Sweat gland nerve fiber density and association with sudomotor function, symptoms, and risk factors in adolescents with type 1 diabetes.

PURPOSE: To quantify sweat gland nerve fiber density in adolescents with diabetes. Additionally, to investigate associations between sudomotor innervation, sweat responses, and possible risk factors for sudomotor neuropathy. METHODS: Cross-sectional study where 60 adolescents with type 1 diabetes (duration > 5 years) and 23 control subjects were included. Clinical data, quantitative sudomotor axon reflex test, and skin biopsies were obtained. Skin tissue was immunostained and imaged by confocal microscopy. Quantification of the sweat gland volume and three-dimensional reconstruction of the nerve fibers was performed using a design-unbiased technique. RESULTS: Adolescents with diabetes had a significant reduction of maximum and mean values of nerve fiber length and nerve fiber density in sweat glands compared to controls (p values < 0.05). No association between nerve fiber density and sweat responses was found (p = 0.21). In cases with reduced sweat gland nerve fiber length, nerve fiber density, and volume, the sweat response was reduced or absent. Height, systolic blood pressure, time in hypoglycemia, and total daily and basal/total insulin dose were positively correlated to sweat response, while low-density lipoprotein, and HbA1c were negatively correlated with sweat response (p values < 0.05). Other microvascular complications and high cholesterol levels increased the relative risk for reduced sweat gland nerve fiber density. CONCLUSION: Our findings of reduced sweat gland innervation in a selected group of adolescents add new knowledge about the structural changes that occur in autonomic nerves due to diabetes. Evaluating both the sweat gland innervation and sweat gland volume was important for understanding the association with sweat responses. Further research is needed to understand its clinical relevance.

Neurofilament light chain in spinal fluid and plasma in multiple system atrophy: a prospective, longitudinal biomarker study.

PURPOSE: There is a critical need for reliable diagnostic biomarkers as well as surrogate markers of disease progression in multiple system atrophy (MSA). Neurofilament light chain (NfL) has been reported to potentially meet those needs. We therefore sought to explore the value of NfL in plasma (NfL-p) in contrast to cerebrospinal fluid (NfL-c) as a diagnostic marker of MSA, and to assess NfL-p and NfL-c as markers of clinical disease progression. METHODS: Well-characterized patients with early MSA (n = 32), Parkinson's disease (PD; n = 21), and matched controls (CON; n = 15) were enrolled in a prospective, longitudinal study of synucleinopathies with serial annual evaluations. NfL was measured using a high-sensitivity immunoassay, and findings were assessed by disease category and relationship with clinical measures of disease progression. RESULTS: Measurements of NfL-c were highly reproducible across immunoassay platforms (Pearson, r = 0.99), while correlation between NfL-c and -p was only moderate (r = 0.66). NfL was significantly higher in MSA compared with CON and PD; the separation was essentially perfect for NfL-c, but there was overlap, particularly with PD, for NfL-p. While clinical measures of disease severity progressively increased over time, NfL-c and -p remained at stable elevated levels within subjects across serial measurements. Neither change in NfL nor baseline NfL were significantly associated with changes in clinical markers of disease severity. CONCLUSIONS: These findings confirm NfL-c as a faithful diagnostic marker of MSA, while NfL-p showed less robust diagnostic value. The significant NfL elevation in MSA was found to be remarkably stable over time and was not predictive of clinical disease progression.

Neurofilament Light Chain in Spinal Fluid and Plasma in Multiple System Atrophy - A Prospective, Longitudinal Biomarker Study.

Purpose There is a critical need for reliable diagnostic biomarkers as well as surrogate markers of disease progression in multiple system atrophy (MSA). Neurofilament light chain (NfL) has been reported to potentially meet those needs. We therefore sought to explore the value of NfL in plasma (NfL-p) in contrast to CSF (NfL-c) as diagnostic marker of MSA, and to assess NfL-p and NfL-c as markers of clinical disease progression. Methods Well-characterized patients with early MSA (n=32), Parkinson's disease (PD, n=21), and matched controls (CON, n=15) were enrolled in a prospective, longitudinal study of synucleinopathies with serial annual evaluations. NfL was measured using a high sensitivity immunoassay, and findings were assessed by disease category and relationship with clinical measures of disease progression. Results Measurements of NfL-c were highly reproducible across immunoassay platforms (Pearson,r=0.99), while correlation between NfL-c and -p was only moderate (r=0.66). NfL was significantly higher in MSA compared to CON and PD; the separation was essentially perfect for NfL-c, but there was overlap, particularly with PD, for NfL-p. While clinical measures of disease severity progressively increased over time, NfL-c and -p remained at stable elevated levels within subjects across serial measurements. Neither change in NfL nor baseline NfL were significantly associated with changes in clinical markers of disease severity. Conclusions These findings confirm NfL-c as faithful diagnostic marker of MSA, while NfL-p showed less robust diagnostic value. The significant NfL elevation in MSA was found to be remarkably stable over time and was not predictive of clinical disease progression.

