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Advancing the IMGGE RD Biobank through BRIDGING-RD Project: Achieving full interoperability of genetic and phenotypic data to enhance participation in transnational research and innovation for human health
No full textIntroduction: The Institute of Molecular Genetics and Genetic Engineering (IMGGE) RD Biobank,
established one decade ago as part of the Horizon Europe Project SERBORDISinn, collects samples from
patient with rare diseases (RD). Currently, over 7.000 samples from patients with confirmed or suspected
RD have been gathered. Given the growing number of samples, there is a need to upgrade the IMGGE RD
Biobank. The goal is to implement best practices that will enhance data handling capabilities and ensure
the biobank meets evolving standards for efficiency, data quality (both genetic and phenotypic), ethics
and interoperability.
Material & methods: A collaboration between IMGGE and Karolinska Institutet was established through
Horizon Europe Project BRIDGING-RD (2024-2027). This initiative involves personnel training, staff and
expert exchanges, and the creation of a report with recommendations for improving the biobank's current
operations.
Results: Recommendations will focus on optimizing the handling, storage, and management of biobank
samples, along with developing improved SOPs for genetic and phenotypic data quality and
interoperability. Ethical, legal and other formal aspects of biobanking will be assessed as well.
Discussion and conclusion: Implementing these recommendations will significantly enhance the IMGGE
RD Biobank’s research potential, enabling more comprehensive studies on RD. This effort will also support
the continued expansion of the biobank's sample collection, ultimately transforming it into a formally
structured resource for collecting, storing and using biological samples for long-term research.
Furthermore, our project will likely set the example that may influence practices of biobanking and ethical
legislation in this area in Serbia.Parallel session 5A: Rare Disease Biobank Insight
IMGGE Annual Research Program 2025
No full textDuring 2025, IMGGE researchers will continue exploring following areas:
- human molecular genetics and genomics,
- microbiology and ecology of microorganisms,
- plant molecular biology.
Research in human molecular genetics and genomics will focus on the identification of genetic
variants (causal, modifier, and pharmacogenetic) through exome and genome sequencing, as
well as the application of OMICS analyses. Bioinformatics tools for big data analysis will be
developed and improved using machine learning and artificial intelligence. Special attention will
be given to the correlation between genotype and phenotype to predict disease progression
and enable personalized therapy.
A key part of the research will involve disease modeling to study cellular processes and
molecular mechanisms that influence therapy response. The development of 2D and 3D model
systems (cell cultures, induced pluripotent stem cells, organoids, xenografts), as well as animal
models (zebrafish, nematodes, rodents), will continue. Additionally, new therapeutics, including
molecular and cellular approaches, will be developed.
Research in microbiology and molecular plant biology will focus on expanding the
microorganism collection (bacteria and bacteriophages) from various habitats. These organisms
will be studied in the context of microbial interactions, antimicrobial resistance, virulence, and
the development of new therapeutic strategies for infections, autoimmune,
neurodegenerative, and neurodevelopmental diseases. Research will also continue on pollutant
degradation and the development of biotechnological processes for the production of bioactive
molecules. In plant molecular biology, researchers will investigate factors influencing tolerance
to abiotic and biotic stress, as well as homologous recombination. The most advanced
molecular biology methods, including OMICS analyses, will be applied, and data will be
integrated using bioinformatics tools and artificial intelligence.Principal Investigator: Dr. Ivana Strahinić, IMGGEDuration period: 202
Inhibition of Pseudomonas aeruginosa quorum sensing-regulated behaviors by mushroom extracts
No full textEthnopharmacological relevanceMicrobial biofilm formation, a quorum sensing (QS) phenomenon based on cell-to-cell communication through ligand-receptor interactions, is one of the major causes of nosocomial and chronic infections, foodborne diseases and associated deaths. Its inhibition is therefore considered a promising approach to fight infections. In this context, Cantharellus cibarius, Trametes versicolor, Lentinula edodes and Pleurotus ostreatus, ethno-medicinal and culinary mushrooms, have been investigated as an antibiofilm and anti-QS agents against Pseudomonas aeruginosa.Aim of the studyThe aim of this study was to identify the most effective solvent system for the extraction of bioactive compounds from medicinal mushrooms with antivirulence activity against P. aeruginosa and to elucidate the potential molecular mechanism. In addition, this work aims to address the urgent need for novel, non-antibiotic