Vitamin E (alpha-tocopherol, VE) is essential to prevent severe neurological symptoms and even death of a genetic form of ataxia associated with vitamin deficiency or AVED. Its essentiality is also proven in secondary deficiencies associated with malnutrition and/or malabsorption syndromes that besides moderate to severe neurological abnormalities can contribute to induce metabolic, musculoskeletal, hematological, and immune dysfunctions, especially in the elderly. VE is the most abundant and ubiquitous fat-soluble nutrient with hydrogen atom donating properties (often described as “antioxidant”) of the plasmalemma; its relative abundance with respect to phospholipid residues is by far the highest among other H donors and its membrane levels influence the flux of lipoperoxyl radicals during both enzymatic and non-enzymatic processes of lipid peroxidation. Consequently, VE directly affects the metabolism and function of membrane fatty acids, also playing a key role in lipid signaling and thus in the indirect control of different enzymes, signal transduction, and transcriptional proteins that connect, under a functional point of view, the VE levels in human tissues with many pathophysiological aspects and deficiency symptoms. Recent evidence strongly supports the participation of the long-chain metabolites of VE in at least some of its “non-antioxidant” properties. Altogether these aspects depict the biological complexity of this vitamin which is far from being comprehensively understood. Last-generation omics technologies make it possible to face such a complexity to represent with unprecedented efficacy both the essentiality aspects and the health-promoting potential of this vitamin in human nutrition studies and clinical trials on deficiency syndromes and other human diseases that may benefit from its biological properties. Transcriptomics and especially metabolomics protocols have been utilized in our laboratories, either separate or in multiomics mode, to develop personalized and precision nutrition (i.e. nutrigenomics) platforms of investigation dedicated to this vitamin, and examples of their potential for innovation in VE research will be given in this presentation, including in vitro studies and clinical trials on hepatic fatty acid metabolism and lipotoxicity, the etiologic factor of non-alcoholic fatty liver disease, and studies in kidney disease patients that develop secondary VE deficiency in the context of severe oxidative stress and lipid peroxidation symptoms.

The selenoperoxidase GPx4, discovered in 1982, plays a pivotal role in preventing ferroptosis. In a moonlighting function, GPx4, in its mitochondrial and nuclear forms, also contributes to spermatogenesis. The critical advantage of Selenium vs. Sulfur catalysis is the stability of the oxidized form of the chalcogen in the catalytic cycle. While the mechanisms of catalytic cycle are understood, its regulation remains largely unknown. Existing evidence supports the notion that ferroptosis is activated when GPx4 is inhibited, glutathione (GSH) concentration is lowered, or the labile iron pool is expanded. The outcome is framed in the context of oxygen toxicity playing the physiological function of controlling cell death. GPx4 stands out as the sole peroxidase indispensable to aerobic life. Moreover, a recent study exploring the role of the residue Arg152 in GPx4, linked to a fatal although not embryonically lethal disease, revealed that the wild-type enzyme exhibits surface-sensing and positive cooperativity in the presence of cardiolipin. This adds complexity to the mechanism of physiological function encompassing the interaction with acidic phospholipids in mitochondrial membranes. Ferroptosis is implicated in both physio-pathological conditions, including embryogenesis, cancer suppression, neurodegenerations, inflammatory disorders, metabolic syndrome, heart and kidney diseases. No antioxidant enzymatic system can substitute for GPx4 in inhibiting ferroptosis, emphasizing the vital role of selenium. Phenolic antioxidants, which reduce lipid hydroperoxyl radicals, can only inhibit lipid peroxidation under physiological conditions, and thus ferroptosis, when the lipid hydroperoxides formed are immediately reduced by GPx4. In contrast, the ferroptosis inhibitor Ferrostatin-1 (Fer-1) proves to be significantly more efficient than phenolic antioxidants. Analytical and computational evidence supports the notion of a pseudo-catalytic cycle where the ferrostatin-iron complex, both produces and reduces lipid alkoxyl radicals from lipid hydroperoxides. This discloses the roadmap for the identification of innovative antioxidants competent for preventing ferroptosis.

