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1.
Article in English | MEDLINE | ID: mdl-35031587

ABSTRACT

BACKGROUND AND OBJECTIVES: Improved biomarkers of neuroprotective treatment are needed in progressive multiple sclerosis (PMS) to facilitate more efficient phase 2 trial design. The MS-STAT randomized controlled trial supported the neuroprotective potential of high-dose simvastatin in secondary progressive MS (SPMS). Here, we analyze serum from the MS-STAT trial to assess the extent to which neurofilament light (NfL) and neurofilament heavy (NfH), both promising biomarkers of neuroaxonal injury, may act as biomarkers of simvastatin treatment in SPMS. METHODS: The MS-STAT trial randomized patients to 80 mg simvastatin or placebo. Serum was analyzed for NfL and NfH using Simoa technology. We used linear mixed models to investigate the treatment effects of simvastatin compared with placebo on NfL and NfH. Additional models examined the relationships between neurofilaments and MRI and clinical measures of disease severity. RESULTS: A total of 140 patients with SPMS were included. There was no evidence for a simvastatin treatment effect on NfL or NfH: compared with placebo, NfL was 1.2% lower (95% CI 10.6% lower to 9.2% higher; p = 0.820) and NfH was 0.4% lower (95% CI 18.4% lower to 21.6% higher; p = 0.969) in the simvastatin treatment group. Secondary analyses suggested that higher NfL was associated with greater subsequent whole brain atrophy, higher T2 lesion volume, and more new/enlarging T2 lesions in the previous 12 months, as well as greater physical disability. There were no significant associations between NfH and MRI or clinical variables. DISCUSSION: We found no evidence of a simvastatin treatment effect on serum neurofilaments. While confirmation of the neuroprotective benefits of simvastatin is awaited from the ongoing phase 3 study (NCT03387670), our results suggest that treatments capable of slowing the rate of whole brain atrophy in SPMS, such as simvastatin, may act via mechanisms largely independent of neuroaxonal injury, as quantified by NfL. This has important implications for the design of future phase 2 clinical trials in PMS. TRIAL REGISTRATION INFORMATION: MS-STAT: NCT00647348. CLASSIFICATION OF EVIDENCE: This study provides class I evidence that simvastatin treatment does not have a large impact on either serum NfL or NfH, as quantified in this study, in SPMS.


Subject(s)
Multiple Sclerosis, Chronic Progressive , Neurofilament Proteins/blood , Neuroprotective Agents/pharmacology , Simvastatin/pharmacology , Adult , Biomarkers , Female , Humans , Male , Middle Aged , Multiple Sclerosis, Chronic Progressive/blood , Multiple Sclerosis, Chronic Progressive/diagnosis , Multiple Sclerosis, Chronic Progressive/drug therapy , Neurofilament Proteins/drug effects , Outcome Assessment, Health Care
2.
J Neuromuscul Dis ; 9(1): 111-119, 2022.
Article in English | MEDLINE | ID: mdl-34776417

ABSTRACT

OBJECTIVE: To retrospectively evaluate the utility of serum and cerebrospinal fluid (CSF) levels of neurofilament light chain (NfL) and phosphorylated neurofilament heavy chain (pNfH) as biomarkers for spinal muscular atrophy (SMA) progression and response to nusinersen treatment. METHODS: NfL and pNfH levels were quantified using single molecular array (SIMOA) in CSF of 33 adult SMA patients (SMN copy number 3-5) before and in response to nusinersen treatment. In 11 of the patients, blood serum samples were also collected. CSF NfL and pNfH from patients were compared to CSF Nfs from age-matched controls without neurological disease (n = 6). For patients, pearson correlation coefficients (r) were calculated to investigate associations between Nf levels and other functional outcome measures. RESULTS: Nf levels were similar between SMA and control adults and showed no change in response to nusinersen treatment in CSF or serum. Cross-sectional analyses showed an increase in CSF NfL and pNfH with age in patients (NfL p = 0.0013; pNfH p = 0.0035) and an increase in CSF NfL in controls (p = 0.002). In non-ambulatory patients, baseline serum pNfH showed a negative correlation with multiple strength and functional assessment metrics including Revised Upper Limb Module (r = -0.822, p = 0.04), upper extremity strength (r = -0.828, p = 0.042), lower extremity strength (r = -0.860, p = 0.028), and total strength (r = -0.870, p = 0.024). CONCLUSIONS: Nf levels did not change in response to nusinersen in adults with SMA and were not different from controls. In patients and controls, we detected an age-related increase in baseline CSF NfL and pNfH levels. Though some associations were identified, our results suggest Nf levels are not preditive or prognostic biomarkers in this population.


Subject(s)
Aging , Muscular Atrophy, Spinal , Neurofilament Proteins , Oligonucleotides/pharmacology , Adult , Biomarkers/blood , Biomarkers/cerebrospinal fluid , Cross-Sectional Studies , Humans , Muscular Atrophy, Spinal/blood , Muscular Atrophy, Spinal/cerebrospinal fluid , Muscular Atrophy, Spinal/diagnosis , Muscular Atrophy, Spinal/drug therapy , Neurofilament Proteins/blood , Neurofilament Proteins/cerebrospinal fluid , Neurofilament Proteins/drug effects , Outcome Assessment, Health Care , Prognosis , Retrospective Studies
3.
J Integr Neurosci ; 20(1): 125-130, 2021 Mar 30.
Article in English | MEDLINE | ID: mdl-33834699

ABSTRACT

This article demonstrates that mannotriose effectively induces the differentiation of mesenchymal stem cells into neuron-like cells in vitro. Rat-derived mesenchymal stem cells were investigated on their potential to differentiate into neuron-like cells induced by mannotriose purified from Radix Rehmanniae Preparata in vitro. The percentage of the neuron-specific enolase positive cells and the Nissl positive cells after mannotriose treatment was increased. The mRNA levels of neurofilament medium and neuron-specific enolase were upregulated in the mannotriose group compared to the control. These findings demonstrate that mannotriose purified from Radix Rehmanniae Preparata can effectively induce differentiation of rat-derived mesenchymal stem cells into neuron-like cells.


