, acting as an interface between the environment and the epigenome.

For the first time in history, an entire statewide public institution of higher education is extending all of its 16 campuses into the online virtual world of Second Life and is creating one of the largest virtual collaborative learning communities in the world. There are three levels of assessment and research for this initial-entry project: (1) at the System level; (2) at the individual campus level; and (3) at the course level. Questions to be addressed include: How can educators use virtual worlds to create large-scale learning communities? How do virtual worlds enhance collaborative and interdisciplinary learning? How are 16 geo-spatially separated campuses collaborating in virtual worlds? What have been major obstacles? How will this project continue to grow and evolve? The creation of the System's virtual collaborative learning community of students, faculty, researchers, and administrators has begun to allow participants to learn, share, collaborate, and grow alongside one another. The ethic of the model is generosity. (Contains 1 figure and 1 table.)

Virtual environments can resolve many practical and pedagogical challenges within higher education. Economic considerations, accessibility issues, and safety concerns can all be somewhat alleviated by creating learning activities in a virtual space. Because of the removal of real-world physical limitations like gravity, durability and scope, virtual space allows for an expansion of possibilities and approaches to knowledge transfer and discovery learning and becomes an "environment for information" rich with collaborative possibilities. Experimentation and participation in conceptual as well as applied projects is encouraged for both students and instructors. One of these virtual environments, Second Life, was used in a cross-disciplinary project for the creation of a Marine Science virtual class environment as an assignment for design students at a major southwestern research university in the United States. This paper reports on the findings from a project that utilized Second Life as a medium for enhancing and extending design education using a process of interdisciplinary collaboration.

This article previews the emergence of "homo virtualis." Drawing on data from seven research studies, peer-reviewed published research articles, and selected excerpts of 30 months of field notes taken in Second Life [SL], the article examines virtual learning environments and embodiment through the lens of interactions of avatars with other avatars, virtual objects, landscapes, sounds, and spatial constructs. Analysis is grounded in the polyvocal evidence provided by select participants who experienced a sense of embodied co-presence and connection with others across geo-physical distances. The discourse ranges from that of high school girls, professional retirees, toxicology and design undergraduates, interdisciplinary graduate students, to educators and researchers from K-12 through university full professors collaborating in SL. In an ecology of virtual contexts, learners inhabit a broader landscape of their own and others' making that allows them to be teachers, designers, researchers, communicators, and collaborators.

Substantial evidence now supports pedagogical applications of virtual worlds; however, most research supporting virtual worlds for education has been conducted using researcher-developed Multi-User Virtual Environments (MUVE). Second Life (SL) is a MUVE that has been adopted by a large number of academic institutions; however, little research has systematically investigated the potential of using SL for higher education. A 2-year pilot study was conducted that included seven SL activities and a mixed-methods evaluation of the SL's affordances, challenges, and limitations. Three SL integration factors emerged, each with sub-factors: pedagogical (relevance, complexity of required SL skills, use of SL affordances); contextual (student prior gaming experience, activity duration, frequency of events); and logistical (SL usability, training, technical support, computer issues). A framework for creating SL instructional activities is presented using the factors. (Contains 5 tables and 19 figures.)

While many reports espouse the potential impact that 3-D virtual worlds are expected to have on teaching and learning in higher education in a few years, there are few empirical studies that inform instructional design and learning assessment in virtual worlds. This study explores the nature and process of learning in Second Life in a graduate interdisciplinary communication course in fall 2007. Literature suggests that 3-D virtual worlds can be well suited for experiential learning environments. In this study, the actual instructional effectiveness of Second Life as an experiential learning environment for interdisciplinary communication is empirically examined using mixed research methods of journal content analysis, surveys, focus group, and virtual world snapshots and video. (Contains 2 figures.)