Alpha-Synuclein Oligomers and Neurofilament Light Chain Predict Phenoconversion of Pure Autonomic Failure.

OBJECTIVE: To explore the role of alpha-synuclein (αSyn) oligomers and neurofilament light chain (NfL) in cerebrospinal fluid (CSF) of patients with pure autonomic failure (PAF) as markers of future phenoconversion to multiple system atrophy (MSA). METHODS: Well-characterized patients with PAF (n = 32) were enrolled between June 2016 and February 2019 at Mayo Clinic Rochester and followed prospectively with annual visits to determine future phenoconversion to MSA, Parkinson's disease (PD), or dementia with Lewy bodies (DLB). ELISA was utilized to measure NfL and protein misfolding cyclic amplification (PMCA) to detect αSyn oligomers in CSF collected at baseline. RESULTS: Patients were followed for a median of 3.9 years. Five patients converted to MSA, 2 to PD, and 2 to DLB. NfL at baseline was elevated only in patients who later developed MSA, perfectly separating those from future PD and DLB converters as well as non-converters. ASyn-PMCA was positive in all but two cases (94%). The PMCA reaction was markedly different in five samples with maximum fluorescence and reaction kinetics previously described in MSA patients; all of these patients later developed MSA. INTERPRETATION: αSyn-PMCA is almost invariably positive in the CSF of patients with PAF establishing this condition as α-synucleinopathy. Both NfL and the magnitude and reaction kinetics of αSyn PMCA faithfully predict which PAF patients will eventually phenoconvert to MSA. This finding has important implications not only for prognostication, but also for future trials of disease modifying therapies, allowing for differentiation of MSA from Lewy body synucleinopathies before motor symptoms develop. ANN NEUROL 2021;89:1212-1220.

Promoting Neuronal Outgrowth Using Ridged Scaffolds Coated with Extracellular Matrix Proteins.

Spinal cord injury (SCI) results in cell death, demyelination, and axonal loss. The spinal cord has a limited ability to regenerate, and current clinical therapies for SCI are not effective in helping promote neurologic recovery. We have developed a novel scaffold biomaterial that is fabricated from the biodegradable hydrogel oligo(poly(ethylene glycol)fumarate) (OPF). We have previously shown that positively charged OPF scaffolds (OPF+) in an open spaced, multichannel design can be loaded with Schwann cells to support axonal generation and functional recovery following SCI. We have now developed a hybrid OPF+ biomaterial that increases the surface area available for cell attachment and that contains an aligned microarchitecture and extracellular matrix (ECM) proteins to better support axonal regeneration. OPF+ was fabricated as 0.08 mm thick sheets containing 100 µm high polymer ridges that self-assemble into a spiral shape when hydrated. Laminin, fibronectin, or collagen I coating promoted neuron attachment and axonal outgrowth on the scaffold surface. In addition, the ridges aligned axons in a longitudinal bipolar orientation. Decreasing the space between the ridges increased the number of cells and neurites aligned in the direction of the ridge. Schwann cells seeded on laminin coated OPF+ sheets aligned along the ridges over a 6-day period and could myelinate dorsal root ganglion neurons over 4 weeks. This novel scaffold design, with closer spaced ridges and Schwann cells, is a novel biomaterial construct to promote regeneration after SCI.

Loss of putative GABAergic neurons in the ventrolateral medulla in multiple system atrophy.

