Establishment of a patient-derived induced pluripotent stem cells with 22q11.2 microdeletion: a model system for studying neurodevelopmental disorders

Background: Neurodevelopmental disorders (NDDs), including autism spectrum disorders, intellectual disability and schizophrenia, represent a public health challenge in modern societies. However, molecular mechanisms underlying NDDs are still unknown. 22q11.2 Deletion Syndrome (22q11.2DS), caused by microdeletion 22q11.2, is one of the syndromes with a high risk for NDDs; it is one of the strongest known risk factors for development of psychiatric illness and one of the highest known genetic risks for schizophrenia. Material and Methods: Peripheral blood mononuclear cells from patients with 22q11.2DS and control subjects were reprogrammed using CytoTune™-iPS 2.0 Sendai Reprogramming Kit. Characterization of generated iPSC lines were done by analyzing their morphology, genomic integrity, pluripotency and the ability to differentiate into three germ layers. Results: Peripheral blood mononuclear cells from seven patients with 22q11.2 microdeletion and three healthy controls were reprogrammed. Genotyping revealed that some of the iPSC lines contain additional CNVs. All generated iPS cell lines express markers of pluripotency and have ability to differentiate into three germ layers. Conclusion: iPSC lines derived from patients with 22q11.2DS and healthy controls were successfully established. They represent a powerful model system for studying molecular mechanisms underlying NDDs.Book of absctracts: European Society of Human Genetics (ESHG) 2025 Conference, May 24–27, 2025. Milan, Ital

Uporedna analiza bioloških aktivnosti divljih i kultivisanih biljaka vrste Helichrysum italicum

Vrsta Helichrysum italicum (Roth) G. Don (Compositae), smilje, visoko je cijenjena biljka u tradicionalnoj medicini Balkana, gdje se koristi u obliku čajeva, tinktura i melema za liječenje kožnih oboljenja, rana, respiratornih i probavnih tegoba, te upala. Cilj ovog istraživanja bio je uporedno ispitivanje antioksidativnih, antidijabetičkih i antiinflamatornih svojstava lista i nadzemnog dijela u fazi cvijetanja divljih i kultivisanih biljaka vrste H. italicum, kao i određivanje koncentracije ukupnih fenolnih jedinjenja. Antioksidativna aktivnost etanolnih ekstrakata određena je spektrofotometrijskim metodama zasnovanim na sposobnosti doniranja elektrona i redukciji metala, antidijabetička kao sposobnost inhibicije aktivnosti α-amilaze i α- glukozidaze, a antiinflamatorna kao sposobnost inhibicije denaturacije proteina. Etanolni ekstrakti divljih biljaka vrste H. italicum sadržavali su znatno veću koncentraciju fenolnih jedinjenja u listu (746 ± 9 mg/gDW) i nadzemnom dijelu (513 ± 8 mg/gDW) u odnosu na kultivisane biljke. Uzorci divljih biljaka pokazali su veći ukupni antioksidativni kapacitet, kao i izraženiju antidijabetičku i antiinflamatornu aktivnost, dok je sposobnost redukcije prelaznih metala bila slična kod obje grupe. Intenzivnija sinteza fenolnih jedinjenja kod H. italicum iz prirodnih staništa doprinosi jačoj biološkoj aktivnosti te ukazuje na moguću primjenu u fitoterapiji.Zbornik sažetaka: V simpozijum biologa i ekologa Republike Srpske sa međunarodnim učešćem - SBERS 2025 Prirodnomatematički fakultet, Univerzitet u Banjoj Luci, 13-15. novembar 202

