Modulation of apoptotic signaling pathways in brain cells of adult male rats after chronic dexamethasone treatment

Deksametazon, jak sintetski glukokortikoid, se dugi niz godina koristi kao lek u tretmanu različitih bolesti poput psorijaze, adrenalne insuficijencije, bakterijalnog meningitisa, moždanih trauma, šloga, alergija, spazma bronhija, reumatidnog artritisa, itd; ali i kao pomoćni lek u hemo- i radioterapiji. Brojne studije ukazuju da deksametazon može regulisati sinaptičku plastičnost, povećati vijabilnost ćelija i njihovu proliferaciju u in vivo i in vitro uslovima. Međutim, uprkos širokoj primeni u terapijske svrhe primećeno je da deksametazon ispoljava i niz negativnih efekata u mozgu, na primer apoptozu u granularnom sloju dentatnog girusa hipokampusa kod mladih i starih pacova, kao i smanjenje kognitivnih funkcija i motornog razvoja. Usled neusklađenosti rezultata mnogih studija, proučavanje sistemskih efekata, kao i efekata deksametazona u mozgu predstavljaju interesantno polje istraživanja prvenstveno jer se efekti niskih doza ovog sintetskog glukokortikoida ne mogu ispitivati centralno usled blokiranja njegovog ulaska dejstvom MDR p-glikoproteina i/ili nekim drugim mehanizmom. Stoga, za potrebe eksperimenta, odrasli mužjaci pacova Wistar soja su podeljeni u dve grupe– kontrolne jedinke i životinje tretirane deksametazonom (100 g/kg/dan) tokom 7 uzastopnih dana. 28 h nakon završetka hroničnog tretmana životinje su žrtvovane. Sistemski efekat deksametazona je praćen promenom biometrijskih parametara (telesna masa, masa timusa i nadbubrežnih žlezdi), kao i koncentracije kortikosterona u serumu i moždanom tkivu. Kako bi se utvrdilo da li male doze deksametazona dovode do apoptoze u hipofizi, hipotalamusu, hipokampusu i prečeonoj kori korišćen je DNK fragmentacioni esej, dok se metodama histološkog bojenja hipokampusa i prečeone kore (fluoro-jade B i krezil ljubičastim) istraživao obim neuralne smrti i morfološke promene. Western blot i RT-PCR analizama su ispitivane promene u ekspresiji proteina i iRNK markera procesa smrti odnosno preživljavanja ćelija. Rezultati ove studije su pokazali da hroničan tretman malim dozama deksametazona uzrokuje hipoaktivnost hipotalamo-hipofizno-adrenalne ose, koja se ogleda u smanjenju telesne mase, mase timusa i nadbubrežnih žlezdi, kao i kortikosterona u serumu...For many years, dexamethasone, a potent synthetic glucocorticoid, has been used as a medication in the treatment of psoriasis, adrenal insufficiency, bacterial meningitis, brain trauma, stroke, allergies, bronchial spasm, rheumatoid arthritis, etc., and as a co-medicament in chemo- and radiotherapy. Numerous studies suggest that dexamethasone is able to regulate synaptic plasticity, enhance cell viability and proliferation in vivo and in vitro. However, dexamethasone exerts a number of adverse reactions in the brain, such as apoptosis in the hippocampal granular layer of dentate gyrus in young and old rats, as well as reduced cognitive and motor development. The dexamethasone-induced systemic effects and dexamethasone-provoked effects in the brain are an interesting field of research, mainly because the effects of low-dose dexamethasone treatment could not be tested directly in brain tissue due to the central blocking action of MDR p-glycoprotein and/or some other mechanism. Therefore, for the purposes of the experiment, adult male Wistar rats were divided into two groups – controls and animals treated with dexamethasone (100 g/kg/day) per 7 days. 28 h upon chronic treatment, rats from both groups were sacrificed. Late systemic effects of dexamethasone were monitored by alterations of biometric parameters (body weight, thymus and adrenal glands mass) and level of corticosterone in serum and brain tissue. Further, using DNA fragmentation assay, present study aimed to determine whether low dose dexamethasone treatment is able to cause apoptosis in the pituitary gland, hypothalamus, hippocampus and prefrontal cortex, while histological staining methods (fluoro-jade B and cresyl violet staining) were applied to investigated the extent of neuronal death and morphological changes in hippocampus and prefrontal cortex. Changes in protein and mRNA expression of cell death and cell survival markers were examined by Western blot and RT-PCR analyzes. The results obtained in this thesis revealed that chronic low dose dexamethasone treatment caused hypoactivity of hypothalamus-pituitary-adrenal axis, reflected in the reduction of body weight, thymus and adrenal glands masses, as well as levels of corticosterone in serum...

