Effect of melatonin supplementation on plasma lipid hydroperoxides, homocysteine concentration and chronic fatigue syndrome in multiple sclerosis patients treated with interferons-beta and mitoxantrone.

UNLABELLED: Multiple sclerosis (MS) prevalence is higher in geographic regions with less sunlight exposure. Melatonin participates in the effects of sunlight in healthy individuals and could play a role in MS pathophysiology. Melatonin crosses the blood-brain barrier and exerts antioxidative, immunomodulatory, and anti-inflammatory effects. Chronic fatigue syndrome concerns 80 - 90% MS patients. The pathophysiology of chronic fatigue syndrome is unknown, however activation of immune, inflammatory, oxidative and nitrosative stress mechanisms and plasma lipid peroxide elevation was reported. Homocysteine increases plasma lipid hydroperoxides levels. The aim was to determine the effect of melatonin supplementation on chronic fatigue syndrome in MS patients and evaluate plasma lipid hydroxyperoxides (LHP) and homocysteine concentrations as a potential biochemical fatigue biomarkers. Into a case-control prospective study 102 MS patients divided according receiving immunomodifying MS treatment into groups: RRMS-pretreated, RRMS-INF-beta, SP/PPMS-mitoxantrone, RRMS-relapse were enrolled. Patients were supplemented with melatonin over 90 days. Plasma LHP, homocysteine concentration, brain MRI and fatigue score were examined. Results show that LHP concentrations were significantly higher in all studied MS groups vs. CONTROLS: In all MS patient groups melatonin application resulted in significant decrease in plasma LHP concentrations. Plasma homocysteine concentration was similar in healthy people, RRMS-pretreated, RRMS-INF-beta and SP/PP-MS-mitoxantrone groups. However, in the RRMS-relapse group plasma levels of homocysteine were significantly higher compared to the RRMS-pretreated group. There were no significant differences in plasma homocysteine concentration in the studied groups before and after melatonin application. The fatigue score was significantly lower in RRMS pretreated group compared to RRMS-INF-beta and SP/PP MS-mitoxantrone treated patients. Plasma lipid hydroxyperoxides could be potential biochemical chronic fatigue syndrome biomarker in MS patients and homocysteine could be a potential marker of acute phase of MS. Melatonin exerts beneficial effects in MS patients based on its' proved antioxidative properties.

Calcium-phosphate metabolism in patients with multiple sclerosis.

INTRODUCTION AND OBJECTIVES: The purpose of this study was to evaluate the concentration of 25-hydroxycholecalciferol and parameters of calcium-phosphate metabolism at different periods of relapsing-remitting multiple sclerosis (RRMS). MATERIALS AND METHODS: Forty-five patients, residents of Poland (49°-50°, N), were enrolled in the study, i.e. 15 immediately after the diagnosis of RRMS, 15 at the early stage and 15 at the advanced stage of RRMS. The results were compared to values obtained in 20 age- and sex-matched controls. RESULTS: Lower serum concentrations of 25-hydroxycholecalciferol and ionised calcium were found in patients compared to the control group. In patients with the disease duration of 5-6 years, concentrations of 25-hydroxycholecalciferol and ionised calcium were lower than in patients in the earlier period of RRMS. The inverse and clearer direction of changes was found in parathormone serum concentration in patients compared to the controls. In patients with a longer disease duration, a significantly lower 25-hydroxycholecalciferol concentration was found in female patients compared to male patients. In patients, more frequent 25-hydroxycholecalciferol and unsaturated fatty acids' supplementation was observed compared to the controls. CONCLUSIONS: In RRMS patients, calcium-phosphate metabolism is disturbed which increases during disease progression.

Proopiomelanocortin but not vasopressin or renin-angiotensin system induces resuscitative effects of central 5-HT1A activation in haemorrhagic shock in rats.

