| 1. |
- Althini, Susanna, et al.
(författare)
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Blocked MAP kinase activity selectively enhances neurotrophic growth responses
- 2004
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Ingår i: Molecular and Cellular Neuroscience. - : Elsevier BV. - 1044-7431 .- 1095-9327. ; 25:2, s. 345-354
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Tidskriftsartikel (refereegranskat)abstract
- Bone morphogenetic proteins (BMPs) 4 and 6 as well as MEK inhibitors PD98059 and U0126 potentiate neurotrophin 3 (NT3)- and neurturin (NTN)-induced neurite outgrowth and survival of peripheral neurons from the E9 chicken embryo. Preexposure to BMP4 or PD98059 was sufficient to prime the potentiation of subsequently added NT3. Phosphorylation of Erk2, induced by NT3, was reduced by MEK inhibition but unaffected by BMP signaling. Real-time PCR showed that neither BMP stimulation nor MEK inhibition increased Trk receptor expression and that the BMP-induced genes Smad6 and Id1 were not upregulated by PD98059. In contrast, both MEK inhibition and BMP signaling suppressed transcription of the serum-response element (SRE)-driven Egr1 gene. A reporter assay using NGF-stimulated PC12 cells demonstrated that MEK/Erk/Elk-driven transcriptional activity was inhibited by Smad1/5 and by PD98059. Thus, suppression of SRE-controlled transcription represents a likely convergence point for pathways regulating neurotrophic responses.
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| 2. |
- Althini, Susanna, et al.
(författare)
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Bone morphogenetic protein signalling in NGF-stimulated PC12 cells
- 2003
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Ingår i: Biochemical and Biophysical Research Communications - BBRC. - 0006-291X .- 1090-2104. ; 307:3, s. 632-639
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Tidskriftsartikel (refereegranskat)abstract
- Bone morphogenetic proteins (BMPs) are shown to potentiate NGF-induced neuronal differentiation in PC12 phaeochromocytoma cells grown on collagen under low-serum conditions. Whereas, cell bodies remained rounded in control medium or with only BMPs present, addition of BMP4 or BMP6 robustly increased the neuritogenic effect of NGF within 2 days. NGF-increased phosphorylation of p44(Erk1) and p42(Erk2) between 2 and 24h was unaffected by addition of BMP6. PC12 cells transfected with the SBE(4x)-luc reporter showed that BMP4 significantly increased receptor-activated Smad activity. Expression of constitutively active BMP receptor ALK2 activating Smad1 and Smad5 resulted in a strong increase in the SBE(4x)-luc reporter response. Adding the inhibitory Smad7 drastically reduced this signal. In contrast to wild-type (wt) Smad5, a Smad5 variant lacking five Erk phosphorylation sites in the linker region (designated Smad5/5SA) showed a strong background transcriptional activity. A fusion construct (Gal4-Smad5/5SA) was also highly transcriptionally active. Addition of the MEK inhibitor U0126 to PC12 cells expressing Gal4-Smad5/wt did not increase background transcriptional activity. However, upon activation by constitutively active ALK2 both Gal4-Smad5/wt and Gal4-Smad5/5SA strongly stimulated transcription. The data show that serine residues of the linker region of Smad5 reduce spontaneous transcriptional activity and that NGF-activated Erk does not antagonise BMP signalling at this site. Hence, NGF and BMP signals are likely to interact further downstream at the transcriptional level in neuronal differentiation of the PC12 cells.
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| 3. |
- Althini, Susanna
(författare)
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Experimental Studies of BMP Signalling in Neuronal Cells
- 2003
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The developing nervous system depends largely on extracellular cues to shape its complex network of neurons. Classically, neurotrophins are known to be important mediators in this process. More recently, Bone Morphogenetic Proteins (BMPs), belonging to the Transforming Growth Factor beta (TGFβ) superfamily of secreted cytokines, have been shown to exert a wide range of effects, such as cellular growth, differentiation, survival and apoptosis, both in the developing and adult nervous system. They signal via serine/threonine kinase receptor essentially to the Smad pathway, which carries the signal to the nucleus where the transcription of target genes is regulated.This thesis investigates the functions of BMPs in the nervous system, using a set of different models. Firstly, a targeted deletion of GDF10 (BMP3b) in the mouse was established to evaluate the role of this growth/differentiation factor in the hippocampal formation, a brain area known to be involved in memory processing. Other members of the TGFβ superfamily likely compensate for the lack of GDF10, since no detectable alterations in hippocampal function or gene transcription profile have been found. Secondly, a mouse model was set up, with the aim to study impaired BMP-signalling in dopaminergic neurons. The tyrosine hydroxylase (TH) locus was used to drive the expression of dominant negative BMP receptors by means of bicistronic mRNAs. TH is the rate-limiting enzyme in the biosynthesis of catecholamine and the mice described, show a graded decrease of TH-activity resulting in severe to mild dopamine deficiency. The contribution of the dominant negative BMP receptors to the phenotype is however secondary to the apparent TH hypomorphism. The final theme of this thesis is the potentiating effects of BMPs on neurotrophin-induced neurite outgrowth as studied in explanted ganglia from chick embryos and in the rat phaeochromocytoma cell line PC12. A number of pharmacological inhibitors of intracellular signalling kinases were applied to the cultures in order to reveal the contribution of different pathways to the enhanced neurite outgrowth. We made the unexpected finding that inhibition of MEK signalling mimicked the potentiating effects of BMP stimulation in the chick system. The underlying mechanisms for the synergistic effects, however, are still an enigma.
