Tag Archives: apoptosis

Action of free and polymer carrier encapsulated doxorubicin towards HCT116 cells of human colorectal carcinoma

Yu. V. Senkiv1,2,4, P. Heffeter2, A. O. Riabtseva3, N. M. Boiko1,
O. S. Zaichenko3, N. Ye. Mitina3, W. Berger2, R. S. Stoika1

1Institute of Cell Biology, National Academy of Sciences of Ukraine, Lviv;
2Institute for Cancer Research, Medical University of Vienna, Austria;
3National University Lviv Polytechnic, Ukraine;
4Ivan Franko Lviv National University, Ukraine;
e-mail: stoika@cellbiol.lviv.ua

Development of novel nanoscale functionali­zed carriers is nowadays one of the most urgent problems in cancer treatment.  The aim of our study was to compare the antineoplastic effect of free doxorubicin and its complex with a nanoscale polymeric carrier towards HTC116 colorectal carcinoma cells. It was established that application of the complex of poly(5-tret-butylperoxy)-5-methyl-1-hexene-3-in-co-glycydyl metacrylat)-graft-polyethyleneglycol (poly(VEP-GMA-PEG)-graft-PEG), where VEP  – 5-tret-butylperoxy)-5-methyl-1-hexene-3-in; GMA – glycydyl metacrylat; graft-PEG – graft-polyethyleneglycol accordingly, functionali­zed with phosphatidylcholine for doxorubicin delivery increased 10 times the efficiency of cytotoxic action of this drug, as compared wich such efficiency in case of the action of free doxorubicin. The encapsulated form of doxorubicin caused more intensive cleavage of the reparation enzyme PARP and longer delay in G2/M cell cycle arrest, compared to such effects of free doxorubicin. The developed carrier itself is non-toxic to the used mammalian cells and does not cause impairment in their cell cycle. A deletion in both alleles of p53 gene did not affect the antineoplastic action of doxorubicin that was immobilized on the nanoscale carrier. Thus, p53-dependent signaling pathways are not involved in the cytotoxic action of doxorubicin-carrier complex. It is suggested that novel nanoscale polymeric carrier poly(VEP-GMA-PEG)-graft-PEG functionalized with phosphatidylcholine could be a promising carrier for targeted delivery of anticancer drugs.

Biochemical effects of combined action of γ-irradiation and paclitaxel on anaplastic thyroid cancer cells

V. M. Pushkarev, O. I. Kovzun, V. V. Pushkarev, M. D. Tronko

State Institution V. P. Komisarenko Institute of Endocrinology and Metabolism,
National Academy of Medical Sciences of Ukraine, Kyiv;
e-mail: pushkarev.vm@gmail.com

The aim of the paper was to describe the biochemical effects of Paclitaxel (Ptx), γ-irradiation (IR) and their combination in undifferentiated thyroid cancer cells (ATC). IR activated common DNA damage-induced signaling and manifested certain mitogenic effect by inactivation of retinoblastoma protein (pRb). There was clear antagonism between Ptx and IR relative to cell cycle regulators – tumor suppressor p53, pRb, CHK2 and c-Abl as well as proapoptotic Bax expression, but combined action of both agents enhanced caspase-3 and, especially, caspase-8 activation. The Ptx at low (1-25 nM) concentrations caused noticeable radioprotective effect.
Thus, in ATC cells the ionizing radiation and Ptx exhibited competitive effects upon phosphorylation of cell cycle controllers: p53, pRb, CHK2, cAbl and expression of Вах. At the same time, the combined effect of radiation and Ptx enhanced antiapoptotic Bcl-2 phosphorylation, caspases activation and survivin expression. The net effect of these events during the first 48-72 h of cells incubation can be considered as antiapoptotic – Ptx attenuated cytotoxic effect of IR.

Inhibitor of the transcription factor NF-κB, DHMEQ, enhances the effect of paclitaxel on cells of anaplastic thyroid carcinoma in vitro and in vivo

V. V. Pushkarev1, D. V. Starenki2, V. M. Pushkarev1,
O. I. Kovzun1, M. D. Tronko1

1State Institution V. P. Komisarenko Institute of Endocrinology and Metabolism,
National Academy of Medical Sciences of Ukraine, Kyiv;
e-mail: pushkarev.vm@gmail.com;
2Department of Biochemistry, Medical College of Wisconsin, USA;
e-mail: dstarenki@mcw.edu

Anticancer drug paclitaxel (Ptx) effect on biochemical mechanisms, regulating apoptosis in anaplastic thyroid carcinoma cells, was studied. It was shown that in addition to apoptotic cell death, Ptx induces signa­ling cascades that ensure cell survival. Paclitaxel-induced activation of nuclear factor kappa B (NF‑κВ) leads to an increase of some antiapoptotic proteins expression such as survivin, cIAP, XIAP. A novel NF‑κВ inhibitor, dehydroxymethylepoxyquinomicin (DHMEQ), was found to enhance cytotoxic effect of Ptx in anaplastic thyroid carcinoma cells. An enhancement of caspase-3 and -9 activation and PARP cleavage as well as the decreased levels of proteins-inhibitors of apoptosis were observed when cells were treated with a combination of both drugs. Mitochondria transmembrane potential (ΔΨm) loss was observed at higher concentrations of Ptx and DHMEQ. NF-κВ inhibition also potentiates paclitaxel effect at tumors formed by xenotransplantation of FRO cells into mice. Tumor mass reduction, significantly different from the effects of each of the compounds alone, was observed in animals, treated with paclitaxel and  NF-κВ inhibitor. Thus, the combined use of paclitaxel and NF-κВ inhibitor inhibits biochemical processes that contribute to the resistance of anaplastic thyroid carcinoma cells to paclitaxel action.

