Heparin-binding epidermal growth factor and fibroblast growth factor 2 rescue Müller glia-derived progenitor cell formation in microglia- and macrophage-ablated chick retinas.

Recent studies have demonstrated the impact of pro-inflammatory signaling and reactive microglia/macrophages on the formation of Müller glial-derived progenitor cells (MGPCs) in the retina. In chick retina, ablation of microglia/macrophages prevents the formation of MGPCs. Analyses of single-cell RNA-sequencing chick retinal libraries revealed that quiescent and activated microglia/macrophages have a significant impact upon the transcriptomic profile of Müller glia (MG). In damaged monocyte-depleted retinas, MG fail to upregulate genes related to different cell signaling pathways, including those related to Wnt, heparin-binding epidermal growth factor (HBEGF), fibroblast growth factor (FGF) and retinoic acid receptors. Inhibition of GSK3ß, to simulate Wnt signaling, failed to rescue the deficit in MGPC formation, whereas application of HBEGF or FGF2 completely rescued the formation of MGPCs in monocyte-depleted retinas. Inhibition of Smad3 or activation of retinoic acid receptors partially rescued the formation of MGPCs in monocyte-depleted retinas. We conclude that signals produced by reactive microglia/macrophages in damaged retinas stimulate MG to upregulate cell signaling through HBEGF, FGF and retinoic acid, and downregulate signaling through TGFß/Smad3 to promote the reprogramming of MG into proliferating MGPCs.

Fatty acid-binding proteins and fatty acid synthase influence glial reactivity and promote the formation of Müller glia-derived progenitor cells in the chick retina.

A recent comparative transcriptomic study of Müller glia (MG) in vertebrate retinas revealed that fatty acid binding proteins (FABPs) are among the most highly expressed genes in chick ( Hoang et al., 2020). Here, we investigate how FABPs and fatty acid synthase (FASN) influence glial cells in the chick retina. During development, FABP7 is highly expressed by retinal progenitor cells and maturing MG, whereas FABP5 is upregulated in maturing MG. PMP2 (FABP8) is expressed by oligodendrocytes and FABP5 is expressed by non-astrocytic inner retinal glial cells, and both of these FABPs are upregulated by activated MG. In addition to suppressing the formation of Müller glia-derived progenitor cells (MGPCs), we find that FABP-inhibition suppresses the proliferation of microglia. FABP-inhibition induces distinct changes in single cell transcriptomic profiles, indicating transitions of MG from resting to reactive states and suppressed MGPC formation, with upregulation of gene modules for gliogenesis and decreases in neurogenesis. FASN-inhibition increases the proliferation of microglia and suppresses the formation of MGPCs. We conclude that fatty acid metabolism and cell signaling involving fatty acids are important in regulating the reactivity and dedifferentiation of MG, and the proliferation of microglia and MGPCs.

Chromatin access regulates the formation of Müller glia-derived progenitor cells in the retina.

Chromatin access and epigenetic control over gene expression play important roles in regulating developmental processes. However, little is known about how chromatin access and epigenetic gene silencing influence mature glial cells and retinal regeneration. Herein, we investigate the expression and functions of S-adenosylhomocysteine hydrolase (SAHH; AHCY) and histone methyltransferases (HMTs) during the formation of Müller glia (MG)-derived progenitor cells (MGPCs) in the chick and mouse retinas. In chick, AHCY, AHCYL1 and AHCYL2, and many different HMTs are dynamically expressed by MG and MGPCs in damaged retinas. Inhibition of SAHH reduced levels of H3K27me3 and potently blocks the formation of proliferating MGPCs. By using a combination of single cell RNA-seq and single cell ATAC-seq, we find significant changes in gene expression and chromatin access in MG with SAHH inhibition and NMDA-treatment; many of these genes are associated with glial and neuronal differentiation. A strong correlation across gene expression, chromatin access, and transcription factor motif access in MG was observed for transcription factors known to convey glial identity and promote retinal development. By comparison, in the mouse retina, inhibition of SAHH has no influence on the differentiation of neuron-like cells from Ascl1-overexpressing MG. We conclude that in the chick the activity of SAHH and HMTs are required for the reprogramming of MG into MGPCs by regulating chromatin access to transcription factors associated with glial differentiation and retinal development.

