alpha-synuclein and Spinal-Cord-Injuries

Following Spinal Cord Injury (SCI), innumerable inflammatory and degenerative fluctuations appear in the injured site, and even remotely in manifold areas of the brain. Howbeit, inflammatory, degenerative, and oscillatory changes of motor cortices have been demonstrated to be due to SCI, according to recent studies confirming the involvement of cognitive areas of the brain, such as hippocampus and prefrontal cortex. Therefore, addressing SCI induced cognitive complications via different sights can be contributory in the treatment approaches.. Herein, we used 16 male Wistar rats (Sham = 8, SCI = 8). Immunohistochemical results revealed that spinal cord contusion significantly increases the accumulation of alpha-synuclein and decreases the expression of Doublecortin (DCX) in the hippocampal regions like Cornu Ammonis1 (CA1) and Dentate Gyrus (DG). Theses degenerative manifestations were parallel with a low expression of Achaete-Scute Family BHLH Transcription Factor 1 (ASCL1), SRY (sex determining region Y)-box 2 (SOX2), and dopaminergic receptors (D1 and D5). Additionally, based on the TUNEL assay analysis, SCI significantly increased the number of apoptotic cells in the CA1 and DG regions. Cognitive function of the animals was assessed, using the O-X maze and Novel Object Recognition (NORT); the obtained findings indicted that after SCI, hippocampal neurodegeneration significantly coincides with the impairment of learning, memory and recognition capability of the injured animals.. Based on the obtained findings, herein SCI reduces neurogenesis, decreases the expression of D1 and D5, and increases apoptosis in the hippocampus, which are all associated with cognitive function deficits.

Topics: alpha-Synuclein; Animals; Cognition; Hippocampus; Male; Neurogenesis; Rats; Rats, Wistar; Spinal Cord Injuries

Synucleinopathies are neurodegenerative diseases in which α-synuclein protein accumulates in neurons and glia. In these diseases, α-synuclein forms dense intracellular aggregates that are disease hallmarks and actively contribute to tissue pathology. Interestingly, many pathological mechanisms, including iron accumulation and lipid peroxidation, are shared between classical synucleinopathies such as Alzheimer's disease, Parkinson's disease and traumatic spinal cord injury (SCI). However, to date, no studies have determined if α-synuclein accumulation occurs after human SCI. To examine this, cross-sections from injured and non-injured human spinal cords were immunolabeled for α-synuclein. This showed robust α-synuclein accumulation in profiles resembling axons and astrocytes in tissue surrounding the injury, revealing that α-synuclein markedly aggregates in traumatically injured human spinal cords. We also detected significant iron deposition in the injury site, a known catalyst for α-synuclein aggregation. Next a rodent SCI model mimicking the histological features of human SCI revealed aggregates and structurally altered monomers of α-synuclein are present after SCI. To determine if α-synuclein exacerbates SCI pathology, α-synuclein knockout mice were tested. Compared to wild type mice, α-synuclein knockout mice had significantly more spared axons and neurons and lower pro-inflammatory mediators, macrophage accumulation, and iron deposition in the injured spinal cord. Interestingly, locomotor analysis revealed that α-synuclein may be essential for dopamine-mediated hindlimb function after SCI. Collectively, the marked upregulation and long-lasting accumulation of α-synuclein and iron suggests that SCI may fit within the family of synucleinopathies and offer new therapeutic targets for promoting neuron preservation and improving function after spinal trauma.

