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Brain lipidomics of B4Galnt1 knock out mouse
No full textGangliozidi su amfipatske molekule stanične membrane koje se nalaze u svim tkivima kralježnjaka, a modeliraju mnoge fiziološke procese u organizmu. Imaju važne uloge u razvoju mozga, posebno u održavanju dugotrajne mijelinizacije aksona budući da su složeni gangliozidi GD1a i GT1b aksonski receptori za mijelinu pridruženi glikoprotein (engl. myelin associated
glycoprotein, MAG). Ako se isključi gen B4Galnt1 cijeli organizam, a osobito mozak, ostaje bez sinteze složenih gangliozida. Ulogu složenih gangliozida preuzimaju jednostavni gangliozidi GM3 i GD3, od kojih je GM3 receptor za MAG pa uzrok demijelinizacije ostaje nepoznata. Cilj ovog rada bio je istražiti razliku u lipidomu mozga miša s isključenim B4Galnt1 (engl. knock out, KO) i miša divljeg tipa (engl. wild type, WT) u mladoj i starijoj dobi pomoću slikovne spektrometrije masa (engl. Mass spectrometry imaging, MSI). Degeneracija mijelina prikazana je imunohistokemijskim dokazivanjem ekspresije MAG-a te
histološkim bojenjem mijelina i sukcinat dehidrogenaze (SDH). Rezultati su pokazali povećan stupanj i opseg mijelinizacije u starijih WT miševa, a smanjen u starijih KO miševa. Aktivnost SDH bila je povećana u starijih i KO i WT miševa. MSI analizom te usporedbom KO i WT miševa starije dobi računalnim analizama identificirano je četiri spoja koji su uključeni u metabolički put feroptoze. Poremećaj metaboličke ravnoteže između oligodendrocita i aksona uslijed slabog vezanja GM3 i MAG-a mogao bi imati za posljedicu feroptozu koja dovodi do odumiranja oligodendrocita te potom demijelinizacije. MSI tehnologija pruža novi uvid u razumijevanje molekularne podloge demijelinizacijskih bolesti što bi moglo otvoriti nove strategije u liječenju i dijagnosticiranju istih.Gangliosides are amphipathic cell molecules found in all vertebrate tissues and model many physiological processes in the organism. They play crucial roles in brain development, especially in maintaining long-term axonal myelination
since the complex gangliosides GD1a and GT1b are axonal receptors for myelin-associated glycoprotein (MAG). If the B4Galnt1 gene is knocked down, the whole organism, and especially the brain, remains without the synthesis
of complex gangliosides. The role of complex gangliosides is taken over by simple gangliosides GM3 and GD3, of which GM3 is a receptor for MAG, so the basis for demyelination remains unknown. This study aimed to investigate the
difference in the brain lipidomics of a B4Galnt1 knock out (KO) mouse and wild-type mouse (WT) at a young and older age using mass spectrometry imaging (MSI). Myelin degeneration was demonstrated by immunohistochemical detection of MAG expression and by histological staining of myelin and succinate dehydrogenase (SDH). The results showed an
increased level and volume of myelination in older WT mice and a decrease in older KO mice. SDH activity was increased in both older KO and WT mice. MSI analysis and comparison of KO and WT mice in elderly age identified four
compounds involved in the metabolic pathway of ferroptosis. Disruption of the metabolic balance between ligodendrocytes and axons due to poor binding of GM3 and MAG could result in ferroptosis leading to oligodendrocyte death
and subsequent demyelination. MSI technology provides new insight into the understanding of the molecular basis of demyelinating diseases, which could open new strategies in their treatment and diagnosis
Investigation of the association of polymorphisms in the circadian CLOCK gene and cancer risk using artificial intelligence
