Brigitte Pettmann

Neuronal Cell Death

We thank J.-F. Brunet, P. G. H. Clarke, O. deLapeyriere, P. Golstein, J.-C. Martinou, C. Mulle, and R. W. Oppenheim for thoughtful comments on the manuscript and C. Waldmann for artwork. In a short review such as this, it is not possible to establish precedence for all ideas or mechanisms, many of which originate from (or are still confined to) studies outside the nervous system. Wherever possible, we have therefore illustrated each topic by citing recent articles involving neurons. We refer the interested reader to the reference lists of those articles, or of the many excellent reviews we have cited, for more ample details. Studies from our laboratory were supported by INSERM, CNRS, Association Francaise contre les Myopathies (AFM), Institut pour la Recherche sur la Moelle Epiniere (IRME), and European Commission BIO4 contract CT960433.

Motoneuron Death Triggered by a Specific Pathway Downstream of Fas: Potentiation by ALS-Linked SOD1 Mutations

Death pathways restricted to specific neuronal classes could potentially allow for precise control of developmental neuronal death and also underlie the selectivity of neuronal loss in neurodegenerative disease. We show that Fas-triggered death of normal embryonic motoneurons requires transcriptional upregulation of neuronal NOS and involves Daxx, ASK1, and p38 together with the classical FADD/caspase-8 cascade. No evidence for involvement of this pathway was found in cells other than motoneurons. Motoneurons from transgenic mice overexpressing ALS-linked SOD1 mutants (G37R, G85R, or G93A) displayed increased susceptibility to activation of this pathway: they were more sensitive to Fas- or NO-triggered cell death but not to trophic deprivation or excitotoxic stimulation. Thus, triggering of a motoneuron-restricted cell death pathway by neighboring cells might contribute to motoneuron loss in ALS.

The brain fibroblast growth factor (FGF) is localized in neurons

The physiological role of fibroblast growth factor (FGF), like many other characterized growth factors, is poorly understood. Therefore, it would be useful to know in which cell type this growth factor is present. Using monoclonal and polyclonal antibodies against FGF, we have localized this molecule in rat brain. Interestingly, FGF is found exclusively in neuronal cells. Since FGF stimulates the proliferation and maturation of glial cells in vitro, its involvement in the interaction between neuronal and glial cells in vivo is likely. A role in the triggering of reactive gliosis can also be hypothesized.

Cardiotrophin-1 requires LIFRβ to promote survival of mouse motoneurons purified by a novel technique

The cytokines ciliary neurotrophic factor (CNTF) and leukemia inhibitory factor (LIF) signal through a receptor complex formed between two transmembrane proteins, gp130 and LIFRβ. In addition, CNTF also uses a ligand‐binding component which is anchored to the cell membrane. In the case of cardiotrophin‐1 (CT‐1), LIFRβ is also required in cardiomyocytes, but this has not been proven in neurons, and published data suggest that motoneurons may use a different receptor complex. We used Lifrβ knockout mice to assess the requirement for this receptor component in the signal transduction of CT‐1 in motoneurons. To study purified motoneurons from such mutants, we have developed a method allowing for isolation of highly purified mouse motoneurons. This protocol is based on the immunoaffinity purification of motoneurons using antibodies against the extracellular domain of the neurotrophin receptor, p75, followed by cell sorting using magnetic microbeads. We show that CNTF, LIF, and CT‐1 are unable to promote the survival of motoneurons derived from homozygous Lifrβ−/− mutant embryos. Thus, LIFRβ is absolutely required to transduce the CT‐1 survival signal in motoneurons. J Neurosci Res 55:119–126, 1999. 

Active killing of neurons during development and following stress: a role for p75(NTR) and Fas?

Evidence for active triggering of neuronal death continues to accumulate. The transmembrane receptors p75(NTR) and Fas can trigger (and in some cases are required for) programmed cell death of the neurons that express them, through signalling pathways that are regulated by a variety of cytoplasmic effectors. Neuronal death induced by trophic deprivation often requires Fas signalling, further blurring the boundaries between naturally occurring and stress-induced neuronal death.

