Community-developed checklists for publishing images and image analyses.

Images document scientific discoveries and are prevalent in modern biomedical research. Microscopy imaging in particular is currently undergoing rapid technological advancements. However, for scientists wishing to publish obtained images and image-analysis results, there are currently no unified guidelines for best practices. Consequently, microscopy images and image data in publications may be unclear or difficult to interpret. Here, we present community-developed checklists for preparing light microscopy images and describing image analyses for publications. These checklists offer authors, readers and publishers key recommendations for image formatting and annotation, color selection, data availability and reporting image-analysis workflows. The goal of our guidelines is to increase the clarity and reproducibility of image figures and thereby to heighten the quality and explanatory power of microscopy data.

Recognising the importance and impact of Imaging Scientists: Global guidelines for establishing career paths within core facilities.

In the dynamic landscape of scientific research, imaging core facilities are vital hubs propelling collaboration and innovation at the technology development and dissemination frontier. Here, we present a collaborative effort led by Global BioImaging (GBI), introducing international recommendations geared towards elevating the careers of Imaging Scientists in core facilities. Despite the critical role of Imaging Scientists in modern research ecosystems, challenges persist in recognising their value, aligning performance metrics and providing avenues for career progression and job security. The challenges encompass a mismatch between classic academic career paths and service-oriented roles, resulting in a lack of understanding regarding the value and impact of Imaging Scientists and core facilities and how to evaluate them properly. They further include challenges around sustainability, dedicated training opportunities and the recruitment and retention of talent. Structured across these interrelated sections, the recommendations within this publication aim to propose globally applicable solutions to navigate these challenges. These recommendations apply equally to colleagues working in other core facilities and research institutions through which access to technologies is facilitated and supported. This publication emphasises the pivotal role of Imaging Scientists in advancing research programs and presents a blueprint for fostering their career progression within institutions all around the world.

Building a FAIR image data ecosystem for microscopy communities.

Bioimaging has now entered the era of big data with faster-than-ever development of complex microscopy technologies leading to increasingly complex datasets. This enormous increase in data size and informational complexity within those datasets has brought with it several difficulties in terms of common and harmonized data handling, analysis, and management practices, which are currently hampering the full potential of image data being realized. Here, we outline a wide range of efforts and solutions currently being developed by the microscopy community to address these challenges on the path towards FAIR bioimaging data. We also highlight how different actors in the microscopy ecosystem are working together, creating synergies that develop new approaches, and how research infrastructures, such as Euro-BioImaging, are fostering these interactions to shape the field.

Planarian regeneration as a model of anatomical homeostasis: Recent progress in biophysical and computational approaches.

Planarian behavior, physiology, and pattern control offer profound lessons for regenerative medicine, evolutionary biology, morphogenetic engineering, robotics, and unconventional computation. Despite recent advances in the molecular genetics of stem cell differentiation, this model organism's remarkable anatomical homeostasis provokes us with truly fundamental puzzles about the origin of large-scale shape and its relationship to the genome. In this review article, we first highlight several deep mysteries about planarian regeneration in the context of the current paradigm in this field. We then review recent progress in understanding of the physiological control of an endogenous, bioelectric pattern memory that guides regeneration, and how modulating this memory can permanently alter the flatworm's target morphology. Finally, we focus on computational approaches that complement reductive pathway analysis with synthetic, systems-level understanding of morphological decision-making. We analyze existing models of planarian pattern control and highlight recent successes and remaining knowledge gaps in this interdisciplinary frontier field.

Nervous system and tissue polarity dynamically adapt to new morphologies in planaria.

The coordination of tissue-level polarity with organism-level polarity is crucial in development, disease, and regeneration. Here, we characterize a new example of large-scale control of dynamic remodeling of body polarity. Exploiting the flexibility of the body plan in regenerating planarians, we used mirror duplication of the primary axis to show how established tissue-level polarity adapts to new organism-level polarity. Characterization of epithelial planar cell polarity revealed a remarkable reorientation of tissue polarity in double-headed planarians. This reorientation of cilia occurs even following irradiation-induced loss of all stem cells, suggesting independence of the polarity change from the formation of new cells. The presence of the two heads plays an important role in regulating the rate of change in overall polarity. We further present data that suggest that the nervous system itself adapts its polarity to match the new organismal anatomy as revealed by changes in nerve transport driving distinct regenerative outcomes. Thus, in planaria tissue-level polarity can dynamically reorient to match the organism-level anatomical configuration.

