RESUMEN
Traumatic spinal cord injury (SCI) most often leads to permanent paralysis due to the inability of axons to regenerate in the adult mammalian central nervous system (CNS). In the past, we have shown that mast cells (MCs) improve the functional outcome after SCI by suppressing scar tissue formation at the lesion site via mouse mast cell protease 6 (mMCP6). In this study, we investigated whether recombinant mMCP6 can be used therapeutically to improve the functional outcome after SCI. Therefore, we applied mMCP6 locally via an intrathecal catheter in the subacute phase after a spinal cord hemisection injury in mice. Our findings showed that hind limb motor function was significantly improved in mice that received recombinant mMCP6 compared with the vehicle-treated group. In contrast to our previous findings in mMCP6 knockout mice, the lesion size and expression levels of the scar components fibronectin, laminin, and axon-growth-inhibitory chondroitin sulfate proteoglycans were not affected by the treatment with recombinant mMCP6. Surprisingly, no difference in infiltration of CD4+ T cells and reactivity of Iba-1+ microglia/macrophages at the lesion site was observed between the mMCP6-treated mice and control mice. Additionally, local protein levels of the pro- and anti-inflammatory mediators IL-1ß, IL-2, IL-4, IL-6, IL-10, TNF-α, IFNγ, and MCP-1 were comparable between the two treatment groups, indicating that locally applied mMCP6 did not affect inflammatory processes after injury. However, the increase in locomotor performance in mMCP6-treated mice was accompanied by reduced demyelination and astrogliosis in the perilesional area after SCI. Consistently, we found that TNF-α/IL-1ß-astrocyte activation was decreased and that oligodendrocyte precursor cell (OPC) differentiation was increased after recombinant mMCP6 treatment in vitro. Mechanistically, this suggests effects of mMCP6 on reducing astrogliosis and improving (re)myelination in the spinal cord after injury. In conclusion, these data show for the first time that recombinant mMCP6 is therapeutically active in enhancing recovery after SCI.
Asunto(s)
Remielinización , Traumatismos de la Médula Espinal , Ratones , Animales , Gliosis/tratamiento farmacológico , Gliosis/metabolismo , Cicatriz/tratamiento farmacológico , Cicatriz/prevención & control , Mastocitos/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Traumatismos de la Médula Espinal/tratamiento farmacológico , Traumatismos de la Médula Espinal/metabolismo , Médula Espinal/metabolismo , Ratones Noqueados , Recuperación de la Función , Modelos Animales de Enfermedad , MamíferosRESUMEN
BACKGROUND: Spinal cord injury (SCI) elicits a robust neuroinflammatory reaction which, in turn, exacerbates the initial mechanical damage. Pivotal players orchestrating this response are macrophages (Mφs) and microglia. After SCI, the inflammatory environment is dominated by pro-inflammatory Mφs/microglia, which contribute to secondary cell death and prevent regeneration. Therefore, reprogramming Mφ/microglia towards a more anti-inflammatory and potentially neuroprotective phenotype has gained substantial therapeutic interest in recent years. Interleukin-13 (IL-13) is a potent inducer of such an anti-inflammatory phenotype. In this study, we used genetically modified Mφs as carriers to continuously secrete IL-13 (IL-13 Mφs) at the lesion site. METHODS: Mφs were genetically modified to secrete IL-13 (IL-13 Mφs) and were phenotypically characterized using qPCR, western blot, and ELISA. To analyze the therapeutic potential, the IL-13 Mφs were intraspinally injected at the perilesional area after hemisection SCI in female mice. Functional recovery and histopathological improvements were evaluated using the Basso Mouse Scale score and immunohistochemistry. Neuroprotective effects of IL-13 were investigated using different cell viability assays in murine and human neuroblastoma cell lines, human neurospheroids, as well as murine organotypic brain slice cultures. RESULTS: In contrast to Mφs prestimulated with recombinant IL-13, perilesional transplantation of IL-13 Mφs promoted functional recovery following SCI in mice. This improvement was accompanied by reduced lesion size and demyelinated area. The local anti-inflammatory shift induced by IL-13 Mφs resulted in reduced neuronal death and fewer contacts between dystrophic axons and Mφs/microglia, suggesting suppression of axonal dieback. Using IL-4Rα-deficient mice, we show that IL-13 signaling is required for these beneficial effects. Whereas direct neuroprotective effects of IL-13 on murine and human neuroblastoma cell lines or human neurospheroid cultures were absent, IL-13 rescued murine organotypic brain slices from cell death, probably by indirectly modulating the Mφ/microglia responses. CONCLUSIONS: Collectively, our data suggest that the IL-13-induced anti-inflammatory Mφ/microglia phenotype can preserve neuronal tissue and ameliorate axonal dieback, thereby promoting recovery after SCI.
