imagine (Institute of molecular genetics and genetic engineering)
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Analysis of TNS3-203 and LRRFIP1-211 Transcripts as Oral Cancer Biomarkers
No full textIntroduction A recent pan-cancer transcriptome analysis indicated differential activity of alternative promoters of genes TNS3 and LRRFIP1 in head and neck squamous cell carcinoma compared to non-cancerous tissue. The promoters upregulated in head and neck squamous cell carcinoma regulate expression of transcripts TNS3-203 and LRRFIP1-211. Objective Our aim was to investigate the biomarker potential of TNS3-203 and LRRFIP1-211 transcripts in oral cancer, the most common type of head and neck cancer. Materials and Methods An in silico approach was used to characterize the promoters and transcripts of interest. Relative expression of TNS3-203 and LRRFIP1-211 transcripts was evaluated by qRT-PCR in a group of 46 oral cancer patients in samples of cancer and adjacent non-cancerous tissue. Results TNS3-203 was significantly overexpressed in oral cancer compared with matched non-cancerous tissue, so this transcript can potentially be used as a diagnostic biomarker. There were no differences in LRRFIP1-211 level between analyzed tissues. None of the investigated transcripts has prognostic potential in oral cancer. Conclusion The results obtained indicate the diagnostic potential of TNS3-203, but not LRRFIP1-211 transcript and its role in oral carcinogenesis.Related to supplementary material:[https://imagine.imgge.bg.ac.rs/handle/123456789/2768
Biodegradable Electrospun PLGA Nanofibers-Encapsulated Trichinella Spiralis Antigens Protect from Relapsing Experimental Autoimmune Encephalomyelitis and Related Gut Microbiota Dysbiosis
No full textPurpose: Trichinella spiralis has evolved complex immunomodulatory mechanisms mediated by excretory-secretory products (ESL1) that enable its survival in the host. Consequently, ESL1 antigens display excellent potential for treating autoimmune diseases such as multiple sclerosis (MS). However, whether timely controlled delivery of ESL1 antigens in vivo, as in natural infections, could enhance its therapeutic potential for MS is still unknown. Methods: To test this, we encapsulated ESL1 antigens into biodegradable poly (lactide-co-glycolic) acid (PLGA) nanofibers by emulsion electrospinning as a delivery system and assessed their release dynamics in vitro, and in an animal MS model, experimental autoimmune encephalomyelitis (EAE), induced 7 days after PLGA/ESL1 subcutaneous implantation. PLGA/ESL1 effects on EAE symptoms were monitored along with multiple immune cell subsets in target organs at the peak and recovery of EAE. Gut barrier function and microbiota composition were analyzed using qPCR, 16S rRNA sequencing, and metabolomic analyses. Results: ESL1 antigens, released from PLGA and drained via myeloid antigen-presenting cells through lymph nodes, protected the animals from developing EAE symptoms. These effects correlated with reduced activation of myeloid cells, increased IL-10 expression, and reduced accumulation of proinflammatory natural killer (NK) cells, T helper (Th)1 and Th17 cells in the spleen and central nervous system (CNS). Additionally, CD4+CD25hiFoxP3+ regulatory T cells and IL-10-producing B cells were expanded in PLGA/ESL1-treated animals, compared to control animals. The migration of ESL1 to the guts correlated with locally reduced inflammation and gut barrier damage. Additionally, PLGA/ESL1-treated animals displayed an unaltered microbiota characterized only by a more pronounced protective mevalonate pathway and expanded short-chain fatty acid-producing bacteria, which are known to suppress inflammation. Conclusion: The delivery of T. spiralis ESL1 antigens via biodegradable electrospun PLGA nanofiber implants efficiently protected the animals from developing EAE by inducing a beneficial immune response in the spleen, gut, and CNS. This platform provides excellent grounds for further development of novel MS therapies
ANKRD2 Knockdown as a Therapeutic Strategy in Osteosarcoma: Effects on Proliferation and Drug Response in U2OS and HOS Cells
