RESUMEN
Toll-like receptors (TLRs) play an important role in the innate immune response after CNS injury. Although TLR4 is one of the best characterized, its role in chronic stages after spinal cord injury (SCI) is not well understood. We examined the role of TLR4 signaling in injury-induced responses at 1â d, 7â d, and 8â weeks after spinal cord contusion injury in adult female TLR4 null and wild-type mice. Analyses include secondary damage, a range of transcriptome and protein analyses of inflammatory, cell death, and extracellular matrix (ECM) molecules, as well as immune cell infiltration and changes in axonal sprouting and locomotor recovery. Lack of TLR4 signaling results in reduced neuronal and myelin loss, reduced activation of NFκB, and decreased expression of inflammatory cytokines and necroptotic cell death pathway at a late time point (8â weeks) after injury. TLR4 null mice also showed reduction of scar-related ECM molecules at 8â weeks after SCI, accompanied by increase in ECM molecules associated with perineuronal nets, increased sprouting of serotonergic fibers, and improved locomotor recovery. These findings reveal novel effects of TLR4 signaling in chronic SCI. We show that TLR4 influences inflammation, cell death, and ECM deposition at late-stage post-injury when secondary injury processes are normally considered to be over. This highlights the potential for late-stage targeting of TLR4 as a potential therapy for chronic SCI.
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Citocinas , Traumatismos de la Médula Espinal , Ratones , Femenino , Animales , Citocinas/metabolismo , Receptor Toll-Like 4/genética , Receptor Toll-Like 4/metabolismo , Neuronas/metabolismo , Inflamación/metabolismo , Ratones Noqueados , Médula Espinal/metabolismo , Recuperación de la Función/fisiologíaRESUMEN
Whereas pathogen-specific T and B cells are a primary focus of interest during infectious disease, we have used COVID-19 to ask whether their emergence comes at a cost of broader B cell and T cell repertoire disruption. We applied a genomic DNA-based approach to concurrently study the immunoglobulin-heavy (IGH) and T cell receptor (TCR) ß and δ chain loci of 95 individuals. Our approach detected anticipated repertoire focusing for the IGH repertoire, including expansions of clusters of related sequences temporally aligned with SARS-CoV-2-specific seroconversion, and enrichment of some shared SARS-CoV-2-associated sequences. No significant age-related or disease severity-related deficiencies were noted for the IGH repertoire. By contrast, whereas focusing occurred at the TCRß and TCRδ loci, including some TCRß sequence-sharing, disruptive repertoire narrowing was almost entirely limited to many patients aged older than 50 y. By temporarily reducing T cell diversity and by risking expansions of nonbeneficial T cells, these traits may constitute an age-related risk factor for COVID-19, including a vulnerability to new variants for which T cells may provide key protection.
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Inmunidad Adaptativa , COVID-19 , Cadenas Pesadas de Inmunoglobulina , Receptores de Antígenos de Linfocitos T alfa-beta , Receptores de Antígenos de Linfocitos T , SARS-CoV-2 , Inmunidad Adaptativa/genética , Anciano , Linfocitos B/inmunología , COVID-19/genética , COVID-19/inmunología , Sitios Genéticos , Humanos , Cadenas Pesadas de Inmunoglobulina/genética , Receptores de Antígenos de Linfocitos T/genética , Receptores de Antígenos de Linfocitos T alfa-beta/genética , SARS-CoV-2/inmunología , Seroconversión , Linfocitos T/inmunologíaRESUMEN
BACKGROUND: The efficacy and safety profiles of vaccines against SARS-CoV-2 in patients with cancer is unknown. We aimed to assess the safety and immunogenicity of the BNT162b2 (Pfizer-BioNTech) vaccine in patients with cancer. METHODS: For this prospective observational study, we recruited patients with cancer and healthy controls (mostly health-care workers) from three London hospitals between Dec 8, 2020, and Feb 18, 2021. Participants who were vaccinated between Dec 8 and Dec 29, 2020, received two 30 µg doses of BNT162b2 administered intramuscularly 21 days apart; patients vaccinated after this date received only one 30 µg dose with a planned follow-up boost at 12 weeks. Blood samples were taken before vaccination and at 3 weeks and 5 weeks after the first vaccination. Where possible, serial nasopharyngeal real-time RT-PCR (rRT-PCR) swab tests were done every 10 days or in cases of symptomatic COVID-19. The coprimary endpoints were seroconversion to SARS-CoV-2 spike (S) protein in patients with cancer following the first vaccination with the BNT162b2 vaccine and the effect of vaccine boosting after 21 days on