Generation of contractile forces by three-dimensional bundled axonal tracts in micro-tissue engineered neural networks.

Axonal extension and retraction are ongoing processes that occur throughout all developmental stages of an organism. The ability of axons to produce mechanical forces internally and respond to externally generated forces is crucial for nervous system development, maintenance, and plasticity. Such axonal mechanobiological phenomena have typically been evaluated in vitro at a single-cell level, but these mechanisms have not been studied when axons are present in a bundled three-dimensional (3D) form like in native tissue. In an attempt to emulate native cortico-cortical interactions under in vitro conditions, we present our approach to utilize previously described micro-tissue engineered neural networks (micro-TENNs). Here, micro-TENNs were comprised of discrete populations of rat cortical neurons that were spanned by 3D bundled axonal tracts and physically integrated with each other. We found that these bundled axonal tracts inherently exhibited an ability to generate contractile forces as the microtissue matured. We therefore utilized this micro-TENN testbed to characterize the intrinsic contractile forces generated by the integrated axonal tracts in the absence of any external force. We found that contractile forces generated by bundled axons were dependent on microtubule stability. Moreover, these intra-axonal contractile forces could simultaneously generate tensile forces to induce so-called axonal "stretch-growth" in different axonal tracts within the same microtissue. The culmination of axonal contraction generally occurred with the fusion of both the neuronal somatic regions along the axonal tracts, therefore perhaps showing the innate tendency of cortical neurons to minimize their wiring distance, a phenomenon also perceived during brain morphogenesis. In future applications, this testbed may be used to investigate mechanisms of neuroanatomical development and those underlying certain neurodevelopmental disorders.

Porcine Models of Neurotrauma and Neurological Disorders.

The translation of therapeutics from lab to clinic has a dismal record in the fields of neurotrauma and neurological disorders [...].

Bedside to bench: the outlook for psychedelic research.

There has recently been a resurgence of interest in psychedelic compounds based on studies demonstrating their potential therapeutic applications in treating post-traumatic stress disorder, substance abuse disorders, and treatment-resistant depression. Despite promising efficacy observed in some clinical trials, the full range of biological effects and mechanism(s) of action of these compounds have yet to be fully established. Indeed, most studies to date have focused on assessing the psychological mechanisms of psychedelics, often neglecting the non-psychological modes of action. However, it is important to understand that psychedelics may mediate their therapeutic effects through multi-faceted mechanisms, such as the modulation of brain network activity, neuronal plasticity, neuroendocrine function, glial cell regulation, epigenetic processes, and the gut-brain axis. This review provides a framework supporting the implementation of a multi-faceted approach, incorporating in silico, in vitro and in vivo modeling, to aid in the comprehensive understanding of the physiological effects of psychedelics and their potential for clinical application beyond the treatment of psychiatric disorders. We also provide an overview of the literature supporting the potential utility of psychedelics for the treatment of brain injury (e.g., stroke and traumatic brain injury), neurodegenerative diseases (e.g., Parkinson's and Alzheimer's diseases), and gut-brain axis dysfunction associated with psychiatric disorders (e.g., generalized anxiety disorder and major depressive disorder). To move the field forward, we outline advantageous experimental frameworks to explore these and other novel applications for psychedelics.

Porcine Astrocytes and Their Relevance for Translational Neurotrauma Research.

Astrocytes are essential to virtually all brain processes, from ion homeostasis to neurovascular coupling to metabolism, and even play an active role in signaling and plasticity. Astrocytic dysfunction can be devastating to neighboring neurons made inherently vulnerable by their polarized, excitable membranes. Therefore, correcting astrocyte dysfunction is an attractive therapeutic target to enhance neuroprotection and recovery following acquired brain injury. However, the translation of such therapeutic strategies is hindered by a knowledge base dependent almost entirely on rodent data. To facilitate additional astrocytic research in the translatable pig model, we present a review of astrocyte findings from pig studies of health and disease. We hope that this review can serve as a road map for intrepid pig researchers interested in studying astrocyte biology.

Porcine Models of Spinal Cord Injury.