No full textCikličke su promjene u brojnim metaboličkim procesima, fiziologiji te ponašanju jedna od glavnih karakteristika koje su specifične za sve žive organizme. Jednim se imenom nazivaju cirkadijalni ritam jer osciliraju u periodu koji traje otprilike 24 sata. Ovaj unutarnji mehanizam ima bitnu ulogu u prilagodbi na konstantne promjene u okolišu, a njegove glavne komponente su cirkadijalni satni geni. Cirkadijalni ritam je na molekularnoj razini, pomoću satnih gena, reguliran putem transkripcijsko
translacijske povratne sprege. Deregulacija u cirkadijalnom ritmu ima veliku ulogu u karcinogenezi te brojnim drugim kliničkim patologijama. Analogno tome, onkogeni procesi direktno remete pravilno funkcioniranje cirkadijalnog ritma te stvaraju okoliš koji pogoduje razvoju karcinoma. Odnedavno sve je veći interes za ispitivanje utjecaja genetičkih varijacija, posebice polimorfizama jednog nukleotida (SNP), u satnim genima na razvoj karcinoma. SNP su jedna od najčešćih genetičkih varijacija u
ljudskom genomu. U ovom istraţivanju prikupljeni su podaci iz studija slučajeva i kontrola koje sadrže frekvencije genotipova određenih polimorfizama na cirkadijalnom CLOCK genu te se na temelju tih podataka istraživala povezanost između genetskih varijacija u navedenom genu te rizika od nastanka karcinoma. Varijacije u genima povezane su sa kompleksnim malignim oboljenjima poput karcinoma te se njihova točna uloga mora još potvrditi dodatnim istraživanjima. Također, algoritmom
umjetne inteligencije (UI), algoritmom k-najbližih susjeda (KNN), predviđala se vrsta karcinoma (karcinom dojke ili klasa - miješani). Algoritam se koristi za klasificiranje ulaznih podataka u određene klase prema najvećoj sličnosti. U našem istraživanju, klasificirali smo prema vrsti karcinoma te je preciznost algoritma bila 99.99%. Temeljem dobivenih rezultata smatramo da bi korištenje UI moglo uvelike olakšati dijagnosticiranje karcinoma, no ovaj pristup tek je u samim počecima te
su potrebna dodatna istraživanja koja će potvrditi vjerodostojnost ove metode.Cyclic changes in numerous metabolic processes, physiology and behavior are one of the main characteristics that are specific to all living organisms. They are called circadian rhythm because they oscillate over a period of approximately 24 hours. This internal mechanism plays an important role in adapting to everchanging environmental stimuli and its main
components are circadian clock genes. On molecular level, circadian rhythtm is controled by clock genes via transcriptional-translational feedback loop. Deregulation in circadian rhythm plays a major role in carcinogenesis and a vast number of clinical pathologies. Analogously, oncogenic processes directly disrupt the proper functioning of the circadian rhythm and create an environment convenient for cancer development. Recently, there has been an emerging interest in examining
the impact of genetic variations in clock genes, particulary single nucleotide polymorphisms (SNPs), on cancer development. SNPs are one of the most common genetic variations in human genome. This study collected data form case-control studies containing the frequencies of genotypes of certain SNPs on circadian CLOCK gene. Based on that data,
relationship between genetic variations and cancer risk was investigated. Variations in genes have been linked to complex malignancies such as cancer, but their exact role needs to be confirmed by additional research. Also, the algorithm of artificial intelligence (AI), the k-nearest neighbours (KNN), was used to predict the type of cancer (breast or mixed). The
algorithm is used to classify the input data into specific classes according to the greatest similarity. In our study, we classified according to the type of cacer and the accuracy of the algorithm was 99.99%. Based on the obtained results, we believe that AI could greatly facilitate the diagnostic of cancer, but this approach is only in its beginnings and further research is needed to confirm the credibility of this method
Antiproliferative and antioxidant effects of structural analogues of a marine natural compound granulatamide B