Asunto(s)
Neuroblastoma , Fármacos Neuroprotectores , Traumatismos de la Médula Espinal , Animales , Femenino , Humanos , Interleucina-13/uso terapéutico , Macrófagos/metabolismo , Ratones , Fármacos Neuroprotectores/uso terapéutico , Traumatismos de la Médula Espinal/patologíaRESUMEN
Mast cells (MCs), which are well known for their effector functions in TH2-skewed allergic and also autoimmune inflammation, have become increasingly acknowledged for their role in protection of health. It is now clear that they are also key modulators of immune responses at interface organs, such as the skin or gut. MCs can prime tissues for adequate inflammatory responses and cooperate with dendritic cells in T-cell activation. They also regulate harmful immune responses in trauma and help to successfully orchestrate pregnancy. This review focuses on the beneficial effects of MCs on tissue homeostasis and elimination of toxins or venoms. MCs can enhance pathogen clearance in many bacterial, viral, and parasitic infections, such as through Toll-like receptor 2-triggered degranulation, secretion of antimicrobial cathelicidins, neutrophil recruitment, or provision of extracellular DNA traps. The role of MCs in tumors is more ambiguous; however, encouraging new findings show they can change the tumor microenvironment toward antitumor immunity when adequately triggered. Uterine tissue remodeling by α-chymase (mast cell protease [MCP] 5) is crucial for successful embryo implantation. MCP-4 and the tryptase MCP-6 emerge to be protective in central nervous system trauma by reducing inflammatory damage and excessive scar formation, thereby protecting axon growth. Last but not least, proteases, such as carboxypeptidase A, released by FcεRI-activated MCs detoxify an increasing number of venoms and endogenous toxins. A better understanding of the plasticity of MCs will help improve these advantageous effects and hint at ways to cut down detrimental MC actions.
Asunto(s)
Inmunidad Innata , Infecciones/inmunología , Mastocitos/inmunología , Animales , Catelicidinas/metabolismo , Degranulación de la Célula , Implantación del Embrión , Femenino , Homeostasis , Humanos , Embarazo , Receptor Toll-Like 2/metabolismoRESUMEN
Pan-histone deacetylase (HDAC) inhibition with valproic acid (VPA) has beneficial effects after spinal cord injury (SCI), although with side effects. We focused on specific HDAC8 inhibition, because it is known to reduce anti-inflammatory mediators produced by macrophages (Mφ). We hypothesized that HDAC8 inhibition improves functional recovery after SCI by reducing pro-inflammatory classically activated Mφ. Specific HDAC8 inhibition with PCI-34051 reduced the numbers of perilesional Mφ as measured by histological analyses, but did not improve functional recovery (Basso Mouse Scale). We could not reproduce the published improvement of functional recovery described in contusion SCI models using VPA in our T-cut hemisection SCI model. The presence of spared fibers might be the underlying reason for the conflicting data in different SCI models.
Asunto(s)
Inhibidores de Histona Desacetilasas/farmacología , Histona Desacetilasas/metabolismo , Traumatismos de la Médula Espinal/metabolismo , Animales , Antiinflamatorios/farmacología , Ácidos Hidroxámicos/farmacología , Indoles/farmacología , Inflamación/tratamiento farmacológico , Macrófagos/inmunología , Macrófagos/metabolismo , Ratones , Recuperación de la Función/efectos de los fármacos , Médula Espinal/patología , Traumatismos de la Médula Espinal/tratamiento farmacológico , Traumatismos de la Médula Espinal/inmunología , Ácido Valproico/farmacologíaRESUMEN
A disintegrin and metalloproteinase 17 (ADAM17) is the major sheddase involved in the cleavage of a plethora of cytokines, cytokine receptors and growth factors, thereby playing a substantial role in inflammatory and regenerative processes after central nervous system trauma. By making use of a hypomorphic ADAM17 knockin mouse model as well as pharmacological ADAM10/ADAM17 inhibitors, we showed that ADAM17-deficiency or inhibition significantly increases clearance of apoptotic cells, promotes axon growth and improves functional recovery after spinal cord injury (SCI) in mice. Microglia-specific ADAM17-knockout (ADAM17flox+/+-Cx3Cr1 Cre+/-) mice also showed improved functional recovery similar to hypomorphic ADAM17 mice. In contrast, endothelial-specific (ADAM17flox+/+-Cdh5Pacs Cre+/-) and macrophage-specific (ADAM17flox+/+-LysM Cre+/-) ADAM17-knockout mice or bone marrow chimera with transplanted ADAM17-deficient macrophages, displayed no functional improvement compared to wild type mice. These data indicate that ADAM17 expression on microglia cells (and not on macrophages or endothelial cells) plays a detrimental role in inflammation and functional recovery after SCI.
