Category Archives: Uncategorized
Therapeutic potential of topical autologous angiostatin application in managing tuberculosis-related corneal injury: a case report
N. Greben1*, I. Gavryliak1, V. Bilous2,
V. Korsa2, A. Tykhomyrov2
1Department of Ophthalmology, Bogomolets National Medical University, Kyiv, Ukraine;
2Department of Enzyme Chemistry and Biochemistry,
Palladin Institute of Biochemistry, Kyiv, Ukraine;
*e-mail: nkgreden@ukr.net
Received: 03 June 2025; Revised: 17 September 2025;
Accepted: 28 November 2025; Available on-line: 29 December 2025
Ocular tuberculosis (TB) is a vision-threatening condition that frequently manifests as corneal neovascularization and stromal keratitis, which triggers a cascade of inflammatory and hypoxia-driven responses. Conventional therapeutic approaches, including corticosteroids and antimicrobial agents, often fail to halt disease progression. Here, we report a case of a 50-year-old patient diagnosed with TB-associated keratitis, unresponsive to standard treatment. The aim of the study was to evaluate the effectiveness of the alternative therapeutic strategy involving topical administration of angiostatin, a natural anti-angiogenic polypeptide derived from the autologous plasminogen. Solution of angiostatin fragment containing the first three kringle domains (K1-3) was applied in a two doses of eye drops (~15 μg per administration) five times daily for 2 months, with a cumulative exposure of approximately 4.5 mg. Treatment efficacy was monitored using both standard ophthalmologic assessments and non-invasive biochemical indicators such as the levels of hypoxia-inducible factor HIF-1α, vascular endothelial growth factor (VEGF), matrix metalloproteinase (MMP-9), fibrinogen/fibrin (Fg/Fb) and lactoferrin measured the in tear fluid across treatment time points (Day 0, 14, and 61) using Western blot analysis. The high intensity of HIF-1 α, VEGF and MMP-9 expression, Fg/Fb accumulation and the presence of low-molecular-weight fragments of lactoferrin were detected in the tear fluid prior to the treatment. Following angiostatin therapy, the patient exhibited marked regression of corneal neovascularization and restoration of corneal transparency, complemented with normalization of HIF-1α, VEGF, and MMP-9 levels, reduced Fg/Fb accumulation and the presence of intact lactoferrin in the tear fluid. The data obtained demonstrated a multifactorial mechanism of angiostatin action that extends beyond classical anti-angiogenic pathways. The convergence of clinical and molecular indicators of recovery underscores the potential of angiostatin application as a safe and effective therapeutic alternative for managing corneal complications in ocular TB, particularly in cases resistant to conventional treatment.
The fatty acid composition of cell lipids in walnut bacterial pathogens
M. I. Zarudniak, L. A. Dankevych*, I. P. Tokovenko, V. P. Patyka
D. K. Zabolotny Institute of Microbiology and Virology,
*e-mail: ldankevich@ukr.net
Received: 23 April 2025; Revised: 25 September 2025;
Accepted: 28 November 2025; Available on-line: 23 December 2025
Walnut (Juglans regia) is the most economically important and widespread nut crop in Ukraine. As bacterial diseases of walnut can reduce the yield of this culture by up to 40%, the monitoring of pathogens in a given crop and their identification are extremely important. The fatty acid composition of cell lipids is used in the taxonomy of plant pathogenic bacteria. The objective of this study was to determine the fatty acid composition of cell lipids of Agrobacterium, Xanthomonas, and Pseudomonas collection strains that can actually infect walnut, and those isolated from affected walnut trees in different regions of Ukraine. Fatty acid methyl esters were obtained by two different methods of extraction, with the use of 5% acetyl chloride in methanol at 100°C for 4 h, or 1.5% sulfuric acid in methanol at 80°C for 1 hour. Fatty acid methyl esters were analyzed using gas chromatography–mass spectrometry system. According to the found similarity of the fatty acid composition, the strains isolated from the affected walnut were related to representative collection strains of A. tumefaciens, X. arboricola and P. syringae. It should be noted that during the isolation of fatty acids with the use of 1.5% solution of H2SO4 in methanol, the amount of individual saturated and unsaturated fatty acids in the studied strains decreased and almost all hydroxyl acids, identified as a key taxonomic markers, disappeared in comparison with the using of 5% solution of acetyl chloride in methanol at the hydrolysis stage.
