Tag Archives: myometrium
Ca(2+)-dependent mechanisms of mitochondria-targeted calix[4]arene C-772 effect on uterine smooth muscle contractile activity
I. S. Forys1 (https://orcid.org/0009-0002-8958-8244)
O. V. Tsymbalyuk2 (https://orcid.org/0000-0002-4524-7627)
Yu. V. Danylovych1 (https://orcid.org/0000-0002-3526-7085)
H. V. Danylovych1* (https://orcid.org/0000-0003-0571-4494)
M. V. Rudnytska1 (https://orcid.org/0000-0001-7766-3900)
R. V. Rodik3 (https://orcid.org/0000-0003-2258-6957)
S. O. Kosterin1 (https://orcid.org/0000-0003-2961-5554)
1Department of Muscle Biochemistry, Palladin Institute of Biochemistry,
National Academy of Sciences of Ukraine, Kyiv;
2Department of Molecular Biotechnology and Bioinformatics, Educational Scientific Institute
of High Technologies, Taras Shevchenko National University of Kyiv, Ukraine;
3Department of Chemistry of Macrocyclic Compounds, Institute of Organic Chemistry,
National Academy of Sciences of Ukraine, Kyiv;
*e-mail: danylovychanna@ukr.net
Received: 06 April 2026; Revised: 06 May 2026;
Accepted: 29 May 2026; Available on-line: June 2026
Background. Identification of non-toxic exogenous compounds capable of selectively influencing intracellular Ca2+-dependent processes and smooth muscle contractility remains an important task of molecular biotechnology. Calix[4]arenes are considered promising modulators of cellular functions, and calix[4]arene C-772 is suggested to selectively effect mitochondrial Ca2+ handling. Objective. The aim of this study was to evaluate the effect of calix[4]arene C-772 on Ca2+ transport in mitochondria of uterine smooth muscle, nitric oxide production in myocytes, and the contractile activity of myometrial strips. Methods. Non-pregnant Wistar rats were used in experiments, confocal microscopy, spectrofluorimetry, flow cytometry, and mechanokinetic analysis were applied. Results. Mitochondrial localization of calix[4]arenes in myocytes was confirmed by colocalization of the сalix[4]arene–FITC conjugate C-1308 with Mitotracker Orange CM-H2TMRos. Calix[4]arene C-772 (10 μM) interacted with cardiolipin in mitochondrial membranes, inhibited nitric oxide synthesis, reduced Ca2+ accumulation in isolated mitochondria and increased the amplitude of rat myometrial smooth muscle spontaneous contractions. Mechanokinetic analysis demonstrated increased force, velocity, and impulse parameters of contraction–relaxation cycles. Conclusions. These findings indicate that calix[4]arene C-772 at micromolar concentrations can serve as an effective modulator of mitochondrial functional activity and uterine smooth muscle contractility.
Department of Muscle Biochemistry: calixarenes as modulators of energy-dependent Са(2+)-transporting pumps in smooth muscles
S. O. Kosterin
Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine,
Department of Muscle Biochemistry, Kyiv;
e-mail: kinet@biochem.kiev.ua
Received: 12 May 2025; Revised: 18 June 2025;
Accepted: 30 October 2025; Available on-line: 02 December 2025
In this scientific-historical review devoted to the recent achievements of the Muscle Biochemistry Department of the Palladin Institute of Biochemistry, the NAS of Ukraine, we synthesize findings from interdisciplinary investigations of intracellular calcium homeostasis in smooth muscle (exemplified by the myometrium) conducted at the interface of biochemistry, physical and organic chemistry, biophysics, and mathematical/computational modeling. We emphasize that the selected calix[4]arenes considered here act selectively as inhibitors of the Mg2+,ATP-dependent calcium and sodium pumps – ion-transporting ATPases (electroenzymes Ca2+,Mg2+-ATPase and Na+,K+-ATPase) – of the plasma membrane of smooth-muscle cells, enabling controlled modulation of intracellular Ca2+ homeostasis and the contractile activity of the myometrium. The data obtained also indicate that the selected calix[4]arenes can be regarded as compounds suitable for efficient investigation of mitochondrial function in smooth-muscle cells, in particular the mechanisms of transmembrane Ca2+ exchange, the principles governing membrane-potential formation, and the contribution of these subcellular structures to the control of the mechanokinetics of the contraction–relaxation cycle. It is shown that some calix[4]arenes act as effectors of the ATPase activity of contractile proteins and protect this activity from the inhibitory influence of heavy-metal ions. Taken together, these results outline biochemical approaches to the fine regulation of calcium fluxes and smooth-muscle contractility and underscore the potential of calix[4]arenes as selective “molecular platforms” useful for addressing fundamental and applied (biomedical) problems in contemporary physico-chemical muscle biology.
