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Contents UBJ, 2024, Volume 96, Issue 1

In the footsteps of a biochemical luminary: the scientific legacy of Yakub Parnas

L. Kobylinska

Department of Biochemistry, Danylo Halytsky Lviv National Medical University, Lviv, Ukraine;
e-mail: Kobylinska_Lesya@meduniv.lviv.ua

Received: 10 January 2024; Revised: 20 February 2024;
Accepted: 21 February 2024; Available on-line: 26 February 2024

The article is dedicated to commemorating Yakub Karol Parnas, an eminent figure in biochemistry and a professor in the Department of Physiological (Biological) Chemistry at Lviv Medical University. It underscores the profound impact of Yakub Parnas on Ukrainian biochemistry, shedding light on intriguing facets­ of his personality during his highly productive two decades at Lviv University. His research significantly contributed to the unveiling of anaerobic glucose metabolism, later acknowledged as the Embden-Meyerhoff-Parnas pathway. Professor Yakub Parnas and his Lviv school conducted pivotal studies on enzymatic transformations associated with anaerobic muscle function and alcoholic fermentation, glycogen phosphorolysis, revelation of ATP synthesis within glycolysis, pioneering use of radioactively labeled phosphorus in biological experiments.

Мixed-ligand complexes of germanium – 3d-metal with 1-hydroxyethane-1,1-diphosphonic acid аnd 2,2′-bipyridine as modulators of Bacillus sp. IMV B-7883 elastase and fibrinogenase activity

O. V. Gudzenko1*, L. D. Varbanets1, I. I. Seifullina2,
О. E. Martsynko2, K. K. Tsymbalyuk2,3

1Zabolotny Institute of Microbiology and Virology,
National Academy of Sciences of Ukraine, Kyiv;
2Odesa Mechnikov National University, Ukraine;
3LLC “INSPECTORAT UKRAINE”, Odesa, Ukraine;
*e-mail:alena.gudzenko81@gmail.com

Received: 07 November 2023; Revised: 11 December 2023;
Accepted: 01 February 2024; Available on-line: 26 February 2024

We have previously shown that Bacillus sp. IMV B-7883 exhibits both elastase and fibrinogenolytic activity. One of the approaches to enhance enzymatic activity is the use of coordination compounds capable to affect enzyme’s activity or synthesis. The purpose of this work was to study the effect of mixed-ligand complexes of Ge(IV) – Co(II) (Ni(II), Cu(II)) with 1-hydroxyethane-1,1-diphosphonic acid аnd 2,2′-bipyridine on the activity of elastase and fibrinogenase purified from Bacillus sp. IMV B-7883. Previously synthesized and characterized mixed-ligand complexes and enzymes purified from the supernatant of the bacterial culture liquid were used in the study. Elastase activity was determined colorimetrically with the use of Congo red, fibrinogenase activity was estimated by fibrinogen hydrolysis measured by absorption at 275 nm. It was shown that complexes 1 (C132H164Co4Ge6N24O68P12) and 2 (C132H148Ge6N24Ni4O60P12) inhibited activity of Bacillus sp. IMV B-7883 elastase by 54 and 71% respectively, while complex 3 (C92H128Cu4Ge6N16O63P12) enhanced it by 30%. Stimulating effect of all three complexes on fibrinogenase activity was revealed. Thus, complex 1 and 2 activated the enzyme by more than 50% and complex 1 – by 19%. The data obtained indicate a complex mechanism of the studied complexes influence on enzymatic activity depending on both their composition and structure.