strategies to combat multidrug-resistant (MDR) pathogens.Materials and methodsFluorescence microscopy (FM), scanning electron microscopy (SEM) and quantitative real-time PCR were employed to explore the antibiofilm and anti-QS potential of mushroom extracts. Chemical characterization of the extracts was performed using Uv–Vis, FTIR and UHPLC Q-ToF MS, while their toxicity was evaluated in an in vivo model using Caenorhabditis elegans.ResultsHot alkali polysaccharide extracts (APE) were the most effective, as they significantly decreased the mRNA levels of all tested QS, especially genes encoding autoinducer synthases (lasI, rhlI and pqsA), by 2- to 17-fold, as well as virulence factor genes of P. aeruginosa. Furthermore, micrographs (FM and SEM) confirmed that all APEs significantly inhibited adhesion and biofilm formation. FTIR and UV–Vis analyses showed that the extracts consisted mainly of polysaccharides, while UHPLC Q-ToF MS analysis revealed the presence of azelaic acid and vanillylacetone in all extracts. The in silico studies showed that vanillylacetone strongly interferes with various QS receptors (LasR, PqsR and RhlR) and could serve as a competitive inhibitor for the autoinducer molecules and possibly induce conformational changes in proteins. APE has also been found to increase the antibacterial efficacy of antibiotics. In vivo toxicity assays with C. elegans demonstrated their safety (with a survival rate of ≥90 %).ConclusionsThe tested mushrooms are promising sources of safe, natural antibiofilm and antivirulence agents, justifying their traditional use in infection control and offering potential for novel anti-infective therapies
Nicotine alters neutrophil chemotaxis without affecting cell numbers in zebrafish embryos
No full textNicotine exposure in utero has been linked to disruption of the normal development and function
of the fetal immune system, causing altered inflammatory responses, immune cell deficiencies,
and immune imbalances that can persist into postnatal life, increasing susceptibility to respiratory
diseases and potentially other immune-related disorders. Zebrafish provide a valuable model to
study in detail the effects of nicotine exposure during development. This study aimed to investigate
the impact of nicotine on neutrophils in developing zebrafish embryos.
Zebrafish embryos Tg(mpx:EGFP) were exposed to 1 and 10 μg/mL nicotine from 6 hours post -
fertilization (hpf) until 72 hpf, while an untreated group served as controls. In order to obtain the
number of neutrophils, flow cytometry was used - embryos were homogenized, and analyzed on a
CyFlow Space cytometer, with a total of 100,000 events per sample. Neutrophil chemotaxis was
assessed in vivo by confocal microscopy (Leica SP8 Confocal Laser Scanning Microscope) by
tracking neutrophil movement and distribution within the embryo.
Flow cytometry revealed no significant difference in total neutrophil numbers between nicotinetreated
and control groups. In contrast, confocal microscopy demonstrated altered neutrophil
chemotaxis in nicotine-exposed embryos, suggesting functional modulation of neutrophil
migration independent of overall cell number.
Our findings suggest that nicotine modulates neutrophil migratory behavior in zebrafish embryos
in a manner consistent with previous observations in humans. This supports the suitability of
zebrafish as a model organism for studying nicotine-induced immune alterations, as well as for
investigating the effects of other inhaled or systemic agents relevant to human pulmonary diseases.BeCELS 2025: Belgrade Conference for Early-Career Life Scientists, taking place on Friday, September 5, 2025, at the Institute of Molecular Genetics and Genetic Engineering (IMGGE) in Belgrad
Exploring the expression and prognostic utility of NISCH in primary and metastatic rectal cancer
No full textBackground: Rectal adenocarcinoma (READ) ranks 8th in terms of incidence and 10th in mortality rates, representing
a significant public health concern worldwide. As a result, new diagnostic and prognostic biomarkers are urgently
needed. NISCH has been identified as both a tumor suppressor and a positive prognostic marker in breast and ovarian
cancers. Recent analyses of NISCH across various tumor types have revealed its context-dependent role. However,
NISCH has not been extensively studied in READ. The aim of this study was to analyze NISCH expression in both primary
and metastatic READ and evaluate its prognostic potential.
Material (Patients) and Methods: NISCH levels in normal and READ samples were obtained through Xena Functional
Genomics Explorer, TIMER, and GEPIA2 online tools. Additionally NISCH was analysed by RT-qPCR in our two cohorts
with paired samples: cohort 1 consisted of 28 patients with primary READ with adjacent normal rectal tissue, and the
cohort 2 with 17 patients with rectal cancer with liver metastasis (RCLM) and adjacent normal liver tissue. GAPDH
levels were used for normalization. Data were analysed using the 2-dCt method. Kaplan–Meier (KM) analysis was used
to evaluate overall survival (OS) and disease-free survival (DFS) in cohort 1 and time to recurrence (TTR) in cohort 2
based on NISCH expression.
