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Background: COVID-19 is a disease in several stages starting with virus replication to dysregulation in immune system response, organ failure and recovery/death. Our aim was to determine the effect of Ganoderma lucidum, lycopene, sulforaphane, royal jelly and resveratrol extract on markers of oxidative stress, inflammation, routine laboratory analyses and duration of symptoms in COVID-19 patients. Methods: The oxidative stress parameters and interleukines 6 and 8 (IL-6, IL-8), tumor necrosis factor alpha (TNF-α) were determined in order to estimate the antioxidant and the anti-inflammatory effect of the product using a spectrophotometric and a magnetic bead-based multiplex assay in serum of 30 patients with mild form of COVID-19. Results: Statistically significant differences were obtained for all investigated parameters between the treated patients and the control group. Moreover, significant differences were observed for leukocytes, neutrophil to leukocyte ratio and iron. The average duration of the symptoms was 9.4±0.487 days versus 13.1±0.483 days in the treatment and the control group, respectively (p=0.0003). Conclusion: Our results demonstrated the promising effect of Ge132+NaturalTM on reducing the oxidative stress and the IL-6, IL-8 and TNF-α levels, and symptoms duration in COVID-19 patients. The evidence presented herein suggest that the combination of Ganoderma lucidum extract, lycopene, sulforaphane, royal jelly and resveratrol could be used as a potent an adjuvant therapy in diseases accompanied by increased oxidative stress and inflammation.

Goals: Arthroscopic ganglionectomy has become an increasingly popular surgical option for dorsal wrist ganglia. The aim of this study was to describe our technique for the arthroscopic treatment of dorsal wrist ganglia and to evaluate the clinical results as well as the recurrence rate in our patients. Methods: In a prospective study, 48 patients who underwent arthroscopic treatment of dorsal ganglion of the wrist were included. Patients were evaluated preoperatively and 3 and 24 months post arthroscopy. A presence of recurrence at 24 months was recorded. The subjective outcome was evaluated with The Patient-Rated Wrist Evaluation (PRWE) and the Visual analog scale (VAS). Objective outcomes included grip strength and range of movement measurements. Results: The mean age was 36 years; the majority were women (36/48). Mean grip strength and the average flexion and extension in the wrist showed improvement after 3 months. The total PRWE score improved from 26.7 preoperatively to 10.2 at three months and 2.4 two years after surgery. After 24 months follow-up, there were only two patients with recurrence (4.2%). Conclusion: The arthroscopic resection of the dorsal wrist ganglia is a procedure with a low recurrence rate and lowest scaring and stiffness; it should be considered as a golden standard for operative treatment of the dorsal wrist ganglia.

Background/aim: Hematological parameters are the starting point in COVID-19 severity classification. The aim of this study was to analyze oxidative stress in hospitalized COVID-19 patients and to determine its association with D-dimer, neutrophil to lymphocyte ratio (NLR), and platelets to lymphocyte ratio (PLR) as markers for disease progression. Materials and methods: 52 patients with moderate and severe forms of COVID-19 were enrolled. A hematological and coagulation profile was performed for each patient. PAT (total antioxidant power, iron-reducing) and d-ROMs (plasma peroxides) were determined in serum at admission and 7 days after hospitalization. Results: The severe group presented parameters that indicated a poor prognosis. Patients that recovered had a significant reduction in d-ROM (t-test, p Conclusion: Our results show that oxidative stress markers can be used as a tool for disease progression in COVID-19. This analysis is easily accessible and affordable in addition to conventional hematological parameters performed for severity classification.

Background/aim: Allergic rhinitis (AR) is characterized by chronic inflammation of the nasal mucosa. Under the influence of exogenous factors - allergens, reactive oxygen species (ROS) are released during cellular metabolism. They induce a series of pathological changes in the mucosa. Oxidative stress is а result of an imbalance between the production of ROS and the ability to neutralize them. The aim of this study is to compare the levels of oxidative stress between healthy children and children with allergic rhinitis. Material and methods: A total number of 60 children were included (30 healthy children and 30 children with AR). The oxidative stress index was determined by using the FRAS 5 (Free Radical Analytical System) Bravo system. Demographic characteristics, medical history, children’s living conditions and eating habits were obtained from the questionnaire. Anthropometric measurements and the absolute number of eosinophils in the peripheral smear were performed on each child. Results: This study showed high oxidative stress index and a significantly higher value of the absolute number of eosinophils in the peripheral smear in children with AR in comparison to healthy children (p Conclusion: Compared to healthy children, children with AR have a high index of oxidative stress, despite of the very high mean value of the concentration of water-soluble antioxidants in serum (PAT test) in the group of children with AR.