Subject(s)
Cell Differentiation/drug effects , Mesenchymal Stem Cells/drug effects , Neurofilament Proteins/drug effects , Neurons , Phosphopyruvate Hydratase/drug effects , Rehmannia , Trisaccharides/pharmacology , Animals , Plant Preparations , Rats , Up-Regulation
4.
Neuroreport ; 32(6): 518-524, 2021 04 07.
Article in English | MEDLINE | ID: mdl-33788819

ABSTRACT

Achyranthes bidentata polypeptide k (ABPPk), a powerful active component from a traditional Chinese medicinal herb-Achyranthes bidentata Bl., has exhibited promising neuroprotective activity due to its multiple-targeting capability. However, the effect of ABPPk on the survival, growth and axonal regeneration of spinal cord motor neurons remains unclear. Here, a modified method, which is more optimized for embryonic cells in ambient carbon dioxide levels, was used for acquisition of rat embryonic spinal cord motor neurons with high survival and purity. ABPPk concentration-dependently enhanced the neuronal viability and promoted the neurite outgrowth. Co-culture of motor neurons and skeletal myocytes model indicated that ABPPk enhanced the neuromuscular junction development and maturation. A microfluidic axotomy model was further established for the axonal disconnection, and ABPPk significantly accelerated the axonal regeneration of motor neurons. Furthermore, we demonstrated that the upregulation of three neurofilament protein subunits in motor neurons might be relevant to the mechanisms of the growth-promoting effect of ABPPk. Our findings provide an experimental and theoretical basis for the development of ABPPk as a potential application in the development of treatment strategy for nerve injury diseases.


Subject(s)
Achyranthes , Axons/drug effects , Motor Neurons/drug effects , Muscle Fibers, Skeletal/drug effects , Nerve Regeneration/drug effects , Neuromuscular Junction/drug effects , Neuronal Outgrowth/drug effects , Plant Extracts/pharmacology , Animals , Axotomy , Cell Survival/drug effects , Coculture Techniques , GAP-43 Protein/drug effects , GAP-43 Protein/metabolism , In Vitro Techniques , Neurofilament Proteins/drug effects , Neurofilament Proteins/metabolism , Peptides/pharmacology , Peripheral Nerve Injuries , Primary Cell Culture , Rats , Rats, Sprague-Dawley , Spinal Cord/cytology
5.
Br J Anaesth ; 125(3): 282-290, 2020 09.
Article in English | MEDLINE | ID: mdl-32536445

ABSTRACT

BACKGROUND: Postoperative neurocognitive disorders may arise in part from adverse effects of general anaesthetics on the CNS, especially in older patients or individuals otherwise vulnerable to neurotoxicity because of systemic disease or the presence of pre-existing neuropathology. Previous studies have documented cytokine and injury biomarker responses to surgical procedures that included general anaesthesia, but it is not clear to what degree anaesthetics contribute to these responses. METHODS: We performed a prospective cohort study of 59 healthy volunteers aged 40-80 yr who did not undergo surgery. Plasma markers of neurological injury and inflammation were measured immediately before and 5 h after induction of general anaesthesia with 1 minimum alveolar concentration of sevoflurane. Biomarkers included interleukin-6 (IL-6), tumour necrosis factor alpha (TNF-α), C-reactive protein (CRP), and neural injury (tau, neurofilament light [NF-L], and glial fibrillary acidic protein [GFAP]). RESULTS: Baseline biomarkers were in the normal range, although NF-L and GFAP were elevated as a function of age. At 5 h after induction of anaesthesia, plasma tau, NF-L, and GFAP were significantly decreased relative to baseline. Plasma IL-6 was significantly increased after anaesthesia, but by a biologically insignificant degree (<1 pg ml-1); plasma TNF-α and CRP were unchanged. CONCLUSIONS: Sevoflurane general anaesthesia without surgery, even in older adults, did not provoke an inflammatory state or neuronal injury at a concentration that is detectable by an acute elevation of measured plasma biomarkers in the early hours after exposure. CLINICAL TRIAL REGISTRATION: NCT02275026.


Subject(s)
Anesthesia, General/methods , Anesthetics, Inhalation/blood , Adult , Aged , Aged, 80 and over , Biomarkers/blood , C-Reactive Protein/drug effects , Cohort Studies , Female , Glial Fibrillary Acidic Protein/blood , Glial Fibrillary Acidic Protein/drug effects , Healthy Volunteers , Humans , Interleukin-6/blood , Male , Middle Aged , Neurofilament Proteins/blood , Neurofilament Proteins/drug effects , Prospective Studies , Reference Values , Tumor Necrosis Factor-alpha/blood , Tumor Necrosis Factor-alpha/drug effects
6.
Article in English | MEDLINE | ID: mdl-32404429

ABSTRACT

OBJECTIVE: To investigate serum neurofilament light chain (sNfL) as a potential biomarker for disease activity and treatment response in pediatric patients with multiple sclerosis (MS). METHODS: In this retrospective cohort study, sNfL levels were measured in a pediatric MS cohort (n = 55, follow-up 12-105 months) and in a non-neurologic pediatric control cohort (n = 301) using a high-sensitivity single-molecule array assay. Association of sNfL levels and treatment and clinical and MRI parameters were calculated. RESULTS: Untreated patients had higher sNfL levels than controls (median 19.0 vs 4.6 pg/mL; CI [4.732, 6.911]), p < 0.001). sNfL levels were significantly associated with MRI activity (+9.1% per contrast-enhancing lesion, CI [1.045, 1.138], p < 0.001; +0.6% per T2-weighted lesion, CI [1.001, 1.010], p = 0.015). Higher values were associated with a relapse <90 days ago (+51.1%; CI [1.184, 1.929], p < 0.001) and a higher Expanded Disability Status Scale score (CI [1.001, 1.240], p = 0.048). In patients treated with interferon beta-1a/b (n = 27), sNfL levels declined from 14.7 to 7.9 pg/mL after 6 ± 2 months (CI [0.339, 0.603], p < 0.001). Patients with insufficient control of clinical or MRI disease activity under treatment with interferon beta-1a/b or glatiramer acetate who switched to fingolimod (n = 18) showed a reduction of sNfL levels from 16.5 to 10.0 pg/mL 6 ± 2 months after switch (CI [0.481, 0.701], p < 0.001). CONCLUSIONS: sNfL is a useful biomarker for monitoring disease activity and treatment response in pediatric MS. It is most likely helpful to predict disease severity and to guide treatment decisions in patients with pediatric MS. This study provides Class III evidence that sNfL levels are associated with disease activity in pediatric MS.