This paper presents an empirical study of how Second Life (SL) was utilized for a highly successful project-based graduate interdisciplinary communication course. Researchers found that an integrated threefold approach emphasizing project-based pedagogy, technical training and support, and assessment/research was effective in cultivating and understanding learning in SL. Based on research findings, a project-based application of SL that fully accommodates student experiential learning is recommended. It is suggested that teachers who are new To Whom It May Concern: SL and are preparing to "take the plunge" adopt a systematic "team approach" to integrate this trio of components. The students participating in this study formed an interdisciplinary team for their required SL project in which they chose to initiate a collaboration with the Basic Initiative and a group of architecture students to create a virtual presence for two green, sustainable, urban housing designs called the Alley Flats. Preliminary survey results suggest that the use of SL substantially enhanced the quality and experiences of student learning. Specific areas for improvement in future use are also identified. (Contains 9 figures.)

This article describes a university-sponsored experiential-based simulation, the NANO SCENARIO, to increase the public's awareness and affect attitudes on the societal implications of nanoscience and nanotechnology by bringing together diverse stakeholders' perspectives in a participatory learning environment. Nanotechnology has the potential for massive societal effects across all sectors, globally. Thus, new forms of education are needed to prepare members of society for making complex decisions about policy, governance, and values. Here, the authors examine the theoretical and conceptual framework of the simulation, which uses improvised role-play and perspective-taking to engage the public in a transformative learning experience. As part of this examination, they describe the uses and constraints of simulations with open-ended goals and without competition and prescribed rules. They believe educational role-play scenarios with active participation of the public can serve as a dynamic method for civic engagement across a range of complex, interdisciplinary topics and new technological dilemmas.

This article describes a university-sponsored experiential-based simulation, the NANO SCENARIO, to increase the public's awareness and affect attitudes on the societal implications of nanoscience and nanotechnology by bringing together diverse stakeholders' perspectives in a participatory learning environment. Nanotechnology has the potential for massive societal effects across all sectors, globally. Thus, new forms of education are needed to prepare members of society for making complex decisions about policy, governance, and values. Here, the authors examine the theoretical and conceptual framework of the simulation, which uses improvised role-play and perspective-taking to engage the public in a transformative learning experience. As part of this examination, they describe the uses and constraints of simulations with open-ended goals and without competition and prescribed rules. They believe educational role-play scenarios with active participation of the public can serve as a dynamic method for civic engagement across a range of complex, interdisciplinary topics and new technological dilemmas. (Contains 1 note.)

The goal of a university's human infrastructure is to make the faculty self-sufficient, life-long learners. Criteria for establishing a baseline faculty development program for technology include scaleability, functionality, sustainability, integration of existing resources, information competence, and access. Design and implementation principles should focus on the underlying long-term goals and the most effective means for reaching them. (AEF)

Findings of a questionnaire evaluating Blacks' and Whites' perceptions of each other indicated congruous views in some areas but suggest that Whites are more negative toward interracial relations than they perceive themselves to be and that both groups respond to relationships according to their degree of sensitivity or group interest. (MK)