No full textAnkrd2, a mechanoresponsive protein primarily studied in muscle physiology, is emerging as a player in cancer progression. This study investigates the functional role of Ankrd2 in osteosarcoma cells, revealing its critical involvement in cell proliferation and response to chemotherapeutic drugs. We showed that Ankrd2 knockdown impairs the activation of PI3K/Akt and ERK1/2 pathways, reduces levels of cell cycle regulators including cyclin D1 and cyclin B, and counteracts the expression of nuclear lamin A and lamin B, disrupting nuclear morphology and DNA integrity. Strikingly, the loss of Ankrd2 enhances the sensitivity of osteosarcoma cells to doxorubicin and cisplatin, highlighting Ankrd2 potential as a therapeutic target to improve chemotherapeutic efficacy. Defining a novel mechanistic role for Ankrd2 in promoting tumor progression, we propose that Ankrd2 reduction could be exploited as an adjuvant strategy to enhance the efficacy of chemotherapy, offering new therapeutic opportunities for OS treatment
High Expression Levels of the Long Non-Coding RNAs Lnc-IRF2-3 and Lnc-KIAA1755-4 Are Markers of Poor Prognosis in Chronic Lymphocytic Leukemia
No full textLong non-coding RNAs (lncRNAs) play complex roles at multiple levels of gene regulation, thus modulating key cellular processes involved in the pathogenesis and progression of cancer. Aberrant expression of lncRNAs has been reported in various malignancies, including chronic lymphocytic leukemia (CLL). We investigated the expression of lnc-IRF2-3 and lnc-KIAA1755-4 in peripheral blood mononuclear cells of 112 previously untreated CLL patients by quantitative reverse-transcriptase polymerase chain reaction. Both lncRNAs were found to be overexpressed in CLL samples in comparison to healthy controls, and their high levels were associated with adverse clinico-biological characteristics of patients at diagnosis. High lnc-IRF2-3 expression was associated with high leukocyte and lymphocyte counts, high β2-microglobulin, advanced Binet stage, unfavorable cytogenetics, CD38-positivity and IGHV-unmutated status. Regarding lnc-KIAA1755-4, its high expression was associated with high leukocyte count, lymphocyte count, β2-microglobulin, lactate dehydrogenase and low hemoglobin, as well as with IGHV-unmutated status. In addition, we observed shorter time to first treatment and overall survival of patients expressing high levels of both lncRNAs in comparison to low-expressing patients. In summary, our study showed that high lnc-IRF2-3 and lnc-KIAA1755-4 expression at diagnosis predicts poor survival in CLL. The mechanisms of their upregulation, as well as their specific targets in CLL cells, remain to be elucidated
Bioaccessibility of Pesticides in Fruits
No full textThe bioaccessibility of pesticide residues is a crucial parameter for the prediction of risks to human health and potential exposure to pesticides. Due to various limitations of in vivo studies, in vitro digestion is imposed as a rapid and cost-effective alternative for assessing the bioaccessibility of pesticides in fruits. In this chapter, a protocol for the assessment of the bioaccessibility of pesticides in fruit is proposed and described in detail, including the collection/preparation of fruit samples, the in vitro digestion protocol, the bioaccessibility assessment, and the health risk assessment. It is a standardized in vitro digestion protocol designed to ensure better comparability of results between studies and a better understanding of the bioaccessibility of different pesticides during digestion
22 Alpha-1 antitrypsin affects neutrophil migration during tail fin regeneration in zebrafish larvae
No full textRegeneration is process very similar to wound healing with shared mechanisms, including
neutrophil response. Studying neutrophil behavior in animal regeneration models could help in
understanding their role in wound healing in humans. Compounds that affect innate immune
response could serve as potential therapeutics for wound healing treatment. Alpha-1 antitrypsin
(AAT) is a serine protease inhibitor with well characterized immunomodulatory activity and
interaction with neutrophils. We aimed to examine AAT’s potential to modulate neutrophils
migration during tail fin regeneration in zebrafish larvae in order to further assess its therapeutic
use in wound healing.
Two- day old zebrafish larvae with fluorescently labeled neutrophils underwent amputation of the
tail fin. After amputation, larvae were incubated for 2 to 6 hours at 28.5ºC with AAT (2mg/ml) or
HBSS, as control. After appropriate time post amputation, larvae were anesthetized and imaged
on Zeiss Axiovert fluorescent microscope. Neutrophils were counted within area of 500 μm from
wound plain, using ZEN software. Statistical analysis was performed in Past4.03 software.
