Expression Of Neurofilament Proteins Research Articles

Expression of dendrin, neurofilament and glial fibrillary acidic protein in the brain of the dogfish Scyliorhinus canicula L.

Background and purpose: Dendrin is a brain and renal protein that is supposed to be involved in cytoskeletal modifications at the synapse and a part of the slit diaphragm and podocytes. Here, we aimed to investigate dendrin expression in dogfish brain since this newly discovered protein was never reported in fish. We compared the expression of dendrin to those of glial (GFAP) and neuronal (NF) proteins, which have already been described in the dogfish brain. Materials and methods: Histological and immunofluorescent techniques were performed on tissue samples. The obtained data were statistically analyzed. Results: Our results have shown that dendrin is expressed in all observed parts of the dogfish brain. In the forebrain, both observed parts (telencephalon and olfactory lobes) expressed dendrin. Regarding the percentage area of dendrin expression, it is expressed more in olfactory lobes than in the telencephalon. Compared with GFAP and NF expression, the expression of dendrin significantly differs in both parts of the forebrain. The highest dendrin expression was noticed in the midbrain. In dogfish midbrain, the difference in expression of dendrin in comparison to those of GFAP and NF was even more significant. The percentage area of dendrin expression in the hindbrain (cerebellum and medulla oblongata) was smaller than those in the forebrain and midbrain, contrary the percentage area of intermediate filaments GFAP and NF were significantly higher. Conclusion: These results are the first report on dendrin expression in the dogfish brain opening the path for future studies on its role and function.

Cortical Layer Markers Expression and Increased Synaptic Density in Interstitial Neurons of the White Matter from Drug-Resistant Epilepsy Patients.

The interstitial neurons in the white matter of the human and non-human primate cortex share a similar developmental origin with subplate neurons and deep-layer cortical neurons. A subset of interstitial neurons expresses the molecular markers of subplate neurons, but whether interstitial neurons express cortical layer markers in the adult human brain remains unexplored. Here we report the expression of cortical layer markers in interstitial neurons in the white matter of the adult human brain, supporting the hypothesis that interstitial neurons could be derived from cortical progenitor cells. Furthermore, we found increased non-phosphorylated neurofilament protein (NPNFP) expression in interstitial neurons in the white matter of drug-resistant epilepsy patients. We also identified the expression of glutamatergic and g-aminobutyric acid (GABAergic) synaptic puncta that were distributed in the perikarya and dendrites of interstitial neurons. The density of glutamatergic and GABAergic synaptic puncta was increased in interstitial neurons in the white matter of drug-resistant epilepsy patients compared with control brain tissues with no history of epilepsy. Together, our results provide important insights of the molecular identity of interstitial neurons in the adult human white matter. Increased synaptic density of interstitial neurons could result in an imbalanced synaptic network in the white matter and participate as part of the epileptic network in drug-resistant epilepsy.

Periurethral and Intravenous Injections of Adipose-Derived Stem Cells to Promote Local Tissue Recovery in a Rat Model of Stress Urinary Incontinence

ObjectiveTo compare the effects of periurethral and intravenous injection of adipose-derived stem cells (ADSCs) on voiding function and tissue recovery in a stress urinary incontinence (SUI) rat model. MethodsSixty-four postpartum rats were randomly allocated to a normal group and the SUI model was established in 48 rats by vagina balloon dilation and bilateral ovariectomy. The SUI rats were randomized into 3 groups and received urethral injection of PBS (SUI group), periurethral injection of ADSCs (PU group), and intravenous injection of ADSCs (IV group) in 10 days after the ovariectomy. After 1, 7, and 14 days, ADSCs were tracked in urethra specimen. The urinary function of the remaining rats was analyzed at day 28, and urethral tissues were harvested for Western blotting and histochemical analyses. ResultsAlpha smooth muscle actin, myosin heavy chain, vascular endothelial growth factor, and neurofilament protein expression was increased in the IV and PU groups. Voiding function was also improved, with no significant differences between the IV and PU groups. The cell retention rate in rat urethral tissues was higher in the PU group than that in the IV group. Compared with the IV group, myosin heavy chain, vascular endothelial growth factor, neurofilament and transforming growth factor-β1 (TGF-β1)/Smad pathway protein expression levels were significantly higher in the PU group, while alpha smooth muscle actin expression was significantly lower (P < .05). ConclusionPeriurethral and intravenous injection of ADSCs induces different degrees of recovery of the urethral sphincter, cytokine secretion levels and cell retention rates in the urethral tissues in SUI rats, however, there was no significant difference in 2 methods.

