Olgica Nedić

Dysregulation of microRNA expression in breast cancer (BC) has been associated with molecular disturbances involved in cancer initiation, progression and metastasis. Specific microRNAs also act as endocrine modulators in BC, thereby influencing the biological behavior of the tumor and drug responses. Our objective was to employ bioinformatics tools to identify and characterize microRNAs acting as candidate players involved in epithelial–mesenchymal transition, migration, invasion, and/or hormonal regulation in BC. We systematically integrated microRNA profiling data from three different studies on BC cell lines with different invasive capabilities and from another study on lymph node metastases and matching primary BC, resulting in five microRNA hits—DE-microRNAs miR-146a-5p, miR-222-3p, miR-205-5p, miR-141-3p and miR-200c-3p. This set of microRNAs was evaluated for clinical significance in BC and subjected to target prediction, microRNA–mRNA network construction, functional enrichment analysis and quantification in BC cell lines by qPCR. An upregulated DE-microRNA, miR-222-3p, displayed distinctive pro-metastatic features, supported by its clinical relevance, as well as by the results of the functional enrichment analysis of its target genes. Downregulation of the members of the miR-200 family and miR-205-5p were significantly associated with negative clinical features, while their targets were enriched with genes that were relevant to cancer aggressiveness. These results are in line with the presumed functional relevance of the selected DE-microRNAs in BC.

Beans (Phaseolus vulgaris L.) are an excellent raw material for developing vegan and gluten-free products because of their high content of proteins, dietary fiber, and bioactive compounds. This study aimed to evaluate the chemical composition, antioxidant capacity, and sensory properties of a fermented Tetovac (var.) bean-based product enriched with different microgreens and spices. Notable differences in phytochemical profiles were observed depending on the type of microgreens used. The contents of total phenolics and carotenoids (fresh weight basis) were highest in the sample with radish microgreens and saffron (1.178 mg GAE/g and 47.56 µg/g, respectively), whereas the highest total flavonoid content was recorded in the sample with pea microgreens (5.243 mg RE/g). In contrast, soluble sugar contents did not differ significantly among the samples (P > 0.05). Phytate content decreased in most samples, compared to the control (P < 0.05). Analysis of the protein profile of the sample containing pea microgreens during different storage periods revealed an increase in total protein content, particularly in the fraction of small proteins and peptides. Flavor intensity, sourness, bitterness, and firmness increased gradually during storage. Sensory evaluation revealed that samples containing basil and chive microgreens achieved high overall acceptability and purchase intention, whereas samples with radish microgreens were less acceptable due to pronounced bitterness and sourness.

Background and Purpose: Dietary interventions are a cornerstone in the management of diabetes mellitus type 2. The efficiency, however, depends on pharmacokinetic factors, including interaction of food ingredients with plasma proteins. The concept of this study was to investigate binding effects of three pronounced antioxidants present in the Mediterranean diet: resveratrol, (dihydro)lipoic acid and oleuropein, with albumin isolated from persons with diabetes (HbA1c 63±7 mmol/mol, or 7.9±0.6%) and healthy persons, carrying its intrinsic ligands. Experimental approach: Spectrofuorometric analysis, native electrophoresis and immunoblotting were performed with albumin before and after the interaction with antioxidants. Key results: Fluorescence spectra of the protein from two study groups were similar, whereas a spectrum of methylglyoxal-modified albumin (in vitro oxidised) was different. Calculated binding constants were also similar for two study groups for all three ligands. Kinetic fluorescence measurements revealed significantly altered activity of albumin-bound (dihydro)lipoic acid in persons with diabetes compared to healthy individuals, and no significant difference in the activity of resveratrol in expressing antioxidant protection of albumin upon its exposure to oxidative stress. Conclusions: Although the findings should be further validated using other antioxidants and glycated albumin derived from persons stratified according to the severity of a disease, the results have documented that in vitro methylglyoxal-oxidised albumin, routinely employed for diabetes-simulated investigations, was shown not to reflect this pathophysiological condition properly and not to be adequate for the assessment of relevant nutritional/biochemical potential of food antioxidants.