Isolation and characterization of collagenase-active preparation from Rapana venosa salivary glands
V. A. Toptikov*, Ye. A. Shesterenko, Yu. A. Shesterenko
Medical Biotechnology and Enzymology Laboratory, Department of Biomedicine,
A.V. Bogatsky Physico-Chemical Institute, National Academy of Sciences of Ukraine, Odesa;
*e-mail: v.a.toptikov@gmail.com
Received: 02 July2025; Revised: 28 August 2025;
Accepted: 28 November 2025; Available on-line: 23 December 2025
Collagenases have found practical applications in both medicine and the food industry, but the search for novel collagenase sources remains an active area of studies. Rapana, a predatory mollusk that primarily feeds on bivalves rich in connective tissue, has emerged as a potential source of collagenolytic enzymes. This study aimed to isolate collagenase-active preparation from Rapana venosa salivary glands and characterize its properties. The salivary gland extract was purified by acetone precipitation followed by ammonium sulfate treatment. Electrophoresis was performed by the Laemmli protocol under both reducing and non-reducing conditions. Proteolytic activity was determined spectrophotometrically using collagen or gelatin as a substrate. The preparation consisted of five protein fractions and exhibited enzymatic polymorphism. A 13.8-fold purification of collagenase activity was achieved, at least 22% of total proteins displayed collagenolytic activity, while 88% showed gelatinolytic activity. The optimum of preparation activity was found in acidic (pH 4.5) and alkaline (-9.5) ranges, with thermal optimum at 46°C. At room temperature, about 90% of activity was maintained for 8 h. Serine protease inhibitors did not affect enzyme activity, metal ion chelators completely inhibited it. Reducing agents enhancing SH-groups increased enzyme activity, disulfide bond regeneration or SH-group modification decreased it. The data obtained showed that the collagenase-active enzyme preparation from Rapana venosa salivary glands consists mainly of metalloproteinases and cysteine proteases, exhibiting high stability.
Enzyme-linked immunosorbent assay for the determination of total prostate-specific antigen
K. M. Shevchuk1, O. B. Besarab1*, Yu. V. Gorshunov1, O. Yu. Galkin1,2
1National Technical University of Ukraine “Igor Sikorsky Kyiv Polytechnic Institute”, Kyiv;
2Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
*e-mail: besarab@lll.kpi.ua
Received: 26 August 2025; Revised: 08 October 2025;
Accepted: 28 November 2025; Available on-line: 23 December 2025
Prostate-specific antigen (PSA) remains the most widely used biomarker for prostate cancer diagnostics and monitoring. The development of highly informative, sensitive, and reproducible immunoassays for PSA determination is essential for improving diagnostic accuracy. The aim of the study was to develop and optimize a non-competitive “sandwich” ELISA for the determination of total PSA, using a panel of monoclonal antibodies (mAbs) of different specificity groups and epitopes. “Sandwich” ELISA configurations were designed using different capture and detecting antibody pairs. Antibody sorption conditions, reagents working concentrations, incubation parameters and buffer composition were optimized. Analytical performance was evaluated using PSA preparations standardized against the WHO International Standard (96/670). The most effective antibody combinations were found among mAbs targeting epitopes P2 (capture: 21B7, 11G5, 26B9) and P3 (detection: 21F4, 23B4, 27C10), with the high-affinity pair 26B9-27C10 showing the best sorption–detection properties. The use of mixed conjugates did not improve sensitivity in the range of 1-10 ng/ml PSA. Hydrophilization of polystyrene plates surface increased the ELISA signal up to 1.39-fold, depending on the antibody isotype and origin. The developed optimized “sandwich” ELISA demonstrates high specificity and sensitivity for total PSA determination, with analytical characteristics suitable for potential clinical diagnostic applications.