In addition to the proinflammatory state, cytokine production, and cell death, SARS-CoV-2 infection is also associated with oxidative stress as demonstrated by increase in reactive oxygen species (ROS) levels and an alteration of antioxidant defense during the infection. Proinflammatory cytokines and chemokines play an important role in respiratory infections caused by viruses including SARS-CoV-2 by activation of the adaptive immune response. In case when the response is not controlled, it can lead to lung tissue involvement in the course of acute respiratory distress syndrome (ARDS) or can result in multiple organ failure. Oxidative stress markers show good correlation with several cytokines, which can be measured at the beginning of the disease in a primary care setting to predict the course of COVID-19.

Background. The aim of this study was to evaluate the ability of severity scoring systems to predict 30-day mortality in patients with severe community-acquired pneumonia. Methods. The study included 98 patients aged ≥18 years with community acquired pneumonia hospitalized at the Intensive Care Unit of the University Clinic for Infectious Diseases in Skopje, Republic of North Macedonia, during a 3-year period. We recorded demographic, clinical and common biochemical parameters. Five severity scores were calculated at admission: CURB 65 (Confusion, Urea, Respiratory Rate, Blood pressure, Age ≥65 years), SCAP (Severe Community Acquired Pneumonia score), SAPS II (Simplified Acute Physiology Score), SOFA (Sequential Organ Failure Assessment Score) and MPM (Mortality Prediction Model). Primary outcome variable was 30-day in-hospital mortality. Results. The mean age of the patients was 59.08 ± 15.76 years, predominantly males (68%). The overall 30-day mortality was 52%. Charlson Comorbidity index was increased in non-survivors (3.72 ± 2.33) and was associated with the outcome. All severity indexes had higher values in patients who died, that showed statistical significance between the analysed groups. The areas under curve (AUC) values of the five scores for 30-day mortality were 0.670, 0.732, 0,726, 0.785 and 0.777, respectively. Conclusion. Widely used severity scores accurately detected patients with pneumonia that had increased risk for poor outcome, but none of them individually demonstrated any advantage over the others.

BACKGROUND: COVID-19 is characterized by the presence of oxidative stress. Vitamin D status has been reviewed as one of the factors that may affect disease severity. The aim of this study was to assess the relationship between serum vitamin D levels, oxidative stress markers and disease severity in hospitalized COVID-19 patients. METHODS: Vitamin D levels were measured in 33 patients with COVID-19. The total antioxidant power and plasma peroxides were determined in serum. RESULTS: Severe COVID-19 patients have lower vitamin D levels (18.39 ± 2.29 ng/mL vs. 28.47 ± 3.05 ng/mL, p < .05) and higher oxidative stress compared to the moderate group. When divided according to serum vitamin D levels, significantly higher values of LDH (604.8 ± 76.98 IU/mL vs. 261.57 ± 47.33 IU/mL) and D-dimer (5978 ± 2028ng/mL vs. 977.7 ± 172 ng/mL) were obtained in the group with vitamin D below 30 ng/mL, followed with significantly higher levels of plasma peroxides (d-ROMs: 414.9 ± 15.82 U.Carr vs. 352.4 ± 18.77 U.Carr; p < .05) and oxidative stress index (OSI: 92.25 ± 6.60 vs. 51.89 ± 6.45; p < .001). CONCLUSION: The presented data provide a justification to consider vitamin D as an important factor that could ameliorate disease severity through its anti-inflammatory and antioxidant effects.