Multifunctional chitosan-based hydrogels: characterization and evaluation of biocompatibility and biodegradability in vitro

N. Manko1, M. Lootsik1, V. Antonyuk1,2, I. Ivasechko1,
N. Skorokhyd1, H. Kosiakova3, O. Mehed’3, T. Horid’ko3,
N. Hula3, O. Klyuchivska1, R. Panchuk1, N. Pokhodylo4,
О. Barabash4, T. Dumych2, R. Stoika1,4*

1Institute of Cell Biology, National Academy of Sciences of Ukraine, Lviv;
2Danylo Halytsky National Medical University of Lviv, Lviv, Ukraine;
3Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
4Ivan Franko National University of Lviv, Lviv, Ukraine;
*e-mail: stoika.rostyslav@gmail.com

Received: 29 November 2023; Revised: 31 December 2023;
Accepted: 01 February 2024; Available on-line: 26 February 2024

Creation of novel remedies efficient in supporting wound healing remains an actual task in pharmacology. Hydrogels showed high efficiency in wound healing and tissue regeneration due to viscosity, elasticity and fluidity that provide them with functional characteristics similar to that in extracellular matrix. The aim of the study was to create chitosan-based hydrogels functionalized with different components (chondroitin-6-sulfate, hyaluronic acid, N-stearoylethanolamine) and to estimate their biocompatibility and biodegradabili­ty in vitro. For the first time, a lipid substance N-stearoylethanolamine (NSE) known as suppressor of pro-inflammatory cytokines expression was used as hydrogel component (1.95 mg/g). FTIR analysis confirmed the complexation of chitosan molecule with hyaluronate, chondroitin-6-sulfate, NSE. MTT-test and Trypan blue exclusion test were used to study hydrogels cytotoxicity towards human cells of different tissue origin. Biodegradability of hydrogels was evaluated using direct hydrogel contact with cells and cell-independent degradation. It was shown that chondroitin-6-sulfate (<2 mg/ml), hyaluronic acid (<2 mg/ml) and NSE (26 µg/ml) did not demonstrate significant toxic effects towards pseudonormal human cells of the MCF10A, HaCat, HEK293 lines and mouse cells of the Balb/3T3 line. The studied hydrogels were stable in saline solution, while in a complete culture medium containing 10% fetal bovine blood serum they underwent degradation in >24 h. The identified biodegradability of the chitosan-based hydrogels is important for the release of noncovalently immobilized NSE into biological medium. Further studies on laboratory animals with experimental wounds are expected to explore the potential of created hydrogels as anti-inflammatory and wound-healing agents.

Free radical processes in the liver mitochondria of rats exposed to diethyl phthalate

O. V. Ketsa*, A. P. Husliakova, M. M. Marchenko

Yuriy Fedkovych Chernivtsi National University, Chernivtsi, Ukraine;
*e-mail: o.ketsa@chnu.edu.ua

Received: 11 October 2023; Revised: 28 November 2023;
Accepted: 01 February 2024; Available on-line: 26 February 2024

Diethyl phthalate (DEF) is a synthetic chemical widely used as plasticizer and additive in personal care and pharmaceutical products. Low-dose exposure to this xenobiotic over a long period contributes to its intake into the human body in a high doses. To date, studies of DEF influence on free radical processes in liver cells, in which it is not only metabolized but can also have a prooxidant effect, remain limited. The aim of our research was to determine ROS formation, the intensity of oxidative modification of proteins (OMP) and proteolytic activity in the mitochondrial fraction of liver tissue of rats exposed to diethyl phtalate. The experimental white outbred rats were divided into three groups: I – intact animals (control), II and III – rats administered DEF orally for three weeks at a doses that reflect the dose levels received by humans, namely 2.5 and 5.4 mg/kg b.w. respectively. The animals were euthanized on the 14th and 21st day after xenobiotic administration. The mitochondrial fraction from the rat liver was isolated and the level of superoxide and hydroxyl radicals, protein carbonyl derivatives, SH-groups and Schiff bases was determined. Proteolytic activity was assessed in the test of hemoglobin cleavage. It was shown that administration of DEF in a dose of 2.5 mg/kg initiated ROS generation and OMP intensification in the rat liver mitochondria only with prolonged administration for 21 days, whereas its administration in a dose of 5.4 mg/kg led to intensification of these processes already on day 14th followed by further amplification on day 21st. The activity of proteolytic enzymes in the mitochondrial fraction was found to be depended on the degree of DEF–induced OMP and was increased with minor OMP intensification but decreased with significant intensification of proteins free radical oxidation.

