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Beta-oksidacija u modelu kvasca Barthovog sindroma
No full textBarth syndrome is a complicated and life-threatening systemic disease. It is characterized by a wide range of conditions, mostly affecting the heart and skeletal muscles. Symptoms include cardiomyopathy, skeletal myopathy, neutropenia, increased urinary excretion of 3-methylglutaconic acid and weaker motor skills. Being a genetic disease, it is currently uncurable and is managed through medications that decrease symptom severity. The syndrome is caused by mutations in the TAZ gene which encodes the tafazzin enzyme. Tafazzin has an important role in the cardiolipin synthesis and remodeling pathway. Cardiolipin is a unique phospholipid that makes up a significant portion of the inner mitochondrial membrane. It goes through a complex and highly conserved synthesis and remodeling pathway which is altered in patients with Barth syndrome. Unlike other phospholipids, cardiolipin has a specific structure that differs from other phospholipids and enables it to interact with various membrane proteins and complexes, playing a crucial role in maintaining mitochondrial energy metabolism. By gaining a deeper understanding of the molecular mechanisms and the significance of cardiolipin synthesis and remodeling, new treatment strategies could be developed. These strategies would target the underlying cause of the disease, unlike currently available therapeutic approaches. One potential target is beta-oxidation. Trimetazidine, a drug commonly used to treat heart conditions, inhibits beta-oxidation and has been shown in previous studies to affect mitochondrial phospholipid turnover. However, its impact on cardiolipin synthesis and turnover remains unknown. In this research, we are interested in investigating if inhibition of beta-oxidation enhances mitochondrial phospholipid turnover and increases the levels of functional cardiolipin in cells with TAZ deficiencies. Saccharomyces cerevisiae serves as an excellent model organism for studying Barth syndrome because all the enzymatic steps involved in cardiolipin synthesis and remodeling are conserved between yeast and humans.Barthov sindrom je složena, po život opasna bolest koja zahvaća cijeli organizam. Karakteriziran je širokim spektrom simptoma, koji uglavnom zahvaćaju srce i tjelesne mišiće. Simptomi uključuju kardiomiopatiju, skeletnu miopatiju, neutropeniju, povećano izlučivanje 3-metilglutakonske kiseline putem urina te slabije motoričke sposobnosti. Kao genetska bolest je neizlječiva, a pacijenti se tretiraju lijekovima koji ublažavaju prisutne simptome. Bolest je uzrokovana mutacijama u TAZ genu koji kodira enzim tafazzin. Tafazzin ima važnu ulogu u sintezi i remodeliranju kardolipina. Kardolipin je specifičan fosfolipid koji čini značajan udio unutarnje membrane mitohondrija. Prilikom sinteze i remodeliranja, kardiolipin prolazi složen i evolucijski očuvan put koji je izmijenjen kod pacijenata s Barthovim sindromom. Za razliku od većine fosfolipida, kardolipin ima specifičnu strukturu koja mu omogućuje interakciju s raznim proteinskim kompleksima u mitohondriju. Ovo svojstvo omogućava ključnu ulogu kardiolipina u održavanju mitohondrijskog energetskog metabolizma. Boljim razumjevnjem značaja i molekularnog mehanizma sinteze kardiolipina, mogle bi se razviti nove strategije u liječenju Barthovog sindroma. Jedna potencijalna strategija je inhibicija beta-oksidacije. Trimetazidin, lijek koji se koristi za liječenje srčanih oboljenja, inhibira beta-oksidaciju te prema prethodnim istraživanjima, utječe na promjenu sastava mitohondrijskih fosfolipida. Međutim, utjecaj trimetazidina na kardiolipin je nepoznat. U ovom istraživanju nastojimo ispitati može li inhibicija beta-oksidacije povećati promjenu mitohondrijskih fosfolipida i povećati razine funkcionalnog kardolipina u stanicama s TAZ mutacijama. Saccharomyces cerevisiae je izvrstan modelni organizam za proučavanje Barthovog sindroma jer su svi enzimski koraci uključeni u sintezu i remodeliranje kardolipina evolucijski očuvani između kvasca i ljudi
The latest drugs in the treatment of oncological diseases
No full textOnkološke bolesti čine veliki zdravstveni problem u svijetu, a rak je među vodećim uzrocima smrti u mnogim zemljama. Kada je u pitanju liječenje raka, poznato je da je proces liječenja jako složen i dugotrajan. Pristupi liječenju kao što su kemoterapija i radioterapija imaju mnogo nedostataka. Srećom, u posljednjih nekoliko godina postignut je velik i značajan napredak na području onkologije. Ovaj napredak uključuje razvoj imunoterapije i ciljane terapije kao potencijalno najboljih mogućnosti za liječenje različitih vrsta raka. Kada se radi o imunoterapiji, najboljima su se pokazali "checkpoint" inhibitori te adoptivna T-stanična terapija, konkretnije CAR T-stanična terapija. Među odobrenim lijekovima za liječenje raka u posljednjih nekoliko godina najviše su zastupljeni