Multiple Sclerosis: Keir G

Freedman MS, Thompson EJ, Deisenhammer F, Giovannoni G, Grimsley G, Keir G, Ohman S, Racke MK, Sharief M, Sindic CJ, Sellebjerg F, Tourtellotte WW. · Department of Medicine (Neurology), University of Ottawa, Multiple Sclerosis Research Clinic, The Ottawa Hospital, Ottawa, Ontario, Canada. · Arch Neurol. · Pubmed #15956157 links to  free full text

Abstract: New criteria for the diagnosis of multiple sclerosis (MS) were published as the result of an internationally formed committee. To increase the specificity of diagnosis and to minimize the number of false diagnoses, the committee recommended the use of both clinical and paraclinical criteria, the latter involving information obtained from magnetic resonance imaging, evoked potentials, and cerebrospinal fluid (CSF) analysis. Although rigorous magnetic resonance imaging requirements were provided, the "new criteria paper" fell short in terms of guidelines as to how the CSF analysis should be performed and simply equated the IgG index with isoelectric focusing, without any justification. The spectrum of parameters analyzed and methods for CSF analysis differ worldwide and often yield variable results in terms of sensitivity, specificity, accuracy, and reliability, with no decided "optimal" CSF test for the diagnosis of MS. To address this question specifically, an international panel of experts in MS and CSF diagnostic techniques was convened and the result was this article, representing a consensus of all the participants. These recommendations for establishing a standard for the evaluation of CSF in patients suspected of having MS should greatly complement the new criteria in ensuring that a correct diagnosis of MS is being made.

Lim ET, Sellebjerg F, Jensen CV, Altmann DR, Grant D, Keir G, Thompson EJ, Giovannoni G. · Department of Neuroinflammation, Institute of Neurology, University College London, Queen Square, London WC1N 3BG, UK. · Mult Scler. · Pubmed #16193890 No free full text.

Abstract: The objectives of this study were (1) to determine how cerebrospinal fluid (CSF) neurofilament heavy chain (NfH(SM134) and NfH(SM135)) levels relate to clinical outcome in optic neuritis (ON) and multiple sclerosis (MS) relapse patients treated with high dose oral methylprednisolone; and (2) to correlate neurofilament and myelin basic protein (MBP) concentrations, particularly as the latter was previously associated with clinical disability. Fifty subjects participated in two double-blind, randomized, placebo-controlled clinical trials. Eight/18 patients in the ON trial and 15/32 subjects in the MS attack trial were treated with oral methylprednisolone. In the MS attack trial group, CSF NfH(SM134) and NfH(SM135) measured at week 3 and deltaCSF NfH(SMI34) levels from baseline to week 3 were predictive of clinical outcome at week 8 and 52. In the ON group, no such association was seen. When both groups were combined, baseline CSF NfH(SHM134) and NfH(SM135) correlated positively with baseline enhancing lesion volume (ELV) (r(s) =0.50, P <0.01 and rS =0.53, P <0.01, respectively). Levels of NfH(SM135) at baseline and week 3 also strongly correlated with the MBP concentration. This study supports the view that acute inflammation in ON and MS results in axonal pathology and that the latter has a role in determining functional impairment.

Lim ET, Petzold A, Leary SM, Altmann DR, Keir G, Thompson EJ, Miller DH, Thompson AJ, Giovannoni G. · Department of Neuroinflammation, Institute of Neurology, University College London, Queen Square, London, WC1N 3BG, UK. · J Negat Results Biomed. · Pubmed #15482599 links to  free full text

Abstract: S100B belongs to a family of calcium-binding proteins implicated in intracellular and extracellular regulatory activities. This study of serum S100B in primary progressive multiple sclerosis (PPMS) is based on data obtained from a randomized, controlled trial of Interferon beta-1a in subjects with PPMS. The key questions were whether S100B levels were associated with either disability or MRI findings in primary progressive MS and whether Interferon beta-1a has an effect on their S100B levels. Serial serum S100B levels were measured using an ELISA method. The results demonstrated that serum S100B is not related to either disease progression or MRI findings in subjects with primary progressive MS given Interferon beta-1a. Furthermore there is no correlation between S100B levels and the primary and secondary outcome measures.