seroconversion. All participants with available data were included in the safety and immunogenicity analyses. Ongoing follow-up is underway for further blood sampling after the delayed (12-week) vaccine boost. This study is registered with the NHS Health Research Authority and Health and Care Research Wales (REC ID 20/HRA/2031). FINDINGS: 151 patients with cancer (95 patients with solid cancer and 56 patients with haematological cancer) and 54 healthy controls were enrolled. For this interim data analysis of the safety and immunogenicity of vaccinated patients with cancer, samples and data obtained up to March 19, 2021, were analysed. After exclusion of 17 patients who had been exposed to SARS-CoV-2 (detected by either antibody seroconversion or a positive rRT-PCR COVID-19 swab test) from the immunogenicity analysis, the proportion of positive anti-S IgG titres at approximately 21 days following a single vaccine inoculum across the three cohorts were 32 (94%; 95% CI 81-98) of 34 healthy controls; 21 (38%; 26-51) of 56 patients with solid cancer, and eight (18%; 10-32) of 44 patients with haematological cancer. 16 healthy controls, 25 patients with solid cancer, and six patients with haematological cancer received a second dose on day 21. Of the patients with available blood samples 2 weeks following a 21-day vaccine boost, and excluding 17 participants with evidence of previous natural SARS-CoV-2 exposure, 18 (95%; 95% CI 75-99) of 19 patients with solid cancer, 12 (100%; 76-100) of 12 healthy controls, and three (60%; 23-88) of five patients with haematological cancers were seropositive, compared with ten (30%; 17-47) of 33, 18 (86%; 65-95) of 21, and four (11%; 4-25) of 36, respectively, who did not receive a boost. The vaccine was well tolerated; no toxicities were reported in 75 (54%) of 140 patients with cancer following the first dose of BNT162b2, and in 22 (71%) of 31 patients with cancer following the second dose. Similarly, no toxicities were reported in 15 (38%) of 40 healthy controls after the first dose and in five (31%) of 16 after the second dose. Injection-site pain within 7 days following the first dose was the most commonly reported local reaction (23 [35%] of 65 patients with cancer; 12 [48%] of 25 healthy controls). No vaccine-related deaths were reported. INTERPRETATION: In patients with cancer, one dose of the BNT162b2 vaccine yields poor efficacy. Immunogenicity increased significantly in patients with solid cancer within 2 weeks of a vaccine boost at day 21 after the first dose. These data support prioritisation of patients with cancer for an early (day 21) second dose of the BNT162b2 vaccine. FUNDING: King's College London, Cancer Research UK, Wellcome Trust, Rosetrees Trust, and Francis Crick Institute.
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Vacunas contra la COVID-19/uso terapéutico , COVID-19/inmunología , Neoplasias/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Anticuerpos Antivirales/sangre , Vacuna BNT162 , COVID-19/sangre , COVID-19/complicaciones , COVID-19/virología , Vacunas contra la COVID-19/inmunología , Relación Dosis-Respuesta Inmunológica , Femenino , Humanos , Inmunogenicidad Vacunal/inmunología , Londres/epidemiología , Masculino , Persona de Mediana Edad , Neoplasias/sangre , Neoplasias/complicaciones , Neoplasias/virología , Estudios Prospectivos , SARS-CoV-2 , GalesRESUMEN
Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder affecting motoneurons (MNs), with no effective treatment currently available. The molecular mechanisms that are involved in MN death are complex and not fully understood, with partial contributions of surrounding glial cells and skeletal muscle to the disease. Neuregulin 1 (NRG1) is a trophic factor highly expressed in MNs and neuromuscular junctions. Recent studies have suggested a crucial role of the isoform I (NRG1-I) in the collateral reinnervation process in skeletal muscle, and NRG1-III in the preservation of MNs in the spinal cord, opening a window for developing novel therapies for neuromuscular diseases like ALS. In this study, we overexpressed NRG1-I widely in the skeletal muscles of the SOD1G93A transgenic mouse. The results show that NRG1 gene therapy activated the survival pathways in muscle and spinal cord, increasing the number of surviving MNs and neuromuscular junctions and reducing the astroglial reactivity in the spinal cord of the treated SOD1G93A mice. Furthermore, NRG1-I overexpression preserved motor function and delayed the onset of clinical disease. In summary, our data indicates that NRG1 plays an important role on MN survival and muscle innervation in ALS, and that viral-mediated overexpression of NRG1 isoforms may be considered as a promising approach for ALS treatment.
