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Dmytro Oleksiyovych Melnychuk

On the 75th anniversary of birthday

Nobel Laureates of the early 20th century E. Behring, I. Mechnikov, P. Ehrlich, C. Richet, J. Bordet, K. Landsteiner and their contribution to the development of molecular immunology

V. M. Danilova, R. P. Vynogradova, S. V. Komisarenko

Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;

The discoveries of immunologists have often been recognized as the most significant in the field of medicine and physiology, since the immune system is extremely vital for the organism, and the study of the principles of its functioning is of fundamental importance to the prevention (vaccination), diagnosis and therapy of many diseases. This article refers to the scientists of the early twentieth century, who received the most prestigious scientific award – the Nobel Prize in Physiology or Medicine and who built the groundwork for the development of immunology as a science. Thus, in 1901, E. von Behring received the first Nobel Prize “for his work on serum thera­py, especially its application against diphtheria, by which he has opened a new road in the domain of medical science and thereby placed in the hands of the physician a victorious weapon against illness and deaths”; in 1908, I. Mechnikov and P. Ehrlich received the Nobel Prize in Physiology or Medicine for the creating of the cellular and humoral theory of immunity; in 1913 – C. Richet – “in reco­gnition of his work on anaphylaxis”; in 1919 – J. Bordet – “for his discoveries relating to immunity (the role of complement, mechanisms of precipitation, agglutination…)”; in 1930 – K. Landsteiner – “for his discove­ry of human blood groups”. Their works spurred the development of modern molecular immunology – the science of the organization and function of the immune system, as an effective defense barrier in the living organism, which recognize and distinguish between “self” and “non-self”.

Long-term hypocholesterolemic effect of amidated alginate in rats

M. Marounek1, Z. Volek1, T. Taubner1, D. Dušková1, L. Kalachniuk2

1Institute of Animal Science, Prague, Czech Republic;
2National University of Life and Environmental Sciences of Ukraine, Kyiv;

The effect of octadecylamide of alginic acid on blood serum and hepatic cholesterol, and the faecal output of fat and sterols was examined in female rats fed diets containing cholesterol and palm fat at 10 and 50 g/kg, respectively for 10 weeks. Amidated alginate, supplied at 10 and 20 g/kg, significantly decreased serum cholesterol from 5.25 to 2.99 and 2.39 µmol/ml, respectively, and decreased hepatic cholesterol from 30.7 to 12.3 and 9.4 µmol/g, respectively. Amidated alginate increased the faecal output of fat and at higher dosing significantly decreased faecal output of bile acids. Faecal output of bile acids and hepatic cholesterol significantly correlated (r = 0.791; P < 0.001). The results of the present experiment showed that hypocholesterolemic effect of amidated alginate persisted within 10 weeks of feeding.

Preventive effect of N-stearoylethanolamine on memory disorders, blood and brain biochemical parameters in rats with experimental scopolamine-induced cognitive impairment

T. M. Horid’ko1, H. V. Kosiakova1, A. G. Berdyshev1, O. F. Meged1,
O. V. Onopchenko1, V. M. Klimashevsky1, О. S. Tkachenko1, V. R. Bazylianska1,
V. O. Kholin2, K. O. Peschana2, S. A. Mykhalskiy2, N. M. Hula1

1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
2Institute of Gerontology, National Academy of Medical Sciences of Ukraine, Kyiv;

The impairment of cognitive functions is the most studied medical and social problem nowadays. The aim of this study was to evaluate the protective effects of N-stearoylethanolamine (NSE) on memory state, blood and brain biochemical parameters in rats under scopolamine-induced cognitive impairment. The results of this study shown that NSE administration to rats per os (5 mg/kg, 5 days, during last 3 days NSE was administrated 20 min prior to scopolamine injection (1 mg/kg, once daily for 3 days, intraperitoneally)) prevented the development of memory impairment. In particular, NSE action was associated with the prevention of increase in acetylcholinesterase activity, changes in phospholipid, free and esterified cholesterol level in hippocampus and frontal cortex, and disruption in pro-/antioxidant balance in blood and studied brain sections. Considering the above mentioned biological effects, NSE is a promising drug candidate for integrative therapy of cognitive impairment of different profiles.

