Project description:Nogo-A localized on myelin adaxonal membrane in the adult CNS is well known for its role as neurite outgrowth inhibitor following a lesion. Nogo-A KO mice show enhanced regenerative/compensatory fiber growth following CNS lesion. However, changes undergoing in their intact CNS have not been studied. Moreover, Nogo-A in the intact adult CNS in also expressed in some neuronal subpopulations, e.g. in the hippocampus, olfactory bulbs and dorsal root ganglia. We compared the intact adult CNS (spinal cord) of Nogo-A KO mice in order to identify: potential compensating molecules which could be interesting new inhibitory neurite outgrowth candidates, possible molecules involved in the up to now not yet clarified downstream signalling pathway of Nogo-A, additional new functions for myelin or neuronal Nogo-A in the intact adult CNS. Keywords: gene expression, Nogo-A KO, spinal cord, adult, naive, unlesioned
Project description:Nogo-A localized on myelin adaxonal membrane in the adult CNS is well known for its role as neurite outgrowth inhibitor following a lesion. Nogo-A KO mice show enhanced regenerative/compensatory fiber growth following CNS lesion. However, changes undergoing in their intact CNS have not been studied. Moreover, Nogo-A in the intact adult CNS in also expressed in some neuronal subpopulations, e.g. in the hippocampus, olfactory bulbs and dorsal root ganglia. We compared the intact adult CNS (spinal cord) of Nogo-A KO mice in order to identify: potential compensating molecules which could be interesting new inhibitory neurite outgrowth candidates, possible molecules involved in the up to now not yet clarified downstream signalling pathway of Nogo-A, additional new functions for myelin or neuronal Nogo-A in the intact adult CNS. Keywords: gene expression, Nogo-A KO, spinal cord, adult, naive, unlesioned Spinal cords from 3 adult C57Bl/6 wild type and Nogo-A KO mice have been explanted. Total RNA has been extracted and processed for hybridization on Mouse 430 2.0 Affymetrix GeneChips. Following scanning and first analysis with MAS 5.0, further analysis was performed by GeneSpring 7.2 (Silicon Genetics, Redwood City, CA). A present call filter (2 out of 3 present calls in at least one out of the different studied conditions) was applied. Normalization was run per chip as well as per gene to the median of the control replicates. Data were statistical restricted through a 1-way Anova (p=0.05). A final threshold of =1.2 folds of increase or decrease in the expression level of each single transcript was applied. Regulated transcripts have been assigned to functional categories according to GeneOntology as well as literature and database mining (Pubmed and Bioinformatics Harvester EMBL Heidelberg).
Project description:We asked what genes are significantly differentially regulated in the spinal cord of SCI trkB.T1 WT and trkB.T1 KO mice. TrkB.T1 is upregulated shortly after SCI although the precise mechanisms underyling this upregulation are poorly understood. In the trkB.T1 null, we show less mechanical allodynia and better locomotor recovery following SCI. The microarray studies helped us to elucidate a signaling pathway that is differently regulated in the WT versus KO mice at 1 day after SCI. In this study, we did not examine gene changes within a genotype after SCI. Rather, we examined DGE by genotype at each time point. Spinal cord tissue from WT and KO mice in a sham condition (intact spinal cord) versus 1D, 3D and 7D following SCI was harvested for microarray analyses.
Project description:We have previously shown that Il1a-knockout (KO) mice exhibit rapid (at day 1) and persistent improvements in locomotion associated with reduced lesion volume compared with Il1b-KO mice and C57BL/6 controls after traumatic spinal cord injury (SCI). To investigate the mechanism by which Il1a mediates its detrimental effect, we analyzed the transcriptome of the injured spinal cord of Il1a-KO, Il1b-KO and C57BL/6 mice at 24 hours after SCI using GeneChip microarrays. Il1a-KO, Il1b-KO and C57BL/6 mice were subjected to a 50-kdyn SCI and a 6-mm spinal cord segment centered over the site of contusion extracted for RNA isolation and microarray analysis.
Project description:We have previously shown that Il1a-knockout (KO) mice exhibit rapid (at day 1) and persistent improvements in locomotion associated with reduced lesion volume compared with Il1b-KO mice and C57BL/6 controls after traumatic spinal cord injury (SCI). To investigate the mechanism by which Il1a mediates its detrimental effect, we analyzed the transcriptome of the injured spinal cord of Il1a-KO, Il1b-KO and C57BL/6 mice at 24 hours after SCI using GeneChip microarrays.
Project description:The neurite outgrowth inhibitory myelin protein Nogo-A has been well studied in the context of central nervous system (CNS) injury and disease. We studied the effects of the application of neutralizing anti-Nogo-A antibodies (11C7 and 7B12) in intact CNS tissue in vitro using rat organotypic hippocampal slice cultures. This study had the purpose of elucidating the role of Nogo-A in the adult intact CNS and determining the consequences of its neutralization through antibody application. In vitro cultures treated with anti-Nogo-A antibody showed an elicited growth response. The results also gave indications that hippocampal circuitry might be altered due to the regulation at the synaptic and neurotransmission level.