Asunto(s)
Proteína ADAM17/metabolismo , Microglía/metabolismo , Traumatismos de la Médula Espinal/metabolismo , Animales , Modelos Animales de Enfermedad , Femenino , Inflamación/metabolismo , Macrófagos/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Fagocitosis/inmunología , Fagocitosis/fisiología , Recuperación de la Función/fisiologíaRESUMEN
Angiogenesis is associated with changes in endothelial cell (EC) proliferation and tube formation, controlled by extracellular receptor-activated kinase (ERK)/mitogen activated protein kinase (MAPK) and Akt signaling. Important regulators of these systems include hormones acting on G-protein-coupled receptors, such as beta 2-adrenoceptors (ß2-ARs). In central nervous system (CNS) trauma, the importance of ß2-AR modulation has been highlighted, although the effects on revascularization remain unclear. Vascular protection and revascularization are, however, key to support regeneration. We have investigated the angiogenic capacity of the specific ß2-AR agonist terbutaline on ECs derived from the CNS, namely bEnd.3-cells. As angiogenesis is a multistep process involving increased proliferation and tube formation of ECs, we investigated the effects of terbutaline on these processes. We show that terbutaline significantly induced bEnd.3 tube formation in a matrigel in vitro assay. Moreover, administration of specific inhibitors of ERK and Akt signaling both inhibited terbutaline-induced tube formation. The proliferation rate of the ECs was not affected. In order to investigate the general effects of terbutaline in an organotypic system, we have used the chick chorioallantoic membrane (CAM)-assay. Most importantly, terbutaline increased the number of blood vessels in this in ovo setting. Although we observed a positive trend, the systemic administration of terbutaline did not significantly improve the functional outcome, nor did it affect revascularization in our spinal cord injury model. In conclusion, these data indicate that terbutaline is promising to stimulate blood vessel formation, underscoring the importance of further research into the angiotherapeutic relevance of terbutaline and ß2-AR signaling after CNS-trauma. J. Cell. Physiol. 232: 298-308, 2017. © 2016 Wiley Periodicals, Inc.
Asunto(s)
Agonistas de Receptores Adrenérgicos beta 2/farmacología , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Neovascularización Fisiológica/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptores Adrenérgicos beta 2/metabolismo , Transducción de Señal/efectos de los fármacos , Terbutalina/farmacología , Animales , Línea Celular , Proliferación Celular/efectos de los fármacos , Pollos , Membrana Corioalantoides/efectos de los fármacos , Membrana Corioalantoides/metabolismo , Células Endoteliales/citología , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Ratones , Modelos Biológicos , Regeneración/efectos de los fármacos , Traumatismos de la Médula Espinal/tratamiento farmacológico , Traumatismos de la Médula Espinal/patología , Terbutalina/uso terapéuticoRESUMEN
An important barrier for axon regeneration and recovery after traumatic spinal cord injury (SCI) is attributed to the scar that is formed at the lesion site. Here, we investigated the effect of mouse mast cell protease (mMCP) 6, a mast cell (MC)-specific tryptase, on scarring and functional recovery after a spinal cord hemisection injury. Functional recovery was significantly impaired in both MC-deficient and mMCP6-knockout (mMCP6(-/-)) mice after SCI compared with wild-type control mice. This decrease in locomotor performance was associated with an increased lesion size and excessive scarring at the injury site. Axon growth-inhibitory chondroitin sulfate proteoglycans and the extracellular matrix components fibronectin, laminin, and collagen IV were significantly up-regulated in MC-deficient and mMCP6(-/-) mice, with an increase in scar volume between 23 and 32%. A degradation assay revealed that mMCP6 directly cleaves fibronectin and collagen IV in vitro In addition, gene expression levels of the scar components fibronectin, aggrecan, and collagen IV were increased up to 6.8-fold in mMCP6(-/-) mice in the subacute phase after injury. These data indicate that endogenous mMCP6 has scar-suppressing properties after SCI via indirect cleavage of axon growth-inhibitory scar components and alteration of the gene expression profile of these factors.-Vangansewinkel, T., Geurts, N., Quanten, K., Nelissen, S., Lemmens, S., Geboes, L., Dooley, D., Vidal, P. M., Pejler, G., Hendrix, S. Mast cells promote scar remodeling and functional recovery after spinal cord injury via mouse mast cell protease 6.