Detecting and Characterizing Inferior Vena Cava Filters on Abdominal Computed Tomography with Data-Driven Computational Frameworks.

Two data-driven algorithms were developed for detecting and characterizing Inferior Vena Cava (IVC) filters on abdominal computed tomography to assist healthcare providers with the appropriate management of these devices to decrease complications: one based on 2-dimensional data and transfer learning (2D + TL) and an augmented version of the same algorithm which accounts for the 3-dimensional information leveraging recurrent convolutional neural networks (3D + RCNN). The study contains 2048 abdominal computed tomography studies obtained from 439 patients who underwent IVC filter placement during the 10-year period from January 1st, 2009, to January 1st, 2019. Among these, 399 patients had retrievable filters, and 40 had non-retrievable filter types. The reference annotations for the filter location were obtained through a custom-developed interface. The ground truth annotations for the filter types were determined based on the electronic medical record and physician review of imaging. The initial stage of the framework returns a list of locations containing metallic objects based on the density of the structure. The second stage processes the candidate locations and determines which one contains an IVC filter. The final stage of the pipeline classifies the filter types as retrievable vs. non-retrievable. The computational models are trained using Tensorflow Keras API on an Nvidia Quadro GV100 system. We utilized a fine-tuning supervised training strategy to conduct our experiments. We find that the system achieves high sensitivity on detecting the filter locations with a high confidence value. The 2D + TL model achieved a sensitivity of 0.911 and a precision of 0.804, and the 3D + RCNN model achieved a sensitivity of 0.923 and a precision of 0.853 for filter detection. The system confidence for the IVC location predictions is high: 0.993 for 2D + TL and 0.996 for 3D + RCNN. The filter type prediction component of the system achieved 0.945 sensitivity, 0.882 specificity, and 0.97 AUC score with 2D + TL and 0. 940 sensitivity, 0.927 specificity, and 0.975 AUC score with 3D + RCNN. With the intent to create tools to improve patient outcomes, this study describes the initial phase of a computational framework to support healthcare providers in detecting patients with retained IVC filters, so an individualized decision can be made to remove these devices when appropriate, to decrease complications. To our knowledge, this is the first study that curates abdominal computed tomography (CT) scans and presents an algorithm for automated detection and characterization of IVC filters.

Traumatic Brain Injury Causes Chronic Cortical Inflammation and Neuronal Dysfunction Mediated by Microglia.

Traumatic brain injury (TBI) can lead to significant neuropsychiatric problems and neurodegenerative pathologies, which develop and persist years after injury. Neuroinflammatory processes evolve over this same period. Therefore, we aimed to determine the contribution of microglia to neuropathology at acute [1 d postinjury (dpi)], subacute (7 dpi), and chronic (30 dpi) time points. Microglia were depleted with PLX5622, a CSF1R antagonist, before midline fluid percussion injury (FPI) in male mice and cortical neuropathology/inflammation was assessed using a neuropathology mRNA panel. Gene expression associated with inflammation and neuropathology were robustly increased acutely after injury (1 dpi) and the majority of this expression was microglia independent. At 7 and 30 dpi, however, microglial depletion reversed TBI-related expression of genes associated with inflammation, interferon signaling, and neuropathology. Myriad suppressed genes at subacute and chronic endpoints were attributed to neurons. To understand the relationship between microglia, neurons, and other glia, single-cell RNA sequencing was completed 7 dpi, a critical time point in the evolution from acute to chronic pathogenesis. Cortical microglia exhibited distinct TBI-associated clustering with increased type-1 interferon and neurodegenerative/damage-related genes. In cortical neurons, genes associated with dopamine signaling, long-term potentiation, calcium signaling, and synaptogenesis were suppressed. Microglial depletion reversed the majority of these neuronal alterations. Furthermore, there was reduced cortical dendritic complexity 7 dpi, reduced neuronal connectively 30 dpi, and cognitive impairment 30 dpi. All of these TBI-associated functional and behavioral impairments were prevented by microglial depletion. Collectively, these studies indicate that microglia promote persistent neuropathology and long-term functional impairments in neuronal homeostasis after TBI.SIGNIFICANCE STATEMENT Millions of traumatic brain injuries (TBIs) occur in the United States alone each year. Survivors face elevated rates of cognitive and psychiatric complications long after the inciting injury. Recent studies of human brain injury link chronic neuroinflammation to adverse neurologic outcomes, suggesting that evolving inflammatory processes may be an opportunity for intervention. Here, we eliminate microglia to compare the effects of diffuse TBI on neurons in the presence and absence of microglia and microglia-mediated inflammation. In the absence of microglia, neurons do not undergo TBI-induced changes in gene transcription or structure. Microglial elimination prevented TBI-induced cognitive changes 30 d postinjury (dpi). Therefore, microglia have a critical role in disrupting neuronal homeostasis after TBI, particularly at subacute and chronic timepoints.