Microglia Influence Neurofilament Deposition in ALS iPSC-Derived Motor Neurons.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease in which upper and lower motor neuron loss is the primary phenotype, leading to muscle weakness and wasting, respiratory failure, and death. Although a portion of ALS cases are linked to one of over 50 unique genes, the vast majority of cases are sporadic in nature. However, the mechanisms underlying the motor neuron loss in either familial or sporadic ALS are not entirely clear. Here, we used induced pluripotent stem cells derived from a set of identical twin brothers discordant for ALS to assess the role of astrocytes and microglia on the expression and accumulation of neurofilament proteins in motor neurons. We found that motor neurons derived from the affected twin which exhibited increased transcript levels of all three neurofilament isoforms and increased expression of phosphorylated neurofilament puncta. We further found that treatment of the motor neurons with astrocyte-conditioned medium and microglial-conditioned medium significantly impacted neurofilament deposition. Together, these data suggest that glial-secreted factors can alter neurofilament pathology in ALS iPSC-derived motor neurons.

Meningeal Hemangiopericytoma: A Rare Intracranial Tumor—About a Case

Introduction: Hemangiopericytoma is a very rare vascular tumor, difficult to diagnose, often confused with a meningioma. Imaging is not very specific and the diagnostic certainty remains histological. Observation: We report a case of meningeal hemangiopericytoma in a 48-year-old patient with a history of active smoking and whose sister died of glioblastoma. He was brought to the emergency room for a fall, confusion, and tonic-clonic seizures. The diagnosis was suspected on imaging (CT and MRI, in front of a syndrome of right temporal intraventricular mass measured at 37 mm in diameter, centered on the choroid plexus, with perilesional vasogenic edema and posterior white commissure. There was homogeneous enhancement of the lesion after gadolinium injection. The diagnosis was confirmed by immunohistochemical study, which showed diffuse expression of STAT6 and partial labeling for CD34, without significant expression of SSTR2. Expression of neurofilament protein was also found in the brain parenchyma. Conclusion: Hemangiopericytoma is a rare tumor, characterized by its malignant potential, its high rate of recurrence and distant metastasis. The management is based on a total microsurgical resection followed by radiotherapy.

Age-related dynamics of morphological changes in the human cortex and corpus callosum: &#x0D; Microgliocytes and neurofilaments as markers of aging

Objectives to analyze the number and sum of diameters of microgliocytes in the precentral gyrus cortex and the corpus callosum of a person in the early adulthood and of an old person and to identify the patterns of neurofilaments expression in these parts of the brain.&#x0D; Material and methods. The results of a sectional study of the cerebral cortex in the precentral gyrus and corpus callosum using hematoxylin and eosin staining were analyzed, and an immunohistochemical study was performed using antibodies to neurofilaments. The dead were divided into two groups depending on their age. The group I included 23 men and 27 women in their early adulthood, aged from 23 to 28 years; the group II included 19 men and 25 women of the old age from 75 to 83 years. We examined the number of microgliocytes, the sum of their diameters, and the expression of neurofilament proteins in the tissue.&#x0D; Results. Morphometric study of autopsy material showed that in the corpus callosum, as well as in the cortex of the precentral gyrus, there was an age-related statistically significant increase in the number of microgliocytes and the sum of their diameters (p 0.001). We found that the expression of neurofilaments in the cortex of the precentral gyrus is uniform both in the early adulthood and in old age. In the corpus callosum tissue, the age-related neurodegenerative changes are more pronounced: by old age, there is a significant decrease in the expression of neurofilaments with the formation of their rarefaction sites in the areas of microgliocyte clusters.&#x0D; Conclusion. The results of morphological study of the precentral gyrus cortex and corpus callosum of two age groups testify that the number and sum of diameters of microgliocytes responsible for the utilization of spent myelin have a certain relationship with age and expression of neurofilaments in the nervous tissue.