upravo lijekovi koji pripadaju nekoj od vrsta imunoterapije. Među najnovijima iz 2024. godine su primjerice tislelizumab i lifileucel. Druga vrsta inovativne terapije, ciljana terapija, također se pokazala vrlo uspješnom. Ova terapija uključuje razne inhibitore tirozin kinaze i monoklonska protutijela od kojih su se obećavajućima pokazala bispecifična protutijela. Najnovije odobrena bispecifična protutijela iz 2023. godine su elranatamab i talquetamab. Ciljana terapija je veliko otkriće iz razloga što ciljano djeluje na specifični antigen na stanicama raka, ne oštećujući previše zdrave ljudske stanice. Naravno, u tijeku je razvoj i mnogih drugih novih pristupa za liječenje raka. Veliki broj terapija još se uvijek nalazi u fazi kliničkih ispitivanja, a u sljedećih par godina mogao bi biti odobren značajan broj novih lijekova.Cancer constitutes a major health problem worldwide and is among the leading causes of death in many countries. When it comes to cancer treatment, it is well known that the process is highly complex and takes time. Traditional approaches such as chemotherapy and radiotherapy have not proven to be the best options. Fortunately, significant progress has been made in the field of oncology in recent years. This progress includes the development of immunotherapy and targeted therapy as potentially the best options for treating various types of cancer. In the realm of immunotherapy, checkpoint inhibitors and adoptive cell therapy, specifically CAR T-cell therapy, have shown the most promise. Among the approved cancer treatments in the past few years, the majority belong to some form of immunotherapy. Notable examples from 2024 include tislelizumab and lifileucel. Another type of innovative therapy, targeted therapy, has also proven to be very successful. This therapy includes various tyrosine kinase inhibitors and monoclonal antibodies, among which bispecific antibodies have shown great promise. The newest approved bispecific antibodies from 2023 are elranatamab and talquetamab. Targeted therapy is a significant breakthrough because it specifically targets antigens on cancer cells, causing minimal damage to healthy human cells. Naturally, the development of many other new approaches to cancer treatment is ongoing. A large number of therapies are still in clinical trial phases, and a significant number of new drugs could be approved in the next few years
Systemic effects of photoactivated 5,10,15,20-tetrakis(N-methylpyridinium-3-yl) porphyrin on healthy Drosophila melanogaster
No full textThe influence of GSK-J4 on KDM6B gene activity in three human cancer cell lines
No full textEpigenetičke promjene su pokazane kao nositelji ključne uloge u tumorigenezi i širenju tumora.
Jedna od glavnih epigentičkih promjena u tumorskim stanicama je promjena razine modifikacija
kromatina poput metilacije lizina 27 u histonu H3 (H3K27) što doprinosi promjeni strukture
kromatina i transkripcijskoj aktivnosti mnogih važnih gena. Važni regulatori stanja modifikacija
histona su enzimi poput EZH2 i JMJD3 koji u stanicama imaju antagnostičko djelovanje te su
povezani s raznim vrstama tumora pa tako predstavljaju važnu terapeutsku metu. Glavni inhibitori tih enzima su spojevi poput GSK-J4, inhibitor JMJD3, i tazemetostat, inhibitor EZH2. Ovi spojevi su pokazali obećavajuće rezultate u zaustavljanju epigenetičkih mehanizama koji podržavaju rast tumora mijenjajući njihov epigenetički okoliš te imaju puno slabiji sistemski učinak na
organizam za razliku od konvencionalnih terapija poput cisplatine. U ovom istraživanju analiziran je utjecaj GSK-J4 i tazemetostata na ispoljenost proteina JMJD3 te transkripcijskih faktora
SP1 i STAT3 u tri stanična modela raka s kombiniranim dodavanjem cisplatine kako bi istražili
mogućnost senzibilizacije. Rezultati pokazuju djelomičan utjecaj primjene GSK-J4 na promjenu
ispoljenosti JMJD3 proteina te SP1 i STAT3, uzimajući u obzir da većina ovih promjena može biti
povezana s korištenjem cisplatine u eksperimentima. Također, otkrili smo značajnu promjenu
ekspresije pSTAT3 proteina u jezgrama stanica raka nakon primjene tazemetostata, iako nije
postignuta statistička značajnost u tim promjenama. Potrebna su daljnja istraživanja kako bi se
bolje razumjeli specifični mehanizmi djelovanja ovih inhibitora i njihov potencijal u terapiji raka,
posebno u kontekstu epigenetičkih promjena koje doprinose razvoju bolesti.Changes in the epigenetic environment have been shown to play a crucial role in tumorigenesis
and the subsequent spread of tumors. One of the primary epigenetic alterations in tumor cells
involves changes in chromatin modifications such as H3K27 methylation, altering chromatin
structure and transcriptional activity of many crucial genes. Key regulators of histone modification states are enzymes like EZH2 and JMJD3, which exhibit antagonistic actions in cells and are
associated with numerous types of tumors, thus representing important therapeutic targets.
