Jean-Pierre Julien

INNATE IMMUNITY : THE MISSING LINK IN NEUROPROTECTION AND NEURODEGENERATION?

Innate immunity was previously thought to be a nonspecific immunological programme that was engaged by peripheral organs to maintain homeostasis after stress and injury. Emerging evidence indicates that this highly organized response also takes place in the central nervous system. Through the recognition of neuronal fingerprints, the long-term induction of the innate immune response and its transition to an adaptive form might be central to the pathophysiology and aetiology of neurodegenerative disorders. Paradoxically, this response also protects neurons by favouring remyelination and trophic support afforded by glial cells.

Progressive neuronopathy in transgenic mice expressing the human neurofilament heavy gene: A mouse model of amyotrophic lateral sclerosis

We generated four transgenic mice with a 34 kb genomic fragment including the complete human neurofilament heavy (NF-H) gene. This human NF-H fragment contained all regulatory elements for tissue-specific expression, and in two transgenic lines, human NF-H proteins were produced at levels up to 2-fold the levels of endogenous mouse NF-H protein. By 3-4 months of age, these NF-H transgenics progressively develop neurological defects and abnormal neurofilamentous swellings that are highly reminiscent of those found in amyotrophic lateral sclerosis (ALS). We propose that a modest up-regulation of NF-H cross-linkers can result in an impairment of neurofilament transport, causing neuronal swellings with ensuing axonopathy and muscle atrophy, a mechanism of pathogenesis pertinent to the possible etiology of ALS.

Minocycline Slows Disease Progression in a Mouse Model of Amyotrophic Lateral Sclerosis

There is currently no effective pharmacological treatment for amyotrophic lateral sclerosis (ALS). Because recent evidence suggests that secondary inflammation and caspase activation may contribute to neurodegeneration in ALS, we tested the effects of minocycline, a second-generation tetracycline with anti-inflammatory properties, in mice expressing a mutant superoxide dismutase (SOD1(G37R)) linked to human ALS. Administration of minocycline into the diet, beginning at late presymptomatic stage (7 or 9 months of age), delayed the onset of motor neuron degeneration, muscle strength decline, and it increased the longevity of SOD1(G37R) mice by approximately 5 weeks for approximately 70% of tested mice. Moreover, less activation of microglia was detected at early symptomatic stage (46 weeks) and at the end stage of disease in the spinal cord of SOD1(G37R) mice treated with minocycline. These results indicate that minocycline, which is clinically well tolerated, may represent a novel and effective drug for treatment of ALS.

Amyotrophic lateral sclerosis. unfolding the toxicity of the misfolded.

inherited loci map to chromosomes 2q and 15q whereas one dominant juvenile-onset locus maps to chromo-However, the genes responsible for these familial ALS-FTD and juvenile ALS cases have not yet been Amyotrophic lateral sclerosis (ALS) is one of the most identified. common adult-onset neurogenerative diseases, having In a small number of sporadic ALS patients (u202b%1فu202c of a prevalence of u202b5فu202c per 100,000 individuals. This human cases), codon deletions or insertion in the KSP repeat disease, first described by Charcot in 1869, is character-motif of the neurofilament NF-H gene have been identi-ized by the selective degeneration of motor neurons, fied (for review see Cleveland, 1999). No such NF-H the large nerve cells connecting the brain to the spinal mutants have been detected in over 1000 control DNA cord and from the spinal cord to muscles, that control samples. The combined data suggest that NF-H variants muscle movement. The loss of motor neurons leads may represent risk factors for ALS disease. to progressive atrophy of skeletal muscles. ALS is a For the vast majority of ALS cases, the factors trig-relentless disease that manifests as progressive decline gering focal initiation and then spreading of motor neu-in muscular function resulting in eventual paralysis, ron degeneration in sporadic ALS remain to be eluci-speech deficits and, ultimately, death due to respiratory dated. Various hypotheses have been suggested as failure in the majority of ALS patients within 2 to 5 years potential contributors of disease such as oxidative dam-of clinical onset. The weakness, which typically begins age, excitotoxicity, mitochondrial defects, and autoim-focally and propagates, is usually associated with the munity but these could be secondary to the neurodegen-degeneration of both lower motor neurons in the brain-eration process. The virus hypothesis as primary stem and spinal cord, and upper motor neurons in the cause of ALS is now being revisited in light of a recent cerebral cortex. Approximately 10% of ALS patients are report of enterovirus (EV) nucleic acids detected by re-familial cases. The majority of ALS cases are sporadic verse transcriptase-PCR in the spinal cord of a high (90%) with no known genetic component. While current percentage of patients with sporadic ALS (Berger et al., evidence suggests that multiple genetic and environ-2000). More studies are needed to confirm these results mental factors may be implicated in ALS pathogenesis, and to demonstrate that EV sequences can play a causal both sporadic and familial ALS cases share …