Large animal models of spinal cord injury may be useful tools in facilitating the development of translational therapies for spinal cord injury (SCI). Porcine models of SCI are of particular interest due to significant anatomic and physiologic similarities to humans. The similar size and functional organization of the porcine spinal cord, for instance, may facilitate more accurate evaluation of axonal regeneration across long distances that more closely resemble the realities of clinical SCI. Furthermore, the porcine cardiovascular system closely resembles that of humans, including at the level of the spinal cord vascular supply. These anatomic and physiologic similarities to humans not only enable more representative SCI models with the ability to accurately evaluate the translational potential of novel therapies, especially biologics, they also facilitate the collection of physiologic data to assess response to therapy in a setting similar to those used in the clinical management of SCI. This review summarizes the current landscape of porcine spinal cord injury research, including the available models, outcome measures, and the strengths, limitations, and alternatives to porcine models. As the number of investigational SCI therapies grow, porcine SCI models provide an attractive platform for the evaluation of promising treatments prior to clinical translation.

Multimodal Neuromonitoring and Neurocritical Care in Swine to Enhance Translational Relevance in Brain Trauma Research.

Neurocritical care significantly impacts outcomes after moderate-to-severe acquired brain injury, but it is rarely applied in preclinical studies. We created a comprehensive neurointensive care unit (neuroICU) for use in swine to account for the influence of neurocritical care, collect clinically relevant monitoring data, and create a paradigm that is capable of validating therapeutics/diagnostics in the unique neurocritical care space. Our multidisciplinary team of neuroscientists, neurointensivists, and veterinarians adapted/optimized the clinical neuroICU (e.g., multimodal neuromonitoring) and critical care pathways (e.g., managing cerebral perfusion pressure with sedation, ventilation, and hypertonic saline) for use in swine. Moreover, this neurocritical care paradigm enabled the first demonstration of an extended preclinical study period for moderate-to-severe traumatic brain injury with coma beyond 8 h. There are many similarities with humans that make swine an ideal model species for brain injury studies, including a large brain mass, gyrencephalic cortex, high white matter volume, and topography of basal cisterns, amongst other critical factors. Here we describe the neurocritical care techniques we developed and the medical management of swine following subarachnoid hemorrhage and traumatic brain injury with coma. Incorporating neurocritical care in swine studies will reduce the translational gap for therapeutics and diagnostics specifically tailored for moderate-to-severe acquired brain injury.

Initial treatment and survival in Danish patients diagnosed with non-small-cell lung cancer (2005-2015): SCAN-LEAF study.

Aim: To describe initial treatment patterns and survival of patients diagnosed with non-small-cell lung cancer (NSCLC) in Denmark, before immune checkpoint inhibitor and later-generation tyrosine kinase inhibitor use. Patients & methods: Adults diagnosed with incident NSCLC (2005-2015; follow-up: 2016). Initial treatments and overall survival (OS) are reported. Results: 31,939 NSCLC patients (51.6% stage IV) were included. Increasing use of curative radiotherapy/chemoradiation for stage I, II/IIIA and IIIB NSCLC coincided with improved 2-year OS. Systemic anticancer therapy use increased for patients with stage IV non-squamous NSCLC (53.0-60.6%) but not squamous NSCLC (44.9-47.3%). 1-year OS improved in patients with stage IV non-squamous NSCLC (23-31%) but not squamous NSCLC (22-25%). Conclusion: Trends indicated improved OS as treatments evolved between 2005 and 2015, but the effect was limited to 1-year OS in stage IV disease.

Unique Astrocyte Cytoskeletal and Nuclear Morphology in a Three-Dimensional Tissue-Engineered Rostral Migratory Stream.