Objectives: Olanzapine is an atypical antipsychotic that is approved across Europe, the USA, and in many other countries for oral treatment of schizophrenia and acute manic episodes in patients with bipolar disorder as well as for maintenance therapy to prevent recurrence in responders. The objective of the present study was to compare the pharmacokinetics of two 10 mg tablet formulations of Olanzapine following a single oral dose in healthy volunteers under fasting conditions, as per the European Medicine Agency (EMA) guidelines to grant marketing authorization. Methods: This study was a randomized, open-label, two-treatment, two-period, two-sequences, single-dose, cross-over design with a washout period of 14 days. Both the test and the reference products were administered as 10 mg tablets with 240 mL of water after an overnight fast in each study period. A total of twenty blood samples were collected before dosing and within 144 hours after drug administration. Adverse events were monitored, recorded, and evaluated by investigators throughout the study. Results: Of the 24 healthy adult male subjects enrolled, all of them completed both study periods. The geometric mean ratio 90% confidence intervals (CI) for fasting Cmax, AUC0-t, and AUC0-infinity were 94.83-113.71%, 95.04-105.69% and 95.94-107.00%, respectively. The 90% CI for the ratios of the three primary pharmacokinetic parameters (using log-transformed data) were within the range of 80-125%, meeting the regulatory criteria for bioequivalence. Conclusions: The generic Olanzapine was bioequivalent to the reference formulation. It was well tolerated and provides an acceptable alternative to the reference drug.

The active metabolite of azathioprine, 6-thioguanine nucleotide (6-TGN) is the main component responsible for the immunosuppressive effect in treatment of inflammatory bowel disease (IBD). The aim of this study was to assess the correlation between the concentration of 6-thioguanine nucleotide and disease activity, azathioprine-related adverse effects and time duration of treatment in patients with inflammatory bowel disease. Thirty-four patients were included in this study. Type of disease, gender, time duration of therapy and adverse effects were recorded. Metabolite concentration was determined by high performance liquid chromatography. Twenty-one percent of patients have experienced an adverse effect, with leucocytopenia most commonly occurring (42.9%). More adverse effects were registered when patients were treated with azathioprine in a period of less than 3 months in comparison to the group of patients that have been under therapy between 3-12 months and more than 12 months (p˂0.05). Most of the patients that presented any adverse effect had high 6-TGN concentration (>450 pmol/8x108 Er). The mean value of 6-TGN metabolite concentration in IBD patients treated with azathioprine was 437.46 pmol/8x108 Er ± 198.82 pmol/8x108. The time duration of azathioprine treatment did not have any significant impact on the achieved 6-TGN concentration (p>0.05).Twenty patients (58.9%) had achieved remission after therapy initiation with azathioprine. More alertness is recommended to clinicians towards patients in the first 3 months of the therapy. Our study demonstrated that higher 6-TGN concentration is associated with azathioprine toxicity.

Clarithromycin is a semi-synthetic macrolide antibiotic, chemically 6-0-methylerythromycin, formulated as immediate-release tablets, extended-release tablets, and granules for oral suspension. The objective of this study was to evaluate and compare the relative bioavailability, and therefore the bioequivalence of Clarithromycin 500 mg test formulation versus a reference Klacid® forte 500 mg formulation, following a single dose administration under fasting conditions. The study was a single center, open, single dose, randomized, two-way crossover study in healthy male volunteers, with a wash-out period of one week between study periods. Twenty-four male healthy volunteers, aged 18-49 years were included into study. Blood samples for determination of clarithromycin and 14-OH clarithromycin concentrations were withdrawn at zero (pre-drug administration), 0.33, 0.66, 1, 1.5, 2, 2.5, 3, 3.5, 4, 5, 6, 8, 12, 24 and 36 hours post-drug administration. The determination of clarithromycin and 14-OH clarithromycin concentrations in plasma was performed using validated LC/MS/MS method and internal standardization after liquid/liquid extraction with methyl t-butyl ether. The test formulation of clarithromycin, dosed at 500 mg is bioequivalent for primary clarithromycin and 14-OH clarithromycin parameters (Cmax, AUC0-t and AUC0-∞) to the reference formulation after a single oral administration of 500 mg clarithromycin. Both medications were well tolerated with no serious adverse events. Thus, in view of the clinical use, both formulations are exchangeable without restrictions. Keywords: clarithromycin, 14-OH clarithromycin, bioavailability, bioequivalence study, single-dose

