Ivana Stojanovic

Single nucleotide polymorphisms (SNPs) rs429358 and rs7412, the most commonly investigated variants in the apolipoprotein E gene (APOE) are crucial for APOEε4 carrier status determination. However, their association with inflammatory cytokine levels in patients with dementia due to Alzheimer’s disease (AD) remains unclear. This study aimed to investigate the influence of the APOEε4 allele on pro-inflammatory cytokine levels in the cerebrospinal fluid of patients with AD dementia. The research was conducted on 36 patients with probable dementia due to AD. APOE rs429358 and rs7412 were analyzed using the Real-Time PCR method with allele-specific TaqMan assays, followed by the analysis of APOEε4 allele carrier status. Patients carrying at least one APOEε4 allele were assigned as APOEε4+. Core biomarkers (Aβ42/40 ratio, t-Tau, p-Tau levels), as well as pro-inflammatory cytokine (Tumor necrosis factor-a (TNF-a) and interleukin-1b (IL-1b)) levels, were determined in the patient’s cerebrospinal fluid (CSF) using Enzyme-linked Immunosorbent Assay (ELISA). Seventeen patients (47.22%) were assigned as APOEε4+. CSF TNF-α levels were significantly higher in APOEε4+ AD dementia patients in comparison to APOEε4- patients (p<0.001), while no significant differences in IL-1b levels between these two groups were obtained. Correlation analysis showed that TNF-α negatively correlated with the Aβ42/40 ratio (p=0.033), while positive correlation with t-Tau and p-Tau was observed (p=0.001, p=0.015, respectively). These findings highlight the potential significance of TNF-α in the context of APOEε4 positivity and its implications in AD pathology.

Background/Aim. Multiple sclerosis (MS) is an immune-mediated disease of the nervous system in which the myelin sheath is destroyed during the process of neurodegeneration. Experimental autoimmune encephalomyelitis (EAE) is an animal model of MS in which preservation of myelin and remyelination of axons can improve neuron survival. The aim of the study was to evaluate the activation capacity of neuronal tissue by autoimmune inflammation and treatment with intermittent (i) theta burst stimulation (TBS)- iTBS or continuous TBS (cTBS) based on the expression profiles of nestin in astrocytes, oligodendrocytes, and neurons. Methods. Two forms of TBS ? iTBS and cTBS ? were used to extend the period during which axons can be remyelinated. It was investigated how iTBS or cTBS protocols affect the expression profiles of nestin with glial fibrillary acidic protein, myelin basic protein (MBP), and neuronal nuclear protein in rat spinal cord. Changes at the molecular level were monitored using the immunofluorescence method. Results. The obtained results showed that both protocols (iTBS and cTBS) increased the expression of nestin and MBP and reduced astrogliosis in the spinal cord of EAE rats. Conclusion. The therapeutic potential of TBS in EAE contributes to the improvement of the intrinsic ability to recover from spinal cord injury.

Although single nucleotide polymorphisms rs429358 and rs7412 in the apolipoprotein E gene (APOE) are widely investigated to analyze APOE alleles, there are no data on their association with Alzheimer?s disease (AD) dementia in a Serbian population. This study aimed to investigate the distribution of APOE ?2/?3/?4 alleles in patients with dementia due to AD and cognitively unimpaired subjects and to assess the association of the APOE ?4 allele with disease risk in the Serbian population. A case-control study included patients with dementia due to AD and cognitively unimpaired individuals. APOE rs429358 and rs7412 were analyzed using the Real-Time PCR method with allele-specific TaqMan assays, followed by APOE ?2/?3/?4 allele carrier status analysis. Patients had a significantly higher frequency of the APOE ?4 allele than the control group (P<0.001). The APOE ?4 allele was found to be associated with a 3-fold higher risk of AD dementia compared to the reference ?3 allele (P<0.001). In conclusion, this is the first study to suggest that carriers of the APOE ?4 allele have a higher risk of developing dementia due to AD than those who carry the APOE ?3 allele in the Serbian population.