Delayed maturation of regenerating myelinated axons in mice lacking neurofilaments.

Using the technique of homologous recombination in embryonic stem cells, we generated mice bearing a targeted disruption of the gene encoding the neurofilament light (NF-L) protein. The absence of NF-L protein in mice resulted in dramatic declines of approximately 20-fold in the levels of neurofilament medium and heavy proteins in the brain and sciatic nerve while increases were detected for other cytoskeletal proteins such as tubulin and GAP-43. Despite a lack of neurofilaments and hypotrophy of axons, the NF-L knockout mice develop normally and do not exhibit overt phenotypes. However, in both NF-L -/- and NF-L +/- mice, the regeneration of myelinated axons following crush injury of peripheral nerves was found to be abnormal. In the second week after axotomy, the number of newly regenerated myelinated axons in the sciatic nerve and facial nerve of NF-L -/- mice corresponded to only approximately 25 and approximately 5% of the number of myelinated axons found in normal mice, respectively. At this early postaxotomy stage, electron microscopy of nerve segments distal to the crush site in NF-L -/- mice revealed abundant clusters of axonal sprouts that were indicative of retarded maturation of regenerating fibers. The analysis of the distal sciatic nerve at 2 months after crush indicated that neurofilament-deficient axons have the capacity to regrow for a long distance and to remyelinate, albeit at a slower rate. These results provide the first direct evidence for a role of neurofilaments in the maturation of regenerating myelinated axons.

Deregulation of Cdk5 in a Mouse Model of ALS: Toxicity Alleviated by Perikaryal Neurofilament Inclusions

Recent studies suggest that increased activity of cyclin-dependent kinase 5 (Cdk5) may contribute to neuronal death and cytoskeletal abnormalities in Alzheimers disease. We report here such deregulation of Cdk5 activity associated with the hyperphosphorylation of tau and neurofilament (NF) proteins in mice expressing a mutant superoxide dismutase (SOD1(G37R)) linked to amyotrophic lateral sclerosis (ALS). A Cdk5 involvement in motor neuron degeneration is supported by our analysis of three SOD1(G37R) mouse lines exhibiting perikaryal inclusions of NF proteins. Our results suggest that perikaryal accumulations of NF proteins in motor neurons may alleviate ALS pathogenesis by acting as a phosphorylation sink for Cdk5 activity, thereby reducing the detrimental hyperphosphorylation of tau and other neuronal substrates.