The aim of this study was to determine the effectory mechanisms: vasopressin, renin-angiotensin system and proopiomelanocortin-derived peptides (POMC), partaking in the effects of serotonin through central serotonin 1A receptor (5-HT1A) receptors in haemorrhagic shock in rats. The study was conducted on male Wistar rats. All experimental procedures were carried out under full anaesthesia. The principal experiment included a 2 hour observation period in haemorrhagic shock. Drugs used - a selective 5-HT1A agonist 8-OH-DPAT (5 µg/5 µl); V1a receptor antagonist [ß-mercapto-ß, ß-cyclo-pentamethylenepropionyl(1),O-me-Tyr(2),Arg(8)]AVP (10 µg/kg); angiotensin type I receptor antagonist (AT1) ZD7155 (0.5 mg/kg, i.v.); angiotensin-converting-enzyme inhibitor captopril (30 mg/kg, i.v.); melanocortin type 4 (MC4) receptor antagonist HS014 (5 µg, i.c.v.). There was no influence of ZD715, captopril or blocking of the V1a receptors on changes in the heart rate (HR), mean arterial pressure (MAP), peripheral blood flow or resistance caused by the central stimulation of 5-HT1A receptors (P≥0.05). However, selective blocking of central MC4 receptors caused a slight, but significant decrease in HR and MAP (P<0.05). POMC derivatives acting via the central MC4 receptor participate in the resuscitative effects of 8-OH-DPAT. The angiotensin and vasopressin systems do not participate in these actions.

Influence of melatonin supplementation on serum antioxidative properties and impact of the quality of life in multiple sclerosis patients.

The relationship between the prevalence of multiple sclerosis (MS) and sunlight's ultraviolet radiation was proved. Oxidative stress plays a role in the pathogenic traits of MS. Melatonin possesses antioxidative properties and regulates circadian rhythms. Several studies have reported that the quality of life is worse in patients with MS than in healthy controls, with a higher prevalence of sleep disturbances, depression and fatigue. The aim of study was to evaluate 5 mg daily melatonin supplementation over 90 days on serum malondialdehyde (MDA) concentration and superoxide dismutase (SOD) activity and its' influence on impact of the quality of life of MS patients. A case-control prospective study was performed on 102 MS patients and 20 controls matched for age and sex. The EDSS, MRI examinations and Multiple Sclerosis Impact Scale (MSIS-29) questionnaire was completed. Marked increase in serum MDA concentration in all MS patients groups was observed and after melatonin treatment decreased significantly in interferons-beta and glatiramer acetate-treated groups, but not in mitoxantrone-treated group. A significant increase in SOD activity compared to controls only in glatiramer acetate-treated group was observed. After 3 months melatonin supplementation the SOD activity increased compared to initial values in interferons beta-treated groups. A significant increase in both MSIS-29-PHYS and MSIS-29-PSYCH items mean scores only in the MX group as compared to other groups was observed. There were no significant differences in mean MSIS-29-PHYS was observed before and after melatonin therapy. Melatonin supplementation caused a decrease in mean MSIS-29-PSYCH scores compared to initial values in interferons beta-treated groups. Finding from our study suggest that melatonin can act as an antioxidant and improves reduced quality in MS patients.

Labeled [³H]--thymidine incorporation in the DNA of 3T3-L1 preadipocytes due to MT2- and not MT3- melatonin receptor.

The discovery of MT1, MT2, and MT3 melatonin receptors on adipose tissue cells gives grounds for considering the possibility of melatonin as a factor which influences energy storage through modulation of metabolism and adipocyte proliferation. To date only a few contradictory studies have been published on the influence of melatonin on preadipocytes. The aim of the present study is to evaluate the influence of melatonin at physiological and supraphysiological concentrations on the proliferation of 3T3-L1 murine preadipocytes after 3 and 24 hours of the experiment and to determine the participation of membrane melatonin MT2 receptors, and for the first time--MT3, in its melatonin action during a 24-hour experiment. The 3T3-L1 murine preadipocyte cell line were cultured with or without melatonin at 10⁻³ and 10⁻9 mol/L, with or without melatonin antagonists luzindole (10⁻4 mol/L) and prazosin (10⁻5 mol/L). Cell proliferation was determined by means of labeled [³H]-thymidine incorporation in the DNA of the cell. Melatonin at both physiological and supraphysiological concentrations has a stimulating effect on the number of 3T3-L1 preadipocytes. The application of luzindole inhibits the above effect of melatonin both at 10⁻³ mol/L and 10⁻9 mol/L concentrations (P<0.05). The presence of prazosin does not have a statistically significant influence on the effects of melatonin action. Summarizing, it has been proven that melatonin exerts a proproliferative effect on 3T3-L1 preadipocytes at physiological and supraphysiological concentrations, partially by MT2, and not by MT3 receptors.