No full textJedna od strategija prilagodbi ekstremnom okolišu koju su razvili morski organizmi je sinteza sekundarnih metabolita. Ti spojevi nisu neophodni za rast, razvoj i razmnožavanje organizma već imaju adaptivnu, obrambenu, signalnu ili neku drugu ulogu. Sjedilački organizmi poput spužvi i koralja izvor su brojnih biološki aktivnih spojeva zbog čega je bioprospecting mora važan za farmaceutsku, kozmetičku i druge industrije. Spojevi izolirani iz koralja Eunicella sp. mogu se klasificirati u tri skupine: terpenoidi, steroli te alkalodi i nukleozidi. Jedan od tih spojeva je indolni alkaloid granulatamid B koji je pokazao potencijalan inhibicijski učinak prema različitim ljudskim tumorskim stanicama. S obzirom da je važnost strukture indolnog prstena već prepoznata od strane farmaceutske industrije, sintetizirano je pet strukturnih analoga granulatamida B kojima su ispitani antioksidativni i antiproliferativni učinci koristeći redom ABTS i DPPH te MTT eseje. Spojevi su sintetizirani konjugacijom triptamina i nezasićenih masnih kiselina s ciljem nastanka amidne veze. Korištene masne kiseline su arahidonska, retinoična, palmitoleinska, oleinska i alfa-linolenska kiselina. Sinteza spojeva je potvrđena infracrvenom (IR) spektroskopijom, ultraljubičastom/vidljivom (UV-Vis) spektrofotometrijom, spektroskopijom nuklearne magnetske rezonancije (NMR) i spektrometrijom masa (MS). ABTS esej je ukazao na smanjenje antioksidativne aktivnosti kao posljedica vezanja masno-kiselinskog lanca osim u slučaju N-retinoiltriptamina. Potonje se može objasniti uz pretpostavku doprinosa sustava konjugiranih dvostrukih veza retinoične kiseline u inhibiciji spomenutog radikala. Nasuprot toga, DPPH esejom nisu uspješno utvrđena antioksidativna svojstva sintetiziranih spojeva. Dodatno, MTT esejom je uočeno kako svi sintetizirani spojevi pokazuju inhibitorni učinak prema staničnim linijama, među kojima je najaktivniji bio N-retinoiltriptamin. S druge strane, derivat alfa-linolenske kiseline je imao najslabiji učinak na normalnu staničnu liniju zbog čega bi oba navedena spoja trebalo dodatno strukturno modificirati.One of the adaption strategies to the extreme environment developed by the marine organism is the synthesis of secondary metabolites. These compounds are not necessary for the growth, development and reproduction of the organisms but have an adaptive, defensive, signaling or some other roles. Sessile marine organisms, such are sponges and corals, present a source of numerous biologically active compounds, making sea bioprospecting important for the pharmaceutical, cosmetic and other industries. Compounds isolated from the corals of Eunicella sp. can be classified into three groups: terpenoids, sterols as well as alkaloids and nucleosides. One of these compounds is the indole alkaloid granulatamide B which has exerted a potential inhibitory activity towards various human cancer cell lines. Since the importance of the indole ring structure has already been recognized by the pharmaceutical industry, five structural analogs of granulatamide B have been synthesized, followed by evaluation of antioxidant and antiproliferative activities by DPPH/ABTS and MTT assays, respectively. The fatty acids which were used are: arachidonic, retinoic, palmitoleic, oleic and alpha-linolenic acids. The synthesis of the compounds was confirmed by infrared (IR) spectroscopy, ultraviolet-visible (UV-Vis) spectrophotometry, nuclear magnetic resonance (NMR) spectrometry and mass (MS) spectrometry. The antioxidant activity of synthesized compounds decreased as shown by ABTS assay as a consequence of fatty acids binding, except for N-retinoyltryptamine. The latter can be explained by the presence of conjugated double bonds in retinoic acid. Contrary, none of the examined analogues has been active in inhibition of DPPH radical. Furthermore, the results of MTT assay revealed that all analogues displayed antiproliferative activity against cancer cell lines, with N-retinoyltryptamine being the most active. However, derivative of alpha-linolenic acid was the least active against normal, non-transformed cell lines, therefore, both mentioned compounds should be considered for the further structural modification