Thiacalix[4]arene С-1193 – a promising inhibitor of the sodium pump in the uterine smooth muscle cells
O. V. Maliuk1*, T. O. Veklich1, O. V. Tsymbalyuk2, O. V. Bevza1,
S. O. Cherenok3, A. I. Selikhova3, V. I. Kalchenko3, S. O. Kosterin1
1Palladin Institute of Biochemistry, National Academy of Sciences
of Ukraine, Kyiv, Ukraine;
*e-mail: sanya2000ua@gmail.com;
2Educational and Scientific Institute of High Technologies,
Taras Shevchenko National University of Kyiv, Kyiv, Ukraine;
3Institute of Organic Chemistry, National Academy of Sciences of Ukraine, Kyiv, Ukraine
Received: 29 May 2025; Revised: 18 July 2025;
Accepted: 12 September 2025; Available on-line: 17 September 2025
Thiacalix[4]arene C-1193 (25,27-dibutoxythiacalix[4]arene-bis-hydroxymethylphosphonic acid) was shown to inhibit the activity of Na+,K+-ATPase with a high efficiency (І0.5 = 42.1 ± 0.6 nM) with no effect on the activity of Mg2+-ATPase, Са2+-ATPase and Са2+,Mg2+-ATPase in the plasma membrane fraction of rat uterine smooth muscle cells. The kinetic regularities of the C-1193 inhibitory effect on Na+,K+-ATPase activity were investigated. It was demonstrated that C-1193 increased the enzyme activation constant by Na+ but not by K+ ions. The contractile activity of the rat uterine horns was investigated by tenzometric methods with the use of longitudinal uterine smooth muscle strips with intact endometrium. С-1193 induced a considerable increase in the amplitude of the acetylcholine-induced contractions as well as the maximal velocity of the contraction and relaxation phases. No effect of С-1193 on contractive activity induced by the selective agonist of М3-cholinoreceptors cevimeline was observed. The results of computer simulation showed that С-1193inhibitory effect must be related to the cooperative action of methylene bisphosphonate fragments on the upper rim of the calixarene platform, and the linker sulfur atoms of calixarene “cup” on the Na+,K+-ATPase macrostructure.
Kinetic regularities of thiacalix[4]arene C-1087 inhibitory effect on the activity of Mg(2+)-dependent Ca(2+)-transporting ATP hydrolase in the plasma membrane of smooth muscle cells
Т. О. Veklich1, О. V. Bevza1, О. V. Maliuk1*, S. О. Kosterin1,
R. V. Rodik2, S. H. Vyshnevskyi2, V. І. Kalchenko2
1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
*e-mail: alexmaliukid@gmail.com;
2Institute of Organic Chemistry, National Academy of Sciences of Ukraine
Received: 05 November 2023; Revised: 04 January 2024;
Accepted: 01 February 2024; Available on-line: 26 February 2024
The experiments with the suspension of plasma membranes of myometrium cells, treated with 0.1% digitonin solution, were used to study kinetic regularities of the inhibitory effect of tetra-N-phenylsulfonyl trifluoroacetamidine-thiacalixarene (С-1087) on the activity of Са2+,Mg2+-ATPase. The studies demonstrated the impact of C-1087 on the cumulative effect and the maximal velocity of ATP hydrolysis. No effect of С-1087 on the affinity between Са2+,Mg2+-ATPase, and АТР, affinity and cumulative effect of Ca ions and activation coefficient for Mg ions was revealed. A considerable decrease in the maximal velocity of ATP hydrolysis evidenced a complete non-competitive mechanism of inhibiting Са2+,Mg2+-АТРase activity with thiacalix[4]arene С-1087. Computer simulation demonstrated that thiacalix[4]arene С-1087 inhibiting effect on Са2+,Mg2+-ATPase may be conditioned by the cumulative effect of four spatially oriented N-sulfonylamidine groups on the upper rim of its macrocyclic platform.
