Non-resolving neuroinflammation regulates axon regeneration in chronic spinal cord injury.

Chronic spinal cord injury (SCI) lesions retain increased densities of microglia and macrophages. In acute SCI, macrophages induce growth cone collapse, facilitate axon retraction away from lesion boundaries, as well as play a key role in orchestrating the growth-inhibitory glial scar. Little is known about the role of sustained inflammation in chronic SCI, or whether chronic inflammation affects repair and regeneration. We performed transcriptional analysis using the Nanostring Neuropathology panel to characterize the resolution of inflammation into chronic SCI, to characterize the chronic SCI microenvironment, as well as to identify spinal cord responses to macrophage depletion and repopulation using the CSF1R inhibitor, PLX-5622. We determined the ability for macrophage depletion and repopulation to augment axon growth into chronic lesions both with and without regenerative stimulation using neuronal-specific PTEN knockout (PTEN-KO). PTEN-KO was delivered with spinal injections of retrogradely transported adeno associated viruses (AAVrg's). Both transcriptional analyses and immunohistochemistry revealed the ability for PLX-5622 to significantly deplete inflammation around and within chronic SCI lesions, with a return to pre-depleted inflammatory densities after treatment removal. Neuronal-specific transcripts were significantly elevated in mice after inflammatory repopulation, but no significant effects were observed with macrophage depletion alone. Axon densities significantly increased within the lesion after PLX-5622 treatment with a more consistent effect observed in mice with inflammatory repopulation. PTEN-KO did not further increase axon densities within the lesion beyond effects induced by PLX-5622. We identified that PLX-5622 increased axon densities within the lesion that are histologically identified as 5-HT+and CGRP+, both of which are not robustly transduced by AAVrg's. Our work identified that increased macrophage/microglia densities in the chronic SCI environment may be actively retained by homeostatic mechanisms likely affiliated with a sustained elevated expression of CSF1 and other chemokines. Finally, we identify a novel role of sustained inflammation as a prospective barrier to axon regeneration in chronic SCI.

Tissue-resident macrophages specifically express Lactotransferrin and Vegfc during ear pinna regeneration in spiny mice.

The details of how macrophages control different healing trajectories (regeneration vs. scar formation) remain poorly defined. Spiny mice (Acomys spp.) can regenerate external ear pinnae tissue, whereas lab mice (Mus musculus) form scar tissue in response to an identical injury. Here, we used this dual species system to dissect macrophage phenotypes between healing modes. We identified secreted factors from activated Acomys macrophages that induce a pro-regenerative phenotype in fibroblasts from both species. Transcriptional profiling of Acomys macrophages and subsequent in vitro tests identified VEGFC, PDGFA, and Lactotransferrin (LTF) as potential pro-regenerative modulators. Examining macrophages in vivo, we found that Acomys-resident macrophages secreted VEGFC and LTF, whereas Mus macrophages do not. Lastly, we demonstrate the requirement for VEGFC during regeneration and find that interrupting lymphangiogenesis delays blastema and new tissue formation. Together, our results demonstrate that cell-autonomous mechanisms govern how macrophages react to the same stimuli to differentially produce factors that facilitate regeneration.

Azithromycin reduces hemoglobin-induced innate neuroimmune activation.

Neonatal intraventricular hemorrhage (IVH) releases blood products into the lateral ventricles and brain parenchyma. There are currently no medical treatments for IVH and surgery is used to treat a delayed effect of IVH, post-hemorrhagic hydrocephalus. However, surgery is not a cure for intrinsic brain injury from IVH, and is performed in a subacute time frame. Like many neurological diseases and injuries, innate immune activation is implicated in the pathogenesis of IVH. Innate immune activation is a pharmaceutically targetable mechanism to reduce brain injury and post-hemorrhagic hydrocephalus after IVH. Here, we tested the macrolide antibiotic azithromycin, which has immunomodulatory properties, to reduce innate immune activation in an in vitro model of microglial activation using the blood product hemoglobin (Hgb). We then utilized azithromycin in our in vivo model of IVH, using intraventricular blood injection into the lateral ventricle of post-natal day 5 rat pups. In both models, azithromycin modulated innate immune activation by several outcome measures including mitochondrial bioenergetic analysis, cytokine expression and flow cytometric analysis. This suggests that azithromycin, which is safe for neonates, could hold promise for modulating innate immune activation after IVH.