Ferroptosis, a form of regulated cell death characterized by iron-dependent lipid peroxidation, is linked to neurodegenerative disorders and cold-induced cell death. SLC7A11 (xCT) plays a crucial role in protecting cells against ferroptosis by maintaining intracellular cysteine and glutathione levels. SLC7A11 requires the chaperone protein SLC3A2 for its localization on the plasma membrane to mediate cystine uptake. Arctic ground squirrels (AGS) are known to be protected from cold tissue temperatures and oxidative stress and to resist neuropathology following cerebral ischemia/reperfusion. This study investigated how ferroptosis is influenced by the hibernation season in AGS hippocampus. RNA-Seq, gene expression, and differential gene expression analysis were conducted on hippocampus tissue samples from male and female AGS collected during the summer active season, torpor, and interbout arousal (IBA). Hippocampus was dissected from partially thawed whole brain prior to RNA extraction.  Total RNA samples were used for cDNA library construction and sequencing by BGI Americas Corporation (Cambridge, MA) and analyzed using CLC Genomics Workbench (QIAGEN). Genes were mapped to the Ictidomys tridecemlineatus reference genome and transcript (HiC_Itri_2, GCF_016881025.1). Results show the highest number of differentially expressed genes (4,042) in torpor compared to summer active animals. Notably, SLC7A11 expression was elevated in torpor compared to summer active animals (fold change: 1.80, FDR-p value: 0.0034). Additionally, SLC3A2 was significantly upregulated in torpor compared to IBA (fold change: 1.24; FDR-p value: 0.030). SLC7A11 transports glutamate(out)/cystine(in). Cystine is rapidly converted into cysteine, a limiting reactant for glutathione synthesis, in the presence of NADPH. These findings suggest that SLC7A11 and SLC3A2 may protect AGS from ferroptosis during the hibernation season. This research provides insights into the molecular mechanisms underlying neuroprotection in hibernating AGS and may have implications for understanding and potentially treating neurodegenerative disorders.

In recent decades, a global increase in the incidence of skin cancer, particularly squamous cell carcinoma (SCC), has been observed. To explore the pathogenesis and potential therapeutic approaches for this cancer type, in vivo studies employing various mouse models and ultraviolet (UV) light have been conducted. A comparative study on skin carcinogenesis across four hairless mouse models subjected to UV light exposure was initiated. The mouse strains utilized in this research were: SKH-hr1, SKH-hr2, SKH-hr2+ApoE, and immunodeficient Nude. Based on the various measured parameters, in contrast to the SKH-hr1, SKH-hr2+apoE and SKH-hr2 models were identified as the most appropriate.  The bark extract of Pinus maritima (PBE) was examined for SCC preventive action. It was evaluated in two different experimental animal tumor models induced by ultraviolet radiation (UVR) and combination of UVR with 7,12-dimethylbenz[a]anthracene. A significant decrease in the number of animals bearing tumors, increase in viability and delayed appearance of tumors were observed. Through immunochemical analysis, the expression of P-glycoprotein, multi-drug resistance-associated protein (MRP), and glucose (GLUT-1) transporters in SCC, SCC adjacent area, and normal skin tissues were examined. It was revealed that all assessed transporters were expressed across all skin tissues; however, expression levels were notably higher in tumor and tumor-adjacent areas compared to normal tissues. Male and female hairless SKH-2 mice were exposed for 10 months to cigarette smoke (CS) and/or UV light after administration or not of French maritime pine bark extract (PBE) to study the SCC induction and possible protection by PBE. The results showed that UV and CS were harmful and act synergistically inducing SCC, whereas PBE seems to protect skin against SCC. Type 1 and 2 diabetic, and nondiabetic male mice were exposed to UV radiation for eight months. Remarkably, Type 1 diabetic mice did not develop squamous cell carcinoma or pigmented nevi, contrary to normal and Type 2 diabetic skin. Type 1 diabetic mice showed protection against oxidative stress.

Ageing is a complex process characterised by the gradual deterioration of physiological functions and increased susceptibility to various age-related diseases. Mitochondrial dysfunction is an important factor contributing to ageing. Sirtuin 3 (Sirt3), a mitochondrial protein essential for energy homeostasis, plays a critical role in maintaining mitochondrial function, as loss of Sirt3 reduces energy and impairs cellular repair, which accelerates ageing. The aim of this study was to investigate the role of Sirt3 in male and female mouse embryonic fibroblasts (MEF) exposed to etoposide-induced DNA damage. We employed state-of-the-art genetic, molecular, and imaging technologies as well as metabolomic analyses to provide insights into the molecular mechanisms underlying these responses. We found that the loss of Sirt3 affected metabolic responses differently depending on sex: while male MEF showed minimal damage, possibly due to earlier stress adaptation, female MEF lacking Sirt3 were more vulnerable, suggesting that Sirt3 plays a critical role in enhancing their ability to withstand such challenges. By focusing on Sirt3 and sex-specific signalling pathways it modulates, this study has a potential for developing new strategies to combat diseases associated with DNA damage — a cornerstone of the ageing process.

