Cervical cancer is one of the leading causes of cancer in women, worldwide. Infection with humanpapillomavirus (HPV) has been accepted as the primary cause for the development of invasive cervicalcancer and its precursor lesions. Despite HPV infection has been proposed as an indispensable factor forcervical cancer development, only a subset of neoplastic lesions with HPV infection persist and progress toinvasive cancer. This suggests us that other molecular events are also involved in cancer progression. Aimof this study was to extract mRNA from cytobrush-collected healthy and HPV infected cervical epithelialcells and investigate various RNA extraction and purification protocols for assessment of RNA yield andquality. Taking into consideration that cervical cancer screening is based on the cytology basedPapanicolaou test (Pap test), main challenge is to investigate whether the samples obtained by regular Paptesting can be used for gene expression analysis. For this purpose, a total of 68 cervical specimens werepreviously tested for HPV infection. Following HPV testing, samples were submitted to RNA extractionand compared to the products after additional purification step involving DNase I. Products obtained afterdifferent RNA extraction and purification methods were visualized using 2% agarose gel electrophoresis.In conclusion, DNase I based RNA purification represents a necessary step for the assurance of a high-quality extracted RNA used for gene expression analysis studies. Reliance on commercial kits for RNAextraction only, without performing additional purification step can lead to errors in drawing finalconclusions and/or to false negative gene expression profiling, affecting the overall diagnostic procedure.According to obtained results, the type of sampling used in this study was not suitable for the subsequentgene expression analysis.

ABSTRACT Background Trajectories of patients receiving kidney replacement therapy (KRT), including transitions between hemodialysis (HD), peritoneal dialysis (PD) and kidney transplantation (KTx), may vary across patient subgroups and have not yet been investigated in Europe. This study aimed to: (i) describe the number of shifts across KRT modalities; and (ii) characterize the most frequent patient trajectories, including the direction of shifts across KRT modalities, the duration spent on each modality, and patient events, including death. Methods Data of adult patients (≥20 years) who initiated KRT between 2004 and 2013 were extracted from the European Renal Association (ERA) Registry database and patients were followed for 10 years from the moment of KRT initiation. Results were stratified by age, sex, primary renal disease (PRD), country and initial KRT modality. Results Among 289 323 patients, more than two-thirds (69.6%) remained on their initial KRT modality until death or the end of 10 years follow-up, but younger patients and patients who initiated with PD had more shifts across KRT modalities than others. Overall, the most frequent patient trajectories were HD followed by death (53.4%), HD to KTx (10.2%), remaining on HD for 10 years (5.6%), PD followed by death (4.4%) and PD to KTx (2.9%). The most frequent patient trajectories differed between age and PRD groups, and across European countries, but not between women and men. Conclusion The number and direction of shifts across KRT modalities varied by age, PRD, country and initial KRT modality, but not by sex. Future research should further investigate trajectories of patient subgroups that stood out, including young HD patients who died on their initial KRT modality and those who remained on HD for 10 years without undergoing KTx.

High welfare standards for animals used in research is as much an ethical issue as it is a cornerstone of high-quality science. Researchers can improve both animal welfare and data reliability by implementing strategies that reduce stress in experimental animals. One modern and effective approach is to monitor animals within their familiar home-cage environment. Home-cage monitoring (HCM) systems integrate multiple approaches to automatically, continuously, and non-invasively monitor the physiology and behaviour of laboratory animals within their home environments. HCM favours the animals’ natural rhythms and behaviours while reducing stress from various sources and the need for human intervention. In this article, we explore how HCM contributes to the 3Rs framework introduced by Russell and Burch and focus particularly on how to select the most appropriate HCM system for specific research needs. We discuss available resources and practical limitations for system choice, and provide a brief outlook on the evolving role of artificial intelligence to analyse HCM data. We also discuss the opportunities and barriers to HCM adoption, particularly in relation to countries with developing research structure and limited funding in Europe. Our central message is clear: use of HCM technologies supports 3Rs and promotes both better science and better animal welfare. Pametne kletke, večja dobrobit: Podpora načelom 3R v raziskavah na živalih s spremljanjem v domači kletki in ustrezno izbiro sistema Izvleček: Visoki standardi dobrobiti živali v raziskavah niso zgolj etična obveznost, temveč tudi temelj visokokakovostne znanosti. Raziskovalci lahko izboljšajo tako dobrobit živali kot tudi zanesljivost podatkov z uvedbo strategij, ki zmanjšujejo stres pri poskusnih živalih. Eden izmed sodobnih in učinkovitih pristopov je spremljanje živali v njihovem domačem okolju. Sistemi za spremljanje v domači kletki (HCM, angl. home-cage monitoring) združujejo več pristopov za samodejno, neprekinjeno in neinvazivno spremljanje fiziologije in vedenja laboratorijskih živali v njihovem domačem okolju. HCM podpira naravne ritme in vedenja živali ter zmanjšuje stres iz različnih virov in potrebo po posegih človeka. V članku opisujemo, kako HCM prispeva k načelom 3R, ki sta ga uvedla Russell in Burch, s posebnim poudarkom na izbiri najprimernejšega sistema HCM za specifične raziskovalne potrebe. Obravnavamo razpoložljive vire in praktične omejitve pri izviri sistema ter podajamo kratek pogled na razvijajočo se vlogo umetne inteligence pri analizi podatkov HCM. Prispevek obravnava tudi priložnosti in ovire pri uvajanju HCM, zlasti v povezavi z državami z manj razvito raziskovalno infrastrukturo in omejenimi sredstvi v Evropi. Naše osrednje sporočilo je jasno: uporaba tehnologij HCM podpira načela 3R ter spodbuja boljšo dobrobit živali in boljšo znanost. Ključne besede: laboratorijske živali; avtomatsko spremljanje vedenja; izboljšave; kontinuirno zbiranje podatkov; stres