Background. Clinical evidence suggests increased oxidative stress in COVID-19 patients and this worsened redox status could potentially contribute to the progression of the disease. Objectives. To investigate the oxidative stress we have measured oxidative stress parameters, namely, PAT (total antioxidant power, iron reducing) and d-ROMs (plasma peroxides). Additionally we have investigated their correlation with the most frequently used clinical parameters CRP, LDH, and NLR in serum from moderate and severe COVID-19 patients hospitalized in a tertiary hospital. Methods. PAT and d-ROMs were determined by analytical photometric metric method in serum from 50 hospitalized patients. For each of them, two samples were collected and analyzed immediately after collection seven days apart. Results. All patients at admission had a much higher value for plasma peroxides and a significant correlation between oxidative stress parameters and CRP, LDH, and NLR. (p Conclusion. We consider that using OSI at the beginning of COVID-19 disease presents a valuable starting point for the general assessment of oxidative stress and hence enabling a better triage of the patients in terms of disease severity.

Introduction: COVID-19 can be worsened by hyper-production of cytokines accompanied by increased level of oxidative stress. The aim of this study was to investigate the correlation between a set of cytokines and the markers of the oxidative stress. Methods: The levels of cytokines IL-2, IL-4, IL-6, IL8, IL-10, VEGF, IFN-γ, TNF-α, IL-1α, MCP-1 and EGF were determined by using High Sensitivity Evidence Investigator™ Biochip Array technology. The oxidative stress parameters (d-ROM, PAT, OS index) were measured in serum on FRAS5 analytical photometric system. Results: IL-6, IL-8, IL-10, VEGF, MCP-1 and EGF were significantly higher (p Conclusion: The oxidative stress markers show good correlation with the tested cytokines which can be measured at the beginning of the disease in a primary care setting to predict the course of COVID-19.

Salt sensitive hypertension is known to be a contributing factor for the progression of kidney disease. This study was undertaken to investigate the role of excessive dietary salt on renal function and to evaluate the effect of valsartan and amlodipin given as a combination therapy on blood pressure and parameters specific to the renal function in salt loaded SHR rats. 48 male SHR rats at age of 20 weeks and body weight ranging between 270-350 g were used. SHR rats were divided into 3 groups: control group of rats -SHRC (n = 16) given tab water ad libitum and two salt treated groups in which tab water was replaced with a solution of NaCl (1%) from age of 8 weeks given ad libitum: SHRVAL+AMLO group (n = 16) where investigated drugs were administered at a dose of 10 mg/kg/ b.w. (valsartan) and 5 mg/kg/ b.w. (amlodipin) by gavage and SHR NaCl group (n = 16) that received saline in the same volume and the same time intervals as the SHRVAL+AMLO group. For a period of 12 weeks we have investigated the effect of the VAL+AMLO drug combination on systolic blood pressure (SBP), body weight and renal function tests. Salt loading with 1% solution in the SHR NaCl group has lead to significant increase of blood pressure, proteinuria and decrease in creatinine clearance. Combined treatment with АТ1-receptor blocker and calcium antagonist has managed to control blood pressure and ameliorated renal damage.

Background/Aim. Matrix metalloproteinases (MMPs) are perceived to play a key role in tumor invasion and metastasis by their capacity to degrade basement membranes and extracellular matrix proteins. The aim of this study was to investigate the expressions of MMP-2, MMP-7 and MMP-9 in tumor tissue and their relation to clinicopathologic features in patients with colorectal cancer. Methods. Specimens of resected colorectal cancer and surrounding normal tissue of 82 patients were immunohistochemically stained for MMP-2, MMP-7 and MMP-9. The results of immunohistochemical expression of MMPs were correlated with some clinical and pathologic parameters. Results. Immunohistochemical expression of MMP-2 was more frequent in the patients with higher preoperative serum levels of carcinoembryonic antigen (CEA) (p = 0.047), MMP-2 (p = 0.018), MMP-9 (p = 0.036) and in those with lymph node metastasis (p = 0.018) and the advanced stage of the disease (p = 0.046). Expression of MMP-7 was more frequent in the patients with elevated preoperative serum levels of: CEA (p = 0.012), MMP-7 (p = 0.036), MMP-9 (p = 0.023) and with deeply invasive neoplasms (p = 0.027). MMP-9 cell expression was in a positive correlation with elevated preoperative serum levels of: CEA (p = 0.013), MMP-2 (p = 0.012), MMP-9 (p = 0.018) and depth of CRC invasion, i.e. Tparameter (p = 0.027). Conclusion. Immunohistochemical expression of MMPs is a useful indicator of the disease development and progression in patients with colorectal cancer.