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Esclerosis Amiotrófica Lateral/terapia , Terapia Genética , Neuronas Motoras/metabolismo , Neurregulina-1/genética , Esclerosis Amiotrófica Lateral/metabolismo , Animales , Modelos Animales de Enfermedad , Ratones Transgénicos , Músculo Esquelético/metabolismo , Neuroglía/metabolismo , Unión Neuromuscular/metabolismo , Médula Espinal/metabolismoRESUMEN
Lysophosphatidic acid (LPA) is an extracellular lipid mediator involved in many physiological functions by signaling through six known G-protein-coupled receptors (LPA1-LPA6). In the central nervous system (CNS), LPA mediates a wide range of effects, including neural progenitor cell physiology, astrocyte and microglia activation, neuronal cell death, axonal retraction, and contributions to pain, schizophrenia and hydrocephalus. We recently reported that LPA-LPA1 signaling mediates functional deficits and myelin loss after spinal cord injury (SCI). Here, we provide clear evidence on the deleterious contribution of another LPA receptor, LPA2, to myelin loss after SCI. We found that LPA2 is constitutively expressed in the spinal cord parenchyma and its transcripts were up-regulated after contusion injury, in part, by microglial cells. We also found that the demyelinating lesion triggered by intraspinal injection of LPA into the undamaged spinal cord was markedly reduced in the lack of LPA2. Similarly, LPA2 deficient mice showed enhanced motor skills and myelin sparing after SCI. To gain insights into the detrimental actions of LPA2 in spinal cord we performed cell culture studies. These experiments revealed that, similar to LPA1, activation of microglia LPA2 led to oligodendrocyte cell death. Moreover, we also found that the cytotoxic effects underlaying microglial LPA-LPA2 axis were mediated by the release of purines by microglia and the activation of P2X7 receptor on oligodendrocytes. Overall, this study provides new mechanistic insights into how LPA contributes to SCI physiopathology, and suggest that targeting LPA2 could be a novel therapeutic approach for the treatment of acute SCI.
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Receptores del Ácido Lisofosfatídico/metabolismo , Traumatismos de la Médula Espinal/metabolismo , Animales , Muerte Celular/fisiología , Enfermedades Desmielinizantes , Femenino , Lisofosfolípidos/metabolismo , Ratones , Ratones Endogámicos C57BL , Microglía/metabolismo , Microglía/patología , Neuroinmunomodulación/fisiología , Oligodendroglía/metabolismo , Transducción de Señal , Médula Espinal/metabolismo , Traumatismos de la Médula Espinal/patologíaRESUMEN
IL-37, a member of the IL-1 family, broadly reduces innate inflammation as well as acquired immunity. Whether the antiinflammatory properties of IL-37 extend to the central nervous system remains unknown, however. In the present study, we subjected mice transgenic for human IL-37 (hIL-37tg) and wild-type (WT) mice to spinal cord contusion injury and then treated them with recombinant human IL-37 (rIL-37). In the hIL-37tg mice, the expression of IL-37 was barely detectable in the uninjured cords, but was strongly induced at 24 h and 72 h after the spinal cord injury (SCI). Compared with WT mice, hIL-37tg mice exhibited increased myelin and neuronal sparing and protection against locomotor deficits, including 2.5-fold greater speed in a forced treadmill challenge. Reduced levels of cytokines (e.g., an 80% reduction in IL-6) were observed in the injured cords of hIL-37tg mice, along with lower numbers of blood-borne neutrophils, macrophages, and activated microglia. We treated WT mice with a single intraspinal injection of either full-length or processed rIL-37 after the injury and found that the IL-37-treated mice had significantly enhanced locomotor skills in an open field using the Basso Mouse Scale, as well as supported faster speed on a mechanical treadmill. Treatment with both forms of rIL-37 led to similar beneficial effects on locomotor recovery after SCI. This study presents novel data indicating that IL-37 suppresses inflammation in a clinically relevant model of SCI, and suggests that rIL-37 may have therapeutic potential for the treatment of acute SCI.