STUDY OBJECTIVES: Multiple system atrophy (MSA) is associated with disturbances in cardiovascular, sleep and respiratory control. The lateral paragigantocellular nucleus (LPGi) in the ventrolateral medulla (VLM) contains GABAergic neurons that participate in control of rapid eye movement (REM) sleep and cardiovagal responses. We sought to determine whether there was loss of putative GABAergic neurons in the LPGi and adjacent regions in MSA. METHODS: Sections of the medulla were processed for GAD65/67 immunoreactivity in eight subjects with clinical and neuropathological diagnosis of MSA and in six control subjects. These putative GABAergic LPGi neurons were mapped based on their relationship to adjacent monoaminergic VLM groups. RESULTS: There were markedly decreased numbers of GAD-immunoreactive neurons in the LPGi and adjacent VLM regions in MSA. CONCLUSIONS: There is loss of GABAergic neurons in the VLM, including the LPGi in patients with MSA. Whereas these findings provide a possible mechanistic substrate, given the few cases included, further studies are necessary to determine whether they contribute to REM sleep-related cardiovagal and possibly respiratory dysregulation in MSA.

Defining Spatial Relationships Between Spinal Cord Axons and Blood Vessels in Hydrogel Scaffolds.

Positively charged oligo(poly(ethylene glycol) fumarate) (OPF+) hydrogel scaffolds, implanted into a complete transection spinal cord injury (SCI), facilitate a permissive regenerative environment and provide a platform for controlled observation of repair mechanisms. Axonal regeneration after SCI is critically dependent upon nutrients and oxygen from a newly formed blood supply. Our objective was to investigate fundamental characteristics of revascularization in association with the ingrowth of axons into hydrogel scaffolds, thereby defining spatial relationships between axons and the neovasculature. A novel combination of stereologic estimates and precision image analysis techniques quantitate neurovascular regeneration in rats. Multichannel hydrogel scaffolds containing Matrigel-only (MG), Schwann cells (SCs), or SCs with rapamycin-eluting poly(lactic co-glycolic acid) microspheres (RAPA) were implanted for 6 weeks following complete spinal cord transection. Image analysis of 72 scaffold channels identified a total of 2494 myelinated and 4173 unmyelinated axons at 10 µm circumferential intervals centered around 708 individual blood vessel profiles. Blood vessel number, density, volume, diameter, intervessel distances, total vessel surface and cross-sectional areas, and radial diffusion distances were compared. Axon number and density, blood vessel surface area, and vessel cross-sectional areas in the SC group exceeded that in the MG and RAPA groups. Individual axons were concentrated within a concentric radius of 200-250 µm from blood vessel walls, in Gaussian distributions, which identified a peak axonal number (Mean Peak Amplitude) corresponding to defined distances (Mean Peak Distance) from each vessel, the highest concentrations of axons were relatively excluded from a 25-30 µm zone immediately adjacent to the vessel, and from vessel distances >150 µm. Higher axonal densities correlated with smaller vessel cross-sectional areas. A statistical spatial algorithm was used to generate cumulative distribution F- and G-functions of axonal distribution in the reference channel space. Axons located around blood vessels were definitively organized as clusters and were not randomly distributed. A scoring system stratifies 5 direct measurements and 12 derivative parameters influencing regeneration outcomes. By providing methods to quantify the axonal-vessel relationships, these results may refine spinal cord tissue engineering strategies to optimize the regeneration of complete neurovascular bundles in their relevant spatial relationships after SCI. Impact statement Vascular disruption and impaired neovascularization contribute critically to the poor regenerative capacity of the spinal cord after injury. In this study, hydrogel scaffolds provide a detailed model system to investigate the regeneration of spinal cord axons as they directly associate with individual blood vessels, using novel methods to define their spatial relationships and the physiologic implications of that organization. These results refine future tissue engineering strategies for spinal cord repair to optimize the re-development of complete neurovascular bundles in their relevant spatial architectures.

Identification of Multicolor Fluorescent Probes for Heterogeneous Aß Deposits in Alzheimer's Disease.