Neural precursor cells (NPCs) are generated in the subventricular zone (SVZ) and travel through the rostral migratory stream (RMS) to replace olfactory bulb interneurons in the brains of most adult mammals. Following brain injury, SVZ-derived NPCs can divert from the RMS and migrate toward injured brain regions but arrive in numbers too low to promote functional recovery without experimental intervention. Our lab has biofabricated a "living scaffold" that replicates the structural and functional features of the endogenous RMS. This tissue-engineered rostral migratory stream (TE-RMS) is a new regenerative medicine strategy designed to facilitate stable and sustained NPC delivery into neuron-deficient brain regions following brain injury or neurodegenerative disease and an in vitro tool to investigate the mechanisms of neuronal migration and cell-cell communication. We have previously shown that the TE-RMS replicates the basic structure and protein expression of the endogenous RMS and can direct immature neuronal migration in vitro and in vivo. Here, we further describe profound morphological changes that occur following precise physical manipulation and subsequent self-assembly of astrocytes into the TE-RMS, including significant cytoskeletal rearrangement and nuclear elongation. The unique cytoskeletal and nuclear architecture of TE-RMS astrocytes mimics astrocytes in the endogenous rat RMS. Advanced imaging techniques reveal the unique morphology of TE-RMS cells that has yet to be described of astrocytes in vitro. The TE-RMS offers a novel platform to elucidate astrocyte cytoskeletal and nuclear dynamics and their relationship to cell behavior and function.

Relationships between injury kinematics, neurological recovery, and pathology following concussion.

Mild traumatic brain injury affects millions of individuals annually primarily through falls, traffic collisions, or blunt trauma and can generate symptoms that persist for years. Closed-head rotational loading is the most common cause of mild traumatic brain injury and is defined by a rapid rotational acceleration of brain tissue within an intact skull. Injury kinematics-the mechanical descriptors of injury-inducing motion-explain movement of the head, which govern energy transfer, and, therefore, determine injury severity. However, the relationship between closed-head rotational injury kinematics-such as angular velocity, angular acceleration, and injury duration-and outcome after mild traumatic brain injury is not completely understood. To address this gap in knowledge, we analysed archived surgical records of 24 swine experiencing a diffuse closed-head rotational acceleration mild traumatic brain injury against 12 sham animals. Kinematics were contrasted against acute recovery outcomes, specifically apnea time, extubation time, standing time, and recovery duration. Compared to controls, animals experiencing a mild traumatic brain injury were far more likely to have apnea (P < 0.001), shorter time to extubation (P = 0.023), and longer time from extubation to standing (P = 0.006). Using least absolute shrinkage and selection operator-based regressions, kinematic parameters, including maximum negative angular velocity and time from peak angular velocity to maximum angular deceleration, were selected to explain variation in apnea time, standing time, and recovery duration. Simplified linear models employing the least absolute shrinkage and selection operator-selected variables explained a modest degree of variation in apnea time (adjusted R 2 = 0.18), standing time (adjusted R 2 = 0.19), and recovery duration (adjusted R 2 = 0.27). Neuropathology was correlated with multiple injury kinematics, with maximum angular acceleration exhibiting the strongest correlation (R 2 = 0.66). Together, these data suggest the interplay between multiple injury kinematics, including maximum negative angular velocity (immediately preceding cessation of head motion) and time from peak angular velocity to maximum angular deceleration, best explain acute recovery metrics and neuropathology after mild traumatic brain injury in swine. Future experiments that independently manipulate individual kinematic parameters could be instrumental in developing translational diagnostics for clinical mild traumatic brain injury.

An implantable human stem cell-derived tissue-engineered rostral migratory stream for directed neuronal replacement.

The rostral migratory stream (RMS) facilitates neuroblast migration from the subventricular zone to the olfactory bulb throughout adulthood. Brain lesions attract neuroblast migration out of the RMS, but resultant regeneration is insufficient. Increasing neuroblast migration into lesions has improved recovery in rodent studies. We previously developed techniques for fabricating an astrocyte-based Tissue-Engineered RMS (TE-RMS) intended to redirect endogenous neuroblasts into distal brain lesions for sustained neuronal replacement. Here, we demonstrate that astrocyte-like-cells can be derived from adult human gingiva mesenchymal stem cells and used for TE-RMS fabrication. We report that key proteins enriched in the RMS are enriched in TE-RMSs. Furthermore, the human TE-RMS facilitates directed migration of immature neurons in vitro. Finally, human TE-RMSs implanted in athymic rat brains redirect migration of neuroblasts out of the endogenous RMS. By emulating the brain's most efficient means for directing neuroblast migration, the TE-RMS offers a promising new approach to neuroregenerative medicine.

Treatment and outcomes for early non-small-cell lung cancer: a retrospective analysis of a Portuguese hospital database.