Colorectal cancer (CRC) is one of the most frequent forms of malignant tumors in the human population. The literature data about the role of arginine metabolism in CRC point out its double-faced role. In three tissue specimens of 50 patients who underwent surgical resection for colon adenocarcinoma (tumor, adjacent and healthy tissues more than 10 cm from the tumor border ? at the incision margin) taken during surgery, polyamines and the concentration of NO2+NO3 and arginase activity were determined. Polyamine levels and arginase activity were significantly increased in cancer and adjacent tissue specimens compared to healthy ones, while the level of NO2+NO3 was significantly lower in cancer compared to both adjacent and healthy tissues. The high polyamine content in the adjacent colonic mucosa indicates a high proliferative potential of tumor-adjacent tissue. Although we found individual correlations indicating the possible prognostic value of arginase, the performed statistical analysis did not show a predictive significance of arginase activity in the examined tissue specimens for five-year survival of the patients. Nevertheless, the obtained results provide the rationale for further studies of arginine metabolism in tissue specimens after surgery in patients with CRC, which could be useful in the evaluation of the risk for tumor growth, recurrence, metastases and survival after surgical intervention.

Although genetic variations rs780094 and rs1260326 of the glucokinase regulatory protein gene (GCKR) could be associated with lipid profile imbalance, their influence on acute ischemic stroke (AIS) risk has not yet been established. The aim of this study was to investigate the influence of GCKR single nucleotide polymorphisms (SNPs) rs780094 and rs1260326 on lipid profile parameters in patients with AIS, and to evaluate the association of these SNPs with the risk of AIS. In a case-control study, a total of 148 subjects were screened for GCKR rs780094 and rs1260326 SNPs using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. The lipid profile was determined based on serum total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C) and triacylglycerol (TG) concentrations. The frequencies of the minor rs780094T allele and the minor rs1260326T allele were significantly lower in AIS patients compared to controls. The rs780094TT genotype and the rs1260326TT genotype were associated with decreased risk of AIS compared to wildtype carriers. In conclusion, this is the first study implying that decreased risk of AIS in rs780094 and rs1260326 homozygous minor allele carriers is not caused by dyslipidemia, but possibly by the lack of coagulation factor glycosylation.

Background/Aim. Experimental autoimmune encephalomyelitis (EAE) is a model of multiple sclerosis (MS), in which we investigated the neuroprotective effect of agmatine (AGM), known as a primary amine produced via the decarboxylation of L-arginine. Methods. Dark Agouti rats were divided into groups: control (C), Complete Freund`s Adjuvant (CFA), EAE rats decapitated on the 13th day post immunization (dpi) (EAE13) and on the 20th dpi (EAE20), EAE animals given three (EAE+AGM13) and 10 (EAE+AGM20) doses of AGM, and healthy animals ad-ministered three/10 doses of AGM (AGM). Thiobarbituric acid-reacting substances (TBARS), SH groups (SH), total glutathione (GSH), glutathione peroxidase activity (GPx), superoxide dismutase activities (tSOD, MnSOD, CuZ-nSOD) and nitrite/nitrate concentration (NO2+NO3) were assessed in plasma and brain structures [whole encephalitic mass (WEM) and brainstem (BS)]. Results. The obtained results showed that AGM treatment successfully attenuated severe clinical deficits in EAE. Applications of AGM in EAE rats induced normalized TBARS, SH, GSH, GPx and NO in WEM. In BS, AGM expressed less prominent effects, inducing normalized TBARS, GPx and NO, but no effect on SH and GSH. In both brain structures, tSOD activity lowered and normalized at the peak and in the remission phase of the disease, post-AGM treatment. The effect of AGM on the MnSOD in EAE was expressed in WEM/BS only in the remission phase as a reduced activity. Conclusion. Milder clinical form of developed EAE in rats indicates promising therapeutic effect of AGM in MS. The activated antioxidant system and suppressed oxidative/nitrosative stress development may denote a successful blockade of neu-roinflammation initiated by EAE immunization. The study implies the capability of AGM to attenuate oxidative/nitrosative damage at the peak of EAE by modulating antioxidative defense capacity during the time-course of the disease. Thus, AGM may be considered as an agent with a beneficial effect on neuroinflammation in EAE.

