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General Physiology and Biophysics

Volume 35, 2016, No. 4


  It takes two T to shape immunity: emerging role for T-type calcium channels in immune cells
Lubica Lacinova 1), Norbert Weiss

1)Institute of Molecular Physiology and Genetics, Slovak Academy of Sciences, Bratislava, Slovak Republic.

General Physiology and Biophysics. Volume 35, 2016, No. 4: 393-396.

  Hemodynamic properties and arterial structure in male rat offspring with fetal hypothyroidism
Mahboubeh Ghanbari 1), Fatemeh Bagheripuor, Abbas Piryaei, Saleh Zahediasl, Mahsa Noroozzadeh, Asghar Ghasemi

1)Endocrine Physiology Research Center, Research Institute for Endocrine Sciences, Shahid Beheshti University of Medical Sciences, Tehran, Iran.

Thyroid hormones (THs) play a crucial role in the development of different systems during fetal life; fetal hypothyroidism (FH) is associated with reduced cardiac function and dimensions in neonates. The aim of this study is to determine whether TH deficiency during fetal life is associated with arterial structural and hemodynamic changes during adulthood. Hypothyroidism was induced by adding 0.025% 6-propyl-2-thiouracil in drinking water throughout pregnancy, while controls consumed only tap water. Hemodynamic parameters, cross-sectional area, intima-media thickness (IMT), and density of nuclei of smooth muscle cells and endothelial cells (ECs) in the aorta and mesenteric arteries were measured. Compared to controls, in the FH group, baseline systolic blood pressure (105.7 ± 3.1 vs. 87.9 ± 3.3 mm Hg, p < 0.01), diastolic blood pressure (64.4 ± 1.7 vs. 53.2 ± 2.1 mm Hg, p < 0.05), and mean arterial pressure (80.9 ± 2.1 vs. 67.1 ± 2.1 mm Hg, p < 0.01) were significantly lower. In addition, in the FH group, intensity and latency of response to phenylephrine were significantly lower and longer, respectively, as were the IMT and density of ECs in the aorta and superior mesenteric arteries. In conclusion, this study showed that TH deficiency during fetal life can have long-lasting functional and histological effects, which can compromise cardiovascular function during adulthood.

General Physiology and Biophysics. Volume 35, 2016, No. 4: 397-405.

  Hidden information in three-axial ECG data of normal subjects: Fractal dimensions of corresponding points from successive QRS loops as a potential sport and age dependent marker
Srdjan Čalošević 1), Kristijan Dinjar 2), Suzana Čalošević 3), Sven Kurbel 4), Robert Steiner 5)

1)Osijek University Hospital Center, J. Huttlera 4, Osijek, Croatia.
2)Osijek University Hospital Center, J. Huttlera 4, Osijek, Croatia.
3)Osijek University Hospital Center, J. Huttlera 4, Osijek, Croatia.
4)Osijek Medical Faculty, J. Huttlera 4, Osijek, Croatia.
5)Osijek Medical Faculty, J. Huttlera 4, Osijek, Croatia.

This study is aimed to estimate spatial variability of normal QRS loops trajectories in successive cardiac cycles analyzed in XYZ phase space among 27 young and 27 older subjects, both male and female. Among young subjects, 10 individuals were professional football players, while the remaining 17 were students without regular physical activities. Modified three-axial leads (Frank's lead system) were used continually to record high-resolution ECG (1 kHz sampling rate) at rest during 200 seconds by Biopac Student Lab System. Variability of all the three spherical coordinates of the ventricular electrical vector in five characteristic positions regarding the R peak was analyzed. It was found that although fractal dimensions were not gender-dependent (p > 0.05), the calculated values in all tested five positions were greater in young than in older individuals (p < 0.05). Besides that, among young subjects, several fractal dimensions were greater in athletes than in physically inactive students (p < 0.05). These results suggest that the availability of possible phase space trajectories for QRS vector motion is related to individual physical activity and ageing, leaving less variability to physically inactive or older individuals. Recognizing noninvasive features of heart activity might help early detection of heart problems caused by aging and sedentary life style.

General Physiology and Biophysics. Volume 35, 2016, No. 4: 407-415.