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Interleucina-1/uso terapéutico , Traumatismos de la Médula Espinal/tratamiento farmacológico , Animales , Quimiocinas/antagonistas & inhibidores , Quimiocinas/metabolismo , Citocinas/antagonistas & inhibidores , Citocinas/metabolismo , Interleucina-1/genética , Ratones , ARN Mensajero/genética , Traumatismos de la Médula Espinal/metabolismoRESUMEN
Resolution of inflammation is defective after spinal cord injury (SCI), which impairs tissue integrity and remodeling and leads to functional deficits. Effective pharmacological treatments for SCI are not currently available. Maresin 1 (MaR1) is a highly conserved specialized proresolving mediator (SPM) hosting potent anti-inflammatory and proresolving properties with potent tissue regenerative actions. Here, we provide evidence that the inappropriate biosynthesis of SPM in the lesioned spinal cord hampers the resolution of inflammation and leads to deleterious consequences on neurological outcome in adult female mice. We report that, after spinal cord contusion injury in adult female mice, the biosynthesis of SPM is not induced in the lesion site up to 2 weeks after injury. Exogenous administration of MaR1, a highly conserved SPM, propagated inflammatory resolution after SCI, as revealed by accelerated clearance of neutrophils and a reduction in macrophage accumulation at the lesion site. In the search of mechanisms underlying the proresolving actions of MaR1 in SCI, we found that this SPM facilitated several hallmarks of resolution of inflammation, including reduction of proinflammatory cytokines (CXCL1, CXCL2, CCL3, CCL4, IL6, and CSF3), silencing of major inflammatory intracellular signaling cascades (STAT1, STAT3, STAT5, p38, and ERK1/2), redirection of macrophage activation toward a prorepair phenotype, and increase of the phagocytic engulfment of neutrophils by macrophages. Interestingly, MaR1 administration improved locomotor recovery significantly and mitigated secondary injury progression in a clinical relevant model of SCI. These findings suggest that proresolution, immunoresolvent therapies constitute a novel approach to improving neurological recovery after acute SCI.SIGNIFICANCE STATEMENT Inflammation is a protective response to injury or infection. To result in tissue homeostasis, inflammation has to resolve over time. Incomplete or delayed resolution leads to detrimental effects, including propagated tissue damage and impaired wound healing, as occurs after spinal cord injury (SCI). We report that inflammation after SCI is dysregulated in part due to inappropriate synthesis of proresolving lipid mediators. We demonstrate that the administration of the resolution agonist referred to as maresin 1 (MaR1) after SCI actively propagates resolution processes at the lesion site and improves neurological outcome. MaR1 is identified as an interventional candidate to attenuate dysregulated lesional inflammation and to restore functional recovery after SCI.
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Antiinflamatorios/administración & dosificación , Ácidos Docosahexaenoicos/administración & dosificación , Fármacos Neuroprotectores/administración & dosificación , Recuperación de la Función/efectos de los fármacos , Traumatismos de la Médula Espinal/tratamiento farmacológico , Animales , Femenino , Locomoción/efectos de los fármacos , Locomoción/fisiología , Macrófagos/efectos de los fármacos , Macrófagos/fisiología , Ratones , Ratones Endogámicos C57BL , Neutrófilos/efectos de los fármacos , Neutrófilos/fisiología , Recuperación de la Función/fisiología , Traumatismos de la Médula Espinal/fisiopatología , Vértebras TorácicasRESUMEN
Macrophages and microglia play a key role in the maintenance of nervous system homeostasis. However, upon different challenges, they can adopt several phenotypes, which may lead to divergent effects on tissue repair. After spinal cord injury (SCI), microglia and macrophages show predominantly pro-inflammatory activation and contribute to tissue damage. However, the factors that hamper their conversion to an anti-inflammatory state after SCI, or to other protective phenotypes, are poorly understood. Here, we show that IL-4 protein levels are undetectable in the spinal cord after contusion injury, which likely favors microglia and macrophages to remain in a pro-inflammatory state. We also demonstrate that a single delayed intraspinal injection of IL-4, 48 hours after SCI, induces increased expression of M2 marker in microglia and macrophages. We also show that delayed injection of IL-4 leads to the appearance of resolution-phase macrophages, and that IL-4 enhances resolution of inflammation after SCI. Interestingly, we provide clear evidence that delayed administration of IL-4 markedly improves functional outcomes and reduces tissue damage after contusion injury. It is possible that these improvements are mediated by the presence of macrophages with M2 markers and resolution-phase macrophages. These data suggest that therapies aimed at increasing IL-4 levels could be valuable for the treatment of acute SCI, for which there are currently no effective treatments. GLIA 2016;64:2079-2092.
