The effect of three therapeutically used drugs and five polyphenolic compounds on the mechanism of oxidative burst was compared in whole blood and isolated neutrophils at cellular and molecular level. In 10 μM concentration, the compounds investigated decreased the oxidative burst of whole blood in the rank order of potency: N-feruloylserotonin (N-f-5HT) > curcumin (CUR) > quercetin (QUER) > arbutin (ARB) > resveratrol (RES) > dithiaden (DIT) > carvedilol (CARV) > brompheniramine (BPA). The ratio between the percentage inhibition of extracellular versus intracellular chemiluminescence (CL) followed the rank order QUER > N-f-5HT > RES > CUR > DIT and is indicative of the positive effect of the compounds tested against oxidative burst of neutrophils, demonstrating suppression of reactive oxygen species extracellularly with minimal alteration of intracellular reactive oxygen species (ROS). Activation of protein kinase C was significantly decreased by DIT, CUR, QUER and N-f-5HT. CARV, DIT, QUER and ARB reduced activated neutrophil myeloperoxidase release more significantly compared with the effect on superoxide anion generation. All compounds tested increased the activity of caspase-3 in cell-free system. It is suggested that other regulatory mechanisms than protein kinase C might participate in the inhibition of neutrophil activation with the compounds tested. Different mechanisms are concerned in controlling the assembly of NADPH oxidase and the regulatory role of calcium ions is suggested. Compounds decreasing the amount of extracellular ROS generation, yet affecting but minimally intracellular ROS generation, are promising for further investigation in vivo., R. Nosáľ, K. Drábiková, V. Jančinová, T. Mačičková, J. Pečivová, T. Perečko, J. Harmatha, J. Šmidrkal., and Obsahuje bibliografii
Autophagy and poly(ADP-ribose) polymerase 1 (PARP-1) are activated and involved in a series of cell processes under oxidative stress, which is associated with pathogenesis of atherosclerosis. Research on their relationship under oxidative stress has been limited. In this study, we aimed to investigate the activation, relationship, and role of autophagy and PARP-1 in vascular smooth muscle cell (VSMC) death under oxidative stress. This study ex-plored the signal molecule PARP-1 and autophagy in VSMCs using gene silencing and the hydrogen peroxide (H2O2)-stimulated oxidative stress model. We observed that H2O2 could induce autophagy in VSMCs, and the inhibition of autophagy could protect VSMCs against oxidative stress-mediated cell death. Meanwhile, PARP-1 could also be activated by H2O2. Additionally, we analysed the regulatory role of PARP-1 in oxidative stress-mediated autophagy and found that PARP-1 was a novel factor involved in the H2O2-induced autophagy via the AMPK-mTOR pathway. Finally, PARP-1 inhibition protected VSMCs against caspase-dependent apoptosis. These data suggested that PARP-1 played a critical role in H2O2-mediated autophagy and both of them were involved in apoptosis of VSMCs. and Corresponding author: Guoxian Qi
Although statins exert non-lipid cardioprotective effects, their influence on cell death is not fully elucidated. For this purpose, we investigated whether simvastatin treatment (S, 10 mg/kg, 5 days) is capable of mitigating ischemia/reperfusion-induced (IR) apoptosis in the isolated rat hearts, which was examined using immunoblotting analysis. In addition, the content of signal transducer and activator of transcription 3 (STAT3) and its active form, phosphorylated STAT3 (pSTAT3-Thr705), was analyzed. Simvastatin induced neither variations in the plasma lipid levels nor alterations in the baseline content of analysed proteins with the exception of upregulation of cytochrome C. Furthermore, simvastatin significantly increased the baseline levels of pSTAT3 in contrast to the control group. In the IR hearts, simvastatin reduced the expression of Bax and non-cleaved caspase-3. In these hearts, phosphorylation of STAT3 did not differ in comparison to the non-treated IR group, however total STAT3 content was slightly increased. The improved recovery of left ventricular developed pressure co-existed with the increased Bcl- 2/Bax ratio. In conclusion, pleiotropic action of statins may ameliorate viability of cardiomyocytes by favouring the expression of anti-apoptotic Bcl-2 and downregulating the proapoptotic markers; however STAT3 does not seem to be a dominant regulator of this anti-apoptotic action of simvastatin., T. Rajtík, ... [et al.]., and Obsahuje seznam literatury
