Investigation of oxidative stress in relapse and remission periods of patients with relapsing-Remitting multiple sclerosis

Samet Oncel; Musa Ozturk; Rabia Gokcen Gozubatik-Celik; Aysun Soysal; Sevim Baybaş

doi:10.4103/nsn.nsn_142_20

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ORIGINAL ARTICLE

Year : 2021 | Volume : 38 | Issue : 1 | Page : 67-72

Investigation of oxidative stress in relapse and remission periods of patients with relapsing-Remitting multiple sclerosis

Samet Oncel¹, Musa Ozturk², Rabia Gokcen Gozubatik-Celik², Aysun Soysal², Sevim Baybaş²
¹ Department of Neurology, Batman Dunya Hospital, Batman, Turkey
² Department of Neurology, Bakirkoy Training and Research Hospital for Psychiatry and Neurological Disorders, University of Health Sciences, Istanbul, Turkey

Date of Submission	16-Aug-2020
Date of Acceptance	13-Nov-2020
Date of Web Publication	26-Mar-2021

Correspondence Address:
Samet Oncel
Department of Neurology, Batman Dunya Hospital, Batman
Turkey

Source of Support: None, Conflict of Interest: None

DOI: 10.4103/nsn.nsn_142_20

Abstract

Introduction: Multiple sclerosis (MS) is a chronic inflammatory demyelinating disease of the central nervous system. The role of oxidative stress has gained importance in the pathogenesis of free radicals in cells such as protein, lipid, and nucleic acids by showing myelin loss and axonal degeneration. In this study, serum total oxidant level total oxidant status (TOS), total antioxidant status (TAS), oxidative stress index (OSI), and paraoxonase (PON) in patients with relapsing-remitting-MS (RRMS) were examined. The hypothesis was that antioxidants might indicate the attack phase or the progression phase. Methods: Twenty-four patients with RRMS known to have undergone new attacks and 24 healthy controls whose demographic data were appropriate were included. PON and TAS/TOS measurements were performed once in the healthy group and twice in the serum samples of patients during relapse and remission periods. The Number Cruncher Statistical System 2007 and Power Analysis and Sample Size 2008 Statistical Software (Utah, USA) program was used for statistical analysis. Results: TAS, TOS, and PON levels were similar in the patient and control groups during the attack. In remission, only a positive, statistically significant relationship between the duration of the disease and TAS measurements was noted (r = 0.435; P = 0.034). No significant relationship was found between sociodemographic characteristics, illness duration, laboratory/imaging findings, and antioxidant parameters in blood. Conclusion: In this study, TOS, TAS, OSI, and PON serum levels had no role in determining attack and remission periods in RRMS. There were only significant differences between disease duration and TAS results during the remission period. Following TAS results may help in monitoring progression in patients with early MS.

Keywords: Pathogenesis, progression, relapsing-remitting-multiple sclerosis, total antioxidant level

How to cite this article:
Oncel S, Ozturk M, Gozubatik-Celik RG, Soysal A, Baybaş S. Investigation of oxidative stress in relapse and remission periods of patients with relapsing-Remitting multiple sclerosis. Neurol Sci Neurophysiol 2021;38:67-72

How to cite this URL:
Oncel S, Ozturk M, Gozubatik-Celik RG, Soysal A, Baybaş S. Investigation of oxidative stress in relapse and remission periods of patients with relapsing-Remitting multiple sclerosis. Neurol Sci Neurophysiol [serial online] 2021 [cited 2023 Mar 23];38:67-72. Available from: http://www.nsnjournal.org/text.asp?2021/38/1/67/309678

Introduction

Multiple sclerosis (MS) is a chronic inflammatory demyelinating disease of the central nervous system (CNS). Although its etiology is still unclear, immunologic mechanisms are prominent in the onset and progression of the disease. Environmental factors, especially oxidative stress, are thought to contribute to this process.^[1]