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| 4. |
- Althini, Sanna, et al.
(författare)
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Normal Nigrostriatal Innervation but Dopamine Dysfunction in Mice Carrying Hypomorphic Tyrosine Hydroxylase Alleles
- 2003
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Ingår i: Journal of Neuroscience Research. - : Wiley. - 0360-4012 .- 1097-4547. ; 72:4, s. 444-453
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Tidskriftsartikel (refereegranskat)abstract
- We investigated the use of the mouse tyrosine hydroxylase (TH) gene to drive knock-in constructs in catecholaminergic neurons. Two targeting constructs representing truncated forms of either of the BMP receptors ALK-2 or BMPR-II preceded by an internal ribosome entry site (IRES) were introduced into the 3' untranslated region of TH. An frt-flanked neomycin-resistance (neo(r)) cassette was placed in the 3' end of the targeting constructs. Mice homozygous for the knock-in alleles showed various degrees of hypokinetic behavior, depending mainly on whether the neo(r) cassette was removed. In situ hybridization and immunohistochemistry showed that TH mRNA and protein were variously down-regulated in these mouse strains. Reduced levels of dopamine and noradrenalin were found in several brain areas. However, number and morphology of neurons in substantia nigra and their projections to striatum appeared normal in the neo(r)-positive TH hypomorphic mice as examined by markers for L-aromatic amino acid decarboxylase and the dopamine transporter. Elimination of the neo(r) cassette from the knock-in alleles partially restored TH and dopamine levels. The present neo(r)-positive TH hypomorphic mice show that nigrostriatal innervation develops independently of TH and should find use as a model for conditions of reduced catecholamine synthesis, as seen in, for example, L-dihydroxyphenylalanine-responsive dystonia/infantile parkinsonism.
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| 5. |
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| 6. |
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| 7. |
- Bedford, Lynn, et al.
(författare)
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Depletion of 26S proteasomes in mouse brain neurons causes neurodegeneration and Lewy-like inclusions resembling human pale bodies
- 2008
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Ingår i: Journal of Neuroscience. - 0270-6474 .- 1529-2401. ; 28:33, s. 8189-98
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Tidskriftsartikel (refereegranskat)abstract
- Ubiquitin-positive intraneuronal inclusions are a consistent feature of the major human neurodegenerative diseases, suggesting that dysfunction of the ubiquitin proteasome system is central to disease etiology. Research using inhibitors of the 20S proteasome to model Parkinson's disease is controversial. We report for the first time that specifically 26S proteasomal dysfunction is sufficient to trigger neurodegenerative disease. Here, we describe novel conditional genetic mouse models using the Cre/loxP system to spatially restrict inactivation of Psmc1 (Rpt2/S4) to neurons of either the substantia nigra or forebrain (e.g., cortex, hippocampus, and striatum). PSMC1 is an essential subunit of the 26S proteasome and Psmc1 conditional knock-out mice display 26S proteasome depletion in targeted neurons, in which the 20S proteasome is not affected. Impairment of specifically ubiquitin-mediated protein degradation caused intraneuronal Lewy-like inclusions and extensive neurodegeneration in the nigrostriatal pathway and forebrain regions. Ubiquitin and alpha-synuclein neuropathology was evident, similar to human Lewy bodies, but interestingly, inclusion bodies contained mitochondria. We support this observation by demonstrating mitochondria in an early form of Lewy body (pale body) from Parkinson's disease patients. The results directly confirm that 26S dysfunction in neurons is involved in the pathology of neurodegenerative disease. The model demonstrates that 26S proteasomes are necessary for normal neuronal homeostasis and that 20S proteasome activity is insufficient for neuronal survival. Finally, we are providing the first reproducible genetic platform for identifying new therapeutic targets to slow or prevent neurodegeneration.
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| 8. |
- Bengtsson, Henrik, et al.