Protein kinases, transcription factors and other apoptosis- and proliferation-related proteins can regulate reproduction, but their involvement in sexual maturation remains to be elucidated. The general aim of the in vivo and in vitro experiments with porcine ovarian granulosa cells was to identify possible intracellular regulators of female sexual maturation. For this purpose, proliferation (expression of proliferating cell nuclear antigen - PCNA, mitogen-activated protein kinases - ERK 1,2 related MAPK and cyclin B1), apoptosis (expression of the apoptotic protein Bax and apoptosis regulator Bcl-2 protein), expression of some protein kinases (cAMP dependent protein kinase - PKA, cGMPdependent protein kinase - PKG, tyrosine kinase - TK) and cAMP responsive element binding protein 1 (CREB-1) was examined in granulosa cells isolated from ovaries of immature and mature gilts. Expression of PCNA, ERK1,2 related MAPK, cyclin B1, Bcl-2, Bax, PKA, CREB-1, TK and PKG in porcine granulosa cells were detected by immunocytochemistry. Sexual maturation was associated with significant increase in the expression of Bcl-2, Bax, PKA, CREB-1 and TK and with decrease in the expression of ERK1,2 related MAPK, cyclin B1 and PKG in granulosa cells. No significant difference in PCNA expression was noted. The present data obtained from in vitro study indicate that sexual maturation in females is influenced by puberty-related changes in porcine ovarian signaling substances: increase in Bcl-2, Bax, PKA, CREB-1, TK and decrease in ERK1,2 related MAPK, cyclin B1 and PKG. It suggests that these signaling molecules could be potential regulators of porcine sexual maturation., A. Kolesarova, A. V. Sirotkin, M. Mellen, S. Roychoudhury., and Obsahuje bibliografii
The action of the medicinal plant Tribulus terrestris (TT) on bovine ovarian cell functions, as well as the protective potential of TT against xylene (X) action, remain unknown. The aim of the present in vitro study was to elucidate the influence of TT, X and their combination on basic bovine ovarian cell functions. For this purpose, we examined the effect of TT (at doses of 0, 1, 10, and 100 ng/ml), X (at 20 μg/ml) and the combination of TT + X (at these doses) on proliferation, apoptosis and hormone release by cultured bovine ovarian granulosa cells. Markers of proliferation (accumulation of PCNA), apoptosis (accumulation of Bax) and the release of hormones (progesterone, testosterone and insulin-like growth factor I, IGF-I) were analyzed by quantitative immunocytochemistry and RIA, respectively. TT addition was able to stimulate proliferation and testosterone release and inhibit apoptosis and progesterone output. The addition of X alone stimulated proliferation, apoptosis and IGF-I release and inhibited progesterone and testosterone release by ovarian cells. TT was able to modify X effects: it prevented the antiproliferative effect of X, induced the proapoptotic action of X, and promoted X action on progesterone but not testosterone or IGF-I release. Taken together, our observations represent the first demonstration that TT can be a promoter of ovarian cell functions (a stimulator of proliferation and a suppressor of apoptosis) and a regulator of ovarian steroidogenesis. X can increase ovarian cell proliferation and IGF-I release and inhibit ovarian steroidogenesis. These effects could explain its antireproductive and cancer actions. The ability of TT to modify X action on proliferation and apoptosis indicates that TT might be a natural protector against some ovarian cell disorders associated with X action on proliferation and apoptosis, but it can also promote its adverse effects on progesterone release.
Excessive production of oxygen free radicals has been regarded as a causative common denominator of many pathological processes in the animal kingdom. Hydroxyl and nitrosyl radicals represent the major cause of the destruction of biomolecules either by a direct reaction or by triggering a chain reaction of free radicals. Scavenging of free radicals may act preventively or therapeutically. A number of substances that preferentially react with free radicals can serve as scavengers, thus increasing the internal capacity/activity of endogenous antioxidants and protecting cells and tissues against oxidative damage. Molecular hydrogen (H2) reacts with strong oxidants, such as hydroxyl and nitrosyl radicals, in the cells, that enables utilization of its potential for preventive and therapeutic applications. H2 rapidly diffuses into tissues and cells without affecting metabolic redox reactions and signaling reactive species. H2 reduces oxidative stress also by regulating gene expression, and functions as an anti-inflammatory and anti-apoptotic agent. There is a growing body of evidence based on the results of animal experiments and clinical observations that H2 may represent an effective antioxidant for the prevention of oxidative stress-related diseases. Application of molecular hydrogen in situations with excessive production of free radicals, in particular, hydroxyl and nitrosyl radicals is relatively simple and effective, therefore, it deserves special attention., J. Slezák, B. Kura, K. Frimmel, M. Zálešák, T. Ravingerová, C. Viczenczová, Ľ. Okruhlicová, N. Tribulová., and Obsahuje bibliografii