Subject(s)
Immunologic Factors/pharmacology , Multiple Sclerosis/blood , Multiple Sclerosis/diagnosis , Multiple Sclerosis/drug therapy , Neurofilament Proteins/blood , Adolescent , Biomarkers/blood , Child , Disease Progression , Female , Follow-Up Studies , Humans , Male , Multiple Sclerosis/physiopathology , Neurofilament Proteins/drug effects , Retrospective Studies , Treatment Outcome
7.
Brain Dev ; 42(3): 311-314, 2020 Mar.
Article in English | MEDLINE | ID: mdl-31889567

ABSTRACT

BACKGROUND: In July 2018, a rare and serious adverse effect (AE), namely, communicating hydrocephalus unrelated to meningitis or bleeding, was reported in relation to five patients treated with nusinersen for spinal muscular atrophy (SMA). Some patients were managed using a ventriculo-peritoneal shunt (VPS) implant and continued to receive nusinersen treatment. However, there is limited information concerning the effectiveness and safety of nusinersen treatment for patients with a VPS. CASE REPORT: A female patient exhibited general hypotonia soon after birth and was diagnosed, using genetic analysis, with spinal muscular atrophy. She required permanent invasive ventilation from 2 months of age. She developed a progressive hydrocephalus and underwent placement of a VPS in infancy. Treatment with nusinersen was initiated when she was 7 years old. The neurofilament light-chain (NfL) concentration in the cerebrospinal fluid (CSF) decreased over time with nusinersen treatment. Twelve months have passed since the start of nusinersen treatment and no AEs have been observed. CONCLUSION: Nusinersen treatment may be effective and safe, even after placement of a VPS. NfL levels in the CSF could be valuable markers of disease activity/treatment response even in advanced stages of SMA.


Subject(s)
Muscular Atrophy, Spinal/drug therapy , Neurofilament Proteins , Oligonucleotides/pharmacology , Ventriculoperitoneal Shunt , Child , Female , Humans , Muscular Atrophy, Spinal/surgery , Neurofilament Proteins/cerebrospinal fluid , Neurofilament Proteins/drug effects , Oligonucleotides/administration & dosage , Oligonucleotides/adverse effects
8.
J Neurol ; 267(1): 36-44, 2020 Jan.
Article in English | MEDLINE | ID: mdl-31552549

ABSTRACT

OBJECTIVE: To determine the diagnostic and monitoring value of serum neurofilament light chain (NfL) in spinal muscular atrophy (SMA). METHODS: We measured serum NfL in 46 SMA patients at baseline and over 14 months of treatment with the antisense-oligonucleotide (ASO) nusinersen using the ultrasensitive single molecule array (Simoa) technology. Serum NfL levels of SMA patients were compared to controls and related to cerebrospinal fluid (CSF) NfL, blood-CSF barrier function quantified by the albumin blood/CSF ratio (Qalb) and motor scores (Hammersmith Functional Motor Scale Expanded, HFMSE; Amyotrophic Lateral Sclerosis Functional Rating Scale-Revised, ALSFRS-R). RESULTS: Serum NfL levels of SMA patients were in the range of controls (p = 0.316) and did not correlate with CSF NfL (ρ = 0.302, p = 0.142) or Qalb (ρ = - 0.160, p = 0.293). During therapy, serum NfL levels were relatively stable with notable concentration changes in single SMA patients, however, within the control range. Higher NfL levels were associated with worse motor performance in SMA (baseline: HFMSE ρ = - 0.330, p = 0.025, ALSFRS-R ρ = - 0.403, p = 0.005; after 10 months: HFMSE ρ = - 0.525, p = 0.008, ALSFRS-R ρ = - 0.537, p = 0.007), but changes in motor scores did not correlate with changes in serum NfL. CONCLUSION: Diagnostic and monitoring performance of serum NfL measurement seems to differ between SMA subtypes. Unlike to SMA type 1, in adolescent and adult SMA type 2 and 3 patients, neurodegeneration is not reflected by increased NfL levels and short-term therapeutic effects cannot be observed. Long-term follow-up has to be performed to see if even low levels of NfL might be good prognostic markers.


Subject(s)
Muscular Atrophy, Spinal/blood , Muscular Atrophy, Spinal/drug therapy , Neurofilament Proteins/blood , Neurofilament Proteins/drug effects , Oligonucleotides/pharmacology , Outcome Assessment, Health Care , Spinal Muscular Atrophies of Childhood/blood , Spinal Muscular Atrophies of Childhood/drug therapy , Adolescent , Adult , Child , Female , Follow-Up Studies , Humans , Male , Middle Aged , Muscular Atrophy, Spinal/physiopathology , Severity of Illness Index , Spinal Muscular Atrophies of Childhood/physiopathology , Young Adult
9.
Mult Scler ; 26(6): 659-667, 2020 05.
Article in English | MEDLINE | ID: mdl-30912689

