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Age-dependent protein aggregation is a conserved phenomenon that is associated with many neurodegenerative diseases, including Alzheimer's disease (AD). AD is characterized by aggregation of Tau, a microtubule-binding protein that is normally soluble and mainly localized to neuronal axons, but can form oligomers and higher-order amyloid-like aggregates that accumulate in soma and dendrites and eventually lead to neuronal death. Although the main risk factor for the onset of AD is aging, the exact causes of Tau protein aggregation are largely unclear. To investigate factors that influence Tau protein aggregation, we expressed human Tau protein fused with fluorescent proteins in yeast Saccharomyces cerevisiae. We examined intracellular localization of Tau in young and aged cells, and in cells under different stress conditions, such as glucose starvation, elevated temperature, hyperosmotic and proteotoxic stress caused by toxic amino acid analogue. Furthermore, to study factors affecting Tau oligomerization, we used luminescent reporter NanoBiT in which protein-protein interaction results in the complementation of the luciferase NanoLuc. Results show basal levels of Tau-NanoBiT reporter signal in logarithmically growing wild-type cells, suggesting that Tau oligomerization doesn't occur under normal growth conditions.

As resident central nervous system (CNS) immune cells, microglia play an important role of active sensor and adaptable effector cells both in healthy brain and pathologic conditions, one of them being cerebral ischemia. Cerebral ischemia is followed by neuroinflammatory response initiated in ischemic penumbra – a metabolically active but neurophysiologically silent region around the infarct core. Part of this response includes the activation of microglia that along with macrophages first infiltrate penumbra where, depending on the surrounding conditions, this activation can lead to protection or exhibit detrimental effects. Direct effect of ischemia on microglia response can be examined in oxygen- glucose deprivation (OGD) model in vitro. What triggers the transformation of microglia from resting to activated state is still unclear. Therefore, we used BV-2 microglial cell line to examine the effect of dimethyoxalylglycine (DMOG), a prolyl hydroxylase inhibitor, and Necrostatin 1 (Nec-1), a RIP1 kinase inhibitor, on the response of ramified i.e. resting BV-2 cells to OGD followed by reoxygenation. 24h prior and during the cells’ exposure to OGD, they were grown in complete RPMI supplemented with 0, 5% FBS, to support ramified microglial phenotype. Either DMOG (10M) or Nec-1 (40M) were present during 8h of oxygen deprivation. Cell viability as well as the presence of apoptotic cells were followed after 16hr of reoxygenation period by MTS and Annexin V/Propidium iodide assay, respectively. Furthermore, changes in the activation of caspase 3 and RIP1K protein level in OGD exposed cells were examined by Western blot. While DMOG, protected BV-2 cells from dying as revealed by MTS assay, Nec-1 exhibited the opposite effect. Flow cytometric analysis has shown that DMOG protective effect is based primarily on reduction in the number of early apoptotic cells while Nec-1 increases their number. Also, DMOG reduced caspase 3 activity but did not affect RIP1 kinase protein level in OGD exposed microglia. The obtained data indicate that these two important enzymatic regulators of cell’s death could have a role in regulation of microglia’s response to OGD.

Age-dependent protein aggregation is a conserved phenomenon that is associated with many neurodegenerative diseases, including Alzheimer's disease (AD). AD is characterized by aggregation of Tau, a microtubule-binding protein that is normally soluble and mainly localized to neuronal axons, but which can form oligomers and higher order amyloid-like aggregates that accumulate in soma and dendrites and eventually lead to neuronal death. Although the main risk factor for the onset of AD is aging, the exact causes of Tau protein aggregation are still largely unclear. To investigate factors that influence Tau protein aggregation, we expressed human Tau protein fused with fluorescent proteins in yeast Saccharomyces cerevisiae. We examined the intracellular localization of Tau in young and aged cells, and in cells under different stress conditions, such as glucose starvation, hyperosmotic stress, elevated temperature and proteotoxic stress caused by a toxic amino acid analogue. Furthermore, to study the factors affecting Tau oligomerization, which is considered to be an early step in Tau pathology, we used luminescent reporter NanoBiT in which protein-protein interaction results in the complementation of the luciferase NanoLuc. Our results show basal levels of Tau-NanoBiT reporter signal in logarithmically growing wild-type cells, suggesting that Tau oligomerization does not occur under normal growth conditions.

Age-dependent protein aggregation is a conserved phenomenon that is associated with many neurodegenerative diseases, including Alzheimer's disease (AD). AD is characterized by aggregation of Tau, a microtubule-binding protein that is normally soluble and mainly localized to neuronal axons, but which can form oligomers and higher-order amyloid-like aggregates that accumulate in soma and dendrites and eventually lead to neuronal death. Although the main risk factor for the onset of AD is aging, the exact causes of tau protein aggregation are still largely unclear. To investigate factors that influence tau protein aggregation, we expressed human tau protein fused with fluorescent proteins in yeast Saccharomyces cerevisiae. We examined its intracellular localization in young, logarithmically growing cells, in chronologically aged cells, and under different stress conditions, such as glucose starvation, hyperosmotic stress, elevated temperature, and proteotoxic stress caused by a toxic amino acid analog. Furthermore, to study the factors affecting Tau oligomerization, which is considered to be an early step in Tau pathology, we used luminescent reporter NanoBiT in which protein-protein interaction results in the complementation of the luciferase NanoLuc. Our results show basal levels of Tau-NanoBiT reporter signal in logarithmically growing wild-type cells, suggesting that Tau oligomerization does not occur under normal growth conditions.

U sklopu diplomskog rada razvijena je metoda sinteze halogeniranih derivata marinoaziridina. Strukturni derivati 8a-8h pripravljeni su reakcijom različito supstituiranih N-tosil imina 7a-7h i akiralne 2-kinolinonske sulfonijeve soli 6 u prisustvu baze, kao smjese cis i trans izomera u dobrom iskorištenju. Ispitan je utjecaj baze, temperature i iminskog supstituenta na iskorištenje i dijastereoselektivnost reakcije. Uočeno je da baza nema bitan utjecaj na dijastereoselektivnost aziridinacije. Dijastereoselektivnost reakcije je različita i ovisna je o iminskom supstituentu. U većini slučajeva prevladava trans izomer, dok je neobična cis dijastereoselektivnost uočena kod derivata 8d. U drugom dijelu istraživanja ispitani su uvjeti odjeljivanja enantiomera novosintetiziranih spojeva na polisaharidnim kiralnim nepokretnim fazama temeljenim na derivatima amiloze i celuloze primjenom visokodjelotvorne tekućinske kromatografije. Chiral ART Cellulose SC nepokretna faza sa selektorom tris-(3,5-diklorfenilkarbamat) celulozom pokazala se najboljim izborom u odjeljivanju enantiomera, ali i u odjeljivanju cis i trans izomera. In this thesis, a method of synthesis of halogenated marinoaziridine derivatives was developed. Structural derivatives 8a-8h were prepared by reaction of variously substituted N-tosyl imines 7a-7h and achiral 2-quinolinone sulfonium salt 6 in the presence of a base, as a mixture of cis and trans isomers in good yield. The effect of base, temperature and imine substituent on the yield and diastereoselectivity of the reaction was investigated. It was observed that the base had no significant effect on the diastereoselectivity of aziridination. The diastereoselectivity of the reaction is different and depends on the imine substituent. In most cases, the trans isomer predominates, while unusual cis diastereoselectivity was observed in derivative 8d. In the second part of this research, the conditions of enantioseparation of newly synthesized compounds on polysaccharide chiral stationary phases based on amylose and cellulose derivatives were investigated using high performance liquid chromatography. Chiral ART Cellulose SC stationary phase with selector cellulose tris-(3,5-dichlorophenylcarbamate) proved to be the best choice in the enantioseparation, but also in the separation of cis and trans isomers.