Pyrogallol is a polyphenol that generates the superoxide anion. In this study, we investigated the influence of pyrogallol on human platelets. Our data showed that exposure of platelets to pyrogallol induced numerous manifestations of apoptosis including depolarization of mitochondrial inner membrane and release of cytochrome c from the mitochondria. Pyrogallol also induced downstream extra-mitochondrial apoptotic responses, including activation of caspase-3 and phosphatidylserine exposure on the outer leaflet of the plasma membrane. Addition of glutathione significantly rescued cells from pyrogallol-induced apoptosis, as evidenced by a decrease of all markers of apoptosis. Thus, pyrogallol appears to produce depletion of intracellular glutathione content in platelets, the main non-protein antioxidant in the cells. Furthermore, inhibition of γ-glutamyl trans-peptidase, an enzyme that plays the main role in the cellular supply of glutathione, reverted the glutathione (GSH) protection over platelet apoptosis. Our results indicate that pyrogallol induces apoptosis by suppressing the natural anti-oxidation in human platelets. and Corresponding author: Mercedes López Zambrano
Flavonoids, polyphenol derivatives of plant origin, possess a broad range of pharmacological properties. A number of studies have found both pro/anti-apoptotic effects for many of these compounds. For these reasons we investigated whether ProvinolsTM, flavonoids obtained from red wine, have anti-apoptotic properties. The investigations have been carried out in rats treated with Cyclosporine A (CsA). In particular, four groups of rats have been treated for 21 days with either olive oil (control group), with CsA, with ProvinolsTM, or with CsA and ProvinolsTM simultaneously. Oxidative stress, systolic blood pressure, body weight, biochemical parameters and different markers of pro/anti-apoptotic pathway were measured. CsA produced an increase of systolic blood pressure, a decrease in body weight, serum creatinine levels, urinary total protein concentration and creatinine clearance. Moreover, CsA induced renal alterations and the translocation of Bax and cytochrome c from cytoplasm to mitochondria and vice versa. These changes activated the caspase cascade pathway, that leads to morphological and biochemical features of apoptosis. ProvinolsTM restored morphological and biochemical alterations and prevented nephrotoxicity. In conclusion, this study may augment our current understanding of the controversial pro-/anti-apoptotic properties of flavonoids and their molecular mechanisms., R. Rezzani ... [et al.]., and Obsahuje seznam literatury
Damage of alveolar-capillary barrier, inflammation, oxidative
injury, and lung cell apoptosis represent the key features of acute
lung injury (ALI). This study evaluated if selective
phosphodiesterase (PDE)-4 inhibitor roflumilast can reduce the
mentioned changes in lavage-induced model of ALI. Rabbits with
ALI were divided into 2 groups: ALI without therapy (A group)
and ALI treated with roflumilast i.v. (1 mg/kg; A+R group). One
group of healthy animals without ALI served as ventilated
controls (C group). All animals were oxygen-ventilated for further
4 h. At the end of experiment, total and differential counts of
cells in bronchoalveolar lavage fluid (BALF) and total and
differential counts of white blood cells were estimated. Lung
edema formation was assessed from determination of protein
content in BALF. Pro-inflammatory cytokines (TNFα, IL-6 and
IL-8) and markers of oxidation (3-nitrotyrosine, thiobarbituricacid reactive substances) were detected in the lung tissue and
plasma. Apoptosis of lung cells was investigated
immunohistochemically. Treatment with roflumilast reduced leak
of cells, particularly of neutrophils, into the lung, decreased
concentrations of cytokines and oxidative products in the lung
and plasma, and reduced lung cell apoptosis and edema
formation. Concluding, PDE4 inhibitor roflumilast showed potent
anti-inflammatory actions in this model of ALI.
Coordinated regulation of apoptosis is critical for development, homeostasis, and immunity in larvae of Metazoa. We determined the full nucleotide sequence of an inhibitor of an apoptosis protein in a lepidopteran insect Mythimna separata (Walker) (MsIAP) and carried out functional analyses of the MsIAP. The full-length cDNA of MsIAP was 1642 bp, which encoded 379 amino acid residues with a calculated molecular mass of 41,834 Da, and two BIR domains and one RING domain revealed using amino acid sequence analysis. In addition, the sequences of these domains were similar to Drosophila IAP1 and those of some other lepidopteran insects. We carried out a functional analyses of MsIAP related to apoptosis regulation using RNA interference. The effects of MsIAP knockdown on adhering hemocytes and non-adhering hemocytes as controls were examined using Hoechst33342/propidium iodide staining, effector caspase activity and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL) staining. A significantly higher number of propidium iodide and TUNEL-positive cells was recorded in adhering hemocytes from MsIAP knockdown larvae than from control larvae, but these differences were not recorded for non-adhering hemocytes. However, higher effector caspase activity was detected in both adhering and non-adhering hemocytes from MsIAP knockdown larvae compared to that in control larvae. These results indicate that the knockdown of MsIAP induces apoptosis in larval adhering hemocytes, which MsIAP negatively and non-redundantly regulate apoptosis, and that IAP function is conserved in M. separata and other insect species including Drosophila and several lepidopteran insects., Masashi Kamezaki, Kakeru Yokoi, Ken Miura., and Obsahuje bibliografii