Emergent White Matter Degeneration in the rTg-DI Rat Model of Cerebral Amyloid Angiopathy Exhibits Unique Proteomic Changes

Cerebral amyloid angiopathy (CAA), characterized by cerebral vascular amyloid accumulation, neuroinflammation, microbleeds, and white matter (WM) degeneration, is a common comorbidity in Alzheimer disease and a prominent contributor to vascular cognitive impairment and dementia. WM loss was recently reported in the corpus callosum (CC) in the rTg-DI rat model of CAA. The current study shows that the CC exhibits a much lower CAA burden compared with the adjacent cortex. Sequential Window Acquisition of All Theoretical Mass Spectra tandem mass spectrometry was used to show specific proteomic changes in the CC with emerging WM loss and compare them with the proteome of adjacent cortical tissue in rTg-DI rats. In the CC, annexin A3, heat shock protein β1, and cystatin C were elevated at 4 months (M) before WM loss and at 12M with evident WM loss. Although annexin A3 and cystatin C were also enhanced in the cortex at 12M, annexin A5 and the leukodystrophy-associated astrocyte proteins megalencephalic leukoencephalopathy with subcortical cysts 1 and GlialCAM were distinctly elevated in the CC. Pathway analysis indicated neurodegeneration of axons, reflected by reduced expression of myelin and neurofilament proteins, was common to the CC and cortex; activation of Tgf-β1 and F2/thrombin was restricted to the CC. This study provides new insights into the proteomic changes that accompany WM loss in the CC of rTg-DI rats.

Melatonin Reduces Neuroinflammation and Improves Axonal Hypomyelination by Modulating M1/M2 Microglia Polarization via JAK2-STAT3-Telomerase Pathway in Postnatal Rats Exposed to Lipopolysaccharide

Microglia activation and associated inflammation are implicated in the periventricular white matter damage (PWMD) in septic postnatal rats. This study investigated whether melatonin would mitigate inflammation and alleviate the axonal hypomyelination in the corpus callosum in septic postnatal rats. We further explored if this might be related to the modulation of microglial polarization from M1 phenotype to M2 through the JAK2/STAT3/telomerase pathway. We reported here that indeed melatonin not only can it reduce the neurobehavioral disturbances in LPS-injected rats, but it can also dampen microglia-mediated inflammation. Thus, in LPS + melatonin group, the expression of proinflammatory mediators in M1 phenotype microglia was downregulated. As opposed to this, M2 microglia were increased which was accompanied by upregulated expression of anti-inflammatory mediators along with telomerase reverse transcriptase or melatonin receptor 1(MT1). In parallel to this was decreased NG2 expression but increased expression of myelin and neurofilament proteins. Melatonin can improve hypomyelination which was confirmed by electron microscopy. In vitro in primary microglia stimulated by LPS, melatonin decreased the expression of proinflammatory mediators significantly; but it increased the expression of anti-inflammatory mediators. Additionally, the expression levels of p-JAK2 and p-STAT3 were significantly elevated in microglia after melatonin treatment. Remarkably, the effect of melatonin on LPS-treated microglia was blocked by melatonin receptor, JAK2, STAT3 and telomerase reverse transcriptase inhibitors, respectively. Taken together, it is concluded that melatonin can attenuate PWMD through shifting M1 microglia towards M2 via MT1/JAK2/STAT3/telomerase pathway. The results suggest a new therapeutic strategy whereby melatonin may be adopted to convert microglial polarization from M1 to M2 phenotype that would ultimately contribute to the attenuation of PWMD.