ABSTRACT

OBJECTIVES: Serum neurofilament light chain (sNfL) has been proposed a potential biomarker in multiple sclerosis (MS) based on mainly cross-sectional observations in Western population. To clarify clinical implication of sNfL, we longitudinally analysed sNfL levels at multiple time points in Korean MS patients undergoing alemtuzumab therapy. METHODS: Between 2016 and 2018, 144 sera from 17 MS patients treated with alemtuzumab at National Cancer Centre and 35 sera from 35 age- and gender-matched healthy controls (HCs) were collected for a longitudinal study with a mean 21-month follow-up. The sera were measured for sNfL levels using single molecule array. Patients were classified into two groups: evidence of disease activity (EDA) or no evidence of disease activity (NEDA). RESULTS: During alemtuzumab therapy, sNfL levels in EDA patients were significantly higher than those in NEDA patients and HCs (p < 0.001). In longitudinal analysis, the sNfL levels were consistently low in NEDA patients, while it consistently increased in radiologically and/or clinically active status in EDA patients. All sNfL levels in radiologically and/or clinically active status samples were higher than those in inactive status samples. CONCLUSION: These results suggest that sNfL is a promising monitoring biomarker for personalized therapeutics in MS patients.


Subject(s)
Alemtuzumab/pharmacology , Disease Progression , Immunologic Factors/pharmacology , Multiple Sclerosis, Relapsing-Remitting/blood , Multiple Sclerosis, Relapsing-Remitting/drug therapy , Neurofilament Proteins/blood , Outcome Assessment, Health Care , Adult , Alemtuzumab/administration & dosage , Biomarkers/blood , Cross-Sectional Studies , Female , Humans , Immunologic Factors/administration & dosage , Longitudinal Studies , Male , Middle Aged , Neurofilament Proteins/drug effects
10.
Ann Clin Transl Neurol ; 6(12): 2437-2447, 2019 12.
Article in English | MEDLINE | ID: mdl-31814335

ABSTRACT

OBJECTIVE: Neuronal ceroid lipofuscinosis type 2 (CLN2 disease) is a rare, progressive, fatal neurodegenerative pediatric disorder resulting from deficiencies of the lysosomal enzyme tripeptidyl peptidase 1 that are caused by mutations in TPP1. Identifying biomarkers of CLN2 disease progression will be important in assessing the efficacy of therapeutic interventions for this disorder. Neurofilament light is an intrinsic component of healthy neurons; elevated circulating extracellular neurofilament light is a biomarker of neuropathology in several adult-onset neurological diseases. Our objective was to assess whether circulating neurofilament light is a biomarker that is responsive to enzyme replacement therapy (ERT) in CLN2 disease. METHODS: Using an ultrasensitive immunoassay, we assessed plasma neurofilament light changes during disease progression in a canine model of CLN2 disease and in ERT clinical trial CLN2 disease patients. RESULTS: In tripeptidyl peptidase 1 (TPP1)-null dogs (N = 11), but not in control dogs [N = 6 (TPP1+/- ) and N = 27 (WT)], neurofilament light levels increased more than tenfold above initial low baseline levels during disease progression. Before treatment in 21 human subjects with CLN2 disease (age range: 1.72-6.85 years), neurofilament light levels were 48-fold higher (P < 0.001) than in 7 pediatric controls (age range: 8-11 years). Pretreatment neurofilament light did not significantly correlate with disease severity or age. In CLN2 disease subjects receiving ERT, neurofilament light levels decreased by 50% each year over more than 3 years of treatment. INTERPRETATION: Our data indicate that circulating neurofilament light is a treatment-responsive biomarker in CLN2 disease and could contribute to understanding of the pathophysiology of this devastating pediatric disorder.


Subject(s)
Aminopeptidases/pharmacology , Dipeptidyl-Peptidases and Tripeptidyl-Peptidases/pharmacology , Disease Progression , Enzyme Replacement Therapy , Neurofilament Proteins/blood , Neuronal Ceroid-Lipofuscinoses/blood , Serine Proteases/pharmacology , Aminopeptidases/genetics , Animals , Animals, Genetically Modified , Biomarkers/blood , Child , Child, Preschool , Dipeptidyl-Peptidases and Tripeptidyl-Peptidases/genetics , Disease Models, Animal , Dogs , Female , Humans , Infant , Male , Neurofilament Proteins/drug effects , Neuronal Ceroid-Lipofuscinoses/drug therapy , Recombinant Proteins/pharmacology , Serine Proteases/genetics , Tripeptidyl-Peptidase 1
11.
J Neurotrauma ; 35(10): 1185-1191, 2018 05 15.
Article in English | MEDLINE | ID: mdl-29415612

ABSTRACT

The primary aim of this study was to examine the effects of valproic acid (VPA) treatment on serum glial fibrillary acidic protein (GFAP) and neurofilament light chain (NF-L) levels. To achieve this aim, we obtained serum samples from: 1) 10 Yorkshire swine subjected to controlled cortical impact traumatic brain injury (CCI TBI) + polytrauma and randomized to receive either normal saline (NS) + VPA (n = 5) or NS alone (n = 5) and 2) five additional swine subjected to CCI TBI without polytrauma and treated with VPA. GFAP and NF-L levels were measured in samples obtained from baseline until 10 days post-injury using a digital immunoassay from Quanterix Corporation. We found that elevated GFAP and NF-L levels were first detected at 2 h post-injury; and peaked at 24 h and 72 h respectively. GFAP levels returned to baseline levels by Day 10, while NF-L remained elevated at Day 10. In TBI + polytrauma swine, the magnitude and duration of biomarker elevation, quantified by the area under the biomarker-concentration-versus-time curve during the first 10 days (AUC0-10days), was higher in the NS group, compared with the VPA group. For GFAP, the AUC0-10days was 45,535 (IQR: 35,741-105,711) and 22,837 (IQR: 8,082-46,627) for the NS and NS+VPA groups, respectively. For NF-L, the AUC0-10days was 43,073 (IQR: 18,739-120,794) and 4,475 (2,868-11,157) for the NS and NS+VPA groups, respectively. Twenty-four hour GFAP and NF-L levels had the strongest correlation with lesion size and time to normalization of behavior. Accordingly, we conclude that treatment with VPA results in significantly lower serum GFAP and NF-L levels. The time-point at which GFAP and NF-L levels have the strongest correlation with outcome is 24 h post-injury.