CNS cells are more susceptible to oxidative stress because of their high oxygen content. Research has revealed multiple immune pathways and their pro-inflammatory products in the pathophysiology of MS. Molecules such as nitric oxide, peroxynitrite, and hydrogen peroxide disrupt cell signal transduction, leading to the death of neurons and oligodendrocytes, and contributing to demyelination. They also cause axonal destruction through phosphorylation, which is essential for protecting axons from proteolysis in the chronic period. In the pathophysiology, reactive oxygen radicals play an important role in damaging lipids and proteins by peroxidant damage to myelin and indirect degradation of myelin basic protein.^{[2],[3],[4],[5]} The factor that is blamed is the toxicity of glutamate. Excess activation of glutamate contributes to blood–brain dysfunction, degradation of oligodendrocytes, and mitochondrial dysfunction by increasing calcium flow through neurons and oligodendrocytes, and by creating free radicals including superoxide and mitochondrial nitrogen.^[6]

Total oxidant status (TOS) is an indicator of total oxidative stress in the body. Reactive oxygen and nitrogen compounds, homocysteines, myeloperoxidase, and lipoxygenases are mainly oxidized substances in plasma.^[7] Most oxidant substances in plasma are reactive oxygen and nitrogen derivatives, homocysteine, myeloperoxidase, and lipoxygenases.^[7] As a result of the increase in total oxidant levels and decrease in total antioxidant levels in patients with MS, it increases blood–brain barrier (BBB) dysfunction, causes mitochondrial dysfunction, and decreases adenosine triphosphate production. It also causes DNA damage by disrupting cell signal transmission. These events result in the death of neurons and oligodendrocytes. Therefore, it is thought to play an important role in disease progression.^[8]

Another indicator of antioxidant capacity is the level of paraoxonase (PON) in the blood. PON is linked to high-density lipoproteins. It protects the nervous system from the neurotoxicity of circulating organophosphates. It inhibits lipid peroxidation of low-density lipoproteins (LDL), neutralizes free radicals such as hydrogen peroxide, and hydrolyzes platelet-activating factor. In the literature, there are different results regarding PON activity during the attack periods of patients with MS.^{[5],[9],[10],[11]}

The oxidative stress index (OSI) was used as another parameter in the evaluation of oxidative stress in previous studies. This is a proportional index obtained when total oxidants are separated into total antioxidants. When the OSI is elevated, it indicates an increase in oxidative stress.^[12],[13]

Different results have been reported in the literature on this subject, and the results in patients with progressive MS suggest that these parameters may play a role in disease progression. In our research, we investigated the role of oxidative stress in the pathophysiology of relapsing-remitting-MS (RRMS), the possible differences in recession cycles, and whether antioxidants could be used as criteria to assess attacks and their role in treatment strategies.

Methods

Clinical features

The study included 24 patients who were diagnosed as having RRMS in the neurology clinic of our hospital between January 2012 and December 2012, in accordance with the McDonald criteria (2010 revision),^[14] and 24 age- and sex-matched healthy volunteers. Patients were evaluated in both attack and remission periods. Patients who were clinically stable at least 3 months after the attack and had no new (enhancing or nonenhancing) lesions on imaging were admitted in remission.

The sociodemographic features, neurologic examinations, expanded disability status scale (EDSS) scores for attack and remission, the blood samples taken on the day of attack, the total number of attacks, the disease duration, the disease's initial symptoms, and the location of the magnetic resonance imaging lesions, were recorded. The exclusion criteria were as follows: use of antilipidemic medications, body mass index ≥30 or smokers, people with diabetes mellitus, oncologic, any other neurologic disorder, or any other known systemic or auto-immune disease. The research was voluntary, and the participating patients gave a written informed consent form.

Laboratory

Plasma, or serum, contains several oxidants and antioxidants. Although measuring these substances can be performed individually, total antioxidant status (TAS) and TOS measurements, which show their additive effects, were preferred instead of measuring each substance for oxidizing and antioxidant effects, as long, costly, and complex tests are needed. Serum TAS and TOS levels, OSI, and PON activity were measured in venous blood once in the healthy control group and twice in the patient group (in relapse and remission periods).

