Konrad Rejdak

A consensus protocol for the standardization of cerebrospinal fluid collection and biobanking

There is a long history of research into body fluid biomarkers in neurodegenerative and neuroinflammatory diseases. However, only a few biomarkers in CSF are being used in clinical practice. One of the most critical factors in CSF biomarker research is the inadequate powering of studies because of the lack of sufficient samples that can be obtained in single-center studies. Therefore, collaboration between investigators is needed to establish large biobanks of well-defined samples. Standardized protocols for biobanking are a prerequisite to ensure that the statistical power gained by increasing the numbers of CSF samples is not compromised by preanalytical factors. Here, a consensus report on recommendations for CSF collection and biobanking is presented, formed by the BioMS-eu network for CSF biomarker research in multiple sclerosis. We focus on CSF collection procedures, preanalytical factors, and high-quality clinical and paraclinical information. The biobanking protocols are applicable for CSF biobanks for research targeting any neurologic disease.

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 ≤ 6.0; EDSS ≤ 4.0; Multiple Sclerosis Severity Score [MSSS] ≤ 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.

Basic mechanisms of antiepileptic drugs and their pharmacokinetic/pharmacodynamic interactions: an update.

This article aims to summarize the current views of AED action and the promising new targets for the pharmacotherapy of epilepsy. In the first section of this paper, a neurobiological basis of epilepsy treatment and brief pharmacological characteristics of classical and new AEDs will be presented. In the second part, the results of experimental studies that have combined AEDs with similar or different mechanisms of action will be discussed.

Axonal degeneration and inflammation in acute optic neuritis

Aims: To investigate whether plasma biomarkers for axonal injury and inflammation are related to loss and recovery of visual function in acute optic neuritis (ON). Methods: Eighteen patients with ON and 14 controls were investigated in a longitudinal, prospective study. Plasma phosphorylated neurofilament heavy chain (NfHSMI35; a surrogate marker of axonal injury), nitric oxide metabolites (NOx), and citrulline (surrogate markers of inflammation) were measured. Results: Patients with ON had higher median plasma NfHSMI35 values than controls (0.17 versus 0.005 ng/ml; p < 0.05) and higher NOx values (49 versus 35.5μM; p < 0.001). Plasma NfHSMI35 values correlated inversely with visual acuity at presentation (R  =  −0.67; p  =  0.01). NfHSMI35 was higher in patients with poor recovery of visual acuity than in those with good recovery (0.25 ng/ml versus 0.09 ng/ml; p < 0.05). Three of four patients with high NfHSMI35 and high NOx values experienced a poor recovery as opposed to only one of five with high NOx but normal NfHSMI35 values. Conclusions: NfHSMI35, a surrogate marker for axonal damage, is a prognostic indicator and should be considered in the design of neuroprotective treatment strategies.

Research advances in basic mechanisms of seizures and antiepileptic drug action

Epilepsy is a common neurological disease but the mechanism of seizure generation has been only partially unraveled. Furthermore, almost 30% of epileptic patients are resistant to pharmacological treatment. Therefore, elucidation of the basic mechanism of seizures and search for new antiepileptics in order to treat the drug-resistant form of epilepsy and to improve the efficacy of current therapies seem justified. The aim of this overview is a brief presentation of some new concepts and research directions in pathogenesis and pharmacotherapy of epilepsy. Development of ideas on the mechanisms of seizures and antiepileptic drugs reflects the progress in our understanding of the central nervous system physiology, particularly of neurotransmission. Hyperactivity of excitatory amino acid systems, insufficient GABAA receptor-mediated neurotransmission, and disturbances in intrinsic properties of neuronal membranes are still regarded as the most important mechanisms of seizures. New data add to the complexity of GABA-glutamate interaction showing both excitatory and inhibitory role of GABA and glutamatergic neurons in the central nervous system. Moreover, besides synaptic NMDA and GABAA receptors, also extrasynaptic receptors for the amino acid transmitters have been recently implicated in the pathomechanism of epilepsy. Changes in expression, polymorphisms, lost- or gain-function mutations as well as cellular energetic imbalance can contribute to the disturbed function of the ligand- and voltage-dependent sodium, potassium, chloride and calcium channels, resulting in epileptiform activity. Voltage-dependent sodium and calcium channel blockers, and GABA mimetics are the most clinically useful groups of antiepileptic drugs and the newest research in this field is focused on more selective and subtle regulations of their molecular targets. Of interest is an emerging role of extrasynaptic GABAA receptors, various kinds of potassium ion channels, hyperpolarization-activated cyclic nucleotide gated (HCN) channels, acid-sensing ion channels, and gap junctions in the regulation of neuronal excitability and seizures. Iono- and metabotropic glutamate receptors used to be viewed as an attractive target for new anticonvulsants, however, opinions are now less enthusiastic, since their competitive and non-competitive antagonists possess undesired side effects. Positive or negative allosteric modulators of glutamate receptors with fewer side-effects can be more promising. The introduction of new compounds acting through novel pharmacological mechanisms gives hope that the proportion of patients with uncontrolled epilepsy will substantially decrease. However, this may be possible if molecular background of the pharmacoresistance in epilepsy is deciphered.

