Endoplasmic reticulum stress, as well as hypoxia and ischemia, are important factors for tumor neovascularization and growth. Cancer cells preferentially utilize glycolysis in order to satisfy their increased energetic and biosynthetic requirements. High glucose metabolism of cancer cells is caused by a combination of hypoxia-responsive transcription factors, activation of oncogenic proteins and the loss of tumor suppressor function and is realized in part by activating a family of regulatory bifunctional 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatases (PFKFB) and hexokinase 2. We have studied the effect of hypoxia and ischemia on the expression of PFKFB and hexokinase genes in glioma cell line U87 under knockdown of endoplasmic reticulum-nuclei-1 (ERN1) sensing and signaling enzyme. It was shown that loss of the signaling enzyme ERN1 function leads to an increase in the expression levels of HK1, HK2, PFKFB3 and PFKFB4 mRNA. Moreover, the expression levels of all studied genes increase under hypoxia in control and ERN1-deficient glioma cells; however knockdown of ERN1 suppresses the effect of hypoxia. Besides, HK2 and PFKFB4 are more sensitive to hypoxia than HK1 and PFKFB3. Glucose or glutamine deprivation conditions have different effects on the expression levels of these genes and its effect depends mainly on ERN1 function. Expression levels of alternative splice variants of PFKFB3 and PFKFB4 mRNA change at used experimental conditions in a fashion similar to the basic PFKFB variants. Thus, the expression of hexokinase and PFKFB genes is mainly dependent on ERN1 signaling enzyme function in normal, hypoxic and ischemic conditions.

The fibrinogen molecule contains multiple binding motifs for different types of cellular receptors, acting as a molecular link between coagulation and cell adhesion. In this study we generated a truncated form of the fibrinogen molecule lacking the Bβ1-42 sequence by site-specific proteolysis and evaluated the role of the fragment in adhesive capabilities of platelets, endothelial and cancer cells. Fibrinogen with the removed Bβ1-42 sequence and fibrin without the Bβ15-42 fragment (desβ1-42 fibrinogen and desABβ15-42 fibrin) were obtained by proteolysis using the specific protease from the venom of Echis multisquamatis. The cleaved fragment was purified by HPLC and was identified using MALDI-TOF. ADP-and collagen-induced aggregation of washed platelets in the presence of fibrinogen desBβ1-42 was studied using an aggregometer. Proliferation of mice aortic endothelial cells (MAEC) and human umbilical vein endothelial cells (HUVEC) was studied using the fibrin desABβ15-42 as the scaffold. Cell viability was quantified by the MTT test (MAEC). Generation time was calculated for the estimation of proliferative activity of HUVEC. Lung cancer cell line Н1299 was used to evaluate cancer cell motility in vitro using the scratch assay. Direct comparison of cellular behavior in the presence of truncated vs native forms demonstrated attenuated cell adhesion in the presence of fibrinogen desBβ1-42 and fibrin desBβ15-42. The platelet aggregation rate was only slightly decreased in the presence of fibrinogen desBβ1-42 but resulted in 15-20% disaggregation of adhered platelets. We also observed the substantial decrease of generation time of HUVEC and inhibition of viability of MAEC cells grown on scaffolds of a desABβ15-42 matrix. Finally, desBβ1-42 modulated the motility of H1299 cells in vitro and suppressed the wound healing by 20% compared to the full-length fibrinogen. We postulate that fragment 1-42 of the BβN-domain of fibrinogen is not sufficient for platelet aggregation, however it may contribute to platelet clot formation in later stages. at the same time, this fragment may be important for establishing proper cell-to-cell contacts and cell viability of endothelial cells. Also, 1-42 amino acid fragment of the BβN-domain supported the migration of cancer cells suggesting that interactions of fibrinogen with cancer cells could be a target for anticancer therapy. The Bβ1-42 fragment of fibrinogen contributes to efficient intracellular interactions of different types of cells, including platelets, endothelial cells and cancer cells. K e y w o r d s: fibrinogen, adhesion, cell migration, endothelium, cell proliferation, platelets.