Miscanthus × giganteus was tested for textile dye removal. Sorption of Direct Blue 78 was achieved slowly by the leaf (63% after 24 h), while sorption of Basic Red 18 was fast by the stem (96% in an hour). Lignocellulose (24.62% in leaf, 41.34% in fresh and 48.05% in old stem) was responsible for the interaction. FTIR spectra and SEM images of native material and with sorbed dye were similar. Negligible quantities of peroxidases (2 μg/g in old stem) pointed to physical forces underlying sorption. pHpzc for stem-BR18 pair was 5.90 and maximum sorption could be achieved in pH interval 4-9. Desorption and repeated sorption defined maximal binding capacity of 20.8 mg BR18/g of stem. BR18 could be desorbed by only 23% with 0.1 M HCl. Small quantities of zinc (0.71-1.13%), copper (0.74-1.43%) and silicon (0.12-0.28%) were detected without significant difference between samples, as well as chlorine (0.24%) in the sample after desorption and in the sample with sorbed 20.8 mg/g BR18. We propose a more thorough investigation of M. × giganteus as a sorbent of a wider pallet of dyes, as it exerts a potential for such purpose.

MicroRNA-based regulatory mechanisms show disturbances related to oxidative stress (OS) interconnected with inflammation (IFM), as well as impairments associated with gestational diabetes (GDM). The aim of this study was to assess the diagnostic and prognostic significance of the OS/IFM-related microRNA in GDM by using peripheral blood mononuclear cells (PBMCs) and serum-derived extracellular vesicles (EVs) as biological samples. We selected the known OS/IFM-associated microRNAs miR-146a-5p, miR-155-5p, and miR-21-5p as candidates for our GDM biomarker analysis. Quantitative RT-PCR was employed for relative quantification of the selected microRNAs from paired samples of PBMCs and EVs derived from patients with GDM and healthy controls (n = 50 per group). The expression levels were analyzed for correlations with lipid and glycemic status indicators; metal ion-related parameters; serum thiol content; protein carbonyl and thiobarbituric acid-reactive substances’ (TBARS) levels; glutathione reductase (GR), Superoxide dismutase (SOD), and catalase (CAT) activity; and NRF2 expression. MiR-146a-5p and miR-21-5p were significantly upregulated in both PBMCs and EVs obtained from GDM patients. EVs-miR-21-5p showed a positive correlation with glycemic status in GDM patients, while miR-155-5p from PBMCs demonstrated correlation with iron-related parameters. The expression of selected microRNAs was found to correlate with NRF2 expression and SOD activity. The level of miR-146a-5p negatively correlated with neonatal anthropometric characteristics, while a higher level of PBMCs-miR-21-5p expression was determined in GDM patients with adverse pregnancy outcomes (p = 0.012). Our data demonstrate a disturbance of OS/IFM-microRNAs in GDM and illustrate their potential to serve as indicators of the associated OS-related changes, neonatal characteristics, and adverse pregnancy outcomes.

Gestational diabetes (GDM) is the most common pregnancy-related metabolic disorder and a major risk factor for both mother and a child, during pregnancy and after delivery. Impairment of the anti-oxidative system acts as the driving force of the devastating consequences in GDM, while microRNA molecules are affected by disturbances related to (glyco)oxidative stress ((g)OS) and emerged as potential sensors and effectors of (g)OS-associated mechanisms. The aim of the present study was to evaluate the potential of OS-related microRNAs miR-27a, miR-222 and miR-340 from peripheral blood mononuclear cells (PBMCs) to serve as OS indicators of in GDM. MicroRNA quantification was conducted in samples of patients with GDM and normoglycemic controls (n = 45 each). Levels of expression were tested for correlations with the activities of glutathione reductase (GR), superoxide dismutase (SOD) and catalase (CAT) and serum thiol content. MiR-27a demonstrated correlation with the expression of other two microRNAs. GR activity during pregnancy weeks 24-28 was increased in GDM, while thiol content showed an opposite direction of change. Similar results were acquired when comparing GDM patients with and without adverse pregnancy outcomes. The expression of tested microRNAs was positively correlated with the activity of GR in GDM group, while the correlation of the expression of miR-27a-3p and miR-340-5p with SOD activity and blood thiol content was negative. Our findings support the relevance of redox dysregulation in mid-pregnancy for developing obstetric and neonatal complications. The results also qualify miR-27a-3p, miR-222-3p and miR-340-5p for redox status indicators in GDM.