Tissue flaps are used to cover large/poorly healing wounds, but involve complex surgery and donor site morbidity. In this study a tissue flap is assembled using the mammalian body as a bioreactor to functionally connect an artery and vein to a human capillary network assembled from induced pluripotent stem cell-derived endothelial cells (hiPSC ECs). In vitro : Porous NovoSorb™ scaffolds (3 mm × 1.35 mm) were seeded with 200,000 hiPSC ECs ± 100,000 human vascular smooth muscle cells (hvSMC), and cultured for 1–3 days, with capillaries formed by 24 h which were CD31+, VE-Cadherin+, EphB4+, VEGFR2+ and Ki67+, whilst hvSMCs (calponin+) attached abluminally. In vivo : In SCID mice, bi-lateral epigastric vascular pedicles were isolated in a silicone chamber for a 3 week 'delay period' for pedicle capillary sprouting, then reopened, and two hiPSC EC ± hvSMCs seeded scaffolds transplanted over the pedicle. The chamber was either resealed (Group 1), or removed and surrounding tissue secured around the pedicle + scaffolds (Group 2), for 1 or 2 weeks. Human capillaries survived in vivo and were CD31+, VE-Cadherin+ and VEGFR2+. Human vSMCs remained attached, and host mesenchymal cells also attached abluminally. Systemically injected FITC-dextran present in human capillary lumens indicated inosculation to host capillaries. Human iPSC EC capillary morphometric parameters at one week in vivo were equal to or higher than the same parameters measured in human abdominal skin. This 'proof of concept' study has demonstrated that bio-engineering an autologous human tissue flap based on hiPSC EC could minimize the use of donor flaps and has potential applications for complex wound coverage. Tissue flaps, used for surgical reconstruction of wounds, require complex surgery, often associated with morbidity. Bio-engineering a simpler alternative, we assembled a human induced pluripotent stem cell derived endothelial cell (hiPSC ECs) capillary network in a porous scaffold in vitro , which when transplanted over a mouse vascular pedicle in vivo formed a functional tissue flap with mouse blood flow in the human capillaries. Therefore it is feasible to form an autologous tissue flap derived from a hiPSC EC capillary network assembled in vitro, and functionally connect to a vascular pedicle in vivo that could be utilized in complex wound repair for chronic or acute wounds. [ABSTRACT FROM AUTHOR]

This article describes a city-level economic development policymaking network. To do this, we identify the key participants in the network, derived both from the scholarly literature and experience. We then outline the network's structure. We refer to that structure as the All-Channel Star Network. An examination of this network structure will involve a consideration of interactions between network participants and leadership within the larger city economic development network. [ABSTRACT FROM AUTHOR]

For public meetings in general, and for the North Omaha Development Project Meeting in particular, a Conversation Analysis (CA) approach asks how participants in public meetings involved in decision-making communicate and display their trust and mistrust of one another. From a CA perspective, while it is important to examine ways to cultivate greater trust in the public arena, research is also warranted that looks closely at people's observable talk and behavior to better understand how the qualities of trust and mistrust are enacted by participants and how others orient to those displays. Power differences are critical, and whether the organizers are controlling the agenda or whether the attendees are attempting to influence the meeting's procedural mechanisms, these are behaviors that must in the end be performed as the proceedings unfold. Displays of observable behavior are subject to empirical analysis and are the focus of what a CA approach can contribute to the study of public meetings. [ABSTRACT FROM AUTHOR]

This special issue of IJP2 attempts to build linkages between public participation scholarship and communication research that emphasizes close attention to naturally-occurring interaction. The essays all investigate different aspects of the communication that occurs during one public meeting: a public forum that focused on issues of economic development in Omaha, Nebraska. Through their investigation of this common case, the essays in this issue provide detailed description of some communication processes common to public meetings such as nonverbal communication, question and answer behavior, storytelling, the use of the term "community," and the terms people use to talk about their own communication. These studies highlight how such interactive practices function in important ways to build or challenge notions of community, frame the purpose and outcome of the meeting, display power differences among participants, and clarify key community values. This collection of essays highlights how close attention to what happens during public meetings can have important implications for both the theory and practice of public participation. A full video of the meeting is available online. [ABSTRACT FROM AUTHOR]

The article critically assessing Joseph S. Himes, a sociologist, major works over the span of his professional career, divides his writings into early and late periods in order to delineate the distinctive theoretical influences on his writings and to disclose the discrete approaches emerging in eath. Although Himes has contributed to the sociological literature on racial and social conflict for ova four decades, it has only been in recent years that his contribution to sociology has gained the attention of his colleagues. His most significant contribution occurs in the tatter phase of the second period, the phase in which his writings reflect racial and social conflicts as principal theoretical questions. In this phase, Himes adopts Simmeliane Coser conflict categories as deductive nomological explanations for his own approach to racial conflict. The final distinctive development in his writing involves his effort to expand and synthesize conflict categories into a more general predictive model of social conflict.