Amyotrophic lateral sclerosis-motor neuron disease, monoclonal gammopathy, hyperparathyroidism, and B12 deficiency: case report and review of the literature

<p>Abstract</p> <p>Introduction</p> <p>Amyotrophic lateral sclerosis (the most common form of motor neuron disease) is a progressive and devastating disease involving both lower and upper motor neurons, typically following a relentless path towards death. Given the gravity of this diagnosis, all efforts must be made by the clinician to exclude alternative and more treatable entities. Frequent serology testing involves searching for treatable disorders, including vitamin B12 deficiency, parathyroid anomalies, and monoclonal gammopathies.</p> <p>Case presentation</p> <p>We present the case of a 78-year-old Caucasian man with all three of the aforementioned commonly searched for disorders during an investigation for amyotrophic lateral sclerosis.</p> <p>Conclusions</p> <p>The clinical utility of these common tests and what they ultimately mean in patients with amyotrophic lateral sclerosis is discussed, along with a review of the literature.</p

Human Umbilical Cord Blood Treatment in a Mouse Model of ALS: Optimization of Cell Dose

Amyotrophic Lateral Sclerosis (ALS) is a multicausal disease characterized by motor neuron degeneration in the spinal cord and brain. Cell therapy may be a promising new treatment for this devastating disorder. We recently showed that a single low dose (10(6) cells) of mononuclear human umbilical cord blood (MNC hUCB) cells administered intravenously to G93A mice delayed symptom progression and modestly prolonged lifespan. The aim of this pre-clinical translation study is to optimize the dose of MNC hUCB cells to retard disease progression in G93A mice. Three different doses of MNC hUCB cells, 10x10(6), 25x10(6) and 50x10(6), were administered intravenously into pre-symptomatic G93A mice. Motor function tests and various assays to determine cell effects were performed on these mice.Our results showed that a cell dose of 25x10(6) cells significantly increased lifespan of mice by 20-25% and delayed disease progression by 15%. The most beneficial effect on decreasing pro-inflammatory cytokines in the brain and spinal cord was found in this group of mice. Human Th2 cytokines were found in plasma of mice receiving 25x10(6) cells, although prevalent human Th1 cytokines were indicated in mice with 50x10(6) cells. High response of splenic cells to mitogen (PHA) was indicated in mice receiving 25x10(6) (mainly) and 10x10(6) cells. Significantly increased lymphocytes and decreased neutrophils in the peripheral blood were found only in animals receiving 25x10(6) cells. Stable reduction in microglia density in both cervical and lumbar spinal cords was also noted in mice administered with 25x10(6) cells.These results demonstrate that treatment for ALS with an appropriate dose of MNC hUCB cells may provide a neuroprotective effect for motor neurons through active involvement of these cells in modulating the host immune inflammatory system response

RNAseq Analyses Identify Tumor Necrosis Factor-Mediated Inflammation as a Major Abnormality in ALS Spinal Cord

ALS is a rapidly progressive, devastating neurodegenerative illness of adults that produces disabling weakness and spasticity arising from death of lower and upper motor neurons. No meaningful therapies exist to slow ALS progression, and molecular insights into pathogenesis and progression are sorely needed. In that context, we used high-depth, next generation RNA sequencing (RNAseq, Illumina) to define gene network abnormalities in RNA samples depleted of rRNA and isolated from cervical spinal cord sections of 7 ALS and 8 CTL samples. We aligned \u3e50 million 2X150 bp paired-end sequences/sample to the hg19 human genome and applied three different algorithms (Cuffdiff2, DEseq2, EdgeR) for identification of differentially expressed genes (DEG’s). Ingenuity Pathways Analysis (IPA) and Weighted Gene Co-expression Network Analysis (WGCNA) identified inflammatory processes as significantly elevated in our ALS samples, with tumor necrosis factor (TNF) found to be a major pathway regulator (IPA) and TNFα-induced protein 2 (TNFAIP2) as a major network “hub” gene (WGCNA). Using the oPOSSUM algorithm, we analyzed transcription factors (TF) controlling expression of the nine DEG/hub genes in the ALS samples and identified TF’s involved in inflammation (NFkB, REL, NFkB1) and macrophage function (NR1H2::RXRA heterodimer). Transient expression in human iPSC-derived motor neurons of TNFAIP2 (also a DEG identified by all three algorithms) reduced cell viability and induced caspase 3/7 activation. Using high-density RNAseq, multiple algorithms for DEG identification, and an unsupervised gene co-expression network approach, we identified significant elevation of inflammatory processes in ALS spinal cord with TNF as a major regulatory molecule. Overexpression of the DEG TNFAIP2 in human motor neurons, the population most vulnerable to die in ALS, increased cell death and caspase 3/7 activation. We propose that therapies targeted to reduce inflammatory TNFα signaling may be helpful in ALS patients

