Increased urinary free immunoglobulin light chain excretion in patients with multiple sclerosis.

BACKGROUND: Plasma and B cells are implicated in multiple sclerosis (MS) and produce free light chains (FLC) that are excreted in urine. OBJECTIVE: To confirm that demyelinating diseases (DD) cause increased urinary FLCs. METHOD: Urinary FLC in 50 patients with DD were compared to 20 patients with posterior uveitis (PU), 19 with AIDS, 34 with rheumatoid arthritis (RA) and 19 normal controls (NC). RESULT: Subjects with DD, PU, RA and AIDS have higher urinary FLCs than NC (p<0.01). Urinary FLCs did not correlate with gadolinium-enhancing lesions on MRI. CONCLUSIONS: Urinary FLCs are raised in DD. Further studies are required to see if they correlate with disease activity.

Seroprevalence of herpes simplex virus type 2 in multiple sclerosis.

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.

Increased urinary excretion of nitric oxide metabolites in longitudinally monitored migraine patients.

This study evaluated a relationship between nitric oxide (NO) and migraine attacks in order to gain insight into migraine pathomechanism. The study groups consisted of 12 migraineurs and eight controls. All subjects collected morning urine samples for 40 consecutive days. Urinary NO metabolites, nitrite/nitrate (NO(x)) levels were measured with the vanadium-based assay, whilst creatinine (Cr) and neopterin were determined with high-performance liquid chromatography. The mean urinary NO(x)/Cr ratio and number of NO(x) peaks was significantly greater in the migraine group compared with controls (P = 0.01 and P = 0.007, respectively). In the second approach, high NO(x) values were re-assessed in relation to raised neopterin, a marker of systemic infection or inflammation, and were excluded. The excretion of NO(x) persisted being pulsatile, and migraineurs had more peaks compared with controls (P = 0.01). In seven patients, NO(x) peaks coincided with headache days. This was more frequent than expected by random association in four patients (Monte-Carlo simulation; odds ratios: 2.16-7.77; no overlap of 95% CI). In four patients, NO(x) peaks preceded or followed headache days. Although there is a difference in the pattern of urinary NO(x) excretion between control and migraine populations, the variable temporal association of NO(x) peaks and headaches suggests a complex role of NO in this condition.

CSF neurofilament levels: a potential prognostic marker in Guillain-Barré syndrome.

Long-term morbidity from Guillain-Barré syndrome (GBS) is caused by axonal damage. This prospective study demonstrated that neurofilaments (NfHs), a biomarker for axonal damage, were of prognostic value in GBS. CSF NfH levels correlated with the F score and Medical Research Council summed score and were higher in patients with neurophysiologic evidence of axonal degeneration compared to those without. Pathologically high CSF NfH levels (>0.73 ng/mL) predicted worse motor and functional outcome.

The new global multiple sclerosis severity score (MSSS) correlates with axonal but not glial biomarkers.

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.

Axonal damage and outcome in subarachnoid haemorrhage.

BACKGROUND: On the basis of preliminary evidence from patients with subarachnoid haemorrhage (SAH), axonal degeneration is thought to be an underestimated pathological feature. METHODS: A longitudinal study in 17 patients with aneurysmal SAH. Ventricular CSF was collected daily for up to 14 days. The neurofilament heavy chain(SMI35) (NfH(SMI35), a biomarker for axonal damage) was quantified using a standard ELISA (upper limit of normal 0.73 ng/ml). The primary outcome measure was the Glasgow Outcome Score (GOS) at 3 months. RESULTS: Of 148 samples from patients with SAH, pathologically high NfH levels in the CSF were found in 78 (52.7%) samples, compared with 20 (5%) of 416 samples from the reference population (p<0.0001). A pathological increase in NfH was observed in all patients with a bad outcome (GOS 1-3) compared with 8% of those with a good outcome (GOS 4-5, p<0.0001). This increase typically became significant 7 days after the haemorrhage (p<0.01). The result was confirmed by analysing the individual mean NfH concentrations in the CSF (3.45 v 0.37 ng/ml, p<0.01), and was reinforced by the inverse correlation of NfH in the CSF with the GOS (r = -0.65, p<0.01). Severity of injury was found to be correlated to NfH(SMI35) levels in the CSF (World Federation of Neurological Surgeons, r = 0.63, p<0.01 and Glasgow Coma Score, r = -0.61, p<0.01). CONCLUSION: Patients with SAH thus have secondary axonal degeneration, which may adversely affect their outcome.

Acute axonal damage predicts clinical outcome in patients with multiple sclerosis.

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.

Anti-myelin antibodies do not allow earlier diagnosis of multiple sclerosis.

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.

Antigenic differences between neurological and diabetic patients with anti-glutamic acid decarboxylase antibodies.

