RESUMEN
Lysophosphatidic acid (LPA) is a unique bioactive lysophospholipid that induces pleiotropic effects in various cell types and organisms by acting on its specific receptors. LPA is mainly synthetised extracellularly by the ectonucleotide pyrophosphatase/phosphodiesterase 2/autotaxin (enpp2). Altered LPA signalling is associated with embryonic abnormalities, suggesting critical roles for LPA during development. However, the role of LPA signalling during early embryogenesis is not well established. We demonstrate that enpp2/LPA signalling in the early zebrafish embryo results in altered axis and midline formation, defects in left right (L-R) patterning, ciliogenesis of the Kupffer's vesicle (KV), through the modulation of cell migration during gastrulation in a lpar1-3 Rho/ROCK-dependant manner. Overall, this study demonstrates an essential role of enpp2/LPA signalling during early embryogenesis.
Asunto(s)
Tipificación del Cuerpo , Embrión no Mamífero/enzimología , Hidrolasas Diéster Fosfóricas/metabolismo , Proteínas de Pez Cebra/metabolismo , Pez Cebra/embriología , Pez Cebra/metabolismo , Animales , Movimiento Celular , Cilios/metabolismo , Desarrollo Embrionario/genética , Regulación del Desarrollo de la Expresión Génica , Organogénesis , Fenotipo , Hidrolasas Diéster Fosfóricas/genética , Pez Cebra/genética , Proteínas de Pez Cebra/genética , Proteínas de Unión al GTP rho/metabolismo , Quinasas Asociadas a rho/metabolismoRESUMEN
Spinal cord injury (SCI) frequently leads to a permanent functional impairment as a result of the initial injury followed by secondary injury mechanism, which is characterised by increased inflammation, glial scarring and neuronal cell death. Finding drugs that may reduce inflammatory cell invasion and activation to reduce glial scarring and increase neuronal survival is of major importance for improving the outcome after SCI. In the present study, we examined the effect of rapamycin, an mTORC1 inhibitor and an inducer of autophagy, on recovery from spinal cord injury. Autophagy, a process that facilitates the degradation of cytoplasmic proteins, is also important for maintenance of neuronal homeostasis and plays a major role in neurodegeneration after neurotrauma. We examined rapamycin effects on the inflammatory response, glial scar formation, neuronal survival and regeneration in vivo using spinal cord hemisection model in mice, and in vitro using primary cortical neurons and human astrocytes. We show that a single injection of rapamycin, inhibited p62/SQSTM1, a marker of autophagy, inhibited mTORC1 downstream effector p70S6K, reduced macrophage/neutrophil infiltration into the lesion site, microglia activation and secretion of TNFα. Rapamycin inhibited astrocyte proliferation and reduced the number of GFAP expressing cells at the lesion site. Finally, it increased neuronal survival and axonogenesis towards the lesion site. Our study shows that rapamycin treatment increased significantly p-Akt levels at the lesion site following SCI. Similarly, rapamycin treatment of neurons and astrocytes induced p-Akt elevation under stress conditions. Together, these findings indicate that rapamycin is a promising candidate for treatment of acute SCI condition and may be a useful therapeutic agent.
Asunto(s)
Inmunosupresores/uso terapéutico , Inflamación/tratamiento farmacológico , Inflamación/etiología , Sirolimus/uso terapéutico , Traumatismos de la Médula Espinal/complicaciones , Traumatismos de la Médula Espinal/patología , Animales , Astrocitos/efectos de los fármacos , Astrocitos/fisiología , Antígeno CD11b/metabolismo , Recuento de Células , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Células Cultivadas , Modelos Animales de Enfermedad , Proteína 3 Similar a ELAV/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Proteína Ácida Fibrilar de la Glía/metabolismo , Humanos , Antígeno Ki-67/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Neuronas/efectos de los fármacos , Ratas , Factores de TiempoRESUMEN
OBJECTIVES: A major impediment for recovery after mammalian spinal cord injury (SCI) is the glial scar formed by proliferating reactive astrocytes. Finding factors that may reduce glial scarring, increase neuronal survival, and promote neurite outgrowth are of major importance for improving the outcome after SCI. Exogenous fibroblast growth factor (Fgf) has been shown to decrease injury volume and improve functional outcome; however, the mechanisms by which this is mediated are still largely unknown. METHODS: In this study, Fgf2 was administered for 2 weeks in mice subcutaneously, starting 30 min after spinal cord hemisection. RESULTS: Fgf2 treatment decreased the expression of TNF-a at the lesion site, decreased monocyte/macrophage infiltration, and decreased gliosis. Fgf2 induced astrocytes to adopt a polarized morphology and increased expression of radial markers such as Pax6 and nestin. In addition, the levels of chondroitin sulfate proteoglycans (CSPGs), expressed by glia, were markedly decreased. Furthermore, Fgf2 treatment promotes the formation of parallel glial processes, "bridges," at the lesion site that enable regenerating axons through the injury site. Additionally, Fgf2 treatment increased Sox2-expressing cells in the gray matter and neurogenesis around and at the lesion site. Importantly, these effects were correlated with enhanced functional recovery of the left paretic hind limb. CONCLUSIONS: Thus, early pharmacological intervention with Fgf2 following SCI is neuroprotective and creates a proregenerative environment by the modulation of the glia response.