Results: In silico analyses using Xena and TIMER online tools revealed no significant difference in NISCH expression
between READ and normal rectal tissue. In contrast, GEPIA2 analysis indicated that NISCH was reduced in READ. Due
to these inconsistent findings across databases, we further evaluated NISCH expression on our cohorts. No significant
difference in NISCH expression was observed between READ and adjacent normal rectal tissue in cohort 1. Also, there
was no difference in NISCH expression between primary and metastatic READ from cohort 2. However, NISCH was
significantly downregulated in RCLM compared to healthy liver tissue. KM analysis showed no association between
NISCH expression and OS or DFS (cohort 1) or TTR (cohort 2).
Conclusions: While NISCH expression did not differ between primary READ, adjacent normal tissue, or metastatic
lesions, it was significantly downregulated in liver metastases compared to healthy liver tissue, suggesting a potential
role in the metastatic liver microenvironment. NISCH expression showed no prognostic value for OS, DFS or recurrence
in the analyzed rectal cancer cohorts
GENE EXPRESSION PROFILING OF ASTROCYTES DERIVED FROM IPSCS OF PATIENTS WITH 22Q11.2 DELETION SYNDROME
No full textDeletion Syndrome (22q11.2DS), caused by microdeletion 22q11.2, is the most
common microdeletion syndrome in humans. t is closely linked to an increased risk of
neurodevelopmental disorders and provides a valuable model for investigating the molecular
mechanisms underlying these disorders, which are still not fully understood. Here we analyzed
transcriptomic profiling of 22q11.2DS astrocytes derived from iPSCs of mother and child with
22q11.2DS and one healthy control. RNA-seq was carried out on Illumina NovaSeq 6000
sequencer. Bioinformatic processing of raw data was conducted via NVIDIA platform, while
differential gene expression analysis was performed in RStudio using DESeq2 R package. The
obtained list of DEGs was used for pathway enrichment analysis by employing EnrichR and
WikiPathways. 125 DEGs with lower expression and 287 DEGs with higher expression in
22q11.2DS astrocytes compared to the control were obtained. For genes with lower expression
in 22q11.2DS astrocytes, 22q11.2 Copy Number Variation Syndrome and Axon Guidance were
the top enriched pathways, while for genes with higher expression in 22q11.2DS astrocytes we
did not identify biological pathways that are enriched in DEG lists more than would be expected
by chance. We found 178 DEGs with lower expression and 205 DEGs with higher expression in
astrocytes of symptomatic child with 22q11.2DS compared to non-symptomatic mother with
22q11.2DS. Employing EnrichR and WikiPathways we did not identify biological pathways that
are enriched in DEG lists more than would be expected by chance. Our findings offer
preliminary evidence of an altered transcriptomic landscape of 22q11.2DS astrocytes.Abstract book: FENS Regional Meeting 2025, Oslo, Norway, 16-19 June 202
The effect of prothrombin Belgrade (FII c.1787G>A) mutation on hemostasis and association with thrombophilia
No full textProtrombin je ključni regulator hemostaze, a mutacije u njegovom genu mogu izazvati poremećaje
zgrušavanja krvi, uključujući trombofiliju. Protrombin Beograd (FII c.1787G>A) pripada grupi
mutacija koje izazivaju rezistenciju na antitrombin–trombotički mehanizam koji još nije do kraja
razjašnjen. Cilj disertacije bio je ispitivanje efekata ove mutacije i razumevanje šireg mehanizma
njenog delovanja.
Za potrebe funkcionalne analize, eksprimirani su rekombinantni wild type i mutirani protrombin.
Ispitivan je uticaj mutacije na koagulaciju i fibrinolizu određivanjem hemostatskog potencijala i
turbiditeta ugruška, kao i analizom fibrinske mreže konfokalnom i elektronskom mikroskopijom.
Proučavani su efekti na aktivaciju i agregaciju trombocita (ELISA i LTA metodama), i uticaj na
proliferaciju i migraciju endotelnih ćelija (MTT i test isceljenja rane). Urađena je integrativna
bioinformatička analiza podataka dobijenih sekvenciranjem celokupnog genoma nosilaca, radi
boljeg razumevanja mehanizma delovanja mutacije i procene njene evolucione starosti. Jedan deo
teze bio je posvećen razvoju eseja za brzu detekciju mutacije.
