@misc{Skórkowska_Alicja_Evaluation_2024,
 author={Skórkowska, Alicja},
 address={Kraków},
 howpublished={online},
 year={2024},
 school={Rada Dyscypliny Nauki farmaceutyczne},
 language={pol},
 abstract={Brain stroke is the second (after heart attack) leading cause of death and the most common cause of disability in adult. The vast majority of stroke cases are caused by arterial occlusion and classified as ischemic stroke. An intravenous thrombolysis with the recombinant tissue plasminogen activator (rt−PA) is the only available pharmacotherapy, additionally limited due to numerous exclusion criteria, thus, only a small percentage of patients receive this treatment. It clearly indicates the need to thoroughly understand the molecular pathomechanism of brain ischemia and to develop new treatment strategies. Hydrogen sulfide (H2S), a gaseous signaling molecule and an endogenous neuromodulator, seems to be a promising agent in the treatment of ischemic stroke. Disturbances in endogenous H2S production have been observed in many neurological diseases, e.g. stroke and depressive disorders (also common in patients suffering from cerebral ischemia), indicating potential therapeutic application of this molecule. However, the administration of gaseous H2S and determining a safe and effective dose for each patient is a very difficult task. An interesting solution to avoid these limitations is the use of the novel, stable and slow-releasing H2S, AP39 compound. It is a mitochondria-targeted H2S donor, which is especially important, because mitochondrial dysfunction is crucial in neurodegen},
 abstract={eration after stroke and restoration of mitochondria function may be a key factor in the prevention of stroke progression. Since ischemic stroke is characterized by a cascade of interrelated processes with the endpoint in neuronal cell death, the aim of the study was to investigate the effect of AP39 on main components of stroke pathomechanism, such as excitotoxicity, disturbances of cellular metabolism and microglia activation. An in vivo model of brain ischemia in rats (middle cerebral artery occlusion, MCAO) as well as in vitro models: oxygen-glucose deprivation (OGD)/reoxygenation in HT-22 neuronal cell line and lipopolysaccharide (LPS)-activated primary cultures of microglia cells from brain cortex were used in the study. Transient 90-min MCAO was used to induce focal cerebral ischemia in Sprague Dawley rats. AP39 was administered in a single dose of 100 nmol/kg b.w. i.v., 10 min after restoration of blood flow or performing sham surgery. Neurological deficit by Phillips’s score and infarct volume in 2,3,5-triphenyltetrazolium chloride (TTC) stainig were evaluated. Liquid chromatography − mass spectrometry was used to determine the extracellular glutamate level in brain microdialysates collected intrasurgically and from freely moving animals 24 hours and 3 days after reperfusion. Brain tissues were isolated 24 hours after reperfusion. The protein level of glutamate transpo},
 abstract={rters, N-methyl-D-aspartate receptor subunits, key enzymes of glycolysis, Krebs cycle, mitochondrial respiratory chain complexes, sulfide-quinone oxidoreductase and markers of microglia activation were investigated by Western-blot analysis in the cortex, striatum and hippocampus of ispilateral hemisphere. Coronal cryosections of brain were immunostained with antibodies against glutamate transporters and visualized by confocal microscopy. HT-22 neuronal cells were subjected to OGD procedure to mimic ischemic conditions. AP39 (10 nM − 250 nM) was added to the culture medium during OGD (21 hours) and reoxygenation (3 hours) periods. The control group was represented by HT-22 cells cultured under standard conditions (normoxia and media containing glucose). Cell viability in MTT reduction test as well as the level of extracellular lactate dehydrogenase (LDH) were determined. Fluorescent dye JC-1 was used to estimate the mitochondrial membrane potential. The process of oxidative phosphorylation was determined by the measurement of oxygen consumption rate using Seahorse-based assays. Primary cultures of microglia were prepared from the cortex of 1-2−day old Sprague Dawley rat pups and isolated from other glial cells by shaking method. Cells were incubated with AP39 (10 nM − 250 nM) for 24 hours and then subjected to LPS (50 ng/ml) stimulation for another 24 hours. Cell viability in MT},
 abstract={T reduction test, nitrite level and concentration of pro- and anti-inflammatory cytokines in culture medium were estimated. The following effects of AP39 were shown: 1) significantly decreased neurological deficit and infarct volume after MCAO in AP39-treated rats; 2) a significant and longitudinal decrease of extracellular glutamate concentration in MCAO+AP39 group, comparing to MCAO group; 3) increased level of GLT-1 transporter responsible for glutamate reuptake from the synaptic cleft into astrocytes; 4) decreased level of VGLUT1 transporter, which regulates glutamate transport to synaptic vesicles; 5) increased level of pyruvate dehydrogenase, the key enzyme linking the process of glycolysis and Krebs cycle, in the striatum of MCAO+AP39 group, comparing to MCAO group; 6) enhanced ATP production in neuronal cells subjected to OGD/reoxygenation in the presence of AP39; 7) decreased level of CD86, the marker of clasically activated microglia, associated with pro-inflammatory responses, in MCAO+AP39 group; 8) decrease of the LPS-stimulated release of pro-inflammatory cytokines in primary cultures of microglia in the presence of AP39. To conclude, the obtained results indicate that AP39 may activate the mechanisms counteracting excitotoxicity, metabolic disturbances and microglia activation, processes following ischemia-reperfusion injury in the brain.},
 title={Evaluation of the neuroprotective effect of the H2S donor, compound AP39 in a model of ischemic stroke : in vivo and in vitro studies},
 type={Praca doktorska},
 keywords={ischemic stroke, hydrogen sulfide donor, AP39 compound, excitotoxicity},
}