Subject(s)
Biomarkers/blood , Brain Injuries, Traumatic/blood , Glial Fibrillary Acidic Protein/blood , Neurofilament Proteins/blood , Neuroprotective Agents/pharmacology , Valproic Acid/pharmacology , Animals , Female , Glial Fibrillary Acidic Protein/drug effects , Neurofilament Proteins/drug effects , Swine
12.
Neurosci Lett ; 668: 24-30, 2018 03 06.
Article in English | MEDLINE | ID: mdl-29307599

ABSTRACT

Histone deacetylase (HDAC) inhibitors, which have an effect on cell homeostasis, cell cycle progression, and terminal differentiation, can act to promote self-renewal and enhance directed differentiation of several lineages of stem cells. However, the roles of HDAC inhibitors on neurogenic differentiation and the mechanisms of Wnt signaling following treatment with HDAC inhibitors remain unclear in stem cells. We hypothesized that HDAC inhibitors regulate downstream Wnt signaling and neurogenic differentiation of mesenchymal stem cells. Following neural induction with supplementary factors, human adipose tissue-derived mesenchymal stem cells (hADSCs) were differentiated into neurogenic cells in vitro. We examined the neurogenic differentiation induced by the HDAC inhibitors, MS-275, sodium butyrate (NaB), trichostatin A (TSA), and valproic acid (VPA), by RT-PCR and western blot analysis. Based on RT-PCR analysis, the expressions of NEUROG2 and NEFL were highly increased following HDAC inhibitor treatment compared with control medium. Most of the neuronal marker genes were expressed when neural-induced hADSCs (NI-hADSCs) were treated with the HDAC inhibitors individually. Interestingly, expression of most of the Wnt-related genes were highly increased following treatment with the HDAC inhibitors, especially with MS-275 treatment. Further, the protein level of Wnt5 was upregulated after neurogenic induction with MS-275 and VPA treatment, based on western blot analysis. Furthermore, we found that c-Jun expression was increased after treatment with the HDAC inhibitors, except with NaB. The protein levels of phosphor-JNK and phosphor-GSK-3ß were upregulated considerably. In conclusion, the HDAC inhibitors could induce neurogenic differentiation of hADSCs by activating canonical Wnt or non-canonical Wnt signaling pathways.


Subject(s)
Adipose Tissue/cytology , Cell Differentiation/drug effects , Cytoskeletal Proteins/metabolism , Histone Deacetylase Inhibitors/pharmacology , Mesenchymal Stem Cells/drug effects , Neurofilament Proteins/metabolism , Neurons/drug effects , Nuclear Proteins/metabolism , Wnt Proteins/drug effects , Wnt Signaling Pathway/drug effects , Basic Helix-Loop-Helix Transcription Factors/drug effects , Basic Helix-Loop-Helix Transcription Factors/metabolism , Cells, Cultured , Cytoskeletal Proteins/drug effects , Humans , MAP Kinase Signaling System/physiology , Nerve Tissue Proteins/drug effects , Nerve Tissue Proteins/metabolism , Neurofilament Proteins/drug effects , Nuclear Proteins/drug effects
13.
Neurotox Res ; 33(2): 316-336, 2018 02.
Article in English | MEDLINE | ID: mdl-28875469

ABSTRACT

In the present study, we screened the sera of subjects chronically exposed to mixtures of pesticides (composed mainly of organophosphorus compounds (OPs) and others) and developed neurological symptoms for the presence of autoantibodies against cytoskeletal neural proteins. OPs have a well-characterized clinical profile resulting from acute cholinergic crisis. However, some of these compounds cause neuronal degeneration and demyelination known as organophosphorus compound-induced delayed neurotoxicity (OPIDN) and/or organophosphorus compound-induced chronic neurotoxicity (OPICN). Studies from our group have demonstrated the presence of autoantibodies to essential neuronal and glial proteins against cytoskeletal neural proteins in patients with chemical-induced brain injury. In this study, we screened the serum of 50 pesticide-exposed subjects and 25 non-exposed controls, using Western blot analysis against the following proteins: neurofilament triplet proteins (NFPs), tubulin, microtubule-associated tau proteins (Tau), microtubule-associated protein-2 (MAP-2), myelin basic protein (MBP), myelin-associated glycoprotein (MAG), glial fibrillary acidic protein (GFAP), calcium-calmodulin kinase II (CaMKII), glial S100-B protein, and alpha-synuclein (SNCA). Serum reactivity was measured as arbitrary chemiluminescence units. As a group, exposed subjects had significantly higher levels of autoantibody reactivity in all cases examined. The folds of increase in of autoantibodies against neural proteins of the subjects compared to healthy humans in descending order were as follows: MBP, 7.67, MAG 5.89, CaMKII 5.50, GFAP 5.1, TAU 4.96, MAP2 4.83, SNCA 4.55, NFP 4.55, S-100B 2.43, and tubulin 1.78. This study has demonstrated the presence of serum autoantibodies to central nervous system-specific proteins in a group of farmers chronically exposed to pesticides who developed neurological signs and symptoms of neural injury. These autoantibodies can be used as future diagnostic/therapeutic target for OP-induced neurotoxicity.