The effects of PDK4 inhibition on AMPK protein levels and PGC-1α gene expression following endurance training in skeletal muscle of Wistar rats

S. Aminizadeh1, Y. Masoumi-Ardakani1, B. Shahouzehi2

1Physiology Research Center, Institute of Basic and Clinical Physiology Sciences, Kerman University of Medical Sciences, Iran;
2Cardiovascular Research Center, Institute of Basic and Clinical Physiology Sciences, Kerman University of Medical Sciences, Iran;

There are regulatory networks in cells which surveil the physiological and environmental states. These cellular regulations are conducted through gene expression modulation. Skeletal muscle is able to adapt shortly and produce ATP at different conditions. AMPK (AMP-activated protein kinase) and PGC-1α (peroxisome proliferator-activated receptor-gamma coactivator-1alpha) are important regulators of cellular energy homeostasis. We designed this study to examine the effects of interactions between endurance training­ and PDK4 (pyruvate dehydrogenase kinase 4) inhibition on AMPK and PGC-1α expression in rat skeletal muscle. Thirty-two male Wistar rats were randomly selected and divided into 4 groups (n = 8); Group 1 control which did not receive any treatment, Group 2 received dichloroacetic acid (DCA) (150 mg/kg/day), Group 3 (endurance training group), Group 4 which received DCA and performed endurance training. AMPK protein expression, PDK4 and PGC-1α gene expression were measured by western blotting and real-time PCR, respectively. Our data showed that PDK4 inhibition caused AMPK protein elevation. Endurance training­ (group 2) and PDK4 inhibition (group 4) induce significant enhancement of PGC-1α gene expression compared to control group. The group which received DCA showed significant elevation of PDK4 gene expression compared to control group (P = 0.001), also other two groups (groups 2 & 3) showed significant elevation of PDK4 gene expression compared to control (P = 0.006). It seems that the combination of endurance training and PDK4 inhibition by up-regulation of PGC-1α expression, effectively improves energy state and performance in skeletal muscle.

Disturbance of the transmembrane phosphatidylserine asymmetry in hepatocytes as an apoptosis marker under the action of xenobiotics on rats

O. A. Nakonechna, L. A. Babijchuk, A. I. Bezrodna

Kharkiv National Medical University, Kharkiv, Ukraine;

It has been reported that unfavorable chemical environmental factors affect the functional state of liver, activate free radical processes against the background of the reduced antioxidant activity, change physico-chemical properties and membrane phospholipid composition of  hepatocytes. The aim of our research was to estimate phosphatidylserine distribution in the phospholipid bilayer of hepatocyte membranes and apoptosis stages in hepatocytes of rats under the influence of surfactants: ethyleneglycol (EG), polyethyleneglycol 400, (PEG-400) and polypropyleneglycol (PPG) at a dose of 1/10 DL50. It was found in the  subacute toxicological experiment on rats that the investigated xenobiotics EG, PEG-400 and PPG at a dose of 1/10 DL50 caused phosphatidylserine translocation to the outer membrane in the phospholipid bilayer of hepatocytes. This is a specific signal for macrophages aiming at recognition and elimination of apoptotic cells. Analysis of cell death modes under the influence of the investigated xenobiotics at a dose of 1/10 DL50 revealed that the intake of xenobiotics was associated with  an increase in the amount of apoptotic/necrotic hepatocytes.

Influence of C(60) fullerene on the ischemia-reperfusion injury in the skeletal muscle of rat limb: mechanokinetic and biochemical analysis

D. O. Zavodovskyi1, S. Yu. Zay2, T. Yu. Matvienko1, Yu. I. Prylutskyy1,
N. Y. Nurishchenko1, S. S. Paradizova3, L. L. Bezuh3, U. Ritter4, P. Scharff4

1ESC Institute of Biology and Medicine, Taras Shevchenko National University of Kyiv, Ukraine;
2Lesya Ukrainka Eastern European National University, Lutsk, Ukraine;
3SI The Territorial Medical Association of the Ministry of Internal Affairs of Ukraine in Kyiv;
4Institute of Chemistry and Biotechnology, Technical University of Ilmenau, Ilmenau, Germany;

Influence of the pristine C60 fullerene aqueous colloidal solution (C60FAS) on the ischemia-reperfusion injury in the skeletal muscle of rat limb was studied. Skeletal muscle damage effects were induced by 3 h lasting­ vascular ischemia. The impact of C60FAS was studied after its intramuscular injection immediately after 1 h of reper­fusion at different doses, namely: 1, 2 and 3 mg/kg of body weight. Changes in the mechanokinetic parame­ters of ischemic skeletal muscle contraction at different modes of functioning and biochemical parame­ters of blood were used as markers of ischemic injury, and analyzed in detail under action of C60FAS. The obtained results indicate on great promise of use of C60FAS to reduce the consequences of ischemic muscle trauma.