Ionizing irradiation affect extracellular nucleotide hydrolysis in brain of rats in different stages of development: ii 30-day-old rats

The effect of acute gamma irradiation (IR) on enzyme activity of rat brain Ecto-Nucleotide Diphosphohydrolase (E-NTPDase), in presence of adenosine triand diphophashates (ATP and ADP) and divalent cations (Ca2+ and Mg2+), has been investigated. The aim of research was to study the influence of low (50 cGy) and therapeutic (2Gy) doses of whole-body irradiation on rat brain E-NTPDase enzyme activity 24h after treatment in prepubertal and adult rats. Our results suggest that whole-body irradiation could induce modulation of neural activity in rat brain, especially in young rats.Physical chemistry 2008 : 9th international conference on fundamental and applied aspects of physical chemistry; Belgrade (Serbia); 24-28 September 200

Time-dependent expression of Bcl-2 and Bax proteins in cortical brain area of adult wistar rats after permanent bilateral occlusions

Model of permanent bilateral occlusion of common carotid arteries (2VO) is generally used to investigate mechanisms of chronic cerebral hypoperfusion that occurs in aging and other neurodegenerative processes. The aim of this study was to determine timedependent modulation of mitochondrial apoptotic signaling in cortical brain area following chronic cerebral hypoperfusion. Using Western blot technique we monitored the changes in the expression of proteins of Bcl-2 family (Bcl-2, Bax) 3, 7 and 90 days following the insult. According to our results the greatest impact of chronic cerebral hipoperfusion occurred on 7th day.Physical chemistry 2012 : 11th international conference on fundamental and applied aspects of physical chemistry; Belgrade (Serbia); 24-28 September 201

L-cysteine modulates the ecto-atpase activity inhibition in presence of cadmium (ii) and mercury (ii) ions

L-cysteine is used as effective oral chelating agent due to property of its sulfhydryl group to bind heavy metal ions. The aim of this study was to investigate ability of L-cysteine to prevent mercury (II) and cadmium (II) induced ecto-ATPase activity inhibition of rat uterus plasma membranes. Results show that 10 mmol/l L-cysteine have protective effect on enzyme activity in the presence of cadmium and mercury ions

Time-Related Sex Diffrences in Cerebral Hypoperfusion-Induced Brain Injury

Although the model of cerebral hypoperfusion in rats has been a matter of many investigations over the years, the exact intracellular and biochemical mechanisms that lead to neuron loss and memory decline have not been clearly identified. In the current study, we examined whether cerebral hypoperfusion causes changes in hippocampal protein expression of apoptotic markers in the synaptosomal fraction and neurodegeneration in a time-dependent and sex-specific manner. Adult male and female Wistar rats were divided into two main groups, controls that underwent sham operation, and animals subjected to permanent bilateral occlusion of common carotid arteries. Both male and female rats were killed 3, 7 or 90 days following the insult. The obtained results indicate that the peak of processes that lead to apoptosis occured on postoperative day 7 and that they were more prominent in males, indicating that neuroprotective effects of certain substances (planned for future experiments), should be tested at this time point

Toxicity assessment of Gentiana lutea L. root extract and its monoterpene compounds

Root of Gentiana lutea commercially available as gentian root, a natural antidote for different types of poisons, possess antioxidative, immunomodulatory, cytoprotective and anti-inflammatory, and adverse, genotoxic and mutagenic effects. It has monoterpenes loganic acid, swertiamarin, gentiopicroside and sweroside as most abundant constituents. In this study, we assessed the toxicity of monoterpenes’ reactive molecular fragments using in silico prediction by VEGA-QSAR platform. Further, we compared the data obtained with in vitro geno- and cyto- toxicity testing of the above monoterpenes and the G. lutea root extract (GE), on human primary unstimulated and mitogen-stimulated peripheral blood mononuclear cells (PBMCs). Viability was assessed by TB and XTT tests after 48 h treatment. DNA damage was evaluated by alkaline comet assay on unstimulated cells, whereas cytokinesis-block micronucleus assay was employed on mitogen-stimulated PBMCs. Stability of compounds throughout treatment was monitored by UPLC. The observed in vitro results had highest compliance with in silico IRFMN/ISSCAN-CGX prediction model. Compounds showed high stability during experiment while treatment with single compounds reduced number of viable cells and increased DNA damage. GE treatment had toxic impact on unstimulated PBMCs but no significant genotoxic influence on mitogen-stimulated PBMCs. In summary, the mild GE effect suggests that the complexity of crude GE extract chemical composition may attenuate the toxicity of the tested monoterpenes loganic acid, swertiamarin, gentiopicroside and sweroside

Theoretical and experimental evaluation of K2Br+ and K3Br+ clusters' ionization energies

In current study, a non-stoichiometric bromine-doped potassium K2Br+and K3Br+clusters are generated by combining a Knudsen effusion cell as a chemical reactor with thermal or surface ionization,and selected by a magnetic sector mass spectrometer. Furthermore, their ionization energies (IEs) are calculated for the first time using B3LYP/9-ve PP(K),cc-pVTZ-PP(Br) level of theory. Herein, presented results indicate that experimentally obtained IEs by Ionov equation, 4.10 ± 0.20 eV for K2Br+, and 4.03 ± 0.20 eV for K3Br+, are in consistence with their theoretically determined IEs.Physical chemistry 2016 : 13th international conference on fundamental and applied aspects of physical chemistry; Belgrade (Serbia); 26-30 September 2016