Petzold A, Brassat D, Mas P, Rejdak K, Keir G, Giovannoni G, Thompson EJ, Clanet M. · Institute of Neurology, Department of Neuroimmunology, Queen Square, London WC1N 3BG, UK. · Mult Scler. · Pubmed #15222692 No free full text.

Abstract: BACKGROUND: This study aimed to investigate if treatment response could retrospectively be related to inflammatory or axonal pathology as measured by plasma surrogate markers. METHODS: In this 1-year observational study 30 multiple sclerosis (MS) patients with relapsing-remitting disease were treated with intramuscular IFNbeta-1a or subcutaneous IFNbeta-1b. Responders and nonresponders were defined according to clinical and magnetic resonance imaging criteria. The control group consisted of 14 healthy subjects. Plasma levels of surrogate markers for inflammation (nitric oxide metabolites (NOx)), astrocytic activation (S100B) and axonal damage (NfH(SM135)) were measured using standard assays. RESULTS: There were 11 nonresponders and 19 responders to IFNbeta treatment. Median S100B levels were elevated in a higher proportion of treatment responders (63%, 42.9 pg/mL) compared to nonresponders (18%, 11.7 pg/mL, P < 0.05, Fisher's exact test) and controls (0%, 2 pg/mL, P < 0.001). Levels of NOx were found to be more frequently elevated in nonresponders (72%, 39 microM) compared to healthy controls (0%, 37 microM, P < 0.05). Levels of NfH(SM135) were more frequently elevated in responders (58%, 300 pg/mL, P < 0.001) and nonresponders (72%, 500 pg/mL, P < 0.001) compared to controls (0%, 4.5 pg/mL). CONCLUSION: Patients with relapsing-remitting MS who had surrogate marker supported evidence for astrocytic activation responded more frequently to treatment with IFNbeta.

Davies G, Keir G, Thompson EJ, Giovannoni G. · Department of Neuroinflammation, Institute of Neurology, University College London. · Neurology. · Pubmed #12682325 No free full text.

Abstract: BACKGROUND: Intrathecal oligoclonal band synthesis occurs in 95% of patients with clinically definite MS but may also occur in the context of CNS infection and other inflammatory conditions. By contrast, the significance of an intrathecal synthesis of a monoclonal band remains uncertain. Previously, an association between a single intrathecal band and CNS lymphoma has been reported but a relationship has also been shown with diagnoses more usually associated with an oligoclonal pattern. At present, it is not known whether a single band will convert to an oligoclonal response with time. METHODS: Data were obtained from patients who had CSF and serum analyzed by isoelectric focusing (IEF) at the authors' institutions over a 6-year period. Clinical details were acquired for those who underwent repeat lumbar puncture after an initial CSF examination revealed an intrathecal monoclonal immunoglobulin G band. RESULTS: Of the 31 patients identified as having an initial intrathecal monoclonal band, clinical details were available for 27. Of those, 9 were found on subsequent lumbar puncture to have developed an intrathecal oligoclonal response. CONCLUSIONS: Among those subjects who developed oligoclonal bands, there was a propensity for either a diagnosis of MS or clinically isolated syndromes due to demyelination. In the 18 subjects who either reverted to having normal CSF IEF or continued to exhibit only the monoclonal band, no cases of MS were encountered. Importantly, one of these had cerebral lymphoma.

Amorini AM, Petzold A, Tavazzi B, Eikelenboom J, Keir G, Belli A, Giovannoni G, Di Pietro V, Polman C, D'Urso S, Vagnozzi R, Uitdehaag B, Lazzarino G. · Institute of Biochemistry and Clinical Biochemistry, Catholic University of Rome Sacro Cuore, Rome, Italy. · Clin Biochem. · Pubmed #19341721 No free full text.

