imagine (Institute of molecular genetics and genetic engineering)
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Polyenes in Medium Chain Length Polyhydroxyalkanoate (mcl-PHA) Biopolymer Microspheres with Reduced Toxicity and Improved Therapeutic Effect against Candida Infection in Zebrafish Model
Get PDFImmobilizing antifungal polyenes such as nystatin (Nys) and amphotericin B (AmB) into biodegradable formulations is advantageous compared to free drug administration providing sustained release, reduced dosing due to localized targeting and overall reduced systemic drug toxicity. In this study, we encapsulated Nys and AmB in medium chain length polyhydroxyalkanoates (mcl-PHA) microspheres (7-8 mu m in diameter). The obtained formulations have been validated for antifungal activity in vitro against a panel of pathogenic fungi including species of Candida, Aspergillus, Microsporum and Trichophyton genera and toxicity and efficacy in vivo using the zebrafish model of disseminated candidiasis. While free polyenes, especially AmB, were highly toxic to zebrafish embryos at the effective (MIC) doses, after their loading into mcl-PHA microspheres, inner organ toxicity and teratogenicity associated with both drugs were not observed, even at 100 x MIC doses. The obtained mcl-PHA/polyene formulations have successfully eradicated C. albicans infection and showed an improved therapeutic profile in zebrafish by enhancing infected embryos survival. This approach is contributing to the antifungal arsenal as polyenes, although the first broad-spectrum antifungals on the market are still the gold standard for treatment of fungal infections
Vitamin B Complex and Experimental Autoimmune Encephalomyelitis -Attenuation of the Clinical Signs and Gut Microbiota Dysbiosis
Get PDFThe present study aimed to investigate the neuroprotective effects of the vitamin B complex (B1, B2, B3, B5, B6, and B12-VBC), by studying the changes in the femoral nerve, quadriceps muscle, popliteal lymph nodes and gut microbiota in the rat model of multiple sclerosis, experimental autoimmune encephalomyelitis (EAE). VBC treatment attenuated clinical signs of EAE during the disease, and reduced the duration of EAE thereby contributing to a faster recovery. In VBC-treated EAE rats, a significant decrease in nerve and muscle nuclear density was revealed during the onset period of the disease, while a marked increase was detected at the end of the disease, compared with untreated EAE rats. In the lymph nodes of VBC-treated EAE rats, a fewer number of lymphoid follicles in the cortical area and smaller epithelioid granulomas were detected. The changes in microbiota composition were examined using 16S rRNA gene sequencing and bioinformatics analysis, which revealed the potential of VBC treatment in establishing and/or maintaining gut microbiota homeostasis. Finally, the present study demonstrated that VBC treatment ameliorated the cellular changes in the affected peripheral nerve, muscles innervated by this nerve, and the gut microbiota dysbiosis which occurred during the EAE
Novel RclSAR three-component system regulates expression of the intI1 gene in the stationary growth phase
No full textThe rapid and appropriate response of Pseudomonas spp. to environmental fluctuations has been enabled by numerous signal transduction regulatory systems. Regulatory systems in Pseudomonas aeruginosa are organized in a complex network which provides quick and fine-tuned cellular response through regulation of virulence and antibiotic resistance determinants production. Mobile integrons represent genetic elements included in the rapid dissemination of multiple antibiotic resistance determinants. The key factor of integron dynamics is enzyme integrase. So far, global regulators LexA, RpoS and PsrA have been recognized as regulators of the intI1 transcription. In this study, we discovered novel activator of the intI1 transcription, sensor kinase RclS, in Pseudomonas putida WCS358. This regulation is limited to stationary growth phase and appears to be indirect, at least through regulation of the rpoS expression. Sensor kinase RclS is a part of novel three-component system Rcl (Roc-like) together with two response regulators, RclR and RclA. RclS acted as a negative regulator of the rclA transcription, while the role in the rclR transcription regulation could not be defined. The RclSAR regulatory system seems to be a part of complex intI1 regulatory network which includes major stress response (SOS and RpoS) regulons