Morphological Coordination: A Common Ancestral Function Unifying Neural and Non-Neural Signaling.

Nervous systems are traditionally thought of as providing sensing and behavioral coordination functions at the level of the whole organism. What is the evolutionary origin of the mechanisms enabling the nervous systems' information processing ability? Here, we review evidence from evolutionary, developmental, and regenerative biology suggesting a deeper, ancestral function of both pre-neural and neural cell-cell communication systems: the long-distance coordination of cell division and differentiation required to create and maintain body-axis symmetries. This conceptualization of the function of nervous system activity sheds new light on the evolutionary transition from the morphologically rudimentary, non-neural Porifera and Placazoa to the complex morphologies of Ctenophores, Cnidarians, and Bilaterians. It further allows a sharp formulation of the distinction between long-distance axis-symmetry coordination based on external coordinates, e.g., by whole-organism scale trophisms as employed by plants and sessile animals, and coordination based on body-centered coordinates as employed by motile animals. Thus we suggest that the systems that control animal behavior evolved from ancient mechanisms adapting preexisting ionic and neurotransmitter mechanisms to regulate individual cell behaviors during morphogenesis. An appreciation of the ancient, non-neural origins of bioelectrically mediated computation suggests new approaches to the study of embryological development, including embryological dysregulation, cancer, regenerative medicine, and synthetic bioengineering.

QUAREP-LiMi: A community-driven initiative to establish guidelines for quality assessment and reproducibility for instruments and images in light microscopy.

A modern day light microscope has evolved from a tool devoted to making primarily empirical observations to what is now a sophisticated , quantitative device that is an integral part of both physical and life science research. Nowadays, microscopes are found in nearly every experimental laboratory. However, despite their prevalent use in capturing and quantifying scientific phenomena, neither a thorough understanding of the principles underlying quantitative imaging techniques nor appropriate knowledge of how to calibrate, operate and maintain microscopes can be taken for granted. This is clearly demonstrated by the well-documented and widespread difficulties that are routinely encountered in evaluating acquired data and reproducing scientific experiments. Indeed, studies have shown that more than 70% of researchers have tried and failed to repeat another scientist's experiments, while more than half have even failed to reproduce their own experiments. One factor behind the reproducibility crisis of experiments published in scientific journals is the frequent underreporting of imaging methods caused by a lack of awareness and/or a lack of knowledge of the applied technique. Whereas quality control procedures for some methods used in biomedical research, such as genomics (e.g. DNA sequencing, RNA-seq) or cytometry, have been introduced (e.g. ENCODE), this issue has not been tackled for optical microscopy instrumentation and images. Although many calibration standards and protocols have been published, there is a lack of awareness and agreement on common standards and guidelines for quality assessment and reproducibility. In April 2020, the QUality Assessment and REProducibility for instruments and images in Light Microscopy (QUAREP-LiMi) initiative was formed. This initiative comprises imaging scientists from academia and industry who share a common interest in achieving a better understanding of the performance and limitations of microscopes and improved quality control (QC) in light microscopy. The ultimate goal of the QUAREP-LiMi initiative is to establish a set of common QC standards, guidelines, metadata models and tools, including detailed protocols, with the ultimate aim of improving reproducible advances in scientific research. This White Paper (1) summarizes the major obstacles identified in the field that motivated the launch of the QUAREP-LiMi initiative; (2) identifies the urgent need to address these obstacles in a grassroots manner, through a community of stakeholders including, researchers, imaging scientists, bioimage analysts, bioimage informatics developers, corporate partners, funding agencies, standards organizations, scientific publishers and observers of such; (3) outlines the current actions of the QUAREP-LiMi initiative and (4) proposes future steps that can be taken to improve the dissemination and acceptance of the proposed guidelines to manage QC. To summarize, the principal goal of the QUAREP-LiMi initiative is to improve the overall quality and reproducibility of light microscope image data by introducing broadly accepted standard practices and accurately captured image data metrics.