Abstract To preserve resources for future generations and promote rural development, supporting ecotourism is essential. This paper provides guidelines for developing ecotourism, highlighting its role in environmental conservation. While mass tourism benefits rural communities, it can cause significant environmental harm. Therefore, this research promotes ecotourism as a sustainable alternative. In rural areas, ecotourism supports development by responsibly using natural resources. The study focuses on the potential of rural settlements in the Semberija region of Bosnia and Herzegovina, assessing their capacity for ecotourism to aid local development. A decision model was developed, considering four main criteria - natural, infrastructure, socio-cultural, and economic - and their sub-criteria. This model evaluates six rural communities’ ecotourism potential. To determine the importance of each criterion, a fuzzy weighting method with the Bonferroni mean operator was used, revealing economic factors as the most influential. The fuzzy ranking method then ranked the settlements, with Amajlije identified as having the highest ecotourism potential. The findings suggest that promoting ecotourism in Amajlije and similar communities can support sustainable rural development, balancing environmental preservation with economic growth.

Background Acute cholecystitis (AC) is one of the most common surgical emergencies with a wide range of clinical outcomes. Early identification of patients at risk for postoperative complications is essential for optimizing surgical decision-making and resource allocation. Hemogram-derived indices such as the systemic immune-inflammation index (SII) and neutrophil-to-lymphocyte ratio (NLR), in addition to biochemical markers, may provide prognostic value beyond traditional risk factors. Materials and methods This retrospective single-center study included 210 patients admitted to the University Clinical Center Tuzla with AC between January 2024 and January 2025. Demographic, clinical, and laboratory data were collected. Receiver operating characteristic (ROC) analysis was performed to identify optimal cut-off values for predicting complications. Multivariate logistic regression was adjusted for age, sex, diabetes mellitus, hypertension, and other baseline comorbidities, in addition to SII, NLR, glucose, and creatinine. Results Four variables emerged as independent predictors of complications: SII > 950 remained an independent predictor after full adjustment (p = 0.002) with a sensitivity of 78% and specificity of 72%. It yielded the highest discriminatory accuracy among the evaluated markers, with an area under the curve (AUC) of 0.81 (95% confidence interval (CI) 0.75-0.87). No formal comparison with TG18 grading was performed. In contrast, baseline comorbidities such as diabetes mellitus and hypertension did not retain significance after adjustment. Conclusion SII, NLR, glucose, and creatinine independently predicted complications in AC, with SII emerging as the strongest predictor among the evaluated variables. These findings suggest that incorporating hemogram-derived indices into preoperative assessment may enhance risk stratification. However, the retrospective single-center design and potential confounding related to the surgical approach warrant cautious interpretation.