Abstract: OBJECTIVES: In this study, the concentrations of uric acid, purine profile and creatinine in samples of cerebrospinal fluid and serum of multiple sclerosis (MS) patients were measured by HPLC and compared with corresponding values recorded in patients without MS (cerebrospinal fluid) and healthy subjects (serum). DESIGN AND METHODS: All samples were deproteinized with ultrafiltration (which ensures minimal sample manipulation and efficient protein removal) and then assayed for the synchronous HPLC separation of uric acid, hypoxanthine, xanthine, inosine, adenosine, guanosine and creatinine. RESULTS: The values of all compounds assayed were significantly higher in both biological fluids of MS patients with respect to values measured in controls. In particular, serum hypoxanthine, xanthine, uric acid and sum of oxypurines were, respectively, 3.17, 3.11, 1.23 and 1.27-fold higher in these patients than corresponding values recorded in controls (p<0.001). CONCLUSIONS: Differently from what previously reported, we here demonstrate that all purine compounds, including uric acid, are elevated in biological fluids of MS patients. Reinforced by the trend observed for creatinine, this corroborates the notion of sustained purine catabolism, possibly due to imbalance in ATP homeostasis, under these pathological conditions. These results cast doubt on the hypothesis that uric acid is depleted in MS because of increased oxidative stress, rather suggesting that this disease causes a generalized increase in purine catabolism. As observed in other pathological states, uric acid, purine compounds and creatinine, can be considered markers of metabolic energy imbalance rather than of reactive oxygen species, even in MS.

Petzold A, Gveric D, Groves M, Schmierer K, Grant D, Chapman M, Keir G, Cuzner L, Thompson EJ. · Department of Neuroinflammation, Institute of Neurology, Queen Square, London, UK. · Exp Neurol. · Pubmed #18619438 links to  free full text

Abstract: AIMS: Axonal pathology extends to the axonal cytoarchitecture leaving its signature on axoskeletal proteins. This study investigated whether neurofilament (NfH) phosphorylation would relate to the dynamics of axonal pathology in multiple sclerosis (MS). METHODS: NfH phosphoforms (SMI32, SMI34, SMI35) were quantified by ELISA from microdissected samples of control and MS brain and spinal cord. Individual axons were analysed by electron microscopy, densitometrically and morphologically in adjacent tissue sections. Experiments were carried out pre- and post enzymatic dephosphorylation. RESULTS: In control tissue a rostro-caudal gradient of NfH indicated an increase in axonal density from the brain gray matter towards the spinal cord. The highest levels of phosphorylated and hyperphosphorylated NfH were found in acute lesions of brain and spinal cord, in contrast to chronic lesions where levels were lower than in white matter, consistent with axonal loss. Dephosphorylated NfH was higher, but less densly packed in MS white matter axons compared to control tissue. CONCLUSIONS: The findings suggest that a less organised/compact axoskeleton or impaired axonal transport may represent an early sign of axonal pathology within the normal appearing white matter in MS. Subsequently a proportional increase of dephosphorylated NfH, aberrant phosphorylation and/or aggregation may occur whilst the protein is transported through the white matter towards the MS plaque, where hyperphosphorylated NfH dominates.

Keir G, Barrio S, Thompson EJ. · Department of Neuroinflammation, Institute of Neurology, Queen Square, London WC1 N 3BG, UK. · Ann Clin Biochem. · Pubmed #18583626 No free full text.