Neural control of body-plan axis in regenerating planaria.

Control of axial polarity during regeneration is a crucial open question. We developed a quantitative model of regenerating planaria, which elucidates self-assembly mechanisms of morphogen gradients required for robust body-plan control. The computational model has been developed to predict the fraction of heteromorphoses expected in a population of regenerating planaria fragments subjected to different treatments, and for fragments originating from different regions along the anterior-posterior and medio-lateral axis. This allows for a direct comparison between computational and experimental regeneration outcomes. Vector transport of morphogens was identified as a fundamental requirement to account for virtually scale-free self-assembly of the morphogen gradients observed in planarian homeostasis and regeneration. The model correctly describes altered body-plans following many known experimental manipulations, and accurately predicts outcomes of novel cutting scenarios, which we tested. We show that the vector transport field coincides with the alignment of nerve axons distributed throughout the planarian tissue, and demonstrate that the head-tail axis is controlled by the net polarity of neurons in a regenerating fragment. This model provides a comprehensive framework for mechanistically understanding fundamental aspects of body-plan regulation, and sheds new light on the role of the nervous system in directing growth and form.

The Role of Early Bioelectric Signals in the Regeneration of Planarian Anterior/Posterior Polarity.

Axial patterning during planarian regeneration relies on a transcriptional circuit that confers distinct positional information on the two ends of an amputated fragment. The earliest known elements of this system begin demarcating differences between anterior and posterior wounds by 6 h postamputation. However, it is still unknown what upstream events break the axial symmetry, allowing a mutual repressor system to establish invariant, distinct biochemical states at the anterior and posterior ends. Here, we show that bioelectric signaling at 3 h is crucial for the formation of proper anterior-posterior polarity in planaria. Briefly manipulating the endogenous bioelectric state by depolarizing the injured tissue during the first 3 h of regeneration alters gene expression by 6 h postamputation and leads to a double-headed phenotype upon regeneration despite confirmed washout of ionophores from tissue. These data reveal a primary functional role for resting membrane potential taking place within the first 3 h after injury and kick-starting the downstream pattern of events that elaborate anatomy over the following 10 days. We propose a simple model of molecular-genetic mechanisms to explain how physiological events taking place immediately after injury regulate the spatial distribution of downstream gene expression and anatomy of regenerating planaria.

Cytoplasmic flows in starfish oocytes are fully determined by cortical contractions.

Cytoplasmic flows are an ubiquitous feature of biological systems, in particular in large cells, such as oocytes and eggs in early animal development. Here we show that cytoplasmic flows in starfish oocytes, which can be imaged well with transmission light microscopy, are fully determined by the cortical dynamics during surface contraction waves. We first show that the dynamics of the oocyte surface is highly symmetric around the animal-vegetal axis. We then mathematically solve the Stokes equation for flows inside a deforming sphere using the measured surface displacements as boundary conditions. Our theoretical predictions agree very well with the intracellular flows quantified by particle image velocimetry, proving that during this stage the starfish cytoplasm behaves as a simple Newtonian fluid on the micrometer scale. We calculate the pressure field inside the oocyte and find that its gradient is too small as to explain polar body extrusion, in contrast to earlier suggestions. Myosin II inhibition by blebbistatin confirms this conclusion, because it diminishes cell shape changes and hydrodynamic flow, but does not abolish polar body formation.

In inferior myocardial infarction, neither ST elevation in lead V1 nor ST depression in lead I are reliable findings for the diagnosis of right ventricular infarction.