The role of serotonergic 5-HT1A receptors in central cardiovascular regulation in haemorrhagic shock in rats.

The bleeding and haemorrhage is strictly related with accidents and many medical procedures. In some conditions it leads to hypovolaemia and further to hypovolaemic shock. Under conditions of haemorrhagic shock, heart rate and blood pressure critically collapse. Reversing the sympathoinhibitory phase of hypovolaemia could be crucial for clinical management of injured patients after haemorrhage. Systemic administration of 5-HT1A agonists seams to produce resuscitating effects. The aim of this study was to investigate the participation of central serotonin and, in particular, 5-HT1A receptors in cardiovascular regulation in haemorrhagic shock in rats. Intracerebroventricular (i.c.v.) administration of serotonin (5-HT) increased the heart rate (HR), mean arterial pressure (MAP) and implicated that all haemorrhaged animals survived for the whole observation time (2 hours). Similar, although significantly more minor, effects were achieved after selective 5-HT1A activation. Moreover, the i.c.v. administration of selective 5-HT1A antagonist before i.c.v. 5-HT injection partially inhibited 5-HT induced changes. The results of the present work indicate that 5-HT plays an important role in the reversal of the haemorrhagic shock in rats. These effects are at least partially dependent on activation of 5-HT1A receptors.

Antioxidative enzymes activity and malondialdehyde concentration during mitoxantrone therapy in multiple sclerosis patients.

Mitoxantrone (MX) is approved for the treatment of aggressive relapsing-remitting, secondary-progressive and progressive-relapsing form of multiple sclerosis (MS). The mechanism of its action is multiaxial, however, it is not free from side effects. The causes of the side effects are still unknown and require further investigation. The aim of this study was to investigate the influence of MX therapy on enzymatic parameters of endogenous antioxidative status: manganese and copper/zinc superoxide dismutase (MnSOD, Cu/ZnSOD), catalase (CAT), glutathione peroxidase (GSH-Px) and lipid peroxidation marker--malondialdehyde (MDA) in blood serum and cerebrospinal fluid (CSF) in patients suffering from MS. After the MX therapy serum and the CSF MDA concentrations increased significantly. We reported that MnSOD activities decrease in serum and the CSF, while, surprisingly, the serum Cu/ZnSOD activity increases after the MX therapy. We also noted a marked decrease in CSF CAT and GSH-Px activity after the MX treatment. Our results strongly suggest the influence of MX therapy on oxidation/antioxidation status of serum and the CSF. These findings open up new opportunities for a better understanding of underlying physiopathological events in MS and provide a new insight into MX's mechanisms of action, especially its potent side effects.

Basal and postprandial plasma levels of PYY, ghrelin, cholecystokinin, gastrin and insulin in women with moderate and morbid obesity and metabolic syndrome.