Decreased level of kynurenic acid in cerebrospinal fluid of relapsing-onset multiple sclerosis patients

The present study was undertaken to measure cerebrospinal fluid (CSF) levels of kynurenic acid (KYNA) in patients with relapsing-onset multiple sclerosis (MS) during remission or not progressing for at least 2 months. In these patients the levels of CSF KYNA were found to be significantly lower compared with subjects with non-inflammatory neurological diseases, as well as those with inflammatory disease (median (interquartile range): 0.41 (0.3-0.5) pmol/ml, n=26 vs. 0.67 (0.5-1.1), n=23, P<0.01 and 1.7 (1.5-2.6), n=16, P<0.001, respectively). These results provide further evidence of the alterations in the kynurenine pathway during remitting-onset MS.

Multiple sclerosis: a practical overview for clinicians

Multiple sclerosis (MS) is the commonest disabling neurological condition to afflict young adults and therefore has a high social burden. Over several decades, there has been a considerable progress in the understanding of the disease pathogenesis as well as in the clinical management of MS patients. The emphasis in managing MS patients has shifted to multidisciplinary teams working in specialist groups. A review of the literature was conducted using MedLine to identify recent advances in MS. The current consensus is that MS is an autoimmune disease triggered by environmental agents acting in genetically susceptible people. Based on that concept, new methods of immune intervention procedures have been introduced into clinical practice. Licensed first-line disease-modifying therapies reduce the MS attack or relapse rate by a third and delaying short-term disease progression. More effective therapies have emerged; however, these are associated with increased risks. New clinical and pathological insights are making us question the aetiology and pathogenesis of MS. The recognition of pathological heterogeneity has raised the question of whether MS is a single disease entity or a syndrome. Recent evidence suggests that the pathological subtype may predict therapeutic response to specific therapies. A new novel auto-antibody has defined a subset of neuromyelitis optica or Devics disease as being distinct from MS. This is an attractive concept that is not widely accepted. The observation that MS progresses despite immunosuppressive therapy suggests that MS may be a neurodegenerative disease with overlapping immune activation possibly in response to the release of central nervous system auto-antigens. The development of neuroprotective therapies for MS is required to prevent the devastating effects of long-term disability as a result of progressive disease.

Chitinase 3-like 1: prognostic biomarker in clinically isolated syndromes.