A comparison of the behaviour and survival of angling vs. trap-sampled Salmo salar smolts.

An experiment was undertaken, using acoustic telemetry, to compare the survival and migratory timing of Salmo salar L. smolts sampled, under optimal conditions, in a traditional fixed Wolf trap against a sample of rod-caught fish captured using a sensitive angling technique. No significant difference was evident in survival with 83% of both samples detected in the river outflow, 67% of the trap and 76% of the rod samples were detected in coastal waters and finally 43% of the trap and 35% of the rod samples were detected on an offshore array c. 50 km from the river outlet. No significant difference was evident in the time taken for trap- and rod-sampled fish to reach either the river outflow, coastal or offshore waters. Angling, if undertaken sensitively, can provide an effective, resource-efficient and ethically justifiable sampling tool for juvenile salmonid age classes.

Cannabinoid signaling promotes the de-differentiation and proliferation of Müller glia-derived progenitor cells.

Endocannabinoids (eCB) are lipid-based neurotransmitters that are known to influence synaptic function in the visual system. eCBs are also known to suppress neuroinflammation in different pathological states. However, nothing is known about the roles of the eCB system during the transition of Müller glia (MG) into proliferating progenitor-like cells in the retina. Accordingly, we used the chick and mouse model to characterize expression patterns of eCB-related genes and applied pharmacological agents to investigate how the eCB system impacts glial reactivity and the capacity of MG to become Müller glia-derived progenitor cells (MGPCs). We queried single cell RNA-seq libraries to identify eCB-related genes and identify cells with dynamic patterns of expression in damaged retinas. MG and inner retinal neurons expressed the eCB receptor CNR1, as well as enzymes involved in eCB metabolism. In the chick, intraocular injections of cannabinoids, 2-Arachidonoylglycerol (2-AG) and Anandamide (AEA), stimulated the formation of MGPCs. Cannabinoid Receptor 1 (CNR1)-agonists and Monoglyceride Lipase-inhibitor promoted the formation of MGPCs, whereas CNR1-antagonist and inhibitors of eCB synthesis suppressed this process. In damaged mouse retinas where MG activate NFkB-signaling, activation of CNR1 decreased and inhibition of CNR1 increased NFkB, whereas levels of neuronal cell death were unaffected. Surprisingly, retinal microglia were largely unaffected by increases or decreases in eCB-signaling in both chick and mouse retinas. We conclude that the eCB system in the retina influences the reactivity of MG and the formation of proliferating MGPCs, but does not influence the reactivity of immune cells in the retina.

Midkine is neuroprotective and influences glial reactivity and the formation of Müller glia-derived progenitor cells in chick and mouse retinas.