Fetuin-A level in patients with untreated thyroid dysfunction

A. S. Idan1*, M. I. Hamzah1, M. S. Khudhair2

1Clinical Biochemistry Department, Al-Nahrain University, Medicine College, Iraq;
2Internal Medicine Department, Al-Nahrain University, Medicine College, Iraq;
*e-mail: alaas3223@gmail.com

Received: 26 August 2023; Revised: 08 November 2023;
Accepted: 01 February 2024; Available on-line: 26 February 2024

Fetuin-A, a plasma glycoprotein, has been demonstrated to play an essential role in the pathogene­sis of several metabolic disorders. This study aimed to estimate fetuin-A serum level in patients with newly diagnosed primary hyperthyroidism (PHT) and subclinical hypothyroidism (SCH) and to examine its correlation with thyroid hormones level, age and sex of patients. The study involved 90 patients with untreated thyroid dysfunction verified with thyroid function test (45 with PHT and 45 with SCH) and 90 control subjects. Triiodo­thyronin (T3), tetraiodothyronin (T4), and thyroid stimulating hormone (TSH) serum concentrations were measured with enzyme-linked fluorescent assay (ELFA), fetuin-A concentration was measured with enzyme-linked immunosorbent assay (ELISA). It was demonstrated that the level of fetuin-A was significantly higher in the PHT group as compared with the control group and showed a significant positive correlation with the T3 level. In the SCH group, the level of fetuin-A was significantly lower and showed a negative correlation with TSH level. Fetuin-A level rose with age in the PHT group and was unaffected by sex in all studied groups. The perfect AUC value obtained for fetuin-A in the comparison between PHT and SCH groups suggests its potential use as a reliable diagnostic marker to differentiate between these two thyroid conditions.

Antioxidants as supplements during drug-induced thrombocytopenia: a comparative analysis of Vanillic acid, L-carnitine and Caripill™

M. Mithun, V. Rajashekaraiah*

Department of Biotechnology, School of Sciences,
JAIN (Deemed-to-be University), Bengaluru, Karnataka, India;
*e-mail: vani.rs@jainuniversity.ac.in

Received: 20 September 2023; Revised: 07 November 2023;
Accepted: 01 February 2024; Available on-line: 26 February 2024

Drug-induced thrombocytopenia (DIT) is a disorder where platelet count declines as an adverse effect of therapeutic drugs. Plant extract of C. papaya Caripill™ is known to elevate platelet count under thrombocytopenic conditions. To evaluate the contribution of supplements with antioxidant potential to treat DIT, the comparative study of Caripill™, vanillic acid L-carnitine effect on platelet count and indices of oxidative stress in a model of rat thrombocytopenia induced through oral administration of hydroxyurea was performed. Wistar rats were grouped into four categories with five animals in each group: control (without any treatment); control + antioxidants; thrombocytopenia; thrombocytopenia + antioxidants. The above-mentioned antioxidants were supplemented orally at 50 mg/kg for 7 days. The level of lipid peroxidation products­, superoxides, protein carbonyls and sulfhydryls, SOD and CAT activity in isolated platelets as oxidative stress markers, and indices of platelets aggregation and ATP secretion as functional markers were used. Vanillic acid was shown to be beneficial, similar to Caripill™, during hydroxyurea-induced thrombocytopenia by maintaining platelet functions, enhancing both the antioxidant capacity of platelets and its number. L-carnitine efficiently up-regulated the enzymatic antioxidants, maintained platelet functions and protected lipids and proteins from oxidation in thrombocytopenic rats, however, it could not improve the platelet count. These findings open new avenues for employing the studied antioxidants as supplements for therapeutic purposes.