Neurofilaments in Health and Disease

This article reviews current knowledge of neurofilament structure, phosphorylation, and function and neurofilament involvement in disease. Neurofilaments are obligate heteropolymers requiring the NF-L subunit together with either the NF-M or the NF-H subunit for polymer formation. Neurofilaments are very dynamic structures; they contain phosphorylation sites for a large number of protein kinases, including protein kinase A (PKA), protein kinase C (PKC), cyclin-dependent kinase 5 (Cdk5), extracellular signal regulated kinase (ERK), glycogen synthase kinase-3 (GSK-3), and stress-activated protein kinase gamma (SAPK gamma). Most of the neurofilament phosphorylation sites, located in tail regions of NF-M and NF-H, consist of the repeat sequence motif, Lys-Ser-Pro (KSP). In addition to the well-established role of neurofilaments in the control of axon caliber, there is growing evidence based on transgenic mouse studies that neurofilaments can affect the dynamics and perhaps the function of other cytoskeletal elements, such as microtubules and actin filaments. Perturbations in phosphorylation or in metabolism of neurofilaments are frequently observed in neurodegenerative diseases. A down-regulation of mRNA encoding neurofilament proteins and the presence of neurofilament deposits are common features of human neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS), Parkinsons disease, and Alzheimers disease. Although the extent to which neurofilament abnormalities contribute to pathogenesis in these human diseases remains unknown, emerging evidence, based primarily on transgenic mouse studies and on the discovery of deletion mutations in the NF-H gene of some ALS eases, suggests that disorganized neurofilaments can provoke selective degeneration and death of neurons. An interference of axonal transport by disorganized neurofilaments has been proposed as one possible mechanism of neurofilament-induced pathology. Other factors that can potentially lead to the accumulation of neurofilaments will be discussed as well as the emerging evidence for neurofilaments as being possible targets of oxidative damage by mutations in the superoxide dismutase enzyme (SOD1); such mutations are responsible for approximately 20% of familial ALS cases.

Neurofilament functions in health and disease

Transgenic approaches have recently been used to investigate the functions of neuronal intermediate filaments. Gene knockout studies have demonstrated that neurofilaments are not required for axogenesis and that individual neurofilament proteins play distinct roles in filament assembly and in the radial growth of axons. The involvement of neurofilaments in disease is supported by the discovery of novel mutations in the neurofilament heavy gene from cases of amyotrophic lateral sclerosis and by reports of neuronal death in mouse models expressing neurofilament and alpha-internexin transgenes. However, mouse studies have shown that axonal neurofilaments are not required for pathogenesis caused by mutations in superoxide dismutase and that increasing perikaryal levels of neurofilament proteins may even confer protection in this disease.

Control of dendritic cell cross-presentation by the major histocompatibility complex class I cytoplasmic domain

Dendritic cells (DCs) can present extracellularly derived antigens in the context of major histocompatibility complex (MHC) class I molecules, a process called cross-presentation. Although recognized to be important for priming of T cell responses to many viral, bacterial and tumor antigens, the mechanistic details of this alternative antigen-presentation pathway are poorly understood. We demonstrate here the existence of an endolysosomal compartment in DCs where exogenously derived peptides can be acquired for presentation to T cells, and show that the MHC class I cytoplasmic domain contains a tyrosine-based targeting signal required for routing MHC class I molecules through these compartments. We also report that transgenic mice expressing H-2Kb with a tyrosine mutation mount inferior H-2Kb-restricted cytotoxic T lymphocyte responses against two immunodominant viral epitopes, providing evidence of a crucial function for cross-priming in antiviral immunity.

Integrators of the Cytoskeleton that Stabilize Microtubules

Sensory neurodegeneration occurs in mice defective in BPAG1, a gene encoding cytoskeletal linker proteins capable of anchoring neuronal intermediate filaments to actin cytoskeleton. While BPAG1 null mice fail to anchor neurofilaments (NFs), BPAG1/NF null mice still degenerate in the absence of NFs. We report a novel neural splice form that lacks the actin-binding domain and instead binds and stabilizes microtubules. This interaction is functionally important; in mice and in vitro, neurons lacking BPAG1 display short, disorganized, and unstable microtubules defective in axonal transport. Ironically, BPAG1 neural isoforms represent microtubule-associated proteins that when absent lead to devastating consequences. Moreover, BPAG1 can functionally account for the extraordinary stability of axonal microtubules necessary for transport over long distances. Its isoforms interconnect all three cytoskeletal networks, a feature apparently central to neuronal survival.