PTEN knockout using retrogradely transported AAVs transiently restores locomotor abilities in both acute and chronic spinal cord injury.

Restoring function in chronic stages of spinal cord injury (SCI) has often been met with failure or reduced efficacy when regenerative strategies are delayed past the acute or sub-acute stages of injury. Restoring function in the chronically injured spinal cord remains a critical challenge. We found that a single injection of retrogradely transported adeno-associated viruses (AAVrg) to knockout the phosphatase and tensin homolog protein (PTEN) in chronic SCI can effectively target both damaged and spared axons and transiently restore locomotor functions in near-complete injury models. AAVrg's were injected to deliver cre recombinase and/or a red fluorescent protein (RFP) under the human Synapsin 1 promoter (hSyn1) into the spinal cords of C57BL/6 PTENFloxΔ/Δ mice to knockout PTEN (PTEN-KO) in a severe thoracic SCI crush model at both acute and chronic time points. PTEN-KO improved locomotor abilities in both acute and chronic SCI conditions over a 9-week period. Regardless of whether treatment was initiated at the time of injury (acute), or three months after SCI (chronic), mice with limited hindlimb joint movement gained hindlimb weight support after treatment. Interestingly, functional improvements were not sustained beyond 9 weeks coincident with a loss of RFP reporter-gene expression and a near-complete loss of treatment-associated functional recovery by 6 months post-treatment. Treatment effects were also specific to severely injured mice; animals with weight support at the time of treatment lost function over a 6-month period. Retrograde tracing with Fluorogold revealed viable neurons throughout the motor cortex despite a loss of RFP expression at 9 weeks post-PTEN-KO. However, few Fluorogold labeled neurons were detected within the motor cortex at 6 months post-treatment. BDA labeling from the motor cortex revealed a dense corticospinal tract (CST) bundle in all groups except chronically treated PTEN-KO mice, indicating a potential long-term toxic effect of PTEN-KO to neurons in the motor cortex which was corroborated by a loss of ß-tubulin III labeling above the lesion within spinal cords after PTEN-KO. PTEN-KO mice had significantly more ß-tubulin III labeled axons within the lesion when treatment was delivered acutely, but not chronically post-SCI. In conclusion, we have found that using AAVrg's to knockout PTEN is an effective manipulation capable of restoring motor functions in chronic SCI and can enhance axon growth of currently unidentified axon populations when delivered acutely after injury. However, the long-term consequences of PTEN-KO on neuronal health and viability should be further explored.

Effects of Acute Ethanol Intoxication on Spinal Cord Injury Outcomes in Female Mice.

Abstract Approximately one in three traumatic spinal cord injuries (SCIs) occurs during or shortly after the consumption of alcohol. A small number of retrospective clinical studies report variable effects of alcohol intoxication on mortality, neurological recovery, and complications after SCI. Some of these studies demonstrate a protective effect of alcohol intoxication on SCI outcomes, whereas others show an increased complication risk. Pre-clinical studies in rat, ferret, and feline SCI models report a detrimental effect of ethanol intoxication on hemorrhage, motor recovery, and biochemical markers of tissue injury. However, no studies to date have investigated the neuropathological consequences of ethanol intoxication at the time of SCI or the reciprocal effect of SCI on ethanol metabolism. Therefore, we combined a pre-clinical mouse model of acute ethanol intoxication and experimental vertebral level T9 contusion SCI to investigate their interactive effects in female mice. We first investigated the effect of SCI on ethanol metabolism and found that T9 SCI does not alter ethanol metabolism. However, we did find that isoflurane anesthesia significantly slowed ethanol metabolism independent of SCI. We also determined how acute ethanol intoxication at the time of SCI alters locomotor recovery and lesion pathology. Using the Basso Mouse Scale (BMS) and CatWalk XT Gait Analysis System, we assessed locomotor recovery for 6 weeks after injury and observed that acute ethanol intoxication at the time of injury did not alter locomotor recovery. We also found no effect of ethanol intoxication on heat hyperalgesia development. There was, however, a detrimental effect of ethanol on tissue sparing after SCI. Therefore, we conclude that acute alcohol intoxication at the time of injury may contribute to the neuropathological consequences of SCI.

