Tag Archives: fibrinogen
Hemostatic system imbalance in the pre-eclampsia model in rats
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1SI “Institute of Pharmacology and Toxicology, National Academy
of Medical Sciences of Ukraine”, Kyiv;
2Palladin Institute of Biochemistry, National Academy
of Sciences of Ukraine, Kyiv;
3Shupyk National Medical Academy of Postgraduate Education, Kyiv, Ukraine.
*e-mail: anna_shayakhmetova@yahoo.com
Received: 03 June 2026; Revised: 17 June 2026;
Accepted: 27 July 2026; Available on-line: 04 August 2026
Background. Preeclampsia (PE) is a multisystem disorder in pregnancy, that results from placental ischemia, oxidative stress-induced placental damage and placental factors dissemination that cause systemic endothelial dysfunction. Patients with preeclampsia often develop a severe prothrombotic state, the mechanisms of which are not fully elucidated. Objectives. The aim of this study was to assess the informativeness of hemostatic system parameters that characterize the degree of coagulation pathway activation and anticoagulant pathway capacity capacity in experimental preeclampsia model in rats for predicting the development of thrombotic complications in PE. Methods. Wistar females rats were used in the study. To induce a preeclampsia-like syndrome, N(ω)-nitro-L-arginine methyl ester (L-NAME), at a concentration of 0.3 g/l in the drinking water was administered ad libitum from the 8th to the 14th day of gestation. On gestational day 21 the rats were anesthetized and blood samples were collected. Urinary and serum concentrations of creatinine, urea, and uric acid were determined using a fully automated Chemistry Analyzer and commercially available kits. Fibrinogen, soluble fibrin monomeric complexes and protein C concentrations, prothrombin time and antithrombin III activity in the blood plasma were estimated with a standard spectrophotometric methods. Results. Increased plasma fibrinogen, significant shortening of plasma clotting time, decrease in both protein C and AT III and increase in SFMCs plasma levels were detected in rats with experimentally L-NAME induced preeclampsia, indicating not only activation of the coagulation system but also disruption of the balance between procoagulant and anticoagulant pathways. Conclusions. L-NAME-induced PE in rats resulted in a pronounced imbalance of the hemostatic system and reproduced key pathophysiological features of PE, including excessive procoagulant shift, intravascular thrombin generation and impairment of anticoagulant mechanisms, making this model a valuable tool for studying mechanisms of disease progression. Analysis of hemostasis parameters in this model, indicates that soluble fibrin, protein C and antithrombin III are the most informative markers for assessing the risk of intravascular coagulation during preeclampsia.
Markers of prothrombotic state of patients recovered after COVID-19
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1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
2Ivan Horbachevsky Ternopil Medical University, Ternopil, Ukraine;
3International Research and Innovation in Medicine Program, CSMC, Los-Angeles, CA, USA;
*e-mail: d.korolova@gmail.com
Received: 08 May 2026; Revised: 18 July 2026;
Accepted: 27 July 2026; Available on-line: 04 August 2026
Background. In recent period the attention of scientists and doctors was focused mainly on methods of COVID-19 treatment. However, clinical course of coronavirus disease strongly affects blood coagulation with the tendency for venous and arterial thrombosis, leading to long-term imbalance between coagulation and anticoagulation systems, which can cause serious danger for health of post-COVID-19 (PCD) patients. Objective. In our work we focused on the analysis of the molecular markers of hemostasis system activation in blood plasma of patients recovered after COVID-19. Methods. Blood plasma samples of 119 seropositive patients recovered from COVID-19 were analysed. The patients were divided into a high-risk group (HRG, n = 163) and a non-risk group (NRG, n = 28) according to WHO recommendations. Patients were examined in a recovery time scale of 1-30; 31-60 days and more than 2 months. Blood plasma samples of seronegative healthy volunteers were collected as the reference group (n = 19). The concentration of soluble fibrin (SF) and D-dimer was measured with sandwich ELISA, generation of activated protein C (PC) and prethrombin-1 (Pre-1)) level with assay using specific chromogenic substrates, and fibrinogen concentration with spectrophotometric method after initiation of coagulation by Ancistron-H. Results. Concentration of fibrinogen, SF and Pre-1 were increased in a blood plasma of patients of NRG and HRG groups in comparison to the reference group. Increasing of fibrinogen and Pre-1 concentrations correlated with COVID-19 stage. The fibrinogen and SF concentrations remained high more than 2 months after recovery in HRG group and elevated after 2 months after recovery in NRG group. D-dimer and protein C levels were elevated in 8% and 11% of PCD patients respectively. Conclusions. Long-term prethrombotic state was more expressed in HRG group of PCD patients. Monitoring of the main coagulation markers can indicate the risk of intravascular thrombus formation in PCD patients. SF, Pre-1 and elevated fibrinogen concentrations were selected as useful thrombosis markers in PCD patients.