Foxo3a induces motoneuron death through the Fas pathway in cooperation with JNK

BackgroundProgrammed cell death of motoneurons in the developing spinal cord is thought to be regulated through the availability of target-derived neurotrophic factors. When deprived of trophic support, embryonic spinal motoneurons in vitro over-express FasL, a ligand activating a Fas-mediated death pathway. How trophic factors regulate the expression of FasL is presently unclear, but two regulators of FasL, FOXO3a (FKHRL1) and JNK have been described to play a role in other cell types. Thus, their potential function in motoneurons was investigated in this study.ResultsWe show here that as a result of removal of neurotrophic factors and the consequent reduction in signalling through the PI3K/Akt pathway, Foxo3a translocates from the cytoplasm to the nucleus where it triggers cell death. Death is reduced in Fas and FasL mutant motoneurons and in the presence of JNK inhibitors indicating that a significant part of it requires activation of the Fas/FasL pathway through JNK.ConclusionsTherefore, in motoneurons as in other cell types, FOXO transcriptional regulators provide an important link between other signalling pathways and the cell death machinery.

Ultrastructural localization of fibroblast growth factor in neurons of rat brain

The distribution of fibroblast growth factor (FGF) at the ultrastructural level in the brain of young (15- and 20-day-old) and adult (3-month-old) rats was investigated by immunocytochemistry. Strong staining was observed in most neurons of the cortex of young rat brain. In the same brain area of adult rat many neurons were also stained intensely, while others were negative. Neurons in the other parts of the brain and especially in the adult rat, were generally more weakly stained. The reaction product was located in the cytoplasm of the neuronal cell bodies and their processes. Astrocytes, oligodendrocytes, microglial cells, meningeal cells, choroïd epithelial cells, ependymal cells and capillary endothelial cells showed no staining.

Signaling by death receptors in the nervous system.

Cell death plays an important role both in shaping the developing nervous system and in neurological disease and traumatic injury. In spite of their name, death receptors can trigger either cell death or survival and growth. Recent studies implicate five death receptors--Fas/CD95, TNFR1 (tumor necrosis factor receptor-1), p75NTR (p75 neurotrophin receptor), DR4, and DR5 (death receptors-4 and -5)--in different aspects of neural development or degeneration. Their roles may be neuroprotective in models of Parkinsons disease, or pro-apoptotic in ALS and stroke. Such different outcomes probably reflect the diversity of transcriptional and posttranslational signaling pathways downstream of death receptors in neurons and glia.

Adenoviral cardiotrophin-1 gene transfer protects pmn mice from progressive motor neuronopathy

Cardiotrophin-1 (CT-1), an IL-6-related cytokine, causes hypertrophy of cardiac myocytes and has pleiotropic effects on various other cell types, including motoneurons. Here, we analyzed systemic CT-1 effects in progressive motor neuronopathy (pmn) mice that suffer from progressive motoneuronal degeneration, muscle paralysis, and premature death. Administration of an adenoviral CT-1 vector to newborn pmn mice leads to sustained CT-1 expression in the injected muscles and bloodstream, prolonged survival of animals, and improved motor functions. CT-1-treated pmn mice showed a significantly reduced degeneration of facial motoneuron cytons and phrenic nerve myelinated axons. The terminal innervation of skeletal muscle, grossly disturbed in untreated pmn mice, was almost completely preserved in CT-1-treated pmn mice. The remarkable neuroprotection conferred by CT-1 might become clinically relevant if CT-1 side effects, including cardiotoxicity, could be circumvented by a more targeted delivery of this cytokine to the nervous system.

Adenovirus-mediated transfer of the neurotrophin-3 gene into skeletal muscle of pmn mice: Therapeutic effects and mechanisms of action

Several neurotrophic factors (CNTF, BDNF, IGF-1) have been suggested for the treatment of motor neuron diseases. In ALS patients, however, the repeated subcutaneous injection of these factors as recombinant proteins is complicated by their toxicity or poor bioavailability. We have constructed an adenovirus vector coding for neurotrophin-3 (AdNT-3) allowing for stable and/or targeted delivery of NT-3 to motoneurons. The intramuscular administration of this vector was tested in the mouse mutant pmn (progressive motor neuronopathy). AdNT-3-treated pmn mice showed prolonged lifespan, improved neuromuscular function, reduced motor axonal degeneration and efficient reinnervation of muscle fibres. NT-3 protein and also adenovirus vectors, when injected into muscle, can be transported by motoneurons via retrograde axonal transport to their cell bodies in the spinal cord. Using ELISA and RT-PCR analyses in muscle, spinal cord and serum of AdNT-3-treated pmn mice, we have investigated the contribution of these processes to the observed therapeutic effects. Our results suggest that most if not all therapeutic benefit was due to the continuous systemic liberation of adenoviral NT-3. Therefore, viral gene therapy vectors auch as adenoviruses, AAVs, lentiviruses and new types of gene transfer not based on viral vectors that allow for efficient in vivo liberation of neurotrophic factors have potential for the future treatment of human motor neuron diseases.