Marinoaziridines A and B are the first aziridine- containing natural products isolated from Gram- negative bacteria from marine sediment.1 Their absolute configuration is so far unknown as well as their total synthesis. Many aziridine- containing compounds demonstrate very useful pharmacological activity including anticancer, antibacterial, antimicrobial activity, etc. strongly indicating that the presence of the aziridine ring in natural as well as synthetic compounds is essential for such activities.2 Due to known activity as pharmacophore, aziridine moiety attracted considerable attention to medicinal and synthetic organic chemists.3 In conducted research the (±)-halogenated marinoaziridine derivatives 1a-1h were prepared by reaction of variously substituted N-tosyl imines and achiral 2-quinolinone sulfonium salt in the presence of a base, as a mixture of cis and trans isomers in good yield. The diastereoselectivity of the reaction is different and depends on the imine substituent. In most cases, the trans isomer predominates, while unusual cis diastereoselectivity was observed in derivative 1d. In the second part of this research, the conditions of enantioseparation of newly synthesized compounds on polysaccharide chiral stationary phases based on amylose and cellulose derivatives were investigated using high performance liquid chromatography. The obtained results showed that the Chiral ART Cellulose SC stationary phase with selector cellulose tris- (3, 5-dichlorophenylcarbamate) proved to be the best choice in the enantioseparation

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by the accumulation of tau protein aggregates in cells of the affected brain regions and the consequent neuronal death. In healthy neurons, tau is a soluble, intrinsically disordered protein that is mostly bound to the microtubules in the axon, however, in the affected neurons, tau accumulates in the soma and dendrites within amyloid-like aggregates, presumably via an intermediary step involving toxic oligomeric structures. The main risk factor for the onset of AD is aging, however, despite a vast number of studies, the causes of tau protein aggregation and toxicity are still largely unclear. Since the ability of a cell to maintain protein homeostasis decreases with aging, impaired protein quality control pathways are considered a possible factor in the development of AD. Yeast is a single cell eukaryotic organism that is amenable to genetic analysis, and many of its cellular pathways, including protein quality control are evolutionarily conserved. Moreover, quiescent yeast can be used to study processes characteristic for non-dividing cells, such as chronological aging. To investigate factors that influence tau protein aggregation and toxicity, in particular cellular aging and proteotoxic stress, we expressed human tau protein in yeast Saccharomyces cerevisiae. We will present a characterization of human tau protein expressed in quiescent yeast. Acknowledgments: Supported by the Research Cooperability Program of the Croatian Science Foundation funded by the European Union from the European Social Fund under the Operational Program Efficient Human Resources 2014-2020 (grant PZS-2019-02-3610) ; Croatian Science Foundation grants IP-2019-04-3584 and DOK- 01-2018/ European Social Fund ; Centre of Excellence for Basic, Clinical and Translational Neuroscience (project “Experimental and clinical research of hypoxic-ischemic damage in perinatal and adult brain” ; GA KK01.1.1.01.0007 funded by the European Union through the European Regional Development Fund).

20-Hydroxyeicosatetraenoic acid (20-HETE), a potent vasoconstrictor, is a cytochrome P-450 (CYP)-derived metabolite of arachidonic acid. Inhibitors of 20-HETE synthesis and/or actions have been reported to protect brain from ischemic stroke in the rat and primate. The neuroprotective effects were not associated with an increase in cerebral blood flow suggesting that these drugs may enhance the survival of neurons after ischemic injury independent of their effect on cerebral blood flow. In this regard, our previous study has demonstrated that 20-HETE directly promote neuronal injury in organotypic hippocampal slices subjected to oxygen-glucose deprivation (OGD) and that inhibitors of 20-HETE synthesis protect neurons from OGD-induced cell death by decreasing ROS formation and activation of caspase-3. In line with our findings, protective effect of 20-HETE inhibition has also been reported in mouse cortical neurons after OGD. In the present study we used N27 rat dopaminergic neuronal cells subjected to OGD followed by reoxygenation to examine whether 20-HETE contributes to ischemic injury through activation of NADPH oxidase and increased superoxide production. The preliminary results indicate that cell viability after OGD increased after treatment with a 20-HETE synthesis inhibitor or an antagonist. Administration of a 20-HETE mimetic had the opposite effect and increased neuronal cell death after OGD. That effect was reversed by coadministration of a NADPH oxidase inhibitor, apocynin, suggesting that 20- HETE amplifies neuronal cell death by increasing oxidative stress through NADPH oxidase- dependent mechanisms. Further studies will be taken to elucidate the molecular mechanisms underlying the protective effect of 20-HETE inhibitors against OGD-induced neuronal injury.

El objetivo de este trabajo es explicar el fenómeno del tabú lingüístico y analizar el uso de los eufemismos en el lenguaje político. Al inicio se introducirá el origen del tabú, su denominación y sus funciones. Se presentará la expansión del tabú por el mundo y su presencia a lo largo de la historia. Entonces, se explicará el fenómeno de la interdicción lingüística relacionada con el tabú. Se pondrá de relieve el eufemismo como el mecanismo lingüístico más usado para evitar las palabras que provocan el rechazo social. A continuación, se presentarán otros procedimientos eufemísticos que se usan para sustituir el tabú lingüístico, en otras palabras, las palabras prohibidas. Más adelante, se explicará el proceso de la lexicalización de los eufemismos y las causas que lo inician. En este trabajo, también se indicarán los ámbitos sociales en los que se usa el eufemismo como instrumento de manipulación. También se destacará el papel importante que tienen los eufemismos en la formación del lenguaje políticamente correcto. Al final, se presentara el análisis de los eufemismos de diferentes ámbitos sociales dentro del cual se observarán sus funciones. El objetivo del análisis es mostrar diversos efectos que se quieren conseguir con el uso de los eufemismos en diferentes situaciones y revisar si esconden una función manipulativa. Cilj ovog rada je objasniti fenomen jezičnog tabua i analizirati upotrebu eufemizma u političkom jeziku. Prvo će se objasniti podrijetlo tabua, njegov naziv i funkcije. Zatim će se objasniti pojava zabrane tabua u jeziku. Stavit će se naglasak na eufemizam kao jezični mehanizam koji se najčešće koristi kako bi se izbjegle riječi koje izazivaju društveno odbacivanje. Zatim će biti predstavljeni drugi eufemistički postupci koji se koriste za zamjenu jezičnih tabua, odnosno zabranjenih riječi. Kasnije će se objasniti postupak leksikalizacije eufemizama te koji su uzroci koji ga pokreću. U ovom će se radu ukazati i društvena područja u kojima se eufemizam koristi kao instrument manipulacije. Također će biti istaknuta važna uloga eufemizma u formuliranju politički korektnog jezika. Na kraju će se iznijeti analiza eufemizama različitih društvenih područja unutar kojih će se promatrati njihove funkcije. Cilj analize je pokazati različite efekte koje čovjek želi postići korištenjem eufemizama u različitim situacijama i provjeriti krije li se iza njih pokušaj manipulacije.