Immunogenicity assay of KatG protein from Mycobacterium tuberculosis in mice: preliminary screening of TB vaccine

P. Purkan1, R. Budiyanto1, R. Akbar1, S. P. A. Wahyuningsih2, W. Retnowati3

1Biochemistry Division, Chemistry Department, Faculty of Sciences and Technology, Airlangga University, Campus C, Jl. Mulyorejo-Surabaya, Indonesia;
2Biology Department, Faculty of Sciences and Technology, Airlangga University, Campus C, Jl. Mulyorejo-Surabaya, Indonesia;
3Microbiology Department, Faculty of Medicine, Airlangga University, Campus C, Jl. Moestopo-Surabaya, Indonesia;

The tuberculosis (TB) disease is still widely found even though BCG vaccine given to many people. Ineffectiveness of the BCG vaccine is one of causes that make the difficulties in preventing TB transmission. Objective of the research was to determine the immunogenicity of KatG protein of M. tuberculosis clinical isolate L19 in mice. The KatG protein as antigen was prepared by expression of the katG gene of M. tuberculosis clinical isolate L19 in Escherichia coli BL21 using pColdII-DNA vector. After purification by affinity chromatography, the KatG was vaccinated to mice to detect its immunogenicity. The expression of katG in E. coli BL21 could result in KatG protein with molecular weight 80 kDa in sodium dodecyl sulfate gel electrophoresis (SDS-PAGE). The pure KatG protein could significantly stimulate the immune response of mice by triggering the antibodies production of IgG1, IgG2a, IgG2b, IgG2c, IgG3, and IgM. The highest antibody level was obtained when the mice were vaccinated by KatG L19 with the dose of 45 μg/ml. Of the antibodies, the IgG2c isotype was dominantly produced in the blood serum. The KatG protein exhibited a high immunogenici­ty in mice, so it is possible to develop as a vaccine candidate for TB. A clinical test should be performed in a future to ensure its safety as a therapeutic protein.

Effects of alpha-ketoglutarate on lifespan and functional aging of Drosophila melanogaster flies

M. P. Lylyk1, M. M. Bayliak1, H. V. Shmihel1,
J. M. Storey2, K. B. Storey2, V. I. Lushchak1

1Vasyl Stefanyk Precarpathian National University, Ivano-Frankivsk, Ukraine;
2Institute of Biochemistry, Carleton University, Ottawa, Canada;

The effects of an alpha-ketoglutarate-supplemented diet on lifespan and functional senescence were evaluated in the Canton S strain of Drosophila melanogaster. The results suggest that effects of dietary alpha-ketoglutarate (AKG) are dose- and gender-dependent. In males, diets containing 1-10 mM AKG did not affect mean and maximum lifespans, except that an increased maximum lifespan observed at 10 mM AKG. Diet with 20 mM AKG shortened median lifespan and had no effect on maximum lifespan of males. In females, diets with low concentrations of AKG (1 and 5 mM) did not affect lifespan, whereas diets supplemented with 10 and 20 mM AKG increased both median and maximum lifespans. At a lifespan-prolonging concentration (10 mM), AKG decreased fecundity, increased cold resistance and had no effect on climbing activity or resistance to oxi­dative stress in flies of either gender at middle (24 days) and old (40 days) ages. Moreover, middle-aged AKG-fed females but not males were more resistant to heat stress that was accompanied by higher levels of HSP90 protein as compared with controls. Middle-aged flies on AKG-supplemented diets showed elevated oxidative stress and had higher total protein and triacylglycerol levels as compared with controls. Hence, anti-aging effects of AKG do not seem to be related to preventing oxidative stress development but involve metabolic rearrangement and synthesis of specific protective proteins, which aid to resist destructive processes with age.