Rezultati pokazuju minimalni efekat na procese koagulacije i fibrinolize, uz detektovano sporije
zgrušavanje i nastanak debljih fibrinskih vlakana kada je mutacija prisutna. Mutacija izaziva
pojačanu aktivaciju trombocita, bez direktnog uticaja na agregaciju. Efekat na proliferaciju i
migraciju endotelnih ćelija nije detektovan. Otkriveni su geni koji potencijalno modulišu
aktivaciju trombocita i trombotički potencijal mutacije, što je u skladu sa dobijenim rezultatima.
Procenjena je starost mutacije na 2000–7000 godina. Uspostavljeni su eksperimentalni protokoli
za razlikovanje uzoraka nosilaca od nenosilaca mutacije, što predstavlja osnovu za klinički test.Prothrombin is a key regulator of hemostasis, and mutations in its gene can cause coagulation
disorders, including thrombophilia. The Prothrombin Belgrade belongs to mutations that causes
antithrombin resistance—a new thrombotic mechanism not yet fully understood. The aim of this
dissertation was to investigate the effects of this mutation and to better understand its broader
mechanism of action.
For functional analysis, recombinant wild-type and mutated prothrombin were first expressed. The
mutation’s impact on coagulation and fibrinolysis was examined using global hemostatic potential
and clot turbidity assays, as well as analyzing fibrin network structure using confocal and electron
microscopy. Effects on platelet activation and aggregation (ELISA and LTA assays) and on
endothelial cell proliferation and migration (MTT and scratch assay) were also studied. An
integrative bioinformatics analysis of whole-genome sequencing data from prothrombin Belgrade
mutation carriers was performed to better understand the mutation’s mechanism and estimate its
age. In addition, part of the thesis focused on developing a rapid assay for mutation detection.
The results showed that the mutation modestly affects coagulation and fibrinolysis, with delayed
clotting and thicker fibrin fibers. It increases platelet activation without affecting aggregation. No
effects on endothelial cell proliferation or migration were observed. Modifier genes possibly
influencing platelet activation and thrombotic risk were identified, in line with experimental
results. The mutation’s age is estimated at 2000–7000 years. Experimental protocols were
established to distinguish carriers from non-carriers, forming a basis for a clinical test
Hypoxia-Induced Disruption of Human Neurogenesis: Insights from a Study on Neural Precursor-Like Culture
No full textDifferent pathological conditions caused by oxygen deprivation in the brain affect
neurogenesis, consequently impairing several cognitive domains and leading to
age-related neurodegenerative disorders. Studying these processes requires the
use of suitable model systems that accurately reproduce these events in
humans. In the present study, experiments were undertaken to further
characterize the potential of using the human NT2/D1 teratocarcinoma cell line
for studying the effect of hypoxia on neuronal development and neuroregeneration.
By modifying the current protocol and reducing the concentration of
inducing agent, all-trans retinoic acid, from 10μM to 1μM we greatly improved the
efficiency of neuronal differentiation of these cells, especially regarding a
population of DCX-positive neurons. The response of NT2/D1-derived neural
precursor-like cells (NP-LCs) to hypoxia mimicking agent CoCl2 simulated hypoxia
by triggering the expression of the HIF-1α protein. A transcriptome study revealed
2,925 genes that were differentially expressed among treatment groups, of
which 1,065 were upregulated and 1,860 were downregulated in NP-LCs treated
with CoCl2. A functional study revealed the enrichment of various metabolic
processes, as well as increased transmembrane passage, in response to hypoxic
conditions. Furthermore, by integrating the results from gene expression and
double immunocytochemistry analysis, we demonstrated that hypoxia had
multiple effects on the neuronal differentiation of NP-LCs: it reduces the number
of differentiated neurons while simultaneously increasing their maturity status.