OBJECTIVE: In the presence of inferior myocardial infarction (MI), ST depression (STD) in lead I has been claimed to be accurate for diagnosis of right ventricular (RV) MI. We sought to evaluate this claim and also whether ST Elevation (STE) in lead V1 would be helpful, with or without STD in V2. METHODS: Retrospective study of consecutive inferior STEMI, comparing ECGs of patients with, to those without, RVMI, as determined by angiographic coronary occlusion proximal to the RV marginal branch. STE and STD were measured at the J-point, relative to the PQ junction. The primary outcomes were sensitivity/specificity of 1) STD in lead I ≥ 0.5 mm and 2) STE in lead V1 ≥ 0.5 mm, stratified by presence or absence of posterior (inferobasal) MI, as determined by ≥0.5 mm STD in lead V2, for differentiating RVMI from non-RVMI. RESULTS: Of 149 patients with inferior STEMI, 43 (29%) had RVMI and 106 (71%) did not. There was no difference in the presence or absence of at least 0.5 mm STD in Lead I between patients with (37/43, 86%) vs. without RVMI (85/106, 80%, p = 0.56). In those with, vs. without, RVMI, (15/43, 35%) had STE in V1, versus (17/106, 16%) (p = 0.015). Specificity of STE in V1 for RVMI was 84%; sensitivity was 35%. Sensitivity was higher without (69%), than with (35%), STD in V2. CONCLUSION: Among inferior STEMI, the presence of any ST depression in lead I does not help to diagnose RVMI. ST elevation ≥0.5 mm in lead V1 is specific for RVMI, and moderately sensitive only if concomitant STD ≥ 0.5 mm in V2 is not present. Although STE in V1 is quite specific, overall the diagnostic characteristics of the standard 12­lead ECG are inadequate to definitively diagnose, or exclude, RVMI, as defined angiographically.

Discharge Glucose Is Not Associated With Short-Term Adverse Outcomes in Emergency Department Patients With Moderate to Severe Hyperglycemia.

STUDY OBJECTIVE: Hyperglycemia is frequently encountered in the emergency department (ED), and there is no consensus on optimal care before discharge. The importance of glucose reduction in the ED is unknown. We seek to determine whether an association exists between discharge glucose and 7-day adverse outcomes. METHODS: A cohort design with retrospective chart review was conducted at a high-volume urban ED. Patients were included if any glucose level was greater than or equal to 400 mg/dL and they were discharged from the ED. Generalized estimating equation models were created for the 7-day outcomes with a primary predictor of discharge glucose. RESULTS: The cohort consisted of 422 patients with 566 ED encounters. Mean arrival and discharge glucose were 491 mg/dL (SD 82 mg/dL) and 334 mg/dL (SD 101 mg/dL), respectively. In the 7-day follow-up period, 62 (13%) and 36 (7%) patients had a repeat ED visit for hyperglycemia and were hospitalized, respectively. Two patients had diabetic ketoacidosis. After adjustment for arrival glucose, whether a chemistry panel was obtained, amount of intravenous fluids administered, and amount of subcutaneous insulin administered, discharge glucose was not associated with repeat ED visit for hyperglycemia (adjusted odds ratio 0.997; 95% confidence interval 0.993 to 1.001) or hospitalization for any reason (adjusted odds ratio 0.998; 95% confidence interval 0.995 to 1.002). CONCLUSION: ED discharge glucose in patients with moderate to severe hyperglycemia was not associated with 7-day outcomes of repeat ED visit for hyperglycemia or hospitalization. Attaining a specific glucose goal before discharge in patients with hyperglycemia may be less important than traditionally thought.

ST depression in lead aVL differentiates inferior ST-elevation myocardial infarction from pericarditis.