Recent studies suggest midkine (MDK) is involved in the development and regeneration of the zebrafish retina. We investigate the expression patterns of MDK and related factors, roles in neuronal survival, and influence upon the formation of Müller glia-derived progenitor cells (MGPCs) in chick and mouse model systems. By using single-cell RNA-sequencing, we find that MDK and pleiotrophin (PTN), a MDK-related cytokine, are upregulated by Müller glia (MG) during later stages of development in chick. While PTN is downregulated, MDK is dramatically upregulated in mature MG after retinal damage or FGF2 and insulin treatment. By comparison, MDK and PTN are downregulated by MG in damaged mouse retinas. In both chick and mouse retinas, exogenous MDK induces expression of cFos and pS6 in MG. In the chick, MDK significantly decreases numbers dying neurons, reactive microglia, and proliferating MGPCs, whereas PTN has no effect. Inhibition of MDK-signaling with Na3 VO4 blocks neuroprotective effects with an increase in the number of dying cells and negates the pro-proliferative effects on MGPCs in damaged retinas. Inhibitors of PP2A and Pak1, which are associated with MDK-signaling through integrin ß1, suppressed the formation of MGPCs in damaged chick retinas. In mice, MDK promotes a small but significant increase in proliferating MGPCs in damaged retinas and potently decreases the number of dying cells. We conclude that MDK expression is dynamically regulated in Müller glia during embryonic maturation, following retinal injury, and during reprogramming into MGPCs. MDK mediates glial activity, neuronal survival, and the re-programming of Müller glia into proliferating MGPCs.

River lamprey present an unusual predation threat to Atlantic salmon smolts in Lough Neagh, Northern Ireland.

A new monitoring programme on the Lough Neagh catchment has documented a high incidence of river lamprey, Lampetra fluviatilis L., predation on Atlantic salmon smolts, Salmo salar L. In total 470 smolts were examined during the 2020 emigration period with 168 fish (36%) exhibiting lamprey scars of which 57 were lightly scarred and 111 were classed as heavily scarred. Lamprey predation was not size selective on Lough Neagh S. salar smolts.

Reactive microglia and IL1ß/IL-1R1-signaling mediate neuroprotection in excitotoxin-damaged mouse retina.

BACKGROUND: Microglia and inflammation have context-specific impacts upon neuronal survival in different models of central nervous system (CNS) disease. Herein, we investigate how inflammatory mediators, including microglia, interleukin 1 beta (IL1ß), and signaling through interleukin 1 receptor type 1 (IL-1R1), influence the survival of retinal neurons in response to excitotoxic damage. METHODS: Excitotoxic retinal damage was induced via intraocular injections of NMDA. Microglial phenotype and neuronal survival were assessed by immunohistochemistry. Single-cell RNA sequencing was performed to obtain transcriptomic profiles. Microglia were ablated by using clodronate liposome or PLX5622. Retinas were treated with IL1ß prior to NMDA damage and cell death was assessed in wild type, IL-1R1 null mice, and mice expressing IL-1R1 only in astrocytes. RESULTS: NMDA-induced damage included neuronal cell death, microglial reactivity, upregulation of pro-inflammatory cytokines, and genes associated with IL1ß-signaling in different types of retinal neurons and glia. Expression of the IL1ß receptor, IL-1R1, was evident in astrocytes, endothelial cells, some Müller glia, and OFF bipolar cells. Ablation of microglia with clodronate liposomes or Csf1r antagonist (PLX5622) resulted in elevated cell death and diminished neuronal survival in excitotoxin-damaged retinas. Exogenous IL1ß stimulated the proliferation and reactivity of microglia in the absence of damage, reduced numbers of dying cells in damaged retinas, and increased neuronal survival following an insult. IL1ß failed to provide neuroprotection in the IL-1R1-null retina, but IL1ß-mediated neuroprotection was rescued when expression of IL-1R1 was restored in astrocytes. CONCLUSIONS: We conclude that reactive microglia provide protection to retinal neurons, since the absence of microglia is detrimental to survival. We propose that, at least in part, the survival-influencing effects of microglia may be mediated by IL1ß, IL-1R1, and interactions of microglia and other macroglia.

Cholesterol Modifies Huntingtin Binding to, Disruption of, and Aggregation on Lipid Membranes.