AIM: This observational study evaluated treatment patterns and survival for patients with stage I-IIIA non-small-cell lung cancer (NSCLC). MATERIALS & METHODS: Adults newly diagnosed with NSCLC in 2012-2016 at IPO-Porto hospital were included. Treatment data were available for patients diagnosed in 2015-2016. RESULTS: 495 patients were included (median age: 67 years). The most common treatments were surgery alone or with another therapy (stage I: 66%) and systemic anticancer therapy plus radiotherapy (stage II: 54%; stage IIIA: 59%). One-year OS (95% CI) for patients with stage I, II and IIIA NSCLC (diagnosed 2012-2016) were 92% (88-96), 71% (62-82) and 69% (63-75), respectively; one-year OS (95% CI) for treated patients with stage I-II or stage IIIA NSCLC (diagnosed 2015-2016) were 89% (81-97) and 86% (75-98) for non-squamous cell and 76% (60-95) and 49% (34-70) for squamous cell NSCLC. CONCLUSION: Treatment advances are strongly needed for stage I-IIIA NSCLC, especially for patients with squamous cell histology.

Trends in the prescription of systemic anticancer therapy and mortality among patients with advanced non-small cell lung cancer: a real-world retrospective observational cohort study from the I-O optimise initiative.

OBJECTIVES: To assess how a decade of developments in systematic anticancer therapy (SACT) for advanced non-small cell lung cancer (NSCLC) affected overall survival (OS) in a large UK University Hospital. DESIGN: Real-world retrospective observational cohort study using existing data recorded in electronic medical records. SETTING: A large National Health Service (NHS) university teaching hospital serving 800 000 people living in a diverse metropolitan area of the UK. PARTICIPANTS: 2119 adults diagnosed with advanced NSCLC (tumour, node, metastasis stage IIIB or IV) between 2007 and 2017 at Leeds Teaching Hospitals NHS Trust. MAIN OUTCOMES AND MEASURES: OS following diagnosis and the analysis of factors associated with receiving SACT. RESULTS: Median OS for all participants was 2.9 months, increasing for the SACT-treated subcohort from 8.4 months (2007-2012) to 9.1 months (2013-2017) (p=0.02); 1-year OS increased from 33% to 39% over the same period for the SACT-treated group. Median OS for the untreated subcohort was 1.6 months in both time periods. Overall, 30.6% (648/2119) patients received SACT; treatment rates increased from 28.6% (338/1181) in 2007-2012 to 33.0% (310/938) in 2013-2017 (p=0.03). Age and performance status were independent predictors for SACT treatment; advanced age and higher performance status were associated with lower SACT treatment rates. CONCLUSION: Although developments in SACT during 2007-2017 correspond to some changes in survival for treated patients with advanced NSCLC, treatment rates remain low and the prognosis for all patients remains poor.

Evolving use of real-world evidence in the regulatory process: a focus on immuno-oncology treatment and outcomes.

In recent years, regulatory bodies have increasingly recognized the utility of real-world evidence (RWE) for supplementing and supporting clinical trial data in new drug applications. Nevertheless, the integration of RWE into established regulatory processes is complex and the generation of 'regulatory-grade' real-world data faces operational, methodological, data-related and policy-related challenges. In parallel with this evolving role for RWE, immuno-oncology therapies have emerged as leading cancer treatments and are expected to continue to play a central role in the future. In this article, we review the current literature on the use of RWE for regulatory submissions, with a focus on novel anticancer immunotherapies, and discuss the utility and current limitations of RWE in the context of drug development and regulatory approvals.

Tissue Engineered Bands of Büngner for Accelerated Motor and Sensory Axonal Outgrowth.

