Polonca Ferk

Polycystic ovary syndrome (PCOS) is one of the most biologically heterogeneous disorders in reproductive endocrinology, extending far beyond the classical features of hyperandrogenism, ovulatory dysfunction, and polycystic ovarian morphology. Although substantial progress has been made in defining the endocrine, metabolic, genetic, epigenetic, and inflammatory mechanisms associated with PCOS, these processes have largely been investigated independently, and their contribution to the marked clinical and biological heterogeneity of the disorder remains incompletely understood. Recent advances in cellular-resolution biology, including single-cell and single-nucleus transcriptomics, spatial omics, and multimodal molecular profiling, together with computational systems biology, enable increasingly detailed characterization of cellular states, spatial organization, and molecular relationships within the ovarian-follicular environment. In this Review, we use the ovarian cellular ecosystem as an operational systems framework to test whether PCOS-associated abnormalities observed in individual ovarian and follicular compartments form reproducible higher-order patterns of multicellular organization. The framework is organized across five measurable domains: cellular composition and states, intercellular signaling relationships, spatial niche organization, cross-compartment molecular coupling, and systemic-ovarian coupling. Current direct human cellular-resolution evidence remains sparse and compartment-specific. Available studies support altered molecular and cellular states within selected ovarian and follicular compartments more strongly than coordinated, spatially organized, or progressive ecosystem-wide remodeling. We critically evaluate how cellular-resolution, spatial, multi-omics, computational and artificial intelligence approaches can be used to test these ecosystem-level hypotheses, explicitly distinguishing direct measurement from computational inference and identifying the validation required for functional or causal interpretation. Finally, we discuss the translational implications of this framework for biologically informed patient stratification, ecosystem-derived biomarker discovery, mechanism-informed therapeutic development, and precision reproductive endocrinology. By integrating current evidence within a falsifiable framework, this Review defines measurable features and validation criteria for testing ovarian ecosystem organization in PCOS and identifies the critical evidence gaps that future cellular-resolution, spatial, and multi-omic studies must address. This provides a practical roadmap for determining whether reproducible ovarian ecosystem states can improve biological stratification, biomarker discovery, or therapeutic target identification beyond compartment-specific abnormalities and established systemic and clinical determinants.

Nanoparticulate TiO₂ (TiO₂ NPs) is a widely used material, whose potential toxicity towards eukaryotic cells has been addressed by multiple studies. TiO₂ NPs are considered toxic due to their production of reactive oxygen species (ROS), which can, among others, lead to cellular damage, inflammatory responses, and differences in gene expression. TiO₂ NPs exhibited toxicity in multiple organs in animals, generating potential health risks also in humans, such as developing tumors or progress of preexisting cancer processes. On the other hand, the capability of TiO₂ NPs to induce cell death has found application in photodynamic therapy of cancers. In aquatic environments, much has been done in understanding the impact of TiO₂ on bivalves, in which an effect on hemocytes, among others, is reported. Adversities are also reported from other aquatic organisms, including primary producers. These are affected also on land and though some potential benefit might exist when it comes to agricultural plants, TiO₂ can also lead to cellular damage and should be considered when it comes to transfer along the food chain towards human consumers. In general, much work still needs to be done to unravel the delicate balance between beneficial and detrimental effects of TiO₂ NPs on eukaryotic cells.

The complete mitochondrial genome of the Carniolan honeybee (Apis mellifera carnica) from Slovenia, a homeland of this subspecies, was acquired in two contigs from WGS data and annotated. The newly obtained mitochondrial genome is a circular closed loop of 16,447 bp. It comprises 37 genes (13 protein coding genes, 22 tRNA genes, and 2 rRNA genes) and an AT-rich control region. The order of the tRNA genes resembles the order characteristic of A. mellifera. The mitogenomic sequence of A. m. carnica from Slovenia contains 44 uniquely coded sites in comparison to the closely related subspecies A. m. ligustica and to A. m. carnica from Austria. Furthermore, 24 differences were recognised in comparison between A. m. carnica and A. m. ligustica subspecies. Among them, there are three SNPs that affect translation in the nd2, nd4, and cox2 genes, respectively. The phylogenetic placement of A. m. carnica from Slovenia within C lineage deviates from the expected position and changes the perspective on relationship between C and O lineages. The results of this study represent a valuable addition to the information available in the phylogenomic studies of A. mellifera—a pollinator species of worldwide importance. Such genomic information is essential for this local subspecies’ conservation and preservation as well as its breeding and selection.

Single cell RNA sequencing (scRNA-seq) represents a new large scale and high throughput technique allowing analysis of the whole transcriptome at the resolution of an individual cell. It has emerged as an imperative method in life science research, uncovering complex cellular networks and providing indices that will eventually lead to the development of more targeted and personalized therapies. The importance of scRNA-seq has been particularly highlighted through the analysis of complex biological systems, in which cellular heterogeneity is a key aspect, such as the immune system. Autoimmune inflammatory rheumatic diseases represent a group of disorders, associated with a dysregulated immune system and high patient heterogeneity in both pathophysiological and clinical aspects. This complicates the complete understanding of underlying pathological mechanisms, associated with limited therapeutic options available and their long-term inefficiency and even toxicity. There is an unmet need to investigate, in depth, the cellular and molecular mechanisms driving the pathogenesis of rheumatic diseases and drug resistance, identify novel therapeutic targets, as well as make a step forward in using stratified and informed therapeutic decisions, which could now be achieved with the use of single cell approaches. This review summarizes the current use of scRNA-seq in studying different rheumatic diseases, based on recent findings from published in vitro, in vivo, and clinical studies, as well as discusses the potential implementation of scRNA-seq in the development of precision medicine in rheumatology.