Expression Profiles of Long Non-Coding RNA GAS5 and MicroRNA-222 in Younger AML Patients
Get PDFAcute myeloid leukemia (AML) is a heterogeneous malignant disease both on clinical and genetic levels. AML has poor prognosis and, therefore, there is a constant need to find new prognostic markers, as well as markers that can be used as targets for innovative therapeutics. Recently, the search for new biomarkers has turned researchers' attention towards non-coding RNAs, especially long non-coding RNAs (lncRNAs) and micro RNAs (miRNAs). We investigated the expression level of growth arrest-specific transcript 5 (GAS5) lncRNA in 94 younger AML patients, and also the expression level of miR-222 in a cohort of 39 AML patients with normal karyotype (AML-NK), in order to examine their prognostic potential. Our results showed that GAS5 expression level in AML patients was lower compared to healthy controls. Lower GAS5 expression on diagnosis was related to an adverse prognosis. In the AML-NK group patients had higher expression of miR-222 compared to healthy controls. A synergistic effect of GAS5(low)/miR-222(high) status on disease prognosis was not established. This is the first study focused on examining the GAS5 and miR-222 expression pattern in AML patients. Its initial findings indicate the need for further investigation of these two non-coding RNAs, their potential roles in leukemogenesis, and the prognosis of AML patients
High risk population screening for Fabry disease in hemodialysis patients in Vojvodina: Pilot study
Get PDFUvod/Cilj Fabrijeva bolest (FB) X-vezana je lizozomna bolest skladištenja koja se razvija kao posledica mutacije gena alfa-galaktozidaze A (GLA). Postoji više od 1080 poznatih varijanti gena GLA. Neki od njih su patogeni, ali većina ih je benigna ili predstavljaju genetsku promenu koja se može klasifikovati kao genetska varijanta nepoznatog značaja ili jednostavno predstavljati genetski polimorfizam. Postoje dve glavne karakteristike FB, klasični oblik i kasnije varijante bolesti. Glavni ciljni organi kod bolesnika sa ovom bolešću su bubrezi, srce i nervni sistem. Imajući u vidu činjenicu da je FB retka bolest, najbolji način za aktivnu pretragu bolesnika je skrining populacije visokog rizika, nakon čega bi trebalo izvršiti porodični skrining za svaki slučaj probanda. Metode U ovom radu predstavljamo rezultate multicentrične pilot studije koja predstavlja nalaze skrininga bolesnika na hemodijalizi na FB u šest hemodijaliznih centara u Vojvodini. Rezultati Identifikovan je jedan bolesnik sa benignom mutacijom i 16 bolesnika sa genetskim polimorfizmom gena GLA. Utvrdili smo da genetske promene na genu GLA mogu biti česte, ali su veoma retko od kliničkog značaja i retko dovode do manifestacija FB. Zaključak Rezultati ove skrining studije će NAM omogućiti uvid u prevalenciju FB u hemodijaliznoj populaciji i usmeriti naš budući rad.Introduction/Objective Fabry disease (FD) is an X-linked lysosomal storage disease that develops as a consequence of mutation in the Alpha-galactosidase A (GLA) gene. There are more than 1080 known variants in the GLA gene. Some of them are pathogenic, but most of them are benign or represent the genetic change that can be classified as a genetic variant of unknown significance or simply be a representation of genetic polymorphism. There are two main features of FD, classic form and late-onset variants of disease. The main target organs in patients with FD are the kidneys, heart, and nervous system. Bearing in mind the fact that FD is a rare disease, the best way for active searching of patients is high-risk population screening, after which family screening for every proband case should be performed. Methods In this paper, we present results of a multicentric pilot study that represents findings from the screening of hemodialysis patients for FD in six hemodialysis units in Vojvodina. Results We have found one patient with benign mutation and 16 patients with genetic polymorphisms in GLA gene. We have learned that genetic changes in GLA gene can be frequent, but very rarely are of clinical significance and lead to manifestations of FD