Metabolic syndrome (MS), defined as central obesity, hyperinsulinemia, insulin resistance, hypertension, dyslipidemia and glucose intolerance, has been associated with inflammatory biomarkers and cardiovascular diseases. This study was carried out on three groups of women; lean controls, moderately obese with MS (OB-MS) and morbidly obese with MS (MOB-MS). The main objectives were: 1. to analyze the plasma levels of total and acylated ghrelin, peptide YY(3-36) (PYY(3-36)), cholecystokinin (CCK), gastrin and insulin levels under basal conditions and in response to a standard mixed meal, and 2. to elucidate the relationship between the plasma levels of these gut peptides and metabolic syndrome parameters. Plasma levels of the gut hormones were measured by radioimmunoassays at time 0 just before the meal and at 30, 60 and 120 min after a meal ingestion. Traditional lipid profile and high-sensitivity C reactive protein (hs-CRP), the strongest biomarker of inflammation were also determined in OB-MS and MOB-MS. When compared to OB-MS, MOB-MS exhibited much higher anthropometric parameters such as waist circumference, higher fat mass and higher plasma levels of low density lipoprotein-cholesterol (LDL-C) and hs-CRP. Both these obese groups revealed significantly higher values of body mass index (BMI), fat mass, total cholesterol (TC), LDL-C, fasting glucose, fasting insulin, insulin resistance (IR) calculated from homeostatic model assessment (HOMA) and hs-CRP compared to the values recorded in lean subjects. Fasting PYY(3-36) level was lower, while fasting acylated ghrelin was higher in MOB-MS than in OB-MS. Plasma total and acylated ghrelin levels were significantly lower in OB-MS compared to lean women. In MOB-MS women the fasting PYY(3-36) levels were lower compared to lean controls and OB-MS, whilst postprandially in both OB-MS and MOB-MS, it was much lower than in lean women. The fasting plasma levels of total and acylated ghrelin and their postprandial decrease were significantly smaller in both obese groups compared to lean subjects. Plasma hs-CRP levels correlated positively with BMI, waist circumference, fat mass, fasting glucose, HOMA IR and fasting active ghrelin, whilst it negatively correlated with plasma fasting and total ghrelin. Moreover, plasma fasting acylated ghrelin correlated positively with fat mass. Fasting total ghrelin correlated positively with BMI, HDL-C and negatively with HOMA IR. We conclude that MS features of obesity are closely related to fasting and postprandial alterations of concentrations of PYY(3-36), CCK and ghrelin, suggesting that determination of gut hormones controlling food intake might be considered as a valuable tool to assess the progression of MS to comorbidities of obesity.

Role of leptin, ghrelin, angiotensin II and orexins in 3T3 L1 preadipocyte cells proliferation and oxidative metabolism.

There is now growing evidence that the reactive oxygen species have an influence on proliferation and antioxidative status of various cell types. The aim of the study was to investigate the effects of different concentrations of leptin, ghrelin, angiotensin II and orexins on proliferation, culture medium malondialdehyde (MDA) levels and antioxidative enzymes activities: superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and catalase (CAT) in 3T3 L1 preadipocytes cell culture. Cell proliferation was measured using [(3)H]tymidine incorporation. In 3T3-L1 cells leptin caused a significant reduction in proliferation (by 36%) compared to control. Ghrelin increased preadipocyte proliferation, and the effect was stronger in higher dose (by 39%), while proproliferatory effect of angiotensin II was stronger in lower doses (by 47%). All used doses of orexin A significantly increased 3T3 L1 cell proliferation (from 21% to 160%), while orexin B caused a marked reduction (from 35% to 70%) of this proliferation. The effects of both orexins were dose-dependent. Leptin and ghrelin increased activity of SOD, CAT, GSH-Px and decreased level of MDA. Angiotensin II treatment stimulated only SOD and CAT activities. Influence of orexins was different on various enzymes. Orexin A increased MDA levels, while orexin B caused a marked decrease in MDA levels. Our results strongly suggest the effects of appetite affecting hormones such as leptin and ghrelin on proliferation and antioxidative enzyme activities of preadipocyte cell lines. Orexin A was found to be the most efficient proliferative-signalling hormone, while orexin B revealed the most significant inhibitory effect on preadipocytes proliferation.

Influence of melatonin on cell proliferation, antioxidative enzyme activities and lipid peroxidation in 3T3-L1 preadipocytes--an in vitro study.