Medicinska kemija je grana kemije koja uključuje organsku sintezu i farmakologiju za proučavanje i razvoj lijekova u korist čovječanstva. Da bi se postigla što bolja aktivnost i specifičnost lijeka koriste se metode za optimizaciju interakcije između lijeka i njegovog veznog mjesta. Neke od metoda su: promjena supstituenata, proširenje strukture, produljivanje/skraćivanje ugljikovodičnog lanca, produljivanje/skraćivanje prstenova, korištenje izostera, itd. Cimetidin je prvi lijek koji je liječio čir na želudcu, otkriven je sedamdesetih godina prošloga stoljeća. Čirevi uzrokuju nagrizanje sluznice membrane želudca pri čemu se luči želučana kiselina. Lijek je koji djeluje kao blokator na H2 receptor, odnosno antagonist histaminskih receptora na sluznici želudca. Razvoj cimetidina kao lijeka protiv čireva doprinijelo je shvaćanju važnosti pKa u kemiji. Na primjer, prethodnik konačnoj molekuli cimetidina bio je gvaninski analog metiamida. pKa vrijednost gvanidina bila je previsoka, što je poticalo agonistična svojstva molekule, odnosno poticalo lučenje kiseline. Kako bi se postigao antagonistični efekt, pKa vrijednost gvanidina smanjena je adicijom elektron odvlačeće skupine.

U ovom radu nastojalo se pružiti teoretsku podlogu za razumijevanje ključnih pojmova vezanih uz politiku zaštite potrošača i okoliša kao i prikupljanjem sekundarnih podataka iz dostupne literature i prethodno provedenih istraživanja te primarnih podataka iz vlastitog istraživanja, doprinijeti stvaranju jasnije slike o svjesnosti potrošačevih učinaka potrošnje na okoliš i prirodu u kojoj prebiva kako bi se donijeli zaključci vezani uz potrošačku politiku kao instrumentarija zaštite okoliša. U radu se obrađuje povezanost potrošačeva ponašanja, njegovih odluka pri kupovini i konzumaciji proizvoda, sa održivim okolišem. Isto tako, istražila se motivacija koja stoji iza odluka potrošača te na koje se načine može na nju utjecati od strane regulatornih tijela i proizvođača. Potrošači često nisu dovoljno moćni da utječu na promjene na tržištu ponude i potražnje i stoga je nužno kroz instrumente potrošačke politike ojačati njihov utjecaj kroz različite mjere predložene u radu. Provedeno je vlastito empirijsko istraživanje na uzorku od 125 ispitanika, pomoću anketnog upitnika kojim su se istražili stavovi potrošača i njihova svijest o utjecaju vlastitih kupovnih navika na okoliš. Upitnik se sastojao od ukupno 18 pitanja. Prva četiri pitanja odnosila su se na demografsko socijalna obilježja ispitanika. Zatim su se u slijedećih 7 pitanja ispitivale navike i informiranost potrošača o ekološki sigurnim proizvodima. Uslijedila su 4 krucijalna pitanja vezana uz potrošačeva očekivanja naspram postupanja države prema tematici zaštite okoliša, kao i stavovi prema proizvođačima i ekološki sigurnim proizvodima koje prodaju i oglašavaju. Posljednja 3 pitanja imala su funkciju osvještavanja potrošača o posljedicama svakodnevnih navika ponašanja koje imaju devastirajući učinak na prirodu i okoliš.

In the adult central nervous system (CNS), ongoing neurogenesis persists in two regions, the subventricular zone (SVZ) of the lateral ventricles and the subgranular zone (SGZ) of the hippocampal dentate gyrus (DG). Newly born neurons generated from neural stem and progenitor cells in the DG integrate locally into the granular layer of the DG as granule cells, which contribute to learning and memory. However, the precise molecular mechanisms involved in the regulation of neurogenesis are still not fully defined. Toll- like receptors (TLRs) are innate immune receptors that prime an inflammatory response and facilitate activation of the adaptive immune response. In addition, recently TLRs have emerged as regulators of multiple processes in the CNS such as neuronal survival, axonal growth, and synaptic plasticity. Thus TLR2, widely expressed within the CNS, is found to be expressed in neural progenitor cells (NPCs) as well. It has been shown that TLR2 expressed by these cells is involved in their differentiation into neurons. In this study, using adult TLR2 deficient mice and their wild- type littermates, we investigated the impact of TLR2 on hippocampal neurogenesis and cognitive function. TLR2 deficiency does not alter the proliferative capacity of NPC in the dentate gyrus, but affects their phenotypic fate by increasing the formation of astrocytes at the expense of neurons. Furthermore, TLR2 deficient mice exhibit reduced dendritic arborization and performed significantly worse in hippocampal- dependent behavioral tasks compared to their control littermates. Thus, our data indicate that dysregulation of the innate immune system by a TLR2 deficiency impairs neurogenesis in the hippocampal DG and cognitive function in adult mice.

Adult neurogenesis, the generation of new neurons, occurs in two regions of the brain, the subventricular zone (SVZ) of the lateral ventricles and subgranular zone (SGZ) of the dentate gyrus (DG) in the hippocampus. Newly born neurons in the SGZ, differentiate and integrate locally into the DG as granule cells, which contribute to both formation and extinction of new memories. However, the mechanisms underlying the regulation of neurogenesis are largely unknown. Tolllike receptors (TLRs) are a family of innate immune system receptors that prime an inflammatory response and facilitate activation of the adaptive immune response. In addition, recent findings suggest that TLRs may also play roles in regulating the multiple processes in the central nervous system (CNS) such as neurogenesis, axonal growth, and neural plasticity. Thus TLR2, one of the most prevalent TLRs in the CNS, is found to be expressed in neural stem progenitor cells (NSPCs). It has been shown that TLR2 expressed by these cells is directly involved in their differentiation into neurons. The aim of this study is to investigate the impact of TLR2 on hippocampal neurogenesis and neural plasticity using TLR2-deficient mice and their wild-type littermates. TLR2 deficiency in mice impairs hippocampal neurogenesis and exhibits reduced dendritic arborization suggesting that TLR2 may be involved in the processes of learning and memory. Further work is required to elucidate roles for TLR2 in cognitive process.