Tanatogenetic role of type 2 diabetes in neurosurgical patients

To define the tanatogenetic significance of 2 type diabetes in neurosurgical patients. Forty-three patients aged 22-75 years, who had died after neurosurgery, were examined. Case histories and the time course of changes in blood glucose levels were analyzed. The manifestations of macroangiopathy were taken into account at autopsy; those of microangiopathy were considered at microscopy of the internal organs and tissues; Van Gieson and periodic acid-Schiff staining was additionally applied. The expression of insulin, glucagon, synaptophysin, S100 proteins, and neurofilament protein was found in the pancreas. The number and volume of islets and an insulin-to-glucagon ratio were morphometrically estimated. Type 2 diabetes mellitus was identified in 28 (65.1%) patients: 11 (25.6%), 8 (18.6%), and 9 (20.9%) with clinically obvious, subtle, and latent types, respectively; while it was as a background underlying disease in 3 (7.0%) patients. Among the immediate causes of death, there was a preponderance of local angiogenic complications (hemorrhages, infarctions, swelling, and displacement of the brain) (64%). Indurative pancreatitis was not inferior to cardiovascular diseases among complications and concomitant diseases and was detected in most diabetic patients (n=24, or 85.7%) and in 6 (a quarter) patients with acute pancreatic necrosis, which was the immediate cause of death in 2 cases. In the places of paravasal sclerosis, there was a reduction in the terminals of nerve fibers as a manifestation of diabetic neuropathy. Type 2 diabetes mellitus was much more frequently encountered in the most critically ill and dead neurosurgical patients than in the general population and was embodied in the pattern of immediate causes of death. Diabetic neuropathy with a reduction in the terminals of nerve fibers can serve as a substantial basis not only for vascular, but also for ductal dystonia and dyskinesia, which can become an important factor in the pathogenesis of chronic pancreatitis in patients with diabetes mellitus.

Pathologic Prognostic Factors of Pineal Parenchymal Tumor of Intermediate Differentiation

Pineal parenchymal tumor of intermediate differentiation (PPTID) is an uncommon tumor of the pineal gland. Although this behaves as a grade II/III tumor, the exact clinical behavior is not well known. There is no well-established pathologic factor that can predict the behavior of PPTID. The aim of this study was to determine the pathologic prognostic factors in PPTID. All PPTID cases diagnosed between 2006 and 2016 were analyzed retrospectively. Immunohistochemistry for synaptophysin, neurofilament protein (NFP), glial fibrillar acid protein, NeuN, and Ki-67 were performed in all cases. Cases were classified arbitrarily into low grade (mitosis <4/10 hpf and Ki-67 <5%) and high grade (mitosis ≥4/10 hpf and Ki-67 ≥5%). Clinical details including follow-up information were retrieved from the patients' files. A total of 16 patients (6 low grade and 10 high grade) were included in this study. The age ranged from 2 to 55 years (average, 28.2) with a mild male preponderance (male:female, 1.67:1). All cases showed strong and diffuse positivity for synaptophysin. Focal NFP positivity was seen in 2 low-grade and 3 high-grade tumors. Only 2 cases showed focal NeuN positivity. Average Ki-67 index was 1.7% and 12.6% in low-grade and high-grade tumors, respectively. All patients with low-grade tumor were alive without recurrence. Among the patients with high-grade tumors, 2 had local recurrence, 1 had spinal metastasis, and 3 patients died. Mitosis and Ki-67 proliferation index are the most important pathologic prognostic factors in PPTID. NFP expression does not carry any prognostic significance.

Neural stem cell transplantation inhibits glial cell proliferation and P2X receptor-mediated neuropathic pain in spinal cord injury rats.