Major inhibitors of these enzymes include compounds like GSK-J4, an inhibitor of JMJD3, and
tazemetostat, an inhibitor of EZH2. These compounds have shown promising results in disrupting epigenetic mechanisms that support tumor growth by altering their epigenetic landscape
and have much less systemic impact on the body compared to conventional therapies like cisplatin. In this study, we analyzed the impact of GSK-J4 and tazemetostat on the expression of
JMJD3 protein and the expression of SP1 and STAT3 transcription factors in three cancer cell
models with combined addition of cisplatin to investigate the potential for sensitization. Our
results show a partial effect of GSK-J4 on changing JMJD3 protein expression and the expression of SP1 and STAT3, considering that most of these changes may be associated with the use
of cisplatin in the experiments. Additionally, we observed a significant change in pSTAT3 protein expression in the nuclei of cancer cells after tazemetostat treatment, although statistical
significance was not achieved in these changes. These findings suggest the need for further research to better understand the specific mechanisms of action of these inhibitors and their potential in cancer therapy, particularly in the context of epigenetic changes contributing to disease development
The role of calpain in diabetes and diabetic complications
No full textKalpaini su klasa koju čine 15 članova kalcijem aktiviranih nelizosomalnih neutralnih proteaza koje utječu na širok raspon staničnih funkcija. Kalpaini su obično lokalizirani u citosolu i unutar mitohondrija. Pretjerana aktivacija kalpaina povezana je s brojnim bolestima mozga, očiju, srca, pluća, gušterače, bubrega, krvožilnog sustava i skeletnih mišića. Stoga kalpain može poslužiti kao potencijalni terapeutski cilj u mnogim bolestima. Obzirom da se kalpaini povezuju s kroničnim komplikacijama šećerne bolesti (DM), kao što su dijabetička kardiomiopatija, dijabetička nefropatija i dijabetička retinopatija, ovaj pregled sadržava dosadašnja saznanja
temeljena na literaturnim podacima o strukturi, aktivaciji i biološkim funkcijama kalpaina, te učinku prekomjerne aktivacije kalpaina u patogenezi neurodegenerativnih bolesti, bolesti krvožilnog sustava, autoimunih bolesti i upali.Calpains are a class of 15 calcium-activated, non-lysosomal, neutral proteases that influence a variety of cellular functions. Calpains are normally localised in the cytosol and mitochondria. Excessive activation of calpain is associated with numerous diseases of the brain, eyes, heart, lungs, pancreas, kidneys, circulatory system and skeletal muscles. Therefore, calpain can serve as a potential therapeutic target for many diseases.Considering that calpains are associated with chronic complications of diabetes (DM) such as diabetic cardiomyopathy, diabetic nephropathy and diabetic retinopathy, this review presents the current state of knowledge based on literature data on the structure, activation and biological functions of calpain, as well as the effects of excessive calpain activation on the pathogenesis of neurodegenerative diseases, diseases of the circulatory
system, autoimmune diseases and inflammation
Utjecaj suplementacije glutaminom i kolinom kloridom tijekom izolacije na fenotip preferencijalne konzumacije metamfetamina kod D. melanogaster
No full textOvisnost je globalni problem koji pogađa mnoge ljude i zajednice, zbog čega je važno istražiti molekularne mehanizme ovisnosti. Jedan od glavnih okolišnih stresora, socijalna izolacija, narušava ravnotežu neurokemikalija u mozgu, što posljedično utječe na ponašanja povezana s ovisnošću. Ova studija ispituje učinke dodavanja prekursora neurotransmitera i socijalne izolacije na sklonost metamfetaminu (METH) kod D. melanogaster. U ovoj studiji, divlji tip mušica starih tri do pet dana izoliran je na pet dana uz dohranu s 10 mg/mL L-glutamina (GLU) prekursora glutamata ili 3 mg/mL kolin klorida (CH.CL) prekursora acetilkolina. Nakon izolacije i predtretmana mušice su prebačene u FlyCAFÉ esej, gdje mi je tri uzastopna ponuđen izbor između obične hrane i hrane s dodatkom METH-a. Preferencija za METH-a je izračunata kao razlika između volumena konzumirane METH-hrane i uobičajene hrane za individualnu mušicu, a podaci su prikazani kao prosjek za svaki dan eksperimenta. Rezultati ove studije su pokazali da suplementacija s GLU ili CH.CL tijekom pet dana izolacije povećava preferenciju METH-a u FlyCAFÉ-u, što je suprotno prethodnim studijama gdje je pokazano da izolacija smanjuje preferenciju za METH u odnosu na neizolirane mušice. Ovo istraživanje otkriva da dohrana s GLU ili CH.CL smanjuje učinak izazvan izolacijom, uzimajući u obzir socijalne i biokemijske aspekte u testu preferencije METH kod mušica. Budući genetski testovi trebaju potvrditi ulogu glutamina i acetilkolina u regulaciji ovog endofenotipa ovisnosti kod mušica.Addiction represents a global challenge to the world, affecting many individuals and communities, making it necessary to examine the mechanisms of addiction. One major environmental stressor, social