BACKGROUND: ST-segment elevation (STE) due to inferior STE myocardial infarction (STEMI) may be misdiagnosed as pericarditis. Conversely, this less life-threatening etiology of ST elevation may be confused for inferior STEMI. We sought to determine if the presence of any ST-segment depression in lead aVL would differentiate inferior STEMI from pericarditis. METHODS: Retrospective study of 3 populations. Cohort 1 included patients coded as inferior STEMI, cohort 2 included patients with a discharge diagnosis of pericarditis who presented with chest pain and at least 0.5 mm of ST elevation in at least 1 inferior lead. We analyzed the presenting electrocardiogram in both populations, with careful assessment of leads II, III, aVF, and aVL. In addition, we retrospectively studied a third cohort of patients with subtle inferior STEMI (<1-mm STE with occluded artery on catheterization) and assessed the sensitivity of ST depression in lead aVL for this group. RESULTS: Of 154 inferior STEMI patients, 154 had some amount of ST depression in lead aVL (100%; confidence interval, 98%-100%). Of the 49 electrocardiograms in the pericarditis group, all 49 had some inferior STE but none had any ST-segment depression in lead aVL (specificity, 100%; confidence interval, 91%-100%). In the third cohort, there were 272 inferior MIs with coronary occlusion, of which 54 were "subtle." Of these, 49 had some ST depression in lead aVL. CONCLUSION: When there is inferior ST-segment elevation, the presence of any ST depression in lead aVL is highly sensitive for coronary occlusion in inferior myocardial infarction and very specific for differentiating inferior myocardial infarction from pericarditis.

The scaling of goals from cellular to anatomical homeostasis: an evolutionary simulation, experiment and analysis.

Complex living agents consist of cells, which are themselves competent sub-agents navigating physiological and metabolic spaces. Behaviour science, evolutionary developmental biology and the field of machine intelligence all seek to understand the scaling of biological cognition: what enables individual cells to integrate their activities to result in the emergence of a novel, higher-level intelligence with large-scale goals and competencies that belong to it and not to its parts? Here, we report the results of simulations based on the TAME framework, which proposes that evolution pivoted the collective intelligence of cells during morphogenesis of the body into traditional behavioural intelligence by scaling up homeostatic competencies of cells in metabolic space. In this article, we created a minimal in silico system (two-dimensional neural cellular automata) and tested the hypothesis that evolutionary dynamics are sufficient for low-level setpoints of metabolic homeostasis in individual cells to scale up to tissue-level emergent behaviour. Our system showed the evolution of the much more complex setpoints of cell collectives (tissues) that solve a problem in morphospace: the organization of a body-wide positional information axis (the classic French flag problem in developmental biology). We found that these emergent morphogenetic agents exhibit a number of predicted features, including the use of stress propagation dynamics to achieve the target morphology as well as the ability to recover from perturbation (robustness) and long-term stability (even though neither of these was directly selected for). Moreover, we observed an unexpected behaviour of sudden remodelling long after the system stabilizes. We tested this prediction in a biological system-regenerating planaria-and observed a very similar phenomenon. We propose that this system is a first step towards a quantitative understanding of how evolution scales minimal goal-directed behaviour (homeostatic loops) into higher-level problem-solving agents in morphogenetic and other spaces.

Community-developed checklists for publishing images and image analyses.

Images document scientific discoveries and are prevalent in modern biomedical research. Microscopy imaging in particular is currently undergoing rapid technological advancements. However for scientists wishing to publish the obtained images and image analyses results, there are to date no unified guidelines. Consequently, microscopy images and image data in publications may be unclear or difficult to interpret. Here we present community-developed checklists for preparing light microscopy images and image analysis for publications. These checklists offer authors, readers, and publishers key recommendations for image formatting and annotation, color selection, data availability, and for reporting image analysis workflows. The goal of our guidelines is to increase the clarity and reproducibility of image figures and thereby heighten the quality and explanatory power of microscopy data is in publications.

Regulation of axial and head patterning during planarian regeneration by a commensal bacterium.

Some animals, such as planaria, can regenerate complex anatomical structures in a process regulated by genetic and biophysical factors, but additional external inputs into regeneration remain to be uncovered. Microbial communities inhabiting metazoan organisms are important for metabolic, immune, and disease processes, but their instructive influence over host structures remains largely unexplored. Here, we show that Aquitalea sp. FJL05, an endogenous commensal bacterium of Dugesia japonica planarians, and one of the small molecules it produces, indole, can influence axial and head patterning during regeneration, leading to regeneration of permanently two-headed animals. Testing the impact of indole on planaria tissues via RNA sequencing, we find that indole alters the regenerative outcomes in planarians through changes in expression to patterning genes, including a downregulation of Wnt pathway genes. These data provide a unique example of the product of a commensal bacterium modulating transcription of patterning genes to affect the host's anatomical structure during regeneration.