No abstract

Amyotrophic lateral sclerosis (ALS), characterized by degeneration of spinal motor neurons, consists of sporadic and familial forms. One cause of familial ALS is missense mutations in the superoxide dismutase 1 (SOD1) gene. Iron accumulation occurs in the CNS of both forms of ALS; however, its contribution to the pathogenesis of ALS is not known. We examined the role of iron in a transgenic mouse line overexpressing the human SOD1 G37R mutant. We show that multiple mechanisms may underlie the iron accumulation in neurons and glia in SOD1 G37R transgenic mice. These include dysregulation of proteins involved in iron influx and sensing of intracellular iron; iron accumulation in ventral motor neurons secondary to blockage of anterograde axonal transport; and increased mitochondrial iron load in neurons and glia. We also show that treatment of SOD1 G37R mice with an iron chelator extends life span by 5 weeks, accompanied by increased survival of spinal motor neurons and improved locomotor function. These data suggest that iron chelator therapy might be useful for the treatment of ALS.

Motoneurons are key points of convergence within motor networks, acting as the "output channels" that directly control sets of muscles to maintain posture and generate movement. Here we use genetic mosaic techniques to reveal the origins and architecture of the leg motoneurons of Drosophila. We show that a small number of leg motoneurons are born in the embryo but most are generated during larval life. These postembryonic leg motoneurons are produced by five neuroblasts per hemineuromere, and each lineage generates stereotyped lineage-specific projection patterns. Two of these postembryonic neuroblasts generate solely motoneurons that are the bulk of the leg motoneurons. Within the largest lineage, lineage 15, we see distinct birth-order differences in projection patterns. A comparison of the central projections of leg motoneurons and the muscles they innervate reveals a stereotyped architecture and the existence of a myotopic map. Timeline analysis of axonal outgrowth reveals that leg motoneurons reach their sites of terminal arborization in the leg at the time when their dendrites are elaborating their subtype-specific shapes. Our findings provide a comprehensive description of the origin, development, and architecture of leg motoneurons that will aid future studies exploring the link between the assembly and organization of connectivity within the leg motor system of Drosophila.

Lipocalin 2 (Lcn2) plays an important role in defense against bacterial infection by interfering with bacterial iron acquisition. Although Lcn2 is expressed in a number of aseptic inflammatory conditions, its role in these conditions remains unclear. We examined the expression and role of Lcn2 after spinal cord injury (SCI) in adult mice by using a contusion injury model. Lcn2 expression at the protein level is rapidly increased 12-fold at 1 d after SCI and decreases gradually thereafter, being three times as high as control levels at 21 d after injury. Lcn2 expression is strongly induced after contusion injury in astrocytes, neurons, and neutrophils. The Lcn2 receptor (Lcn2R), which has been shown to influence cell survival, is also expressed after SCI in the same cell types. Lcn2-deficient (Lcn2 ؊/؊ ) mice showed significantly better locomotor recovery after spinal cord contusion injury than wild-type (Lcn2 ؉/؉ ) mice. Histological assessments indicate improved neuronal and tissue survival and greater sparing of myelin in Lcn2 ؊/؊ mice after contusion injury. Flow cytometry showed a decrease in neutrophil influx and a small increase in the monocyte population in Lcn2 ؊/؊ injured spinal cords. This change was accompanied by a reduction in the expression of several pro-inflammatory chemokines and cytokines as well as inducible nitric oxide synthase early after SCI in Lcn2 ؊/؊ mice compared with wild-type animals. Our results, therefore, suggest a role for Lcn2 in regulating inflammation in the injured spinal cord and that lack of Lcn2 reduces secondary damage and improves locomotor recovery after spinal cord contusion injury.