Colorectal cancer presents a significant global health challenge, with a high mortality rate. It is the third most commonly diagnosed cancer and is therefore a major cause for concern. The development of colorectal cancer is multifaceted, involving a combination of genetic predispositions and lifestyle factors. The redox and metabolic states may influence the intricate process of colon cancer development. To gain a deeper understanding of the redox-metabolic profiles associated with colon cancer, a human study was conducted. In biopsies from patients with colon cancer, the antioxidant status: copper, zinc superoxide dismutase (CuZnSOD), manganese superoxide dismutase (MnSOD), catalase (CAT), glutathione peroxidase (GSH-Px), glutamate-cysteine ligase (GCL), thioredoxin (Trx) and lactate metabolism were examined in tumor and unaffected colon tissue (remote 15-20 cm) as well as in adipose tissue: proximal (near the tumour tissue), distal (remote 6 cm) and unaffected (remote over 6 cm). The protein levels of CuZnSOD, MnSOD, GSH-Px, and Trx are increased in the tumor tissue compared to the unaffected colon tissue. In addition, the expression of the lactate dehydrogenase (LDH) A isoform, the total activity of LDH and the lactate concentration are higher in transformed tumor tissue than in normal colon tissue. On the other hand, lactate concentration increases and several AD components (CuZnSOD, MnSOD, CAT, GSH-Px, GCL and Trx) decrease in adipose tissue with tumor proximity. Shifts in redox and lactate metabolism in tumor tissue associated with spatial changes in lactate and antioxidant enzymes gradients in adjacent adipose tissue clearly indicate a local redox metabolic interaction between tumor and tumor-associated adipose tissue in shaping the malignant phenotype in human colorectal cancer.

Ionising radiation damages DNA directly, or indirectly, causing water radiolysis and formation of free radicals. Indirect irradiation effects could be diminished in the presence of free radical scavengers, such as dimethyl sulfoxide (DMSO). Such conditions would allow the evaluation of direct radiation effects and provide a better understanding of cellular response to irradiation-induced damages. The goal of this study was to investigate the effects of low (γ-rays) and high linear energy transfer (LET) radiation (carbon ions) in non-small lung cancer cells HTB177. Cells were pre-treated with DMSO and irradiated with 60Co γ-rays and 62 MeV/u carbon ions, with doses ranging from 1-5 Gy. Results obtained by clonogenic survival and γ-H2AX foci assay showed that DMSO increased cell survival and decreased number of DNA damages, which points to radioprotective effect of DMSO. The contribution of direct and indirect radiation effects was estimated by the degree of protection (DP) in presence of DMSO. The values of DP rose in a concentration-dependent manner in all irradiated samples. In cells irradiated with γ-rays, 35% of damages were caused directly, while 65% of lesions could be attributed to indirect radiation actions. In presence of carbon ions, contribution of direct effects was 49%, while 51% of damage resulted from indirect radiation effects, showing that free radicals attain an important role in both low and high LET irradiations. The obtained results showed that DMSO can be used as a free radical scavenger to examine the direct and indirect effects on human cancer cells. The numerical Monte Carlo simulations allow modelling of direct and indirect irradiation actions in cancer cells with photons and hadrons. Therefore, this data will be used for validation and further improvement of numerical simulations in comparison to the data collected on different cell lines and irradiation energies, with the goal to improve therapeutic protocols.