Relationship between genetic variation of the circadian NPAS2 gene and cancer risk using artificial intelligence
No full textCirkadijalni ritam je prirodan ritam u svim živim organizmima koji se ponavlja približno svaka 24 sata. Uloga ovoga ritma je regulacija ciklusa spavanja i budnosti, te psiholoških, metaboličkih i fizioloških procesa. Na molekularnoj razini kontroliran je satnim genima pomoću autoregulacijskih povratnih sprega. Deregulacija ovog ritma dovodi do različitih patoloških stanja uključujući dijabetes, metaboličke sindrome, depresiju i rak. Ekspresija satnih gena ili disrupcija cirkadijalnog ritma novi su i potencijalno promjenjivi čimbenici koji utječu na rizik od razvoja raka. Polimorfizam jednog nukleotida (engl. Single nucleotide polymorphism - SNP) označava promjenu jedne baze u DNA sekvenci što znači da se na jednome položaju u sekvenci mogu naći dva nukleotida. U ovome su istraživanju prikupljeni podaci iz 12 studija slučajeva i kontrola koje su obuhvaćale 7791 pacijenta koji boluje od raka i 7938 zdravih kontrola. U studijama su se nalazili podaci o frekvencijama genotipova SNP-ova NPAS2 gena. Istraživanje je bilo fokusirano na dva SNP-a: rs2305160 i rs17024926. Na temelju prikupljenih podataka, statističkim je metodama istražena povezanost između genetskih varijacija NPAS2 gena i rizika za razvoj raka. Rezultati su pokazali statistički značajnu poveznicu između rs2305160 i raka. Detaljnija analiza provedena prema vrsti raka pokazala je da rs2305160 može imati protektivnu ulogu od pojave raka dojke, dok rezultati kod raka prostate nisu bili statistički značajni. Za rs17024926 polimorfizam nije pronađena značajna povezanost kod niti jednog ispitanog modela nasljeđivanja. Konačno, razvijen je algoritam umjetne inteligencije koji na temelju unesenih podataka predviđa vrstu raka o kojoj se radi (rak dojke ili ostale vrste).Circadian rhythm is a natural process in all living organisms which repeats approximately every 24 hours. The role of this rhythm is to regulate the sleep–wake cycle, our psychological, metabolic and physiological processes. On molecular level, circadian rhythm is controlled by so-called clock genes via autoregulatory feedback loops. Deregulation of these
rhythms leads to a number of pathological conditions including diabetes, metabolic syndrome, depression and cancer. For that reason, expression of clock genes or disruption of circadian rhythms is emerging as a novel and potentially modifiable cancer risk factor. Single nucleotide polymorphism (SNP) stands for single base change in a DNA sequence, with a usual
alternative of two possible nucleotides at a given position. In this study, the data from 12 case-control studies with 7791 cancer patients and 7938 cancer-free controls were collected. Those studies contained available genotype frequencies of the SNPs in NPAS2 clock gene. The study was focused on two SNPs: rs2305160 and rs17024926. Based on that data, the
association between genetic variations in NPAS2 gene and the risk of developing cancer was investigated using statistical tools. The results showed statistically significant associations of rs2305160 with cancer risk. Stratified analysis by cancer type indicated that rs2305160 may decrease the risk of breast cancer, whereas results for prostate cancer did not have
statistical importance. For rs17024926 polymorphism, there was no significant association under any genetic model. Furthermore, Artificial Intelligence (AI) algorithm was developed to predict the type of cancer (breast or mixed)