(författare)
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Generation and characterization of a Gdf1 conditional null allele
- 2008
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Ingår i: Genesis. - : Wiley. - 1526-954X .- 1526-968X. ; 46:7, s. 368-372
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Tidskriftsartikel (refereegranskat)abstract
- Growth differentiation factor-1 (GDF1), a TGF-beta superfamily member, participates in early embryo patterning. Later functions are implied by the Gdf1 expression in the peripheral and central nervous system. Such roles of the gene have been difficult to study, because Gdf1 null mice die during late embryogenesis. Here, we report the production of a mouse carrying a conditional Gdf1 allele, with exon 2 flanked by loxP sites. Crossing these mice with CaMKIIalpha-Cre mice resulted in Gdf1 ablation in the forebrain postnatally. Such mice displayed no behavioral changes or altered expression levels in a set of hippocampal genes examined. However, excision of the floxed Gdf1 exon caused increased expression of the remaining part of the bicistronic Uog1-Gdf1 transcript in the hippocampus. This indicates that the transcript level is regulated by a negative feedback-loop, sensing presence of either the protein or the mRNA region encoded by Gdf1 exon 2.
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| 9. |
- Chou, J., et al.
(författare)
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Nigrostriatal alterations in bone morphogenetic protein receptor II dominant negative mice
- 2008
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Ingår i: Reconstructive Neurosurgery. - Wien : Springer. - 9783211782040 - 9783211782057 ; , s. 93-98
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Bokkapitel (övrigt vetenskapligt/konstnärligt)abstract
- BACKGROUND: We previously demonstrated that exogenous application of bone morphogenetic protein 7 (BMP7) reduced 6-hydroxydopamine-mediated neurodegeneration in a rodent model of Parkinson's disease. The purpose of this study is to examine the endogenous neurotrophic properties of BMP Receptor II in dopaminergic neurons of the nigrostriatal pathway. METHODS: Adult male BMPRII dominant negative (BMPRIIDN) mice and their wild type controls (WT) were placed in the activity chambers for 3 days to monitor locomotor activity. Animals were sacrificed for tyrosine hydroxylase (TH) immunostaining. A subgroup of BMPRIIDN and WT mice were injected with high doses of methamphetamine (MA) and were sacrificed for terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) histochemistry at 4 days after injection. RESULTS: BMPRIIDN mice had lower locomotor activity than the WT. There is a significant decrease in TH neuronal number in substantia nigra compacta, TH fiber density in the substantia nigra reticulata, and TH immunoreactivity in striatum in the BMPRIIDN mice, suggesting that deficiency in endogenous BMP signaling reduces dopaminergic innervation and motor function in the nigrostriatal pathway. Administration of MA increased TUNEL labeling in the substantia nigra in the BMPRIIDN mice. CONCLUSIONS: Endogenous BMPs have trophic effects on nigrostriatal dopaminergic neurons. Deficiency in BMP signaling increases vulnerability to insults induced by high doses of MA.
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| 10. |
- Clausen, Fredrik, et al.
(författare)
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Neutralization of interleukin-1 beta reduces cerebral edema and tissue loss and improves late cognitive outcome following traumatic brain injury in mice
- 2011
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Ingår i: European Journal of Neuroscience. - : Wiley. - 0953-816X .- 1460-9568. ; 34:1, s. 110-123
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Tidskriftsartikel (refereegranskat)abstract
- Increasing evidence suggests that interleukin-1 beta (IL-1 beta) is a key mediator of the inflammatory response following traumatic brain injury (TBI). Recently, we showed that intracerebroventricular administration of an IL-1 beta-neutralizing antibody was neuroprotective following TBI in mice. In the present study, an anti-IL-1 beta antibody or control antibody was administered intraperitoneally following controlled cortical injury (CCI) TBI or sham injury in 105 mice and we extended our histological, immunological and behavioral analysis. First, we demonstrated that the treatment antibody reached target brain regions of brain-injured animals in high concentrations (> 11 nm) remaining up to 8 days post-TBI. At 48 h post-injury, the anti-IL-1b treatment attenuated the TBI-induced hemispheric edema (P < 0.05) but not the memory deficits evaluated using the Morris water maze (MWM). Neutralization of IL-1 beta did not influence the TBI-induced increases (P < 0.05) in the gene expression of the Ccl3 and Ccr2 chemokines, IL-6 or Gfap. Up to 20 days post-injury, neutralization of IL-1 beta was associated with improved visuospatial learning in the MWM, reduced loss of hemispheric tissue and attenuation of the microglial activation caused by TBI (P < 0.05). Motor function using the rotarod and cylinder tests was not affected by the anti-IL-1 beta treatment. Our results suggest an important negative role for IL-1 beta in TBI. The improved histological and behavioral outcome following anti-IL-1 beta treatment also implies that further exploration of IL-1 beta-neutralizing compounds as a treatment option for TBI patients is warranted.
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