Background: Distribution of CYP2C9 and VKORC1 gene polymorphisms may vary significantly among different ethnic groups, and eventually influence the variation in drug metabolism or even failure.Objective: The aim of this study was to evaluate the prevalence of CYP2C9 and VKORC1 alleles in the healthy population of Republic of Macedonia compared to the global geographic data reported from different ethnic populations. Also, to genotype CYP2C9 and VKORC1 genes and eventually to divide individuals in poor, extensive, or intermediate metabolizer.Material and Methods: Blood samples were collected after signing written consent, DNA was isolated from peripheral blood, and CYP2C9 and VKORC1 genes were typed (n=124). Genotyping was performed by commercially available kits (GeneID GmbH, Strassberg, Germany, AID Diagnostica), based on the method of polymerase chain reaction with a subsequent hybridization. The population genetics analysis package, PyPop ver. 0.6.0, was used for analysis of the data.Results: The frequency of alleles varies from 0.931 for CYP2C9*3 to 0.109 for CYP2C9*2 indicating common “wild type” allele in those genes. The frequency ranges spanned ~50% for each allele of VKORC1 gene, indicating no common “wild type” allele in this gene. Test of neutrality showed significant negative value for VKORC1 polymorphism that indicates balancing selection operating on the alleles at that locus. All polymorphisms of CYP2C9*2, CYP2C9*3 and VKORC1 showed a good fit with Hardy-Weinberg expectations.Conclusion: The results of polymorphic alleles of CYP2C9 and VKORC1 genes in Macedonian population can be used for the variation in drug metabolism studies as well for adapting dosage regimes for oral anticoagulant therapies.

Aim. To investigate the effect of the peroxisome proliferators-activator receptor gamma agonist, rosiglitazone, on metabolic parameters and adiponectin levels in an animal model of the metabolic syndrome. Material and methods. Metabolic syndrome was induced in 32 male Wistar rats by adding a fructose in drinking water for 12 weeks. During the last 4 weeks, 16 rats were treated with rosiglitazone (5 mg/kg/day), while the remaining 16 did not receive any medication (fructose group). Another control group consumed standard rat chow and water for 12 weeks. Results. Chronic fructose administration induced a significant increase in systolic blood pressure (SBP), body weight, serum triglycerides (TG), free fatty acids (FFA), insulin, glucose AUC 0-120 (during oral glucose tolerance test) and decreased serum high density lipoprotein (HDL) cholesterol and adiponectin concentrations compared with the control group. Treatment with rosiglitazone over the final 4 weeks reversed these effects and significantly reduced SBP, TG, FFA, insulin concentrations and glucose AUC 0-120 compared with the fructose group. In addition, rosiglitazone increased serum levels of adiponectin twofold from 3.44 ± 0.46 to 7.03 ± 1.30 μg/ ml. Conclusion. This study indicates that rosiglitazone treatment improves the components of the metabolic syndrome, which is accompanied with an increase in adiponectin concentrations. OPEN ACCESS

Clinical efficacy of cisplatin as antitumorous drug is indisputable; but its use is dose-limited due to its association with severe nephrotoxicity. The latest researches have demonstrated that epoetin alpha might have an important role not only in therapeutic aims for correction of different types of anemia, but in could also be efficient as neuroprotective, hepatoprotective, cardioprotective and especially as nephroprotective drug in neprhotoxicity induced by agents based on platinum. The main aim of this study was to determine the effect of epoetin alpha in prevention of nephrotoxicity induced experimentally in Wistar rats by long-term administration of cisplatin at dose of 2 mg/kg/b.w./per week during a period of 8 weeks. The results obtained have shown that epoetin alpha significantly alleviates the functional renal failure induced by long-term administration of cisplatin and improves the general condition as well as reduces the mortality in examined animals.

