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Neurofilament light as a marker for neurodegenerative diseases
Umeå University, Faculty of Medicine, Department of Clinical Microbiology, Immunology/Immunchemistry.
2004 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Neurofilaments are the main cytoskeletal constituents in neuronal cells. They are belived to be important for maintaining the structural integrity and calibre of axons and dendrites thereby influencing the conduction velocity of nerve impulses.The neurofilament chains are divided into three groups according to their molecular size, neurofilament light (NF-L), neurofilament medium (NF-M) and neurofilament heavy (NF-H). The neurofilaments are obligate heteropolymers in vivo in which NF-L forms the backbone to which the heavier chains copolymerize to form the 10 nm neurofilament fibre.

Different degenerative processes in the brain raise significant interest owing to the increasing mean age in the western world. Such diseases include amyotrophic lateral sclerosis, vascular dementia, frontal lobe dementia, progressive supra-nuclear paralysis, multiple system atrophy, low pressure hydrocephalus, and multiple sclerosis (MS).

We have been able to generate six highly specific monoclonal antibodies for NF-L, and four independent epitopes were elucidated using Biacore and V8 protease degradation. Antibody 2:1 and 47:3 were selected components in a two-site ELISA assay for detection of NF-L in body fluids owing to their outstanding abililty to bind the antigen. The assay has a least detectable dose of 60 ng/l and a standard range of 60 to 64 000 ng/l. The assay was validated on its ability to detect changes of NF-L levels in CSF in patients with different neurological diseases. These were cerebral infarction, amyotrophic lateral sclerosis, relapsing remitting MS, extrapyramidal symptoms, and late onset Alzheimer’s disease. All the patient groups displayed significantly elevated NF-L levels as compared to the controls. We also tested the assay’s ability to monitor the amount of axonal breakdown in an animal model of MS. The NF-L levels were found to be elevated in rodents with chronic experimental autoimmune encephalomyelitis, giving a possible tool for monitoring new treatment strategies for axonal protection in MS. When studying a large population based MS material, we found axonal breakdown to be present early in the disease course and the breakdown was observed both in active relapse and clinically stable disease, indicative of ongoing neurodegeneration. NF-L levels were correlated to progression index, that is, high NF-L levels detected early in disease predict a fast progression of the disease. The amount of glial fibrillary acidic protein, a cytoskeletal protein found in astrocytes, was also quantified and was shown to be a good marker for the more progressive MS subtypes, that is, primary progressive and secondary progressive disease, indicating formation of astrocytic scars and activation of astrocytes.

The test dealt with in this thesis has the potential to identify the slow chronic degenerative diseases with progressive disappearance of nerve cells and their large myelinated axons. There is a significant need clinically to be able to quantify such types of cell degeneration in relation to the progressive disappearance of nerve functions and to relate these different conditions to treatment regimens, disease progress, and prognosis.

Place, publisher, year, edition, pages
Umeå: Klinisk mikrobiologi , 2004. , 69 p.
Series
Umeå University medical dissertations, ISSN 0346-6612 ; 930
Keyword [en]
Immunology, Neurofilament protein, ELISA, Cerebrospinal fluid, Neurodegenerative diseases
Keyword [sv]
Immunologi
National Category
Immunology in the medical area
Research subject
Immunology
Identifiers
URN: urn:nbn:se:umu:diva-357ISBN: 91-7305-769-X (print)OAI: oai:DiVA.org:umu-357DiVA: diva2:143223
Public defence
2004-12-03, E04, 6E, Norrlands Universitetssjukhus, Umeå, 09:00
Opponent
Supervisors
Available from: 2004-11-09 Created: 2004-11-09 Last updated: 2010-08-10Bibliographically approved
List of papers
1. Monoclonal antibodies selective for low molecular weight neurofilaments
Open this publication in new window or tab >>Monoclonal antibodies selective for low molecular weight neurofilaments
2002 (English)In: Hybridoma and Hybridomics, ISSN 1536-8599, Vol. 21, no 1, 53-59 p.Article in journal (Refereed) Published
Abstract [en]

Neurofilaments are necessary for the maintenance of axonal caliber and structural organization of nerve cells. The low molecular weight form of neurofilament, the neurofilament light protein, which serves as the core of the filament, was used as immunogen for generation of hybridomas with selective reactivity with this form of the filament. Six hybridomas, out of approximately 100 tested clones, were highly discriminatory. All involved epitopes were localized to the rod region of the antigen, as determined by alpha-chymotrypsin digestion of the purified filament and enzymatic peptide mapping. Synthetic peptides (20 mers) covering the entire rod region did not react with the antibodies. A phage display peptide library was used to identify four consensus sequences for the antibodies. The results indicate that all epitopes were of conformational type. Pair-wise BIAcore data furthermore indicated that the epitopes were independent. The access to such specific reagents is a prerequisite for further elucidation of the biology of the low molecular weight form of neurofilaments proteins.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:umu:diva-4216 (URN)10.1089/15368590252917647 (DOI)11991817 (PubMedID)
Available from: 2004-11-09 Created: 2004-11-09 Last updated: 2010-08-10Bibliographically approved
2. Elevated neurofilament levels in neurological diseases
Open this publication in new window or tab >>Elevated neurofilament levels in neurological diseases
2003 (English)In: Brain Research, ISSN 0006-8993, E-ISSN 1872-6240, Vol. 987, no 1, 25-31 p.Article in journal (Refereed) Published
Abstract [en]