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Interleucina-4/farmacología , Macrófagos/metabolismo , Microglía/metabolismo , Recuperación de la Función/fisiología , Traumatismos de la Médula Espinal/metabolismo , Animales , Arginasa/metabolismo , Citocinas/metabolismo , Potenciales Evocados Motores/efectos de los fármacos , Potenciales Evocados Motores/fisiología , Femenino , Citometría de Flujo , Interleucina-4/metabolismo , Ratones , Ratones Endogámicos C57BL , Músculo Esquelético/fisiopatología , Óxido Nítrico Sintasa de Tipo II/genética , Óxido Nítrico Sintasa de Tipo II/metabolismo , Receptores de Superficie Celular/metabolismo , Recuperación de la Función/efectos de los fármacos , Traumatismos de la Médula Espinal/tratamiento farmacológico , Factores de TiempoRESUMEN
BACKGROUND: It has recently become evident that activating/inhibitory cell surface immune receptors play a critical role in regulating immune and inflammatory processes in the central nervous system (CNS). The immunoreceptor CD300f expressed on monocytes, neutrophils, and mast cells modulates inflammation, phagocytosis, and outcome in models of autoimmune demyelination, allergy, and systemic lupus erythematosus. On the other hand, a finely regulated inflammatory response is essential to induce regeneration after injury to peripheral nerves since hematogenous macrophages, together with resident macrophages and de-differentiated Schwann cells, phagocyte distal axonal and myelin debris in a well-orchestrated inflammatory response. The possible roles and expression of CD300f and its ligands have not been reported under these conditions. METHODS: By using quantitative PCR (QPCR) and CD300f-IgG2a fusion protein, we show the expression of CD300f and its ligands in the normal and crush injured sciatic nerve. The putative role of CD300f in peripheral nerve regeneration was analyzed by blocking receptor-ligand interaction with the same CD300f-IgG2a soluble receptor fusion protein in sciatic nerves of Thy1-YFP-H mice injected at the time of injury. Macrophage M1/M2 polarization phenotype was also analyzed by CD206 and iNOS expression. RESULTS: We found an upregulation of CD300f mRNA and protein expression after injury. Moreover, the ligands are present in restricted membrane patches of Schwann cells, which remain stable after the lesion. The lesioned sciatic nerves of Thy1-YFP-H mice injected with a single dose of CD300f-IgG2a show long lasting effects on nerve regeneration characterized by a lower number of YFP-positive fibres growing into the tibial nerve after 10 days post lesion (dpl) and a delayed functional recovery when compared to PBS- or IgG2a-administered control groups. Animals treated with CD300f-IgG2a show at 10 dpl higher numbers of macrophages and CD206-positive cells and lower levels of iNOS expression than both control groups. At later time points (28 dpl), increased numbers of macrophages and iNOS expression occur. CONCLUSIONS: Taken together, these results show that the pair CD300f ligand is implicated in Wallerian degeneration and nerve regeneration by modulating both the influx and phenotype of macrophages.
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Inflamación/patología , Macrófagos/patología , Regeneración Nerviosa , Receptores Inmunológicos/genética , Animales , Axones/patología , Células CHO , Cricetinae , Cricetulus , Femenino , Inmunoglobulina G/biosíntesis , Inmunoglobulina G/genética , Lectinas Tipo C/metabolismo , Ligandos , Masculino , Receptor de Manosa , Lectinas de Unión a Manosa/metabolismo , Ratones , Compresión Nerviosa , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Nervios Periféricos/patología , Fagocitosis , Fenotipo , Receptores de Superficie Celular/metabolismo , Células de Schwann/patología , Neuropatía Ciática/patología , Degeneración Walleriana/patologíaRESUMEN
αB-crystallin is a member of the heat shock protein family that exerts cell protection under several stress-related conditions. Recent studies have revealed that αB-crystallin plays a beneficial role in a mouse model of multiple sclerosis, brain ischemia, and Alexander disease. Whether αB-crystallin plays a role in modulating the secondary damage after CNS trauma is not known. We report here that αB-crystallin mediates protective effects after spinal cord injury. The levels of αB-crystallin are reduced in spinal cord tissue following contusion lesion. In addition, administration of recombinant human αB-crystallin for the first week after contusion injury leads to sustained improvement in locomotor skills and amelioration of secondary tissue damage. We also provide evidence that recombinant human αB-crystallin modulates the inflammatory response in the injured spinal cord, leading to increased infiltration of granulocytes and reduced recruitment of inflammatory macrophages. Furthermore, the delivery of recombinant human αB-crystallin promotes greater locomotor recovery even when the treatment is initiated 6 h after spinal cord injury. Our findings suggest that administration of recombinant human αB-crystallin may be a good therapeutic approach for treating acute spinal cord injury, for which there is currently no effective treatment.