Thiacalix[4]arene С-1087 is the selective inhibitor of the calcium pump of smooth muscle cells plasma membrane
Т. О. Veklich1*, R. V. Rodik2, О. V. Tsymbalyuk3,
О. V. Shkrabak1, O. V. Maliuk1, S. O. Karakhim1,
S. H. Vyshnevskyi3, V. І. Kalchenko3, S. O. Kosterin1
1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
*e-mail: veklich@biochem.kiev.ua;
2Institute of Organic Chemistry, National Academy of Sciences of Ukraine, Kyiv;
3Educational and Scientific Institute of High Technologies,
Taras Shevchenko National University of Kyiv, Kyiv, Ukraine;
Received: 01 September 2023; Revised: 23 October 2023;
Accepted: 01 December 2023; Available on-line: 18 December 2023
The enzymatic and kinetic analyses were used to demonstrate that 5,11,17,23-tetra(trifluoro)methyl(phenylsulfonylimino)methylamino-25,27-dihexyloxy-26,28-dihydroxythiacalix[4]arene С-1087 effectively inhibited the Са2+,Mg2+-АТРase activity of the rat myometrium cells plasma membrane (І0.5 = 9.4 ± 0.6 µM) with no effect on the relative activity of other membrane ATPases. With the use of confocal microscopy and Ca2+-sensitive fluorescent probe fluo-4, it was shown that the application of thiacalix[4]arene С-1087 to the immobilized uterus myocytes increased the cytosolic concentration of Ca2+. Tenzometric studies of rat uterus smooth muscles with the subsequent mechanokinetic analysis revealed that thiacalix[4]arene С-1087 considerably decreased the maximal velocity of the relaxation of both spontaneous contractile response and contraction induced by hyperpotassium solution.
A new affine inhibitor of sodium pump thiacalix[4]arene С-1193 increases the intracellular concentration of Ca ions and modifies myometrium contractility
Т. О. Veklich1*, S. О. Cherenok2, О. V. Tsymbalyuk3, О. A. Shkrabak1,
S. O. Karakhim1, A. I. Selihova2, V. І. Kalchenko2, S. O. Kosterin1
1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
*e-mail: veklich@biochem.kiev.ua;
2Institute of Organic Chemistry, National Academy of Sciences of Ukraine, Kyiv;
3Educational and Scientific Institute of High Technologies,
Taras Shevchenko National University of Kyiv, Ukraine
Received: 16 June 2023; Revised: 31 September 2023;
Accepted: 27 October 2023; Available on-line: 06 November 2023
The methods of enzymatic and kinetic analysis were used to demonstrate that thiacalix[4]arene-bis-hydroxymethylphosphonic acid С-1193 had the inhibitory effect (І0.5 = 42.1 ± 0.6 nM) on Na+,K+-ATPase activity in the plasma membrane of myometrium cells with no effect on the relative activity of other ATPases localized in this subcellular structure. The method of confocal microscopy and Са2+-sensitive fluorescent probe fluo-4 were used to demonstrate that thiacalix[4]arene С-1193 increased the intracellular concentration of Ca ions in the immobilized uterine myocytes. The tenzometric studies proved that С-1193 (10 and 100 μМ) increased the isometric phasic contractions, induced via the paths of both electromechanical (depolarization with high-potassium solution) and pharmacomechanical (application of uterotonic hormone oxytocin, neurotransmitter acetylcholine or selective agonist of muscarinic acetylcholine receptors cevimeline) coupling. Application of thiacalix[4]arene С-1193 as a selective and effective inhibitor of Nа+,K+-ATPase may be useful both for studyng the regulation of ion homeostasis in smooth muscle cells and creation of new uterotonics based on the calixarene core.