Clear Cell Sarcoma (CCS) are ultra-rare fusion-translocated soft-tissue tumours occurring mainly in young adults with poor prognosis. Their aggressiveness and resistance to conventional chemotherapy, especially in a metastatic setting, characterize these tumours. A functional drug screen consisting of 80 drugs was performed on patient derived CCS cell lines. Top candidates were validated in 3D cell culture and in vivo models, allowing comparison of drug responses among CCS cell lines, including one matched pair (MUG Lucifer cell lines) representing primary and metastatic disease from the same patient. Underlying mechanisms were evaluated with RNA Seq, fluorescence/luminescence based assays, western blot and IHC. In vitro experiments in CCS spheroids highlighted the transcription inhibitor lurbinectedin to be the best overall candidate which was further confirmed in mouse xenografts, albeit to a lesser extent than observed in vitro, especially in the metastatic CCS model. Transcriptional and functional analyses revealed heterogeneous drug responses and differences in cell death mechanisms across CCS lines, reflecting patient-specific variability. Furthermore, we explored combinational treatment of lurbinectedin with selinexor. Synergistic effects were confirmed in a novel autologous co-culture model incorporating cancer-associated fibroblasts, providing a more physiologically relevant system for drug testing. This study identified promising therapeutic avenues by highlighting key vulnerabilities in CCS, considering both inter tumour heterogeneity, tumour plasticity and the stromal influence on drug response.

Heavy metals are persistent environmental pollutants with well-documented toxic, genotoxic, and bioaccumulative effects across ecosystems. This study evaluates blood cell morphology as a potential biomarker of environmental pollution in Miniopterus schreibersii (Kuhl, 1819), a migratory bat species. Individuals were captured from a contaminated site (Dardagani underground quarry) and a reference site (Mokra Megara Cave) in Bosnia and Herzegovina. Environmental sampling included guano, soil, and water. Guano was analyzed for nine heavy metals (Cr, Cu, Mn, Fe, Co, Ni, Cd, Pb, Zn) to assess site-specific contamination. Distinct spatial differences in metal accumulation were observed. Elevated Cu and Mn concentrations in guano from the contaminated site indicated anthropogenic input, whereas higher Fe, Ni, and Pb at the reference site reflected natural lithogenic enrichment. Hematological and cytological examinations revealed morphological alterations in blood cells of bats from the contaminated site, including neutrophil hypersegmentation, polychromatophilia, atypical granulation, and nuclear abnormalities in lymphocytes. The total lymphocyte count differed significantly, and these qualitative changes suggest early physiological adjustments or potential indicators of sublethal toxic exposure. By integrating environmental (soil and guano) and biological (blood morphology) parameters, this study demonstrates a non-lethal and ecologically relevant approach to biomonitoring. Blood cell alterations, combined with metal analyses in guano, provide a sensitive tool for detecting potential chronic environmental stress. M. schreibersii is reaffirmed as a valuable sentinel species for ecological monitoring in karst and other vulnerable habitats; however, the limited number of high-quality blood smears obtainable under field conditions still remains a constraint to broader generalization of the findings.

microRNA-27a is a promising candidate for miRNA mimic therapy to combat obesity, but its clinical application is hindered by enzymatic degradation and low membrane permeability. To address these challenges, we developed cationic nanostructured lipid carriers (cNLCs) via high-pressure homogenization as non-viral carriers for miRNA-27a. However, the formation of a protein corona in biologically-relevant media altered the particle size and surface charge, significantly reducing cellular uptake. To mitigate this issue, we hypothesized that coating miRNA/cNLC complexes with human serum albumin (HSA) will prevent protein corona formation and enhance cellular uptake. The HSA-coated miRNA/cNLC complexes, termed albuplexes, were characterized for particle size, zeta potential, morphology, and stability in various media. The integrity of the HSA coat was assessed using circular dichroism and UV/Vis spectroscopy. We also evaluated the biocompatibility and cellular uptake of albuplexes in 3T3-L1 cells. The biological effects of miRNA-27a on adipocyte development were analyzed through light microscopy and absorbance measurements of Oil-red-O dye in lipid droplets. Results indicated that albuplexes possess favourable physicochemical properties and enhanced stability in serum. Notably, albuplexes were rapidly taken up by 3T3-L cells via endocytosis, although 20 % HSA in the culture medium completely inhibited uptake. Furthermore, albuplexes exhibited an anti-adipogenic effect by reducing the lipid droplet accumulation, suggesting their potential as a therapeutic strategy for miRNA replacement in obesity treatment.