Zdravstvena njega bolesnika sa dekubitusom

Hyperbaric oxygen treatment (HBOT) is a beneficial therapy of ischemic stroke in animal models and humans. The objective of this paper is to describe the possible underlying mechanisms of neural and vascular protection induced by HBOT. We searched the Medline using key words: stroke, neuroprotection, hyperbaric oxygen, microcirculation, vasoreactivity, angiogenesis, COX, NOS, arachidonic acid metabolites, reactive oxygen species (ROS), 20-HETE was conducted. Ischemia induces necrosis, autophagocytosis and apoptosis of brain tissue. After stroke, cerebrovascular resistance increases, this could be due - in part - to the increased release of the vasoconstrictor 20-hydroxyeicosatetraenoic acid (20-HETE. Inhibition of 20-HETE and/or increasing of epoxyeicosatrienoic acids (EETs) levels may decrease cerebral damage following stroke. Neuroprotection by HBOT improves brain metabolism by increasing ATP and decreasing lactate production. Also, HBOT may reduce blood–brain barrier (BBB) permeability and brain edema, thereby decreasing intracranial pressure, inflammatory responses and apoptotic cell death. For example, HBOT decreases hypoxia-inducible factor-lα (HIF-lα) and its targets: erythropoietin and vascular endothelial growth factor (VEGF). Also, HBOT prevents lipid peroxidation of membranes, reduces COX-2 expression and leukocyte adhesion during reperfusion and infiltration of neutrophils into the injured brain. It has also been shown that HBOT improves tissue healing by collagen matrix formation and angiogenesis. Interestingly, HBOT in normal tissues elicits significant vasoconstriction ; in contrast, it improves microcirculation of ischemic tissues by inhibiting post-ischemic vasoconstriction. Our recent studies showed that in diabetic rats HBOT significantly improves dilation of cerebral vessels in response to ACh and hypoxia. Thus HBOT likely elicits neuroprotection via two main pathways: one is improved targeted perfusion to increase oxygen delivery to ischemic tissue and two is, preventing apoptosis due to various changes in the metabolism of brain tissues ; both could be due to altered production of vasoprotective and neuroprotective metabolites of arachidonic acid, such as 20-HETE, and EETs. Support: HUHR Bilateral Collaboration Grant 2009-2011, Drenjancevic and Koller

New adamantane-dipyrromethanes (AdD 1-4) were synthesized and their anion binding properties investigated. AdD 1-3 form 2:1 complexes with F- (AdD:F=2:1) characterized by high association constants, and 1:1 complexes with Cl-, Br-, HSO4- and H2PO4-. The binding of Cl-, Br-, HSO4- and H2PO4- by AdD 1-3 is 2-3 times stronger than for the reference compound, meso-phenyldipyrromethane (5). However, AdD 4 forms complexes with F- characterized by 1:1 and 1:2 stoichiometry (AdD:F = 1:2).

Previously we have shown that liver cells of mice intoxicated with high dose of acetaminophen (Paracetamol, APAP) synthesize increased quantities of prostaglandin E2 (PGE2), prostaglandin I2 (PGI2) and thromboxane (TXA2). The aim of study was to determine which of these prostanoids contribute to defensive reaction of host to toxic effect of APAP (i.e. is hepatoprotective) and those which increase the toxic effect of APAP (i.e. is pathogenic). The toxicity of APAP was determined by observing the survival of mice given a lethal dose of APAP by gastric lavage and concentration of serum aminotransferases (AST and ALT) in mice given a sublethal dose of APAP. The concentration of particular prostanoid in body fluid of mice was modulated either by giving exogenously its stabile analog or giving specific anti-PG antibodies or inhibitors of specific receptors. In previous investigations we have shown that 16-16-dimethyl-PGE2 (a stable analogue of PGE2) had a strong protective effect if given to mice before or after APAP ; in contrast, application of specific anti- PGE2 antibodies increased toxic effect of APAP. Here we have shown that anti-TXB2 antibodies have opposite effect than anti-PGE2 antibodies, i.e., when given to APAP intoxicated mice, they decreased significantly serum AST and ALT levels. Recombinant TXB2 (0, 2 mg/kg) had no significant effect on AST and ALT levels in same mice. Presently we are testing the effect of various prostanoids agonists and antagonist on survival of intoxicated mice. The effect protective PG-s on synthesis of hepatotrophic cytokine, IL-6 and induction of Stat3 protein will also be tested.

INTRODUCTION. Previously we have shown that inflammatory cytokines (IL-1α /β and IL-6) have hepatoprotective effect if given to mice before administration of acetaminophen (APAP) or D-Galactozamine + LPS and that this effect is partially mediated by PGE2. Since many inflammatory cytokines stimulate the synthesis of cAMP as well as PGE2, we investigated the effect of agonists, antagonists and inhibitors of cAMP degradation on APAP toxicity. MATHERIALS AND METHODS. APAP (300 mg/kg) was administered by gastric lavage to mice which were given i.p., 1-2 hours before or up to 3 hours after a stable agonist (dibutyryl-cAMP – Db-cAMP) or antagonist (2, 3-dideoxyadenosine – DDA) of cAMP or inhibitor of phoshodiesterase IV (Rolipram). The survival of mice was followed for 72 hours and level of serum aminotransferases (AST and ALT) were determined 18-24 hours after administration of APAP. RESULTS. Db-cAMP (25 mg/kg) as well as Rolipram (8 mg/kg) significantly increased the survival of mice and reduced AST and ALT serum concentration if given 1-2 hours before or up to 2 hours after AAP administration. On the contrary, DDA (160 μ g/kg) decreased the survival of mice and increased serum aminotransferase concentration if given 2 hours before or ½ ; ; hours after APAP, but only effect on aminotransferase level was statistically significant. The treatment of animals with APAP greatly decreased the level of cAMP (20 or more times) in liver in vivo. IL-1β , which was previously shown to have protective effect on APAP toxicity, if given before APAP, greatly increased the synthesis of cAMP in liver in vivo in comparison to saline control (to approximately 2/3 of level in normal mice), and this effect could be blocked almost completely with DDA, given after IL-1α but before APAP. Similar results with Db-cAMP were obtained in mice intoxicated with D-galactosamine + lipopolysaccharide. Presently, we are investigating signaling ways of this protective effect, using more specific inhibitors of cAMP or its downstream intracellular mediators. Specially, we are investigating the role of NF-κ B and protein kinase A (PKA) in protective action of cAMP. CONCLUSIONS. cAMP has a significant protection on liver toxicity induced by xenobiotics.