Subject(s)
Autoantibodies/immunology , Biomarkers/blood , Neurofilament Proteins/blood , Pesticides/toxicity , Adult , Aged , Autoantibodies/blood , Cytoskeletal Proteins/metabolism , Female , Glial Fibrillary Acidic Protein/blood , Humans , Male , Middle Aged , Myelin Basic Protein/metabolism , Neurofilament Proteins/drug effects , Neurotoxicity Syndromes/blood , Neurotoxicity Syndromes/diagnosis , Organophosphorus Compounds/pharmacology , Peripheral Nervous System Diseases/chemically induced
14.
Folia Med (Plovdiv) ; 58(4): 241-249, 2016 12 01.
Article in English | MEDLINE | ID: mdl-28068280

ABSTRACT

BACKGROUND: Alcohol-induced cerebellar neurodegeneration is a neuroadaptation that is associated with chronic alcohol abuse. Conventional drugs have been largely unsatisfactory in preventing neurodegeneration. Yet, multimodal neuro-protective therapeutic agents have been hypothesised to have high therapeutic potential for the treatment of CNS conditions; there is yet a dilemma of how this would be achieved. Contrarily, medicinal botanicals are naturally multimodal in their mechanism of action. AIM: The effect of L. owariensis was therefore assessed in alcohol-induced neurodegeneration of the cerebellar cortex in rats. MATERIALS AND METHODS: Two groups of rats were oro-gastrically fed thrice daily with 5 g/kg ethanol (25% w/v), and 5 g/kg ethanol (25% w/v) plus L. owariensis (100 mg/kg body weight) respectively in diluted nutritionally complete diet (50% v/v). A control group was correspondingly fed a nutritionally complete diet (50% v/v) made isocaloric with glucose. Cytoarchitectural study of the cerebellar cortex was examined with H&E. Immunocytochemical analysis was carried out with the use of monoclonal antibody anti-NF in order to detect alterations in the neuronal cytoskeleton. RESULTS: After 4 days of binge alcohol treatment, we observed that L. owariensis supplementation significantly lowered the levels of histologic and biochemical indices of neurodegeneration. The level of neurodegeneration and cytoarchitecture distortion of the cerebellar cortex of rats exposed to ethanol was reduced by L. owariensis. Neurofilament-immunoreactivity (NF-IR) was evoked in the Purkinje cells of rats that received L. owariensis supplement. CONCLUSIONS: L. owariensis attenuates alcohol-induced cerebellar degeneration in the rat by alleviating oxidative stress and alteration of NF protein expression in the Purkinje cells.


Subject(s)
Apocynaceae , Central Nervous System Depressants/toxicity , Cerebellum/drug effects , Ethanol/toxicity , Neurofilament Proteins/drug effects , Plant Preparations/pharmacology , Purkinje Cells/drug effects , Alcohol-Induced Disorders, Nervous System , Animals , Cerebellar Diseases , Cerebellum/metabolism , Disease Models, Animal , Immunohistochemistry , Male , Neurodegenerative Diseases , Neurofilament Proteins/metabolism , Purkinje Cells/metabolism , Rats , Rats, Wistar
15.
J Intern Med ; 273(5): 478-89, 2013 May.
Article in English | MEDLINE | ID: mdl-23331301

ABSTRACT

A number of neurotoxic chemicals induce accumulation of neurofilaments in axonal swellings that appear at varying distances from the cell body. This pathology is associated with axonal degeneration of different degrees. The clinical manifestation is most commonly that of a mixed motor-sensory peripheral axonopathy with a disto-proximal pattern of progression, as in cases of chronic exposure to n-hexane and carbon disulphide. It has been demonstrated that protein adduct formation is a primary molecular mechanism of toxicity in these axonopathies, but how this mechanism leads to neurofilament accumulation and axonal degeneration remains unclear. Furthermore, little is known regarding the mechanisms of neurofilamentous axonopathy caused by 3,3'-iminodipropionitrile, an experimental toxin that induces proximal axon swelling that is strikingly similar to that found in early amyotrophic lateral sclerosis. Here, we review the available data and main hypotheses regarding the toxic axonopathies and compare them with the current knowledge of the biological basis of neurofilament transport. We also review recent studies addressing the question of how these axonopathies may cause axonal degeneration. Understanding the mechanisms underlying the toxic axonopathies may provide insight into the relationship between neurofilament behaviour and axonal degeneration, hopefully enabling the identification of new targets for therapeutic intervention. Because neurofilament abnormalities are a common feature of many neurodegenerative diseases, advances in this area may have a wider impact beyond toxicological significance.


Subject(s)
Adhesives/toxicity , Amyotrophic Lateral Sclerosis/chemically induced , Axons/drug effects , Carbon Disulfide/toxicity , Hexanes/toxicity , Neurofilament Proteins/drug effects , Nitriles/toxicity , Amyotrophic Lateral Sclerosis/metabolism , Amyotrophic Lateral Sclerosis/pathology , Axons/metabolism , Axons/pathology , Humans , Neurofilament Proteins/metabolism , Neurotoxins/toxicity
16.
Synapse ; 67(4): 189-98, 2013 Apr.
Article in English | MEDLINE | ID: mdl-23184880

ABSTRACT

Minocycline has been recently implicated in protection against focal cerebral ischemia reperfusion (I/R), but the protective effects on neurobehavioral abnormalities remains contradictory. In the present study, we investigate whether minocycline improves axonal regeneration and neurological function recovery by inhibiting the expression of the repulsive guidance molecular A (RGMa) after focal cerebral ischemia reperfusion. Male Sprague-Dawley (SD) rats were subjected to occlusion of the right middle cerebral artery (MCAO) for 2 h and 3 mg kg⁻¹ minocycline was injected intravenously immediately after reperfusion twice a day for 14 days. The staircase test and modified neurological severity score (mNSS) were performed to evaluate functional outcome and blood-brain barrier (BBB) permeability was assessed by Evan's blue dye extravasation (EB) at the expected time point. The expression of RGMa in ischemic cortex was measured by immunohistochemical staining and Western blot 2 weeks after MCAO. Neurofilament protein 200 (NF-200) immunohistochemical staining was used to assess axonal damage. Treatment with minocycline at a dose of 3 mg kg⁻¹ via the caudal vein significantly reduced the extravasation of EB, elevated mNSS and improved forelimb motor function as assessed by the staircase test when compared to the I/R group (P < 0.05). Moreover, axonal regrowth was enhanced in the minocycline treatment group when compared to the I/R group (P < 0.05). In addition, minocycline significantly reduced the expression of RGMa protein 2 weeks after MCAO as assessed by both immunostaining and Western blot. Our studies suggest that early minocycline treatment promotes neurological functional recovery and axonal regeneration in rats after MCAO, which might be mediated by down-regulating RGMa expression.