Conducting a survey among the general population on vaccination with the future vaccine against COVID-19
No full textCOVID-19 je bolest uzrokovana SARS-CoV-2 virusom koji je prvi puta
detektiran u kineskoj provinciji Wuhan 2019. godine. S obzirom na činjenicu
da je riječ o virusu koji se vrlo brzo širi, nekoliko farmaceutskih tvrtki razvilo
je cjepiva. Cilj ovog rada je provesti anketu kojom se procjenjuje
zabrinutost i stopa prihvaćanja COVID-19 cjepiva među općom populacijom
na području Republike Hrvatske. Također, želi se provjeriti je li potrebno
dodatno informirati javnost o sigurnosti dostupnih cjepiva. Analiza o
zainteresiranosti za cijepljenje provodila se ovisno o dobi i spolu, stupnju
obrazovanja, statusu zaposlenosti i poslovnom sektoru. Idući set pitanja
odnosio se na COVID-19, odnosno jesu li ispitanici preboljeli ovu bolest ili
netko od njihovih bližnjih. Posljednjim setom pitanja ispitivali su se stavovi
o cijepljenju i dostupnim cjepivima. Istraživanje je pokazalo da su mišljenja
ispitanika podijeljena točnije 49,1% se želi cijepiti i 50,9% odbija
cijepljenje. Najviše ih brinu nuspojave te brzina razvoja cjepiva, a na
cijepljenje bi pristali tek nakon određenog vremenskog perioda kad bi se
uvjerili da su cjepiva sigurna. Najviše sumnjaju oni sa završenom srednjom
školom, a zaposleni su u trgovačkom sektoru. Analizom pristiglih anketa
moglo se uočiti kako ispitanici žele sami birati vrstu cjepiva te najviše
ispitanika izabralo bi cjepivo tvrtke Pfizer/BioNTech, točnije 48,8%. Dok,
građani Republike Hrvatske imaju najmanje povjerenja u cjepiva tvrtki
AstraZeneca te Johnson & Johnson.COVID-19 is a disease caused by the SARS-CoV-2 virus that was first
detected in the Chinese province of Wuhan in 2019. Due to the fact that it
is a virus that spreads very quickly, several pharmaceutical companies have
developed vaccines. The aim of this paper is to conduct a survey to assess
the concerns and acceptance rates of COVID-19 vaccines among the
general population in the Republic of Croatia. It also wants to check whether
it is necessary to further inform the public about the safety of available
vaccines. Vaccination interest analysis was conducted depending on age
and gender, level of education, employment status, and business sector.
The next set of questions referred to COVID-19, whether the respondents
overcame this disease or someone close to them. The last set of questions
examined attitudes about vaccination and available vaccines. The research
showed that the opinions of the respondents were divided, namely 49.1%
want to be vaccinated and 50.9% refuse to be vaccinated. They are most
concerned about the side effects and the speed of vaccine development,
and they would only agree to the vaccination after a certain period of time
when they would be convinced that the vaccines are safe. Those who have
finished high school, and are employed in the trade sector, are the most
suspicious. The analysis of the received surveys showed that the
respondents want to choose the type of vaccine themselves, and most
respondents would choose the Pfizer / BioNTech vaccine, more precisely
48.8%. While, the citizens of the Republic of Croatia have the least trust in
vaccines from AstraZeneca and Johnson & Johnson
The role of redox balance in development of chemoresistance