In the scope of our research on the synthesis of the macrocyclic compounds capable of selective recognitions of different guest molecules (1), we turned our attention to preparation of anion sensors. Anions play key roles in wide range of chemical and biological processes and it is of great importance to design suitable molecular receptors that can recognize and bind anions. A significant number of molecules wich are used as anion sensors are composed of pyrroles which can serve sa binding subunit due to presence of acidic NH proton to which anion can be attached by the hydrogen bonds. Since it was shown that the presence of two NH protons is enough to accomplish binding of an anion with sufficient binding constants (2), we choose to investigate the applicability of dipyrromethanes as anion binding agents. Herein we report the synthesis of four new adamantane-dipyrromethane derivatives (AdD) 1-4, as well as investigation of their anion binding properties. It was anticipated that incorporation of the bulky adamantane would hinder rotational mobility of the pyrrole moietes in the AdD resulting in the increase of the stability of the complexes with anion, as compared to the dipyrromethane bearing aromatic phenyl subdtituent in the meso position (5-phenyldipyrromethane). Moreover, adamantane derivatives are characterized by lipophilicity which may provide the potentional applicability of the AdD molecules as anion extracting agents. In this study we also present the result s of the binding with 5 anions: F-, Cl-, Br-, HSO4- and H2PO4-. Stability constants for the complexes with F- anion are much larger then for the other anions.

Diazo spojevi su česti prekursori za pripravu karbena, kratkoživućih vrsta koje sadrže vrlo reaktivni ugljikov atom. Mogučnost istodobnog generiranja dvaju karbenskih centara na pentaciklo[5.4.0.0(2, 6).0(3, 10).5(5, 9)]undekanu objavljena je nedavno. Nastavljajući naš rad na divalentnim ugljikovim intermedijerima adamantan-1, 3-dikarbenima pripravljen je 1, 3-bis(diazobenzoil)adamantan te je proučavano nastajanje i reaktivnost odgovarajučeg dikarbena. Teoretski računi pokazali su da uvođenje fenilnog supstituenta osim steričke osigurava i elektronsku stabilizaciju karbenskog centra. U ovom radu je prikazana sinteza, te uspoređeni rezultati fotokemijske i pirolitičke dekompozicije diazo spoja.

It is known that proinflammatory cytokines influence the effect of hepatotoxic chemicals on liver (1). Previously we have shown that IL-1alpha and IL-6 have hepatoprotective effect if given to mice before administration of acetaminophen (APAP) and that this effect is partially mediated by PGE2 (1, 2). Since many inflammatory cytokines stimulate the synthesis of cAMP as well as PGE2, we investigated the effect of agonists, antagonists and inhibitors of cAMP degradation on APAP toxicity. APP (300 mg/kg) was administered intragastrically to mice which were given one to two hours before or up to 3 hours after a stable agonist of cAMP (dibutyryl-cAMP – Db-cAMP), antagonist (2, 3-dideoxyadenosine – DDA) or inhibitor of phoshodiesterase IV (Rolipram). All agents were given i.p. The survival of mice was followed for 72 hours and level of serum aminotransferases (AST and ALT) were determined 18-24 hours after administration of APAP. Db-cAMP (25 mg/kg) as well as Rolipram (8 mg/kg) significantly increased the survival of mice and reduced AST and ALT serum concentration if given 1-2 hours before or up to 2 hours after AAP administration. Db-cAMP had the same effect if given 3 hours after AAP, but effect was not statistically significant. On the contrary, DDA (160 microg/kg) decreased the survival of mice and increased serum aminotransferase concentration if given 2 hours before or ½ ; hours after AAP, but only effect on aminotransferase level was statistically significant. The treatment of animals with APAP greatly decreased the level of cAMP (20 or more times) in liver in vivo (the degradation of cAMP was blocked by rolipram). IL-1alpha, which was previously shown to have protective effect on APAP toxicity, if given before APAP, greatly increased the synthesis of cAMP in liver in vivo in comparison to saline control (to approximately 2/3 level in normal mice), and this effect could be blocked almost completely with DDA, given after IL-1α but before AAP. Presently, we are investigating signaling ways of this protective effect, using more specific inhibitors of cAMP or its downstream intracellular mediators.

U ovom radu opisana je kristalizacija navedenih pentadekapeptida i oktapeptida metodom uparavanja, metodom difuzije para sjedeće i viseće kapi.

The aim of this study was to determine the level of endogenous prostanglandin E2 (PGE2), prostanglandin F2alpha (6-keto-PGF1alpha) and thromboxane B2 (TXB2) in the gastric and duodenal mucosa of patients with duodenal ulcer and duodenitis. Besides, the investigation aimed at determining the effect of smoking and infection by Helicobacter pylori on prostanglandin synthesis. The investigation comprised 62 patients with duodenal ulcer, 46 patients with duodenitis and 44 controls. The result of our investigation indicate that the decreased prostanglandin synthesis in gastric and duodenal mucosa determined in patients with duodenal ulcer may have a considerable role in development of duodenal ulcer. Futhermore, the harmful effects of smoking on the gastric and duodenal mucosa may be mediated by the decreased prostanglandin synthesis in the gastric and duodenal mucosa. However, Helicobacter pylori seems to affect the development of duodenal ulcer through other mechanisms.

Introduction: It is known that Kupffer and other non-parenchymal liver cells are activated in hepatic injury, even if this is induced acutely by administration of high doses of hepatotoxic drugs. These cells secrete proinflammatory cytokines, prostaglandins (PG) and other products, which may contribute to pathogenic or repair processes. In these experiments we investigated the role of IL-1, which is known for its cytoprotective effect, in mice intoxicated with a lethal dose of acetaminophen (AAP). Materials and Methods: AAP (350 mg/kg) was given to mice by gastric lavage. The recombinant cytokines, PGs and antibodies to these agents were given to mice i.p. before or after AAP. The concentration of IL-1 and IL-6 in body fluids was determined in biossays with sensitive lines and concentration of PGs by RIA. Serum aminotransferase levels (AST and ALT) were determined by standard clinical laboratory methods. Results: IL-1a and IL-1b decreased mortality and serum aminotransferase levels when given to mice 2-20 hours before intoxication with AAP, and were without effect when given after AAP. Maximal effect was obtained with about 3 ng of IL-1a per mouse. Pretreatment with anti-IL-1 antibodies had an opposite effect. IL-1a stimulates IL-6 production in vivo in normal and AAP-intoxicated mice. It also stimulates the production of PGE2 and PGI2 and reduces the production of thromboxane A2 (TXA2) by liver fragments from AAP-intoxicated mice . Pretreatment of mice with polyclonal anti-PGE2 or monoclonal anti-IL-6 antibodies before IL-1a administration decreased its protective effect. Therefore, we investigated the direct protective effect of these two agents. 16-16-dimethyl-PGE 2 (a stable analogue of PGE2 ) had a strong protective effect if given to mice before or up to two hours after AAP. Similarly, recombinant mouse IL-6 (rmIL-6) had also significant protective effect, which was dose-dependent (400 to 10 000 U per mouse), but only if given before AAP. On the contrary, monoclonal antibodies to mouse rmIL-6, given to mice 3 hours before AAP, increased the mortality of mice and serum AST and ALT levels. IL-1a was also protective in mice with hepatoxicity induced with D-galactosamine and lipopolysaccharide. Presently, we are investigating the influence of IL-1 and PGE2 on synthesis of IL-1 receptor antagonist (IL-1Ra) and cyclic nucleotides (cAMP and cGMP), and effect of these agents on hepatotoxicity of AAP. Conclusion: Protective effect of IL-1 in mice with acute hepatotixicity induced with AAP is partly mediated by IL-6 and PGE2. These agents may further act by inducing the synthesis of IL-1Ra and/or cyclic nucleotides.