Antibodies to glutamic acid decarboxylase (GADAb) are found in Stiff-Person syndrome, type 1 diabetes, cerebellar ataxia and other neurological disorders (such as epilepsy and myoclonus) involving the GABAergic ways. GADAb are usually detected by immunohistochemistry (IHC), radioimmunoassay (RIA) or enzyme-linked immunosorbent assay (ELISA). This study analysed the serum of 14 patients with neurological disorders who were positive by IHC for GADAb. The performance of a commercial RIA was compared with in-house immunoblotting and ELISA methods using recombinant GAD65 (rGAD65). RIA was positive in 14 of 14, immunoblotting was positive in seven of 14 and ELISA in 12 of 14. There was no correlation between the RIA result and the ELISA optical densities. Using a sodium thiocyanate chaotrope system with ELISA to determine antibody affinity, we found no significant correlation between antibody affinity and the RIA result. A consensus should be defined concerning which assay could be used as the gold standard for detecting GADAb. The most intriguing finding was that GAD antibodies from uncomplicated diabetics do not appear to recognize GAD in frozen sections from the rat cerebellum, whereas GAD antibodies from neurologically compromised diabetics do. A working proposal is therefore that type 1 diabetic patients with unusual neurological symptoms should be tested for GADAb both by RIA and IHC.

Lack of serum oligoclonal antibody responses to Chlamydophila pneumoniae in multiple sclerosis.

Chlamydophila pneumoniae has been proposed as an aetiological agent in MS via a mechanism involving molecular mimicry. We undertook to investigate whether the presence of CSF oligoclonal IgG OCB or oligoclonal bands correlated with serum IgG raised against C. pneumoniae. Paired serum and CSF of 19 MS patients and 27 control patients with other neurological diseases were studied by IEF and Western blotting. Only 1 of 19 MS patients had serum antibodies against C. pneumoniae compared with 2 of the 26 control patients. This was not significant, leading us to conclude that this study does not support the theory of an association between C. pneumoniae and MS.

Extracellular fluid S100B in the injured brain: a future surrogate marker of acute brain injury?

The authors describe the measurement of S100B protein in brain extracellular fluid (ECF) of patients with acute brain injury (traumatic brain injury and subarachnoid haemorrhage) using the technique of microdialysis. To our knowledge, this is the first report of S100B measurement in the human brain. Acute Brain Injury (ABI) is a leading cause of death and disability and the need for a practical and sensitive biochemical marker for monitoring these patients is urgent. The calcium binding astrocyte protein, S100B, may be a candidate for this role. Previous serum studies have shown S100B to be a sensitive predictor of mortality and rise in intracranial pressure in ABI, but it has never before been measured directly within the brain. The ECF reflects the local biochemistry of the brain parenchyma, and the use of intracerebral microdialysis opens up the possibility of studying many novel surrogate markers of injury in the laboratory, in addition to the conventional markers it measures at the bedside (lactate, pyruvate, glucose, and glycerol). In this preliminary report of two cases, the authors demonstrate the quantification of S100B in ECF microdialysate, and investigate whether changes in hourly S100B profile can be related to secondary brain injury. It is shown that extracellular concentrations of S100B change markedly in response to secondary brain injury. Further investigation is required to determine whether extracellular S100B measurement in ABI could assist in patient management.

Axonal pathology in subarachnoid and intracerebral hemorrhage.

Electrically active axons degenerate in the presence of nitric oxide (NO) in vitro. High CSF NO concentrations have been observed in patients with hemorrhagic brain injury such as subarachnoid hemorrhage (SAH) and intracerebral hemorrhage (ICH). This study investigated the evidence for axonal injury in SAH and ICH and related this to CSF NO levels. In this study, neurofilament phosphoforms (NfH(SMI34), NfH(SMI35), NfH(SMI38), NfH(SMI310)), surrogate markers for axonal injury, and NO metabolites (nitrate, nitrite = NOx) were measured by ELISA in cerebrospinal fluid (CSF) from patients with SAH and ICH and from a group of controls. Injury severity was classified using the Glasgow Coma Scale, and survival was used as the outcome measure. Compared to the control group, a higher proportion of patients with SAH and ICH had elevated NfH(SMI34) levels from day 0 to day 6 (p < 0.001), elevated NfH(SMI35) levels from day 1 to 6 (p < 0.001), and elevated NfH(SMI310) levels at day 0, 1, 4, and 6 (p < 0.001). The NOx levels were higher in the SAH and ICH patients than in the controls (p < 0.05) and distinguished the non-survivors from the survivors (p < 0.05). No direct correlation was found for NOx with any of the NfH phosphoforms. This study provides evidence for primary and secondary axonal injury in patients with SAH and ICH, with non-survivors also having higher NOx levels. CSF NfH phosphoforms might emerge as a putative surrogate marker for monitoring the development for secondary axonal degeneration in neurocritical care and guiding targeted neuroprotective strategies.

Axonal damage accumulates in the progressive phase of multiple sclerosis: three year follow up study.

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.

CSF nitric oxide metabolites are associated with activity and progression of multiple sclerosis.