Measurement of paraoxonase activity

Serum PON basal activity was measured in the absence of sodium chloride. The rate of poroxone hydrolysis (diethyl-p-nitrophenylphosphate) was calculated from the molar absorption coefficient (18.290 mol/L/cm) of p-nitrophenol at pH 8.5, which had absorbance at 37°C and 412 nanometers.^[15]

Total antioxidant level (total antioxidant status) measurement

TAS was evaluated using an internationally accepted method developed by Erel in 2002 and published in Clinical Biochemistry in 2004.^[12]

Total oxidant level (total oxidant status) measurement

TOS was evaluated using an internationally accepted method developed by Erel in 2002 and published in Clinical Biochemistry in 2005.^[13]

Oxidative stress ındex: Oxidative stress index (arbitrary unit)

OSI was calculated using the following formula: TOS (μmol H₂O₂ equivalent/L)/TAS (μmol Trolox equivalent/L).^[12],[13]

Statistical analysis

The Number Cruncher Statistical System 2007 and Power Analysis and Sample Size 2008 statistical software (Utah, USA) was used for statistical analysis.

Ethics committee approval was obtained for the study (Date: April 03, 2012, No.: B.10.4. ISM. 04.34.26.08–185).

Results

Twenty-four patients (14 women [58.3%]) and 24 healthy volunteers (13 women [54.2%]) were included in the study. The mean age of the patients was 36.04 ± 10.21 (minimum–maximum: 18–49) years, and the mean age of the control group was 33.96 ± 9.65 (minimum–maximum: 18–50) years (P = 0.999). The clinical and laboratory information of the patients are summarized in [Table 1].{Table 1}

The mean TAS, TOS, OSI, and PON values of the patients (n = 24) during the attack period were 1.44 ± 0.21 (mean deviation [MD]: 1.49); 5.99 ± 4.10 (MD: 4.95); 0.41 ± 0.23 (MD: 0.33); and 192.17 ± 133.22 (MD: 110.50), respectively. The mean TAS, TOS, OSI, PON results of the patients in the remission periods were 1.44 ± 0.27 (MD: 1.45); 6.99 ± 4.95 (MD: 5.26); 0.48 ± 0.28 (MD: 0.35); and 189.29 ± 127 ± 68 (MD: 100.50), respectively. There were no differences serum findings between attack and remission periods (P = 0.841; P = 0.590; P = 0.305; P = 0.658, respectively).

The mean TAS, TOS, OSI, and PON results of the control group were 1.48 ± 0.52 (MD: 1.58); 6.82 ± 4.01 (MD: 6.04); 0.43 ± 0.17 (MD: 0.43); and 196.17 ± 145.84 (MD: 128.00), respectively. No significant result was obtained in the mean TAS, TOS, OSI, and PON results of the patients during the attack period and remission period compared with the control group (for remission periods: P = 0.252, P = 0.635, P = 0.959, P = 0.765; for the attack period: 0.190, P = 0.317, P = 0.394, P = 0.869, respectively).

The TAS, TOS, OSI, and PON results of the patient group in relapse and remission were evaluated in terms of age, blood draw time, and the number of attacks and no significant result was shown [Table 2]. In addition, no significant difference was observed in the comparison of sex, lesion location (cranial, spinal, cranial and spinal), and initial symptom characteristics (sensory, visual, motor) during the attack period, and also the remission period (P > 0.05).{Table 2}

Similar to the other results, no statistically significant relationship was found between disease duration and the TAS, TOS, OSI, and PON measurements of the patients at the time of attack (P = 0.554, P = 0.849, P = 0.778, P = 0.586). However, a positive correlation of 43.5% (TAS value increases as the duration of the disease increases) between disease duration and the TAS measurements of the patients in remission was noted (r = 0.435; P = 0.034) [Table 2] and [Figure 1].{Figure 1}

While there was no significant difference between the attack EDSS score and TAS (P = 0.348), TOS (P = 0.348), OSI (P = 0.272), and PON (P = 0.861) measurements during the attack, the same result was also valid for serum antioxidant levels in the remission period (TAS: P = 0.967, TOS: P = 0.652, OSI: P = 0.698, PON: P = 0.386).

Discussion

In our study, there were no statistically significant differences in the attack period, RRMS remission, and control group for TAS, TOS, OSI, and PON values. In determining the EDSS scores during attacks and remission, age, and patient lesion locations, no relevant findings were found for the parameters studied, but a statistically significant positive difference between TAS remission measures and disease duration was identified when disease duration was investigated (r = 0.435; P = 0.034). In this case, the remission TAS value increases with increased disease duration and was tested as a compensatory response for oxidative stress.