Asunto(s)
Cicatriz/metabolismo , Mastocitos/fisiología , Traumatismos de la Médula Espinal/metabolismo , Triptasas/metabolismo , Cicatrización de Heridas/fisiología , Animales , Citocinas/genética , Citocinas/metabolismo , Matriz Extracelular , Regulación Enzimológica de la Expresión Génica/fisiología , Ratones , Ratones Noqueados , ARN Mensajero/genética , ARN Mensajero/metabolismo , Triptasas/genéticaRESUMEN
Many researchers have attempted to pharmacologically modulate the adrenergic system to control locomotion, pain, and spasms after central nervous system (CNS) trauma, although such efforts have led to conflicting results. Despite this, multiple studies highlight that α-adrenoceptors (α-ARs) are promising therapeutic targets because in the CNS, they are involved in reactivity to stressors and regulation of locomotion, pain, and spasms. These functions can be activated by direct modulation of these receptors on neuronal networks in the brain and the spinal cord. In addition, these multifunctional receptors are also broadly expressed on immune cells. This suggests that they might play a key role in modulating immunological responses, which may be crucial in treating spinal cord injury and traumatic brain injury as both diseases are characterized by a strong inflammatory component. Reducing the proinflammatory response will create a more permissive environment for axon regeneration and may support neuromodulation in combination therapies. However, pharmacological interventions are hindered by adrenergic system complexity and the even more complicated anatomical and physiological changes in the CNS after trauma. This review is the first concise overview of the pros and cons of α-AR modulation in the context of CNS trauma.
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Dolor/metabolismo , Parálisis/metabolismo , Receptores Adrenérgicos alfa/metabolismo , Espasmo/metabolismo , Traumatismos del Sistema Nervioso/metabolismo , Animales , Lesiones Encefálicas/complicaciones , Lesiones Encefálicas/metabolismo , Humanos , Dolor/complicaciones , Parálisis/complicaciones , Espasmo/complicaciones , Traumatismos del Sistema Nervioso/complicacionesRESUMEN
Mast cells (MCs) are found abundantly in the central nervous system and play a complex role in neuroinflammatory diseases such as multiple sclerosis and stroke. In the present study, we show that MC-deficient Kit(W-sh/W-sh) mice display significantly increased astrogliosis and T cell infiltration as well as significantly reduced functional recovery after spinal cord injury compared to wildtype mice. In addition, MC-deficient mice show significantly increased levels of MCP-1, TNF-α, IL-10 and IL-13 protein levels in the spinal cord. Mice deficient in mouse mast cell protease 4 (mMCP4), an MC-specific chymase, also showed increased MCP-1, IL-6 and IL-13 protein levels in spinal cord samples and a decreased functional outcome after spinal cord injury. A degradation assay using supernatant from MCs derived from either mMCP4(-/-) mice or controls revealed that mMCP4 cleaves MCP-1, IL-6, and IL-13 suggesting a protective role for MC proteases in neuroinflammation. These data show for the first time that MCs may be protective after spinal cord injury and that they may reduce CNS damage by degrading inflammation-associated cytokines via the MC-specific chymase mMCP4.