Accumulation of amyloid-beta (Aß) into amyloid plaques and hyperphosphorylated tau into neurofibrillary tangles (NFTs) are pathological hallmarks of Alzheimer's disease (AD). There is a significant intra- and inter-individual variability in the morphology and conformation of Aß aggregates, which may account in part for the extensive clinical and pathophysiological heterogeneity observed in AD. In this study, we sought to identify an array of fluorescent dyes to specifically probe Aß aggregates, in an effort to address their diversity. We screened a small library of fluorescent probes and identified three benzothiazole-coumarin derivatives that stained both vascular and parenchymal Aß deposits in AD brain sections. The set of these three dyes allowed the visualization of Aß deposits in three different colors (blue, green and far-red). Importantly, two of these dyes specifically stained Aß deposits with no apparent staining of hyperphosphorylated tau or α-synuclein deposits. Furthermore, this set of dyes demonstrated differential interactions with distinct types of Aß deposits present in the same subject. Aß aggregate-specific dyes identified in this study have the potential to be further developed into Aß imaging probes for the diagnosis of AD. In addition, the far-red dye we identified in this study may serve as an imaging probe for small animal imaging of Aß pathology. Finally, these dyes in combination may help us advance our understanding of the relation between the various Aß deposits and the clinical diversity observed in AD.

Alpha-Synuclein Oligomers and Neurofilament Light Chain in Spinal Fluid Differentiate Multiple System Atrophy from Lewy Body Synucleinopathies.

OBJECTIVE: To explore the role of alpha-synuclein (αSyn) oligomers and neurofilament light chain (NFL) in cerebrospinal fluid (CSF) as markers of early multiple system atrophy (MSA) and to contrast findings with Lewy body synucleinopathies. METHODS: In a discovery cohort of well-characterized early MSA patients (n = 24) and matched healthy controls (CON, n = 14), we utilized enzyme-linked immunosorbent assay to measure NFL and protein misfolding cyclic amplification (PMCA) to detect αSyn oligomers in CSF. We confirmed findings in a separate prospectively enrolled cohort of patients with early MSA (n = 38), Parkinson disease (PD, n = 16), and dementia with Lewy bodies (DLB, n = 13), and CON subjects (n = 15). RESULTS: In the discovery cohort, NFL was markedly elevated in MSA patients, with perfect separation from CON. αSyn-PMCA was nonreactive in all CON, whereas all MSA samples were positive. In the confirmatory cohort, NFL again perfectly separated MSA from CON, and was significantly lower in PD and DLB compared to MSA. PMCA was again nonreactive in all CON, and positive in all but 2 MSA cases. All PD and all but 2 DLB samples were also positive for αSyn aggregates but with markedly different reaction kinetics from MSA; aggregation occurred later, but maximum fluorescence was higher, allowing for perfect separation of reactive samples between MSA and Lewy body synucleinopathies. INTERPRETATION: NFL and αSyn oligomers in CSF faithfully differentiate early MSA not only from CON but also from Lewy body synucleinopathies. The findings support the role of these markers as diagnostic biomarkers, and have important implications for understanding pathophysiologic mechanisms underlying the synucleinopathies. ANN NEUROL 2020;88:503-512.

Discriminating α-synuclein strains in Parkinson's disease and multiple system atrophy.

Synucleinopathies are neurodegenerative diseases that are associated with the misfolding and aggregation of α-synuclein, including Parkinson's disease, dementia with Lewy bodies and multiple system atrophy1. Clinically, it is challenging to differentiate Parkinson's disease and multiple system atrophy, especially at the early stages of disease2. Aggregates of α-synuclein in distinct synucleinopathies have been proposed to represent different conformational strains of α-synuclein that can self-propagate and spread from cell to cell3-6. Protein misfolding cyclic amplification (PMCA) is a technique that has previously been used to detect α-synuclein aggregates in samples of cerebrospinal fluid with high sensitivity and specificity7,8. Here we show that the α-synuclein-PMCA assay can discriminate between samples of cerebrospinal fluid from patients diagnosed with Parkinson's disease and samples from patients with multiple system atrophy, with an overall sensitivity of 95.4%. We used a combination of biochemical, biophysical and biological methods to analyse the product of α-synuclein-PMCA, and found that the characteristics of the α-synuclein aggregates in the cerebrospinal fluid could be used to readily distinguish between Parkinson's disease and multiple system atrophy. We also found that the properties of aggregates that were amplified from the cerebrospinal fluid were similar to those of aggregates that were amplified from the brain. These findings suggest that α-synuclein aggregates that are associated with Parkinson's disease and multiple system atrophy correspond to different conformational strains of α-synuclein, which can be amplified and detected by α-synuclein-PMCA. Our results may help to improve our understanding of the mechanism of α-synuclein misfolding and the structures of the aggregates that are implicated in different synucleinopathies, and may also enable the development of a biochemical assay to discriminate between Parkinson's disease and multiple system atrophy.