Angiotensin, vascular endothelial growth factor and caspase-3 levels in blood serum of smoking students
I. A. A. K. Al-Samarai1, A. J. Al Samer1, H. N. Mohammed2
1Department of Applied Chemistry, College of Applied Science,
University of Samarra, Saleh Aden, Iraq;
2Department of Biotechnology, College of Applied Science,
University of Samarra, Saleh Aden, Iraq;
e-mail: israa.a@uosamarra.edu.iq
Received: 18 July 2025; Revised: 18 September 2025;
Accepted: 28 November 2025; Available on-line: 23 December 2025
Smoking cigarettes is currently considered a widespread behavioral habit among university students due to psychological, social, and behavioral factors. Smoking is believed to impair renin-angiotensin system, blood pressure regulation, endothelial function, and cells viability, particularly in the lungs or blood vessels. The study aimed to assess the level of angiotensin, vascular endothelial growth factor (VEGF), caspase-3 and the activity of antioxidants glutathione S-transferase (GST) and superoxide dismutase (SOD) in the blood serum of students at Samarra University (Iraq). The study lasted from 20/2 /2025 to 20/4/ 2025 and involved 100 male students aged 18-28 years. The first group consisted of 30 nonsmoker students and the second included 70 smoker students, whose daily cigarette consumption ranged between 60-100 cigarettes. The results showed a significant increase in angiotensin, VEGF and caspase-3 levels, measured by ELISA, and a significant decrease in GST and SOD activity in the blood serum of smoker students compared to nonsmokers. A high negative correlation between angiotensin, GST and SOD activity in both smokers and nonsmokers, and a positive correlation between angiotensin and caspase-3 levels in smokers were observed, indicating the promising use of studied parameters as indicators of adverse effects caused by smoking.
Inflammatory cytokines profile and oxidative stress markers in the serum of albino rats injected with macrophage migration inhibitory factor
N. T. Guliyeva1, S. V. Guliyeva2*, R. A. Akhundov2,
N. R. Jabbarova3, T. A. Eyvazov2
1Department of Cytology, Embryology and Histology,
Azerbaijan Medical University, Baku, Azerbaijan;
2Research Center, Azerbaijan Medical University, Baku, Azerbaijan;
3Department of Health Care Organization,
Azerbaijan Medical University, Baku, Azerbaijan;
*e-mail: quliyevasevda789@gmail.com
Received: 09 July 2025; Revised: 28 August 2025;
Accepted: 28 November 2025; Available on-line: 23 December 2025
Macrophage migration inhibitory factor (MIF) is a pleiotropic cytokine involved in the regulation of inflammation, immune responses, and redox homeostasis. However, its metabolic effects in experimental models remain insufficiently characterized. The aim of the work was to estimate the effect of recombinant MIF on cytokines profile, antioxidant defense markers and LPO indicators at different time points following its single intraperitoneal administration to albino rats. Animals were divided into a control group (n = 20) and three experimental groups (n = 10 each) assessed in 2, 3, and 14 days after MIF administration (10 µg/kg of b.w.), respectively. Serum samples were analyzed for IL-6, IL-10, TNF-α, IL-4, antioxidant markers and LPO products levels by ELISA and standard biochemical assays. It was shown that MIF administration induced time-dependent pro-inflammatory and pro-oxidant effects. Early compensatory anti-inflammatory responses were marked by increased IL-10 and decreased IL-6 levels. However, at the later stages (days 3 and 14), IL-6 and TNF-α elevation, along with IL-4 suppression, indicated a shift toward chronic inflammation. Antioxidant parameters progressively declined, with maximal suppression observed on day 14. Concurrently, a significant accumulation of LPO products confirmed sustained oxidative stress and membrane damage. These findings underscore the potential of MIF as a pharmacological target for the treatment of chronic inflammatory and metabolic disorders.