Asunto(s)
Citocinas/metabolismo , Mastocitos/metabolismo , Serina Endopeptidasas/metabolismo , Traumatismos de la Médula Espinal/metabolismo , Animales , Astrocitos/patología , Femenino , Mediadores de Inflamación/metabolismo , Locomoción/fisiología , Ratones Endogámicos C57BL , Ratones Noqueados , Traumatismos de la Médula Espinal/patología , Linfocitos T/metabolismo , Vértebras Torácicas/lesionesRESUMEN
Spinal cord injury (SCI) is a life-changing event that severely impacts the patient's quality of life. Modulating neuroinflammation, which exacerbates the primary injury, and stimulating neuro-regenerative repair mechanisms are key strategies to improve functional recovery. Cyclic adenosine monophosphate (cAMP) is a second messenger crucially involved in both processes. Following SCI, intracellular levels of cAMP are known to decrease over time. Therefore, preventing cAMP degradation represents a promising strategy to suppress inflammation while stimulating regeneration. Intracellular cAMP levels are controlled by its hydrolyzing enzymes phosphodiesterases (PDEs). The PDE4 family is most abundantly expressed in the central nervous system (CNS) and its inhibition has been shown to be therapeutically relevant for managing SCI pathology. Unfortunately, the use of full PDE4 inhibitors at therapeutic doses is associated with severe emetic side effects, hampering their translation toward clinical applications. Therefore, in this study, we evaluated the effect of inhibiting specific PDE4 subtypes (PDE4B and PDE4D) on inflammatory and regenerative processes following SCI, as inhibitors selective for these subtypes have been demonstrated to be well-tolerated. We reveal that administration of the PDE4D inhibitor Gebr32a, even when starting 2 dpi, but not the PDE4B inhibitor A33, improved functional as well as histopathological outcomes after SCI, comparable to results obtained with the full PDE4 inhibitor roflumilast. Furthermore, using a luminescent human iPSC-derived neurospheroid model, we show that PDE4D inhibition stabilizes neural viability by preventing apoptosis and stimulating neuronal differentiation. These findings strongly suggest that specific PDE4D inhibition offers a novel therapeutic approach for SCI.
RESUMEN
L-arginine is a semi-essential amino acid involved in a variety of physiological processes in the central nervous system (CNS). It is essential in the survival and functionality of neuronal cells. Nonetheless, L-arginine also has a dark side; it potentiates neuroinflammation and nitric oxide (NO) production, leading to secondary damage. Therefore, modulating the L-arginine metabolism is challenging because both detrimental and beneficial effects are dependent on this semi-essential amino acid. After spinal cord injury (SCI), L-arginine plays a crucial role in trauma-induced neuroinflammation and regenerative processes via the two key enzymes: nitric oxide synthase (NOS) and arginase (ARG). Studies on L-arginine metabolism using ARG and NOS inhibitors highlighted the conflicting role of this semi-essential amino acid. Similarly, L-arginine supplementation resulted in both negative and positive outcomes after SCI. However, new data indicate that arginine depletion substantially improves spinal cord regeneration after injury. Here, we review the challenging characteristics of L-arginine metabolism as a therapeutic target after SCI.
Asunto(s)
Enfermedades Neuroinflamatorias , Traumatismos de la Médula Espinal , Humanos , Arginina/metabolismo , Arginina/farmacología , Traumatismos de la Médula Espinal/tratamiento farmacológico , Traumatismos de la Médula Espinal/metabolismo , Óxido Nítrico Sintasa/metabolismo , Óxido Nítrico Sintasa/farmacología , Sistema Nervioso Central/metabolismo , Médula EspinalRESUMEN
In recent years, the gut-central nervous system axis has emerged as a key factor in the pathophysiology of spinal cord injury (SCI). Interleukin-13 (IL-13) has been shown to have anti-inflammatory and neuroprotective effects in SCI. The aim of this study was to investigate the changes in microbiota composition after hemisection injury and to determine whether systemic recombinant (r)IL-13 treatment could alter the gut microbiome, indirectly promoting functional recovery. The gut microbiota composition was determined by 16S rRNA gene sequencing, and correlations between gut microbiota alterations and functional recovery were assessed. Our results showed that there were no changes in alpha diversity between the groups before and after SCI, while PERMANOVA analysis for beta diversity showed significant differences in fecal microbial communities. Phylogenetic classification of bacterial families revealed a lower abundance of the Bacteroidales S24-7 group and a higher abundance of Lachnospiraceae and Lactobacillaceae in the post-SCI group. Systemic rIL-13 treatment improved functional recovery 28 days post-injury and microbiota analysis revealed increased relative abundance of Clostridiales vadin BB60 and Acetitomaculum and decreased Anaeroplasma, Ruminiclostridium_6, and Ruminococcus compared to controls. Functional assessment with PICRUSt showed that genes related to glyoxylate cycle and palmitoleate biosynthesis-I were the predominant signatures in the rIL-13-treated group, whereas sulfolactate degradation super pathway and formaldehyde assimilation-I were enriched in controls. In conclusion, our results indicate that rIL-13 treatment promotes changes in gut microbial communities and may thereby contribute indirectly to the improvement of functional recovery in mice, possibly having important implications for the development of novel treatment options for SCI.