Topics: Adult; Aged; Aged, 80 and over; alpha-Synuclein; Animals; Astrocytes; Biomarkers; Cell Death; Disease Models, Animal; Dopamine; Female; Gene Knockdown Techniques; Humans; Inflammation; Inflammation Mediators; Iron; Male; Mice; Middle Aged; Neurons; Organ Size; Rats; Rodentia; Signal Transduction; Spinal Cord; Spinal Cord Injuries; Young Adult

Topics: alpha-Synuclein; Humans; Lewy Bodies; Spinal Cord; Spinal Cord Injuries; Synucleinopathies

Endogenous α-synuclein (α-Syn) is involved in many pathophysiological processes in the secondary injury stage after acute spinal cord injury (SCI), and the mechanism governing these functions has not been thoroughly elucidated to date. This research aims to characterize the effect of α-Syn knockdown on transcriptional levels after SCI and to determine the mechanisms underlying α-Syn activity based on RNA-seq.. The establishment of a rat model of lentiviral vector-mediated knockdown of α-Syn in Sprague-Dawley rats with T3 spinal cord contusion (LV_SCI group). The results of the RNA-seq analysis showed that there were 337 differentially expressed genes (DEGs) between the SCI group and the LV_SCI group, and 153 DEGs specific to LV_SCI between the (SCI vs LV_SCI) and (SCI vs CON) comparisons. The top 20 biological transition terms were identified by Gene ontology (GO) analysis. The Kyoto Gene and Genomic Encyclopedia (KEGG) analysis showed that the LV_SCI group significantly upregulated the cholinergic synaptic & nicotine addiction and the neuroactive ligand receptor interaction signaling pathway. Enriched chord analysis analyzes key genes. Further cluster analysis, gene and protein interaction network analysis and RT-qPCR results showed that Chrm2 and Chrnb2 together significantly in both pathways. The proliferation of muscarinic cholinergic receptor subtype 2 (Chrm2) and nicotinic cholinergic receptor subtype β2 (Chrnb2), and the neurogenesis were elevated in the injury site of LV_SCI group by immunofluorescence. Further by subcellular localization, the LV_SCI group enhanced the expression of Chrnb2 at the cell membrane.. Knockdown of α-Syn after SCI enhance motor function and promote neurogenesis probably through enhancing cholinergic signaling pathways and neuroreceptor interactions. This study not only further clarifies the understanding of the mechanism of knockdown of α-Syn on SCI but also helps to guide the treatment strategy for SCI.

Topics: alpha-Synuclein; Animals; Animals, Genetically Modified; Biomarkers; Cholinergic Neurons; Computational Biology; Gene Expression Profiling; Gene Expression Regulation; Gene Knockdown Techniques; Gene Ontology; Gene Regulatory Networks; Neurogenesis; Rats; RNA, Messenger; Signal Transduction; Spinal Cord Injuries; Transcriptome

The prognosis of spinal cord injury (SCI) is closely related to secondary injury, which is dominated by neuroinflammation. There is evidence that α-synuclein aggregates after SCI and that inhibition of α-synuclein aggregation can improve the survival of neurons after SCI, but the mechanism is still unclear. This study was designed to investigate the effects of α-synuclein on neuroinflammation after SCI and to determine the underlying mechanisms.. A T3 spinal cord contusion model was established in adult male Sprague-Dawley rats. An SNCA-shRNA-carrying lentivirus (LV-SNCA-shRNA) was injected into the injury site to block the expression of α-synuclein (forming the SCI+KD group), and the SCI and sham groups were injected with an empty vector. Basso-Beattie-Bresnahan (BBB) behavioural scores and footprint analysis were used to detect motor function. Inflammatory infiltration and myelin loss were measured in the spinal cord tissues of each group by haematoxylin-eosin (HE) and Luxol Fast Blue (LFB) staining, respectively. Immunohistochemistry, Western blot analysis, and RT-qPCR were used to analyse protein expression and transcription levels in the tissues. Immunofluorescence was used to determine the morphology and function of glial cells and the expression of matrix metalloproteinase-9 in the central canal of the spinal cord. Finally, peripheral serum cytokine levels were determined by enzyme-linked immunosorbent assay.. Compared with the SCI group, the SCI+KD group exhibited reduced inflammatory infiltration, preserved myelin, and functional recovery. Specifically, the early arrest of α-synuclein inhibited the pro-inflammatory factors IL-1β, TNF-α, and IL-2 and increased the expression of the anti-inflammatory factors IL-10, TGF-β, and IL-4. The neuroinflammatory response was regulated by reduced proliferation of Iba1+ microglia/macrophages and promotion of the shift of M1-polarized Iba1+/iNOS+ microglia/macrophages to M2-polarized Iba1+/Arg1+ microglia/macrophages after injury. In addition, compared with the SCI group, the SCI+KD group also exhibited a smaller microglia/astrocyte (Iba1/GFAP) immunostaining area in the central canal, lower MMP-9 expression, and improved cerebrospinal barrier function.. Lentivirus-mediated downregulation of α-synuclein reduces neuroinflammation, improves blood-cerebrospinal barrier function, promotes functional recovery, reduces microglial activation, and promotes the polarization of M1 microglia/macrophages to an M2 phenotype to confer a neuroprotective immune microenvironment in rats with SCI.