P2X4 and P2X7 receptors play an important role in neuropathic pain after spinal cord injury. Regulation of P2X4 and P2X7 receptors can obviously reduce pain hypersensitivity after injury. To investigate the role of neural stem cell transplantation on P2X receptor-mediated neuropathic pain and explore related mechanisms, a rat model of spinal cord injury was prepared using the free-falling heavy body method with spinal cord segment 10 as the center. Neural stem cells were injected into the injured spinal cord segment using a micro-syringe. Expression levels of P2X4 and P2X7 receptors, neurofilament protein, and glial fibrillary acidic protein were determined by immunohistochemistry and western blot assay. In addition, sensory function was quantitatively assessed by current perception threshold. The Basso-Beattie-Bresnahan locomotor rating scale was used to assess neuropathological pain. The results showed that 4 weeks after neural stem cell transplantation, expression of neurofilament protein in the injured segment was markedly increased, while expression of glial fibrillary acidic protein and P2X4 and P2X7 receptors was decreased. At this time point, motor and sensory functions of rats were obviously improved, and neuropathic pain was alleviated. These findings demonstrated that neural stem cell transplantation reduced overexpression of P2X4 and P2X7 receptors, activated locomotor and sensory function reconstruction, and played an important role in neuropathic pain regulation after spinal cord injury. Therefore, neural stem cell transplantation is one potential option for relieving neuropathic pain mediated by P2X receptors.

Macular peeling-induced retinal damage: clinical and histopathological evaluation after using different dyes.

To describe functional and histopathological findings after macular peeling using different dyes. Prospective, randomized, comparative, interventional, and immunohistochemical study. Forty-five eyes from 45 patients with idiopathic epiretinal membrane (ERM) underwent pars plana chromovitrectomy with ERM and inner limiting membrane (ILM) using trypan blue 0.15% + brilliant blue 0.05% + lutein 2% in group 1 (15 eyes), trypan blue 0.15% + brilliant blue 0.025% + polyethylene glycol 3350 4% in group 2 (15 eyes), and indocyanine green 0.05% in group 3 (15 eyes). We evaluated visual acuity (VA) and macular sensitivity (MS) preoperatively, 1, 3, and 6months after surgery. The expression of glial fibrillary acidic protein (GFAP) and neurofilament protein (NF) was assessed immunohistochemically on the ILMs peeled as markers of glial and neuronal cells. In group 1, both mean VA and MS were significantly better at 1 and 3months after surgery (P < 0.05), whereas no significant difference was found after 6months. GFAP and NF expression was significantly lower in group 1 (P < 0.05). The ERM/ILM peeling is thought to rip off the intraretinal tissue, based on the amounts of GFAP and NF in the specimens. The use of lutein dyes reduces iatrogenic stress to the retinal tissue and allows a faster functional recovery in the first 3months after surgery, suggesting a less iatrogenic adhesion to the retinal tissue.

Effect of Vitamin D in HN9.10e Embryonic Hippocampal Cells and in Hippocampus from MPTP-Induced Parkinson's Disease Mouse Model.

It has long been proven that neurogenesis continues in the adult brains of mammals in the dentatus gyrus of the hippocampus due to the presence of neural stem cells. Although a large number of studies have been carried out to highlight the localization of vitamin D receptor in hippocampus, the expression of vitamin D receptor in neurogenic dentatus gyrus of hippocampus in Parkinson’s disease (PD) and the molecular mechanisms triggered by vitamin D underlying the production of differentiated neurons from embryonic cells remain unknown. Thus, we performed a preclinical in vivo study by inducing PD in mice with MPTP and showed a reduction of glial fibrillary acidic protein (GFAP) and vitamin D receptor in the dentatus gyrus of hippocampus. Then, we performed an in vitro study by inducing embryonic hippocampal cell differentiation with vitamin D. Interestingly, vitamin D stimulates the expression of its receptor. Vitamin D receptor is a transcription factor that probably is responsible for the upregulation of microtubule associated protein 2 and neurofilament heavy polypeptide genes. The latter increases heavy neurofilament protein expression, essential for neurofilament growth. Notably N-cadherin, implicated in activity for dendritic outgrowth, is upregulated by vitamin D.

Sustained-release of FGF-2 from a hybrid hydrogel of heparin-poloxamer and decellular matrix promotes the neuroprotective effects of proteins after spinal injury.

