Category Archives: Uncategorized
Fibrinolysis regulation by platelets retaining plasminogen and tissue-type plasminogen activator on their surface
T. Grinenko, О. Yusova, O. Revka, I. Patalakh, T. Yatsenko
Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: sedrickedel@gmail.com
Received: 22 July 2019; Accepted: 18 October 2019
Platelets play a key role in hemostasis as cofactors of thrombin generation, fibrin polymerization centers, and initiators of clot retraction, while their ability to modulate clot dissolution remains less understood. The aim of this study was to investigate the interaction of plasminogen and tissue plasminogen activator with native and activated platelets, to determine the amount of plasmin generated by various activators in the presence of platelets, and the ability of platelets to modulate the rate of polymer fibrin hydrolysis. Spectrometric and immunofluorometric methods were used in the study. It was shown that intact circulating platelets carry a small amount of plasminogen on their surface, whereas thrombin-induced activation led to the exposure of plasminogen-binding sites on their plasma membrane. Activated platelets stimulated plasminogen activation by tissue plasminogen activator, urokinase, and streptokinase. Components of prothrombin complex enhanced plasminogen activation by tissue plasminogen activator on the surface of activated platelets. Model system with desAB-fibrin revealed the ability of platelets to stimulate fibrinolysis. These results suggest that the regulation of fibrinolysis by platelets is provided by the binding of plasminogen and plasminogen activators on their surface, acceleration of plasmin generation and, consequently, acceleration of the onset of fibrin lysis and reducing of the clot lifetime, which is important to maintain hemostatic balance.
Comparative analysis of the influence of chlorine and fluorine anions on the fibrin polymerization
L. V. Pyrogova1, G. K. Bereznitsky1, G. K. Gogolinskaya1, T. M. Platonova1,
I. M. Kolesnikova1, O. O. Masenko1, R. Yu. Marunich1, P. Yu. Tsap1,
Yu. V. Ushenin2, Y. M. Makogonenko1, E. V. Lugovskoi1
1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
2V.E. Lashkaryov Institute of Semiconductor Physics, National Academy of Sciences of Ukraine, Kyiv;
e-mail: ymakogonenko@gmail.com
Received: 25 April 2019; Accepted: 18 October 2019
The effect of NaCl and NaF salts in the range of 0.1-0.225 M concentrations on individual stages of fibrin polymerization was investigated, namely: the rate of fibrinogen activation by thrombin, the rate of protofibrils formation, the rate of lateral association of protofibrils, and the maximum clot absorbance value at 350 nm. It was found that the chlorine and fluorine anions equally inhibit the rate of formation of fibrin from fibrinogen and the formation of protofibrils activated with thrombin. Chlorine anions were shown to be significantly more effective than fluorine anions, inhibiting the rate of lateral association and maximal clot absorbance level from fibrin desA and desAB. A component of the inhibitory action of chlorine anions, not related to the ionic strength of the solution, was identified and its effect on the individual polymerization steps was shown. Chlorine anions were found to bind to a fibrin clot. Using the surface plasmon resonance method and fibrin-specific mAb FnI-3c, it was established that the rate of exposure of neoantigenic determinants of mAb in the hinge regions of the fibrinogen molecule during its transformation into fibrin under the action of thrombin is inhibited by chlorine anions in correlation with inhibition of the protofibril lateral association rate. It has been suggested that the inhibitory effect of chlorine anions consists of an ionic component and a component that blocks the conformational mobility of the molecule by the chlorine anions binding to its hinge regions and polymerization sites.
Analysis of Aurora kinases genes expression points on their distinct roles in prostate cancer development
O. Mankovska1, G. Gerashchenko1, E. Rozenberg1, E. Stakhovsky2,
O. Kononenko2, Yu. Bondarenko3, V. Kashuba1,4
1Institute of Molecular Biology and Genetics, National Academy of Sciences of Ukraine, Kyiv;
2National Cancer Institute, Ministry of Health of Ukraine, Kyiv;
3Institute of Urology National Academy of Medical Sciences of Ukraine, Kyiv;
4Department of Microbiology, Tumor and Cell Biology (MTC), Karolinska Institutet, Stockholm, Sweden;
e-mail: mankovsska@gmail.com
Received: 28 May 2019; Accepted: 18 October 2019
Aurora kinases A and B play a crucial role in the regulation of mitosis, while Aurora C controls meiotic division. These proteins showed controversial behavior upon the development of epithelial tumors. Our aim was to examine if there are any differences in expression of Aurora kinases genes in malignant and non-malignant tumors and non-tumor tissues; to compare their expression with clinical characteristics of patients and expression of other prostate cancer-associated genes. Quantitative RT-PCR was used to determine Aurora A-C genes expression in 33 prostate adenocarcinomas (T), paired conventionally normal tissues (N), and 17 adenomas (A). Relative expression values (RE) for genes studied were estimated using 2-ΔCt and 2-ΔΔCt method. The Kruskal-Wallis with correction on multiple comparisons, according to the Benjamini-Hochberg procedure with FDR = 0.2, Dunn-Bonferroni post hoc test and Spearman rank correlation analysis were used for statistical analysis. As turned out, RE values for AURKA were found to be significantly lower in samples of T group in comparison with A group. Moreover, significant up-regulation of AURKC expression levels were detected for T 3-4 stage compared to T 1-2 stage. RE values of AURKC in T group were positively correlated with the tumor stage, while AURKB RE demonstrated a negative correlation with the tumor stage. We also found significant correlations between AURK genes expression levels and prostate cancer-associated genes in T group. We suppose that all these data point to probable involvement of Aurora kinases’ genes in prostate carcinogenesis.