  Effect of low frequency pulsed magnetic field on gravitropic response and cell elongation in coleoptiles of maize seedlings
Anna Kościarz-Grzesiok 1), Karolina Sieroń-Stołtny 2), Malgorzata Polak 3), Aleksander Sieroń 4), Waldemar Karcz 5)

1)Chair and Clinic of Internal Diseases, Angiology and Physical Medicine, Silesian Medical University, Bytom, Poland.
2)Chair and Clinic of Internal Diseases, Angiology and Physical Medicine, Silesian Medical University, Bytom, Poland.
3)Department of Cell Biology, Faculty of Biology and Environmental Protection, University of Silesia, Jagiellońska 28, Katowice, Poland.
4)Chair and Clinic of Internal Diseases, Angiology and Physical Medicine, Silesian Medical University, Bytom, Poland.
5)Department of Plant Physiology, Faculty of Biology and Environmental Protection, University of Silesia, Jagiellońska 28, Katowice, Poland.

The effect of pulsed magnetic field (PMF) on gravitropic response, endogenous growth and growth in the presence of indole-3-acetic acid (IAA) was studied in coleoptiles of maize (Zea mays L.) seedlings. Medium pH changes, measured simultaneously with growth of coleoptile segments, were also determined. In seedlings grown in the presence of PMF, elongation growth of coleoptiles was inhibited by 16%, while growth of roots and mesocotyls did not depend on PMF. Magnetic field also inhibited (by 36%) the gravitropic response of maize seedlings. However, when PMF was applied only during gravistimulation (within 6 h), it suppressed the gravitropic reaction only by 8% at 6 h. It was also found that endogenous growth and IAA-induced growth of maize coleoptile segments excised from seedlings treated with the PMF was stimulated by 52% and 30%, respectively, as compared to control (segments untreated with the PMF). Values of medium pH, measured simultaneously with growth, indicated that PMF-treated coleoptile segments extruded much more protons than untreated segments. In contrast, coleoptile segments treated with the PMF and subsequently incubated in the presence of IAA extruded 2.5-fold less protons as compared to segments treated with IAA only. The data presented here have been discussed with consideration of mechanisms by which auxin (IAA) regulates plant cell growth.

General Physiology and Biophysics. Volume 35, 2016, No. 4: 417-424.

  L-arginine supplementation attenuates capillary regression without increasing integrated succinate dehydrogenase activity and VEGF expression in skeletal muscle during hindlimb unloading
Kensaku Uchida 1), Minoru Tanaka, Hiroyo Kondo, Akihiko Ishihara, Hidemi Fujino

1)Department of Rehabilitation Science, Kobe University, Graduate School of Health Sciences, 7-10-2 Tomogaoka, Suma-ku, Kobe 654-0142, Japan.

Decreased capillary number is observed in atrophied muscle. The change in capillary number is regulated by angiogenic factors. L-arginine enhances expression of endothelial nitric oxide synthase (eNOS), angiogenic factor, in skeletal muscle. Therefore, the aim of this study was to evaluate the effects of L-arginine supplementation on capillary regression during hindlimb unloading. Twenty-four male Wistar rats were divided into four treatment groups: (1) control, (2) L-arginine supplementation, (3) hindlimb unloading, and (4) hindlimb unloading with L-arginine supplementation. Hindlimb unloading resulted in a decrease of capillary-to-muscle fibre (C/F) ratio, eNOS expression, and integrated succinate dehydrogenase (SDH) activity. L-arginine supplementation attenuated the decrease in both eNOS expression and C/F ratio, although it did not increase integrated SDH activity in skeletal muscle. These results indicate that L-arginine supplementation is effective for maintaining capillary number in atrophied muscle, and that elevation of eNOS expression may be one mechanism associated with these responses.

General Physiology and Biophysics. Volume 35, 2016, No. 4: 425-432.

  A comparative study between the effect of 17-β estradiol and angiotensin converting enzyme inhibitor on osteoporosis in ovariectomized rats
Mona Alaam 1), Noha Hussien

1)Department of Physiology, Faculty of Medicine, Benha University, Benha, Egypt.

Osteoprotegerin (OPG) and receptor activator of nuclear factor-kappa B ligand (RANKL) have been revealed in the pathogenesis of primary osteoporosis and other metabolic bone diseases. This study was designed to assess the effect of 17-β estradiol (E2) treatment and angiotensin converting enzyme inhibitor (ACEI), captopril, on osteoporosis induced by ovariectomy in rats and discussing the role of OPG/RANKL ratio in their action. Thirty two adult female rats were divided into four equal groups. Group I: control group, Group II: ovariectomized (OVX) non treated group, Group III: OVX rats treated with E2, Group IV: OVX rats treated with captopril. OVX rats showed a significant decrease in serum Ca2+ and OPG levels with significant increase in serum RANKL, osteocalcin, alkaline phosphatase activity and urinary hydroxyproline levels. Treatment with captopril as well as E2 led to a significant improvement in bone markers levels with a significant increase in OPG/RANKL ratio. Image analysis technique revealed that there was a significant improvement in cortical bone thickness (CBT) and mean trabecular bone density (TBD) in OVX rats treated with either E2 or ACEI. So, we can conclude that the protective effect of E2 and ACEI on osteoporosis may be mediated by influencing OPG/RANKL signaling.