isolation, disrupts the balance of neurochemicals in the brain that subsequently influence the behaviors related to addiction. This study examines the effects of neurotransmitter precursor supplementation and social isolation on methamphetamine (METH) preference in D. melanogaster. Wild-type three to five day-old male flies were isolated for five days and supplemented with either 10 mg/mL L-Glutamine (GLU) precursor of glutamate, or 3 mg/mL choline chloride (CH.CL) precursor of acetylcholine. After isolation and pretreatment, we used the FlyCAFÉ assay where flies were offered a choice between regular food and METH-supplemented food for three consecutive days. METH preference was calculated as difference between METH-food and regular food for induvial fly, and data were presented as average for every day of the experiment. The results of this study have shown that supplementation with GLU or CH.CL during five days of isolation increased METH preference in FlyCAFÉ, what contrasts with previous studies where it was shown that isolation reduces METH preference relative to non-isolated flies. This research reveals that GLU or CH.CL supplementation reduced effect induced by isolation, considering social and biochemical aspects in test for METH preference in flies. Future genetic tests are needed to confirm role of glutamine and acetylcholine in the regulation of this addiction endophenotype in flies
Karakterizacija staničnih linija za istraživanje agregacije proteina kod mentalnih bolesti
No full textMental health is an essential part to of our overall well-being and
human rights, ranging from complete wellbeing to severe emotional
distress. Several risk factors contribute to these diseases, including
psychological and biological components such as genetics and
emotional abilities, which can often be altered by changes in brain
structure and function. Common mental health disorders, including
schizophrenia, bipolar disorder, and major depressive disorder, cause
significant disturbances in behaviour, cognition, and emotional
control, resulting in distress and functional impairment.
Proteins are essential for cellular functions and must fold correctly so
they can perform their functions. However, proteins can misfold and
aggregate, especially under certain conditions, resulting in toxic
aggregates. Normally, cellular quality control systems, such as the
ubiquitin-proteasome system and autophagy, manage these
aggregates. When these mechanisms fail, protein misfolding
diseases, often known as proteinopathies, develop, including
neurodegenerative disorders such as Alzheimer's, Parkinson's, and
Huntington's. These disorders are characterized by toxic protein
aggregation which leads to neuronal dysfunction and cell death,
linking proteinopathies to cognitive decline and psychiatric
symptoms.
The correlation between mental illness and proteinopathies is
becoming increasingly recognised. Proteins such as DISC1 and
TRIOBP-1 have been observed to aggregate in mental illnesses,
suggesting that they play important roles in the pathophysiology of
these conditions.
This thesis focused on providing an insight into the behaviour of
TRIOBP-1 and DISC1 in neuronal cells, allowing us to investigate the
behaviour of these proteins and their potential connection to mental
diseases. The primary aim was to determine the most optimal dose
of doxycycline and period of treatment for protein expression,
followed by the detection of protein aggregation by Western blotting,
fluorescent microscopy, and insolubility assays. By demonstrating
these proteins' aggregation tendencies, trying to contribute to the
understanding of how protein misfolding may connect to
neurodevelopmental diseasesMentalno zdravlje je ključni dio općeg blagostanja i ljudskih prava,
kreće se od potpunog blagostanja do teške emocionalne patnje.
Nekoliko čimbenika rizika doprinosi ovim bolestima, uključujući
psihološke i biološke komponente poput genetike i emocionalnih
sposobnosti, koje se često mogu promijeniti zbog promjena u
strukturi i funkciji mozga. Uobičajeni poremećaji mentalnog zdravlja,
uključujući shizofreniju, bipolarni poremećaj i veliki depresivni
poremećaj, uzrokuju značajne poremećaje u ponašanju, kogniciji i
emocionalnoj kontroli, što rezultira stresom i funkcionalnim
oštećenjima.
Proteini su ključni za stanične funkcije i moraju se pravilno saviti kako
bi mogli obavljati svoje funkcije. Međutim, proteini se mogu
nepravilno saviti i agregirati, osobito u određenim uvjetima, što
rezultira stvaranjem toksičnih agregata. Normalno, stanični sustavi
kontrole kvalitete, poput sustava ubikvitin-proteasoma i autofagije,
upravljaju ovim agregatima. Kada ti mehanizmi zakažu, razvijaju se
bolesti uzrokovane nepravilnim savijanjem proteina, poznate kao
proteinopatije, uključujući neurodegenerativne poremećaje poput
Alzheimerove, Parkinsonove i Huntingtonove bolesti. Ovi poremećaji
karakterizirani su toksičnim agregacijama proteina koje dovode do
disfunkcije neurona i stanične smrti, povezujući proteinopatije s
kognitivnim padom i psihijatrijskim simptomima.
