MicroRNA expression profiling in a mouse spinal cord injury model [P90, 7d]
ABSTRACT: Spinal cord injury (SCI) is a devastating condition resulting in permanent and irreversible deficits. Despite regeneration attempts, the neurons fail due to several dysfunctions. A number of studies have already revealed that, following SCI, microRNAs show two opposite kinds of alterations, one detrimental and one protective. However, there is still little evidence of specific microRNAs involved in axon regrowth. The aim of this project is the characterization of microRNA expression changes in sensorimotor cortex and corticospinal motor neurons, whose axons are severed by SCI. Overall design: miRNA profiling in SCI mouse model (P90) at a specific time point (7 days). Complete transection of the spinal cord was induced at the C5-C6 level in adult mice (3 WT vs. 3 SCI per each group). At 7 days post injury, the sensorimotor cortex was dissected, collected for RNA extraction and the HD sRNA library was sequenced.
Project description:Spinal cord injury (SCI) is a devastating condition resulting in permanent and irreversible deficits. Despite regeneration attempts, the neurons fail due to several dysfunctions. A number of studies have already revealed that, following SCI, microRNAs show two opposite kinds of alterations, one detrimental and one protective. However, there is still little evidence of specific microRNAs involved in axon regrowth. The aim of this project is the characterization of microRNA expression changes in sensorimotor cortex and corticospinal motor neurons, whose axons are severed by SCI. Overall design: miRNA profiling in SCI mouse model (P15 and P90) at different time points (12h and 3 days). Complete transection of the spinal cord was induced at the C5-C6 level in postnatal day 15 C57BL/6J male mice at 12h and 3 days after injury (3 WT vs. 3 SCI) and in adult mice at 12h and 3 days post injury (3 WT vs. 3 SCI per each group). The sensorimotor cortex was dissected, collected for RNA extraction and the HD sRNA library was sequenced.
Project description:Summary: Spinal cord injury (SCI) is a damage to the spinal cord induced by trauma or disease resulting in a loss of mobility or feeling. SCI is characterized by a primary mechanical injury followed by a secondary injury in which several molecular events are altered in the spinal cord often resulting in loss of neuronal function. Analysis of the areas directly (spinal cord) and indirectly (raphe and sensorimotor cortex) affected by injury will help understanding mechanisms of SCI. Hypothesis: Areas of the brain primarily affected by spinal cord injury are the Raphe and the Sensorimotor cortex thus gene expression profiling these two areas might contribute understanding the mechanisms of spinal cord injury. Specific Aim: The project aims at finding significantly altered genes in the Raphe and Sensorimotor cortex following an induced moderate spinal cord injury in T9.
Project description:Summary: Spinal cord injury (SCI) is a damage to the spinal cord induced by trauma or disease resulting in a loss of mobility or feeling. SCI is characterized by a primary mechanical injury followed by a secondary injury in which several molecular events are altered in the spinal cord often resulting in loss of neuronal function. Hypothesis: Spinal cord injury (SCI) induces a cascade of molecular events including the activation of genes associated with transcription factors, inflammation, oxidative stress, ionic imbalance, apoptosis and neuroregeneration which suggests the existance of endogenous reparative attempts. However, not all mechanisms following SCI are well known. Specific Aim: The goal of this project is to analyze the molecular events following spinal cord injury 1 cm above, below, and at the site of injury (T9), aiming at finding potential new targets to improve recovery and therapy.
Project description:The aneurysm clip impact-compression model of spinal cord injury (SCI) in animals mimics the primary mechanism of SCI in human, i.e. acute impact and persisting compression; and its histo-pathological and behavioural outcomes are extensively similar to the human SCI. In order to understand the distinct molecular events underlying this injury model, an analysis of global gene expression of the acute, subacute and chronic stages of a moderate to severe injury to the rat spinal cord was conducted using a microarray gene chip approach. Rat thoracic spinal cord (T7) was injured using aneurysm clip impact-compression injury model and the epicenter area of injured spinal cord was isolated for RNA extraction and processing and hybridization on Affymetrix GeneChip arrays.
Project description:To determine whether the expression levels of circular RNAs were altered and lay a foundation for future work, we used high-throughput microarray analysis to screen circular RNAs expression patterns in the spinal cord of adult rats after traumatic spinal cord injury (SCI), finally to evaluate the potential rat models as a platform for the development of novel therapeutic targets for spinal cord injury in future clinical studies. Overall six rats at 3 days post-SCI in two groups were used to perform the microarray. Overall design: Six rats were randomly assigned to two groups: rats in the sham control group (n=3) were treated with laminectomy alone without contusion; rats in the SCI group (n=3) were subjected to laminectomy plus contusion. Rats were anesthetized at 3 days post-SCI, and a 1cm long segment of spinal cord, including the injury epicenter, was dissected and collected for the experiment.