PTEN knockout using retrogradely transported AAVs restores locomotor abilities in both acute and chronic spinal cord injury.

Restoring function in chronic stages of spinal cord injury (SCI) has often been met with failure or reduced efficacy when regenerative strategies are delayed past the acute or sub-acute stages of injury. Restoring function in the chronically injured spinal cord remains a critical challenge. We found that a single injection of retrogradely transported adeno-associated viruses (AAVrg) to knockout the phosphatase and tensin homolog protein (PTEN) in chronic SCI can effectively target both damaged and spared axons and restore locomotor functions in near-complete injury models. AAVrg's were injected to deliver cre recombinase and/or a red fluorescent protein (RFP) under the human Synapsin 1 promoter (hSyn1) into the spinal cords of C57BL/6 PTEN FloxΔ / Δ mice to knockout PTEN (PTEN-KO) in a severe thoracic SCI crush model at both acute and chronic time points. PTEN-KO improved locomotor abilities in both acute and chronic SCI conditions over a 9-week period. Regardless of whether treatment was initiated at the time of injury (acute), or three months after SCI (chronic), mice with limited hindlimb joint movement gained hindlimb weight support after treatment. Interestingly, functional improvements were not sustained beyond 9 weeks coincident with a loss of RFP reporter-gene expression and a near-complete loss of treatment-associated functional recovery by 6 months post-treatment. Treatment effects were also specific to severely injured mice; animals with weight support at the time of treatment lost function over a 6-month period. Retrograde tracing with Fluorogold revealed viable neurons throughout the motor cortex despite a loss of RFP expression at 9 weeks post-PTEN-KO. However, few Fluorogold labeled neurons were detected within the motor cortex at 6 months post-treatment. BDA labeling from the motor cortex revealed a dense corticospinal tract (CST) bundle in all groups except chronically treated PTEN-KO mice indicating a potential long-term toxic effect of PTEN-KO to neurons in the motor cortex. PTEN-KO mice had significantly more ß - tubulin III labeled axons within the lesion when treatment was delivered acutely, but not chronically post-SCI. In conclusion, we have found that using AAVrg's to knockout PTEN is an effective manipulation capable of restoring motor functions in chronic SCI and can enhance axon growth of currently unidentified axon populations when delivered acutely after injury. However, the long-term consequences of PTEN-KO may exert neurotoxic effects.

Microglia as drivers of neurodegeneration: The role of innate-adaptive immune signaling.

Microglia contribute to neurodegeneration through numerous mechanisms. In this issue of Neuron, Shi et al.1 identify a maladaptive innate-adaptive immune axis with CD8+ T cells, mediated through microglial CCL2/8 and CCR2/5, in radiation-induced brain injury and stroke. Their findings across species and injuries suggest broader implications for neurodegenerative conditions.

Challenges in Translating Regenerative Therapies for Spinal Cord Injury.