Dependence of the mitochondrial adaptive capacity of hepatocytes on the oxidative substrates availability
H. M. Mazur, V. M. Merlavsky, B. O. Manko, V. V. Manko
Ivan Franko National University of Lviv, Ukraine;
e-mail: Volodymyr.Manko@lnu.edu.ua
Received: 11 February 2019; Accepted: 18 October 2019
The ability of the mitochondria to compensate for energy expenditure of cells largely depends on the availability of the oxidative substrates, transported across the intact plasma membrane with molecular carriers of limited affinity. The aim of this study was to investigate the dependence of adaptive respiratory responses of mitochondria of intact hepatocytes on the oxidative substrates. Basal and FCCP-stimulated respiration rates were determined with Clark electrode. After 15-minute incubation in the medium with the oxidative substrates or their combinations (glutamine, pyruvate, succinate, monomethyl succinate, α-ketoglutarate, dimethyl-α-ketoglutarate (2 mM) or glucose (10 mM)), isolated hepatocytes were added into the respiratory chamber. FCCP concentration was 0.25, 0.5 and 1 μM. The adaptive capacity of mitochondria was characterized by the maximal uncoupled respiration rate (the highest respiration rate among all tested FCCP concentrations), the optimal FCCP concentration (the concentration at which the maximal rate is achieved) and the area under the curve (AUC) of the dependence of the uncoupled respiration rate on FCCP concentration. The adaptive capacity of mitochondria, evaluated by AUC, increases in this order of substrates: glucose (0.063 r.u.), endogenous substrates (0.067 r.u.), glutamine (0.092 r.u.), pyruvate (0.113 r.u.), α-ketoglutarate (0.113 r.u.), succinate (0.152 r.u.), dimethyl-α -ketoglutarate (0.156 r.u.), and monomethyl succinate (0.172 r.u.). The adaptive capacity of mitochondria of hepatocytes seems to be partly dependent on plasma membrane transporters affinities (Km) to the oxidative substrates. The presence of glucose in the medium does not improve the adaptive capacity of hepatic mitochondria.
The effect of quercetin on oxidative stress markers and mitochondrial permeability transition in the heart of rats with type 2 diabetes
N. I. Gorbenko1, O. Yu. Borikov2, O. V. Ivanova1, E. V. Taran1,
Т. S. Litvinova1, T. V. Kiprych1, A. S. Shalamai3
1V. Danilevsky Institute of Endocrine Pathology Problems, National Academy of Medical Sciences of Ukraine, Kharkiv;
2V. N. Karazin Kharkiv National University, Ukraine;
3PJSC SIC “Borshchahivskiy Chemical-Pharmaceutical Plant”, Kyiv, Ukraine;
е-mail: Gorbenkonat58@ukr.net
Received: 24 June 2019; Accepted: 13 August 2019
Increasing evidence suggests that oxidative stress and induction of mitochondrial permeability transition in cardiomyocytes are linked to tissue damage and the development of diabetic cardiovascular complications. The aim of this study was to assess the effects of quercetin (Q) on oxidative stress and mitochondrial permeability transition in the heart of rats with type 2 diabetes mellitus (DM). Type 2 DM was induced in 12-week-old male Wistar rats by intraperitoneal injections of 25 mg/kg streptozotocin twice per week followed by a high-fat diet during four weeks. The rats were divided into three groups: control intact group (C, n = 8), untreated diabetic group (Diabetes, n = 8) and diabetic rats treated with Q (50 mg/kg/day per os for 8 weeks) after diabetes induction (Diabetes+Q, n = 8). Administration of Q increased insulin sensitivity and normalized the functional state of cardiac mitochondria due to increased aconitase and succinate dehydrogenase activities in rats with type 2 DM. Q also ameliorated oxidative stress, decreasing the level of advanced oxidation protein products and increasing the activity of thioredoxin-reductase in heart mitochondria of diabetic rats. In addition, Ca2+-induced opening of the mitochondrial permeability transition pore was significantly inhibited in diabetic rats treated with Q in comparison with the untreated diabetic group. These data demonstrate that Q can protect against oxidative stress, mitochondrial permeability transition induction and mitochondrial dysfunction in cardiomyocytes of diabetic rats. We suggest that the use of Q may contribute to the amelioration of cardiovascular risk in type 2 DM.