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Traumatismos de la Médula Espinal/patología , Traumatismos de la Médula Espinal/terapia , Cadena B de alfa-Cristalina/uso terapéutico , Animales , Inhibición de Migración Celular/fisiología , Regulación hacia Abajo/fisiología , Femenino , Granulocitos/patología , Humanos , Mediadores de Inflamación/uso terapéutico , Macrófagos/patología , Ratones , Ratones Endogámicos C57BL , Ratas , Proteínas Recombinantes/uso terapéutico , Traumatismos de la Médula Espinal/metabolismo , Resultado del Tratamiento , Regulación hacia Arriba/fisiología , Cadena B de alfa-Cristalina/antagonistas & inhibidores , Cadena B de alfa-Cristalina/biosíntesisRESUMEN
Spinal cord injury (SCI) leads to irreversible functional deficits due to the disruption of axons and the death of neurons and glial cells. The inflammatory response that occurs in the injured spinal cord results in tissue degeneration; thus, targeting inflammation after acute SCI is expected to ameliorate histopathological evidence indicative of damage and, consequently, reduce functional disabilities. Interleukin 1 beta (IL-1ß) and interleukin 18 (IL-18) are pro-inflammatory cytokines members of the IL-1 family that initiate and propagate inflammation. Here, we report that protein levels of IL-1ß and IL-18 were increased in spinal cord parenchyma after SCI, but with different expression profiles. Whereas levels of IL-1ß were rapidly increased reaching peak levels at 12 h after the injury, levels of IL-18 did not increase until 7 days after the injury. Since activation of the NLRP3 inflammasome is required for the processing and release of IL-1ß and IL-18, we intraperitoneally administered OLT1177, a selective inhibitor of the NLRP3 inflammasome, to reduce the contribution of these cytokines to SCI. At a dose of 200 mg/kg, OLT1177 protected against neurological deficits and histological evidence of damage. OLT1177 also reduced the levels of IL-1ß in the spinal cord after contusion injury and diminished the accumulation of neutrophils and macrophages at later time points. These data suggest that targeting the NLRP3 inflammasome with OLT1177 could be a novel therapeutic strategy to arrest neuroinflammation and reduce functional impairments after acute SCI in humans.
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Antiinflamatorios/farmacología , Inflamasomas/efectos de los fármacos , Vaina de Mielina/patología , Nitrilos/farmacología , Traumatismos de la Médula Espinal/inmunología , Sulfonas/farmacología , Animales , Femenino , Ratones , Proteína con Dominio Pirina 3 de la Familia NLR/inmunología , Enfermedades Neuroinflamatorias/inmunología , Enfermedades Neuroinflamatorias/patología , Traumatismos de la Médula Espinal/patologíaRESUMEN
Chondroitin sulfate proteoglycans (CSPGs) act as potent inhibitors of axonal growth and neuroplasticity after spinal cord injury (SCI). Here we reveal that CSPGs also play a critical role in preventing inflammation resolution by blocking the conversion of pro-inflammatory immune cells to a pro-repair phenotype in rodent models of SCI. We demonstrate that enzymatic digestion of CSPG glycosaminoglycans enhances immune cell clearance and reduces pro-inflammatory protein and gene expression profiles at key resolution time points. Analysis of phenotypically distinct immune cell clusters revealed CSPG-mediated modulation of macrophage and microglial subtypes which, together with T lymphocyte infiltration and composition changes, suggests a role for CSPGs in modulating both innate and adaptive immune responses after SCI. Mechanistically, CSPG activation of a pro-inflammatory phenotype in pro-repair immune cells was found to be TLR4-dependent, identifying TLR4 signalling as a key driver of CSPG-mediated immune modulation. These findings establish CSPGs as critical mediators of inflammation resolution failure after SCI in rodents, which leads to prolonged inflammatory pathology and irreversible tissue destruction.