Intrathecal administration of autologous mesenchymal stem cells in multiple system atrophy.

OBJECTIVE: This phase I/II study sought to explore intrathecal administration of mesenchymal stem cells (MSCs) as therapeutic approach to multiple system atrophy (MSA). METHODS: Utilizing a dose-escalation design, we delivered between 10 and 200 million adipose-derived autologous MSCs intrathecally to patients with early MSA. Patients were closely followed with clinical, laboratory, and imaging surveillance. Primary endpoints were frequency and type of adverse events; key secondary endpoint was the rate of disease progression assessed by the Unified MSA Rating Scale (UMSARS). RESULTS: Twenty-four patients received treatment. There were no attributable serious adverse events, and injections were generally well-tolerated. At the highest dose tier, 3 of 4 patients developed low back/posterior leg pain, associated with thickening/enhancement of lumbar nerve roots. Although there were no associated neurologic deficits, we decided that dose-limiting toxicity was reached. A total of 6 of 12 patients in the medium dose tier developed similar, but milder and transient discomfort. Rate of progression (UMSARS total) was markedly lower compared to a matched historical control group (0.40 ± 0.59 vs 1.44 ± 1.42 points/month, p = 0.004) with an apparent dose-dependent effect. CONCLUSIONS: Intrathecal MSC administration in MSA is safe and well-tolerated but can be associated with a painful implantation response at high doses. Compelling dose-dependent efficacy signals are the basis for a planned placebo-controlled trial. CLASSIFICATION OF EVIDENCE: This phase I/II study provides Class IV evidence that for patients with early MSA, intrathecal MSC administration is safe, may result in a painful implantation response at high doses, and is associated with dose-dependent efficacy signals.

Refined Quantitation of Sweat Gland Innervation.

Skin biopsies have gained increasing popularity as a tool to evaluate disorders affecting small nerve fibers. While reports on sweat gland nerve fiber density (SGNFD) to quantitate sudomotor innervation have been promising, methodologies vary significantly. Although conventional stereology is commonly used, no standard technique has been established. We sought to develop an accurate and reproducible technique to quantify SGNFD. Skin punch biopsies from healthy individuals were cut and stained. Images of sweat glands (SGs) were acquired using confocal and widefield microscopes, and optimized using deconvolution. Nerve fibers were reconstructed and nerve fiber length (NFL) was quantified using three-dimensional (3D) automated software. SGNFD was obtained by dividing NFL by SG volume. SGNFD was also assessed using stereology for comparison. Ninety-two SGs from 10 healthy subjects were analyzed by independent observers. Using confocal microscopy, the software reliably traced nerve fibers. In contrast, rendering of nerve fibers was inferior using widefield microscopy. Interobserver reliability was suboptimal using widefield images compared to confocal (ICC = 0.82 vs ICC = 0.98). Correlation between 3D-reconstruction and stereology was poor (ICC = 0.38). The newly developed technique of SGNFD quantitation using 3D reconstruction of SG innervation with confocal microscopy reliably traces nerve fibers, shows outstanding reproducibility, is almost completely unbiased, and superior to conventional stereology methods.

Combinatorial tissue engineering partially restores function after spinal cord injury.

Hydrogel scaffolds provide a beneficial microenvironment in transected rat spinal cord. A combinatorial biomaterials-based strategy provided a microenvironment that facilitated regeneration while reducing foreign body reaction to the three-dimensional spinal cord construct. We used poly lactic-co-glycolic acid microspheres to provide sustained release of rapamycin from Schwann cell (SC)-loaded, positively charged oligo-polyethylene glycol fumarate scaffolds. The biological activity and dose-release characteristics of rapamycin from microspheres alone and from microspheres embedded in the scaffold were determined in vitro. Three dose formulations of rapamycin were compared with controls in 53 rats. We observed a dose-dependent reduction in the fibrotic reaction to the scaffold and improved functional recovery over 6 weeks. Recovery was replicated in a second cohort of 28 animals that included retransection injury. Immunohistochemical and stereological analysis demonstrated that blood vessel number, surface area, vessel diameter, basement membrane collagen, and microvessel phenotype within the regenerated tissue was dependent on the presence of SCs and rapamycin. TRITC-dextran injection demonstrated enhanced perfusion into scaffold channels. Rapamycin also increased the number of descending regenerated axons, as assessed by Fast Blue retrograde axonal tracing. These results demonstrate that normalization of the neovasculature was associated with enhanced axonal regeneration and improved function after spinal cord transection.