Inhibition of Na(+),K(+)-ATPase and activation of myosin ATPase by calix[4]arene C-107 cause stimulation of isolated smooth muscle contractile activity
T. O. Veklich1, R. D. Labyntseva1, O. A. Shkrabak1, O. V. Tsymbalyuk2,
R. V. Rodik3, V. I. Kalchenko3, S. O. Kosterin1
1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
2Institute of High Technologies, Taras Shevchenko National University of Kyiv, Ukraine;
3Institute of Organic Chemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: veklich@biochem.kiev.ua; otsymbal@bigmir.net; manli@ioch.kiev.ua
Received: 04 Jule 2019; Accepted: 29 November 2019
The discovery of compounds that might modify myometrial contractility is an important area of researches. In our previous experiments, we found that some representatives of macrocyclic compounds family – calix[4]arenes – can modify the enzymatic and transport activity of membrane-bound cation-transport ATP hydrolases. The aim of this work was to study and compare the effect of calix[4]arene C-107 on the enzymatic activities of Mg2+-dependent ATPases of the uterine smooth muscle, namely: ouabain-sensitive Na+,K+-ATPase, plasma membrane Ca2+-independent “basal” Mg2+-ATPase, ATPase of the actomyosin complex and myosin subfragment-1, with effect on the contractile activity of the myometrium. It was shown that calix[4]arene C-107 efficiently inhibited myometrium Na+,K+-ATPase (I50 = 54 ± 6 nM) selectively to other ATP-hydrolases of the plasma membrane and simultaneously activated the enzymatic activity of the myosin ATPase of smooth muscles (A50 = 9.6 ± 0.7 μM). Such reciprocal biochemical effects led to the stimulation of the smooth muscle contractile activity that was demonstrated by the tensometric method using different isolated smooth muscles. Calix[4]arene С-107 was shown to stimulate the increase of the tonic component of myometrium contractions induced by oxytocin, as well as contractions of the caecum muscles induced by high-potassium solution or acetylcholine, and to maintain increased tension for a long time. Thus, calix[4]arene C-107 is a prospective compound for enhancing the smooth muscle basal tone and/or contraction in case of hypotonic dysfunctions.
Сalix[4]arene chalcone amides effects on myometrium mitochondria
S. G. Shlykov1, A. M. Kushnarova-Vakal1, A. V. Sylenko1,
L. G. Babich1, О. Yu. Chunikhin1, O. A. Yesypenko2,
V. I. Kalchenko2, S. O. Kosterin1
1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: sshlykov@biochem.kiev.ua;
2Institute of Organic Chemistry, National Academy of Sciences of Ukraine, Kyiv
Received: 19 November 2018; Accepted: 14 March 2019
Mitochondria are a key player in a wide range of the most important functions of the cell. Calixarenes are supramolecular compounds that have been widely used in bioorganic chemistry and biochemistry. The aim of this work was to study the effects of calix[4]arenes with two (С-1012, С-1021), three (С-1023, С-1024) and four (С-1011) chalcone amide groups on the myometrial mitochondria membranes polarization, Ca2+ concentration in the matrix of these organelles ([Ca2+]m ) and on the average hydrodynamic diameter of mitochondria. It was shown that permeabilized myometrium cells incubation with calix[4]arenes containing two or more chalcone amide groups, was accompanied by an increased level of myometrial mitochondria membranes polarization. All studied calix[4]arenes increased [Ca2+]m values in the absence and in the presence of exogenous Ca2+. The values of [Ca2+]m in the absence of exogenous Ca2+ were higher at mitochondria incubation in Mg2+-containing, than in Mg2+,ATP-containing medium. Incubation of isolated mitochondria with the studied calix[4]arenes resulted in changes of mitochondria volume: at incubation with С-1012, С-1021, C-1023 the average hydrodynamic diameter was decreased, while with С-1011 it was increased. Thus, we have shown that a short-term (5 min) incubation of mitochondria in the presence of 10 µM calix[4]arenes, which contain from two to four chalcone amide groups, increased the level of mitochondria membranes polarization, ionized Ca concentration in the matrix and had different effects on the mitochondrial volume.