Povezanost između mentalnih bolesti i proteinopatija sve se više
prepoznaje. Primijećeno je da proteini poput DISC1 i TRIOBP-1
agregiraju u mentalnim bolestima, što sugerira da oni imaju važnu
ulogu u patofiziologiji ovih stanja.
Ovaj se rad usredotočio na pružanje uvida u ponašanje TRIOBP-1 i
DISC1 u neuronskim stanicama, što nam omogućuje da istražimo
ponašanje tih proteina i njihovu potencijalnu povezanost s mentalnim
bolestima. Primarni cilj bio je utvrditi optimalnu dozu doksiciklina i
razdoblje tretmana za ekspresiju proteina, nakon čega slijedi
detekcija agregacije proteina pomoću Western blottinga,
fluorescentne mikroskopije i ispitivanja netopivosti. Dokazivanjem
sklonosti agregaciji ovih proteina, pokušava doprinijeti razumijevanju
kako nepravilno savijanje proteina može biti povezano s
neurodegenerativnim bolestima
Uloga gena povezanih s redoksom u regulaciji fenotipova izazvanih metamfetaminom u mušici Drosophila melanogaster
No full textAddiction is a multifactorial neuropsychiatric disorder marked by compulsive drug-seeking behaviors despite adverse consequences. Neuroplasticity, the ability of the brain to adapt and reorganize, plays a crucial role in addiction, involving changes in gene expression, protein modifications, and synaptic organization. Alterations in redox homeostasis, particularly dysregulation of reactive oxygen species (ROS) signaling, are implicated in these neuroplastic changes. Drosophila melanogaster, with its well-characterized genome and genetic tools, serves as a valuable model for studying addiction-like behaviors such as drug self-administration (SA) and locomotor sensitization (LS). This research aims to identify genes and proteins involved in methamphetamine (METH) addiction using flies and approach integrating behavioral assays, genetic manipulations (selective breeding and genetic screening), and proteomic analyses.
We developed the FlyCafe assay to measure METH consumption and demonstrated flies' preferential SA of METH. Through 30 generations of selective breeding, we established fly strains with high (HP) and low (LP) preferences for METH, revealing distinct phenotypic profile of LP flies, including increased activity and body weight, decreased sleep and negative geotaxis and increased dopamine, tyramine and glutamate relative to HP flies. Proteomic analysis of these strains identified differential proteins linked to metabolic processes, structural integrity, and protein turnover, including Bacchus, negative regulator of conversion of tyramine to octopamine, which was uniquely present in LP flies. Next, we showed that flies exhibiting LS displayed reduced preference for METH SA, and vice versa, suggesting shared molecular mechanisms through the period gene. We conducted a genetic screen to identify redox-related genes that regulate LS to volatilized METH (vMETH), uncovering several critical genes such as Cat, Sod1, Sod2, Gapdh1, and Men, which play functional roles in the regulation of LS. Remarkably, Sod1, Gapdh1, and Men are necessary in dopaminergic and serotonergic neurons for both LS and SA. Proteomic analysis of brain tissues from flies that did or did not develop LS after two administrations of vMETH identified a set of proteins unique to the LS phenotype, highlighting significant changes in redox-related proteins. This included the upregulation of several antioxidative, glycolytic, and tricarboxylic acid (TCA) cycle enzymes (Cat, Prx3, Prx6c, Jafrac1, Gapdh1, Gapdh2, mAcon1, Mdh1, Mdh2) and the downregulation of enzymes related to oxidative phosphorylation (NADH dehydrogenases).
Combining these results, our study reveals that both LS and SA of METH are modulated by interconnected pathways involving peroxide regulation, glucose metabolism, NADPH production, and neurotransmitter systems. Together, these processes contribute to the development and maintenance of addiction-related neuroplasticity evident as LS and SA. Additionally, our findings highlight the importance of the neurotransmitter tyramine in SA behavior, indicating that elevated tyramine levels, influenced by the Bacchus protein, may modulate the neural circuits involved substance preference and consumption. This research can serve as a foundation for exploring dietary and lifestyle interventions to maintain redox balance and metabolic health. For example, specific dietary supplements that enhance antioxidant defenses or support metabolic pathways, such as antioxidants, metabolic modulators, or NADPH boosters, could be investigated as adjunctive therapies for addiction.Ovisnost je složen neuropsihijatrijski poremećaj karakteriziran kompulzivnim uzimanjem droge unatoč negativnim posljedicama. Neuroplastičnost, sposobnost mozga da se prilagodi i reorganizira kao odgovor na iskustva, ima temeljnu ulogu u razvoju ovisnosti. Taj proces uključuje kompleksne molekularne mehanizme, uključujući promjene u ekspresiji gena, modifikacije proteina i reorganizaciju sinapsi. Promjene u redoks homeostazi, posebice promjene u regulaciji signalizacije putem reaktivnih kisikovih vrsta (ROS), povezane su s neuroplastičnim promjenama i ovisnošću. Drosophila melanogaster, s obzirom na njen dobro karakteriziran genom te lako dostupne sofisticirane genetske alate, pokazala se kao vrijedan modelni organizam za proučavanje ponašanja sličnih ovisnosti, poput samoadministracije droge (SA) i lokomotorne senzitizacije (LS), koja se lako mogu kvantificirati kod vinskih mušica. Glavni cilj ovog istraživanja je identificirati nove gene i proteine uključene u ovisnost o metamfetaminu (METH) koristeći Drosophilu kao model i pristup koji integrira bihevioralne testove, genetske manipulacije (selektivni uzgoj i genetski probir) te proteomske analize.