Brownian motion, electrophoresis, chromatography, and macromolecular chemistry: how it all unites Nobel laureates of the first half of the 20th century – T. Svedberg, A. Tiselius, R. Synge and H. Staudinger
M. V. Grigorieva, V. M. Danilova, S. V. Komisarenko
Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kуiv;
e-mail: mvgrigorieva@biochem.kiev.ua
Received: 31 July 2019; Accepted: 13 August 2019
It is hard to imagine how chemistry, biology and medicine would develop without such techniques as ultracentrifugation, electrophoresis and chromatography. At present innovative hi-tech laboratory ultracentrifuges are widely used in various fields of fundamental science and practice, including colloid chemistry, biochemical analysis, virology, clinical diagnostics, pharmacy, nanotechnology, to name a few. Electrophoresis enables to detect protein abnormalities with high probability, and, therefore, has wide application for the diagnosis of infectious-inflammatory diseases, genetic and immune disorders, malignant tumors and others. Chromatography is widely used for biochemical research and analytical detection and control of drugs and food. But where did these methods, which have had a significant impact on the development of various fields of scientific and economic activity, come from? Who were the pioneers in this field and whose work influenced the formation of the next generation of researchers? These are the questions we address in this article.
Alpha-fetoprotein as a biochemical diagnostic and prognostic marker for prolonged jaundice in newborns
O. H. Mazur, O. S. Yablon, O. S. Rubina,
M. M. Puhach, A. P. Konoplitska
National Pirogov Memorial Medical University, Vinnytsia, Ukraine;
e-mail: alena523@ukr.net
Received: 14 January 2019; Accepted: 13 August 2019
Prolonged jaundice of newborns is a common pathology during the neonatal period. Recently, there has been a tendency toward an increased number of newborns with prolonged jaundice with duration longer than 14 days of life. According to the polyetiologic nature of neonatal jaundice, it is necessary to investigate new diagnostic signs that can predict the development of prolonged jaundice in newborns and, allow identification of new methods of differential diagnosis of neonatal jaundice, as well as decrease the frequency of this pathology. The parameters of serum alpha-fetoprotein (AFP) content in newborns with prolonged jaundice were studied. The content of AFP in blood serum was 1.7 times higher in newborns with prolonged jaundice than in newborns with jaundice for up to 14 days of life: Me = 671.1 [Q1 = 479.9; Q3 = 815.03] ng/ml and Me = 401.0 [Q1 = 284.9; Q3 = 684.0] ng/ml, respectively (P < 0.05). The content of blood serum AFP in newborns with prolonged jaundice was higher for total serum bilirubin greater than 250 μmole/l: Me = 626.2 [Q1 = 454.7; Q3 = 793.2] ng/ml which was confirmed by a strong direct correlation relationship (rxy = 0.64, P < 0.05). The results of the ROC analysis confirmed that AFP ≥ 571.7 ng/ml can identify newborns with prolonged jaundice with a sensitivity of 74.2% and a specificity of 74.5%. The area under the curve (AUC) was 0.870 (95% CI 0.804, 0.937), which confirms the good quality of the AFP model ≥ 571.7 ng/ml.