Calix[4]arene С-956 selectively inhibits plasma membrane Са(2+),Mg(2+)-АТРase in myometrial cells
Т. O. Veklich1, O. A. Skrabak1, Yu. V. Nikonishyna1, R. V. Rodik2, V. I. Kalchenko2, S. O. Kosterin1
1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
2Institute of Organic Chemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: veklich@biochem.kiev.ua; manli@ioch.kiev.ua
Using enzymatic assays and kinetic analysis, we demonstrated that 100 µM calix[4]arene C-956 (5,11,17,23-tetra(trifluoro)methyl-(phenylsulfonylimino) methylamino-25,27-dioctyloxy-26,28-dipropoxycalix[4]arene) had the most significant inhibitory effect on the plasma membrane Са2+,Mg2+-АТРase activity compared to effects of other calix[4]arenes, and had no effect on specific activities of other membrane ATPases. Using confocal microscopy and fluorescent probe fluo-4, we observed an increase of the intracellular level of Ca2+ after application of calix[4]arene C-956 to immobilized myocytes. Analysis of the effect of calix[4]arene C-956 on the hydrodynamic diameter of myocytes demonstrated that application of calix[4]arene C-956 solution decreased this parameter by 45.5 ± 9.4% compared to control value similarly to the action of uterotonic drug oxytocin.
The relationship between the ionized Ca concentration and mitochondrial functions
L. G. Babich1, S. G. Shlykov1, A. M. Kushnarova-Vakal1, N. I. Kupynyak2, V. V. Manko2, V. P. Fomin3, S. O. Kosterin1
1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
e-mail: babich@biochem.kiev.ua;
2Ivan Franko National University of Lviv, Ukraine;
3University of Delaware, Newark, USA
The aim of the study was to show the relationships between ionized Ca concentration ([Ca2+]m) in the mitochondria matrix and functional activity of this organelle. [Ca2+]m was determined using the fluorescent probe Fluo-4, AM. Total level of Ca2+ accumulation in mitochondria was monitored using 45Ca2+ as radioactive tracer. It was shown that incubation of myometrium mitochondria with 3 mM Mg2+ resulted in the low level of [Ca2+]m. Subsequent addition of 100 µM Ca2+ resulted in 8 times increase of [Ca2+]m but in low level of total calcium accumulation. Normalized fluorescence of Ca2+-sensitive probe Fluo-4 in response to the Ca2+ addition was higher than 2.5. At the same time, [Ca2+]m was considerably higher in the medium containing 3 mМ АТР and 3 mМ Mg2+. Subsequent addition of 100 µM Ca2+ to the incubation medium resulted in only 2.4 times increase of [Ca2+]m but considerably higher level of total calcium accumulation was observed. Normalized fluorescence of Fluo-4 in response to the Ca2+ addition was lower than 1.3. In liver mitochondria higher rate of oxygen consumption was detected in the presence of an oxidative substrate succinate than of pyruvate or α-ketoglutarate. At the presence of an oxidative substrate succinate normalized fluorescence of Fluo-4 in liver mitochondria in response to the Ca2+ addition was lower than 1.3. It was concluded that low level of [Ca2+]m was correlated with low functional activity of this organelle and, vise versa, high level of [Ca2+]m was correlated with high functional activity. It was suggested that normalized fluorescence changes in response to the Са2+ addition could be used as a test of the mitochondrial functional activity: lower normalized fluorescence values − higher functional activity.