Three novel unsymmetrically substituted 2,2'-bipyridine ligands were prepared by introducing a 2-hydroxyphenyl group at the 6-position and either a 4-methoxyphenyl, 3,4-dimethoxyphenyl or 3,4-methylenedioxyphenyl group at the 4-position, using a modified Kröhnke protocol. Their corresponding rhenium(I) tricarbonyl iodido complexes, fac-[Re(L)(CO)3I], 1-3, were synthesized and comprehensively characterized by single-crystal X-ray diffraction (SCXRD), 1H and 13C nuclear magnetic resonance (NMR) spectroscopy, infrared (IR) spectroscopy, ultraviolet-visible (UV-Vis) spectroscopy, cyclic voltammetry (CV), high-resolution mass spectrometry (HRMS), and elemental analysis. The common 6-(2-hydroxyphenyl) moiety predominantly influences the spectroscopic and redox properties of the complexes. SCXRD confirms the facial arrangement of the fac-[Re(CO)3N2I] core in all three cases, although solid-state conformational isomerism was observed only in complex 1. In contrast, two NMR-distinguishable isomers are observed in solution for all three complexes. The cytotoxicity of 1-3 was evaluated by MTT assay after 48 h of continuous exposure in several human tumor cell lines (HeLa, PANC-1, MDA-MB-231, A549) and in non-malignant human lung fibroblasts (MRC-5). Notably, all three complexes displayed low-micromolar IC50 values comparable to cisplatin, with the highest activity observed in HeLa cells (5.11-7.45 μM). However, significant activity was also recorded in MRC-5 cells (IC50 = 8.19-8.95 μM), suggesting higher overall toxicity and weaker selectivity compared to cisplatin. Analysis in HeLa cells using bright-field microscopy confirmed a substantial antiproliferative effect.

Simple Summary African swine fever is a deadly viral disease of pigs and wild boar that causes major losses for farmers and threatens food security. The disease does not affect people, but its rapid spread and high fatality in pigs make it one of the most serious challenges for animal health in Europe. Since 2019, the disease has been present in Serbia, and in 2023, it was first reported in Bosnia and Herzegovina. In this study, we examined virus samples collected from pigs and wild boar during outbreaks between 2023 and 2025 to better understand how the virus is spreading in the region. By looking at several important parts of the virus genome, we found that all the samples belonged to the same group, known as cluster 19. This shows that the same type of virus has been circulating for several years without major changes. The results suggest that the disease is being maintained locally, mainly through contact between wild boar and pigs kept on small farms with little or no protection. The discovery of the same virus type in Bosnia and Herzegovina highlights that the disease crosses borders, making regional cooperation and continued monitoring essential for controlling its spread.

Dexketoprofen/tramadol is a fixed-dose multimodal combination analgesic that significantly controls multiple acute pain states, and may have an important clinical application in providing pain control adequate to prevent the transition from acute to chronic postsurgical and low back pain. A consensus is needed to quantify and define the actual burden of postsurgical pain (PSP) and low back pain (LBP), which can support efforts toward effective approaches to manage potential pain chronification. This study utilized a modified Delphi approach. A Scientific Committee set forth 28 statements on six themes about the burden of acute PSP and LBP, their potential transition to chronic pain, their pathophysiology, therapeutic approaches to stop this transition, and the role of multimodal analgesia in this context, specifically a fixed-dose combination oral product of dexketoprofen/tramadol. An international panel of healthcare professionals from various regions and relevant medical specialties participated in a Delphi study and were surveyed for consensus on a 5-point Likert scale with consensus defined as > 70% concordance. A round of online voting lasting 3 months and using an online survey platform was permitted for each participant. A total of 100 experts completed the Delphi survey. All the 28 proposed statements reached consensus > 70% in the first round of voting. A fixed-dose combination product, specifically dexketoprofen/tramadol was recognized as a multimodal analgesic which could effectively relieve acute pain and act to prevent its transition to chronic pain. The high global burden of chronic PSP (CPSP) and chronic LBP (CLBP) was identified as well. Healthcare professionals who deal with pain recognize the burden of acute pain, the risks of acute pain transitioning to chronic pain, and inspire to avert the transition by providing effective multimodal control of acute pain. The role of fixed-dose combination analgesics, in particular dexketoprofen/tramadol, was recognized by consensus as an efficacious and safe therapy option for these acute pain syndromes. 7KDL7wDHZ5GDvGppW1iD89 A Video Abstract is available for this article. To view, please see the online version of the manuscript or follow the ‘Digital Features’ link. A Video Abstract for The Role of Dexketoprofen/Tramadol in Multimodal Therapy to Prevent Acute Postsurgical and Acute Low Back Pain from Developing into Chronic Pain: A Delphi Consensus Study (MP4 112565 KB) A Video Abstract is available for this article. To view, please see the online version of the manuscript or follow the ‘Digital Features’ link. A Video Abstract for The Role of Dexketoprofen/Tramadol in Multimodal Therapy to Prevent Acute Postsurgical and Acute Low Back Pain from Developing into Chronic Pain: A Delphi Consensus Study (MP4 112565 KB)