It is known that Kupffer and other non-parenchymal liver cells are activated in hepatic injury, even if this is induced acutely by administration of high doses of hepatotoxic drugs. These cells secrete prostaglandins, proinflammatory cytokines and other products, which may contribute to pathogenic or repair processes. In these experiments we investigated the role of prostaglandin E2 (PGE2), prostaglandin I2 (PGI2) and thromboxane (TXA2) in acute liver toxicity induced in mice by Paracetamol (acetaminophen, AAP). Mice were given a lethal dose of AAP by gastric lavage and production of PGs was determined in supernatants of short term culture of livers slices, taken from mice at various times (2-24 hours) after AAP administration. Several agents which are known to have hepatoprotective effect and/or modulate PG synthesis (IL-1 , ketoconazole and dipyiridamole) were given to mice 1-2 hours before AAP. The synthesis of all prostanoids was greatly increased in liver slices of mice given AAP in comparison to mice given saline. Pretreatment of mice with IL-1 before administration of AAP further increased the synthesis of PGE2 and PGI2 and reduced the synthesis of TXA2 in liver slices. These changes in level of PG synthesis were significant at most time observed time intervals after AAP administration. The effect of ketoconazole and dipyridamole on PG synthesis was measured only at 6 hour interval after AAP administration. Both agents had similar effect as IL-1 , i.e., they increased the synthesis of PGE2 or PGI2 and reduced the synthesis of TXA2. All agents had a significant protective effect in vivo when given before AAP, i.e., they reduced the mortality of mice and level of serum aminotransferases in survived mice (ALT and AST). 16-16-dimethyl-PGE 2 (a stable analogue of (PGE 2 ) had a strong protective effect if given to mice before or up to two hours after AAP. These data suggest that PGE2 and PGI2 might have a protective and TXA2 noxious effect in acute liver injury.

Background and Purpose. We have shown previously that IL-la is highly protective in mice with acute hepatototoxicity induced by AAP In these experiments we investigated whether PCE2 and IL-6 are involved in IL-la-induced hepatoprotection. In addition, we investigated the effect of spontaneously produced IL-6 on AAP-induced hepatotoxicity, and the ef&not ; fect of IL-la on heptotoxicity induced by LPS in D-galactosamine sensi&not ; tized mice. Materials and Methods. The mice received toxic dose of AAP by gas&not ; tric route. Recombinant human IL-lα or recombinant mouse IL-6 were given to mice i.p. 2 hours before AAP polycolnal anti-PGE2 and mono&not ; clonal anti-IL-6 antibodies were given 3 hours before IL-lα , or, when given alone, 5 hours before AAP The mortality of mice was followed, and serum aminotransferase levels (AST and ALT) were determined 24 hours after the AAP administration. Results. IL-lα significantly reduced and the serum ALT and AST levels in the AAP-treated mice. It also decreased the mortality of mice which re&not ; ceived LPS after D-galactosamine. Pretreatment of mice with anti-PGE2 or anti-IL-6 antibodies before IL-lα administration decreased its protective effect in those with AAP-induced hepatotoxicity. Administration of anti-IL-6 antibodies alone before AAP increased the mortality of mice and the se&not ; rum aminotransferase levels, while, in contrast, administration of recombinant mouse IL-6 before AAP had the opposite, i.e. beneficial effect. Conclusion. Both PGE2 and IL-6 mediate, perhaps in mutually interde&not ; pendent way, the hepatoprotective effect of IL-lα .

The protective effect of interleukin lα (IL-lα ) in mice with acetaminophen (AAP)-induced hepatitis was investigated. IL-lα had a significant protective effect if given 2 or more hours (up to 24 hours) before AAP ; it significantly reduced mortality of mice and decreased serum transaminase level. The maximal effect was obtained with the dose of 1000 U (166 ng/kg)IL-lalpha. Pretreatment with IL-1 significantly increased the synthesis of prostaglandin E, (PGE, )in samples of liver tissue from AAP-treated mice, but had no effect on the synthesis of leukotriene C, (LTC4). Pretreatment with indomethacin (IMC) did not abrogate significantly the protective effect of IL-l. Thus, the hepatoprotective effect of IL-lα can not be entirely explained by the stimulation of prostaglandin (PG) synthesis.

Acetaminophen (AAP) administered to mice via gastric tube in a dose of 300-350 mg/kg induced heavy liver injury and killed 55%- 80% mice within 48 hours. Dimethyl-16-16prostaglandin E2 (PGE2), given intraperitoneally (0.025-0.2 mg/kg) 15 to 30 minutes before AAP, ameliorated the liver injury and reduced the mortality in mice to 20%-30%. PGE2 also significantly reduced the increase in plasma transamina.se (ALT, AST) levels induced by AAP. In the same doses, PGE1 had a slight but not significantly protective effect. When given 2 hours after AAP administration, PGE2 delayed the death of mice but had no significant effect on total mortality. Pretreatment of mice with indomethacin (4 mg/kg i.p.) had no effect on AAP toxicity, while pretreatment of mice with specific anti-PGE2 antibodies increased this toxicity. Thus, PGE2 was concluded to have a protective role in AAP-induced hepatotoxicity.