Project description:Traumatic spinal cord injury (SCI) often leads to loss of locomotor function. Neuroplasticity of spinal circuitry underlies some functional recovery and therefore represents a therapeutic target to improve locomotor function following SCI. However, the cellular and molecular mechanisms mediating neuroplasticity below the lesion level are not fully understood. The present study performed a gene expression profiling in the rat lumbar spinal cord at 1 and 3 weeks after contusive SCI at T9. The below-level gene expression profiles were compared with those of animals that were subjected to treadmill locomotor training. Rat lumbar spinal cords were taken for the microarray analysis at 1 and 3 weeks after contusive spinal cord injury at the T9 level. Another group of rats received treadmill locomotor training for 3 weeks, and theirs spinal cords were harvested for the microarray. The changes in gene expression after spinal cord injury were analyzed at the two time points. The influence of treadmill locomotor training was evaluated by comparing gene expression profiles between animals with or without treadmill training.
Project description:Sensorimotor dysfunction following incomplete spinal cord injury (SCI) is often characterized by paralysis, spasticity and pain. Previously, we showed that intrathecal (i.t.) administration of the albumin-oleic acid (A-OA) complex in rats with SCI produced partial improvement of these symptoms and that oral 2-hydroxyoleic acid (HOA), a non-hydrolyzable OA analogue), was efficacious in the modulation and treatment of nociception and pain-related anxiety, respectively. Here we observed that intrathecal treatment with the complex albumin-HOA (A-HOA) every 3 days following T9 spinal contusion injury promoted significant recovery in locomotor function and marked an inhibition of TA noxious reflex activity (i.e., nociception) in Wistar rats. To investigate the mechanism of action of A-HOA, microarray analysis was carried out in the spinal cord lesion area. Representative genes involved in pain and neuroregeneration were selected to validate the changes observed in the microarray analysis by quantitative real-time RT-PCR. Comparison of the expression between healthy rats, SCI rats, and SCI treated with A-HOA rats revealed relevant changes in the expression of genes associated with neuronal morphogenesis and growth, neuronal survival, pain and inflammation. Thus, treatment with A-HOA not only induced a significant overexpression of growth and differentiation factor 10 (GDF10), tenascin C (TNC), aspirin (ASPN) and sushi-repeat-containing X-linked 2 (SRPX2), but also a significant reduction in the expression of prostaglandin E synthase (PTGES) and phospholipases A1 and A2 (PLA1/2). Currently, SCI has very important unmet clinical needs. A-HOA proved to downregulate genes involved in inflammation and upregulate genes involved in neuron growth, which balanced the important body response to medular lesion and allowed recovery from paralysis and pain. Overall design: We use four different animals for each experimental group which were extracted and processed separately
Project description:Spinal cord injury (SCI) represents a major debilitating health issue with a direct socioeconomic burden on the public and private sectors worldwide. Although several studies have been conducted to identify the molecular progression of injury sequel due from the lesion site, still the exact underlying mechanisms and pathways of injury development have not been fully elucidated. In this work, based on OMICs, 3D MALDI imaging, cytokines arrays, confocal imaging we established for the first time that molecular and cellular processes occurring after spinal cord injury (SCI) are altered between the lesion proximity, i.e., rostral and caudal segments nearby the lesion (R1-C1) whereas segments distant from R1-C1, i.e., R2-C2 and R3-C3 levels co-expressed factors implicated in neurogenesis. Delay in T regulators recruitment between R1 and C1 favor discrepancies between the two segments. This is also reinforced by presence of neurites outgrowth inhibitors in C1, absent in R1. Moreover, the presence of immunoglobulins (IgGs) in neurons at the lesion site at 3 days, validated by mass spectrometry, may present additional factor that contributes to limited regeneration. Treatment in vivo with anti-CD20 one hour after SCI did not improve locomotor function and IgG expression. These results open the door of a novel view of the SCI treatment by considering the C1 as the therapeutic target.
Project description:Spinal cord injury (SCI) is a devastating neurological condition for which there are currently no effective treatment options to restore function. A major obstacle to the development of new therapies is our fragmentary understanding of the coordinated pathophysiological processeses triggered by damage to the human spinal cord. An additional challenge to translation of preclinical therapies is the reliance of clinical trials on standardized neurological assessments to enrol and stratify patients, rather than objective injury biomarkers. Here, we describe a systems biology approach to integrate decades of small-scale experiments with unbiased, genome-wide gene expression from the human spinal cord, revealing a gene regulatory network signature of the pathophysiological response to SCI. Our integrative analyses converge on an evolutionarily conserved gene subnetwork enriched for genes associated with the response to SCI by small-scale experiments, and whose expression is upregulated in a severity-dependent manner following injury and downregulated in functional recovery. We validate the severity-dependent upregulation of this subnetwork in prospective transcriptomic and proteomic studies. Our analysis provides a systems-level view of the coordinated molecular processes activated in response to SCI. Further, our results nominate quantitative biomarkers of injury severity and functional recovery, with the potential to facilitate development and translation of novel therapies.
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.