MITOCHONDRIAL MYOPATHY CAUSED BY MT-ND5 VARIANT: INTEGRATING WES AND MITOCHONDRIAL DNA ANALYSIS

Mitochondrial disorders caused by pathogenic variants in mitochondrial genome represent a heterogeneous group of disorders, including mitochondrial myopathy, Leigh syndrome, MELAS and others. We report a patient with mitochondrial myopathy caused by a pathogenic variant in the MT-ND5 gene. The patient is a male child born after an uneventful full-term pregnancy from healthy nonconsanguineous parents. He presented with failure to thrive with psychomotor delay and recurrent infections. Neurologic examination at 11 months revealed axial hypotonia with limb hypertonia, hyperreflexia, head tremor, weak supporting and postural responses, hypomimic face, bilateral eyelid semiptosis, convergent strabismus and inability to visually track objects. Elevated serum lactate prompted further metabolic analyses which indicated a defect in oxidative phosphorylation. Whole exome sequencing (WES) analysis aimed at nuclear-encoded mitochondrial genes was negative, but the analysis of mitochondrial DNA revealed a pathogenic variant c.758T>C (p.Val253Ala) in the MT-ND5 gene, with a heteroplasmy level of 54%. Variant was confirmed by Sanger sequencing, while the analysis of maternal sample showed that the variant arose de novo. This study highlights the importance of integrating mitochondrial DNA analysis with WES to enable precise diagnosis and treatment of different mitochondrial disorders.Book of abstract: 15th Balkan congress of human genetics and 3rd Alpe Adria meeting of human genetics, 9 - 11 October 2025, Rikli Balance Hotel ,Bled, Sloveni

Analiza kvaliteta sekvenciranja celog genoma korišćenjem humane DNK izolovane iz krvi i pljuvačke

Секвенцирање ДНК је процес којим се одређује редослед нуклеотида у ДНК молекулу. Прва метода за секвенцирање ДНК, метода секвенцирања ДНК по Сангеру, објављена је 1977. године и од тада је у широкој примени (1). Ова метода секвенцирања представља прву генерацију секвенцирања и заснива се на ензимској синтези ДНК, коришћењу обележених (радиоактивно или флуоресцентно) дидеокси нуклеотида који представљају терминаторе синтезе и електрофоретском раздвајање фрагмента ДНК. У периоду између 2004. и 2006. године дошло је до развоја метода нове генерације секвенцирања које се заснивају на паралелној анализи великог броја сегмената ДНК (масовно паралелно секвенцирање) и потом повезивању добијених података биоинформатичком анализом података (2). Методе друге генерације секвенцирања заснивају се на масовном паралеленом секвенцирању милиона кратких секвенци величине од 250 до 800 базних парова (2). Процес обухвата припрему библиотеке, секвенцирање и обраду добијених података. Недостатак методе друге генерације секвенцирања је немогућност детектовања више од 70% хуманих структурних геномских варијанти (2). Методе треће генерације секвенцирања заснивају на масовном паралеленом секвенцирању дугих секвенци (енг. long-read sequencing) дужине од једне килобазе до неколико мегабаза (2). У зависности да ли је материјал који ће бити секвенциран ДНК или РНК, секвенцирање нове генерације може се најшире поделити на секвенцирање ДНК (енг. DNA-seq) и секвенцирање РНК (енг. RNA-seq). Секвенцирање ДНК се надаље дели на секвенцирање целог генома (енг. Whole Genome Sequencing - WGS), секвенцирање целог егзома (енг. Whole Exome Sequencing - WES), секвенцирање епигенома (енг. Epigenome Sequencing) и циљано (таргетно) секвенцирање (енг. Targeted Sequencing) (2).Sekvenciranje DNK je proces kojim se određuje redosled nukleotida u DNK molekulu. Prva metoda za sekvenciranje DNK, metoda sekvenciranja DNK po Sangeru, objavljena je 1977. godine i od tada je u širokoj primeni (1). Ova metoda sekvenciranja predstavlja prvu generaciju sekvenciranja i zasniva se na enzimskoj sintezi DNK, korišćenju obeleženih (radioaktivno ili fluorescentno) dideoksi nukleotida koji predstavljaju terminatore sinteze i elektroforetskom razdvajanje fragmenta DNK. U periodu između 2004. i 2006. godine došlo je do razvoja metoda nove generacije sekvenciranja koje se zasnivaju na paralelnoj analizi velikog broja segmenata DNK (masovno paralelno sekvenciranje) i potom povezivanju dobijenih podataka bioinformatičkom analizom podataka (2). Metode druge generacije sekvenciranja zasnivaju se na masovnom paralelenom sekvenciranju miliona kratkih sekvenci veličine od 250 do 800 baznih parova (2). Proces obuhvata pripremu biblioteke, sekvenciranje i obradu dobijenih podataka. Nedostatak metode druge generacije sekvenciranja je nemogućnost detektovanja više od 70% humanih strukturnih genomskih varijanti (2). Metode treće generacije sekvenciranja zasnivaju na masovnom paralelenom sekvenciranju dugih sekvenci (eng. long-read sequencing) dužine od jedne kilobaze do nekoliko megabaza (2). U zavisnosti da li je materijal koji će biti sekvenciran DNK ili RNK, sekvenciranje nove generacije može se najšire podeliti na sekvenciranje DNK (eng. DNA-seq) i sekvenciranje RNK (eng. RNA-seq). Sekvenciranje DNK se nadalje deli na sekvenciranje celog genoma (eng. Whole Genome Sequencing - WGS), sekvenciranje celog egzoma (eng. Whole Exome Sequencing - WES), sekvenciranje epigenoma (eng. Epigenome Sequencing) i ciljano (targetno) sekvenciranje (eng. Targeted Sequencing) (2)