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Proteoglicanos Tipo Condroitín Sulfato , Traumatismos de la Médula Espinal , Animales , Proteoglicanos Tipo Condroitín Sulfato/metabolismo , Inflamación , Roedores , Traumatismos de la Médula Espinal/patología , Receptor Toll-Like 4/genéticaRESUMEN
Research on microglia has established the differentiation between the so-called M1 and M2 phenotypes. However, new frameworks have been proposed attempting to discern between meaningful microglia profiles. We have set up an in vitro microglial activation model by adding an injured spinal cord (SCI) lysate to microglial cultures, obtained from postnatal rats, in order to mimic the environment of the spinal cord after injury. We found that under the presence of the SCI lysate microglial cells changed their phenotype, developing less ramified but longer processes, and proliferated. The SCI lysate also led to upregulation of pro-inflammatory cytokines, such as IL-1ß, IL-6, and TNF-α, downregulation of the anti-inflammatory cytokines IL-10 and IL-4, and a biphasic profile of iNOS. In addition, a latex beads phagocytosis assay revealed the SCI lysate stimulated the phagocytic capacity of microglia. Flow cytometry analysis indicated that microglial cells showed a pro-inflammatory profile in the presence of SCI lysate. Finally, characterization of the microglial activation in the spinal cord on day 7 after contusion injury, we showed that these cells have a pro-inflammatory phenotype. Overall, these results indicate that the use of SCI lysates could be a useful tool to skew microglia towards a closer phenotype to that observed after the spinal cord contusion injury than the use of LPS or IFNγ.
RESUMEN
Background: Microglia and macrophages adopt a pro-inflammatory phenotype after spinal cord injury (SCI), what is thought to contribute to secondary tissue degeneration. We previously reported that this is due, in part, to the low levels of anti-inflammatory cytokines, such as IL-4. Since IL-13 and IL-4 share receptors and both cytokines drive microglia and macrophages towards an anti-inflammatory phenotype in vitro, here we studied whether administration of IL-13 and IL-4 after SCI leads to beneficial effects. Methods: We injected mice with recombinant IL-13 or IL-4 at 48 h after SCI and assessed their effects on microglia and macrophage phenotype and functional outcomes. We also performed RNA sequencing analysis of macrophages and microglia sorted from the injured spinal cords of mice treated with IL-13 or IL-4 and evaluated the metabolic state of these cells by using Seahorse technology. Results: We observed that IL-13 induced the expression of anti-inflammatory markers in microglia and macrophages after SCI but, in contrast to IL-4, it failed to mediate functional recovery. We found that these two cytokines induced different gene signatures in microglia and macrophages after SCI and that IL-4, in contrast to IL-13, shifted microglia and macrophage metabolism from glycolytic to oxidative phosphorylation. These findings were further confirmed by measuring the metabolic profile of these cells. Importantly, we also revealed that macrophages stimulated with IL-4 or IL-13 are not deleterious to neurons, but they become cytotoxic when oxidative metabolism is blocked. This suggests that the metabolic shift, from glycolysis to oxidative phosphorylation, is required to minimize the cytotoxic responses of microglia and macrophages. Conclusions: These results reveal that the metabolic fitness of microglia and macrophages after SCI contributes to secondary damage and that strategies aimed at boosting oxidative phosphorylation might be a novel approach to minimize the deleterious actions of microglia and macrophages in neurotrauma.