Four hours post amputation (hpa) lower number of neutrophils in region of interest was observed
in treated group compared to control (p<0.05). Additionally, higher number of neutrophils was
presented in proximity of the wound edge in controls than in treatments. For other time points (2
and 6 hpa) there was no statistically significant difference between treated and control groups.
AAT exhibits potential to modulate neutrophils recruitment and infiltration at wound site. These
preliminary results suggest that treatment with AAT could promote tail fin regeneration and
warrant further investigation.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
A novel type of autoaggregation in lactic acid bacteria promoted by new AggS aggregation factor from Streptococcus thermophilus CC40-4S
No full textAutoaggregation in lactic acid bacteria is considered a beneficial probiotic trait and can be used in the food and medical industries to enhance the properties of utilized microorganisms. Currently, a group of aggregation-promoting factors (APFs) in lactic acid bacteria, known as Snowflake Forming Collagen Binding Aggregation Factors (SFCBAFs), is well described. These are large proteins with a molecular mass of over 170 kDa, containing collagen-binding domains and a repeat region, forming a unique autoaggregation phenotype. Here we describe a new type of autoaggregation in lactic acid bacteria found in Streptococcus thermophilus CC40-4S. The whole genome of the autoaggregation-positive strain S. thermophilus CC40-4S was sequenced, and bioinformatic analysis predicted a putative gene aggS involved in autoaggregation, located on the chromosome and flanked by insertion sequences. The aggS gene disruption by homologous recombination using the temperature-sensitive vector pSC led to the loss of the aggregation phenotype. Cloning and heterologous expression in Lactococcus lactis subsp. cremoris MG1363 confirmed the role of the AggS protein in autoaggregation, given that a strong aggregation phenotype was obtained. Like SFCBAF-type APFs, AggS is a large protein (237 kDa) with a repeat region, but it does not contain collagen-binding domains and forms an autoaggregation phenotype with small aggregates. To our knowledge, this is the first report on a gene coding for an aggregation-promoting factor in S. thermophilus
Shaping Up’ Bacterial Biopolymers for Biomedical and Food Applications
No full textBacterial biopolymers are increasingly valued as sustainable, versatile materials with
transformative potential in both biomedical and food industries. Advanced but also
simple engineering and processing strategies are being used to “shape up” these
natural polymers—tailoring their physical, chemical, and functional properties for
specific applications. From biofilms and hydrogels to nanofibers and edible coatings,
bacterial-derived materials such as polyhydroxyalkanoates (PHAs), bacterial
cellulose, and exopolysaccharides are being reimagined as scaffolds for tissue
regeneration, vehicles for drug delivery, and barriers for food preservation. Integrating
materials science, bioprocessing, and innovative fabrication techniques allow
molecular customization, broadening the performance and application range of these
biopolymers. Case studies will highlight recent developments in achieving tunable
mechanical properties, enhanced biocompatibility, and functional integration
(antimicrobial, antioxidant, and functional food applications). By shaping up bacterial
biopolymers—both literally and conceptually—we can unlock new pathways toward
safer, smarter, and more sustainable materials for human health and nutrition.12th ESBP will be held from October 1st to 3rd, 2025, in Lisbon, Portuga
Germline BRCA1 and BRCA2 Testing in the Context of Personalized Therapy: National Experience from Serbia in Breast Cancer Management
No full textBackground: Breast cancer is the most common cancer in women, and germline BRCA1/2 mutations significantly
increase lifetime risk, guiding both risk assessment and treatment. PARP inhibitors, like olaparib, exploit defective
DNA repair in BRCA-mutated cells, improving progression-free survival in HER2-negative patients. This study aimed to
implement nationwide BRCA testing in Serbia, enabling personalized therapy for eligible patients. It also marked the
first collaborative effort of the three major Serbian institutions to introduce a unified NGS protocol for BRCA testing.