Role of fibrinogen and fibrin in initiation of thrombin generation
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Palladin Institute of biochemistry, National Academy of Sciences of Ukraine, Kyiv, Ukraine;
*e-mail: nudovenko26@gmail.com
Received: 07 May 2026; Revised: 27 June 2026;
Accepted: 27 July 2026; Available on-line: 04 August 2026
Background. Thrombin is the key enzyme of blood coagulation that converts fibrinogen into fibrin and activates multiple coagulation factors. However, the temporal relationship between thrombin-mediated fibrin formation and the availability of thrombin for other physiological substrates during the initiation of coagulation remains insufficiently understood. Objective. To investigate the role of fibrinogen and fibrin in regulating thrombin activity during the initiation phase of thrombin generation in plasma. Methods. Thrombin generation was initiated in human plasma using activated partial thromboplastin time (APTT) reagent or Thromborel. Fibrin formation was monitored turbidimetrically at 405 nm. Thrombin amidolytic activity toward the chromogenic substrate S2238 was measured spectrophotometrically at 405 nm. Fibrin polymerization was inhibited with the synthetic peptide GPRP. The effects of plasma dilution and inhibition of fibrin polymerization on the temporal profiles of fibrin formation and thrombin activity were analyzed. Results. During initiation of thrombin generation with both APTT-reagent and Thromborel, the beginning of cleavage of S2238 by thrombin coincided with the end of fibrin clot formation, marked by turbidimetric curve. At 1:3 plasma dilution, fibrin formation lag time was 210 s, amidase activity lag phase was 410 s. At 1:10 dilution, the lag time for fibrin formation was 73 s, amidase activity was 200 s, indicating that dilution (and lower fibrinogen) accelerates thrombin generation. With 1 mM GPRP, fibrin and amidase activity lag times increased by an order of magnitude. Conclusions. Thrombin generated during initiation of thrombin generation acts primarily on fibrinogen, remaining bound to the molecule throughout its conversion into fibrin and clot formation. Its amidase activity emerges only after protofibrils undergo lateral association into fibrin clot. Once the binding sites in the fibrin clot are saturated, thrombin becomes available for other substrates (FV, FVIII, FXI) and physiological functions.
Protein mysterious structure and numerous functions department
V. O. Chernyshenko*, V. I. Gryshchuk
Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine,
Department of Protein Structure and Function, Kyiv;
*e-mail: bio.cherv@gmail.com
Received: 02 July 2025; Revised: 30 July 2025;
Accepted: 30 October 2025; Available on-line: 02 December 2025
This overview is dedicated to the history of the Department of Protein Structure and Function at the Palladin Institute of Biochemistry of the National Academy of Sciences of Ukraine. It outlines the department’s main scientific objectives and highlights its key achievements. In particular, it describes research related to patient blood management, fundamental studies of fibrinogen structure and function, the development of next-generation antithrombotic prototypes, as well as the creation and clinical testing of diagnostic assays. Special attention is given to inventions and studies aimed at addressing pressing social issues, including post-traumatic stress disorder and the consequences of COVID-19. The importance of maintaining continuity in hemostasis research is emphasized, as it enables the generation of unique scientific results and their implementation in medical practice.
Validation of the diagnostics algorithm to monitor coagulation parameters in pregnant women
D. S. Korolova1, A. O. Pavlenko1, A. Altorjay2, S. I. Zhuk3,
I. V. Us3, Y. Tsaryk1, A. Suranyi2, V. O. Chernyshenko1*
1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
2Albert Szent-Györgyi Medical School, University of Szeged, Hungary;
3P. L. Shupyk National Medical Academy of Postgraduate Education, Kyiv, Ukraine;
*e-mail: bio.cherv@gmail.com
Received: 19 May 2023; Revised: 05 June 2023;
Accepted: 07 June 2023; Available on-line: 11 July 2023
Thrombotic events are among the most dangerous complications of pregnancy. Therefore, selection of appropriate tests and standardization of techniques used for accurate diagnostics of blood coagulation system state is of great importance. In this present study, we monitored several molecular markers of the dangers of intravascular thrombus formation and estimated the platelet function in pregnant women during gestation. We performed independent measurements using the same methodology for different cohorts of patients recruited in Kyiv (Ukraine) and in Szeged (Hungary). D-dimer and soluble fibrin were measured using ELISA. Protein C (PC) level was estimated using chromogenic substrate assay. Fibrinogen concentration was measured by spectrophotometry using thrombin-like enzyme. Platelet function was estimated by aggregometry. Statistical data analysis was performed using the Kruskal-Wallis test. Statistically significant increases of fibrinogen concentration from first to third gestational trimester was shown for both studied cohorts of patients (5-6 mg/ml at third trimester on average). Applied methods allowed us to detect the same tendencies of decreases in PC level as well as the appearance of moderate amounts of D-dimer (up to 300 ng/ml) and SF (up to 10-15 ug/ml). Platelet function was increased on the first trimester of pregnancy and decreased during following trimesters slightly. Results indicated the changes in the blood coagulation system of pregnant women during gestation with the same effectiveness independently of the selected cohorts, time and place of measurements. The application of the proposed diagnostics algorithm may allow estimating the risk of thrombotic complications during pregnancy.