Huntington's disease (HD) is an inherited neurodegenerative disease caused by abnormally long CAG-repeats in the huntingtin gene that encode an expanded polyglutamine (polyQ) domain near the N-terminus of the huntingtin (htt) protein. Expanded polyQ domains are directly correlated to disease-related htt aggregation. Htt is found highly associated with a variety of cellular and subcellular membranes that are predominantly comprised of lipids. Since cholesterol homeostasis is altered in HD, we investigated how varying cholesterol content modifies the interactions between htt and lipid membranes. A combination of Langmuir trough monolayer techniques, vesicle permeability and binding assays, and in situ atomic force microscopy were used to directly monitor the interaction of a model, synthetic htt peptide and a full-length htt-exon1 recombinant protein with model membranes comprised of total brain lipid extract (TBLE) and varying amounts of exogenously added cholesterol. As the cholesterol content of the membrane increased, the extent of htt insertion decreased. Vesicles containing extra cholesterol were resistant to htt-induced permeabilization. Morphological and mechanical changes in the bilayer associated with exposure to htt were also drastically altered by the presence of cholesterol. Disrupted regions of pure TBLE bilayers were grainy in appearance and associated with a large number of globular aggregates. In contrast, morphological changes induced by htt in bilayers enriched in cholesterol were plateau-like with a smooth appearance. Collectively, these observations suggest that the presence and amount of cholesterol in lipid membranes play a critical role in htt binding and aggregation on lipid membranes.

Advances in Image-Guided Ablation Therapies for Solid Tumors.

Image-guided solid tumor ablation methods have significantly advanced in their capability to target primary and metastatic tumors. These techniques involve noninvasive or percutaneous insertion of applicators to induce thermal, electrochemical, or mechanical stress on malignant tissue to cause tissue destruction and apoptosis of the tumor margins. Ablation offers substantially lower risks compared to traditional methods. Benefits include shorter recovery periods, reduced bleeding, and greater preservation of organ parenchyma compared to surgical intervention. Due to the reduced morbidity and mortality, image-guided tumor ablation offers new opportunities for treatment in cancer patients who are not candidates for resection. Currently, image-guided ablation techniques are utilized for treating primary and metastatic tumors in various organs with both curative and palliative intent, including the liver, pancreas, kidneys, thyroid, parathyroid, prostate, lung, breast, bone, and soft tissue. The invention of new equipment and techniques is expanding the criteria of eligible patients for therapy, as now larger and more high-risk tumors near critical structures can be ablated. This article provides an overview of the different imaging modalities, noninvasive, and percutaneous ablation techniques available and discusses their applications and associated complications across various organs.

Understanding ChatGPT for evidence-based utilization in interventional radiology.

Advancement in artificial intelligence (AI) has the potential to improve the efficiency and accuracy of medical care. New techniques used in machine learning have enhanced the functionality of software to perform advanced tasks with human-like capabilities. ChatGPT is the most utilized large language model and provides a diverse range of communication tasks. Interventional Radiology (IR) may benefit from the implementation of ChatGPT for specific tasks. This review summarizes the design principles of ChatGPT relevant to healthcare and highlights activities with the greatest potential for ChatGPT utilization in the practice of IR. These tasks involve patient-directed and physician-directed communications to convey medical information efficiently and act as a medical decision support tool. ChatGPT exemplifies the evolving landscape of new AI tools for advancing patient care and how physicians and patients may benefit with strategic execution.

Value of interventional radiology and their contributions to modern medical systems.

Interventional radiology (IR) is a unique specialty that incorporates a diverse set of skills ranging from imaging, procedures, consultation, and patient management. Understanding how IR generates value to the healthcare system is important to review from various perspectives. IR specialists need to understand how to meet demands from various stakeholders to expand their practice improving patient care. Thus, this review discusses the domains of value contributed to medical systems and outlines the parameters of success. IR benefits five distinct parties: patients, practitioners, payers, employers, and innovators. Value to patients and providers is delivered through a wide set of diagnostic and therapeutic interventions. Payers and hospital systems financially benefit from the reduced cost in medical management secondary to fast patient recovery, outpatient procedures, fewer complications, and the prestige of offering diverse expertise for complex patients. Lastly, IR is a field of rapid innovation implementing new procedural technology and techniques. Overall, IR must actively advocate for further growth and influence in the medical field as their value continues to expand in multiple domains. Despite being a nascent specialty, IR has become indispensable to modern medical practice.