Abstract: BACKGROUND: Detection of local synthesis of IgG within the central nervous system is important for the diagnosis of brain inflammatory diseases such as multiple sclerosis. This is typically done by comparing the amounts of IgG in serum and parallel cerebrospinal fluid (CSF). Although there have been well-described problems with qualitative versus quantitative measurements of abnormal IgG, such as in myeloma paraproteins, similar difficulties are also found with CSF IgG. METHODS: Traditional quantitative analysis of IgG by rate nephelometry was followed by separation of the IgG using isoelectric focusing and then either silver stain or immunofixation. Finally, quantitative analysis was performed by scanning densitometry using public domain software downloaded from the National Institutes of Health. RESULTS: We report here the major discrepancies that can occur with CSF IgG between the silver stain versus the IgG stain. CONCLUSIONS: We concur with the earlier recommendation that qualitative separation followed by densitometric estimation of enzyme-linked immunofixation is also more useful than simple quantitative nephelometric analysis followed by silver staining in the detection of local synthesis of IgG, analogous to the earlier work on paraproteins.

Hawkes CH, Giovannoni G, Keir G, Cunnington M, Thompson EJ. · Essex Neuroscience Centre, Oldchurch Hospital, Essex, UK. · Acta Neurol Scand. · Pubmed #17083334 No free full text.

Abstract: BACKGROUND: It has been proposed that multiple sclerosis (MS) might be a sexually transmitted disorder. There is evidence that seropositivity to herpes simplex virus type 2 (HSV-2) correlates well with the number of sexual partners. Accordingly, a raised overall HSV-2 seroprevalence in MS would lend support to this theory. MATERIALS AND METHODS: Serum from 497 UK subjects with clinically definite MS was tested for antibodies to HSV-2 and compared with matched historical controls from within and outside London, blood donors and genito-urinary medicine (GUM) clinics. RESULTS: The unadjusted MS seropositivity rate was 14%. HSV-2 seroprevalence in MS patients aged 35-64 years was significantly higher overall compared with a non-London general population in an unadjusted comparison. HSV-2 seroprevalence in London MS patients compared with London blood donors was significantly greater irrespective of age, but the MS seropositive rate was lower than GUM clinic attenders. In a logistic regression analysis, increased age, female sex and MS diagnosis all independently increased the odds of seropositivity after adjustment for each other. CONCLUSION: It is concluded that there is increased likelihood of HSV-2 exposure in patients with MS and this may indicate a higher than average number of partners.

Petzold A, Eikelenboom MI, Keir G, Polman CH, Uitdehaag BM, Thompson EJ, Giovannoni G. · Department of Neuroinflammation, Institute of Neurology, Queen Square, London, UK. · Mult Scler. · Pubmed #16764346 No free full text.

Abstract: This study investigated whether the new Global Multiple Sclerosis Severity Scale (MSSS) correlated with cerebrospinal fluid biomarkers for axonal and glial pathology. The MSSS correlated with the phosphorylated neurofilament heavy chain (NfH-SM135, R=0.44, P=0.016). The degree of neurofilament phosphorylation (ratio NfH-SM134 to NfH-SM135) was 8-fold higher in severely (median MSSS 6.5) versus mildly (MSSS 3.2) disabled patients (7.3 versus 0.9, P = 0.03). The MSSS may provide a statistically powerful tool for comparing overall disease severity and be useful for validating the biomarker concept in MS.

Lim ET, Berger T, Reindl M, Dalton CM, Fernando K, Keir G, Thompson EJ, Miller DH, Giovannoni G. · Department of Neuroinflammation, Institute of Neurology, University College London, Queen Square, London WC1N 3BG, UK. · Mult Scler. · Pubmed #16042235 No free full text.

Abstract: This study investigates whether the presence of serum and plasma anti-myelin oligodendrocyte glycoprotein (MOG) and anti-myelin basic protein (MBP) in patients presenting with a clinically isolated syndrome compatible with demyelination (CIS) predicts early conversion to multiple sclerosis (MS). Forty-seven patients with CIS (46 with optic neuritis) had anti-MOG and anti-MBP antibodies analysed at baseline, and clinical and magnetic resonance imaging assessments. There was no evidence that the MS status based on either the McDonald or Poser criteria relates to the antibody status.