Topics: alpha-Synuclein; Animals; Down-Regulation; Genetic Vectors; Inflammation; Lentivirus; Male; Rats; Rats, Sprague-Dawley; Recovery of Function; RNA, Small Interfering; Spinal Cord Injuries

Topics: alpha-Synuclein; Animals; Humans; Spinal Cord Injuries

Growing evidence suggests that oxidative stress plays a critical role in neuronal destruction characteristic of Parkinson's disease (PD). However, the molecular mechanisms of oxidative stress-mediated dopaminergic cell death are far from clear. In the current investigation, we tested the hypothesis that acrolein, an oxidative stress and lipid peroxidation (LPO) product, is a key factor in the pathogenesis of PD. Using a combination of in vitro, in vivo, and cell free models, coupled with anatomical, functional, and behavioral examination, we found that acrolein was elevated in 6-OHDA-injected rats, and behavioral deficits associated with 6-OHDA could be mitigated by the application of the acrolein scavenger hydralazine, and mimicked by injection of acrolein in healthy rats. Furthermore, hydralazine alleviated neuronal cell death elicited by 6-OHDA and another PD-related toxin, rotenone, in vitro. We also show that acrolein can promote the aggregation of alpha-synuclein, suggesting that alpha-synuclein self-assembly, a key pathological phenomenon in human PD, could play a role in neurotoxic effects of acrolein in PD models. These studies suggest that acrolein is involved in the pathogenesis of PD, and the administration of anti-acrolein scavengers such as hydralazine could represent a novel strategy to alleviate tissue damage and motor deficits associated with this disease.

Topics: Acrolein; alpha-Synuclein; Animals; Cell Death; Cell Line; Dopamine; Dopaminergic Neurons; Oxidative Stress; Parkinson Disease; Rats; Spinal Cord Injuries; Substantia Nigra

Spinal cord injury (SCI) often causes neurological deficits with poor recovery; the treatment, however, is far from satisfaction, and the mechanisms remain unclear. Using immunohistochemistry and western blotting analysis, we found α-synuclein (SNCA) was significantly up-regulated in the spinal caudal segment of rats subjected to spinal cord transection at 3 days post-operation. Moreover, the role of SNCA on neuronal growth and apoptosis in vitro was determined by using overexpressing and interfering SNCA recombined plasmid vectors, and the underlying mechanism was detected by QRT-PCR and western blotting. Spinal neurons transfected with SNCA-shRNA lentivirus gave rise to an optimal neuronal survival, while it results in cell apoptosis in SNCA-ORF group. In molecular level, SNCA silence induced the up-regulation of CNTF and down-regulation of Caspase7/9. Together, endogenous SNCA plays a crucial role in spinal neuronal survival, in which the underlying mechanism may be linked to the regulation both apoptotic genes (Caspase7/9) and CNTF. The present findings therefore provide novel insights into the role of SNCA in spinal cord and associated mechanism, which may provide novel cue for the treatment of SCI in future clinic trials.