Overall hemostasis potential of blood plasma and its connection to molecular markers of the hemostasis system in patients with stenosis of coronary artery
N. V. Storozhuk1, L. V. Pyrogova2, Т. М. Chernyshenko2,
O. P. Kostyuchenko2, T. M. Platonova2, O. B. Storozhuk1,
B. G. Storozhuk1, R. Yu. Marunich2,
G. K. Bereznytsky2, E. M. Makogonenko2*
1MI Pirogov Vinnytsia National Medical University, Vinnytsia, Ukraine;
2Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
*e-mail: ymakogonenko@gmail.com
Received: 01 April 2021; Accepted: 22 September 2021
The correlation relationships between hemostatic potential parameters and concentrations of molecular markers of the hemostasis system: soluble fibrin (sf), D-dimer (DD), fibrinogen (Fg) and protein C (PC) in patients with stenosis of coronary artery 6 months after coronary angioplasty have been investigated. It was found three directions of changes in the state of the patients hemostasis system: an increasing in fibrinolytic activity (C) ~18% of patients; an increasing in coagulative activity (B) ~31% of patients; and maintaining of the balance between coagulation and fibrinolysis (A) ~51% of patients. In patients with signs of stenosis without angina pectoris, a strong Pearson correlation was shown between the half-life of the clot and the overall hemostatic potential (OHP) (r = 0.75, P << 0.05), a moderate relationship between concentrations of sf and D-dimer (r = 0.67, P <0.05), almost complete connection between coagulation potential (CP) and OHP (r = 0.975, P << 0,05) and strong connection between CP and fibrinolytic potential (FP) (r = 0.80, P << 0.05). In patients with signs of stable angina pectoris, almost complete connection was found between the concentration of sf and D-dimer (r = 0.981, P << 0.05), CP and OHP (r = 0.979, P << 0.05) and a strong connection between CP and FP (r = 0.846, P << 0.05). Possible functional mechanisms of connection between these parameters are discussed.
Thromboelastographic study of fibrin clot and molecular basis of maximum clot firmness
D. S. Korolova1*, Y. M. Stohnii1, V. I. Gryshchuk1, S. I. Zhuk2,
I. V. Us2, T. M. Chernyshenko1, O. P. Kostiuchenko1, K. P. Klymenko1,
O. M. Platonov1,3, O. I. Ivashchenko3, V. O. Chernyshenko1
1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
2Shupyk National Medical Academy of Postgraduate Education, Kyiv, Ukraine;
3ESC “Institute of Biology and Medicine”, Taras Shevchenko National University of Kyiv, Ukraine;
e-mail: d.korolova@gmail.com
Received: 29 January 2021; Accepted: 23 April 2021
Maximum clot firmness (MCF) is the main parameter of thromboelastography (TEG) reflecting the stability of a clot. In this work, we looked for markers that can influence the enhancement of MCF and detected molecular markers and blood clotting parameters that can be involved in such mechanisms. Blood samples of pregnant women with placental disorders were collected in the Kyiv Perinatal Center. TEG was performed on whole blood in EXTEM and INTEM tests. APTT, INR, fibrinogen concentration and platelet aggregation were measured using traditional laboratory approaches. D-dimer was detected in sandwich ELISA using monoclonal antibodies III-3B and II-4D. The relative cross-linking activity of factor XIIIa was measured by the direct quantification of the cross-linked γ-chain of fibrin using Western-Blotting with monoclonal antibody II-4D. D-dimer and fibrinogen concentrations, clotting time in the APTT test, INR and rate of platelet aggregation did not correlate with the MCF. However, we found positive correlations of MCF with factor XIIIa activity: 0.51 and 0.87 for EXTEM and INTEM, respectively. These data indicate that for normal and slightly increased fibrinogen concentrations, fibrin clot firmness will depend mostly on the activity of factor XIIIa. Thus the direct determination of factor XIIIa activity in blood plasma of patients can be relevant for predicting the risk of intravascular coagulation. Evaluation of the content and activity of individual clotting factors or other components of the coagulation system can be useful additions to the TEG diagnostics and should not be neglected.