General Physiology and Biophysics. Volume 35, 2016, No. 4: 433-441.

  Swimming training affects apoptosis-related microRNAs and reduces cardiac apoptosis in mice
Yongcai Zhao 1), Zhiyong Ma

1)Department of Physical Education, Tangshan Normal University, Hebei, PR China.

We sought to investigate effects of exercise training on apoptosis-related microRNAs (miRs) and their validated targets, discussing molecular mechanism of the exercise-induced benefit in heart. Male C57BL/6 mice were randomly assigned to three groups: sedentary (SE), exercise training 1 (ET1) and exercise training 2 (ET2). ET1 swam for 8 weeks, once a day and 5 days per week with incremental load. ET2 performed the same work as ET1 and switched to twice a day by the end of the 5th week. In ET2, positive cell rate (%) tested by TUNEL assay decreased significantly (p < 0.05), and the load decreased miR-1 level by 29% (p < 0.01), also increased miR-30b and miR-21 levels by 32% (p < 0.01) and 18% (p < 0.05) respectively. In addition, Bcl-2 expression was increased by 98% (p < 0.01). p53, PDCD4 and Drp-1 expressions were decreased by 45% (p < 0.01), 6% (p > 0.05) and 36% (p < 0.01) respectively, compared with SE. In ET1, only miR-30b level was increased by 22% (p < 0.05) with a 48% decrease in p53 level (p < 0.01). Both swimming groups increased Bcl-2/Bax ratio significantly (p < 0.01). This study indicated that apoptosis-related miRs and their downstream proteins in heart can be influenced by swimming training that may be responsible for the exercise-induced cardioprotection.

General Physiology and Biophysics. Volume 35, 2016, No. 4: 443-450.

  Apoptotic mechanisms of N1-acetylacetone, N4-4-methoxy-salicylidene-thiosemicarbazide chelating with Nickel(II) on HL60 leukemia cells
Büşra Kaya 1), Belkis Atasever-Arslan, Zeynep Kalkan, Hazal Gür, Bahri Ülküseven

1)Department of Chemistry, Faculty of Engineering, İstanbul University, Istanbul, Turkey.

Thiosemicarbozone complexes that have a broad spectrum of biological activity are formed as a result of condensation reaction between thiosemicarbazide [H2N(C=S)-NH-NH2] and carbonyl-containing compounds. A new Nickel(II) complexes with N1-acetylacetone, N4-4-methoxy-salicylidene-thiosemicarbazidato ligand was synthesized and characterized. We studied the antileukemic activity of the Ni(II) thiosemicarbazone compound and assessed their potential for drug development. Specifically, the effects of this Ni(II) thiosemicarbazone compound on intracellular signal nodes and apoptotic pathways were investigated. According to our results, the Ni(II) thiosemicarbazone compound has apoptotic activity against HL60 cells. Moreover, while Ni(II) thiosemicarbazone compound significantly increased levels of p53 and cleaved caspase-3 proteins, it decreased level of Phospho-Akt1 protein in HL60 cells. The Ni(II) thiosemicarbazone compound could induce HL60 cell apoptosis through inhibiting of PI3K/Akt pathway. These results showed that Ni(II) thiosemicarbozone compound might be an antileukemic agent.

General Physiology and Biophysics. Volume 35, 2016, No. 4: 451-458.

  Hypericin fluorescence kinetics in the presence of low density lipoproteins: study on quail CAM assay for topical delivery
Monika Buríková 1), Boris Bilčík, Mariana Máčajová, Pavel Výboh, Jozef Bizik, Anton Mateašík, Pavol Miškovský, Ivan Čavarga

1)Institute of Animal Biochemistry and Genetics, SAS, Ivanka pri Dunaji, Slovak Republic.