Mechanisms and factors that lead to the formation of lipomas, benign tumors of adipose tissue, are still insufficiently elucidated. Mesenchymal stem cells (MSCs) isolated from lipomas have some similar characteristics to MSCs isolated from white adipose tissue but differ at the molecular level and in their differentiation potential. Considering histological appearance of lipomas, it is not clear to what extent lipomas share common characteristics with other adipose tissue type, brown adipose tissue. Therefore, the aim of this study was to examine the level of uncoupling protein 1 (UCP1), a marker of brown adipose tissue, expression in lipoma tissue as well as in MSCs isolated from lipomas, i.e. lipoma-derived mesenchymal stem cells (LDSCs). LDSCs were grown in standard cell culture conditions and subjected to adipogenic differentiation. UCP1 expression was examined at the RNA level, using Real-Time PCR, and at the protein level, using immunohistochemistry and immunogold staining. Expression of UCP1 in lipoma tissue and LDSCs was compared with the expression of UCP1 in subcutaneous white adipose tissue (scWAT) and adipose-derived mesenchymal stem cells (ADSCs) grown and differentiated in the same cell culture conditions. Differences were observed in UCP1 expression at both RNA and protein levels in lipomas compared to scWAT directing the future research towards the potential of browning mechanisms of adipose tissue involved in lipoma tissue formation.

This research was financially supported by the Science Fund of the Republic of Serbia, PROMIS, #6066747, WARMED and the Ministry of Science, Technological Development and Innovations of the Republic of Serbia, Contract No. 451-03-65/2024-03/200113.

Multiple sclerosis (MS) is a chronic, immune-mediated inflammatory disease of the central nervous system (CNS). Relapsing-remitting multiple sclerosis (RRMS) is the most common clinical subtype of MS. MS is characterized by demyelination and myelin is mainly composed of lipids. Lipids play many roles in the CNS including signaling, structural support, mediating inflammation, and membrane biogenesis. Omega-3 polyunsaturated fatty acids (PUFA) are central to maintaining health and they are present in a wide array of tissues with broad functions including the active component of phospholipid cell membranes and substrate for molecular signaling pathways. This study aimed to evaluate fatty acids (FA) profiles of patients with RRMS (n=30) compared to healthy people (n=20). Analysis of total lipids was performed from erythrocyte samples. The total lipid extracts from erythrocytes were prepared by adding chloroform/methanol (2:1, v/v) mixture containing butylated hydroxytoluene (0.05% BHT weight/volume). FA methyl esters were prepared by transmethylation with 3N HCl in methanol. FA profiles were determined by gas chromatography (GC). The content of individual FA was expressed as a percentage of the total FA. Results of this study revealed that total saturated fatty acids (SFA) are significantly higher in MS patients compared to controls. While total PUFAs, total n-3 PUFAs, and omega-3 index are statistically lower in MS patients. The n-6/n-3 ratio is significantly higher in MS patients compared to controls. Also, the AA/EPA ratio is significantly lower in the control group compared to MS patients. Conversely, the EPA/AA index is significantly reduced in MS patients. Omega-3 lipids, which have a protective role by preserving the blood-brain barrier, are significantly reduced in the erythrocytes of patients with MS. Increased n-3 PUFA and decreased SFA intake could counteract inflammation, energy storage and utilization imbalance and, overall state in patients with MS.

Amphibians are the most threatened vertebrate class. Exposure to pesticides and climate change are considered pivotal factors in the global decline of their populations. Glyphosate-based formulations are the most widely used herbicides, but increasing evidence of their harmful effects, including oxidative stress in exposed organisms, has sparked a heated debate. Current climate prediction models assume a global temperature rise of 3 °C to 5 °C in the coming decades. In poikilotherms, any changes in ambient temperature are directly translated into changes in physiological and biochemical processes. Furthermore, elevated temperatures could intensify the toxic effects of pesticides present in the environment. The aim of this study was to examine the effects of low, environmentally realistic concentrations of glyphosate-based herbicides (30 µg/L active ingredient) and elevated temperature (optimal t1=19°C and increased t2=23°C) on glutathione content (GSH), antioxidant enzyme activities (SOD, CAT, GSH-Px, GR and GST), activity of acetylcholinesterase (AChE) and levels of oxidative damage (TBARS - lipid peroxidation and PC - protein carbonylation) in larvae of the Balkan crested newt (T. ivanbureschi). Our findings revealed that glyphosate had a significant effect on the activity of all antioxidative enzymes, with the exception of SOD. Herbicide and elevated temperature led to a significant increase in the activities of CAT, GSH-Px, GST, and GR, as well as GSH concentration. This response of the antioxidative defense system prevented oxidative damage to lipids and proteins. Glyphosate exhibited a neurotoxic effect by inhibiting AChE only at elevated temperatures, while no significant change occurred at the optimal temperature. The findings suggest the importance of examining the potentially harmful effects of glyphosate in different ecological contexts, such as an increase in average temperatures by several degrees predicted by future climate scenarios.