Postishemijska upala značajan je faktor u razvoju ozljede nakon moždanog udara, a TLR2 receptor jedan je od njezinih glavnih medijatora. Glavni cilj ovog doktorata bio je odrediti utjecaj upale posredovane TLR2 receptorom na apoptozu u ishemijskom okolišu. Utjecaj TLR2 receptora istražen je na modelu ishemijskog oštećenja mišjeg mozga u životinja s normalnim (CAG-luc) i onih s onemogućenim Tlr2 genom (CAG-luc-Tlr2-/-). Oba soja ubikvitarno su izražavala luciferazni transgen. Razmjer apoptoze pratio se bioluminiscencijom zatočenim Z-DEVD-aminoluciferinom koji se cijepa i postaje bioluminiscentno aktivan nakon interakcije s pocijepanom kaspazom-3. Ishemijska ozljeda bila je uzrokovana okluzijom srednje moždane arterije, a njena progresija praćen je magnetskom rezonancom. Stanična smrt je potvrđena metodama imunofluorescencije pocijepanom kaspazom-3, protočne citometrije aneksinom-V i TUNEL esejom. Funkcionalno oštećenje izmjereno je testom neurološkog deficita. Zatočeni Z-DEVD-aminoluciferin validiran je i procijenjen kao adekvatan supstrat za praćenje apoptoze. Longitudinalno in vivo mjerenje apoptoze izazovno je te zahtijeva kvalitetnu validaciju i normalizaciju. Nije bilo ukupne razlike u apoptozi između sojeva, ali u nedostatku TLR2 receptora postojao je izraženiji mehanizam nekroze. U akutnom periodu, CAG-luc- Tlr2-/- miševi bolje su preživljavali od CAG-luc miševa, dok u kroničnom periodu nije bilo razlike u preživljenju. Ipak, magnetna rezonanca je pokazala kako su CAG-luc-Tlr2-/- miševi su u kroničnom periodu izgubili više tkiva ipsilateralne hemisfere od CAG-luc miševa. Stoga, iako smanjena upala u akutnom periodu može biti protektivna, dugoročno narušava oporavak., Postischemic inflammation is a significant contributor to ischemic injury development. The TLR2 receptor is one of its main mediators. The main aim of this thesis was to determine the effect of TLR2-mediated inflammation on apoptotic cell death in the ischemic environment. The effect of TLR2 was investigated on the ischemic injury model in animals with normal Tlr2 (CAG-luc) and those with knock-out Tlr2 gene (CAG-luc-Tlr2-/-). Both strains expressed the firefly luciferase transgene ubiquitously. The scope of apoptosis was determined by the utilization of bioluminescence imaging with caged Z-DEVD-aminoluciferin, which becomes available for the bioluminescence reaction after cleavage with activated caspase-3. Middle cerebral artery occlusion was performed to produce the ischemic injury. Its progression was followed with magnetic resonance imaging. Cell death was further confirmed using immunofluorescence with activated caspase-3, flow cytometry with annexin-V and the TUNEL assay. Functional outcomes were assessed using a neurological deficit test. Caged Z-DEVD-aminoluciferin was validated and assessed as an adequate tool for monitoring apoptosis. Longitudinal in vivo measurement of apoptosis is challenging and requires thorough validation and normalization. No total difference in apoptosis between the used strains was found. However, in the absence of the TLR2 receptor, a more pronounced mechanism of necrosis arose. In the acute period, CAG-luc-Tlr2-/- mice had better survival than CAG-luc mice. In the chronic period, there was no difference in survival between the strains. However, magnetic resonance imaging showed that CAG-luc-Tlr2-/- lost more ipsilateral hemisphere tissue than CAG-luc mice during the chronic period. Therefore, although lower levels of inflammation may be protective in the acute period post-stroke, long-term recovery is impaired.

Hipoksijsko-ishemijska ozljeda mozga u perinatalnom periodu je jedan od najčešćih uzroka cjeloživotnog invaliditeta. U ovom radu smo istražili ulogu stanične smrti po tipu nekroptoze u hipoksijsko-ishemijskom oštećenju stanica živčanog sustava, te ulogu živčanih matičnih stanica u oporavku. Prvi cilj je bio uspostaviti pouzdan model nezrelih stanica živčanog sustava, što je postignuto osmodnevnom diferencijacijom živčanih matičnih stanica miša. Usporedba diferencijacije stanica u ambijentalnoj (20 % kisika) i tkivnoj normokosiji (5 % kisika) je pokazala kako tkivna normoksija povećava izražaj nekih gena vezanih uz matičnost, te onih koji određuju identitet neurona i astrocita. Također smo pokazali kako hipoksija (1 % kisika) u trajanju od 24 h nije dovoljna za pokretanje nekroptoze u nezrelim stanicama živčanog sustava, već je za aktivaciju tog oblika stanične smrti potrebna istovremena deprivacija glukoze (OGD). Budući da inhibitor Necrostatin-1s nije utjecao na nekroptozu nezrelih stanica živčanog sustava, zaključili smo kako je nekroptoza u analiziranom modelu neovisna o kinaznoj aktivnosti RIPK1. Iako matične stanice, niti njihov kondicionirani medij nisu direktno smanjili intenzitet nekroptoze nezrelih stanica živčanog sustava, pokazane su mjerljive dobrobiti: dodane matične stanice su djelomično povratile izražaj biljega neurona TUBB3, dok je kondicionirani medij djelomično povratio izražaj SOX2, jednog od proteina važnog za oporavak živčanog sustava., Hypoxic-ischemic encephalopathy is one of the most common causes of lifelong disability. Here, we have investigated the role of necroptosis, a type of cell death, in hypoxic-ischemic insult of cells of the nervous system, and the role of neural stem cells (NSC) in recovery. The first goal was to establish a model of immature cells of the nervous system, which was achieved through the eight-day differentiation of mouse NSC. A comparison of cell differentiation in ambient (20 % oxygen) and tissue (5 % oxygen) normoxia revealed that tissue normoxia increases the expression of some of the genes involved in stemness and neuronal and astrocytic identity. Since activation of necroptosis in immature cells couldn't be achieved by hypoxia (1 % oxygen) during 24 h, we caused necroptosis by oxygen-glucose deprivation. Since inhibitor Necrostatin-1s didn't influence necroptosis, we have concluded that necroptosis in the analyzed model is not dependent on the kinase activity of RIPK1. Although NSC, nor their conditioned medium didn't directly reduce the intensity of necroptosis, we have discovered other benefits: NSC partly recovered the expression of the neuronal marker TUBB3, while the conditioned medium partly recovered the expression of SOX2, a protein important for recovery of the nervous system.