Medullary neuronal loss is not associated with α-synuclein burden in multiple system atrophy.

BACKGROUND: Accumulation of α-synuclein in multiple system atrophy (MSA) affects medullary autonomic and respiratory control areas, including the rostral ventrolateral medulla and raphe nuclei. Relative neuronal vulnerability and its relationship to α-synuclein accumulation in these areas are unknown. The aim of this study was to determine the extent of loss of adrenergic neurons in the rostral ventrolateral medulla and serotonergic neurons in the ventrolateral medulla and raphe nuclei and its relationship with α-synuclein accumulation. METHODS: Medullary sections from 7 MSA and 6 control subjects were processed for tyrosine hydroxylase, tryptophan hydroxylase, and α-synuclein immunoreactivity. Neuronal counts were performed stereologically, whereas α-synuclein burden in oligodendrocytes and neurons was quantified using object detection density (area/mm2). RESULTS: All MSA cases had orthostatic hypotension; 5 had laryngeal stridor. There was marked neuronal loss in the rostral ventrolateral medulla and medullary raphe in all cases. Most severely affected were tyrosine hydroxylase ventrolateral medulla (C1) neurons (83% reduction), followed by tryptophan hydroxylase neurons in the ventrolateral medulla (70%), raphe obscurus (56%), pallidus (57%), and magnus (47%). α-Synuclein accumulation occurred predominantly as glial cytoplasmic inclusions with rare α-synuclein accumulation occurring within the remaining neurons. Density of α-synuclein did not correlate with neuronal loss in any of the areas analyzed, and there was no correlation between α-synuclein density and disease duration for any regions of interest. CONCLUSIONS: These findings indicate that in MSA adrenergic neurons are more susceptible than serotonergic neurons in the medulla. Further, loss of medullary monoaminergic neurons may progress independently from α-synuclein accumulation in MSA. © 2016 International Parkinson and Movement Disorder Society.

Histaminergic tuberomammillary neuron loss in multiple system atrophy and dementia with Lewy bodies.

BACKGROUND: Histaminergic neurons of the tuberomammillary nucleus are critical for maintenance of wakefulness and participate in basal ganglia modulation and sympathoexcitation. All of these functions are affected in multiple system atrophy and dementia with Lewy bodies. OBJECTIVES: To determine the involvement of histaminergic neurons in these synucleinopathies. METHODS: Immunocytochemical staining for histidine decarboxylase and stereological cell counts in the posterolateral hypothalamus from six cases with multiple system atrophy, six with dementia with Lewy bodies, and five healthy controls. RESULTS: Compared with healthy controls, significant reduction of histaminergic neurons was found both in multiple system atrophy (total estimated cell count, 94,677 ± 5,521 vs. 169,579 ± 12,443; P < 0.001) and in dementia with Lewy bodies (91,110 ± 13,021 vs. 169,579 ± 12,443; P < 0.001). CONCLUSIONS: Loss of tuberomammillary histaminergic neurons could potentially contribute to motor, sleep, and autonomic manifestations of both multiple system atrophy and dementia with Lewy bodies.

Expanding the spectrum of neuronal pathology in multiple system atrophy.