Asunto(s)
Astrocitos/efectos de los fármacos , Factor 2 de Crecimiento de Fibroblastos/farmacología , Gliosis/tratamiento farmacológico , Células-Madre Neurales/efectos de los fármacos , Recuperación de la Función/efectos de los fármacos , Traumatismos de la Médula Espinal/tratamiento farmacológico , Regeneración de la Medula Espinal/efectos de los fármacos , Animales , Masculino , Ratones , Ratones Endogámicos C57BLRESUMEN
We previously reported that lysophosphatidic acid (LPA) inhibits the neuronal differentiation of human embryonic stem cells (hESC). We extended these studies by analyzing LPA's effects on the expansion of neural stem/progenitor cells (NS/PC) derived from hESCs and human induced pluripotent stem cells (iPSC), and we assessed whether data obtained on the neural differentiation of hESCs were relevant to iPSCs. We showed that hESCs and iPSCs exhibited comparable mRNA expression profiles of LPA receptors and producing enzymes upon neural differentiation. We demonstrated that LPA inhibited the expansion of NS/PCs of both origins, mainly by increased apoptosis in a Rho/Rho-associated kinase (ROCK)-dependent mechanism. Furthermore, LPA inhibited the neuronal differentiation of iPSCs. Lastly, LPA induced neurite retraction of NS/PC-derived early neurons through Rho/ROCK, which was accompanied by myosin light chain (MLC) phosphorylation. Our data demonstrate the consistency of LPA effects across various sources of human NS/PCs, rendering hESCs and iPSCs valuable models for studying lysophospholipid signaling in human neural cells. Our data also highlight the importance of the Rho/ROCK pathway in human NS/PCs. As LPA levels are increased in the central nervous system (CNS) following injury, LPA-mediated effects on NS/PCs and early neurons could contribute to the poor neurogenesis observed in the CNS following injury.
Asunto(s)
Lisofosfolípidos/farmacología , Neuronas/citología , Proteínas de Unión al GTP rho/metabolismo , Quinasas Asociadas a rho/metabolismo , Diferenciación Celular , Humanos , Células Madre Pluripotentes Inducidas/citología , Células Madre Pluripotentes Inducidas/efectos de los fármacos , Células Madre Pluripotentes Inducidas/metabolismo , Cadenas Ligeras de Miosina/metabolismo , Células-Madre Neurales/citología , Células-Madre Neurales/efectos de los fármacos , Células-Madre Neurales/metabolismo , Neuronas/metabolismo , Fosforilación , Receptores del Ácido Lisofosfatídico/metabolismo , Transducción de Señal/efectos de los fármacos , Proteínas de Unión al GTP rho/genética , Quinasas Asociadas a rho/genéticaRESUMEN
Evidence suggests a proinflammatory role of lysophosphatidic acid (LPA) in various pathologic abnormalities, including in the central nervous system. Herein, we describe LPA as an important mediator of inflammation after spinal cord injury (SCI) in zebrafish and mice. Furthermore, we describe a novel monoclonal blocking antibody raised against LPA that potently inhibits LPA's effect in vitro and in vivo. This antibody, B3, specifically binds LPA, prevents it from interacting with its complement of receptors, and blocks LPA's effects on the neuronal differentiation of human neural stem/progenitor cells, demonstrating its specificity toward LPA signaling. When administered systemically to mice subjected to SCI, B3 substantially reduced glial inflammation and neuronal death. B3-treated animals demonstrated significantly more neuronal survival upstream of the lesion site, with some functional improvement. This study describes the use of anti-LPA monoclonal antibody as a novel therapeutic approach for the treatment of SCI.
Asunto(s)
Lisofosfolípidos/antagonistas & inhibidores , Recuperación de la Función , Transducción de Señal , Traumatismos de la Médula Espinal/patología , Animales , Anticuerpos Monoclonales/farmacología , Apoptosis/efectos de los fármacos , Células CHO , Muerte Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Cricetinae , Modelos Animales de Enfermedad , Ensayo de Inmunoadsorción Enzimática , Humanos , Inflamación/complicaciones , Inflamación/patología , Lisofosfolípidos/metabolismo , Lisofosfolípidos/farmacología , Ratones , Microglía/efectos de los fármacos , Microglía/patología , Actividad Motora/efectos de los fármacos , Neuritas/efectos de los fármacos , Neuritas/metabolismo , Fármacos Neuroprotectores/farmacología , Receptores del Ácido Lisofosfatídico/metabolismo , Recuperación de la Función/efectos de los fármacos , Transducción de Señal/efectos de los fármacos , Traumatismos de la Médula Espinal/complicaciones , Traumatismos de la Médula Espinal/fisiopatología , Pez CebraRESUMEN
Lysophosphatidic acid (LPA) is a bioactive lipid that regulates a broad range of cellular effects in various cell types, leading to a variety of responses in tissues, including in the nervous system. LPA and its receptors are found in the nervous system, with different cellular and temporal profiles. Through its ability to target most cells of the nervous system and its induction of pleiotropic effects, LPA mediates events during neural development and adulthood. In this review, we summarize the current knowledge on the effects of LPA in the nervous system, during development and adulthood, and in various pathologies of the nervous system. We also explore potential LPA intervention strategies for anti-LPA therapeutics.