Razvili smo FlyCafe test za mjerenje konzumacije METH-a i pokazali da mušice preferencijalno samoadministriraju METH. Kroz 30 generacija selektivnog uzgoja, uspostavili smo sojeve mušica s visokom (HP) i niskom (LP) preferencijom za METH, otkrivajući različit fenotipski profil LP mušica, uključujući povećanu aktivnost i tjelesnu težinu, reducirano spavanje i negativnu geotaksiju te povećane razine dopamina, tiramina i glutamata. Proteomska analiza ovih sojeva identificirala je diferencijalne proteine povezane s metaboličkim procesima, strukturnim integritetom te sintezom i degradacijom proteina, uključujući Bacchus, negativni regulator pretvorbe tiramina u oktopamin, koji je jedinstveno prisutan kod LP mušica. Zatim smo pokazali da mušice koje pokazuju LS imaju smanjenu preferenciju za METH SA, i obratno, sugerirajući zajedničke molekularne mehanizme koji uklljučuju period gen. Proveli smo genetski probir kako bismo identificirali redoks-gene koji reguliraju LS na volatilizirani METH, otkrivajući nekoliko ključnih gena kao što su Cat, Sod1, Sod2, Gapdh1 i Men, koji imaju funkcionalne uloge u regulaciji LS-a. Sod1, Gapdh1 i Men geni nužni su u dopaminskim i serotoninskim neuronima za oba fenotipa, LS i SA. Proteomska analiza moždanih tkiva mušica koje su razvile LS nakon dvije administracije volatiliziranog METH-a identificirala je set proteina jedinstvenih za LS fenotip, naglašavajući značajne promjene u redoks-proteinima. To uključuje povećanu ekspresiju nekoliko antioksidativnih i glikolitičkih enzima te enzima ciklusa trikarboksilnih kiselina (Cat, Prx3, Prx6c, Jafrac1, Gapdh1, Gapdh2, mAcon1, Mdh1, Mdh2) i smanjenje enzima povezanih s oksidativnom fosforilacijom (NADH dehidrogenaze).
Kombinirajući ove rezultate, naše istraživanje pokazalo je da su LS i SA METH-a modulirani međusobno povezanim putevima koji uključuju regulaciju peroksida, metabolizam glukoze, proizvodnju NADPH-a i neurotransmiterske sustave. Zajedno, ovi procesi doprinose razvoju i održavanju neuroplastičnosti povezane s ovisnošću, evidentne kao LS i SA. Dodatno, naši rezultati ističu važnost neurotransmitera tiramina u SA, ukazujući na to da povećane razine tiramina, pod utjecajem Bacchus proteina, mogu modulirati neuronske mreže uključene u preferenciju i konzumaciju droge. Ovo istraživanje može poslužiti kao osnova za istraživanje prehrambenih intervencija s ciljem održavanja redoks ravnoteže i metaboličkog zdravlja. Na primjer, specifični dodaci prehrani koji pojačavaju antioksidativnu obranu ili podržavaju metaboličke puteve, kao što su antioksidansi, metabolički modulatori ili NADPH pojačivači, mogli bi se koristiti kao dodatne terapije za ovisnost
Adsorption of glucose on the zeolite clinoptilolite surface in water and model stomach and gut solutions in vitro and in vivo
No full textZeoliti su prirodni ili sintetski alumosilikatni minerali porozne strukture. Pore su
međusobno povezane kanalima u kojima su smješteni hidratizirani kationi te molekule
vode. Primarna ili osnovna građevna jedinica zeolita je tetraedar TO4 gdje je T oznaka
za četverovalentni silicij (Si(IV)) ili trovalentni aluminij (Al(III)) pri čemu su kod različitih
zeolitnih materijala omjeri Al(III) naspram Si(IV) različiti. Ovi se elementi u prostoru
povezuju preko zajedničkih atoma kisika (O) gradeći sekundarne poliedarske jedinice
čijim povezivanjem nastaje kristalna rešetka. Upravo je ova rešetka odgovorna za
glavno svojstvo zeolita, ionsku izmjenu kationa iz šupljina s kationima iz okoline. Drugo
važno svojstvo zeolita je svojstvo adsorpcije organskih molekula na površinu zeolita.