Following peripheral nerve injury comprising a segmental defect, the extent of axon regeneration decreases precipitously with increasing gap length. Schwann cells play a key role in driving axon re-growth by forming aligned tubular guidance structures called bands of Büngner, which readily occurs in distal nerve segments as well as within autografts - currently the most reliable clinically-available bridging strategy. However, host Schwann cells generally fail to infiltrate large-gap acellular scaffolds, resulting in markedly inferior outcomes and motivating the development of next-generation bridging strategies capable of fully exploiting the inherent pro-regenerative capability of Schwann cells. We sought to create preformed, implantable Schwann cell-laden microtissue that emulates the anisotropic structure and function of naturally-occurring bands of Büngner. Accordingly, we developed a biofabrication scheme leveraging biomaterial-induced self-assembly of dissociated rat primary Schwann cells into dense, fiber-like three-dimensional bundles of Schwann cells and extracellular matrix within hydrogel micro-columns. This engineered microtissue was found to be biomimetic of morphological and phenotypic features of endogenous bands of Büngner, and also demonstrated 8 and 2× faster rates of axonal extension in vitro from primary rat spinal motor neurons and dorsal root ganglion sensory neurons, respectively, compared to 3D matrix-only controls or planar Schwann cells. To our knowledge, this is the first report of accelerated motor axon outgrowth using aligned Schwann cell constructs. For translational considerations, this microtissue was also fabricated using human gingiva-derived Schwann cells as an easily accessible autologous cell source. These results demonstrate the first tissue engineered bands of Büngner (TE-BoBs) comprised of dense three-dimensional bundles of longitudinally aligned Schwann cells that are readily scalable as implantable grafts to accelerate axon regeneration across long segmental nerve defects.

Real-world treatment patterns and survival outcomes for advanced non-small cell lung cancer in the pre-immunotherapy era in Portugal: a retrospective analysis from the I-O Optimise initiative.

BACKGROUND: As part of the multinational I-O Optimise research initiative, this retrospective cohort study of patients with advanced non-small cell lung cancer (NSCLC) evaluated real-world treatment patterns and survival prior to immunotherapy reimbursement in Portugal. METHODS: This study utilized a database held by IPO-Porto, Portugal's largest oncology hospital. Adult patients diagnosed with stage IIIB or IV NSCLC from January 2012 to December 2016 at IPO-Porto, with follow-up to June 2017, were included. Treatment analyses were performed from 2015 onwards. Kaplan-Meier methods were used for overall survival (OS). Factors associated with OS and systemic anti-cancer therapy (SACT) treatment were assessed using multivariate statistical models. RESULTS: Of 1524 patients diagnosed with NSCLC at IPO-Porto, 1008 patients had advanced disease (stage IIIB: 10.1%, 154/1524, stage IV: 56.0%, 854/1524). For those with advanced disease, median age was 65 years (range: 21-92) and 75.6% (762/1008) were male. Median OS (interquartile range [IQR]) was 11.4 (5.2-26.9) months for stage IIIB and 6.3 (2.4-15.0) months for stage IV. Factors associated with decreased risk of death included female sex and epidermal growth factor receptor gene (EGFR)/anaplastic lymphoma kinase gene (ALK) mutations/rearrangements; factors associated with increased risk of death included older age and stage IV disease. Among patients diagnosed in 2015 or 2016, 75.8% (297/392) received ≥1 line of SACT. Platinum-based chemotherapy was the most common first-line therapy (non-squamous cell carcinoma [NSQ]: 72.9%; squamous cell carcinoma [SQ] 87.3%, 55/63; patients with EGFR/ALK mutations/rearrangements primarily received tyrosine kinase inhibitors). The likelihood of receiving SACT was lower in older patients and those diagnosed with stage IV disease. Patients not receiving SACT had poor survival outcomes (median OS [IQR]: NSQ, 1.8 [1.1-3.1] months; SQ, 2.3 (1.3-3.4) months), while median OS (IQR) in SACT-treated patients was 12.6 (6.1-24.5) months for NSQ and 10.3 (5.7-15.9) months for SQ. CONCLUSIONS: This real-world data analysis from a large Portuguese oncology hospital demonstrates a high disease burden for advanced NSCLC in the pre-immunotherapy era, with nearly one-quarter of patients not receiving SACT. Even in patients receiving SACT, median survival was only about 1 year.

Tissue Engineering and Biomaterial Strategies to Elicit Endogenous Neuronal Replacement in the Brain.