No full textKemoterapeutski lijekovi imaju sposobnost stvaranja reaktivnih vrsta koje se vežu za određene strukture unutar stanice raka i potiču staničnu smrt. Redoks homeostaza stanica raka poremećena je zbog kontinuirane izloženosti kemoterapijskim sredstvima, što može rezultirati promjenom sposobnosti stanice da se nosi s prekomjernom razinom reaktivnih vrsta. Kako bi se nosile s oksidativnim stresom, stanice raka kontroliraju povišenu razinu reaktivnih vrsta pojačavajući aktivnost staničnih antioksidativnih sustava. Antioksidativni strojevi sprječavaju staničnu smrt i potiču razvoj tumora dok su istodobno stanice otpornije na egzogeni stimulans poput kemoterapije i radioterapije. Pojačani antioksidativni kapacitet stanica raka ključna je odrednica razvoja rezistencije na kemoterapiju. Otpornost na lijekove razvija se u gotovo svih bolesnika s rakom debelog crijeva, što dovodi do smanjenja terapijske učinkovitosti antikancerogenih sredstava. Nuklearni faktor eritroid 2 povezan s faktorom 2 (engl. nuclear factor erythroid 2-related factor 2, Nrf2) glavni je regulator neutraliziranja staničnih reaktivnih kisikovih vrsta (engl. reactive oxygen species, ROS) i obnavljanja redoks ravnoteže. Nrf2 regulira ekspresiju antioksidativnih gena, gena za metabolizam i detoksikaciju te gena koji kodiraju transportere. Aktivacija Nrf2 može dovesti do progresije raka, invazije, metastaza i kemorezistencije. Nrf2 igra ključnu ulogu u rezistenciji na kemoterapiju potičući metabolizam lijekova ili izbacivanje lijekova iz stanice. Ekspresija Nrf2 često je povećana u različitim vrstama kemorezistentnih stanica raka, stoga blokiranje aktivnosti Nrf2 u malignim stanicama predstavlja značajan pristup za prevenciju raka.Chemotherapeutic drugs have the ability to produce reactive species that bind to certain structures within a cancer cell and promote cell death. The redox homeostasis of cancer cells is disrupted due to continuous exposure to chemotherapeutic agents. To overcome oxidative stress, cancer cells control elevated levels of reactive species by enhancing the activity of cellular antioxidant systems. Highly compatible antioxidant machinery prevent cell death and promote tumor development, while at the same time cells are more resistant to exogenous stimulants such as chemotherapy and radiotherapy. Enhanced antioxidant capacity of cancer cells is a key determinant of the development of resistance to chemotherapy. Drug resistance develops in almost all patients with colon cancer leading to a decrease in the therapeutic efficacy of anticancer agents. Nuclear factor erythroid 2-related factor 2 (Nrf2) is a major regulator of neutralizing cellular ROS and restoring redox balance. Nrf2 regulates the expression of antioxidant genes, metabolic and detoxification genes, and transporters. Due to the dual role of Nrf2 in cancer cells, activation of Nrf2 can lead to cancer progression, invasion, metastasis, and chemoresistance. Nrf2 plays a key role in chemotherapy resistance by stimulating drug metabolism or drug efflux. Expression of Nrf2 is often increased in several types of chemoresistant cancer cells, so blocking Nrf2 activity in malignant cells represents a significant approach to cancer prevention
Intellectual property and access to medicines
No full textMilijuni ljudi širom svijeta nemaju pristup lijekovima koji su im potrebni za liječenje bolesti i očuvanje kvalitete života. Strogi patentni režimi otežavaju dostupnost lijekova u svijetu stvaranjem monopola kojim farmaceutske tvrtke održavaju visoke cijene lijekova koje su izvan dosega velikom broju ljudi. Kriza bolesti HIV/AIDS, koja je krajem devedesetih godina prošlog
stoljeća pogodila najmanje razvijene zemlje i time izazvala smrt velikog broja ljudi za koje HIV lijekovi nisu bili dostupni, izazvala je val promjena. Članice Svjetske trgovinske organizacije (WTO) nastojale su potpisivanjem Sporazuma TRIPS, a kasnije i Deklaracijom iz Dohe, definirati odredbe kojima se nastojalo uskladiti potrebe javnih i komercijalnih zdravstvenih interesa, a da se pritom smanji negativan utjecaj intelektualnog vlasništva na dostupnost lijekova. Primjenjujući mehanizme obveznog licenciranja, paralelnog uvoza te LDC farmaceutskog odricanja, postignut je impresivan napredak u pristupu lijekovima za HIV/AIDS što daje nadu da se odredbe definirane Sporazumom TRIPS mogu koristiti za pristup i ostalim lijekovima.