Melatonin, acting via MT1, MT2 and MT3 membrane receptors, influences central and peripheral regulatory mechanisms of energy homeostasis in mammals. In peripheral tissues, it evokes the pro-proliferative effect in a number of normal cells. Moreover, this hormone inhibits lipolysis in subcutaneous adipocytes in vitro and reduces free oxygen metabolites-induced damage acting directly, as a free radical scavenger, and indirectly, by stimulation of antioxidative enzyme activities. The aim of the study was to examine the effects of melatonin on cell proliferation, antioxidative enzyme activities and malondialdehyde (MDA) concentration in 3T3-L1 preadipocyte cell culture. We found that melatonin (10(-3) and 10(-6) M/L) stimulated cell proliferation in dose- and time-depending manner, and this effect was inhibited by a relatively selective MT2 receptor antagonist - luzindole (10(-4) M/L). Melatonin, increased activities of manganese containing and copper-zinc containing superoxide dismutase (MnSOD and Cu/ZnSOD) isoenzymes, catalase, glutathione reductase and glutathione peroxidase after 24 h of incubation. In contrast, after 48 h of incubation, activities of all studied enzymes were lower than in the control group. There were no changes in MDA concentrations after 24 h of incubation, whereas, in melatonin-treated media, after 48 h of the experiment, MDA level was significantly decreased. Our results demonstrate that melatonin, acting via MT2 receptors, stimulates proliferation of 3T3-L1 preadipocytes and this action could be due to the enhancement in antioxidative enzyme activities and attenuation of lipid peroxidation by this indole.

Effect of extremely low frequency of electromagnetic fields on cell proliferation, antioxidative enzyme activities and lipid peroxidation in 3T3-L1 preadipocytes--an in vitro study.

The exposure to extremely low frequency electromagnetic field (ELF-MF, frequencies less than 200-300 Hz) can alter the transcription and translation of genes, influence the cell proliferation rate and affect enzyme activities. Moreover, the hypothesis that ELF-MF increases free oxygen metabolites generation has been proposed. Since recent in vivo studies suggest that electric and magnetic fields are able to affect adipose cells metabolism. The aim of the study was to examine the effects of ELF-MF (frequency of basic impulse 180-195 Hz, induction 120 microT) on cell proliferation, antioxidative enzyme activities and malondialdehyde (MDA) concentration in 3T3-L1 preadipocyte cell culture. We found that ELF-MF application lasting 36 minutes daily failed to influence cell count after 24h and 48 h of incubation. After 24 h, in the ELF-MF treated group, manganese- and copper-zinc-containing superoxide dismutase (MnSOD and Cu/ZnSOD) isoenzymes media activities were decreased, catalase activity was increased, whereas there were no significant differences in glutathione peroxidase (GSH-Px) and glutathione reductase (GSSG-Rd) activities in comparison to the control. After 48 h of incubation, all enzyme activities were reduced, except for GSSG-Rd, in which no changes were noticed. MDA concentration at 24 h after incubation with the exposure to ELF-MF was significantly higher in comparison to the control, without ELF-MF. After 48 h of incubation, MDA levels were significantly lower in both groups with no differences between the groups without and with ELF-MF. We conclude that ELF-MF influences antioxidative enzyme activities and increases lipid peroxidation in 3T3-L1 preadipocyte cultures.

Influence of extremely-low-frequency magnetic field on antioxidative melatonin properties in AT478 murine squamous cell carcinoma culture.

Effects of melatonin, extremely-low-frequency magnetic field (ELF-MF), and their combination on AT478 murine squamous cell carcinoma line were studied. Manganese superoxide dismutase (MnSOD), copper-zinc superoxide dismutase (Cu/ZnSOD), and glutathione peroxidase (GSH-Px) were used as markers of cells antioxidative status, and malondialdehyde (MDA) level was used as a marker of lipid peroxidation. After melatonin treatment, antioxidative enzyme activities were increased and MDA level was decreased. Application of ELF-MF on treated cells caused an increase of both superoxide dismutases activity and MDA level, but influence of ELF-MF on GSH-Px activity was negligible. All enzyme activity in culture medium containing melatonin (10(-3), 10(-4), 10(-5) M) after exposure to ELF-MF were significantly diminished compared to cells treated only with melatonin. Also MDA levels after combined treatment with melatonin and ELF-MF were significantly decreased. Observed changes were statistically significant (p<0.05). These results strongly suggest that ELF-MF attenuates antioxidative actions of melatonin on cellular level.