U medicinske je svrhe do sada korišten netoksični prirodni zeolitni materijal klinoptilolit.
U ovom su se doktorskom radu za istraživanje koristili različito mikronizirani
klinoptilolitni materijali: TMAZ (tribomehanički aktivirani klinoptilolit), PMA (dvostruko
tribomehanički aktivirani zeolit) i PMAO2 (dvostruko tribomehanički aktivirani zeolit
obogaćen medicinskim kisikom), te kao kontrolni spoj korišten je sintetski Zeolit A. Za
navedene je materijale je u prvom dijelu istraživanja utvrđen omjer Si/Al u vodi i
modelnim otopinama želucai crijeva. Nakon inkubacije u vodi ili modelnim otopinama
u trajanju 24 h, izdvojene su frakcije centrifugiranjem pri 4000 rpm, 14000 rpm i
140000 rpm. Supernatanti svake frakcije analizirani su uz pomoć elektronskog
mikroskopa (skenirajući elektronski mikroskop, SEM) i elektronske disperzijske
spektroskopije (EDS). Oblik čestica supernatanta zeolitnog materijala i omjer Si/Al
uspoređen je sa zeolitnim materijalima u krutom stanju. U drugom dijelu istraživanja,
istražen je i kvantificiran postotak interakcije ili adsorpcije D-glukoze na površinu
zeolitnih materijala uz pomoć elektronske mikroskopije SEM u kombinaciji s
elementnom analizom EDS te kvantitativne metode tekućinske kromatografije
ultravisoke razlučivosti (UHPLC). Adsorptivna svojstva šećera validirana su na
zdravim miševima in vivo za zeolit PMA koji je pokazao najbolja svojstva interakcije s
D-glukozom in vitro pri čemu su rezultati ukazali na smanjenje razina glukoze u krvi u
kod miševa pred tretiranih sa zeolitom PMA. Dobiveni rezultati pokazuju potencijal
korištenja zeolitnih materijala u patološkim stanjima poput metaboličkog sindroma ili
dijabetesa što bi trebalo dodatno istražiti.Zeolites are natural or synthetic aluminosilicate minerals with a porous structure. The
pores are interconnected by channels in which hydrated cations and water molecules
reside. The primary or basic building block of zeolite is the TO4 tetrahedron where T
stays for tetravalent (Si(IV)) or trivalent aluminum (Al(III)), where the ratios of Al(III) to
Si(IV) are different in different zeolite materials. These elements are connected
through common oxygen atoms (O) and build secondary polyhedral units, the
connection of which creates a crystal lattice. It is this lattice that is responsible for the
main property of zeolite, the ion exchange of cations from the cavity with cations from
the environment. Another important property of zeolite is adsorption of organic
molecules on the surface of zeolite. So far, the non-toxic natural zeolite material
clinoptilolite is used for medical purposes. In this doctoral thesis, differently micronized
clinoptilolite materials were studied: TMAZ (tribomechanically activated clinoptilolite),
PMA (double tribomechanically activated zeolite) and PMAO2 (double
tribomechanically activated zeolite enriched with medical oxygen), and synthetic
zeolite was used as a control compound A. In the first part of the research, the Si/Al
ratio in water and model solutions of the stomach and intestines was determined for
the mentioned materials. After incubation in water or model solutions for 24 h, fractions
were separated by centrifugation at 4000 rpm, 14000 rpm and 140000 rpm. The
supernatants of each fraction were analyzed using electron microscope (scanning
electron microscope, SEM) and electron dispersion spectroscopy (EDS). The particle
shape of the zeolite material supernatant and the Si/Al ratio were consistent with the
zeolite materials in the solid state. In the second part of research, the percentage of
interaction or adsorption of D-glucose on the surface was investigated with SEM in
combination with EDS and quantified with ultra-high resolution liquid chromatography
(UHPLC). The adsorptive properties for D-glucose were validated on healthy mice in
vivo for the zeolite PMA, which showed the best properties of interaction with D
glucose in vitro. The results indicated a decrease in blood glucose levels in mice
pretreated with zeolite PMA. The obtained results show the potential of using zeolite
materials in pathological conditions such as metabolic syndrome or diabetes, which
should be further investigated
Association of pre-treatment CA-125 serum levels and p53 expression with clinicopathological factors and survival of high-grade serous ovarian cancer patients
No full textBackground: Ovarian cancer (OC) it the 7th most common cancer among
women worldwide, while its 5-year survival rate in Croatia amounts to 39%.