Asunto(s)
Microbioma Gastrointestinal , Traumatismos de la Médula Espinal , Humanos , Ratones , Animales , Interleucina-13/genética , Filogenia , ARN Ribosómico 16S/genética , Traumatismos de la Médula Espinal/tratamiento farmacológico , Disbiosis/microbiologíaRESUMEN
BACKGROUND: Spinal cord injury (SCI) elicits robust neuroinflammation that eventually exacerbates the initial damage to the spinal cord. L-arginine is critical for the responsiveness of T cells, which are important contributors to neuroinflammation after SCI. Furthermore, L-arginine is the substrate for nitric oxide (NO) production, which is a known inducer of secondary damage. METHODS: To accomplish systemic L-arginine depletion, repetitive injections of recombinant arginase-1 (rArg-I) were performed. Functional recovery and histopathological parameters were analyzed. Splenic immune responses were evaluated by flow cytometry. Pro-inflammatory gene expression and nitrite concentrations were measured. RESULTS: We show for the first time that systemic L-arginine depletion improves locomotor recovery. Flow cytometry and immunohistological analysis showed that intraspinal T-cell infiltration was reduced by 65%, and peripheral numbers of Th1 and Th17 cells were suppressed. Moreover, rArg-I treatment reduced the intraspinal NO production by 40%. Histopathological analyses revealed a 37% and 36% decrease in the number of apoptotic neurons and neuron-macrophage/microglia contacts in the spinal cord, respectively. CONCLUSIONS: Targeting detrimental T-cell responses and NO-production via rArg-I led to a reduced neuronal cell death and an improved functional recovery. These findings indicate that L-arginine depletion holds promise as a therapeutic strategy after SCI.
RESUMEN
A mounting body of evidence suggests that stress plays a major role in the injury progression after spinal cord injury (SCI). Injury activates the stress systems; this in turn may augment the generation of pro-inflammatory cytokines, stimulate pro-inflammatory immune cells, and alter the balance between the pro- and anti-inflammatory immune response. As a result, it is suggested that stress pathways may augment neuronal damage and loss after SCI. Considering these potential detrimental effects of stress after SCI, we hypothesized that inhibition of stress pathways immediately after SCI may offer protection from damage and improve recovery. To investigate the relevance of stress responses in SCI recovery, we investigated the effects of blocking three well-studied stress response axes in a mouse model of SCI. Propranolol, RU-486, and CP-99994 were administered to inhibit the sympathetic axis, the hypothalamus-pituitary-adrenal axis, and the neuropeptide axis, respectively. Surprisingly, assessing functional recovery by the Basso Mouse Scale revealed that RU-486 and CP-99994 did not affect functional outcome, indicating that these pathways are dispensable for neuroprotection or repair after SCI. Moreover, the beta-blocker propranolol worsened functional outcome in the mouse SCI model. In conclusion, immediate inhibition of three major stress axes has no beneficial effects on functional recovery after SCI in mice. These results suggest that injury-induced stress responses do not interfere with the healing process and hence, pharmacological targeting of stress responses is not a recommended treatment option for SCI. These findings are of great importance for other researchers to avoid unnecessary and potentially futile animal experiments.