In the autopsy study conducted by Fischer et al . on 48 cadavers (30 died with MS and 18 dies with nonneurologic diseases), it was reported that the expression of mitochondrial enzymes that cause the production of reactive oxygen radicals was increased in active MS lesions, oxidized DNA and lipids were found in large amounts in MS lesions where tissue damage was intense, and the importance of oxidative stress in the pathophysiology of MS was emphasized.^[16] Significant oxidative stress findings in autopsy studies reported different results in serum examinations. It is worth noting that related research provided varying results in the literature. For instance, Kurban et al . examined total oxidant level (TOS) and PON activity in 50 patients with RRMS and 35 healthy people, and no significant difference was found between the patient and control group.^[9] Similarly, in the study of Hadžović-Džuvo et al ., TAS was evaluated in 33 patients with MS and 24 healthy people, and serum TAS levels were found to be significantly lower in the patient group. The authors emphasized that antioxidant deficiency was important in the disease process.^[8] However, in the study of Acar et al ., in which they measured TAS, TOS, and OSI levels in 35 patients with RRMS and 32 healthy people, TOS and OSI levels were found to be significantly higher in the patient group, and TAS levels were found to be low.^[17]

Several studies examined PON activities and gene polymorphisms and indicated that genetic polymorphisms were not important in the development of MS.^{[10],[18],[19]} In the study of Sdoti et al ., with 209 RRMS and 213 healthy people, it was shared that only the PON-1 55 LM/MM genotype was a risk factor for MS development.

In another study by Ferretti et al . comparing patients with MS in remission and controls, PON activity was found to be significantly lower in patients than in controls, and lipid peroxidation products were found to be higher. In addition, a negative correlation was found between EDSS and PON activity.^[5] However, the most important problem in the study is that the mean age of the patient group was significantly older than the control group. It is the thought by Ferretti et al . that the high average age and low PON activity in the MS group may be the cause because PON activity is known to decrease with age.{Table 3}

Total antioxidant level (TAS) was calculated in another study in patients with secondary progressive MS to best represent the organism's antioxidant status. Antibodies against oxidized LDL in the patients were found to be significantly higher than in controls, whereas total antioxidant levels were significantly lower. These findings indicate that oxidative stress plays a more important role in the progression of the disease in patients with secondary progressive MS.^[3] Antioxidants are thought to use free radicals to exercise their protective effects, reduce BBB permeability, prevent leucocyte migration, and inhibit myelin phagocytoses.^[20]

When the results of similar studies conducted analyzing antioxidant molecules are considered individually and together with our study, the overall level of antioxidants is generally statistically nonsignificant, but individual antioxidant molecules often produce important results. Therefore, in addition to overall antioxidant levels, levels of some individual antioxidant molecules may carry more importance in monitoring the progression and neurodegeneration of the disease. In several experiments testing oxidant and antioxidant molecules individually, oxidative stress rises, and antioxidants are decreased in patients with RRMS. However, no study in the literature has long-term follow-up of blood antioxidant levels. In our research, this elevation of TAS in the early period of the disease may indicate a protective antioxidant response. As the duration of the disease increases, it may be useful in terms of following the progression of the disease, considering the significant decrease in TAS levels in the late period. These findings and follow-up studies of serum oxidative stress parameters indicate that antioxidants may also help current MS therapies.

Patients were randomly and voluntarily chosen, with low average EDSS ratings. The average EDSS scores are higher in literature with high TAS rates in studies conducted on this topic. This was observed as a limitation of our research, and TAS levels were expected to decrease as the disease continued for longer periods.

Conclusion

Consequently, our study shows agreement with other studies in the literature in which serum TAS, TOS, and PON in patients RRMS were studied. TOS and OSI, which were found to be nonsignificant in patients with RRMS, are high in patients with SPMS and PPMS where the neurodegeneration process is more prominent, and TAS and PON were low in various studies, suggesting that oxidative stress is more prominent in progressive forms of MS.

As in the literature, our results showed that oxidative stress parameters, being significant in progressive forms of MS, were not significant in the early MS-period. Our findings show that TAS levels are elevated at the onset of the disease, and other studies have shown that they are low in advanced MS; when all findings are evaluated together, it can be said that TAS levels decrease as the disease progresses. This may be useful in terms of progression follow-up for patients with MS.

Acknowledgment

We would like to thank Bakırköy Mental Health Hospital Foundation for financial support to our study.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.

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