Neurogenesis in the postnatal mammalian brain is known to occur in the dentate gyrus of the hippocampus and the subventricular zone. These neurogenic niches serve as endogenous sources of neural precursor cells that could potentially replace neurons that have been lost or damaged throughout the brain. As an example, manipulation of the subventricular zone to augment neurogenesis has become a popular strategy for attempting to replace neurons that have been lost due to acute brain injury or neurodegenerative disease. In this review article, we describe current experimental strategies to enhance the regenerative potential of endogenous neural precursor cell sources by enhancing cell proliferation in neurogenic regions and/or redirecting migration, including pharmacological, biomaterial, and tissue engineering strategies. In particular, we discuss a novel replacement strategy based on exogenously biofabricated "living scaffolds" that could enhance and redirect endogenous neuroblast migration from the subventricular zone to specified regions throughout the brain. This approach utilizes the first implantable, biomimetic tissue-engineered rostral migratory stream, thereby leveraging the brain's natural mechanism for sustained neuronal replacement by replicating the structure and function of the native rostral migratory stream. Across all these strategies, we discuss several challenges that need to be overcome to successfully harness endogenous neural precursor cells to promote nervous system repair and functional restoration. With further development, the diverse and innovative tissue engineering and biomaterial strategies explored in this review have the potential to facilitate functional neuronal replacement to mitigate neurological and psychiatric symptoms caused by injury, developmental disorders, or neurodegenerative disease.

I-O Optimise: a novel multinational real-world research platform in thoracic malignancies.

Aim: To describe I-O Optimise, a multinational program providing real-world insights into lung cancer management. Materials & methods: Real-world data source selection for I-O Optimise followed a structured approach focused on population coverage, key variable capture, continuous/consistent data availability, record duration and data latency, and database expertise. Results: As of 31 October 2018, seven real-world data sources were included in I-O Optimise, providing data on characteristics, treatment patterns and clinical outcomes from more than 45,000 patients/year with non-small-cell lung cancer, small-cell lung cancer and mesothelioma across Denmark, Norway, Portugal, Spain, Sweden and the UK. Conclusion: The ongoing I-O Optimise initiative has the potential to provide a broad, robust and dynamic research platform to continually address numerous research objectives in the lung cancer arena.

Challenges and demand for modeling disorders of consciousness following traumatic brain injury.

Following severe traumatic brain injury (TBI), many patients experience coma - an unresponsive state lacking wakefulness or awareness. Coma rarely lasts more than two weeks, and emergence involves passing through a state of wakefulness without awareness of self or environment. Patients that linger in these Disorders of Consciousness (DoC) undergo clinical assessments of awareness for diagnosis into Unresponsive Wakefulness Syndrome (no awareness, also called vegetative state) or Minimally Conscious State (periodic increases in awareness). These diagnoses are notoriously inaccurate, offering little prognostic value. Recovery of awareness is unpredictable, returning within weeks, years, or never. This leaves patients' families with difficult decisions and little information on which to base them. Clinical studies have made significant advancements, but remain encumbered by high variability, limited data output, and a lack of necessary controls. Herein we discuss the clear and present need to establish a preclinical model of TBI-induced DoC, the significant challenges involved, and how such a model can be applied to support DoC research.

A tissue-engineered rostral migratory stream for directed neuronal replacement.

New neurons are integrated into the circuitry of the olfactory bulb throughout the lifespan in the mammalian brain-including in humans. These new neurons are born in the subventricular zone and subsequently mature as they are guided over long distances via the rostral migratory stream through mechanisms we are only just beginning to understand. Regeneration after brain injury is very limited, and although some neuroblasts from the rostral migratory stream will leave the path and migrate toward cortical lesion sites, this neuronal replacement is generally not sustained and therefore does not provide enough new neurons to alleviate functional deficits. Using newly discovered microtissue engineering techniques, we have built the first self-contained, implantable constructs that mimic the architecture and function of the rostral migratory stream. This engineered microtissue emulates the dense cord-like bundles of astrocytic somata and processes that are the hallmark anatomical feature of the glial tube. As such, our living microtissue-engineered rostral migratory stream can serve as an in vitro test bed for unlocking the secrets of neuroblast migration and maturation, and may potentially serve as a living transplantable construct derived from a patient's own cells that can redirect their own neuroblasts into lesion sites for sustained neuronal replacement following brain injury or neurodegenerative disease. In this paper, we summarize the development of fabrication methods for this microtissue-engineered rostral migratory stream and provide proof-of-principle evidence that it promotes and directs migration of immature neurons.