Lenka Fialová

Oxidative stress, metabolism of ethanol and alcohol-related diseases.

Alcohol-induced oxidative stress is linked to the metabolism of ethanol. Three metabolic pathways of ethanol have been described in the human body so far. They involve the following enzymes: alcohol dehydrogenase, microsomal ethanol oxidation system (MEOS) and catalase. Each of these pathways could produce free radicals which affect the antioxidant system. Ethanol per se, hyperlactacidemia and elevated NADH increase xanthine oxidase activity, which results in the production of superoxide. Lipid peroxidation and superoxide production correlate with the amount of cytochrome P450 2E1. MEOS aggravates the oxidative stress directly as well as indirectly by impairing the defense systems. Hydroxyethyl radicals are probably involved in the alkylation of hepatic proteins. Nitric oxide (NO) is one of the key factors contributing to the vessel wall homeostasis, an important mediator of the vascular tone and neuronal transduction, and has cytotoxic effects. Stable metabolites--nitrites and nitrates--were increased in alcoholics (34.3 +/- 2.6 vs. 22.7 +/- 1.2 micromol/l, p < 0.001). High NO concentration could be discussed for its excitotoxicity and may be linked to cytotoxicity in neurons, glia and myelin. Formation of NO has been linked to an increased preference for and tolerance to alcohol in recent studies. Increased NO biosynthesis also via inducible NO synthase (NOS, chronic stimulation) may contribute to platelet and endothelial dysfunctions. Comparison of chronically ethanol-fed rats and controls demonstrates that exposure to ethanol causes a decrease in NADPH diaphorase activity (neuronal NOS) in neurons and fibers of the cerebellar cortex and superior colliculus (stratum griseum superficiale and intermedium) in rats. These changes in the highly organized structure contribute to the motor disturbances, which are associated with alcohol abuse. Antiphospholipid antibodies (APA) in alcoholic patients seem to reflect membrane lesions, impairment of immunological reactivity, liver disease progression, and they correlate significantly with the disease severity. The low-density lipoprotein (LDL) oxidation is supposed to be one of the most important pathogenic mechanisms of atherogenesis, and antibodies against oxidized LDL (oxLDL) are some kind of epiphenomenon of this process. We studied IgG oxLDL and four APA (anticardiolipin, antiphosphatidylserine, antiphosphatidylethanolamine and antiphosphatidylcholine antibodies). The IgG oxLDL (406.4 +/- 52.5 vs. 499.9 +/- 52.5 mU/ml) was not affected in alcoholic patients, but oxLDL was higher (71.6 +/- 4.1 vs. 44.2 +/- 2.7 micromol/l, p < 0.001). The prevalence of studied APA in alcoholics with mildly affected liver function was higher than in controls, but not significantly. On the contrary, changes of autoantibodies to IgG oxLDL revealed a wide range of IgG oxLDL titers in a healthy population. These parameters do not appear to be very promising for the evaluation of the risk of atherosclerosis. Free radicals increase the oxidative modification of LDL. This is one of the most important mechanisms, which increases cardiovascular risk in chronic alcoholic patients. Important enzymatic antioxidant systems - superoxide dismutase and glutathione peroxidase - are decreased in alcoholics. We did not find any changes of serum retinol and tocopherol concentrations in alcoholics, and blood and plasma selenium and copper levels were unchanged as well. Only the zinc concentration was decreased in plasma. It could be related to the impairment of the immune system in alcoholics. Measurement of these parameters in blood compartments does not seem to indicate a possible organ, e.g. liver deficiency.

Elevated intrathecal antibodies against the medium neurofilament subunit in multiple sclerosis.

Neurofilaments are cytoskeletal proteins localized within axons, which may interact with the immune system during and following tissue destruction in multiple sclerosis (MS). Antibodies against the medium neurofilament subunit synthesized intrathecally may reflect axonal damage in MS patients. Both immunoglobulin G (IgG) and M (IgM) responses against the purified native medium subunit of neurofilaments (NFM) using enzyme-linked immunosorbent assay (ELISA) were determined in paired serum and cerebrospinal fluid samples obtained from 49 MS patients, 16 normal controls (CN), 21 control patients with miscellaneous diseases (CD) and 14 patients with neurodegenerative disorders (CDEG). Intrathecal production of IgM and IgG antibodies to NFM were elevated in MS patients compared with the CN or CD groups (p < 0.04 for IgM, p < 0.01 for IgG). The increase was present in all the MS courses (relapsing-remitting, primary and secondary progressive). Similar local anti-NFM IgG and IgM synthesis occurred in the MS and CDEG groups. MS patients with short and long disease duration did not differ in terms of their anti-NFM IgM and IgG responses. Repeated examinations showed stable intrathecal anti-NFM production. Intrathecal IgG and IgM antibodies against NFM were increased in MS patients and may serve as a potential marker for axonal pathology. The extent of anti-NFM levels did not correspond to any individualized clinical profiles of MS patients.

Antibodies against light neurofilaments in multiple sclerosis patients.

Objectives –  Axonal damage in multiple sclerosis (MS) may be reflected by antibodies against axon‐specific proteins – the light subunit of neurofilaments (NFL).

Relationship between ALS and the degree of cognitive impairment, markers of neurodegeneration and predictors for poor outcome. A prospective study

Introduction:  Amyotrophic lateral sclerosis (ALS) is a common neurodegenerative disease affecting motor neurons and may be associated with impaired cognition. Reliable prognostic factors for ALS patients are still missing.

Relationship between increased body iron stores, oxidative stress and insulin resistance in healthy men.