Topics: alpha-Synuclein; Animals; Antigens, Nuclear; Apoptosis; Cell Survival; Ciliary Neurotrophic Factor; Disease Models, Animal; Female; Lentivirus; Nerve Growth Factors; Nerve Tissue Proteins; Neurons; Open Reading Frames; Rats, Sprague-Dawley; RNA, Small Interfering; Spinal Cord; Spinal Cord Injuries; Transfection; Virus Assembly

Microglial cell migration and infiltration plays a critical role in spinal cord injury after thoracoabdominal aortic surgery. In our previous study, α-synuclein, a presynaptic protein was shown to be released from injured neurons and cause microglial cell activation. Here, we aimed to explore the effect of α-synuclein on microglial cell migration. Primary microglial cells were isolated from Sprague-Dawley rats and then exposed different doses (0.2, 0.4, and 0.6 μM) of α-synuclein oligomers. The mRNA and protein levels of HIF-1α were then analyzed by qRT-PCR and Western blot. Cell migration was examined by a 96-well Boyden chamber. Moreover, toll-like receptor (TLR) 2-expression as well as TLR7/8-expression was inhibited by specific siRNA transfection. HIF-1α was overexpressed by Ad-HIF-1α transfection. In the results, α-synuclein was found to stimulate HIF-1α accumulation in microglial cells in a dose-dependent manner. Silencing HIF-1α expression dampened α-synuclein induced microglial cell migration. Furthermore, blockade of TLR7/8 expression but not TLR2 expression reduced HIF-1α accumulation in microglial cells. In addition, overexpressed HIF-1α, along with Src, prompted caveolin-1 expression and phosphorylation, as well as migration in microglial cells. Α-synuclein acts via TLR7/8 and enhances HIF-1α expression, which might play a regulatory role in microglial cell migration. © 2017 Wiley Periodicals, Inc.

Topics: alpha-Synuclein; Animals; Aorta; Cell Movement; Cells, Cultured; Hypoxia-Inducible Factor 1, alpha Subunit; Male; Microglia; Rats; Rats, Sprague-Dawley; Spinal Cord Injuries; Toll-Like Receptor 7; Toll-Like Receptor 8; Vascular Surgical Procedures

Spinal cord injury (SCI) often causes severe functional impairment with poor recovery. The treatment, however, is far from satisfaction, and the mechanisms remain unclear. By using proteomics and western blot, we found spinal cord transection (SCT) resulted in a significant down-regulation of α-synuclein (SNCA) in the motor cortex of SCT rats at 3 days post-operation. In order to detect the role of SNCA, we used SNCA-ORF/shRNA lentivirus to upregulate or knockdown SNCA expression. In vivo, SNCA-shRNA lentivirus injection into the cerebral cortex motor area not only inhibited SNCA expression, but also significantly enhanced neurons' survival, and attenuated neuronal apoptosis, as well as promoted motor and sensory function recovery in hind limbs. While, overexpression SNCA exhibited the opposite effects. In vitro, cortical neurons transfected with SNCA-shRNA lentivirus gave rise to an optimal neuronal survival and neurite outgrowth, while it was accompanied by reverse efficiency in SNCA-ORF group. In molecular level, SNCA silence induced the upregulation of Bcl-2 and the downregulation of Bax, and the expression of NGF, BDNF and NT3 was substantially upregulated in cortical neurons. Together, endogenous SNCA play a crucial role in motor and sensory function regulation, in which, the underlying mechanism may be linked to the regulation of apoptosis associated with apoptotic gene (Bax, Bcl2) and neurotrophic factors expression (NGF, BDNF and NT3). These finds provide novel insights to understand the role of SNCA in cerebral cortex after SCT, and it may be as a novel treatment target for SCI repair in future clinic trials.

Topics: alpha-Synuclein; Animals; Apoptosis; bcl-2-Associated X Protein; Brain-Derived Neurotrophic Factor; Cell Survival; Cells, Cultured; Cerebral Cortex; Female; Nerve Growth Factor; Nerve Growth Factors; Proto-Oncogene Proteins c-bcl-2; Rats; Rats, Sprague-Dawley; Recovery of Function; RNA Interference; RNA, Small Interfering; Spinal Cord; Spinal Cord Injuries