There has been increasing interest in fluorescence-based imaging techniques in clinical practice, with the aim to detect and visualize the tumour configuration and the border with healthy tissue. Strong photodynamic activity of hypericin (Hyp) can be improved by various molecular transport systems (e.g. LDL). Our aim was to examine pharmacokinetics of Hyp in the presence of LDL particles on ex ovo chorioallantoic membrane (CAM) of Japanese quail with implanted TE1 tumour spheroids (human squamocellular carcinoma). Spheroids were implanted on CAM surface on embryonal day 7 and after 24 hours formulations of free Hyp and Hyp:LDL 100:1 and 200:1 were topically applied. All experimental formulations in the fluorescent image very well visualized the tumour spheroid position, with gradual increase of fluorescence intensity in 6-h observation period. LDL transportation system exhibited clear superiority in fluorescence pharmacokinetics than free Hyp formulation by increasing tumour-normal difference. Our experimental results confirm that Hyp and Hyp:LDL complex is potent fluorophore for photodynamic diagnosis of squamocellular carcinoma.

General Physiology and Biophysics. Volume 35, 2016, No. 4: 459-468.

  Influance of regular swimming on serum levels of CRP, IL-6, TNF-α in high-fat diet-induced type 2 diabetic rats
Rafighe Ghiasi 1), Farhad Ghadiri Soufi, Gisou Mohaddes, Alireza Alihemmati, Mohammad Somi, Hadi Ebrahimi, Fariba Mirzaie Bavil, Mohammad Alipour

1)Liver and Gastrointestinal Diseases Research Center, Tabriz University of Medical Sciences, Tabriz, Iran.

Due to key role of inflammation in pathogenesis of type 2 diabetes mellitus (T2DM), aim of this study was evaluating the influance of regular swimming on serum levels of C-reactive protein (CRP), interlukin-6 (IL-6), tumor necrosis factor-α (TNF-α) in high-fat diet-induced diabetic rats. Fourty male Wistar rats were randomly divided into control, diabetic, exercise and diabetic-exercise groups (n = 10). Diabetes was induced by high-fat diet and streptozotocin (35 mg/kg, i.p.). In exercise groups, after induction of diabetes, animals were subjected to swimming (60 min/5 days a week) for 10 weeks. At the end of training, rats were anestatized and blood samples and pancreatic tissues were collected and used for evaluation of CRP, IL-6, TNF-α and pancreatic histopatholology. Our results showed significantly increase in lymphocytes, monocytes and decrease in neutrophils in diabetic rats (p < 0.01), which these parameters significantly reversed to control levels by induction of swimming (p < 0.01). In diabetic group, the levels of CRP, IL-6 and TNF-α increased (p < 0.01), and swimming decreased these factors significantly. Histopathological results of this study also showed that swimming can prevent damage induced by diabetes. The present study indicates that swim training is associated with improved inflammation and inflammatory mediators and pancreatic damage.

General Physiology and Biophysics. Volume 35, 2016, No. 4: 469-476.

  Ultrastructural remodelling of slow skeletal muscle fibres in creatine kinase deficient mice: a quantitative study
Marta Novotová 1), Bohumila Tarabová, Lucia Tylková, Renée Ventura-Clapier, Ivan Zahradník

1)Institute of Molecular Physiology and Genetics, Slovak Academy of Sciences, Bratislava, Slovak Republic.

Creatine kinase content, isoform distribution, and participation in energy transfer are muscle type specific. We analysed ultrastructural changes in slow muscle fibres of soleus due to invalidation of creatine kinase (CK) to reveal a difference in the remodelling strategy in comparison with fast muscle fibres of gastrocnemius published previously. We have employed the stereological method of vertical sections and electron microscopy of soleus muscles of wild type (WT) and CK-/- mice. The mitochondrial volume density was 1.4× higher but that of sarcoplasmic reticulum (SR) was almost 5× lower in slow CK-/- muscles fibres than in WT fibres. The volume density of terminal cisterns and of t-tubules was also lower in CK-/- than in WT fibres. The analysis of organelle environment revealed increased neighbourhood of mitochondria and A-bands that resulted from the decreased volume density of SR, from relocation of mitochondria along myofibrils, and from intrusion of mitochondria to myofibrils. These processes direct ATP supply closer to the contractile machinery. The decreased interaction between mitochondria and SR suggests reduced dependence of calcium uptake on oxidative ATP production. In conclusion, the architecture of skeletal muscle cells is under control of a cellular program that optimizes energy utilization specifically for a given muscle type.

General Physiology and Biophysics. Volume 35, 2016, No. 4: 477-486.

  Effects of exposure to extremely low-frequency electromagnetic fields on the differentiation of Th17 T cells and regulatory T cells
Yun-Jung Lee 1), , Kyeong Eun Hyung, Jong-Sun Yoo, Ye Won Jang, Soo Jeong Kim, Do Ik Lee, Sang Joon Lee, So-young Park, Ji Hoon Jeong, Kwang Woo Hwang

1)Laboratory of Host Defense Modulation, College of Pharmacy, Chung-Ang University, Seoul 06974, Korea.