Multiple effects of angiostatins in injured cornea

V. L. Bilous*, A. O. Tykhomyrov

Department of Enzyme Chemistry and Biochemistry, Palladin Institute of Biochemistry,
National Academy of Sciences of Ukraine, Kyiv;
*e-mail: basil.bilous@gmail.com

Received: 07 October 2023; Revised: 01 November 2023;
Accepted: 01 February 2024; Available on-line: 26 February 2024

Prolonged inflammation and excessive neovascularization of the cornea due to severe injury can impair optical clarity and lead to vision impairment. Plasminogen kringle (K) fragments, known as angiostatins (AS), play a well-established role as inhibitors of neovascularization by suppressing pro-angiogenic signaling­. However, AS effects in the cornea, beyond inhibiting the angiogenesis, are still unexplored. In this study, we estimate the protective effect of two AS variants (K1-3 and K5) against alkali burn injury induced in rabbit and rat corneas. AS K1-3 in the single doses of 0.075 or 0.75 μg (0.1 or 1.0 μM, respectively) or 0.3 μg of AS K5 (1.0 μM) were applied locally as eye drops daily for 14 days after the injury. A significant regression of corneal vessels in-growth in injured eyes treated with AS was revealed. Western blot analysis of corneal tissue lysates revealed that injury-induced overexpression of protein markers­ of hypoxia (HIF-1α), angiogenesis (VEGF), tissue remodeling and fibrosis (MMP-9), autophagy (beclin-1) and endoplasmic reticulum stress (GRP-78) was significantly reduced under AS treatment. Besides, the level of tight junctions protein ZO-1 was shown to be up-regulated after the treatment of the damaged cornea with AS K1-3. Summarizing, our study uncovered novel biological functions of the kringle-containing plasminogen fragments indicating its beneficial effects during corneal healing in the experimental model of alkali burn. The data obtained can be helpful for the development of novel efficient formulations to manage complications of ocular surface injuries.

Kinetic regularities of thiacalix[4]arene C-1087 inhibitory effect on the activity of Mg(2+)-dependent Ca(2+)-transporting ATP hydrolase in the plasma membrane of smooth muscle cells

Т. О. Veklich1, О. V. Bevza1, О. V. Maliuk1*, S. О. Kosterin1,
R. V. Rodik2, S. H. Vyshnevskyi2, V. І. Kalchenko2

1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
*e-mail: alexmaliukid@gmail.com;
2Institute of Organic Chemistry, National Academy of Sciences of Ukraine

Received: 05 November 2023; Revised: 04 January 2024;
Accepted: 01 February 2024; Available on-line: 26 February 2024

The experiments with the suspension of plasma membranes of myometrium cells, treated with 0.1% digitonin solution, were used to study kinetic regularities of the inhibitory effect of tetra-N-phenylsulfonyl trifluoroacetamidine-thiacalixarene (С-1087) on the activity of Са2+,Mg2+-ATPase. The studies demonstrated the impact of C-1087 on the cumulative effect and the maximal velocity of ATP hydrolysis. No effect of С-1087 on the affinity between Са2+,Mg2+-ATPase, and АТР, affinity and cumulative effect of Ca ions and activation coefficient for Mg ions was revealed. A considerable decrease in the maximal velocity of ATP hydrolysis evidenced a complete non-competitive mechanism of inhibiting Са2+,Mg2+-АТРase activity with thiacalix[4]arene С-1087. Computer simulation demonstrated that thiacalix[4]arene С-1087 inhibiting effect on Са2+,Mg2+-ATPase may be conditioned by the cumulative effect of four spatially oriented N-sulfonylamidine groups on the upper rim of its macrocyclic platform.

Phosphatidic acid formation and signaling in plant cells

Y. S. Kolesnikov, S. V. Kretynin, V. S. Kravets, Y. K. Bukhonska*

V.P. Kukhar Institute of Bioorganic Chemistry and Petrochemistry,
National Academy of Sciences of Ukraine, Kyiv;
*e-mail: yasya.yaroslavka@gmail.com

Received: 07 November 2023; Revised: 18 December 2023;
Accepted: 01 February 2024; Available on-line: 26 February 2024

This review conteins updated information on the structure, localization and regulation of phosphatidic acid (PA)-producing enzymes phospholipase D, phosphoinositide-specific and non-specific phospholipases C and diacylglycerol kinases is analyzed. The specific role of PA and PA-producing enzymes in plant stress signaling is discussed.