Glycans in Sera of Amyotrophic Lateral Sclerosis Patients and Their Role in Killing Neuronal Cells

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease caused by degeneration of upper and lower motor neurons. To date, glycosylation patterns of glycoproteins in fluids of ALS patients have not been described. Moreover, the aberrant glycosylation related to the pathogenesis of other neurodegenerative diseases encouraged us to explore the glycome of ALS patient sera. We found high levels of sialylated glycans and low levels of core fucosylated glycans in serum-derived N-glycans of patients with ALS, compared to healthy volunteer sera. Based on these results, we analyzed the IgG Fc N297-glycans, as IgG are major serum glycoproteins affected by sialylation or core fucosylation and are found in the motor cortex of ALS patients. The analyses revealed a distinct glycan, A2BG2, in IgG derived from ALS patient sera (ALS-IgG). This glycan increases the affinity of IgG to CD16 on effector cells, consequently enhancing Antibody-Dependent Cellular Cytotoxicity (ADCC). Therefore, we explore whether the Fc-N297-glycans of IgG may be involved in ALS disease. Immunostaining of brain and spinal cord tissues revealed over-expression of CD16 and co-localization of intact ALS-IgG with CD16 and in brain with activated microglia of G93A-SOD1 mice. Intact ALS-IgG enhanced effector cell activation and ADCC reaction in comparison to sugar-depleted or control IgG. ALS-IgG were localized in the synapse between brain microglia and neurons of G93A-SOD1 mice, manifesting a promising in vivo ADCC reaction. Therefore, glycans of ALS-IgG may serve as a biomarker for the disease and may be involved in neuronal damage

CNS-targeted glucocorticoid reduces pathology in mouse model of amyotrophic lateral sclerosis

Hallmarks of CNS inflammation, including microglial and astrocyte activation, are prominent features in post-mortem tissue from amyotrophic lateral sclerosis (ALS) patients and in mice overexpressing mutant superoxide dismutase-1 (SOD1 G93A ). Administration of non-targeted glucocorticoids does not significantly alter disease progression, but this may reflect poor CNS delivery. Here, we sought to discover whether CNS-targeted, liposomal encapsulated glucocorticoid would inhibit the CNS inflammatory response and reduce motor neuron loss. SOD1 G93A mice were treated with saline, free methylprednisolone (MP, 10 mg/kg/week) or glutathione PEGylated liposomal MP (2B3-201, 10 mg/kg/week) and compared to saline treated wild-type animals. Animals were treated weekly with intravenous injections for 9 weeks from 60 days of age. Weights and motor performance were monitored during this period. At the end of the experimental period (116 days) mice were imaged using T 2-weighted MRI for brainstem pathology; brain and spinal cord tissue were then collected for histological analysis

Evidence of Compromised Blood-Spinal Cord Barrier in Early and Late Symptomatic SOD1 Mice Modeling ALS