Petzold A, Eikelenboom MJ, Keir G, Grant D, Lazeron RH, Polman CH, Uitdehaag BM, Thompson EJ, Giovannoni G. · Institute of Neurology, Department of Neuroinflammation, Queen Square, London WC1N 3BG, UK. · J Neurol Neurosurg Psychiatry. · Pubmed #15654034 links to  free full text

Abstract: BACKGROUND: Neurofilament phosphoforms (Nf) are principal components of the axoskeleton released during axonal injury. Cerebrospinal fluid (CSF) levels of Nf phosphoforms might be useful surrogate markers for disability in multiple sclerosis (MS), aid in distinguishing clinical subtypes, and provide valuable prognostic information. METHOD: Thirty four patients with MS were included in a three year follow up study along with 318 controls with other non-inflammatory neurological diseases. CSF levels of two Nf heavy chain (NfH) phosphoforms (NfH(SMI35), NfH(SMI34)) were quantified at baseline and three year follow up using new ELISA techniques. Levels of NfH phosphoforms, the degree of phosphorylation (NfH(SMI34):NfH(SMI35) ratio), and changes in NfH levels between baseline and follow up (Delta NfH) were related to the clinical phenotype (RR or SP/PP), to three clinical scales (Kurtzke's EDSS, ambulation index (AI), and nine hole peg test (9HPT)), and to progression of disability. RESULTS: A significantly higher proportion (59%) of patients with SP/PPMS experienced an increase in NfH(SMI35) levels between baseline and follow up compared with those with RRMS (14%, p<0.05). CSF NfH(SMI34) levels at baseline were higher in patients with SP/PP (11 pg/ml) compared with RR (7 pg/ml, p<0.05) and NfH(SMI35) levels were higher at follow up in SP/PP (129 pg/ml) compared with levels below assay sensitivity in RR (p<0.05). NfH(SMI35) correlated with the EDSS (r(s) = 0.54, p<0.01), the AI (r(s) = 0.42, p<0.05), and the 9HPT (r(s) = 0.59, p<0.01) at follow up. CONCLUSION: The increase in NfH during the progressive phase of the disease together with the correlation of NfH(SMI35) with all clinical scales at follow up suggests that cumulative axonal loss is responsible for sustained disability and that high NfH(SMI35) levels are a poor prognostic sign.

Lim ET, Grant D, Pashenkov M, Keir G, Thompson EJ, Söderström M, Giovannoni G. · Department of Neuroinflammation, Institute of Neurology, Queen Square, London WC1N 3BG, UK. · Mult Scler. · Pubmed #15222688 No free full text.

Abstract: This study evaluates levels of cerebrospinal fluid (CSF) brain-specific proteins (BSP) in subjects with optic neuritis (ON) who are at high risk of progression to multiple sclerosis (MS). Forty-one subjects had acute ON and 17 subjects with other neurological diseases (OND) served as controls. Twenty-one subjects with ON had white matter lesions on magnetic resonance imaging (MRI) and intrathecal synthesis of oligoclonal IgG bands (OB) consistent with being at high risk of progression to MS; eight of whom later were diagnosed with clinically definite MS (CDMS). Levels of S100B, ferritin and two neurofilament heavy chain phosphoforms (NfH(SM134) and NfH(SM135)) were analysed using ELISA technique. A putative index of 'axonal health' was expressed as a ratio of NfH(SM134) to NfH(S135). NfH(SM134) and the NfH(SM134:SM135) were significantly elevated in subjects with ON compared to controls. No significant differences in levels of CSF BSP were seen between ON subjects with CDMS plus those at high risk of progression to MS and ON subjects with normal MRI and negative CSF analysis. In conclusion, there is evidence of axonal damage in subjects who present with ON, which is independent of the diagnosis of CDMS.