Epithelial ovarian cancer (EOC) represents 90% of all ovarian cancers, while
high-grade serous ovarian cancer (HGSOC) is the most common EOC
histological subtype and accounts for 75% of all EOCs. Its lethality is
attributable to the majority of patients being diagnosed at an advanced
stage. There are discrepancies between studies that investigated preoperative serum CA-125 levels as an outcome predictor, while the TP53
gene is ubiquitously mutated in HGSOC. Based on these findings, I have
decided to evaluate pre-treatment serum CA-125 levels together with p53
mutation status and pattern in HGSOC, as well as their correlation. Pretreatment serum CA-125 was also correlated with clinicopathological
variables, while their impact on patient overall survival was also examined.
Methods: A retrospective cohort study was undertaken on 28 HGSOC
patients diagnosed between July 2016 and December 2018 at the
Gynecology and Obstetrics Clinic in Rijeka (Croatia). Pre-treatment serum
CA-125 levels were assessed for their correlation with FIGO stage, p53
mutation pattern, residual tumor (RT) size and type of cytoreductive
surgery. Furthermore, pre-treatment serum CA-125, RT, type of
cytoreductive surgery, chemotherapy regimen and FIGO stage were
examined as potential prognostic factors.
Results: Pre-treatment CA-125 serum levels did not significicantly
correlate with FIGO stage, p53 mutation, RT nor the type of cytoreductive
surgery. The pattern of p53 mutation also had no significant association
with FIGO stage. While pre-treatment CA-125 serum levels, FIGO stage
and RT size were not classified as significant survival predictors, reduced
survival was noted for patients who underwent interval cytoreduction (p =
0.002) and neoadjuvant chemotherapy (p < 0.0001).
Conclusion: Despite pre-treatment CA-125 being abnormal in the majority
of HGSOC patients, it showed no prognostic value, nor significant
association with clinicopathological factors. Similarly, an aberrant p53 is
indeed a characteristic of this HGSOC cohort.Povod i značaj: Karcinom jajnika sedmi je najčešći karcinom kod žena u
svijetu, a petogodišnje preživljenje Hrvatica iznosi 39%. Epitelni karcinom
jajnika predstavlja 90% svih slučajeva karcinoma jajnika, dok serozni
karcinom visokog gradusa čini najčešći histološki podtip epitelnog
karcinoma jajnika (75% svih slučajeva). Stopa smrtnosti vrlo je visoka
zbog toga što se najčešće dijagnosticira u uznapredovalim stadijima.
Prijašnje studije pokušale su evaluirati prognostički značaj serumskih
razina tumorskog markera CA-125 prije ikakvih liječničkih tretmana i
zahvata, ali se njihovi rezultati ne podudaraju. Međutim, mutacije tumor
supresorskog gena TP53 karakteristika su seroznog karcinoma jajnika
visokog gradusa. Bazirajući se na rezultatima prijašnjih istraživanja, u
ovom diplomskom radu evaluirane su pre-operativne razine serumskog CA125 te je analizirana ekspresija proteina p53 u slučajevima seroznog
karcinoma jajnika visokog gradusa. Osim toga, analizirana je i korelacija
ovih dviju varijabli. Nadalje, pre-operativna razina serumskog CA-125
korelirana je i s kliničkopatološkim varijablama te je razmatran njihov
učinak na sveukupno preživljenje pacijentica.
Metode: Provedeno je retrospektivno kohortno istraživanje, koje je
uključivalo 28 pacijentica s dijagnozom seroznog karcinoma jajnika visokog
gradusa koje su liječene između srpnja 2016. i prosinca 2018.godine u
Klinici za ginekologiju i opstetriciju, KBC-a Rijeka. Razmatrana je statistička
korelacija pre-operativnih razina CA-125 s FIGO stadijem, vrstom mutacije
p53, veličinom rezidualnog tumora te s vrstom citoreduktivnog zahvata.
Također, analiziran je i značaj pre-operativnih razina CA-125, veličine
rezidualnog tumora, vrste citoreduktivnog zahvata te kemoterapijskog
režima u prognozi bolesnica.
Rezultati: Pre-operativne razine CA-125 nisu značajno korelirale s FIGO
stadijem, vrstom p53 mutacije, veličinom rezidualnog tumora ni s vrstom
citoreduktivnog zahvata. Vrsta p53 mutacije također nije pokazala
značajnu korelaciju s FIGO stadijem. Dok pre-operativne razine CA-125,
FIGO stadij i veličina rezidualnog tumora nisu svrstani kao značajne
prediktorske varijable, indikacija je suprotna za vrstu citoreduktivnog
zahvata (p = 0.002) te za kemoterapijski režim (p < 0.0001).
Zaključak: Unatoč tome što je pre-operativna CA-125 razina abnormalno
povišena u većini pacijentica sa seroznim karcinomom jajnika visokog
gradusa, nema prognostički značaj niti značajnu korelaciju s
kliničkopatološkim varijablama. Slično, p53 je abnormalno eksprimiran u
većini pacijentica te se radi o potvrđenoj karakteristici ovog histološkog
podtipa