The present study reveals the mechanism of severe hypoxia NP-LCs at the
transcriptional level and provides a valuable source for investigating the energy
metabolism mechanism of the hypoxia response in the human brain.Book of abstract: 9 Congress of the Serbian Neuroscience th Society
with international participation, October 6-8, 2025. Belgrade, Serbi
Cosmetic Safety Evaluation of Cultivated Serbian Plant Petals via Danio rerio Toxicity Test
No full textFlower petals are a sustainable and eco-friendly source of bioactive compounds, as their harvesting does not compromise plant survival or ecosystem balance. Rich in phenolics, flavonoids, and other valuable metabolites [1], petals remain largely underexplored compared to leaves, roots, or aerial parts, making them a promising source of bioactive ingredients for innovative cosmetic applications. To ensure their safe use, four methanolic petal extracts (PEs) with strong cosmetic potential—prairie rose (Rosa setigera Michx.), common peony (Paeonia officinalis L.), common lilac (Syringa vulgaris L.), and ivy geranium (Pelargonium peltatum (L.) L'Hér. ex Aiton)—were evaluated for toxicity using zebrafish (Danio rerio) embryos across six concentrations (15.62–500 μg/mL), assessing survival and hatchability. Notably, rose caused only 20% lethality at the highest concentration but significantly inhibited hatching at ≥31.25 μg/mL, whereas geranium exhibited the highest lethality, with 100% mortality at ≥125 μg/mL. Hatchability assays revealed species-specific embryotoxicity: rose completely inhibited hatching at ≥62.5 μg/mL, peony at 125 and 62.5 μg/mL, lilac showed minimal impact up to 125 μg/mL, and geranium affected hatching only at 31.25 μg/mL. Overall, in vivo results confirmed safe concentrations of ≤31.25 μg/mL for peony, lilac, and ivy geranium, and ≤15.62 μg/mL for rose. These findings establish safe concentration ranges for the tested PEs, providing guidance for their sustainable and safe incorporation into cosmetic formulations.Book of Abstracts : 11th Conference of Young Chemists of Serbia Kragujevac, 25th October 202
Ispitivanje potencijalnog antitumorskog dejstva ekstrakata biljke Geum urbanum na ćelijske linije poreklom od glioblastoma in vitro
No full textGlioblastom (GBM) je najčešći maligni tumor mozga i predstavlja 46,1 % malignih tumora
Centralnog Nervnog Sistema (CNS) (1). Može se javiti bilo gde u CNS-u, ali najčešće nastaje
u beloj masi velikog mozga. Uglavnom se razvija de novo, bez dokaza o prethodno postojećoj
leziji. Kliničku sliku karakteriše kratka istorija simptoma, brza progresija bolesti i kratko
preživljavanje. GBM nastaje iz glijalnih ćelija ili njihovih prekursora usled poremećene
kontrole ćelijske proliferacije, koja dovodi do nekontrolisanog rasta tumorskih ćelija (2).
Svetska zdravstvena organizacija (SZO) je 2021. godine objavila peto izdanje klasifikacije
tumora CNS-a, koje predstavlja šestu verziju međunarodnog standarda za klasifikaciju
tumora mozga i kičmene moždine i uvodi značajne promene koje unapređuju ulogu
molekularne dijagnostike u klasifikaciji CNS tumora (3). Korišćenjem savremenih
dijagnostičkih tehnologija i molekularnih markera u ovom izdanju uvedeni su novi tipovi i
podtipovi tumora. Kada je reč o gliomima, glavne razlike uključuju: podelu između glioma
tipičnih za odrasle i glioma tipičnih za decu, kombinaciju histoloških i molekularnih nalaza u
klasifikaciji glijalnih neoplazmi, prepoznavanje novih neoplastičnih entiteta i reviziju
nomenklature. Prema ovom izdanju GBM spada u difuzne gliome tipične za odrasle (4).
Postoje brojni faktori koji su povezani sa povećanim rizikom od nastanka GBM-a, kao što su
brojne genetske bolesti i nasledni poremećaji, povreda glave poput potresa mozga, virusi,
poput HCMV-a (eng. Human Cytomegalovirus-humani citomegalovirus) i izlaganje visokim
dozama jonizujućeg zračenja. Samo 1% opisanih slučajeva GBM-a predstavlja familijarnu
formu tumora (5). Osnovne karakteristike GBM tumora predstavljaju genetske mutacije,
neograničen proliferativni kapacitet, nekontrolisan rast i replikativni potencijal, visoka stopa
invazije i metastaziranja, indukcija angiogeneze, heterogenost, prisustvo tumorskih matičnih
ćelija, epitelno-mezenhimalna tranzicija (EMT), kao i otpornost na radio- i hemio-terapiju
(Slika 1) (6). Selektivnost krvno-moždane barijere (eng. blood-brain barier, BBB)
ograničava prodor leka i njegovu efikasnost unutar normalnog mozga i tumora i predstavlja
jedan od mehanizama otpornosti GMB-a na postojeće terapijske protokole. Ovakve
karakteristike i mehanizmi otpornosti GBM-a ograničavaju napredak u stopi preživljavanja
pacijentata sa GBM u poslednje tri decenije (7)