A fast and simple biotech method is presented for the simultaneous isolation and purification of transferrin (Tf) and immunoglobulin G (IgG) from the same pool-sample of human serum, yielding >98% pure proteins. Serum sample preparation was achieved by precipitation with ethacridine lactate (rivanol). Protein purification was performed with AKTA Avant 150 FPLC, using a Resource Q column. Three different buffers at pH 6.2 (MES, phosphate, and Bis-Tris) were tested. Isolated and purified proteins retained their native 3D structure, as shown by spectrofluorimetric measurements. Tf functionality was preserved, as confirmed by the retention of both the iron binding capacity and its ability to interact with the transferrin receptor (immunofluorescent staining), as well as the immunogenicity of IgG, as shown by Western blot analysis with immunodetection. The formation of IgG aggregates was avoided. This biotech method is a rapid, simple, and time-saving alternative to other methods for the isolation of extremely pure IgG and Tf, while it is also the only method so far described for their simultaneous isolation.

Poor bioavailability and reduced stability are the main drawbacks to efficiently utilizing many naturally occurring antioxidants, so their binding to circulatory proteins is essential. This work investigated whether major human circulatory proteins, besides albumin, including transferrin, alpha-2-macroglobulin, and fibrinogen, bind widely consumed antioxidants and food supplements, including quercetin, trans-resveratrol, and dihydrolipoic acid, thus filling the gap of detailed pharmacokinetic properties of these food supplements. Detailed examination of the protein structural and functional changes that occur upon ligand binding was analyzed by spectroscopic methods and in silico docking and molecular dynamic simulation studies on the model that consists of the protein/antioxidant pair with the highest affinity constant. It was found that alpha-2-macroglobulin binds trans-resveratrol with the highest affinity (K_a of 4.5 x 10⁴ M^-1). In silico results revealed four potential binding sites between trans-resveratrol and alpha-2-macroglobulin, with hydrogen bonds being crucial for binding, while other observed interactions (primarily aromatic interactions) are of secondary importance. The binding of trans-resveratrol to alpha-2-macroglobulin leads to mutual protection of both molecules from oxidative stress and significantly increased hidrosolubility of resveratrol, both of which could serve to increase the bioavailability and bioactivity of resveratrol in circulation.

Recent findings highlighted the potential of long non-coding RNAs (lncRNAs) as novel indicators of gestational diabetes mellitus (GDM), as they demonstrate altered expression in metabolic disorders, oxidative stress (OS) and inflammation (IFM). The aim of this study was to evaluate the diagnostic potential and prognostic significance of the OS/IFM-related lncRNAs H19, MALAT1 and MEG3 in GDM and their correlations with redox status-related parameters. The relative quantification of selected lncRNAs from peripheral blood mononuclear cells (PBMCs) of GDM patients and controls (n = 50 each) was performed by qPCR. The expression levels were tested for correlations with metal ion concentrations, NRF2 expression, activities of glutathione reductase (GR), superoxide dismutase (SOD), catalase (CAT), serum thiol content, protein carbonyl level and thiobarbituric acid reactive substances. MALAT1 and H19 were significantly downregulated in GDM patients (p = 0.0095 and p = 0.012, respectively). A correlation was observed between H19 expression and zinc levels in both GDM patients and controls. MALAT1 expression positively correlated with NFE2L2 levels in GDM patients (p = 0.026), while H19 exhibited a positive correlation with GR activity in controls (p = 0.018) and an inverse correlation with SOD activity (p = 0.048). Our data show the disturbance of OS/IFM-lncRNAs in GDM pathogenesis and illustrate the biomarker potential of the analyzed lncRNAs, as well as of certain redox status parameters.