Regenerating the injured spinal cord is a substantial challenge with many obstacles that need to be overcome to achieve robust functional benefits. This abundance of hurdles can partly explain the limited success when applying regenerative intervention treatments in animal models and/or people. In this article, we elaborate on a few of these obstacles, starting with the applicability of animal models and how they compare to the clinical setting. We then discuss the requirement for combinatorial interventions and the associated problems in experimental design, including the addition of rehabilitative training. The article expands on differences in lesion sizes and locations between humans and common animal models, and how this difference can determine the success or failure of an intervention. An additional and frequently overlooked problem in the translation of interventions that applies beyond the field of neuroregeneration is the reporting bias and the lack of transparency in reporting findings. New data mandates are tackling this problem and will eventually result in a more balanced view of the field. Finally, we will discuss strategies to negotiate the challenging course of successful translation to facilitate successful translation of regeneration promoting interventions.

Improving translatability of spinal cord injury research by including age as a demographic variable.

Pre-clinical and clinical spinal cord injury (SCI) studies differ in study design, particularly in the demographic characteristics of the chosen population. In clinical study design, criteria such as such as motor scores, neurological level, and severity of injury are often key determinants for participant inclusion. Further, demographic variables in clinical trials often include individuals from a wide age range and typically include both sexes, albeit historically most cases of SCI occur in males. In contrast, pre-clinical SCI models predominately utilize young adult rodents and typically use only females. While it is often not feasible to power SCI clinical trials to test multi-variable designs such as contrasting different ages, recent pre-clinical findings in SCI animal models have emphasized the importance of considering age as a biological variable prior to human experiments. Emerging pre-clinical data have identified case examples of treatments that diverge in efficacy across different demographic variables and have elucidated several age-dependent effects in SCI. The extent to which these differing or diverging treatment responses manifest clinically can not only complicate statistical findings and trial interpretations but also may be predictive of worse outcomes in select clinical populations. This review highlights recent literature including age as a biological variable in pre-clinical studies and articulates the results with respect to implications for clinical trials. Based on emerging unpredictable treatment outcomes in older rodents, we argue for the importance of including age as a biological variable in pre-clinical animal models prior to clinical testing. We believe that careful analyses of how age interacts with SCI treatments and pathophysiology will help guide clinical trial design and may improve both the safety and outcomes of such important efforts.

Advanced Age and Neurotrauma Diminish Glutathione and Impair Antioxidant Defense after Spinal Cord Injury.

Advanced age at the time of spinal cord injury (SCI) exacerbates damage from reactive oxygen species (ROS). Mechanisms underlying this age-dependent response are not well understood and may arise from decreased antioxidant defense. We investigated how spinal cord levels of the antioxidant glutathione (GSH), and its regulation, change with age and SCI. GSH is used by GSH peroxidase to sequester ROS and is recycled by GSH reductase. Male and female, 4- and 14-month-old (MO) mice received a 60 kDyn contusion SCI, and the levels of GSH and its regulatory enzymes were evaluated at one and three days post-injury (dpi). The mice with SCI were treated with N-acetylcysteine-amide (NACA; 150 mg/kg), a cysteine supplement that increases GSH, to determine effects on functional and histological outcomes. GSH was decreased with older age in sham mice, and an SCI-dependent depletion was observed in 4-MO mice by three dpi. Neither age nor injury affected the abundance of proteins regulating GSH synthesis or recycling. GSH peroxidase activity, however, increased after SCI only in 4-MO mice. In contrast, GSH peroxidase activity was increased in 14-MO sham mice, indicating that spinal cords of older mice have an elevated oxidative state. Indeed, 14-MO sham mice had more oxidized protein (3-nitrotyrosine [3-NT]) within their spinal cords compared with 4-MO sham mice. Only 4-MO mice had significant injury-induced increases in 3-NT at three dpi. NACA treatment restored GSH and improved the redox environment in injured 4- and 14-MO mice at one dpi; however, three days of NACA delivery did not improve motor, sensory, or anatomical deficits at 28 dpi in 4-MO mice and trended toward toxicity in all outcomes in 14-MO mice. Our observation suggests that GSH levels at acute stages of SCI play a minimal role in age-dependent outcomes reported after SCI in mice. Collective results implicate elements of injury occurring after three dpi, such as inflammation, as key regulators of age-dependent effects.