Tijekom nastanka moždanog udara agonisti gvanilat ciklaze (GC)-A djeluju neuroprotektivno. Ovaj učinak nije dokazan za agoniste GC-B. S obzirom da do danas ne postoji istraživanje koje proučava ulogu GC-C receptora i njegovih agonista u razvoju ishemične ozljede mozga, cilj je ove doktorske disertacije utvrditi ulogu urogvanilina (UGN) te aktivacije GC-C ovisnog i GC-C neovisnog signalnog puta u razvoju moždanog udara. Istraživanje je provedeno na mišjem modelu moždanog udara (začepljenje srednje moždane arterije u trajanju od 60 min, MCAO) i to u životinja kojima nedostaje GC-C ili UGN te divljim tipovima (WT) životinja iz istog legla. Mozak miševa sniman je magnetnom rezonancom prije te 24 sata nakon MCAO-a. 48 sati nakon MCAO-a, sniman je Ca2+ odgovor na podražaj UGN-om te je tkivo podvrgnuto imunohistokemijskom bojanju. Nedostatak GC-C-a dovodi do razvoja manje ishemijske lezije. Nedostatak UGN-a nije doveo do jednakih rezultata. U WT i životinja kojima nedostaje UGN, Ca2+ odgovor nakon davanja UGN-a je veći u astrocitima peri-ishemijskog korikalnog područja velikog mozga kada se usporedi s odgovarajućim područjem korteksa neoštećene hemisfere. To nije zabilježeno i kod životinja kojima nedostaje GC-C. Jedini način kako bi GC-C mogao utjecati na Ca2+ stanični signalni sustav astrocita peri-ishemijskog područja je da ove stanice izražavaju GC-C nakon MCAO-a, što nije slučaj u fiziološkim uvjetima. Mogući razlog nastanka manje ishemijske lezije je smanjen Ca2+ odgovor astrocita u peri-ishemijskom području. Kako u životinja kojima nedostaje UGN nije primijećena promjena u veličini lezije u odnosu na divlji tip životinja, možemo pretpostaviti moguću ulogu još uvijek neotkrivenih agonista signalnog sustava gvanilinskih peptida u mozgu ili ulogu drugih, do sada nepoznatih čimbenika regulacije kalcijskog signalnog sustava astrocita. Ovi rezultati ukazuju na mogućnost primjene GC-C inhibitora u smanjenju moždanog oštećenja nakon pojave ishemičnog moždanog udara., After ischaemic stroke, agonists of guanylate cyclase (GC)-A have a neuroprotective effect, whereas the same has not been confirmed for agonists of GC-B. Considering that there has been a lack of research into the effect of GC-C and its agonist uroguanylin (UGN) on ischemic stroke development, the aim of this doctoral thesis is to determine the role of UGN and activation of GC-C dependent and GC-C independent signaling pathway on the development of stroke. In this study a mouse model of ischemic stroke (middle cerebral artery occlusion for 60 min, MCAO) was used in GC-C KO and UGN KO mice and their wild type (WT) littermates. MR images of mouse brains were acquired before and 24 h after MCAO. 48 h after MCAO, the Ca2+ response to UGN stimulation was recorded and immunohistochemical staining was performed. Absence of GC-C resulted in the development of smaller ischaemic lesions while the same effect was not present in the absence of UGN. Upon UGN stimulation, WT and UGN KO animals showed a stronger Ca2+ response in astrocytes of the peri-ischaemic cerebral cortex compared with the same cortical region of the unaffected hemisphere. This was not observed in GC-C KO animals. The only reason why GC-C might affect Ca2+ signalling in peri-ischaemic astrocytes is that GC-C is expressed in these cells after MCAO, which is not the case in physiological conditions. Possible reason for the development of smaller ischaemic lesions is the reduced astrocyte's Ca2+ response in the peri-ischaemic area. Considering that lesion size did not differ between animals lacking UGN and WT, we can assume either the involvement of still unknown agonists of the guanylin peptide signaling system in the brain or the effect exerted by yet undiscovered regulatory factors of the astrocytic Ca2+ signaling system. These results suggest a possible application of GC-C inhibitors in decreasing brain damage after ischaemic stroke.

Budući da nije poznato kako se regulira likvorski tlak unutar kraniospinalnog prostora, u ovom radu ispitali smo utjecaj promjene položaja tijela i volumena likvora na tlak likvora u kranijskom i spinalnom prostoru kod 19 bolesnika sa normotenzivnim hidrocefalusom. Korištenjem dviju volumetrijskih tehnika (CIVET i volBrain) uočeno je kako volumen likvora u kranijskom prostoru iznosi oko 300 ml a metodom poluautomatske segmentacije u spinalnom prostoru prosječno 114 ml, što je ukupno znatno veći volumen likvora nego što se do sada smatralo. Smanjenje volumena likvora lumbalnom punkcijom za 15 ml dovodi do pada likvorskog tlaka za prosječno 8,5 cm H2O što ukazuje da volumen cerebrospinalne tekućine u spinalnom prostoru ima velik utjecaj na visinu tlaka likvora. Mjerenje tlaka likvora u kranijskom i lumbalnom subarahnoidalnom prostoru u horizontalnom položaju ukazuje da je tlak likvora u čitavom kraniospinalnom prostoru na istoj hidrostatskoj razini jednak, dok pri podizanju tijela za 15° tlak u kranijskom prostoru pada a u spinalnom raste. Opažena razlika tlakova u kraniju i spinalno odgovara upravo hidrostatskoj razlici. Po prvi put je opaženo kako bolesnici koji su imali pozitivan klinički odgovor na produženu eksternu lumbalnu drenažu imaju značajno veći volumen likvora u spinalnom prostoru (prosječno oko 120 ml) što ukazuje na novi prediktivni čimbenik u liječenju bolesnika sa normotenzivnim hidrocefalusom. Rezultati ukazuju kako se likvorski tlak ponaša kao hidrostatski tlak i da ne ovisi o sekreciji, cirkulaciji i apsorpciji likvora što je u skladu sa novim shvaćanjima fiziologije i patofiziologije likvora., Since it is unknown how cerebrospinal fluid (CSF) pressure within the craniospinal space is regulated, in this paper we examined the influence of body position and CSF volume on CSF pressure in the cranial and spinal space in 19 patients with normal pressure hydrocephalus. Using two volumetric techniques for cranial (CIVET and volBrain) and semi-automatic segmentation for spinal space, it was observed that the cranial CSF volume is about 300 ml and the spinal CSF volume is 114 ml, which is a much larger volume of CSF than was previously thought. Withdrawal of 15 ml of CSF volume by lumbar puncture leads to a decrease in CSF pressure by average 8.5 cm H2O, which indicates that the CSF volume in the spinal space has a great influence on the level of CSF pressure. The measurement of CSF pressure in the horizontal position in the cranial and lumbar subarachnoid spaces indicates that the CSF pressure in the entire craniospinal space at the same hydrostatic level is same, while lifting the body by 15° decreases the pressure in the cranial space and increases the pressure in the spinal space. The observed difference in pressure between cranial and spinal space corresponds exactly to the hydrostatic difference. For the first time, it was observed that patients who had a positive clinical response to prolonged external lumbar drainage had a significantly higher CSF spinal volume (around 120 ml on average) indicating a new predictive factor in the treatment of patients with normal pressure hydrocephalus. The results indicate that CSF pressure behaves like hydrostatic pressure and does not depend on the secretion, circulation and absorption of cerebrospinal fluid, which is consistent