CNS injury-induced hemorrhage and tissue damage leads to excess iron, which can cause secondary degeneration. The mechanisms that handle this excess iron are not fully understood. We report that spinal cord contusion injury (SCI) in mice induces an "iron homeostatic response" that partially limits iron-catalyzed oxidative damage. We show that ceruloplasmin (Cp), a ferroxidase that oxidizes toxic ferrous iron, is important for this process. SCI in Cp-deficient mice demonstrates that Cp detoxifies and mobilizes iron and reduces secondary tissue degeneration and functional loss. Our results provide new insights into how astrocytes and macrophages handle iron after SCI. Importantly, we show that iron chelator treatment has a delayed effect in improving locomotor recovery between 3 and 6 weeks after SCI. These data reveal important aspects of the molecular control of CNS iron homeostasis after SCI and suggest that iron chelator therapy may improve functional recovery after CNS trauma and hemorrhagic stroke.

Abnormal iron homeostasis is increasingly thought to contribute to the pathogenesis of several neurodegenerative disorders. We have previously reported impaired iron homeostasis in a mouse model of spinal cord injury and in a mouse model of amyotrophic lateral sclerosis. Both these disorders are associated with CNS inflammation. However, what effect inflammation, and in particular, inflammatory cytokines have on iron homeostasis in CNS glia remains largely unknown. Here we report that the proinflammatory cytokine TNF-α, and the anti-inflammatory cytokine TGF-β1 affect iron homeostasis in astrocytes and microglia in distinct ways. Treatment of astrocytes in vitro with TNF-α induced the expression of the iron importer "divalent iron transporter 1" (DMT1) and suppressed the expression of the iron exporter ferroportin (FPN). However, TGF-β1 had no effect on DMT1 expression but increased the expression of FPN in astrocytes. In microglia, on the other hand, both cytokines caused induction of DMT1 and suppression of FPN expression. Iron influx and efflux assays in vitro confirmed that iron homeostasis in astrocytes and microglia is differentially regulated by these cytokines. In particular, TNF-α caused an increase in iron uptake and retention by both astrocytes and microglia, while TGF-β1 promoted iron efflux from astrocytes but caused iron retention in microglia. These data suggest that these two cytokines, which are expressed in CNS inflammation in injury and disease, can have profound and divergent effects on iron homeostasis in astrocytes and microglia.

Spinal cord injury (SCI) triggers inflammatory responses that involve neutrophils, macrophages/microglia and astrocytes and molecules

Regeneration of injured central nervous system axons is highly restricted, causing neurological impairment. To date, although the lack of intrinsic regenerative potential is well described, a key regulatory molecular mechanism for the enhancement of both axonal regrowth and functional recovery after central nervous system injury remains elusive. While ubiquitin ligases coordinate neuronal morphogenesis and connectivity during development as well as after axonal injury, their role specifically in axonal regeneration is unknown. Following a bioinformatics network analysis combining ubiquitin ligases with previously defined axonal regenerative proteins, we found a triad composed of the ubiquitin ligases MDM4, MDM2 and the transcription factor p53 (encoded by TP53) as a putative central signalling complex restricting the regeneration program. Indeed, conditional deletion of MDM4 or pharmacological inhibition of MDM2/p53 interaction in the eye and spinal cord promote axonal regeneration and sprouting of the optic nerve after crush and of supraspinal tracts after spinal cord injury. The double conditional deletion of MDM4-p53 as well as MDM2 inhibition in p53-deficient mice blocks this regenerative phenotype, showing its dependence upon p53. Genome-wide gene expression analysis from ex vivo fluorescence-activated cell sorting in MDM4-deficient retinal ganglion cells identifies the downstream target IGF1R, whose activity and expression was found to be required for the regeneration elicited by MDM4 deletion. Importantly, we demonstrate that pharmacological enhancement of the MDM2/p53-IGF1R axis enhances axonal sprouting as well as functional recovery after spinal cord injury. Thus, our results show MDM4-MDM2/p53-IGF1R as an original regulatory mechanism for CNS regeneration and offer novel targets to enhance neurological recovery.media-1vid110.1093/brain/awv125_video_abstractawv125_video_abstract.