The role of resistin in the genesis of metabolic disorders in pathological pregnancy
S. O. Ostafiichuk
Ivano-Frankivsk National Medical University, Ukraine;
e-mail: svitlana.ostafijchuk@gmail.com
Received: 12 March 2019; Accepted: 13 August 2019
Pathological gestational weight gain (GWG) is a risk factor for obstetric and perinatal complications. High metabolic activity of adipose tissue and the placenta during pregnancy manifests as an increased production of adipokines that are involved in glucose regulation and insulin sensitivity. The aim of this study was to determine the role of resistin in the genesis of metabolic disorders in pathological GWG pregnancies. The 163 pregnant women were examined in the study: 97 (59.5%) had normal, 18 (11.0%) had insufficient and 48 (29.4%) had excessive prepregnancy weight and obesity. GWG was the recommended level in 56 (34.4%), insufficient in 33 (20.2%), and excessive in 74 (45.4%) women. Anthropometry was performed in each trimester of pregnancy, the weight gain was measured, and the percentage of body fat mass, concentrations of resistin, glucose, insulin, and the HOMA-IR were evaluated. Positive associations were found between hyperresistinemia in the second trimester of pregnancy, and subsequent weight gain (r = 0.27, P = 0.0006), percentage of body fat mass (r = 0.93, P = 0.000) and insulin resistance (r = 0.89, P = 0.000) in late pregnancy; these associations were especially evident in excessive GWG. Determination of predictors of insulin resistance, associated with endocrine activity of adipose tissue, such as the adipokine resistin, in the second trimester of pregnancy may help to predict the severity of metabolic shifts during pregnancy and the risk of developing obstetric and perinatal complications.
Citicoline affects serum angiostatin and neurospecific protein levels in patients with atrial fibrillation and ischemic stroke
A. A. Tykhomyrov1, Yu. S. Kushnir2, V. S. Nedzvetsky3,
T. V. Grinenko1, O. V. Kuryata2
1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
2State Establishment “Dnipropetrovsk Medical Academy of Health Ministry of Ukraine”, Dnipro;
3Bingöl University, Bingöl, Turkey;
e-mail: artem_tykhomyrov@ukr.net
Received: 22 May 2019; Accepted: 13 August 2019
Ischemic stroke is considered as one of the most frequent and severe complications of atrial fibrillation. The present study was undertaken to examine whether post-insult treatment with cytidine diphosphate-choline (CDP-choline, or citicoline) affects serum levels of the angiogenesis inhibitor angiostatin and neurospecific proteins as markers of brain damage in patients with cerebral ischemia associated with atrial fibrillation. Thirty-three patients with a diagnosis of acute ischemic stroke received citicoline sodium by intravenous infusions (1,000 mg daily for 14 days) in addition to the standard treatment (basic group). Twenty-five patients with the same pathologies, who received only standard therapy, were enrolled in the study as a control group. Serum content of angiostatin and neurospecific proteins, namely neurofilament heavy subunit (NF-H) and glial fibrillary acidic protein (GFAP), was measured by immunoblotting at the basal level and after the treatment. Citicoline treatment caused significant decreases in serum levels of angiostatin (by 40% vs. basal level, P < 0.05), GFAP (by 61%, P < 0.01), and the NF-H subunit (by 19%, P < 0.05) and had no effect on the serum albumin content. In contrast, there were no statistically significant differences between baseline levels of the studied protein markers and their content after the treatment period in the control group. These findings indicate for the first time that CDP-choline protects both astrocytes and neurons and improves angiogenic capacity through down-regulation of angiostatin in post-ischemic patients with atrial fibrillation after acute ischemic stroke. Further studies are needed to test associations between serum levels of these biomarkers, clinical outcomes, and treatment efficacy of stroke.
Influence of human HB-EGF secreted form on cells with different EGFR and ErbB4 quantity
O. I. Krynina, N. V. Korotkevych, A. J. Labyntsev,
S. I. Romaniuk, D. V. Kolybo, S. V. Komisarenko
Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: olyakrynina@gmail.com
Received: 18 July 2019; Accepted: 13 August 2019
HB-EGF is one of the most potent ligands of EGFR and ErbB4 receptors. This growth factor plays a pivotal role in many cellular processes, but its effect differs from one cell type to another and remains not fully understood. The aim of this work was to investigate the dependence between the rate of HB-EGF mediated cell proliferation and activation of EGFR and ErbB4 receptors. Therefore, the effects of human recombinant sHB-EGF (rsHB-EGF) on the proliferation of cell lines with different EGFR and ErbB4 quantity and ratio, as well as activation of the MARK-cascade p38 and ERK1/2 (p42/44) kinases, were analyzed. For comparison, a similar study of the effect of native sHB-EGF secreted by human histiocytic lymphoma cells U937 during co-cultivation with different cell lines was performed.
It was proved that cell proliferation in response to sHB-EGF depends not only on the quantity but also on the ratio of EGFR and ErbB4. It was shown that signaling through ErbB4 is associated with activation of p38 kinase and signaling through EGFR associated with activation of ERK1/2 (p42/44) kinase. We assume the existence of two different mechanisms for sHB-EGF-mediated stimulation of cell proliferation, and the simultaneous launch of these mechanisms provides a maximal proliferative response. The results of this study support the feasibility of creating anti-proliferative drugs that target ErbB4.