Asunto(s)
Sistema Hipotálamo-Hipofisario/metabolismo , Neuropéptidos/metabolismo , Recuperación de la Función/fisiología , Traumatismos de la Médula Espinal/metabolismo , Estrés Fisiológico/fisiología , Sistema Nervioso Simpático/metabolismo , Antagonistas Adrenérgicos beta/farmacología , Animales , Citocinas/antagonistas & inhibidores , Citocinas/metabolismo , Femenino , Sistema Hipotálamo-Hipofisario/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Neuropéptidos/antagonistas & inhibidores , Recuperación de la Función/efectos de los fármacos , Traumatismos de la Médula Espinal/fisiopatología , Estrés Fisiológico/efectos de los fármacos , Sistema Nervioso Simpático/efectos de los fármacos , Vértebras Torácicas/lesionesRESUMEN
Spinal cord injury (SCI) triggers the formation of a glial and fibrotic scar, which creates a major barrier for neuroregenerative processes. Previous findings indicate that mast cells (MCs) protect the spinal cord after mechanical damage by suppressing detrimental inflammatory processes via mouse mast cell protease 4 (mMCP4), a MC-specific chymase. In addition to these immunomodulatory properties, mMCP4 also plays an important role in tissue remodeling and extracellular matrix degradation. Therefore, we have investigated the effects of mMCP4 on the scarring response after SCI. We demonstrate that the decrease in locomotor performance in mMCP4-/- mice is correlated with excessive scar formation at the lesion. The expression of axon-growth inhibitory chondroitin sulfate proteoglycans was dramatically increased in the perilesional area in mMCP4-/- mice compared to wild type mice. Moreover, the fibronectin-, laminin-, and collagen IV-positive scar was significantly enlarged in mMCP4-/- mice at the lesion center. A degradation assay revealed that mMCP4 directly cleaves collagen IV in vitro. On the gene expression level, neurocan and GFAP were significantly higher in the mMCP4-/- group at day 2 and day 28 after injury respectively. In contrast, the expression of fibronectin and collagen IV was reduced in mMCP4-/- mice compared to WT mice at day 7 after SCI. In conclusion, our data show that mMCP4 modulates scar development after SCI by altering the gene and protein expression patterns of key scar factors in vivo. Therefore, we suggest a new mechanism via which endogenous mMCP4 can improve recovery after SCI.
Asunto(s)
Cicatriz/metabolismo , Serina Endopeptidasas/metabolismo , Traumatismos de la Médula Espinal/metabolismo , Animales , Proteoglicanos Tipo Condroitín Sulfato/metabolismo , Quimasas/metabolismo , Cicatriz/patología , Fibronectinas/metabolismo , Fibrosis , Proteína Ácida Fibrilar de la Glía/metabolismo , Mastocitos/metabolismo , Ratones , Regeneración Nerviosa/fisiología , Neuroglía/patología , Serina Endopeptidasas/genética , Serina Endopeptidasas/fisiología , Médula Espinal/metabolismoRESUMEN
After spinal cord injury (SCI), monocyte derived macrophages play a detrimental role. Histone deacetylases (HDACs) are central epigenetic regulators of macrophage-polarization. We hypothesized that HDAC3 inhibition suppresses the pro-inflammatory macrophage phenotype (M1), promotes the anti-inflammatory phenotype (M2) and improves functional recovery after SCI. Therefore, two inhibitors of HDAC3 were selected, namely scriptaid and RGFP966. The impact on macrophage polarization was studied by investigating the effect on gene and protein expression of selected M1 and M2 markers. We show that scriptaid differentially influences M1 and M2 markers. It increases CD86 and iNOS gene expression and decreases GPR18, CD38, FPR2 and Arg-1 gene expression as well as the production of IL-6 and NO. RGFP966 primarily increased the expression of the M2 markers Arg-1 and Ym1 and reduced the production of IL-6 (M1). RGFP966 and scriptaid reduced the formation of foamy macrophages. Finally, to investigate the impact of HDAC3 inhibition on functional recovery after SCI, we studied the effects of RGFP966 and scriptaid in an in vivo T-cut hemisection SCI model. Histological analyses were performed on spinal cord sections to determine lesion size and astrogliosis, demyelinated area and selected infiltrating immune cells. RGFP966 and scriptaid did not affect functional recovery or histopathological outcome after SCI. In conclusion, these results indicate that specific HDAC3 inhibition with RGFP966 promotes alternative activation of macrophages and reduces the formation of foamy macrophages, but does not lead to a better functional recovery after SCI.
RESUMEN
Basophils are the smallest population of granulocytes found in the circulation. They have crucial and nonredundant roles in allergic disorders, in protection from parasite infections, in autoimmunity, and in the regulation of type 2 immunity. They share phenotypic and functional properties with mast cells, which exert substantial protective effects after traumatic brain injury and spinal cord injury, although they are considered one of the most proinflammatory cell types in the body. In contrast, the in vivo functions of basophils in central nervous system trauma are still obscure and not well studied. In this study, we show that by comparing spinal cord injury in wild type vs. basophil-deficient Mcpt8Cre transgenic mice, the locomotor recovery is not affected in mice depleted in basophils. In addition, no substantial differences were observed in the lesion size and in the astrocytic and macrophage/microglia reaction between both mouse strains. Hence, despite the multiple properties shared with mast cells, these data show, for the first time, to our knowledge, that basophils are dispensable for the functional recovery process after hemisection injury to the spinal cord in mice.