Exploring the expression and prognostic utility of NISCH in primary and metastatic rectal cancer

Background: 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

FTIR analysis of biopolymer degradation using Streptomyces microflavus DG19 in liquid cultures and compost

The data and files contained in this dataset are related to the scientific publication "Degradation of biomaterials by Streptomyces microflavus DG19: depolymerization activity, genome mining, and soil burial assessment". This study evaluates the capacity of the bacterium Streptomyces microflavus DG19 to degrade a selection of biopolymers, highlighting its potential as a whole- cell biocatalyst for enhancing composting and developing sustainable remediation strategies for bioplastic waste. This dataset provides the raw Fourier-transform infrared spectroscopy (FTIR) data used to characterize the structural changes on the surface of various biomaterials after being subjected to degradation by S. microflavus DG19. The analyzed materials include poly(3- hydroxybutyrate) (PHB), poly(ε-caprolactone) (PCL), and post-consumer items made of poly(3- hydroxybutyrate-co-3-hydroxyhexanoate) (PHBH) and PHB. The data consists of spectra collected from control samples and samples treated with the bacterium in two different experimental conditions: liquid culture and simulated soil burial (compost). The FTIR analysis confirmed the degradation process by revealing changes in characteristic vibrational bands, particularly the carbonyl (C=O) and ester (C-O-C) stretching regions, which correspond to the hydrolysis of the polymer backbone. Overall, these spectral data provide key molecular-level evidence supporting the paper's findings on the depolymerization activity of S. microflavus DG19.readme.txt (6.928Kb)***Dataset contents*** DG19_compost_FTIR.png (553.7Kb) DG19_liquid_cultures_FTIR.png (543.4Kb) PCL_compost_control.csv (137.0Kb) PCL_compost_DG19.csv (136.9Kb) PCL_film_DG19.csv (136.9Kb) PCL_film_control.csv (136.9Kb) PCL_film_DG19.csv (136.9Kb) PHB_compost_control.csv (136.9Kb) PHB_compost_DG19.csv (136.9Kb) PHB_cup_compost_control.csv (136.9Kb) PHB_cup_compost_DG19.csv (136.9Kb) PHB_film_control.csv (136.9Kb) PHB_film_DG19.csv (136.9Kb) PHBH_card_compost_control.csv (136.9Kb) PHBH_card_compost_DG19.csv (136.9Kb)File readme.txt (6.928KbKb) is under licence public domain CC0Dataset for: Agapov, A., Pantelić, B., Ponjavić, M., Herrera, D. A. G., Nicevic, M.,& Nikodinović-Runić, J.. (2025). Degradation of biomaterials by Streptomyces microflavus DG19: depolymerization activity, genome mining, and soil burial assessment. in Biotechnology for the Environment Springer Nature., 2(1), 8. [https://doi.org/10.1186/s44314-025-00024-7

DEVELOPMENT OF IPSC-BASED MODEL SYSTEM FROM PATIENTS WITH 22Q11.2 DUPLICATION SYNDROME FOR STUDYING NEURODEVELOPMENTAL DISORDERS