Background: The blood-brain barrier (BBB), blood-spinal cord barrier (BSCB), and blood-cerebrospinal fluid barrier (BCSFB) control cerebral/spinal cord homeostasis by selective transport of molecules and cells from the systemic compartment. In the spinal cord and brain of both ALS patients and animal models, infiltration of T-cell lymphocytes, monocyte-derived macrophages and dendritic cells, and IgG deposits have been observed that may have a critical role in motor neuron damage. Additionally, increased levels of albumin and IgG have been found in the cerebrospinal fluid in ALS patients. These findings suggest altered barrier permeability in ALS. Recently, we showed disruption of the BBB and BSCB in areas of motor neuron degeneration in the brain and spinal cord in G93A SOD1 mice modeling ALS at both early and late stages of disease using electron microscopy. Examination of capillary ultrastructure revealed endothelial cell degeneration, which, along with astrocyte alteration, compromised the BBB and BSCB. However, the effect of these alterations upon barrier function in ALS is still unclear. The aim of this study was to determine the functional competence of the BSCB in G93A mice at different stages of disease. Methodology/Principal Findings: Evans Blue (EB) dye was intravenously injected into ALS mice at early or late stage disease. Vascular leakage and the condition of basement membranes, endothelial cells, and astrocytes were investigated in cervical and lumbar spinal cords using immunohistochemistry. Results showed EB leakage in spinal cord microvessels from all G93A mice, indicating dysfunction in endothelia and basement membranes and confirming our previous ultrastructural findings on BSCB disruption. Additionally, downregulation of Glut-1 and CD146 expressions in the endothelial cells of the BSCB were found which may relate to vascular leakage. Conclusions/Significance: Results suggest that the BSCB is compromised in areas of motor neuron degeneration in ALS mice at both early and late stages of the disease

Effects of peripheral nerve injury on parvalbumin expression in adult rat dorsal root ganglion neurons

Background: Parvalbumin (PV) is a calcium binding protein that identifies a subpopulation of proprioceptive dorsal root ganglion (DRG) neurons. Calcitonin gene-related peptide (CGRP) is also expressed in a high proportion of muscle afferents but its relationship to PV is unclear. Little is known of the phenotypic responses of muscle afferents to nerve injury. Sciatic nerve axotomy or L5 spinal nerve ligation and section (SNL) lesions were used to explore these issues in adult rats using immunocytochemistry. Results: In naive animals, the mean PV expression was 25 % of L4 or L5 dorsal root ganglion (DRG) neurons, and this was unchanged 2 weeks after sciatic nerve axotomy. Colocalization studies with the injury marker activating transcription factor 3 (ATF3) showed that approximately 24 % of PV neurons expressed ATF3 after sciatic nerve axotomy suggesting that PV may show a phenotypic switch from injured to uninjured neurons. This possibility was further assessed using the spinal nerve ligation (SNL) injury model where injured and uninjured neurons are located in different DRGs. Two weeks after L5 SNL there was no change in total PV staining and essentially all L5 PV neurons expressed ATF3. Additionally, there was no increase in PV-ir in the adjacent uninjured L4 DRG cells. Co-labelling of DRG neurons revealed that less than 2 % of PV neurons normally expressed CGRP and no colocalization was seen after injury. Conclusion: These experiments clearly show that axotomy does not produce down regulation of PV protein in the DRG. Moreover, this lack of change is not due to a phenotypic switch in PV immunoreactive (ir) neurons, or de novo expression of PV-ir in uninjured neurons after nerve injury. These results further illustrate differences that occur when muscle afferents are injured as compared to cutaneous afferents

Advances, challenges and future directions for stem cell therapy in amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is a rapidly progressive neurodegenerative condition where loss of motor neurons within the brain and spinal cord leads to muscle atrophy, weakness, paralysis and ultimately death within 3–5 years from onset of symptoms. The specific molecular mechanisms underlying the disease pathology are not fully understood and neuroprotective treatment options are minimally effective. In recent years, stem cell transplantation as a new therapy for ALS patients has been extensively investigated, becoming an intense and debated field of study. In several preclinical studies using the SOD1G93A mouse model of ALS, stem cells were demonstrated to be neuroprotective, effectively delayed disease onset and extended survival. Despite substantial improvements in stem cell technology and promising results in preclinical studies, several questions still remain unanswered, such as the identification of the most suitable and beneficial cell source, cell dose, route of delivery and therapeutic mechanisms. This review will cover publications in this field and comprehensively discuss advances, challenges and future direction regarding the therapeutic potential of stem cells in ALS, with a focus on mesenchymal stem cells. In summary, given their high proliferation activity, immunomodulation, multi-differentiation potential, and the capacity to secrete neuroprotective factors, adult mesenchymal stem cells represent a promising candidate for clinical translation. However, technical hurdles such as optimal dose, differentiation state, route of administration, and the underlying potential therapeutic mechanisms still need to be assessed