Discovery of 1-Benzhydryl-Piperazine-Based HDAC Inhibitors with Anti-Breast Cancer Activity: Synthesis, Molecular Modeling, In Vitro and In Vivo Biological Evaluation
Get PDFIsoform-selective histone deacetylase (HDAC) inhibition is promoted as a rational strategy to develop safer anti-cancer drugs compared to non-selective HDAC inhibitors. Despite this presumed benefit, considerably more non-selective HDAC inhibitors have undergone clinical trials. In this report, we detail the design and discovery of potent HDAC inhibitors, with 1-benzhydryl piperazine as a surface recognition group, that differ in hydrocarbon linker. In vitro HDAC screening identified two selective HDAC6 inhibitors with nanomolar IC50 values, as well as two non-selective nanomolar HDAC inhibitors. Structure-based molecular modeling was employed to study the influence of linker chemistry of synthesized inhibitors on HDAC6 potency. The breast cancer cell lines (MDA-MB-231 and MCF-7) were used to evaluate compound-mediated in vitro anti-cancer, anti-migratory, and anti-invasive activities. Experiments on the zebrafish MDA-MB-231 xenograft model revealed that a novel non-selective HDAC inhibitor with a seven-carbon-atom linker exhibits potent anti-tumor, anti-metastatic, and anti-angiogenic effects when tested at low micromolar concentrations
Усавршавање протокола за изолацију ДНК високе молекуларне масе из кичице (Centaurium erythraea)
Get PDFBiljka kičica (Centaurium erythraea) je interesantna ne samo zbog svojih
lekovitih svojstava, već i zbog izuzetne razvojne plastičnosti i regenerative
sposobnosti u in vitro kulturi. Kako bi identifikovali gene uključene u
morfogenetske procese, sekundarni metabolizam i odgovore na stres, pored
sekvenciranih transkriptoma1 je potreban i sekvenciran genom. Za sekvenciranje
genoma najmodernijom PacBio tehnologijom, neophodna je veoma čista
visokomolekularna DNK, čijih 90% čine lanci > 20 kb. DNK je izolovana iz
mladih listova diploidnog varijeteta kičice gajene in vitro, korišćenjem četiri
različita protokola: (1) Quick-DNA HMW MagBead Kit (Zymo Research, #D6060);
(2) modifikovani CTAB protokol;2 (3) protokol za izolaciju bakterijske DNK
prilagođen biljnim tkivima i (4) izmenjen protokol za izolaciju RNK iz bora.3
Kvalitet DNK izolata je proveravan spektrofotometrijski Nano Drop aparatom,
običnom elektroforezom, pulsnom elektroforezom i Femto Pulsnom analizom.
Protokol (1) daje veoma niske prinose, bilo kad se koristi samostalno ili kao
vid prečišćavanja DNK izolovane na drugi način. Protokolom (3) se dobija visok
prinos DNK nezadovoljavajućeg kvaliteta. Najkvalitetniju DNK u količinama
dovoljnim za sekvenciranje su dali protokoli (2) i (4).Биљка кичица (Centaurium erythraea) је интересантна не само због својих
лековитих својстава, већ и због изузетне развојне пластичности и регенеративе
способности у in vitro култури. Како би идентификовали гене укључене у
морфогенетске процесе, секундарни метаболизам и одговоре на стрес, поред
секвенцираних транскриптома1 је потребан и секвенциран геном. За секвенцирање
генома најмодернијом PacBio технологијом, неопходна је веома чиста
високомолекуларна ДНК, чијих 90% чине ланци > 20 kb. ДНК је изолована из
младих листова диплоидног варијетета кичице гајене in vitro, коришћењем четири
различита протокола: (1) Quick-DNA HMW MagBead Kit (Zymo Research, #D6060);
(2) модификовани ЦТАБ протокол;2 (3) протокол за изолацију бактеријске ДНК
прилагођен биљним ткивима и (4) измењен протокол за изолацију РНК из бора.3
Квалитет ДНК изолата је провераван спектрофотометријски Nano Drop апаратом,
обичном електрофорезом, пулсном електрофорезом и Фемто Пулсном анализом.
Протокол (1) даје веома ниске приносе, било кад се користи самостално или као
вид пречишћавања ДНК изоловане на други начин. Протоколом (3) се добија висок
принос ДНК незадовољавајућег квалитета. Најквалитетнију ДНК у количинама
довољним за секвенцирање су дали протоколи (2) и (4).Knjiga sažetaka: Treći Kongres biologa Srbije, Zlatibor, Srbija 21 - 25. 9. 2022
Natural products as a base for ecological sustainable preparations (NATBIOPREP)
Get PDFPollution is a reality of modern life and affects every individual. The skin, the largest human
organ, is the first line of defense against air pollution, the harmful effects of the sun’s
ultraviolet (UV) radiation, and pathogenic microorganisms.