OBJECTIVE: To investigate the relationship of CSF and the serum nitric oxide metabolites nitrite and nitrate (NOx) to disease activity and progression in patients with multiple sclerosis (MS). METHODS: The study was divided into cross-sectional and follow-up. In the cross-sectional study, 20 patients with relapsing-remitting (RR), 21 with secondary progressive (SP), and 10 with primary progressive (PP) MS and 14 control subjects were included. Patients were assessed on clinical (Expanded Disability Status Scale [EDSS], Ambulation Index [AI], 9-Hole Peg Test [9-HPT]) and MRI measurements. In the follow-up study, 34 MS patients from the cross-sectional study agreed to be assessed again after an average of 3.0 +/- 0.5 years. NOx was measured using a vanadium-based assay. RESULTS: In the cross-sectional study, CSF NOx was raised in patients with RR-MS (p = 0.001) and PP-MS (p = 0.02) vs controls. Higher CSF NOx levels were found in patients with mild disability (AI < or = 6.0; EDSS < or = 4.0; Multiple Sclerosis Severity Score [MSSS] < or = 4.8) vs patients with advanced disease (AI > 6.0 [p = 0.002]; EDSS > 4.0 [p = 0.02]; MSSS > 4.8 [p = 0.01]). In the subgroup of patients having Gd-enhancing MRI lesions (n = 11), correlation between the volume of enhancement and CSF NOx was found (r = 0.74, p = 0.01). In the follow-up study, patients with disability progression had higher baseline CSF NOx levels than those who were stable on EDSS (p = 0.02) or AI (p = 0.03). A positive correlation was found between baseline CSF NOx and the change in MR T2-weighted lesion load (r = 0.4, p = 0.03). CONCLUSIONS: CSF nitrite and nitrate levels were increased in mildly disabled patients with MS and found to correlate with the volume of Gd-enhanced lesions on MRI. Raised baseline CSF NOx was associated with clinical and MRI progression in MS patients over 3-year follow-up.

Cerebrospinal fluid levels of brain specific proteins in optic neuritis.

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.

Treatment response in relation to inflammatory and axonal surrogate marker in multiple sclerosis.

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.

The spectrum of antineuronal autoantibodies in a series of neurological patients.

The aim of the present study is to identify the range of neurological disorders expressing antineuronal antibodies, evaluate the number of different patterns of reactivity that can be detected, and analyse the contribution of these studies to the identification of subgroups of patients. The records of 882 patients were reviewed and their sera and cerebrospinal fluids tested for antineuronal antibodies. Patients were initially divided into four groups according to suspected clinical diagnosis. Autoantibodies were detected by immunohistochemistry, Western blot of gradient-separated neuronal and recombinant proteins and by RIA. Cerebellar degeneration and sensory neuropathies were the most common neurological disorders in which paraneoplastic-related anti-neuronal antibodies were detected. However, in addition to PCA1/anti-Yo and ANNA1/anti-Hu antibodies, we found other reactivities in six patients with cerebellar degeneration: anti-GAD in three females and atypical in the other cases. The widest range of different anti-neuronal antibodies was detected in patients with peripheral sensory neuropathy. Few patients with Stiff-Person syndrome, temporal lobe epilepsy and myoclonus harboured anti-GAD antibodies. Atypical antibodies were detected in single cases with motor neuron disorder and multiple system atrophy. No anti-neuronal antibodies were detected in patients with neurological complications of connective tissue disorders other than Sjögren's syndrome, or in neurological diseases other than motor neuron disease and multiple system atrophy. Our study shows that the spectrum of neurological disorders in which anti-neuronal antibodies can be detected is wider than previously thought. In addition, we found patterns of neuronal staining and Western blot reactivity that differed from those so far reported. This may permit identification of subgroups of patients in whom strategies directed at removing and/or suppressing antibody production could be of some benefit.

Herpes simplex encephalitis: an audit of the use of laboratory diagnostic tests.

BACKGROUND: The combination of both PCR and intrathecal antibody studies is recommended to confirm or refute the diagnosis of herpes simplex encephalitis (HSE). AIM: To investigate the pattern of use of laboratory tests in the diagnosis of suspected cases of HSE, and to determine the final diagnosis in cases proven not to be HSE. DESIGN: Structured audit. METHODS: We reviewed the case-notes of all patients who, over a five-year time period, presented with suspected encephalitis; and/or were prescribed aciclovir. Clinical and laboratory criteria were used to categorize the likelihood of HSE. RESULTS: We identified 222 patients: 10 (5%) had definite HSE, 24 (10%) possible HSE, and 144 (65%) a definite alternative diagnosis. In 44 (20%), no final diagnosis was made, but the diagnosis of HSE was excluded. PCR was performed in 68 (31%), intrathecal antibody studies in 24 (11%), and brain biopsy in 17 (8%). A wide range of diseases mimicked HSE, but most common were inflammatory diseases and other infections of the central nervous system. DISCUSSION: Laboratory tests, particularly intrathecal antibody assays, are under-used in the diagnosis of HSE. Although early empirical treatment of suspected HSE is essential, confirmation or exclusion of the diagnosis is equally important to avoid overlooking alternative diagnoses. Identification of the aetiology of encephalitis is of particular importance, given the current concerns of emerging infections and bioterrorism.