Aim: The aim of our cross-sectional study was to assess the relationships between body iron stores, oxidative stress, impaired insulin sensitivity and carotid atherosclerosis in a cohort of healthy men in primary prevention of cardiovascular disease. Methods: We examined 151 volunteers, aged 35– 60 years. Anthropometric parameters, markers of metabolic syndrome, insulin resistance, inflammatory markers, parameters of oxidative stress and intima-media thickness of common carotid artery were measured. Results: Ferritin correlated positively with waist circumference, body mass index, impaired insulin sensitivity, plasma triglycerides and inversely with high-density lipoprotein cholesterol. We observed positive correlations between ferritin, oxidized lowdensity lipoprotein and advanced oxidation protein products after adjustment for age, waist circumference, body mass index and measured inflammatory markers (high-sensitivity C-reactive protein, fibrinogen, interleukin-6 and tumor necrosis factor-α). There were no significant associations between ferritin and intima-media thickness or markers of endothelial dysfunction. In a stepwise multiple regression analysis, triglycerides, waist circumference and elevated transaminases were independent determinants of the serum ferritin level. Conclusion: Our results provide evidence for a relationship between plasma ferritin and oxidative modification of lipids as well as proteins in vivo. Higher body iron stores may contribute to impaired insulin sensitivity through increased oxidative stress in a cohort of healthy men.

Increased levels of pregnancy-associated plasma protein-A in patients with hypercholesterolemia: the effect of atorvastatin treatment

BACKGROUND Serum levels of pregnancy-associated plasma protein-A (PAPP-A) have recently been linked to plaque instability and are increased in acute coronary syndromes. The relation between PAPP-A levels and coronary risk factors, namely blood lipids, has not been studied to date. We have therefore investigated whether serum PAPP-A levels are increased in asymptomatic hypercholesterolemic subjects and whether PAPP-A levels are influenced by atorvastatin therapy. METHODS We examined 27 subjects with isolated hypercholesterolemia free of manifest vascular disease and 29 age-matched healthy control subjects. Patients were examined at baseline and after 10 weeks of atorvastatin treatment (20 mg/d). RESULTS In untreated hypercholesterolemic subjects, PAPP-A levels were significantly higher than in control subjects (8.02 +/- 1.86 mU/L vs 6.50 +/- 2.54 mU/L, P =.018). There was no correlation between PAPP-A levels and serum lipid levels. Atorvastatin treatment reduced total and LDL-cholesterol by 31% and 40%, respectively. Despite this profound lipid lowering, there was no significant change in the serum PAPP-A levels. CONCLUSIONS PAPP-A levels are elevated in hypercholesterolemic subjects without clinical signs of atherosclerosis. PAPP-A may therefore not only reflect plaque instability but also serve as a marker of total atherosclerotic burden in asymptomatic subjects with hyperlipidemia. However, PAPP-A levels are not influenced by atorvastatin treatment.

Cerebrospinal fluid and serum antibodies against neurofilaments in patients with amyotrophic lateral sclerosis

Background:  The aim of the study was to assess autoimmune involvement in amyotrophic lateral sclerosis (ALS).

Patients with Alzheimer disease have elevated intrathecal synthesis of antibodies against tau protein and heavy neurofilament

The role of humoral immunity related to neuron- and disease-specific cytoskeletal proteins patients with Alzheimer disease (AD) is unknown. We measured antibodies against tau protein, light and heavy (NFH) neurofilaments using ELISA in 80 paired serum and cerebrospinal fluid (CSF) samples from patients with AD, with other dementias (OD), with neuro-inflammatory diseases (IC) and 25 controls (NC). We estimated intrathecal synthesis according to the formula (CSF/serum anti-neurocytoskeletal IgG)/(CSF/serum total IgG). AD patients had significantly higher intrathecal anti-tau and anti-NFH antibodies than the other groups. These innovative findings may hint at specific alterations in humoral anti-neurocytoskeletal immunity and selectivity in AD, which could have diagnostic and immunotherapeutic implications.

Serum and cerebrospinal fluid light neurofilaments and antibodies against them in clinically isolated syndrome and multiple sclerosis

A release of light neurofilament subunits (NFL) into cerebrospinal fluid (CSF) and serum in multiple sclerosis (MS) may induce an immune response. We examined CSF and serum NFL levels and IgG antibodies against NFL in 19 patients with a clinically isolated syndrome (CIS) early converted into MS, 20 CIS-non-converters, 23 MS patients and 32 controls. CSF NFL levels were significantly higher in all patient groups. The highest CSF or intrathecally (IT) synthesized anti-NFL antibodies and CSF/serum ratios of anti-NFL antibodies were observed in CIS-converters. CSF NFL and CSF or IT anti-NFL antibodies could be surrogate biomarkers of axonal injury in early MS.

Increased Intrathecal High-Avidity Anti-Tau Antibodies in Patients with Multiple Sclerosis

Background Antibodies against tau protein indicate an interaction between the immune system and the neurocytoskeleton and therefore may reflect axonal injury in multiple sclerosis (MS). Methodology/Principal Findings The levels and avidities of anti-tau IgG antibodies were measured using ELISA in paired cerebrospinal fluid (CSF) and serum samples obtained from 49 MS patients and 47 controls. Anti-tau antibodies were significantly elevated intrathecally (p<0.0001) in the MS group. The CSF anti-tau antibody levels were lower in MS patients receiving therapy than those without treatment (p<0.05). The avidities of anti-tau antibodies were higher in the CSF than in the serum (MS group p<0.0001; controls p<0.005). Anti-tau avidities in the CSF were elevated in MS patients in comparison with controls (p<0.05), but not in serum. Conclusions MS patients have higher levels of intrathecal anti-tau antibodies. Anti-tau antibodies have different avidities in different compartments with the highest values in the CSF of MS patients.