NATBIOPREP is focused on the development of innovative products for (1) healthy skin damage
prevention, and (2) prevention of infection development in already damaged skin providing
solutions based on a combination of purified enzymes originating from beneficial
environmental bacteria and novel technologies of supercritical extraction of medicinal plants.
Combination of these advanced technologies will result in products with a synergistic effect in
skin protection. Long-acting topical formulations will allow stable enzyme action and gradual
release of plant extracts.Principal Investigator: Dr Lidija Senerovic, IMGGEDuration period: 2022-202
Molecular profile of thymoma
Get PDFTimomi, timusni karcinomi (TC) i timusni neuroendokrini kanceri su najčešce neoplazije lokalizovane
u timusu i pripadaju retkim formama tumora. Timomi su indolentne forme tumora, a klasifikovani su u A,
AB, B1,B2, B3 kategorije. Novije genetičke studije timoma pokazale su da je jedan od dominantnih markera
ove bolesti , gen GTF2I sa najčešćim ’drajver’ varijantama Chr7 c.1211T> A, p. Leu404His i c.1271T>A, p.
Leu424His. Pored GTF2I, u patogenezu timoma su uključeni EGFR, TP53, kao i Ras signalni putevi. Molekularna
pozadina timoma jos uvek nije dovoljno istražena, međutim sa sve većim napretkom novih tehnologija,
poput sekvenciranja nove generacije (eng. NGS) došlo se do novih informacija koje ukazuju na
uzrok, detektuju nove molekularne markere, a samim tim omogućuju uspešnije terapijske pristupe ovoj
patologiji.Thymoma, thymic carcinoma (TC), and thymic neuroendocrine carcinoma are rare thymic epithelial
tumors (TETs) and the most frequent thymus-specific neoplasia. Thymomas are indolent or the
less aggressive forms of TETs classified into the following subgroups A, AB, B1, B2, and B3. The most
recent studies suggest thymoma specific gene GTF2I, with Chr7 c.1211T> A, p. Leu404His and
c.1271T>A, p. Leu424His, is considered driver variants for this disease. Additionally, in thymoma pathogenesis
are included EGFR, TP53 and Ras signaling pathways. The molecular background of thymoma
is still obscure, but the advance of new technologies such as next-generation sequencing (eng.
NGS) brings new information related to the molecular milieu of thymoma and other cancers. New information
help to explore the cause of the disease, detection of new molecular markers, and better
therapy approaches for thymoma
Comparative study of physiological, biochemical and morphological parameters in two tomato genotypes, wild type cv. Ailsa Craig and its ABA-deficient mutant flacca
Get PDFThe objective of this study was to determine the constitutive differences in physiological, biochemical and morphological parameters between two tomato genotypes with different levels of abscisic acid (ABA) – wild type Ailsa Craig (WT) and ABA deficient mutant flacca. Due to reduced ABA biosynthesis, flacca mutant is a suitable model system for investigating the influence of endogenous leaf ABA level in response to drought in plants. Within this research, plants were grown under controlled conditions at 800 μmol m-2s-1, until the end of the vegetative phase when samples were taken. The accumulation of the most abundant soluble sugars, sorbitol, phenolic compounds, and ascorbate in the leaves, as well as the cell walls compounds, were analyzed. Lower constitutive ABA content in flacca was accompanied by two times higher stomatal conductance and similar leaf water potential. Higher content of phenolic compounds (HBAs, HCAs, flavonoids) was determined in WT plants, which was in contrast with the elevated accumulation of the epidermal flavonoids in flacca. Larger accumulation of sorbitol in WT, and of the most abundant soluble sugars (glucose, fructose and sucrose) indicate that flacca accumulates lower content of osmolytes which was opposite to the condition at low light. However, an alternative mechanism related to cell wall modulation imposed its importance in the development of plant acclimation mechanisms under stressful environmental conditions in tomato deficient in ABA. On the other hand, an elevated ascorbate redox state in flacca indicates a higher sensitivity to oxidative stress of the mutant compared to WT even in optimal environmental conditions.Book of Abstracts: 4th International Conference on Plant Biology and 23rd SPPS Meeting, 6-8 October 2022, Belgrad