Nakon što je prošla gotovo trećina vremenskog razdoblja namijenjenog za ostvarivanje ciljeva održivog razvoja svijet se na početku 2020. godine suočio s pandemijom bolesti COVID-19. Unatoč činjenici da su cjepiva za ovu bolest vrlo brzo pronašla put do tržišta njihova je raspodjela u svijetu izuzetno neravnomjerna. Hoće li se farmaceutske tvrtke odreći patentnih
prava u svrhu javnog zdravstva ili će pandemiju iskoristiti za ostvarivanje profita tek treba vidjeti. No, jedno je sigurno: uspije li se pobijediti COVID19 kriza globalnim udruživanjem zdravstvenih sustava i ravnomjernom raspodjelom cjepiva to se može primijeniti i na pitanje dostupnosti lijekova za ostale bolesti u najmanje razvijenim zemljama svijeta.Millions of people around the world do not have access to the medicines they need to treat diseases and maintain the quality of life. Strict patent protection regimes hinder access to medicines in the world by creating a
monopol by which pharmaceutical companies maintain high drug prices that are beyond the reach of a large number of people in the least developed countries (LDC). The HIV/AIDS crisis, which hit the LDCs in the late 1990s and caused death
of a large number of people for whom HIV drugs were not available, has triggered a wave of change. The World Trade Organization (WTO) members sought to define provisions by the TRIPS Agreement, and later the Doha Declaration, to reconcile the needs of public and commercial health interests while reducing the negative impact of intellectual property on the availability of medicines. Implementation of mandatory licensing, parallel import and LDC pharmaceutical waiver mechanisms, allowed an impressive progress in access to HIV/AIDS medicines. That gives hope that the provisions defined by the TRIPS Agreement can be used to access other medicines. After almost a third of the time period dedicated to achieving the Sustainable Development Goals, the world faced a pandemic of the COVID19 disease in early 2020. Despite the fact that vaccines for this disease have found the fast way to the market, their distribution in the world is
extremely uneven. Whether pharmaceutical companies will give up patent rights for public health purposes or use the pandemic to make a profit remains to be seen. But one thing is certain: if the COVID-19 crisis can be defeated by globally pooling health systems and distributing the vaccines evenly, this can also be applied to the problem of the availability of
medicines for other diseases in LDC around the world
Establishment of Long-Term Primary Cortical Neuronal Cultures From Neonatal Opossum Monodelphis domestica
No full textPrimary dissociated neuronal cultures have become a standard model for studying central nervous system (CNS) development. Such cultures are predominantly prepared from the hippocampus or cortex of rodents (mice and rats), while other mammals are less used. Here, we describe the establishment and extensive characterization of the primary dissociated neuronal cultures derived from the cortex of the gray South American shorttailed opossums, Monodelphis domestica. Opossums are unique in their ability to fully regenerate their CNS after an injury during their early postnatal development. Thus, we used cortex of postnatal day (P) 3–5 opossum to establish long-surviving and nearly pure neuronal cultures, as well as mixed cultures composed of radial glia cells (RGCs) in which their neurogenic and gliogenic potential was confirmed. Both types of cultures can survive for more than 1 month in vitro. We also prepared neuronal cultures from the P16–18 opossum cortex, which were composed of astrocytes and microglia, in addition to neurons. The long-surviving opossum primary dissociated neuronal cultures represent a novel mammalian in vitro platform particularly useful to study CNS development and regeneration
Integrative analysis of squamous cell lung cancer by using omics approach
No full textTumor pluća je među najčešće dijagnosticiranim malignim oboljenjima na svijetu, sa
visokom stopom smrtnosti. Tumor pluća skvamoznih stanica je relativno česti podtip tumora
pluća koji je dijagnosticiran u 20-30 % slučajeva, a od njega je češće dijagnosticirani samo
adenokarcinom pluća. Pacijenti oboljeli od tumora pluća skvamoznih stanica ne pokazuju
uniformnost u kliničkom ishodu, i liječe se na jednak način. Obzirom da profil genskih
varijanti, kao i specifični profil ekspresije gena, još uvijek nije u potpunosti razjašnjen u
ovom tipu tumora, pacijenti se još uvijek uglavnom liječe sistemskom kemoterapijom.
Pretpostavka je da bi bolja molekularna karakterizacija tumora pluća skvamoznih stanica
prema biološkim karakteristikama, mogla dovesti do personaliziranog pristupa liječenju
pacijenata, boljeg predviđanja odgovora na terapiju i u konačnici boljeg ishoda liječenja ove
vrste tumora. Stoga su potrebna daljnja istraživanja koja će doprinijeti boljoj molekularnoj
karakterizaciji ove vrste tumora.
