Project description:To profile downstream gene-expression changes regulated by DLK, we profiled motoneurons in the lumbar spinal cord using RiboTag, following sciatic nerve crush in both control and Dlk conditional knockout (Dlk ΔMN) mice. We found that DLK regulates the expression of many secreted proteins, which have the potential to influence the behavior of other cells, including the immune system.
Project description:Sciatic nerve ligation was performed on cohorts of 2-month and 24-month old animals. Resulting gene-expression data were generated from sciatic nerve 1 and 4 days after injury compared to naïve animals. Results show differences in sciatic nerve responses with normal aging. Total RNA taken from sciatic nerves from 2-month and 24-month old animals at either day 0, 1 and 4 after sciatic nerve crush injury.
Project description:Dual Leucine-zipper Kinase (DLK)-dependent stress signaling is a critical determinant of neuronal survival and regenerative potential following axon damage, but it remains uncertain whether injury-activated DLK is adequate to initiate and maintain a pro-regenerative transcriptional response in the CNS. Using a drug-activatable DLK construct, we stimulated stress signaling for comparison of the retinal transcriptional response to, and in addition to, the response stimulated by mouse optic nerve injury in wildtype mice and in the context of partial axon regeneration enabled by disruption of the tumor suppressor PTEN.
Project description:Sciatic nerve crush (SNC) triggers sterile inflammation within the distal nerve and de-afferented dorsal root ganglia (DRGs). In the nerve, neutrophils and pro-inflammatory Ly6Chigh monocytes appear first and rapidly give way to Ly6Clow resolving macrophages. Transcriptional profiling of injured nerve tissue identifies six macrophage subpopulations, repair Schwann cells and mesenchymal cells as the main cell types. Macrophages at the nerve crush site are distinct from macrophages associated with degenerating nerve fibers. Monocytes and macrophages in the injured nerve “eat” apoptotic cell corpses of leukocytes and thereby contribute to an anti-inflammatory milieu. Studies with chimeric mice show that following SNC few blood-derived immune cells enter DRGs. Myeloid cells in the injured nerve, but not DRGs, express the receptor for the chemokine GM-CSF. In the absence of GM-CSF, conditioning-lesion induced regeneration of DRG neuron central projections is abrogated. Thus, a carefully orchestrated immune response in the nerve is required for conditioning-lesion induced neurorepair.
Project description:Reactive gliosis is a complex process that involves profound changes in gene expression. We used microarray to indentify differentially expressed genes and to investigate the molecular mechanisms of reactive gliosis in optic nerve head in response to optic nerve crush injury. C57Bl/6 female mice were 6-8 weeks old at the time of optic nerve crush surgery. The optic nerve in the left eye was crush 1 mm behind the globe for 10 seconds and the right eye served as contralateral control. The animals were allowed to recover for 1 day, 3 day, 1 week, 3 weeks and 3 months before the optic nerve heads were collected. The naive control mice did not receive any surgery in either eye. Due to the small tissue size of the mouse optic nerve head, two optic nerve heads were pooled together for each microarray chip. The left eyes and the right eyes of two mice were combined respectively to form one pair of experiment and control samples. There were five biological replicates (10 mice) for each condition.
Project description:We used two groups of C57BL/6J mice, one with optic nerve crush on one eye, and another with no crush as control. Three mice were subjected to optic nerve crush, with sample names 121, 113, 114 and two were used as control with sample names 118 and 119. For the optic nerve crush, a surgical peritomy was made behind and above the eyeball and the eye muscles were gently retracted to expose the optic nerve. Dumont #5 forceps (FST) were used to crush the optic nerve approximately 0.5-1 mm behind the globe without damaging retinal vessels or affecting the blood supply.
Project description:Sciatic nerve axon segments from adult mouse were isolated. Following enzymatic digestion, lysate was subjected to incubation with beads conjugated with either clathrin heavy chain antibody or control IgG antibody for clathrin immunoprecipitation. MudPIT analysis was subsequently performed to identify proteins co-precipitating with clathrin.
Project description:In this study, we analyzed the transcriptome profiles of mouse sciatic nerves subjected to crush injuries after inducible deletion of Raptor conditionally in Schwann cells (using a PLPCreERT2-driven recombination of floxed alleles) as compared to controls (floxed Raptor homozygous, PLPCreERT2-negative). The transcriptome profiles of the contralateral uninjured nerves were also analyzed. Differentially expressed genes, defined as genes with a fold change>1.2 and fold discovery rate <0.05, in injured and contralateral nerves of mutants compared to controls were subjected to gene ontology analysis. Additionally, differentially expressed genes in injured mutants nerves as compared to injured control nerves were further analyzed for enrichment of transcription factor binding motifs in the corresponding promoter regions using the bioinformatic tool Homer version 4.9 (Heinz et al., Molecular Cell, 2010)
Project description:BackgroundCytokines are essential cellular modulators of various physiological and pathological activities, including peripheral nerve repair and regeneration. However, the molecular changes of these cellular mediators after peripheral nerve injury are still unclear. This study aimed to identify cytokines critical for the regenerative process of injured peripheral nerves.MethodsThe sequencing data of the injured nerve stumps and the dorsal root ganglia (DRGs) of Sprague-Dawley (SD) rats subjected to sciatic nerve (SN) crush injury were analyzed to determine the expression patterns of genes coding for cytokines. PCR was used to validate the accuracy of the sequencing data.ResultsA total of 46, 52, and 54 upstream cytokines were differentially expressed in the SNs at 1 day, 4 days, and 7 days after nerve injury. A total of 25, 28, and 34 upstream cytokines were differentially expressed in the DRGs at these time points. The expression patterns of some essential upstream cytokines are displayed in a heatmap and were validated by PCR. Bioinformatic analysis of these differentially expressed upstream cytokines after nerve injury demonstrated that inflammatory and immune responses were significantly involved.ConclusionsIn summary, these findings provide an overview of the dynamic changes in cytokines in the SNs and DRGs at different time points after nerve crush injury in rats, elucidate the biological processes of differentially expressed cytokines, especially the important roles in inflammatory and immune responses after peripheral nerve injury, and thus might contribute to the identification of potential treatments for peripheral nerve repair and regeneration.