Background/Aim. Nowadays therapy with fixed orthodontic appliances is widely spread, having in mind positive effects it has in malocclusion treatments The side effect is increased gingivial inflammation in treated patients. The aims of this research are to estimate the inflammatory condition of gingiva in the first 6 months of orthodontic therapy on the basis of clinical parameters of sulcus bleeding index, plaque index, gingival crevicular fluid and salivary nitric oxide concentration, and to investigate role of nitric oxide as predicting factor of gingival inflammation in orthodontic patients. Methods. In this study, 30 patients of the Dental Clinic treated with fixed orthodontic appliances (11 males and 19 females), aged 15-22 years, were examined. Clinical parameters were evaluated and gingival crevicular fluid and saliva were collected, before the beginning of orthodontics therapy, and then, three and six months after it. Results. The approximate values of clinical parameters, gingival crevicular fluid and salivary nitric oxide concentration progressively increased. Low statistical significance of correlations among gingival crevicular fluid and salivary nitric oxide concentration and the measured clinical parameters were found. There is a statistically significant correlation between gingival crevicular fluid and salivary nitric oxide concentration. Conclusion. According to the obtained results, we can conclude that gingival crevicular fluid and saliva are reliable mediums for monitoring of the gingival inflammatory condition. More studies are needed to investigate a potential role of nitric oxide as predicting factor of gingival inflammation in orthodontic patients.

Background/Aim. Acute myocardial infarction (AMI) is an important cause of mortality/morbidity worldwide. Biomarkers improve diagnostic and prognostic accuracy in AMI. The aim of this study was to investigate an increase of markers of endothelial dysfunction in AMI, measured on the 3rd day after the initial event and to investigate their association with short- and long-term (3-year) prognosis (outcome). Methods. The prospective study included 108 patients with AMI in the experimental group and 50 apparently healthy subjects in the control group. Endothelin-1 (ET-1) and nitric oxide degradation products (NOx) were determined. Results. The average age of the participants in the experimental group was 62 ? 10 years and 59 ? 9 years in the control group; 74.1% of the patients in experimental group were males and 68.8% in the control group. In 74.1% of the patients, ST-elevation myocardial infarction (STEMI) was diagnosed, and 25.9% of the patients presented with non-ST-elevation myocardial infarction (NSTEMI). Thirteen (5.6%) patients died during 3 years and they had significantly higher ET-1 levels compared to survivors [4.02 (2.72?5.93) vs 3.06 (2.23?3.58) pg/mL; p = 0.015]. Endothelin- 1 in 46 (42.6%) patients with composite endpoint (3- year mortality and rehospitalization) was significantly increased compared to other patients [3.14 (2.54?4.41) vs 3.05 (2.18?3.56) pg/mL; p = 0.035]. Intrahospital complications were found in 41 (48%) patients. Participants with echocardiographically detected complications (ventricular dyskinesia, left ventricular thrombus and papillary muscle rupture) had higher ET-1 levels compared to other patients [4.02 (2.78?5.57) vs 3.06 (2.29?3.66) pg/mL; p = 0.012]. Endothelin- 1 concentration above the 75th percentile (> 3.77 pg/mL) was associated with the increased risk for composite endpoint [Log Rank (?2 = 13.44; p < 0.001)]. Patients who were rehospitalized had significantly lower NOx concentration [125.5 (111.4?143.6) vs 139.3 (116.79?165.2) ?mol/L; p = 0.04]. Endothelin-1 positively correlated with high sensitivity troponin I (hsTnI), brain natriuretic peptide (BNP) and a number of leukocytes. Conclusion. Endothelin- 1 and NOx were increased on the 3rd day after AMI, and they were predictors of worse short- and long-term (3- year) prognosis (outcome). Endothelin-1 positively correlated with conventional prognostic markers in AMI.