Men of the molecules
In memoriam of Prof. Russell Doolittle,
Prof. Eduard Lugovskoi and their friendship that outlive both of them
V. O. Chernyshenko
Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: bio.cherv@gmail.com
Received: 02 July 2020; Accepted: 21 July 2020
In memoriam Eduard Lugovskoi and Russell Doolittle we are referring to several episodes of their life and work. Russell Doolittle an American biochemist and his friend and colleague Ukrainian scientist Eduard Lugovskoi, both studied fibrinogen structure and functions and finally united their efforts in the revealing of the new mechanism of intramolecular interactions of fibrin molecule through coiled-coil region. The results of their common work and discussions were included to the article “The fibrin Bβ125-135 site is involved in the lateral association of protofibrils”. Valuable part of the communication dedicated to the poetry of Eduard Lugovskoi that inspired both of scientists in work and life. We are providing some remembrance of their collaboration, their letters sent to each other, fragments of handwriting and common photo of Russell Doolittle and Eduard Lugovskoi.
Novel monoclonal antibody to fibrin(ogen) αC-region for detection of the earliest forms of soluble fibrin
N. E. Lugovska1, I. M. Kolesnikova1, Ye. M. Stohnii1, V. O. Chernyshenko1*,
A. V. Rebriev1, O. P. Kostiuchenko1, G .K. Gogolinska1, N. A. Dziubliuk2,
L. D. Varbanets2, T. M. Platonova1, S. V. Komisarenko1
1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
2Zabolotny Institute of Microbiology and Virology,National Academy of Sciences of Ukraine, Kyiv;
*e-mail: bio.cherv@gmail.com
Received: 08 May 2020; Accepted: 30 June 2020
Obtaining new monoclonal antibodies (mAbs) towards fibrin(ogen) and its fragments is an important task for studying mechanisms of blood clot formation, searching for novel antithrombotic agents and developing immunodiagnostics. The aim of the present work was to create and characterize a new mAb towards the fibrin(ogen) αС-region. We surmise that having a specific mAb towards this flexible part of the molecule will allow us to study the role of the αС-region in fibrin polymerization and also to develop an approach for detecting the earliest forms of soluble fibrin by sandwich ELISA. Using hybridoma technology we оbtained mAb 1-5A to the αC-region of fibrinogen.. It was characterized using several variations of ELISA and Western blot. Application of specific proteases together with MALDI-TOF analysis allowed us to localize its epitope that is located in fragment 537-595 of the Aα-chain of fibrin(ogen). МAb 1-5A can be used as a detecting tag-antibody in sandwich ELISA for the quantification of the earliest forms of soluble fibrin which are uncleaved by plasmin and preserved C-terminal portions of αC-regions. These earliest forms of soluble fibrin are direct evidence of blood coagulation system activation, thrombin generation and the danger of intravascular thrombus formation. Their determination will provide additional, more accurate information about the state of the blood coagulation system and the risk of blood clotting, which is very important for the timely and correct selection of adequate antithrombotic therapy. MAb 1-5A effectively binds the αC-containing molecules of fibrinogen and fibrin in blood plasma. It also can be used for studying protein-protein and protein-cellular interactions of the αC-regions of fibrin(ogen).
Identification of the binding site for plasminogen kringle 5 in the α-chain of fibrin(ogen) D-fragment
L. G. Kapustianenko*, T. V. Grinenko, A. V. Rebriev,
O. I. Yusova, A. A. Tykhomyrov
Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
*e-mail: kapustyanenko@biochem.kiev.ua
Received: 17 May 2020; Accepted: 30 June 2020
The interaction of the fifth kringle of Glu-plasminogen with fibrin triggers activation and initiation of fibrinolysis, yet the site on fibrin that binds kringle 5 remains unknown. The aim of our work was to determine an amino acid sequence in the D-fragment of fibrin(ogen) molecule, which is complementary to the lysine-binding site (LBS) in kringle 5. We studied the interaction between kringle 5 of plasminogen with polypeptide chains of the D-fragments of fibrin and cyanogen bromide fragments FCB-2 and t-NDSK and showed that kringle 5 bound specifically to α- and γ-chains of the D-fragment and the α-chain of FCB-2. Tryptic peptides of D-fragment α-chain were obtained, separated by their ability to bind with the immobilized kringle 5, and then all studied peptides were characterized by MALDI-TOF analysis. The critical amino acid residues of the α-chain of D-fragment, which provide its interaction with kringle 5, turned out to be α171Arg and/or α176Lys. The binding site of Glu-plasminogen complementary to the LBS of kringle 5 is located within Аα168Ala−183Lys, a sequence in a weakly structured loop between two supercoils in the α-chain of the D-fragment of the fibrin(ogen) molecule.