BACKGROUND: The aim of this study is estimation of pharmacokinetic parameters: Cmax, tmax, t1/2, AUC0-t and AUC0-∞ with the two-way analysis of variance, single observation (ANOVA) for two preparations containing acyclovir.OBJECTIVE: In order to evaluate pharmacokinetic study of acyclovir, method for quantitative determination of acyclovir in human plasma should be simple, rapid and reproducible. Therefore, the method is developed, validated and applied for analysis of acyclovir in plasma samples obtained from healthy volunteers.MATERIAL AND METHODS: High performance liquid chromatographic (HPLC) method with UV-detection for the determination of acyclovir in human plasma is presented. This method involves protein precipitation with 20 % (V/V) perchloric acid. The chromatographic separation was accomplished on a reversed phase C8 column with a mobile phase composed of 0.1 % (v/v) triethylamine in water (pH 2.5). No internal standard is required. UV detection was set at 255 nm. The method was successfully applied for the evaluation of pharmacokinetic profiles of acyclovir tablets in 24 healthy volunteers.RESULTS: The validation results shows that proposed method is rugged, precise (RSDs for intra- and inter-day precision ranged from 1.02 to 8.37 %) and accurate (relative errors are less than 6.66 %). The calibration curve was linear in the concentration range of 0.1-2.0 µg/ml and the limit of quantification was 0.1 µg/ml. The Cmax, tmax and AUCs for the two products were not statistically different (p>0.05), suggesting that the plasma profiles generated by Zovirax were comparable to those produced by acyclovir manufactured by Jaka 80 company.CONCLUSION: Good precision, accuracy, simplicity, sensitivity and shorter time of analysis of the method makes it particularly useful for processing of multiple samples in a limited period of time for pharmacokinetic study of acyclovir.

This study was initiated to refine and characterize a nongenetic experimental model of type 2 diabetes mellitus and to follow up various metabolic parameters up to six weeks after diabetes induction. Male Wistar rats were divided into 4 groups: CON group--consumed standard rat chow and served as control; HFD group--consumed high-fat diet (45% calories as fat); STZ group-was injected once intraperitoneally with streptozotocin (35 mg/kg) on day 14, and DM-2 group--consumed high-fat diet and was injected with streptozotocin. The metabolic parameters were measured one week after streptozotocin injection (week 3) and at the end of the study (week 9). Our results confirm that HFD-group developed dyslipidaemia, obesity and insulin resistance. All metabolic parameters remained largely unaltered in STZ-group during the study. Only the combination of high-fat diet and streptozotocin (DM-2 group) induced type 2 diabetes that was characterized with moderate hyperglycaemia, insulin resistance, hypertriglyceridaemia, elevated free fatty acids, hypercholesterolaemia and increased plasma glucagon levels at the time of diabetes onset (week 3). The observed changes of the metabolic parameters after six additional weeks demonstrated an aggravated diabetic state, as confirmed from significantly increased fasting plasma glucose values, insufficient insulin secretion, severe hyperlipidaemia, increased glucagon levels, decreased serum adiponectin concentrations and significantly elevated urinary protein excretion. These results indicate that apart from its utility as a model of diabetes aetiology, this model could also be used for elucidating the role of the hormones adiponectin and glucagon in the progression of type 2 diabetes, as well as for investigating the diabetic complications.

Introduction: Matrix metalloproteinases are produced by tumour cells, hence, they may be associated with tumour progression including invasion, migration, angiogenesis and metastasis. Finding prognostic markers to better identify patients with higher risk for poor survival would be valuable in order to customize pre- and postoperative treatment as well as to enable closer follow-up of these patients. Aim of our study was to examineMMP-2, MMP-7 and MMP-9 serum levels and correlated them with pathological data such as stage of the colorectal cancer (CRC) and outcome.Methods: The investigation included 82 patients with operable CRC without distant metastases, who had underwent blood tests in order to determine the MMP-2, MMP-7 and MMP-9 serum levels in the following time periods: preoperatively, 3, 6, 9 and 12 months postoperatively.Results: The values of the investigated MMPs decrease postoperatively and start to increase 6 month later in patients of all stages of the disease, reaching the highest value 12 month postoperatively with statistically important differences of MMP-2, MMP-7 and MMP-7 serum levels in terms of disease staging and defined points of time. Analysis of the results showed that the MMP-2 serum levels obtained 3 and 12 months postoperatively,than MMP-7 serum levels 12 months postoperatively and the MMP-9 serum levels in all analyzed points in time were in significant association with the CRC patients’outcome.Conclusion: The MMP-2, MMP-7 and especially MMP-9 serum values could be important indicators for diagnosis of the patients with CRC and for monitoring of disease progression.