Petzold A, Eikelenboom MJ, Gveric D, Keir G, Chapman M, Lazeron RH, Cuzner ML, Polman CH, Uitdehaag BM, Thompson EJ, Giovannoni G. · Department of Neuroinflammation, Institute of Neurology, Queen Square, London, UK. · Brain. · Pubmed #12076997 links to  free full text

Abstract: Disease progression in multiple sclerosis occurs within the interface of glial activation and gliosis. This study aimed to investigate the relationship between biomarkers of different glial cell responses: (i) to disease dynamics and the clinical subtypes of multiple sclerosis; (ii) to disability; and (iii) to cross-validate these findings in a post-mortem study. To address the first goal, 51 patients with multiple sclerosis [20 relapsing remitting (RR), 21 secondary progressive (SP) and 10 primary progressive (PP)] and 51 neurological control patients were included. Disability was assessed using the ambulation index (AI), the Expanded Disability Status Scale score (EDSS) and the 9-hole PEG test (9HPT). Patients underwent lumbar puncture within 7 days of clinical assessment. Post-mortem brain tissue (12 multiple sclerosis and eight control patients) was classified histologically and adjacent sites were homogenized for protein analysis. S100B, ferritin and glial-fibrillary acidic protein (GFAP) were quantified in CSF and brain-tissue homogenate by ELISA (enzyme-linked immunosorbent assay) techniques developed in-house. There was a significant trend for increasing S100B levels from PP to SP to RR multiple sclerosis (P < 0.05). S100B was significantly higher in RR multiple sclerosis than in control patients (P < 0.01), whilst ferritin levels were significantly higher in SP multiple sclerosis than in control patients (P < 0.01). The S100B : ferritin ratio discriminated patients with RR multiple sclerosis from SP, PP or control patients (P < 0.05, P < 0.01 and P < 0.01, respectively). Multiple sclerosis patients with poor ambulation (AI > or =7) or severe disability (EDSS >6.5) had significantly higher CSF GFAP levels than less disabled multiple sclerosis or control patients (P < 0.01 and P < 0.001, respectively). There was a correlation between GFAP levels and ambulation in SP multiple sclerosis (r = 0.57, P < 0.01), and between S100B level and the 9HPT in PP multiple sclerosis patients (r = -0.85, P < 0.01). The post-mortem study showed significantly higher S100B levels in the acute than in the subacute plaques (P < 0.01), whilst ferritin levels were elevated in all multiple sclerosis lesion stages. Both GFAP and S100B levels were significantly higher in the cortex of multiple sclerosis than in control brain homogenate (P < 0.001 and P < 0.05, respectively). We found that S100B is a good marker for the relapsing phase of the disease (confirmed by post-mortem observation) as opposed to ferritin, which is elevated throughout the entire course. GFAP correlated with disability scales and may therefore be a marker for irreversible damage. The results of this study have broad implications for finding new and sensitive outcome measures for treatment trials that aim to delay the development of disability. They may also be considered in future classifications of multiple sclerosis patients.

Villar LM, González-Porqué P, Masjuán J, Alvarez-Cermeño JC, Bootello A, Keir G. · Department of Immunology, Hospital Ramón y Cajal, Madrid, Spain. · J Immunol Methods. · Pubmed #11684132 No free full text.

Abstract: We have developed a sensitive and specific isoelectrofocusing (IEF) method for the detection of oligoclonal (OGC) IgM in the cerebrospinal fluid (CSF) and sera. In this procedure, ampholytes, in the range pH 5-8, were used to improve the resolution, and serum was diluted in saline to avoid IgM precipitation. Samples were treated with 50 mM dithiothreitol (DTT) at pH 9.5 to reduce IgM and improve the migration of the protein. Using an anti-IgM labelled with alkaline phosphatase, the limit of detection was found to be 0.1 ng in 5 microl of sample.

Giovannoni G, Morris PR, Keir G. · No affiliation provided · Ann Neurol. · Pubmed #10805348 No free full text.

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