The potential risks that electromagnetic fields (EMF) pose to human physiology have been debated for several decades, especially considering that EMF is almost omnipresent and some occupations involve regular exposure to particularly strong fields. In the present study, the effects of 60 Hz 0.3 mT EMF on CD4+ T cells were evaluated. Production of T cell related cytokines, IFN-γ and IL-2, was not altered in CD4+ T cells that were exposed to EMF, and cell proliferation was also unaffected. The expression of genes present in a subset of Th17 cells was upregulated following EMF exposure, and the production of effector cytokines of the IL-17A subset also increased. To determine signaling pathways that underlie these effects, phosphorylation of STAT3 and SMAD3, downstream molecules of cytokines critical for Th17 induction, was analyzed. Increased SMAD3 phosphorylation level in cells exposed to EMF, suggesting that SMAD3 may be at least in part causing the increased Th17 cell production. Differentiation of Treg, another CD4+ T cell subset induced by SMAD3 signaling, was also elevated following EMF exposure. These results suggest that 60 Hz 0.3 mT EMF exposure amplifies TGF-β signaling and increases the generation of specific T cell subsets.

General Physiology and Biophysics. Volume 35, 2016, No. 4: 487-495.

  The expression of P-gp in leukemia cells is associated with cross-resistance to protein N-glycosylation inhibitor tunicamycin
Lucia Pavlikova 1), Mário Šereš, Denisa Imrichova, Milan Hano, Andrej Rusnak, Martina Zamorova, Jaroslav Katrlik, Albert Breier, Zdena Sulová

1)Institute of Molecular Physiology and Genetics, Slovak Academy of Sciences, Dubravska cesta 9, 840 05 Bratislava, Slovak Republic.

In P-gp-positive cell variants obtained from L1210 cells either by selection with vincristine (L1210/R) or by transfection with the human gene encoding P-gp (L1210/T), we have previously described cross-resistance to tunicamycin (TNM), a protein N-glycosylation inhibitor. Here we studied whether this cross-resistance also underlies P-gp-positive variants of human acute myeloid leukemia cells (AML) derived from SKM-1 and MOLM-13 cells (SKM-1/VCR, SKM-1/LEN, MOLM-13/VCR) by selection with vincristine (VCR) and lenalidomide (LEN). While SKM-1/LEN cells were P-gp positive, no P-gp was detected in MOLM-13/LEN cells. P-gp-positive cells could be repeatedly passaged in medium containing TNM. In contrast, more than 90% of P-gp-negative cells were entering and progressing through cell death mechanisms after the third passage in medium containing TNM. Combined apoptosis/necrosis cell death was detected in L1210 cells after exposure to TNM. Passaging of P-gp-negative AML cells in medium containing TNM induced preferentially apoptosis. Damage to P-gp-negative cells induced with TNM was associated with arrest in the G1 phase of the cell cycle. P-gp-positive leukemia cells differed from P-gp-negative cells in the composition of plasma membrane glycoproteins, which we monitored with the aid of different lectins. The application of TNM to cells induced additional changes in membrane-linked glycosides.

General Physiology and Biophysics. Volume 35, 2016, No. 4: 497-510.

  Inhibition of VEGF mediated post receptor signalling pathways by recently developed tyrosine kinase inhibitor in comparison with sunitinib
Roman Moravčík 1), Katarína Stebelová, Andrej Boháč, Michal Zeman

1)Department of Animal Physiology and Ethology, Faculty of Natural Sciences, Comenius University in Bratislava, Mlynská dolina, Ilkovičova 6, 842 15 Bratislava, Slovak Republic.

Inhibition of angiogenesis involves blocking of tyrosine kinases (TK) implicated in signalling of vascular endothelial growth factor receptors (VEFGR). The inhibition of TK results in a disruption of Ras/Raf/MEK/ERK1/2 and PI3K/Akt signalling pathways. We evaluated recently developed TK inhibitor 22SYM and compared its anti-angiogenic effects with an approved multitargeted TK inhibitor sunitinib L-malate (sunitinib). Both compounds significantly inhibited migration and proliferation of human umbilical vein endothelial cells and ERK1/2 and Akt phosphorylation induced by VEGF. The lower inhibitory activity of 22SYM probably reflects its lower bioavailability and higher specific binding to VEGFR2 TK, which may decrease its potential side effects and toxicity in comparison with sunitinib.

General Physiology and Biophysics. Volume 35, 2016, No. 4: 511-514.