Project description:Neonatal spinal cord tissues exhibit remarkable regenerative capabilities than adult tissues after injury, but the role of extracellular matrix (ECM) in this process has remained elusive. Here we found that early developmental spinal cord had higher levels of ECM proteins associated with neural development and axon growth, but fewer inhibitory proteoglycans compared to adult spinal cord. Decellularized spinal cord ECM from neonatal (DNSCM) and adult (DASCM) rabbits preserved these differences. DNSCM promoted proliferation, migration, and neuronal differentiation of neural progenitor cells (NPCs), and facilitated axonal outgrowth and regeneration of spinal cord organoids than DASCM. Pleiotrophin (PTN) and Tenascin (TNC) in DNSCM were identified as contributors to these abilities. Furthermore, DNSCM demonstrated superior performance as a delivery vehicle for NPCs and organoids in rats with spinal cord injury (SCI). It suggests that ECM cues from early development stages might significantly contribute to the prominent regeneration ability in spinal cord.
Project description:Neonatal spinal cord tissues exhibit remarkable regenerative capabilities compared to adult tissues following injury. Although some cellular signaling pathways involved in the process have been identified, the specific role of extracellular matrix (ECM) responsible for neonatal spinal cord regeneration has remained elusive. Here we revealed that early developmental spinal cord contained a higher abundance of ECM proteins associated with neural development and axon growth but fewer inhibitory proteoglycans compared to adult spinal cord. Decellularized spinal cord ECM from neonatal (DNSCM) and adult (DASCM) rabbits preserve the major difference of native spinal cord tissues in both stages. Compared to DASCM, DNSCM promoted proliferation, migration, and neuronal differentiation of neural progenitor cells (NPCs), as well as facilitated the long-distance axonal outgrowth and axon regeneration of spinal cord organoids. Pleiotrophin (PTN) and Tenascin (TNC) in DNSCM were identified as contributors to the remarkable neural regeneration ability. Furthermore, DNSCM demonstrated superior performance when used as a delivery vehicle for NPCs and organoids in rats with spinal cord injury (SCI). It suggests that ECM cues derived from different development stage might contribute to the distinct regeneration ability of spinal cord.
Project description:The neurite outgrowth inhibitory myelin protein Nogo-A has been well studied in the context of central nervous system (CNS) injury and disease. We studied the effects of the application of neutralizing anti-Nogo-A antibodies (11C7 and 7B12) in intact CNS tissue in vitro using rat organotypic hippocampal slice cultures. This study had the purpose of elucidating the role of Nogo-A in the adult intact CNS and determining the consequences of its neutralization through antibody application. In vitro cultures treated with anti-Nogo-A antibody showed an elicited growth response. The results also gave indications that hippocampal circuitry might be altered due to the regulation at the synaptic and neurotransmission level. Experiment Overall Design: Nogo-A function in the intact CNS tissue is not well known, but its neutralization in vivo produced a transitory growth response of Purkinje axons and of the corticospinal tract in intact adult rats (Buffo et al., 2000; Bareyre et al., 2002; Gianola et al., 2003). Nogo-A is relatively highly expressed in oligodendrocytes and some neurons of the hippocampus (Huber et al., 2002; Meier et al., 2003; Gil et al., 2006; Trifunovski et al., 2006). Organotypic hippocampal slice cultures are a good in vitro model to study hippocampal function and structure (Stoppini et al., 1991; 1993; Bahr, 1995; Gahwiler et al., 1997; Hakkoum et al., 2006). They mature in vitro and retain many in vivo features from a structural and functional perspective. We chose this model to study the effects of acute Nogo-A neutralization, using two function blocking monoclonal antibodies, 11C7 and 7B12 (Oertle et al., 2003; Wiessner et al., 2003; Liebscher et al., 2005), exclusively targeted against the Nogo-A specific region. Hippocampal slices from P7 Wistar rats were cultured for 21 DIV. Control untreated cultures where cultured for additional 5days for a total of 26DIV, while control IgG, and 11C7 and 7B12 were added to 21DIV cultures which were then further cultured for 5days, changing medium every 2 days. All the conditions were repeated in triplicates with separate cultures from different animals. For each condition and experimental replicate 24 cultures were pooled together before being processed for RNA extraction. Data analysis was performed by GeneSpring 7.2 (Silicon Genetics, Agilent, CA, US) comparing 11C7 and 7B12 treated samples versus Not treated and IgG treated, as controls. A present call filter (2 out of 3 present calls in at least one out of the 3 experimental replicates) was applied. Normalization was run per chip as well as per gene to the median of the control replicates. Data were statistical restricted through a 1-way Anova (pâ?¤0.05). A final threshold of â?¥1.2 fold of increase or decrease in the expression level of each single transcript was applied. Regulated transcripts have been assigned to functional categories according to GeneOntology as well as literature and database mining (Pubmed; Bioinformatics Harvester EMBL Heidelberg; Rat Genome Database).
Project description:We compare transcriptomic profiles of human induced pluripotent stem cells (iPSCs), motor neurons (MNs) in vitro differentiated from iPSCs or human ESCs containing a HB9::GFP reporter for MNs, and human fetal spinal cords. The purpose of this comparison is to assess the extent of molecular similarities between in vitro differentiated MNs and in vivo fetal or adult spinal cord MNs. Data for adult spinal cord MNs are published from other studies: GSE3526, GSE19332, GSE20589, and GSE40438.