Formation of Müller glia-derived progenitor cells in retinas depleted of microglia.

Recent studies have demonstrated the complex coordination of pro-inflammatory signaling and reactive microglia/macrophage on the formation Müller glial-derived progenitor cells (MGPCs) in the retinas of fish, birds and mice. We generated scRNA-seq libraries to identify transcriptional changes in Müller glia (MG) that result from the depletion of microglia from the chick retina. We found significant changes in different networks of genes in MG in normal and damaged retinas when the microglia are ablated. We identified a failure of MG to upregulate Wnt-ligands, Heparin binding epidermal growth factor (HBEGF), Fibroblast growth factor (FGF), retinoic acid receptors and genes related to Notch-signaling. Inhibition of GSK3ß, to simulate Wnt-signaling, failed to rescue the deficit in formation of proliferating MGPCs in damaged retinas missing microglia. By comparison, application of HBEGF or FGF2 completely rescued the formation of proliferating MGPCs in microglia-depleted retinas. Similarly, injection of a small molecule inhibitor to Smad3 or agonist to retinoic acid receptors partially rescued the formation of proliferating MGPCs in microglia-depleted damaged retinas. According to scRNA-seq libraries, patterns of expression of ligands, receptors, signal transducers and/or processing enzymes to cell-signaling via HBEGF, FGF, retinoic acid and TGFß are rapidly and transiently upregulated by MG after neuronal damage, consistent with important roles for these cell-signaling pathways in regulating the formation of MGPCs. We conclude that quiescent and activated microglia have a significant impact upon the transcriptomic profile of MG. We conclude that signals produced by reactive microglia in damaged retinas stimulate MG to upregulate cell signaling through HBEGF, FGF and retinoic acid, and downregulate signaling through TGFß/Smad3 to promote the reprogramming on MG into proliferating MGPCs.

Nuclear Factor I in neurons, glia and during the formation of Müller glia-derived progenitor cells in avian, porcine and primate retinas.

The regenerative potential of Müller glia (MG) is extraordinary in fish, poor in chick and terrible in mammals. In the chick model, MG readily reprogram into proliferating Müller glia-derived progenitor cells (MGPCs), but neuronal differentiation is very limited. The factors that suppress the neurogenic potential of MGPCs in the chick are slowly being revealed. Isoforms of Nuclear Factor I (NFI) are cell-intrinsic factors that limit neurogenic potential; these factors are required for the formation of MG in the developing mouse retina and deletion of these factors reprograms MG into neuron-like cells in mature mouse retina. Accordingly, we sought to characterize the patterns of expression of NFIs in the developing, mature and damaged chick retina. In addition, we characterized patterns of expression of NFIs in the retinas of large mammals, pigs and monkeys. Using a combination of single-cell RNA-sequencing (scRNA-seq) and immunolabeling, we probed for patterns of expression. In embryonic chick, levels of NFIs are very low in early E5 (embryonic day 5) retinal progenitor cells (RPCs), upregulated in E8 RPCs, further upregulated in differentiating MG at E12 and E15. NFIs are maintained in mature resting MG, microglia and neurons. Levels of NFIs are reduced in activated MG in retinas treated with NMDA and/or insulin+FGF2, and further downregulated in proliferating MGPCs. However, levels of NFIs in MGPCs were significantly higher than those seen in RPCs. Immunolabeling for NFIA and NFIB closely matched patterns of expression revealed in different types of retinal neurons and glia, consistent with findings from scRNA-seq. In addition, we find expression of NFIA and NFIB through progenitors in the circumferential marginal zone at the far periphery of the retina. We find similar patterns of expression for NFIs in scRNA-seq databases for pig and monkey retinas. Patterns of expression of NFIA and NFIB were validated with immunofluorescence in pig and monkey retinas wherein these factors were predominantly detected in MG and a few types of inner retinal neurons. In summary, NFIA and NFIB are prominently expressed in developing chick retina and by mature neurons and glia in the retinas of chicks, pigs and monkeys. Although levels of NFIs are decreased in chick, in MGPCs these levels remain higher than those seen in neurogenic RPCs. We propose that the neurogenic potential of MGPCs in the chick retina is suppressed by NFIs.