Cilj ove doktorske disertacije je analiza i razumijevanje profila ekspresije molekula
mRNA, proteina i miRNA u tumoru pluća skvamoznih stanica s krajnjom svrhom
identifikacije ključnih molekula u tumorigenezi ove bolesti.Lung cancer is among the most commonly diagnosed malignancies in the world, and
has a high mortality rate. Squamous cell lung cancer is a relatively common subtype of lung
cancer, diagnosed in 20-30% of cases, of which only lung adenocarcinoma is more
commonly diagnosed. Patients with squamous cell lung cancer do not show uniformity in
clinical outcome, and are treated in the same way, regardless of morphological subtype.
Because the profile of gene variants, as well as the specific profile of gene expression, is still
not fully elucidated in this type of tumor, patients are still mostly treated with systemic
chemotherapy. It is hypothesized that better molecular characterization of squamous cell lung
cancer by biological characteristics could lead to a personalized approach to patient
treatment, better prediction of response to therapy, and ultimately a better treatment outcome
for this type of tumor. Therefore, further research is needed that will contribute to better
molecular characterization of this type of tumor. The aim of this doctoral dissertation is to
analyze and understand the expression profile of mRNA, protein and miRNA molecules in
squamous cell lung cancer with the ultimate goal of identifying key molecules in the
tumorigenesis of this disease
Effect of murine cytomegalovirus on cellular protein expression for endosomal tubulation regulation
No full textCitomegalovirus (CMV) u inficiranim stanicama uzrokuje opsežnu reorganizaciju sekretornih i endosomalnih membrana u svrhu stvaranja citoplazmatskog odjeljka za sklapanje viriona – cVAC (AC) (eng. cytoplasmic viral assembly compartment). Tijekom tog procesa, virus u stanici mijenja dinamiku i smještaj membranskih struktura djelujući na izražaj i aktivaciju regulatornih proteina koji se mobiliziraju na njihovu površinu. Rab10 GTPaza je jedan od mnogih takvih regulatornih proteina, a nalazimo ga na sučelju ranih endosoma, EE (eng. early endosomes) i endoplazmatskog reciklirajućeg odjeljka, ERC (eng. endoplasmic recycling compartment). U ovom diplomskom radu smo istražili učinak infekcije mišjim citomegalovirusom (MCMV) na izražaj i unutarstanični smještaj Rab10 i njegovih efektora, odnosno regulatora (Rab5, MICAL-L1, EHBP1, ACAP1, ACAP2, EHD1, EHD3) na inficiranim mišjim fibroblastima. Pokazali smo da se izražaj ispitivanih proteina ne mijenja tijekom infekcije, ali se Rab10 GTPaza i njegov efektor EHBP1 u kasnoj fazi infekcije nalaze nakupljeni u unutrašnjem dijelu AC (iAC). Prema tome, možemo zaključiti da MCMV infekcija uzrokuje reorganizaciju endosomalnih membrana na kojima se nalazi Rab10, ali ne mijenjajući njihov izražaj.Cytomegalovirus (CMV) causes extensive reorganization of the secretory and endosomal membranes in infected cells with a purpose of cytoplasmic viral assembly compartmens (cVAC/AC) formation. During that proces, CMV rearrange and change the dynamics of cellular membranes. One of the possible mechanisms involves targeting the expression of regulatory proteins that are activated and mobilized at surfaces of intracellular compartments. Rab10 GTPase represents one of many such proteins and can be found at the interface of early endosomes, EE and endoplasmic recycling compartment, ERC. In this thesis we have investigated the impact of infection with murine CMV (MCMV) on the Rab10 GTPase and its effectors and regulators (Rab5, MICAL-L1, EHBP1, ACAP1, ACAP2, EHD1, EHD3) expression and localization in the infected mouse fibroblasts. We have shown that their total expression do not change in the comparison with uninfected cells, but Rab10, and its effector, EHBP1, can be found in pericentriolar clusters inside AC (iAC) in the late phase of infection. Therefore, we can conclude that MCMV infection cause reorganization of Rab10 positive membranes, but without influence on their total expression