Subject(s)
Axons/drug effects , Brain Ischemia/metabolism , Membrane Proteins/drug effects , Minocycline/pharmacology , Nerve Regeneration/drug effects , Nerve Tissue Proteins/drug effects , Neuroprotective Agents/pharmacology , Reperfusion Injury/metabolism , Animals , Axons/physiology , Cerebral Cortex/drug effects , Cerebral Cortex/metabolism , GPI-Linked Proteins , Male , Membrane Proteins/metabolism , Middle Cerebral Artery , Nerve Regeneration/physiology , Nerve Tissue Proteins/metabolism , Neurofilament Proteins/drug effects , Neurofilament Proteins/metabolism , Rats , Rats, Sprague-Dawley , Recovery of Function/drug effects
17.
Article in Chinese | MEDLINE | ID: mdl-22931767

ABSTRACT

OBJECTIVE: To investigate the effect of 2,5-hexanedione (HD) on degradation of low-molecular-weight neurofilaments (NF-L) in nervous tissue of rats, and to explore the molecular mechanism of n-hexane neuropathy. METHODS: Fifty male Wistar rats were randomly divided into one-week poisoning group (n = 10), two-week poisoning group (n = 10), three-week poisoning group (n = 10), four-week poisoning group (n = 10), and control group (n = 10). In the four poisoning groups, a rat model of n-hexane neuropathy was established by intraperitoneal injection of HD (400 mg/kg/d). The change in the sciatic nerve ultrastructure of each rat was observed under an electron microscope. The progression of HD-induced peripheral neuropathy was evaluated using a gait scoring system. The degradation rates of NF-L in the sciatic nerve and spinal cord of each rat were measured by Western Blotting. RESULTS: The rats showed decrease in muscle strength and abnormal gait after two weeks of HD poisoning and mild or moderate paralysis after four weeks of HD poisoning. The sciatic nerve showed degenerative change, according to electron microscope observation. Compared with the control group, the two-week poisoning group, three-week poisoning group, and four-week poisoning group had the NF-L degradation rates decreased by 25.8%, 70.4%, and 69.7%, respectively, in the supernatant fraction of sciatic nerve, and by 14.7%, 64.6%, and 67.3%, respectively, in the sediment fraction of sciatic nerve, all showing a significant difference (P < 0.01). Compared with the control group, the one-week poisoning group had the NF-L degradation rate decreased by 33.87% in the supernatant fraction of spinal cord, the four-week poisoning group had the NF-L degradation rate increased by 16.2% in the supernatant fraction of spinal cord, and the one-week poisoning group and two-week poisoning group had the NF-L degradation rates decreased by 46.3% and 13.0% in the sediment fraction of spinal cord, all showing a significant difference (P < 0.01). CONCLUSION: HD poisoning significantly inhibits NF-L degradation in the sciatic nerve, which may be associated with NF degeneration and accumulation in the axons of patients with n-hexane neuropathy.


Subject(s)
Hexanes/poisoning , Hexanones/pharmacology , Nerve Tissue/physiopathology , Neurofilament Proteins/metabolism , Sciatic Nerve/physiopathology , Animals , Male , Nerve Tissue/drug effects , Nerve Tissue/metabolism , Neurofilament Proteins/drug effects , Rats , Rats, Wistar , Sciatic Nerve/drug effects , Sciatic Nerve/metabolism
18.
Glia ; 60(3): 422-31, 2012 Mar.
Article in English | MEDLINE | ID: mdl-22161990

ABSTRACT

Neuroaxonal degeneration is a pathological hallmark of multiple sclerosis (MS) contributing to irreversible neurological disability. Pathological mechanisms leading to axonal damage include autoimmunity to neuronal antigens. In actively demyelinating lesions, myelin is phagocytosed by microglia and blood-borne macrophages, whereas the fate of degenerating or damaged axons is unclear. Phagocytosis is essential for clearing neuronal debris to allow repair and regeneration. However, phagocytosis may lead to antigen presentation and autoimmunity, as has been described for neuroaxonal antigens. Despite this notion, it is unknown whether phagocytosis of neuronal antigens occurs in MS. Here, we show using novel, well-characterized antibodies to axonal antigens, that axonal damage is associated with HLA-DR expressing microglia/macrophages engulfing axonal bulbs, indicative of axonal damage. Neuronal proteins were frequently observed inside HLA-DR(+) cells in areas of axonal damage. In vitro, phagocytosis of neurofilament light (NF-L), present in white and gray matter, was observed in human microglia. The number of NF-L or myelin basic protein (MBP) positive cells was quantified using the mouse macrophage cell line J774.2. Intracellular colocalization of NF-L with the lysosomal membrane protein LAMP1 was observed using confocal microscopy confirming that NF-L is taken up and degraded by the cell. In vivo, NF-L and MBP was observed in cerebrospinal fluid cells from patients with MS, suggesting neuronal debris is drained by this route after axonal damage. In summary, neuroaxonal debris is engulfed, phagocytosed, and degraded by HLA-DR(+) cells. Although uptake is essential for clearing neuronal debris, phagocytic cells could also play a role in augmenting autoimmunity to neuronal antigens.