Patients and Methods: From April 2024 to June 2025, a total of 1,739 patients were tested across three institutions:
Institute for Oncology and Radiology of Serbia, Oncology Institute of Vojvodina, and Institute of Molecular Genetics
and Genetic Engineering. BRCA1/2 mutation analysis was performed using next-generation sequencing on the Illumina
platform with the Devyser BRCA NGS kit. Genomic DNA extracted from peripheral blood was quality-controlled and
subjected to massive parallel sequencing. Data analysis was conducted using Amplicon Suite software.
Results: Most patients (72.5%) underwent testing at an early stage of disease. HR+/HER2− breast cancer was the
predominant subtype (63.7%). Pathogenic or likely pathogenic variants were detected in 97 patients (5.6%), including
56 BRCA1 (3.2%) and 41 BRCA2 (2.4%) mutations. Variants of uncertain significance were identified in 42 cases (2.4%).
Among BRCA1-positive patients, 76.8% presented with triple-negative breast cancer (age range: 32–70 years; mean
age: 50 years). In contrast, BRCA2-positive cases were predominantly HR+/HER2− (65.8%; age range: 34–78 years;
mean age: 53.5 years). A notable variation in the geographical distribution of BRCA1/2 mutations was observed, with
prevalence rates of 3.2% in southern Serbia, 5.0% in central Serbia, and 8.35% in Vojvodina.
Conclusions: The introduction of olaparib for breast cancer treatment in Serbia strengthens precision oncology
by offering an additional targeted therapy. Nationwide collaboration among major institutions has enabled the
implementation of germline BRCA testing, which is crucial for identifying patients eligible for PARP inhibitors and
for bringing personalized medicine into routine clinical care. Ongoing research should assess real-world outcomes in
patients treated with olaparib, while this germline testing will also provide valuable insights into the genetic landscape
of the Serbian population.We acknowledge AstraZeneca for donating the BRCA tests used in this nationwide
testing. The study was designed, initiated, and conducted independently by Institute for Oncology and Radiology
of Serbia, Oncology Institute of Vojvodina, and Institute of Molecular Genetics and Genetic Engineering without
sponsorship or involvement from AstraZeneca
Short- and Long-Term Studies of a 3D Osteosarcoma Cell Culture Model for Doxorubicin Drug Screening
No full textBackground: Osteosarcoma is an aggressive malignant bone tumor that mainly develops in children and adolescents
during periods of rapid bone growth and has a high tendency to metastasize. Despite intensive treatment involving
neoadjuvant multi-agent chemotherapy, surgical resection, and adjuvant multi-agent chemotherapy, the tumor recurs
in over 40% of cases. One major obstacle in improving treatment outcomes is the lack of effective preclinical models
that can better reflect the complex tumor environment found in patients. A promising approach is to develop 3D
osteosarcoma cell culture models using bone-mimicking scaffolds combined with biomimetic perfusion bioreactors to
ensure efficient mass transport and adequate biophysical signals. This study aimed to evaluate a previously developed
3D culture model for osteosarcoma cells based on macroporous composite scaffolds (2 wt.% alginate and 2 wt.%
hydroxyapatite) in conjunction with a perfusion bioreactor for anticancer drug screening.
Materials and methods: Murine osteosarcoma cells (K7M2-wt) were seeded onto the scaffolds (15x106 cells cm-3 of
scaffold volume) and cultivated under static conditions for one day. Three experimental studies were performed using
the anticancer drug doxorubicin. In the first, following 1 day of static cultivation, the drug (1 μg cm⁻³) was applied for
1 day under continuous flow in the “3D Perfuse” bioreactor (0.27 cm³ min⁻¹; 40 μm s⁻¹). In the second, scaffolds were
cultured for 7 days under the same conditions to allow spontaneous formation of spheroid-like structures, followed
by 3-day doxorubicin treatment. Additionally, a long-term model was introduced to better reflect clinical settings.
Spheroid-like structures were exposed to the drug for 3 days, followed by a 21-day recovery period under perfusion,
simulating intervals between chemotherapy cycles in patients. The cultivated scaffolds in all studies were then assessed
by performing histological analysis and MTT assay to measure cell metabolic activity.
Results: Osteosarcoma cells cultured in a 3D environment were less sensitive to the drug compared to those in 2D,
reflecting the chemotherapy resistance observed in patients.
Conclusions: This 3D osteosarcoma cell culture model is suitable for short-term drug screening and potentially for long-term studies