IntroductionThe short lifetime of protein-based therapies has largely limited their therapeutic efficacy in injured nervous post-spinal cord injury (post-SCI).MethodsIn this study, an affinity-based hydrogel delivery system provided sustained-release of proteins, thereby extending the efficacy of such therapies. The affinity-based hydrogel was constructed using a novel polymer, heparin-poloxamer (HP), as a temperature-sensitive bulk matrix and decellular spinal cord extracellular matrix (dscECM) as an affinity depot of drug. By tuning the concentration of HP in formulation, the cold ternary fibroblast growth factor-2 (FGF2)-dscECM-HP solution could rapidly gelatinize into a hydrogel at body temperature. Due to the strong affinity for FGF2, hybrid FGF2-dscECM-HP hydrogel enabled sustained-release of encapsulated FGF2 over an extended period in vitro.ResultsCompared to free FGF2, it was observed that both neuron functions and tissue morphology after SCI were clearly recovered in rats treated with FGF2-dscECM-HP hydrogel. Moreover, the expression of neurofilament protein and the density of axons were increased after treatment with hybrid FGF2-dscECM-HP. In addition, the neuroprotective effects of FGF2-dscECM-HP were related to inhibition of chronic endoplasmic reticulum stress-induced apoptosis.ConclusionThe results revealed that a hybrid hydrogel system may be a potential carrier to deliver macromolecular proteins to the injured site and enhance the therapeutic effects of proteins.

Huangqin flavonoid extraction for spinal cord injury in a rat model.

Flavonoids from Huangqin (dried roots of Scutellaria baicalensis Georgi) have anti-inflammatory effects, and are considered useful for treatment of spinal cord injury. To verify this hypothesis, the T9-10 spinal cord segments of rats were damaged using Allen's method to establish a rat spinal cord injury model. Before model establishment, Huangqin flavonoid extraction (12.5 g/kg) was administered intragastrically for 1 week until 28 days after model establishment. Methylprednisolone (30 mg/kg) was injected into the tail vein at 30 minutes after model establishment as a positive control. Basso, Beattie, and Bresnahan locomotor scale scores were used to assess hind limb motor function. Hematoxylin-eosin staining was used to detect pathological changes in the injured spinal cord. Immunofluorescence and western blot assays were performed to measure immunoreactivity and expression levels of brain-derived neurotrophic factor, neuronal marker neurofilament protein, microglial marker CD11b and astrocyte marker glial fibrillary acidic protein in the injured spinal cord. Huangqin flavonoid extraction markedly reduced spinal cord hematoma, inflammatory cell infiltration and cavities and scars, and increased the Basso, Beattie, and Bresnahan locomotor scale scores; these effects were identical to those of methylprednisolone. Huangqin flavonoid extraction also increased immunoreactivity and expression levels of brain-derived neurotrophic factor and neurofilament protein, and reduced immunoreactivity and expression levels of CD11b and glial fibrillary acidic protein, in the injured spinal cord. Overall, these data suggest that Huangqin flavonoid extraction can promote recovery of spinal cord injury by inducing brain-derived neurotrophic factor and neurofilament protein expression, reducing microglia activation and regulating reactive astrocytes.

Modified insulin-like growth factor 1 containing collagen-binding domain for nerve regeneration.

Insulin-like growth factor 1 (IGF-1) is a potential nutrient for nerve repair. However, it is impractical as a therapy because of its limited half-life, rapid clearance, and limited target specificity. To achieve targeted and long-lasting treatment, we investigated the addition of a binding structure by fusing a collagen-binding domain to IGF-1. After confirming its affinity for collagen, the biological activity of this construct was examined by measuring cell proliferation after transfection into PC12 and Schwann cells using a 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide assay. Immunofluorescence staining was conducted to detect neurofilament and microtubule-associated protein 2 expression, while real time-polymerase chain reaction was utilized to determine IGF-1 receptor and nerve growth factor mRNA expression. Our results demonstrate a significant increase in collagen-binding activity of the recombinant protein compared with IGF-1. Moreover, the recombinant protein promoted proliferation of PC12 and Schwann cells, and increased the expression of neurofilament and microtubule-associated protein 2. Importantly, the recombinant protein also stimulated sustained expression of IGF-1 receptor and nerve growth factor mRNA for days. These results show that the recombinant protein achieved the goal of targeting and long-lasting treatment, and thus could become a clinically used factor for promoting nerve regeneration with a prolonged therapeutic effect.