Purification and physico-chemical properties of Bacillus sp. L9 protease with fibrin(ogen)olytic activity
O. V. Gudzenko1*, L. D. Varbanets1, V. O. Chernyshenko2, E. M. Stognii2
1D.K. Zabolotny Institute of Microbiology and Virology,
National Academy of Sciences of Ukraine, Kyiv;
2Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
*e-mail: alena.gudzenko81@gmail.com
Received: 18 September 2025; Revised: 13 October 2025;
Accepted: 28 November 2025; Available on-line: 23 December 2025
Previously, we isolated a number of Bacillus sp. strains from the dry grass of the coastal zone of the Kinburn Spit, which may be promising for further research as producers of proteases with fibrinolytic and fibrinogenolytic activity. The aim of the work was to isolate, purify and study the properties of fibrin(ogen)ase from the Bacillus sp. L9 strain. The enzyme preparation was isolated from the supernatant of the Bacillus sp. L9 culture liquid. The yield of the purified enzyme was 1.8%, the specific fibrinogenolytic and fibrinolytic activities were 483 and 383 U/mg protein, respectively, the molecular weight of the enzyme was about 40 kDa, the optimum pH was 8.0, and the thermooptimum was 40°C. Bacillus sp. L9 fibrin(ogen)ase is a serine protease, in the active center of which is the carboxyl group of the C-terminal (aspartic or glutamic) amino acid. At some distance from the active site are localized sulfhydryl groups that do not participate in catalysis, but play an important role in maintaining the catalytically active conformation of the protein molecule. The enzyme from Bacillus sp. L9 hydrolyzed fibrin molecules much more slowly than fibrinogen, and showed the greatest specificity in the hydrolysis of bonds formed by the Aα-chain of fibrinogen. According to the specificity of action on fibrinogen, the enzyme was identified as α-fibrinogen(ogen)ase.
Identification of different subtypes of K(+) channels in the mitochondria of rat myometrium using K(+) channels modulators
M. V. Rudnytska, H. V. Danylovych*, M. R. Pavliuk, Yu. V. Danylovych
Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
*e-mail: danylovychanna@ukr.net
Received: 17 July 2025; Revised: 25 September 2025;
Accepted: 28 November 2025; Available on-line: 23 December 2025
Potassium ions affect Ca2+ transport in mitochondria, the magnitude of the electric potential on the inner mitochondrial membrane, metabolic processes in the matrix, and osmoregulation. The aim of this study was to identify different subtypes of K+ channels in the mitochondria of rat myometrium. Isolated mitochondria were obtained from the myometrium of non-pregnant Wistar rats by differential centrifugation. Potassium ion accumulation was studied by spectrofluorimetry using the K+-sensitive fluorescent probe PBFI-AM. Myometrial mitochondria effectively accumulate potassium ions within the concentration range of 25–150 mM. No increase in PBFI fluorescence was observed when K+ ions were replaced by choline in equimolar concentrations. In the presence of voltage-operated K+ channels inhibitor 4-aminopyridine, Ca2+-dependent K+ channels blockers charybdotoxin or paxilline, mitoKATP channels inhibitors glibenclamide, 5-hydroxydecanoic acid, or 200 μM ATP, a significant decrease in the PBFI fluorescence signal was observed. Conversely, application of Ca2+-dependent K+ channels specific activators NS11021 and NS1619, as well as of mitoKATP-specific activator cromakalim, resulted in increased mitochondrial K+ accumulation. The efficiency of K+ uptake increased further with the addition of 25–100 μM Ca²⁺ in the presence of 4-aminopyridine and ATP. The results obtained indicate the presence of voltage-operated and Ca2+-dependent subtypes of K+ channels, as well as of H+/K+ exchange system in myometrial mitochondria in addition to mitoKATP channels.