Microglia coordinate cellular interactions during spinal cord repair in mice.

Traumatic spinal cord injury (SCI) triggers a neuro-inflammatory response dominated by tissue-resident microglia and monocyte derived macrophages (MDMs). Since activated microglia and MDMs are morphologically identical and express similar phenotypic markers in vivo, identifying injury responses specifically coordinated by microglia has historically been challenging. Here, we pharmacologically depleted microglia and use anatomical, histopathological, tract tracing, bulk and single cell RNA sequencing to reveal the cellular and molecular responses to SCI controlled by microglia. We show that microglia are vital for SCI recovery and coordinate injury responses in CNS-resident glia and infiltrating leukocytes. Depleting microglia exacerbates tissue damage and worsens functional recovery. Conversely, restoring select microglia-dependent signaling axes, identified through sequencing data, in microglia depleted mice prevents secondary damage and promotes recovery. Additional bioinformatics analyses reveal that optimal repair after SCI might be achieved by co-opting key ligand-receptor interactions between microglia, astrocytes and MDMs.

Gene regulatory networks controlling vertebrate retinal regeneration.

Injury induces retinal Müller glia of certain cold-blooded vertebrates, but not those of mammals, to regenerate neurons. To identify gene regulatory networks that reprogram Müller glia into progenitor cells, we profiled changes in gene expression and chromatin accessibility in Müller glia from zebrafish, chick, and mice in response to different stimuli. We identified evolutionarily conserved and species-specific gene networks controlling glial quiescence, reactivity, and neurogenesis. In zebrafish and chick, the transition from quiescence to reactivity is essential for retinal regeneration, whereas in mice, a dedicated network suppresses neurogenic competence and restores quiescence. Disruption of nuclear factor I transcription factors, which maintain and restore quiescence, induces Müller glia to proliferate and generate neurons in adult mice after injury. These findings may aid in designing therapies to restore retinal neurons lost to degenerative diseases.

Bismaleimide cross-linked anthrax toxin forms functional octamers with high specificity in tumor targeting.

In recent years, anthrax toxin has been reengineered to act as a highly specific antiangiogenic cancer therapeutic, shown to kill tumors in animal models. This has been achieved by modifying protective antigen (PA) so that its activation and toxicity require the presence of two proteases, matrix metalloproteinase (MMP) and urokinase plasminogen activator (uPA), which are upregulated in tumor microenvironments. These therapeutics consist of intercomplementing PA variants, which are individually nontoxic, but form functional toxins upon complementary oligomerization. Here, we have created a dual-protease requiring PA targeting system which utilizes bismaleimide cross-linked PA (CLPA) rather than the intercomplementing PA variants. Three different CLPA agents were tested and, as expected, found to exclusively form octamers. Two of the CLPA agents have in vitro toxicities equal to those of previous intercomplementing agents, while the third CLPA agent had compromised in vitro cleavage and was significantly less cytotoxic. We hypothesize this difference was due to steric hindrance caused by cross-linking two PA monomers in close proximity to the PA cleavage site. Overall, this work advances the development and use of the PA and LF tumor-targeting system as a practical cancer therapeutic, as it provides a way to reduce the drug components of the anthrax toxin drug delivery system from three to two, which may lower the cost and simplify testing in clinical trials. HIGHLIGHT: Previously, anthrax toxin has been reengineered to act as a highly specific antiangiogenic cancer therapeutic. Here, we present a version, which utilizes bismaleimide cross-linked protective antigen (PA) rather than intercomplementing PA variants. This advances the development of anthrax toxin as a practical cancer therapeutic as it reduces the components of the drug delivery system to two, which may lower the cost and simplify testing in clinical trials.