Immunoglobulin G Is Increased in the Injured Spinal Cord in a Sex- and Age-Dependent Manner.

There are limited studies examining age and sex as biological variables in the pathophysiology of spinal cord injury (SCI). The use of older animals and sex-balanced groups in SCI models is increasingly prioritized to better match clinical demographics. Including older animals in SCI studies is technically challenging, and outcomes are unpredictable with respect to biological and treatment responses. Incidental discoveries that are unrelated to the question under investigation often emerge while including age and sex as biological variables. When probing tissue homogenates on Western blots of 4- and 14-month-old (MO) mice, we identified a sex- and age-dependent increase in immunoglobulin G (IgG) within the spinal cords of older, 14-MO mice acutely after SCI, with females having more IgG compared with males. We further probed to determine whether differences in hemorrhage exist between sexes or ages by evaluating hemoglobin within spinal homogenates. Differences in hemoglobin between sexes and ages were not consistently observed. Because IgG was elevated in an age- and sex-dependent manner without of evidence of differences in hemorrhage, our findings point to potential pre-existing differences in IgG within mouse plasma in an age- and sex-dependent manner. This report has identified age- and sex-dependent differences in infiltrating IgG into the injured spinal cord environment that may affect injury and recovery processes. Our findings highlight that systemic contributions to SCI can be sex- and age-dependent and illustrate the value of reporting incidental discoveries.

Reflections on Data Sharing Practices in Spinal Cord Injury Research.

There are few pharmacological therapeutics available for spinal cord injury despite years of preclinical and clinical research. This brief editorial discusses some of the shortcomings of translational research efforts. In addition, we comment on our previous experiences with data curation and highlight evolving efforts by the spinal cord injury research community to improve prospects for future therapeutic development, especially pertaining to preclinical data sharing through the Open Data Commons for Spinal Cord Injury (ODC-SCI).

Promoting FAIR Data Through Community-driven Agile Design: the Open Data Commons for Spinal Cord Injury (odc-sci.org).

The past decade has seen accelerating movement from data protectionism in publishing toward open data sharing to improve reproducibility and translation of biomedical research. Developing data sharing infrastructures to meet these new demands remains a challenge. One model for data sharing involves simply attaching data, irrespective of its type, to publisher websites or general use repositories. However, some argue this creates a 'data dump' that does not promote the goals of making data Findable, Accessible, Interoperable and Reusable (FAIR). Specialized data sharing communities offer an alternative model where data are curated by domain experts to make it both open and FAIR. We report on our experiences developing one such data-sharing ecosystem focusing on 'long-tail' preclinical data, the Open Data Commons for Spinal Cord Injury (odc-sci.org). ODC-SCI was developed with community-based agile design requirements directly pulled from a series of workshops with multiple stakeholders (researchers, consumers, non-profit funders, governmental agencies, journals, and industry members). ODC-SCI focuses on heterogeneous tabular data collected by preclinical researchers including bio-behaviour, histopathology findings and molecular endpoints. This has led to an example of a specialized neurocommons that is well-embraced by the community it aims to serve. In the present paper, we provide a review of the community-based design template and describe the adoption by the community including a high-level review of current data assets, publicly released datasets, and web analytics. Although odc-sci.org is in its late beta stage of development, it represents a successful example of a specialized data commons that may serve as a model for other fields.

Myeloid Arginase 1 Insufficiency Exacerbates Amyloid-ß Associated Neurodegenerative Pathways and Glial Signatures in a Mouse Model of Alzheimer's Disease: A Targeted Transcriptome Analysis.