22q11.2 Duplication Syndrome (22q11.2DupS) is associated with an elevated risk of developing neurodevelopmental disorders (NDDs), such as autism spectrum disorders (ASD) and attention-deficit/hyperactivity disorder. ASD is detected in 14-25% of patients, making 22q11.2DupS one of the genetic syndromes with the highest prevalence of ASD. To better understand the molecular mechanisms underlying the pathophysiology of NDDs, we have generated induced pluripotent stem cells (iPSCs) from patients with 22q11.2DupS. Mononuclear cells from 22q11.2DupS patients and healthy individuals were reprogrammed into iPSC lines using the CytoTuneTM-iPS 2.0 Sendai Reprogramming Kit. The established iPSC lines were characterized by examining the expression of stem cell markers through RT-PCR and their ability to differentiate into cells of three germ layers using the STEMdiffTM Trilineage Differentiation Kit. We differentiated the iPSCs into neurons using the dual SMAD inhibition method together with retinoic acid. The expression of neural and neuronal-specific markers in neural progenitors and mature neurons was analyzed using RT-PCR and immunocytochemistry. The established iPSC lines from three patients with an inherited form of 22q11.2DupS, their mothers who carry the microduplication, and three healthy individuals demonstrated the expression of stem cell markers and the ability to differentiate into the cells of all three germ layers. Both iPSCs from carriers of the 22q11.2 microduplication and healthy controls successfully differentiated into neurons, which was confirmed by the expression of neuronal markers. The generated 22q11.2DupS iPSC lines provide a valuable resource for understanding the molecular mechanisms underlying NDDs associated with 22q11.2 microduplication.Abstract book: FENS Regional Meeting 2025, Oslo, Norway, 16-19 June 202

GENE EXPRESSION PROFILING OF ASTROCYTES DERIVED FROM IPSCS OF PATIENTS WITH 22Q11.2 DELETION SYNDROME

Deletion 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

MATRIX METALLOPROTEINASE 9 GENOTYPE MODULATES ASTHMA CONTROL IN PEDIATRIC ASTHMA PATIENTS

Matrix metalloproteinases, particularly MMP9, play a pivotal role in asthma pathology by influencing extracellular matrix remodeling and inflammation. This study examined 100 Serbian pediatric asthma patients to explore the correlation between MMP9 3’ UTR polymorphisms and MMP9 protein levels, and their impact on therapy response and asthma control. The analysis revealed two key polymorphisms (rs13925 and rs20544) in the MMP9 gene's 3’UTR, with higher frequencies of the rs20544 T allele and TT genotype in patients with well controlled asthma. Positive correlations were found between MMP9 serum levels and blood leukocyte count, and CRP levels. Patients with not well controlled disease exhibited significantly higher MMP9 levels than those with well controlled asthma (p=0.027), indicating MMP9's potential role in asthma therapy response

Structural characteristics and terminal galactose expression of monoclonal IgA paraproteins from sera of multiple myeloma patients

IgA myeloma accounts for approximately 20% of all myeloma cases and is generally associated with a less favorable prognosis than other subtypes. Some complications specific to IgA myeloma might be linked to the structural features of monoclonal IgA. These include a high degree of polymerization, which can cause hyperviscosity syndrome, and reduced expression of galactose (Gal) residues on the glycans of the IgA hinge region, leading to IgA deposition and nephropathy. To assess the level of polymerization and galactose expression, we isolated IgA from the sera of 15 myeloma patients using affinity chromatography on Protein M agarose. Non-reducing SDSPAGE revealed two or more fractions, with molecular weights of 160 kDa and higher. Western blot analysis confirmed the presence of IgA monomers and dimers in all samples. Galactose expression was evaluated by lectin blotting using Ricinus communis agglutinin I (RCA I), a Dgalactose-binding lectin. RCA I blotting demonstrated that both monomeric and polymeric IgA forms expressed galactose, although the intensity notably varied among samples. Our results suggest that the structural properties of myeloma IgA are heterogeneous. To determine whether these differences influence disease progression, we plan to correlate IgA structural features with patients’ clinical data. Further, in-depth glycoprotein analyses will be carried out using mass spectrometry in collaboration with the Horizon CanSERVBeCELS 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