Multiple system atrophy is a sporadic alpha-synucleinopathy that typically affects patients in their sixth decade of life and beyond. The defining clinical features of the disease include progressive autonomic failure, parkinsonism, and cerebellar ataxia leading to significant disability. Pathologically, multiple system atrophy is characterized by glial cytoplasmic inclusions containing filamentous alpha-synuclein. Neuronal inclusions also have been reported but remain less well defined. This study aimed to further define the spectrum of neuronal pathology in 35 patients with multiple system atrophy (20 male, 15 female; mean age at death 64.7 years; median disease duration 6.5 years, range 2.2 to 15.6 years). The morphologic type, topography, and frequencies of neuronal inclusions, including globular cytoplasmic (Lewy body-like) neuronal inclusions, were determined across a wide spectrum of brain regions. A correlation matrix of pathologic severity also was calculated between distinct anatomic regions of involvement (striatum, substantia nigra, olivary and pontine nuclei, hippocampus, forebrain and thalamus, anterior cingulate and neocortex, and white matter of cerebrum, cerebellum, and corpus callosum). The major finding was the identification of widespread neuronal inclusions in the majority of patients, not only in typical disease-associated regions (striatum, substantia nigra), but also within anterior cingulate cortex, amygdala, entorhinal cortex, basal forebrain and hypothalamus. Neuronal inclusion pathology appeared to follow a hierarchy of region-specific susceptibility, independent of the clinical phenotype, and the severity of pathology was duration-dependent. Neuronal inclusions also were identified in regions not previously implicated in the disease, such as within cerebellar roof nuclei. Lewy body-like inclusions in multiple system atrophy followed the stepwise anatomic progression of Lewy body-spectrum disease inclusion pathology in 25.7% of patients with multiple system atrophy, including a patient with visual hallucinations. Further, the presence of Lewy body-like inclusions in neocortex, but not hippocampal alpha-synuclein pathology, was associated with cognitive impairment (P = 0.002). However, several cases had the presence of isolated Lewy body-like inclusions at atypical sites (e.g. thalamus, deep cerebellar nuclei) that are not typical for Lewy body-spectrum disease. Finally, interregional correlations (rho ≥ 0.6) in pathologic glial and neuronal lesion burden suggest shared mechanisms of disease progression between both discrete anatomic regions (e.g. basal forebrain and hippocampus) and cell types (neuronal and glial inclusions in frontal cortex and white matter, respectively). These findings suggest that in addition to glial inclusions, neuronal pathology plays an important role in the developmental and progression of multiple system atrophy.

Positively Charged Oligo[Poly(Ethylene Glycol) Fumarate] Scaffold Implantation Results in a Permissive Lesion Environment after Spinal Cord Injury in Rat.

Positively charged oligo[poly(ethylene glycol) fumarate] (OPF+) scaffolds loaded with Schwann cells bridge spinal cord injury (SCI) lesions and support axonal regeneration in rat. The regeneration achieved is not sufficient for inducing functional recovery. Attempts to increase regeneration would benefit from understanding the effects of the scaffold and transplanted cells on lesion environment. We conducted morphometric and stereological analysis of lesions in rats implanted with OPF+ scaffolds with or without loaded Schwann cells 1, 2, 3, 4, and 8 weeks after thoracic spinal cord transection. No differences were found in collagen scarring, cyst formation, astrocyte reactivity, myelin debris, or chondroitin sulfate proteoglycan (CSPG) accumulation. However, when scaffold-implanted animals were compared with animals with transection injuries only, these barriers to regeneration were significantly reduced, accompanied by increased activated macrophages/microglia. This distinctive and regeneration permissive tissue reaction to scaffold implantation was independent of Schwann cell transplantation. Although the tissue reaction was beneficial in the short term, we observed a chronic fibrotic host response, resulting in scaffolds surrounded by collagen at 8 weeks. This study demonstrates that an appropriate biomaterial scaffold improves the environment for regeneration. Future targeting of the host fibrotic response may allow increased axonal regeneration and functional recovery.

Degeneration of brainstem respiratory neurons in dementia with Lewy bodies.

BACKGROUND: Respiratory dysfunction, including sleep disordered breathing, is characteristic of multiple system atrophy (MSA) and may reflect degeneration of brainstem respiratory nuclei involved in respiratory rhythmogenesis and chemosensitivity, including the pre-Bötzinger complex (preBötC), nucleus raphe pallidus (RPa), and nucleus raphe obscurus (ROb). However, impaired ventilatory responses to hypercapnia have also been reported in dementia with Lewy bodies (DLB), suggesting that these nuclei may also be affected in DLB. OBJECTIVES: To determine whether there is involvement of the preBötC, RPa, and ROb in DLB. DESIGN: We applied stereological methods to analyze sections immunostained for neurokinin-1 receptor and tryptophan hydroxylase in neuropathologically confirmed cases of DLB, MSA, and controls. RESULTS: Reduction of neuronal density occurred in all three nuclei in DLB, as well as in MSA. The magnitude of neuronal depletion in ROb was similar in DLB and MSA (49% versus 56% respectively, compared to controls, P < 0.05), but neuronal loss in the preBötC and RPa was less severe in DLB than in MSA (40% loss in preBötC of DLB, P < 0.05 and 68% loss in MSA, P < 0.0001, compared to controls; 46% loss in RPa of DLB, P < 0.05 and 73% loss in MSA P < 0.0001, compared to controls). CONCLUSIONS: Medullary respiratory nuclei are affected in dementia with Lewy bodies but less severely than in multiple system atrophy. This may help explain differences in the frequency of sleep disordered breathing in these two disorders.