Neurofilaments, a major cytoskeletal constituent of neuronal cells, can be released into the cerebrospinal fluid during several neurodegenerative diseases. By means of a new sensitive ELISA capable of measuring 60 ng/l of neurofilament light, significant elevations were observed for different neurological disorders. Cerebral infarction presented levels of 19800+/-9100 ng/l, amyothropic lateral sclerosis 3600+/-1200 ng/l, 'relapsing-remitting' MS 2500+/-1500 ng/l, extrapyramidal symptoms 1100+/-300 ng/l, late onset AD 300+/-100 ng/l and vascular dementia 1400+/-800 ng/l. In patients with no signs of neurological diseases the upper normal level and cut-off values was determined to be below 100 ng/l. NF-L determinations will be a valuable complement in identifying neuronal degradation and can be used clinically for diagnostic and monitoring purposes.

Keyword
Neurofilament protein, ELISA, Cerebrospinal fluid, Neurodegenerative disease
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:umu:diva-4217 (URN)10.1016/S0006-8993(03)03219-0 (DOI)14499942 (PubMedID)
Available from: 2004-11-09 Created: 2004-11-09 Last updated: 2017-12-14Bibliographically approved
3. Cerebrospinal fluid levels of neurofilament light in chronic experimental autoimmune encephalomyelitis
Open this publication in new window or tab >>Cerebrospinal fluid levels of neurofilament light in chronic experimental autoimmune encephalomyelitis
2005 (English)In: Brain Research Bulletin, ISSN 0361-9230, E-ISSN 1873-2747, Vol. 67, no 4, 264-268 p.Article in journal (Refereed) Published
Abstract [en]

Experimental autoimmune encephalomyelitis (EAE) induced by myelin oligodendrocyte glycoprotein (MOG) is a chronic relapsing-remitting animal model of multiple sclerosis (MS). Neurofilament light (NF-L), a structural protein expressed in neuronal cells can be used to quantify the amount of neuronal damage in MS patients. An immunoassay was used to measure levels of neurofilament light in cerebrospinal fluid (CSF) in rats with myelin oligodendrocyte glycoprotein-induced EAE. Significantly increased levels of neurofilament were found in the immunized animals compared to the controls, strengthening the similarities in the diseases and the progression pattern between the animal model and MS. The turnover of NF-L during this disease is increased since significantly elevated levels also were identified in the spinal cord of the diseased animals and immunohistochemistry gave support for this observation. Monitoring neurofilament levels in EAE can be used to follow disease progression and effects of therapy.

Keyword
Encephalomyelitis, Cerebrospinal fluid, Autoimmune
National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:umu:diva-4218 (URN)10.1016/j.brainresbull.2005.06.031 (DOI)16182933 (PubMedID)
Available from: 2004-11-09 Created: 2004-11-09 Last updated: 2017-12-14Bibliographically approved
4. Neurofilament light and glial fibrillary acidic protein in multiple sclerosis
Open this publication in new window or tab >>Neurofilament light and glial fibrillary acidic protein in multiple sclerosis
Show others...
2004 (English)In: Neurology, ISSN 0028-3878, E-ISSN 1526-632X, Vol. 63, no 9, 1586-1590 p.Article in journal (Refereed) Published
Abstract [en]

Objective: To evaluate levels of neurofilament light (NFL) and glial fibrillary acidic protein (GFAP) in CSF from patients with multiple sclerosis (MS) in relation to clinical progress of the disease.

Methods: CSF levels of NFL and GFAP were determined by sensitive ELISAs in 99 patients with different subtypes of MS, classified in terms of “ongoing relapse” or “clinically stable disease,” and 25 control subjects. Levels were compared with paraclinical data such as immunoglobulin G index and inflammatory cell count in the CSF, and the levels were related to Expanded Disability Status Scale score and progression index at clinical follow-up evaluations later in the disease course.

Results: NFL and GFAP levels were elevated in MS patients as compared with control subjects (p < 0.001). The NFL levels were higher at relapses, whereas GFAP levels were unaffected. High NFL levels correlated with progression in patients with an active relapse (r = 0.49; p < 0.01) and in clinically stable patients (r = 0.29; p < 0.05). GFAP correlated to progression in the total patient cohort (r = 0.24; p < 0.05). Moreover, a strong correlation between NFL levels and inflammatory cell counts was evident in the group of patients with an ongoing relapse (r = 0.52; p = 0.001).

Conclusions: CSF levels of neurofilament light and glial fibrillary acidic protein may have prognostic value in multiple sclerosis.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:umu:diva-4219 (URN)15534240 (PubMedID)
Available from: 2004-11-09 Created: 2004-11-09 Last updated: 2017-12-14Bibliographically approved

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