Chitinase 3-like 1 (CHI3L1) has been proposed as a biomarker associated with the conversion to clinically definite multiple sclerosis in patients with clinically isolated syndromes, based on the finding of increased cerebrospinal fluid CHI3L1 levels in clinically isolated syndrome patients who later converted to multiple sclerosis compared to those who remained as clinically isolated syndrome. Here, we aimed to validate CHI3L1 as a prognostic biomarker in a large cohort of patients with clinically isolated syndrome. This is a longitudinal cohort study of clinically isolated syndrome patients with clinical, magnetic resonance imaging, and cerebrospinal fluid data prospectively acquired. A total of 813 cerebrospinal fluid samples from patients with clinically isolated syndrome were recruited from 15 European multiple sclerosis centres. Cerebrospinal fluid CHI3L1 levels were measured by enzyme-linked immunosorbent assay. Multivariable Cox regression models were used to investigate the association between cerebrospinal fluid CHI3L1 levels and time to conversion to multiple sclerosis and time to reach Expanded Disability Status Scale 3.0. CHI3L1 levels were higher in patients who converted to clinically definite multiple sclerosis compared to patients who continued as clinically isolated syndrome (P = 8.1 × 10(-11)). In the Cox regression analysis, CHI3L1 levels were a risk factor for conversion to multiple sclerosis (hazard ratio = 1.7; P = 1.1 × 10(-5) using Poser criteria; hazard ratio = 1.6; P = 3.7 × 10(-6) for McDonald criteria) independent of other covariates such as brain magnetic resonance imaging abnormalities and presence of cerebrospinal fluid oligoclonal bands, and were the only significant independent risk factor associated with the development of disability (hazard ratio = 3.8; P = 2.5 × 10(-8)). High CHI3L1 levels were associated with shorter time to multiple sclerosis (P = 3.2 × 10(-9) using Poser criteria; P = 5.6 × 10(-11) for McDonald criteria) and more rapid development of disability (P = 1.8 × 10(-10)). These findings validate cerebrospinal fluid CHI3L1 as a biomarker associated with the conversion to multiple sclerosis and development of disability and reinforce the prognostic role of CHI3L1 in patients with clinically isolated syndrome. We propose that determining cerebrospinal fluid chitinase 3-like 1 levels at the time of a clinically isolated syndrome event will help identify those patients with worse disease prognosis.

Cerebrospinal fluid brain specific proteins in relation to nitric oxide metabolites during relapse of multiple sclerosis

This study investigated the cerebrospinal fluid (CSF) levels of ferritin, S100B as biomarkers for glial activation and NfHSM135 — a biomarker of axonal damage — in relation to nitric oxide (NO) metabolites: nitrate and nitrite (NOx) during acute multiple sclerosis (MS) relapse. Thirty-four relapsing—remitting MS (RR-MS) patients during acute relapse and 12 controls were enrolled. Patients were assessed on Expanded Disability Status Scale (EDSS) and underwent lumbar puncture within two weeks following relapse. Twenty patients were available for further follow-up and were assessed on EDSS 6—8 weeks since the relapse onset. The CSF NOx (P < 0.0001), NfHSM135 ( P = 0.01) and S100B (P = 0.009) but not ferritin (P > 0.05) were significantly raised in MS group. There was a significant correlation between CSF ferritin and S100B in RR-MS group (P = 0.004). CSF NOx did not correlate with S100B and ferritin in study groups. RR-MS patients with detectable NfHSM135 levels had higher NOx compared with subjects having undetectable NfHSM135 (P = 0.03). In the follow-up study, raised baseline levels of NOx (P = 0.016) or NfHSM135 (P = 0.04) inversely correlated with the clinical recovery grade expressed as relative EDSS change between baseline and follow-up. In conclusion, NO metabolites were increased and because of their correlation with a biomarker of axonal degeneration (neurofilaments) and a measure for clinical disability (EDSS), relapse-related nitrosative stress is likely to be relevant to the development of sustained disability in an individual patient. Multiple Sclerosis 2008; 14: 59—66. http://msj.sagepub.com

Consensus Guidelines for CSF and Blood Biobanking for CNS Biomarker Studies.

There is a long history of research into body fluid biomarkers in neurodegenerative and neuroinflammatory diseases. However, only a few biomarkers in cerebrospinal fluid (CSF) are being used in clinical practice. Anti-aquaporin-4 antibodies in serum are currently useful for the diagnosis of neuromyelitis optica (NMO), but we could expect novel CSF biomarkers that help define prognosis and response to treatment for this disease. One of the most critical factors in biomarker research is the inadequate powering of studies performed by single centers. Collaboration between investigators is needed to establish large biobanks of well-defined samples. A key issue in collaboration is to establish standardized protocols for biobanking to ensure that the statistical power gained by increasing the numbers of CSF samples is not compromised by pre-analytical factors. Here, consensus guidelines for CSF collection and biobanking are presented, based on the guidelines that have been published by the BioMS-eu network for CSF biomarker research. We focussed on CSF collection procedures, pre-analytical factors and high quality clinical and paraclinical information. Importantly, the biobanking protocols are applicable for CSF biobanks for research targeting any neurological disease.