Background: The plant Viscum album (Mistletoe) is a known source of biologically active substances, used in traditional medicine in Europe and Asia.Aim: The goal was to study cytotoxic/cytostatic effect of mistletoe extract on tumour and normal cells and its influences on certain proteins of the blood coagulation system.Setting: Mistletoe was collected in Ukraine in November 2020.Methods: Water extract of V. album, both leaves and stems, was obtained and fractionated; it was characterised using high-performance liquid chromatography (HPLC) and mass spectrometry and tested on cancer cells and blood plasma.Results: Extract demonstrated the presence of viscotoxins and carbohydrates and thermolabile compounds that enhance the activity of thrombin and factor X in plasma in the presence of calcium ions and increased the activated partial thromboplastin time (APTT) by 2.7 times. The cytotoxic/cytostatic action of the mistletoe fractions, the total extract and the fraction 10 000 Da were higher in relation to Lewis lung carcinoma (LLC) cells than with Vero cells. The IC50 of total extract for LLC cells was 35% (p 0.05) lower than that for Vero cells (1.12 ± 0.2 and 1.71 ± 0.15 mg/ml). Thus, the IC50 of the fraction 10 000 Da for LLC cells was more than by 44% (p 0.05) lower than that for Vero cells (2.07 ± 0.26 and 3.67 ± 0.41 mg/ml).Conclusion: It was shown that both extracts exhibited a cytotoxic/cytostatic effect, more pronounced against tumour cells than normal cells and enhanced the clotting of blood plasma by thermolabile compounds by facilitating plasma coagulation.Contribution: This research makes it possible to study mistletoe as a light and cheap anticancer therapy plant and cure blood coagulopathy or construct antibleeding bandages.

Chelidonium majus is a well-known source of biologically active compounds. Most of them are alkaloids, which are used in researches and for tradition medicine. In this study, we explored the influence of C. majus crude total extract onto blood coagulation system in vitro, primary and secondary hemostasis. Aim. To study influence of C. majus extract onto blood coagulation process. Methods. Turbidimetry of blood plasma, APTT test with chromogenic substrates, and platelet aggregation were used in the work. Results. We demonstrated moderate stimulating effect of the extract on platelets (the rate of platelet aggregation increased up to 10%, followed by disaggregation). Extract also increased the rate of platelet aggregation stimulated by 12.5 mcM of ADP. We observed the increase in the plasma clotting time in the presence of the extract, that corresponded to the 274, 411, 685, 1370 mcg of dry extract, from 70 s in control to 80, 90, 170 and 180 s, respectively, in turbidimetry test of plasma stimulated by APTT-reagent. However, APTT test with the chromogenic substrate of thrombin (S2238) did not show significant influence of this plant’s extract on thrombin activity. Conclusions. Thus, we can conclude that anticoagulant activity of Chelidonium majus extract corresponded to the direct inhibition of fibrin polymerization.

Гіпоксія є одним із потужних індукторів експресії великої групи генів, у тому числі генів, які контролюють гліколіз, ангіогенез і процеси проліферації, що сприяє виживанню клітин за умов зниженого рівня кисню. Але гіпоксія, що має місце у клітинах злоякісних пухлин, відбувається за умов нормального парціального тис-ку кисню внаслідок зниженого його споживання. Більше того, гіпоксія є обов'язковим компонентом процесів злоякісного росту і значною мірою контролює процеси гліко-лізу, ангіогенезу та проліферації. Аналізуються дані про молекулярні механізми індукції залежного від гіпоксії транскрипційного фактора HIF у клітинах за гіпоксії і у злоякісних пухлинах, а також його роль у регуляції експресії генів, механізми взаємодії цього транскрипційного фактора зі специфічними регуляторними послі-довностями промоторів генів, що активуються за гіпоксії.Ключові слова: гіпоксія, молекулярні механізми регуляції, HIF, експресія ге-нів, злоякісні пухлини. ВСТУПВідомо, що гіпоксія істотно змінює характер метаболічних процесів у організмі, значно посилює інтенсивність гліколізу, що сприяє забезпеченню клітин організму енергією в умовах зниженого рівня кисню або недостатнього його використання, а також є надзвичайно важливим фактором росту і прогресії злоякісних пухлин. У даній роботі проаналізовані й узагальнені наявні в літературі дані про молеку-лярні механізми регуляції експресії генів у клітинах за гіпоксії та у злоякісних пух-линах, про механізми індукції процесів проліферації, ангіогенезу та гліколізу, а та-кож про роль ключових ензимів і факторів, зокрема роль HIF, транскрипційного фактора, що індукується гіпоксією, у регуляції цих процесів і в механізмах адапта-ції клітин до умов гіпоксії, як у нормі, так і за різноманітних патологічних станів.