Subject(s)
Microglia/physiology , Multiple Sclerosis/pathology , Neurons/pathology , Phagocytosis/physiology , Adult , Aged , Aged, 80 and over , Animals , Cathepsin D/pharmacology , Cathepsins/pharmacology , Cells, Cultured , Dose-Response Relationship, Drug , Female , HLA-DR Antigens/metabolism , Humans , Male , Mice , Microglia/drug effects , Microscopy, Confocal , Middle Aged , Multiple Sclerosis/cerebrospinal fluid , Myelin Basic Protein/cerebrospinal fluid , Myelin Basic Protein/metabolism , Nerve Fibers, Myelinated/metabolism , Nerve Fibers, Myelinated/pathology , Neurofilament Proteins/cerebrospinal fluid , Neurofilament Proteins/drug effects , Neurofilament Proteins/metabolism , Neurons/drug effects , Phagocytosis/drug effects , Time Factors
19.
Ann Neurol ; 69(1): 83-9, 2011 Jan.
Article in English | MEDLINE | ID: mdl-21280078

ABSTRACT

OBJECTIVE: The impact of present disease-modifying treatments (DMTs) in multiple sclerosis (MS) on nerve injury and reactive astrogliosis is still unclear. Therefore, we studied the effect of natalizumab treatment on the release of 2 brain-specific tissue damage markers into cerebrospinal fluid (CSF) in MS patients. METHODS: CSF samples from 92 patients with relapsing forms of MS were collected in a prospective manner prior to natalizumab treatment and after 6 or 12 months. In 86 cases, natalizumab was used as second-line DMT due to breakthrough of disease activity. The levels of neurofilament light (NFL) and glial fibrillary acidic protein (GFAP) were determined using highly sensitive in-house developed enzyme-linked immunosorbent assays. RESULTS: Natalizumab treatment led to a 3-fold reduction of NFL levels, from a mean value of 1,300 (standard deviation [SD], 2,200) to 400 (SD, 270) ng/l (p < 0.001). The later value was not significantly different from that found in healthy control subjects (350 ng/l; SD, 170; n = 28). Subgroup analysis revealed a consistent effect on NFL release, regardless of previous DMT or whether patients had relapses or were in remission within 3 months prior to natalizumab treatment. No differences between pre- and post-treatment levels of GFAP were detected. INTERPRETATION: Our data demonstrate that natalizumab treatment reduces the accumulation of nerve injury in relapsing forms of MS. It is anticipated that highly effective anti-inflammatory treatment can reduce axonal loss, thereby preventing development of permanent neurological disability.


Subject(s)
Antibodies, Monoclonal/therapeutic use , Axons/pathology , Glial Fibrillary Acidic Protein/cerebrospinal fluid , Integrin alpha4/therapeutic use , Multiple Sclerosis, Relapsing-Remitting/cerebrospinal fluid , Multiple Sclerosis, Relapsing-Remitting/drug therapy , Neurofilament Proteins/cerebrospinal fluid , Adolescent , Adult , Antibodies, Monoclonal, Humanized , Enzyme-Linked Immunosorbent Assay , Female , Glial Fibrillary Acidic Protein/drug effects , Gliosis/chemically induced , Gliosis/prevention & control , Humans , Longitudinal Studies , Male , Middle Aged , Multiple Sclerosis, Chronic Progressive/cerebrospinal fluid , Multiple Sclerosis, Chronic Progressive/drug therapy , Multiple Sclerosis, Relapsing-Remitting/pathology , Natalizumab , Nerve Degeneration/pathology , Nerve Degeneration/prevention & control , Neurofilament Proteins/drug effects , Recurrence , tau Proteins/cerebrospinal fluid , tau Proteins/drug effects
20.
J Neurosci Methods ; 195(2): 143-50, 2011 Feb 15.
Article in English | MEDLINE | ID: mdl-21134399

ABSTRACT

INTRODUCTION: Neurofilament (Nf) aggregates are a common pathological feature of neurodegenerative disorders. Although Nf levels have been investigated as a potential disease biomarker, Nf aggregates may mask Nf epitopes, preventing accurate quantification by immunoassay. Using the SOD1(G93A) mouse model of amyotrophic lateral sclerosis, we developed a method to disrupt Nf aggregates, allowing optimal immunoassay performance. METHODS: Phosphorylated (NfH(SMI35)) and hyperphosphorylated (NfH(SMI34)) Nf levels in plasma from 120-day SOD1(G93A) mice were quantified using an in-house ELISA modified for use with small volumes. Different pre-analytical methods were tested for their ability to solubilize Nf aggregates and immunoblotting was used for qualitative analysis. RESULTS: A 'hook effect' was observed for serially diluted plasma samples quantified using an ELISA originally developed for CSF samples. Immunoblotting confirmed the existence of high molecular-weight NfH aggregates in plasma and the resolving effect of timed urea on these aggregates. Thermostatic (pre-thawing) and chemical (calcium chelators, urea) pre-analytical processing of samples had variable success in disrupting NfH aggregates. Timed urea-calcium chelator incubation yielded the most consistent plasma NfH levels. A one hour sample pre-incubation with 0.5M urea in Barbitone-EDTA buffer at room temperature resolved the "hook effect" without compromising the ELISA. In SOD1(G93A) mice, median levels of NfH(SMI34) were over 10-fold and NfH(SMI35) levels 5-fold greater than controls. CONCLUSION: NfH aggregates can be solubilised and the "hook effect" abolished by a one-hour sample pre-incubation in a urea-calcium chelator-enriched buffer. This method is applicable for quantification of NfH phosphoforms in experimental and disease settings where Nf aggregate formation occurs.


Subject(s)
Immunoassay/methods , Neurofilament Proteins/blood , Plasma/metabolism , Superoxide Dismutase/blood , Age Factors , Animals , Chelating Agents/pharmacology , Dose-Response Relationship, Drug , Edetic Acid/pharmacology , Egtazic Acid/pharmacology , Enzyme-Linked Immunosorbent Assay/methods , Humans , Mice , Mice, Transgenic , Mutation , Neurofilament Proteins/drug effects , Plasma/drug effects , Superoxide Dismutase/genetics , Urea/pharmacology
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