Expression of RGMb in brain tissue of MCAO rats and its relationship with axonal regeneration

ObjectiveTo investigate the changes in the expression of repulsive guidance molecule b (RGMb) in brain tissue of rats with ischemic cerebral infarction and determine its relationship with axonal regeneration, synapse remodeling and magnetic resonance imaging (MRI) parameters with magnetic resonance diffusion tensor imaging as the dynamic continuous monitoring method in vivo, so as to explore the pathophysiological mechanism of the occurrence, development and prognosis of cerebral infarction. MethodsNinety Sprague-Dawley (SD) rats were randomly divided into six groups, namely control group, middle cerebral artery occlusion (MCAO) 12-h group, MCAO 24-h group, MCAO 48-h group, MCAO 7-day group and MCAO 10-day group, each of 15 animals. Rats were examined by head MRI at corresponding time points, followed by measurement of apparent diffusion coefficient (ADC) and fractional anisotropy (FA) values. Subsequently, brain tissues were taken to detect the expression of RGMb, axonal regeneration and synapse remodeling. ResultsAfter infarction, ADC and FA values of the infarcted core area were lower in rats of each group compared to those in rats of normal group (P<0.05), which was lowest at 12h. The positive expressions of RGMb and synaptophysin were continuously increased from the 12th hour after operation, which reached a peak at 48h; while axonas was gradually reduced after operation in each group, which revealed the most obvious damage in the MCAO 24-h group. The protein expression of RGMb was negatively correlated with MRI parameters and axon growth. ConclusionAfter supratentorial cerebral infarction, the expressions of RGMb and synaptophysin were up-regulated in rats, neurofilament protein (NF-200) expression was decreased, and MRI parameters (ADC and FA values) were reduced, indicating that RGMb protein may be involved in the regeneration and remodeling of axons and synapses, and exert an important role in pathophysiological processes such as nerve regeneration disturbance and neuron apoptosis after cerebral ischemia injury. In vivo MRI can be a noninvasive technique to monitor the areas of cerebral infarction and the recovery of neurological function.

Altered neurofilament protein expression in the lateral vestibular nucleus in Parkinson's disease.

A major cause of morbidity in Parkinson's disease (PD) is postural instability. The neuropathology underlying postural instability is unknown. Postural control is mediated by Deiters' neurons of the lateral vestibular nucleus (LVN), which are the brainstem origin of descending vestibulospinal reflexes. Deiters' neurons express the cytostructural protein, non-phosphorylated neurofilament protein (NPNFP). In PD, reduced expression of NPNFP in substantia nigra (SN) neurons is believed to contribute to dysfunction. It was the aim of this study to determine if there is altered expression of NPNFP in the LVN in PD. We immunolabeled NPNFP in brainstem sections of six aged controls (mean age 92yo) and six PD donors (mean age 83yo). Our results show there was a ~50% reduction in NPNFP-positive Deiters' neurons compared to controls (13±2.0/section vs 25.7±3.0/section; p<0.01, repeated measures ANOVA). In contrast, there was no difference in NPNFP-positive counts in the facial nucleus between control and PD. The normalized intensity of NPNFP labeling in LVN was also reduced in PD (0.87±0.05 vs 1.09±0.03; p<0.01). There was a 35% concurrent reduction in NPNFP-positive neuropil in PD relative to controls (p<0.01). We also show there was an 84% increase (p<0.05) in somatic lipofuscin in PD patients compared to control. Lipofuscin aggregation has been shown to increase not only with age but also with neurodegeneration. Furthermore, decreased NPNFP intensity was strongly correlated with increasing lipofuscin autofluorescence across all cases (R 2=0.81, p<0.01). These results show two alterations in cellular content with PD, reduced expression and intensity of NPNFP and increased lipofuscin aggregation in Deiter's neurons. These changes may contribute to degeneration of postural reflexes observed in PD.

Role of the GM1 ganglioside oligosaccharide portion in the TrkA-dependent neurite sprouting in neuroblastoma cells.