Multiple molecular forms of adaptor protein Ruk/CIN85 specifically associate with different subcellular compartments in human breast adenocarcinoma MCF-7 cells

B. O. Vynnytska-Myronovska1, Ya. P. Bobak1, G. V. Pasichnyk2,
N. I. Igumentseva1, A. A. Samoylenko2, L. B. Drobot2

1Institute of Cell Biology, National Academy of Sciences of Ukraine, Lviv;
2Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: drobot@biochem.kiev.ua

Ruk/CIN85 is a receptor-proximal ‘signalling’ adaptor that possesses three SH3 domains, Pro- and Ser-rich regions and C-terminal coiled-coil domain. It employs distinct domains and motifs to act as a transducer platform in intracellular signalling. Based on cDNA analysis, various isoforms of Ruk/CIN85 with different combination of protein-protein interaction domains as well as additional Ruk/CIN85 forms that are the products of post-translational modifications have been demonstrated. Nevertheless, there is no precise information regarding both the subcellular distribution and the role of Ruk/CIN85 multiple molecular forms in cellular responses. Using MCF-7 human breast adenocarcinoma cells and cell fractionation technique, specific association of Ruk/CIN85 molecular forms with different subcellular compartments was demonstrated. Induction of apoptosis of MCF-7 cells by doxorubicin treatment or by serum deprivation resulted in the system changes of Ruk/CIN85 molecular forms intracellular localization as well as their ratio. The data obtained provide a new insight into potential physiological significance of Ruk/CIN85 molecular forms in the regulation of various cellular functions.

Proapoptotic properties of total phytohemagglutinine and its individual isolectins in human cell culture 4BL

T. O. Kochubei, O. V. Maksymchuk, L. L. Маcewicz, О. O. Piven, L. L. Lukash

Institute of Molecular Biology and Genetics, National Аcademy of Sciences of Ukraine, Кyiv;
е-mail: kochubei2009@ukr.net

Phytohemagglutinine (PHA) is widely investigated lectin with mitogenic properties. Recently it was shown that  PHA is not only cell proliferation inducer, but also has a toxic or cytostatic effect. However concentration dependence and molecular mecha­nisms of this effect are not enough investigated­. To study proapoptotic properties of total phytohemagglutinine and its isolectins in human cell culture of not tumor origin 4BL we observed a change in the frequency of apoptotic cells in the tested cell culture under the influence of the total phytohemagglutinine and erythroagglutinin by the method of specific color luminescent dye. The activation of caspases-3 and -8 and induction of protein Bax expression under the influence of lectins were detected by Western blot analysis. It was revealed that erythroagglutinin induced apoptosis with the highest efficiency compared with leukoagglutinin and total phytohemagglutinine. The induction of apoptosis in human cell culture of not tumor origin 4BL is probably caused by activating caspase-dependent and mitochondrial signalling.

Trace elements storage peculiarities and metallothionein content in human thyroid gland under iodine deficiency euthyroid nodular goiter

H. I. Falfushynska1,2, L. L. Gnatyshyna1, O. Osadchuk2, V. O. Shidlovski2, О. B. Stoliar1

1Ternopil Volodymyr Hnatiuk National Pedagogical University, Ukraine;
2I.Ya. Horbachevski Ternopil State Medical University, Ukraine;
e-mail: halynka.f@gmail.com

Accumulation of iodine and copper in the node, paranodular and contralateral (not affected tissue by node) tissues of thyroid gland in relation to the level of metal-binding proteins, potential antioxidants and oxidative changes in tissue was investigated. To assess the severity of the pathological process the molecular markers of cytotoxicity were used. The reduction of total iodine (by 19.5%), increase of inorganic iodine fraction (by 82.4%) and total copper content (twice) in paranodular and nodular tissues compared with contrlateral part have been established. Excess of copper in goitrous-changes tissue was partially accumulated in the metallothioneins. The level of metal-binding form of metallothioneins and reserve of free thiols of these proteins was higher two-three times and lower content of reduced glutathione in node-affected tissue compared to the contralateral part. Signs of cytotoxicity among them: higher cathepsine D free activity (up to 84.6% and 134.4% in paranodular tissue and node respectively) and higher level of DNA strand breaks in the node (up to 22.6%) were observed. In paranodular tissue the range of indices variability compared with parenchyma of contralateral part is shorter than in the node. Thus, under low level of iodine organification and high copper level in goitrous-modified tissue of thyroid gland metallothionein may provide a partial compensatory effect on prooxidative processes.

Calpains: general characteristics and role in various states of the organism

N. F. Starodub1, L. M. Samokhina2, S. N. Koval2, I. A. Snegurskaya2

1National University of Life and Environmental Sciences of Ukraine, Kyiv;
e-mail: nikstarodub@yahoo.com;
2GD L. T. Malaya National Institute of Therapy of NAMS of Ukraine, Kharkov;
e-mail: lub.samokhina@yandex.ua

Calpains are a family of cytoplasmic calcium-dependent proteinases with papain-like activity. They participate in a variety of processes in the body: age changes, functioning of endothelium and pulmonary system, regulation of apoptosis and necrosis, development of various hypometabolic states, arterial hypertension, diabetes and chronic kidney disease, tumor growth. It is concluded that calpains, causing limited proteolysis of substrates, play an important role in a wide range of biological phenome­na. Their activity is associated with the response to the calcium-dependent signaling and the effects of aging. Inhibition of calpains activity contributes to inhibition of endothelial dysfunction, cardiovascular disease, formation of structural and functional changes in the kidney tissue, has neuroprotective effect, preventing sarcopenia, reduces inflammatory reactions caused by hyperventilation of the lungs.