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Interleucina-13/metabolismo , Interleucina-4/metabolismo , Traumatismos de la Médula Espinal/metabolismo , Animales , Antiinflamatorios/farmacología , Citocinas/metabolismo , Modelos Animales de Enfermedad , Femenino , Interleucina-13/inmunología , Interleucina-13/farmacología , Interleucina-4/inmunología , Interleucina-4/farmacología , Macrófagos/metabolismo , Ratones , Ratones Endogámicos C57BL , Microglía/metabolismo , Recuperación de la Función/fisiología , Médula Espinal/metabolismo , Traumatismos de la Médula Espinal/inmunología , Traumatismos de la Médula Espinal/fisiopatología , Resultado del TratamientoRESUMEN
Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease affecting the neuromuscular system for which currently there is no effective therapy. Motoneuron (MN) degeneration involves several complex mechanisms, including surrounding glial cells and skeletal muscle contributions. Neuregulin 1 (NRG1) is a trophic factor present particularly in MNs and neuromuscular junctions. Our previous studies revealed that gene therapy overexpressing the isoform I (NRG1-I) in skeletal muscles as well as overexpressing the isoform III (NRG1-III) directly in the central nervous system are both effective in preserving MNs in the spinal cord of ALS mice, opening novel therapeutic approaches. In this study, we combined administration of both viral vectors overexpressing NRG1-I in skeletal muscles and NRG1-III in spinal cord of the SOD1G93A mice in order to obtain a synergistic effect. The results showed that the combinatorial gene therapy increased preservation of MNs and of innervated neuromuscular junctions and reduced glial reactivity in the spinal cord of the treated SOD1G93A mice. Moreover, NRG1 isoforms overexpression improved motor function of hindlimb muscles and delayed the onset of clinical disease. However, this combinatory gene therapy did not produce a synergic effect compared with single therapies, suggesting an overlap between NRG1-I and NRG1-III activated pathways and their beneficial effects.
RESUMEN
Given the immune system's importance for cancer surveillance and treatment, we have investigated how it may be affected by SARS-CoV-2 infection of cancer patients. Across some heterogeneity in tumor type, stage, and treatment, virus-exposed solid cancer patients display a dominant impact of SARS-CoV-2, apparent from the resemblance of their immune signatures to those for COVID-19+ non-cancer patients. This is not the case for hematological malignancies, with virus-exposed patients collectively displaying heterogeneous humoral responses, an exhausted T cell phenotype and a high prevalence of prolonged virus shedding. Furthermore, while recovered solid cancer patients' immunophenotypes resemble those of non-virus-exposed cancer patients, recovered hematological cancer patients display distinct, lingering immunological legacies. Thus, while solid cancer patients, including those with advanced disease, seem no more at risk of SARS-CoV-2-associated immune dysregulation than the general population, hematological cancer patients show complex immunological consequences of SARS-CoV-2 exposure that might usefully inform their care.
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COVID-19/inmunología , Neoplasias/inmunología , Neoplasias/virología , Síndrome Respiratorio Agudo Grave/inmunología , Adulto , Anciano , Anciano de 80 o más Años , COVID-19/etiología , COVID-19/mortalidad , Femenino , Neoplasias Hematológicas/inmunología , Neoplasias Hematológicas/mortalidad , Neoplasias Hematológicas/terapia , Neoplasias Hematológicas/virología , Humanos , Inmunofenotipificación , Masculino , Persona de Mediana Edad , Nasofaringe/virología , Neoplasias/mortalidad , Neoplasias/terapia , Síndrome Respiratorio Agudo Grave/etiología , Síndrome Respiratorio Agudo Grave/mortalidad , Síndrome Respiratorio Agudo Grave/virología , Linfocitos T/virología , Esparcimiento de Virus , Adulto JovenRESUMEN
A Correction to this paper has been published: https://doi.org/10.1038/s41591-020-01186-5.
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An amendment to this paper has been published and can be accessed via a link at the top of the paper.
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Improved understanding and management of COVID-19, a potentially life-threatening disease, could greatly reduce the threat posed by its etiologic agent, SARS-CoV-2. Toward this end, we have identified a core peripheral blood immune signature across 63 hospital-treated patients with COVID-19 who were otherwise highly heterogeneous. The signature includes discrete changes in B and myelomonocytic cell composition, profoundly altered T cell phenotypes, selective cytokine/chemokine upregulation and SARS-CoV-2-specific antibodies. Some signature traits identify links with other settings of immunoprotection and immunopathology; others, including basophil and plasmacytoid dendritic cell depletion, correlate strongly with disease severity; while a third set of traits, including a triad of IP-10, interleukin-10 and interleukin-6, anticipate subsequent clinical progression. Hence, contingent upon independent validation in other COVID-19 cohorts, individual traits within this signature may collectively and individually guide treatment options; offer insights into COVID-19 pathogenesis; and aid early, risk-based patient stratification that is particularly beneficial in phasic diseases such as COVID-19.