Brain myeloid cells, include infiltrating macrophages and resident microglia, play an essential role in responding to and inducing neurodegenerative diseases, such as Alzheimer's disease (AD). Genome-wide association studies (GWAS) implicate many AD casual and risk genes enriched in brain myeloid cells. Coordinated arginine metabolism through arginase 1 (Arg1) is critical for brain myeloid cells to perform biological functions, whereas dysregulated arginine metabolism disrupts them. Altered arginine metabolism is proposed as a new biomarker pathway for AD. We previously reported Arg1 deficiency in myeloid biased cells using lysozyme M (LysM) promoter-driven deletion worsened amyloidosis-related neuropathology and behavioral impairment. However, it remains unclear how Arg1 deficiency in these cells impacts the whole brain to promote amyloidosis. Herein, we aim to determine how Arg1 deficiency driven by LysM restriction during amyloidosis affects fundamental neurodegenerative pathways at the transcriptome level. By applying several bioinformatic tools and analyses, we found that amyloid-ß (Aß) stimulated transcriptomic signatures in autophagy-related pathways and myeloid cells' inflammatory response. At the same time, myeloid Arg1 deficiency during amyloidosis promoted gene signatures of lipid metabolism, myelination, and migration of myeloid cells. Focusing on Aß associated glial transcriptomic signatures, we found myeloid Arg1 deficiency up-regulated glial gene transcripts that positively correlated with Aß plaque burden. We also observed that Aß preferentially activated disease-associated microglial signatures to increase phagocytic response, whereas myeloid Arg1 deficiency selectively promoted homeostatic microglial signature that is non-phagocytic. These transcriptomic findings suggest a critical role for proper Arg1 function during normal and pathological challenges associated with amyloidosis. Furthermore, understanding pathways that govern Arg1 metabolism may provide new therapeutic opportunities to rebalance immune function and improve microglia/macrophage fitness.

Acute inflammatory profiles differ with sex and age after spinal cord injury.

BACKGROUND: Sex and age are emerging as influential variables that affect spinal cord injury (SCI) recovery. Despite a changing demographic towards older age at the time of SCI, the effects of sex or age on inflammation remain to be elucidated. This study determined the sex- and age-dependency of the innate immune response acutely after SCI. METHODS: Male and female mice of ages 4- and 14-month-old received T9 contusion SCI and the proportion of microglia, monocyte-derived macrophages (MDM), and neutrophils surrounding the lesion were determined at 3- and 7-day post-injury (DPI) using flow cytometry. Cell counts of microglia and MDMs were obtained using immunohistochemistry to verify flow cytometry results at 3-DPI. Microglia and MDMs were separately isolated using fluorescence-activated cell sorting (FACS) at 3-day post-injury (DPI) to assess RNA expression of 27 genes associated with activation, redox, and debris metabolism/clearance. RESULTS: Flow cytometry revealed that being female and older at the time of injury significantly increased MDMs relative to other phagocytes, specifically increasing the ratio of MDMs to microglia at 3-DPI. Cell counts using immunohistochemistry revealed that male mice have more total microglia within SCI lesions that can account for a lower MDM/microglia ratio. With NanoString analyses of 27 genes, only 1 was differentially expressed between sexes in MDMs; specifically, complement protein C1qa was increased in males. No genes were affected by age in MDMs. Only 2 genes were differentially regulated in microglia between sexes after controlling for false discovery rate, specifically CYBB (NOX2) as a reactive oxygen species (ROS)-associated marker as well as MRC1 (CD206), a gene associated with reparative phenotypes. Both genes were increased in female microglia. No microglial genes were differentially regulated between ages. Differences between microglia and MDMs were found in 26 of 27 genes analyzed, all expressed higher in MDMs with three exceptions. Specifically, C1qa, cPLA2, and CD86 were expressed higher in microglia. CONCLUSIONS: These findings indicate that inflammatory responses to SCI are sex-dependent at both the level of cellular recruitment and gene expression.

Immunomodulatory Effects of Azithromycin Revisited: Potential Applications to COVID-19.