Implantation of cauda equina nerve roots through a biodegradable scaffold at the conus medullaris in rat.

BACKGROUND CONTEXT: Traumatic injuries occurring at the conus medullaris of the spinal cord cause permanent damage both to the central nervous system and to the cauda equina nerve roots. PURPOSE: This proof-of-concept study was to determine whether implanting the nerve roots into a biodegradable scaffold would improve regeneration after injury. METHODS: All experimental works involving rats were performed according to the approved guidelines by the Mayo Clinic Institutional Animal Care and Use Committee. Surgical procedures were performed on 32 Sprague-Dawley rats. Four ventral cauda equina nerve roots were reimplanted either directly into the ventral cord stump or through a poly(lactic-co-glycolic acid) (PLGA) scaffold. These experimental groups were compared with a control group in which the nerves were inserted into a muscle fascia barrier that was placed between the spinal cord and the nerve roots. Animals were sacrificed at 4 weeks. RESULTS: There was no difference in motor neuron counts in the spinal cord rostral to the injury in all treatment groups, implying equal potential for the regeneration into implanted nerve roots. One-way analysis of variance testing, with Tukey post hoc test, showed a statistically significant improvement in axon regeneration through the injury in the PLGA scaffold treatment group compared with the control (p<.05, scaffold n=11, control n=11). CONCLUSIONS: This pilot study demonstrated that a PLGA scaffold improved regeneration of axons into peripheral nerve roots. However, the number of regenerating axons observed was limited and did not lead to functional recovery. Future experiments will employ a different scaffold material and possible growth factors or enzymes to increase axon populations.

Autopsy confirmed multiple system atrophy cases: Mayo experience and role of autonomic function tests.

BACKGROUND: Multiple system atrophy (MSA) is a sporadic progressive neurodegenerative disorder characterised by autonomic failure, manifested as orthostatic hypotension or urogenital dysfunction, with combinations of parkinsonism that is poorly responsive to levodopa, cerebellar ataxia and corticospinal dysfunction. Published autopsy confirmed cases have provided reasonable neurological characterisation but have lacked adequate autonomic function testing. OBJECTIVES: To retrospectively evaluate if the autonomic characterisation of MSA is accurate in autopsy confirmed MSA and if consensus criteria are validated by autopsy confirmation. METHODS: 29 autopsy confirmed cases of MSA evaluated at the Mayo Clinic who had undergone formalised autonomic testing, including adrenergic, sudomotor and cardiovagal functions and Thermoregulatory Sweat Test (TST), from which the Composite Autonomic Severity Score (CASS) was derived, were included in the study. PATIENT CHARACTERISTICS: 17 men, 12 women; age of onset 57±8.1 years; disease duration to death 6.5±3.3 years; first symptom autonomic in 18, parkinsonism in seven and cerebellar in two. Clinical phenotype at first visit was MSA-P (predominant parkinsonism) in 18, MSA-C (predominant cerebellar involvement) in eight, pure autonomic failure in two and Parkinson's disease in one. Clinical diagnosis at last visit was MSA for 28 cases. Autonomic failure was severe: CASS was 7.2±2.3 (maximum 10). TST% was 65.6±33.9% and exceeded 30% in 82% of patients. The most common pattern was global anhidrosis. Norepinephrine was normal supine (203.6±112.7) but orthostatic increment of 33.5±23.2% was reduced. Four clinical features (rapid progression, early postural instability, poor levodopa responsiveness and symmetric involvement) were common. CONCLUSION: The pattern of severe and progressive generalised autonomic failure with severe adrenergic and sudomotor failure combined with the clinical phenotype is highly predictive of MSA.