Metformin is an oral antidiabetic drug with potentially beneficial effects in patients with polycystic ovary syndrome (PCOS). Different pharmacological responses to met- formin have been reported in PCOS patients, which may be attributable to various clinical, biochemical, genetic and other factors. Here, we have outlined a short review of the results from current clinical studies on metformin efficacy in PCOS patients. Additionally, preliminary results from a retrospective pharmacogenetic study by our group on a sample of Slovene PCOS patients are presented. Pharmacogenetics is an emerging science that should pro- vide options for individualized medicine, the future of healthcare.

Molecular associations between the cannabinoid system and estrogens have recently been discovered. This research field is particularly interesting because it could lead to new pharmacotherapeutic approaches, especially in diseases where both molecular systems contribute to the pathophysiology. This paper presents an overview of the role of the cannabinoid system and estrogens in osteoporosis, atherosclerosis and cancer, and discusses the potential benefits of the modulating both systems in the pharmacotherapy of these diseases. Although an extensive literature review points to likely interactions between the cannabinoid system and estrogens, appropriate molecular studies have not yet been carried out. Our recent results in primary human osteoblasts are the first to demonstrate a possible interaction between the two systems at a molecular level. Further studies are needed to carefully examine molecular interactions as well as the efficacy and safety of various combinations of estrogens and cannabinoids in the pathogenesis and pharmacotherapy of osteoporosis, atherosclerosis, cancer and other common diseases.

Much biomedical research focuses on the effects of UV light on human cells. UV light sources are a prerequisite for such research. This paper presents the design and achieved performance of a UVA (Ultraviolet A: 320–400 nm) and a UVB (Ultraviolet B: 290–320 nm) LED-based lamp suitable for use in bioassays, as well as inside an incubator. Numerical simulations were used to optimise the number, layout and output power of LEDs to achieve good irradiance homogeneity while maintaining low costs. Design was optimised for the efficient transfer of generated heat away from the irradiated samples through the heatsink at the back of the lamps. The average irradiance of the target surface by the UVA lamp was 70.1 W/m² with a maximum deviation of 4.9%, and the average irradiance by the UVB lamp was 3.1 W/m² with a maximum deviation of 4.8%. With the UVA and UVB lamps, the temperature of samples undergoing irradiation in the incubator rises from 37 to 42 °C within 40 and 67 min, respectively. This by far exceeds the required UV irradiation time in most cases. Tests on Jurkat and HEK-293 cell cultures confirmed the suitability of our lamps for biomedical research.

Octocrylene (OCT) is one of the most widespread chemical UV filters used in sunscreens and cosmetic products. Despite the use of sunscreens and personal care products over decades, melanoma as the most serious and aggressive form of skin cancer is still a cause of concern. Hence the aim of this study was to investigate any potential influence of OCT on metabolic activity, cytotoxicity and ABCB5 mRNA expression in melanoma cells. The ABCB5 transmembrane protein was tested due to its well-known role in the initiation, invasion and metastatic spread of various cancers, including melanoma. Metastatic melanoma cell line WM-266-4 (ATCC) was incubated with selected concentrations of OCT and for different time intervals. The MTT and LDH assays to measure the cells' metabolic activity and cytotoxicity were used respectively. Target gene (ABCB5) expression was detected by quantitative real-time PCR (qRT-PCR), using TaqMan® chemistry. Our results suggest decreased metastatic melanoma cells' metabolic activity, increased cytotoxicity and increased ABCB5 mRNA expression (P<0.05) with longer time of exposure to OCT as compared to control cells. Accordingly, we suspect that the surviving cells are more invasive and aggressive, which might explain their microscopically observed cannibalistic activity. With this study, we elucidate a new promising field for further research to contribute to etiology and prevention of melanoma.

Titanium dioxide (TiO₂) is widely used as an inorganic UV-filter in cosmetic products; however, it has been classified as possibly carcinogenic to humans. While numerous studies demonstrated cytotoxic and genotoxic effects of nano-sized TiO₂ in different cell lines, including human skin cells, studies investigating the effects of micro-TiO₂ on human keratinocytes and melanocytes, in healthy and cancer cells, are scarce. Adenosine triphosphate (ATP) binding cassette subfamily B member 5 (ABCB5) is a plasma membrane protein known for its role in the tumorigenicity, progression, and recurrence of melanoma. Here, we investigated the effect of micro-TiO₂ (average particle size ≤5 µm) on the metabolic activity, cytotoxicity and ABCB5 mRNA expression in metastatic melanoma cells. Metastatic melanoma cell line WM-266-4 was treated with different concentrations of micro-TiO₂ for different incubation times to obtain dose- and time-dependent responses. Untreated WM-266-4 cells, cultured under the same conditions, were used as control. The cell metabolic activity was determined by MTT assay. Cytotoxicity of micro-TiO₂ was analyzed by lactate dehydrogenase (LDH) cytotoxicity assay. The ABCB5 mRNA expression in melanoma cells was analyzed using quantitative reverse transcription polymerase chain reaction (RT-qPCR). After 120 hours of exposure to micro-TiO₂ the metabolic activity of melanoma cells decreased, especially at the two highest micro-TiO₂ concentrations. Comparably, the cytotoxicity of micro-TiO₂ on melanoma cells increased after 48 and 120 hours of exposure, in a time-dependent manner. The ABCB5 mRNA expression in micro-TiO₂-treated melanoma cells also decreased significantly after 24 and 48 hours, in a time-dependent manner. Overall, our results suggest inhibitory effects of micro-TiO₂ on the metabolic activity and ABCB5 mRNA expression in metastatic melanoma cells, indicating its potential use as an anticancer agent.