The rapid advancement of the COVID-19 pandemic has prompted an accelerated pursuit to identify effective therapeutics. Stages of the disease course have been defined by viral burden, lung pathology, and progression through phases of the immune response. Immunological factors including inflammatory cell infiltration and cytokine storm have been associated with severe disease and death. Many immunomodulatory therapies for COVID-19 are currently being investigated, and preliminary results support the premise of targeting the immune response. However, because suppressing immune mechanisms could also impact the clearance of the virus in the early stages of infection, therapeutic success is likely to depend on timing with respect to the disease course. Azithromycin is an immunomodulatory drug that has been shown to have antiviral effects and potential benefit in patients with COVID-19. Multiple immunomodulatory effects have been defined for azithromycin which could provide efficacy during the late stages of the disease, including inhibition of pro-inflammatory cytokine production, inhibition of neutrophil influx, induction of regulatory functions of macrophages, and alterations in autophagy. Here we review the published evidence of these mechanisms along with the current clinical use of azithromycin as an immunomodulatory therapeutic. We then discuss the potential impact of azithromycin on the immune response to COVID-19, as well as caution against immunosuppressive and off-target effects including cardiotoxicity in these patients. While azithromycin has the potential to contribute efficacy, its impact on the COVID-19 immune response requires additional characterization so as to better define its role in individualized therapy.

The effects of myelin on macrophage activation are phenotypic specific via cPLA2 in the context of spinal cord injury inflammation.

Spinal cord injury (SCI) produces chronic, pro-inflammatory macrophage activation that impairs recovery. The mechanisms driving this chronic inflammation are not well understood. Here, we detail the effects of myelin debris on macrophage physiology and demonstrate a novel, activation state-dependent role for cytosolic phospholipase-A2 (cPLA2) in myelin-mediated potentiation of pro-inflammatory macrophage activation. We hypothesized that cPLA2 and myelin debris are key mediators of persistent pro-inflammatory macrophage responses after SCI. To test this, we examined spinal cord tissue 28-days after thoracic contusion SCI in 3-month-old female mice and observed both cPLA2 activation and intracellular accumulation of lipid-rich myelin debris in macrophages. In vitro, we utilized bone marrow-derived macrophages to determine myelin's effects across a spectrum of activation states. We observed phenotype-specific responses with myelin potentiating only pro-inflammatory (LPS + INF-γ; M1) macrophage activation, whereas myelin did not induce pro-inflammatory responses in unstimulated or anti-inflammatory (IL-4; M2) macrophages. Specifically, myelin increased levels of pro-inflammatory cytokines, reactive oxygen species, and nitric oxide production in M1 macrophages as well as M1-mediated neurotoxicity. PACOCF3 (cPLA2 inhibitor) blocked myelin's detrimental effects. Collectively, we provide novel spatiotemporal evidence that myelin and cPLA2 play an important role in the pathophysiology of SCI inflammation and the phenotype-specific response to myelin implicate diverse roles of myelin in neuroinflammatory conditions.

Macrophage-Engineered Vesicles for Therapeutic Delivery and Bidirectional Reprogramming of Immune Cell Polarization.

Macrophages, one of the most important phagocytic cells of the immune system, are highly plastic and are known to exhibit diverse roles under different pathological conditions. The ability to repolarize macrophages from pro-inflammatory (M1) to anti-inflammatory (M2) or vice versa offers a promising therapeutic approach for treating various diseases such as traumatic injury and cancer. Herein, it is demonstrated that macrophage-engineered vesicles (MEVs) generated by disruption of macrophage cellular membranes can be used as nanocarriers capable of reprogramming macrophages and microglia toward either pro- or anti-inflammatory phenotypes. MEVs can be produced at high yields and easily loaded with diagnostic molecules or chemotherapeutics and delivered to both macrophages and cancer cells in vitro and in vivo. Overall, MEVs show promise as potential delivery vehicles for both therapeutics and their ability to controllably modulate macrophage/microglia inflammatory phenotypes.