GM1 ganglioside (II3 NeuAc-Gg4 Cer) is known to promote neurite formation in neuroblastoma cells by activating TrkA-MAPK pathway. The molecular mechanism by which GM1 is involved in the neurodifferentiation process is still unknown, however, invitro and invivo evidences have suggested that the oligosaccharide portion of this ganglioside could be involved. Here, we report that, similarly to the entire GM1 molecule, its oligosaccharide II3 NeuAc-Gg4, rather than its ceramide (Cer) portion is responsible for the neurodifferentiation process by augmenting neurite elongation and increasing the neurofilament protein expression in murine neuroblastoma cells, Neuro2a. Conversely, asialo-GM1, GM2 and GM3 oligosaccharides are not effective in neurite elongation on Neuro2a cells, whereas the effect exerted by the Fuc-GM1 oligosaccharide (IV2 αFucII3 Neu5Ac-Gg4 ) is similar to that exerted by GM1 oligosaccharide. The neurotrophic properties of GM1 oligosaccharide are exerted by activating the TrkA receptor and the following phosphorylation cascade. By photolabeling experiments performed with a nitrophenylazide containing GM1 oligosaccharide, labeled with tritium, we showed a direct interaction between the GM1 oligosaccharide and the extracellular domain of TrkA receptor. Moreover, molecular docking analyses confirmed that GM1 oligosaccharide binds the TrkA-nerve growth factor complex leading to a binding free energy of approx. -11.5kcal/mol, acting as a bridge able to increase and stabilize the TrkA-nerve growth factor molecular interactions.

Impairment of toll-like receptors 2 and 4 leads to compensatory mechanisms after sciatic nerve axotomy.

BackgroundPeripheral nerve injury results in retrograde cell body-related changes in the spinal motoneurons that will contribute to the regenerative response of their axons. Successful functional recovery also depends on molecular events mediated by innate immune response during Wallerian degeneration in the nerve microenvironment. A previous study in our lab demonstrated that TLR 2 and 4 develop opposite effects on synaptic stability in the spinal cord after peripheral nerve injury. Therefore, we suggested that the better preservation of spinal cord microenvironment would positively influence distal axonal regrowth. In this context, the present work aimed to investigate the influence of TLR2 and TLR4 on regeneration and functional recovery after peripheral nerve injury.MethodsEighty-eight mice were anesthetized and subjected to unilateral sciatic nerve crush (C3H/HeJ, n = 22, C3H/HePas, n = 22; C57Bl6/J, n = 22 and TLR2−/−, n = 22). After the appropriate survival times (3, 7, 14 days, and 5 weeks), all mice were killed and the sciatic nerves and tibialis cranialis muscles were processed for immunohistochemistry and transmission electron microscopy (TEM). Gait analysis, after sciatic nerve crushing, was performed in another set of mice (minimum of n = 8 per group), by using the walking track test (CatWalk system).ResultsTLR4 mutant mice presented greater functional recovery as well as an enhanced p75NTR and neurofilament protein expression as compared to the wild-type strain. Moreover, the better functional recovery in mutant mice was correlated to a greater number of nerve terminal sprouts. Knockout mice for TLR2 exhibited 30 % greater number of degenerated axons in the distal stump of the sciatic nerve and a decreased p75NTR and neurofilament protein expression compared to the wild type. However, the absence of TLR2 receptor did not influence the overall functional recovery. End-point equivalent functional recovery in transgenic mice may be a result of enhanced axonal diameter found at 2 weeks after lesion.ConclusionsAltogether, the present results indicate that the lack of TLR2 or the absence of functional TLR4 does affect the nerve regeneration process; however, such changes are minimized through different compensatory mechanisms, resulting in similar motor function recovery, as compared to wild-type mice. These findings contribute to the concept that innate immune-related molecules influence peripheral nerve regeneration by concurrently participating in processes taking place both at the CNS and PNS.Electronic supplementary materialThe online version of this article (doi:10.1186/s12974-016-0579-6) contains supplementary material, which is available to authorized users.