Purpose: Aging is a complex biological process that begins at the cellular level, disrupting energy homeostasis. This study investigated the role of Sirt3, major mitochondrial deacetylase involved in metabolic pathways, in sex-dependent changes in energy homeostasis during aging in kidney of Sirt3 WT and KO mice.

Methods: Enzymatic activity, lipid peroxidation, protein carbonylation with Western blot and metabolomic analyses were performed to assess physiological and metabolic parameters

Results: Higher Sirt3 expression in male WT mice leads to increased vulnerability to its deficiency, as reflected in the shorter lifespan of male KO mice. This is further supported by distinct metabolomic clustering in male KO mice, highlighting significant metabolic disruptions. Male-specific declines in metabolites such as creatine, phosphorylcholine, trimethylamine-N-oxide, and L-carnitine, along with reduced trifunctional multienzyme complex subunit β (HADHB) expression, point to impaired fatty acid metabolism and mitochondrial dysfunction.

Conclusions: The findings emphasize the sex-specific function of Sirt3 in regulating mitochondrial activity, energy metabolism, and oxidative stress in the murine kidney, with male mice exhibiting a greater reliance on Sirt3 for metabolic stability.

A high proliferation rate and the malignancy of cancer cells are favoured by redox and metabolic plasticity, which is determined by the co-evolution of cancer cells with their host microenvironment. The tight functional connections between the mammary glands' epithelium and adipose tissue (AT) allow breast cancer cells to subjugate the AT and form a protumorigenic cancer-associated adipose tissue (CAAT). Our findings in luminal invasive ductal carcinomas in premenopausal women confirmed key cancer cell strategies - the Warburg effect, increased mitochondrial metabolism and redox adaptability, which are associated with a specific shift in the metabolic and redox phenotype of CAAT. Notably, the upregulated master redox-sensitive transcription factor Nrf2 appears to be responsible for the cancer cell-induced redox and metabolic shift of CAAT. We also investigated the role of Nrf2 in the metabolic co-evolution of cancer cells and CAAT during disease progression. Our results in the orthotopic breast cancer mouse model and in the co-culture of breast cancer cells with adipocytes confirmed the different spatiotemporal redox and metabolic properties of cancer cells and CAAT, established with respect to the Nrf2-coupled/uncoupled tumour microenvironment. The uncovered metabolic and redox strategies adopted by breast cancer cells according to CAAT properties and at different disease stages have helped to better understand the biology of the aggressive disease and to identify breast cancer vulnerabilities that could become therapeutic targets.

This research was supported by the Science Fund of the Republic of Serbia, #7750238, Exploring new avenues in breast cancer research: Redox and metabolic reprogramming of cancer and associated adipose tissue - REFRAME.

Since the discovery of the thermogenic role of brown adipocytes, there was consensus that the biochemical and metabolic function of their mitochondria is uniform. By switching the ATP production between glycolytic pathway and oxidative phosphorylation, brown adipocytes are able to produce heat in mitochondria through uncoupling protein 1 (UCP1). Thermogenically active brown adipocyte mitochondria are characterized by clear morphological features (long, tightly packed cristae). The process of their biogenesis includes an increased number of mitochondria (by division), increase of their surface area, and incorporation of UCP1 as well as specific structural organization of the cristae. But, is it true that all BA mitochondria within one cell are structurally and functionally the same? Do they harbor the same set of enzymes? Actually, the very first cell mosaicism, e.g. Harlequin appearance was shown in brown adipose tissue. This unique uneven UCP1 expression suggests that brown adipocyte’s mitochondria may be heterogeneous regarding production of ATP (bioenergetic) vs. heat (thermogenic) role. This presentation deals with structural and functional mitochondrial mosaicism and changes caused by insulin.

This research was supported by the Science Fund of the Republic of Serbia, #7750238, Exploring new avenues in breast cancer research: Redox and metabolic reprogramming of cancer and associated adipose tissue - REFRAME.

A deep understanding of the roles of redox metabolites and pathways in physiology and pathology requires molecular tools that enable both visualization of these processes and their selective modulation. Over the last two decades, a number of genetically encoded fluorescent biosensors for key redox metabolites have been developed, allowing real-time detection in living systems of varying complexity. Recent developments in this area include the ultrasensitive probe HyPer7 and a new fluorogenic probe, HyPerFAST, which enables even more sensitive H2O2 detection across any chosen optical range, from blue to near-infrared. Complementary to imaging with biosensors, chemogenetics offers tunable substrate-dependent modulation of metabolic pathways, allowing the study of normal cell functioning and modeling dysfunctions caused by abnormal pathway activity and/or metabolite levels. We will present recent developments in this area that include insights on oxidative stress brought about by the use of D-amino acid oxidase (DAO) and intriguing details of the Warburg effect brought about by a new mitochondrial "booster," Grubraw, based on bacterial D-amino acid dehydrogenase.

Oxidative biochemistry centered about 35 years ago on the one-electron reduction of H₂O₂ by Fe²⁺, the Fenton reaction, to yield HO^· and a Fe(III)-complex. The discovery that NO^· is formed in vivo and that it reacts with O₂^·− at a diffusion-controlled rate led to ONOO⁻ as an additional oxidant. The rate constant of the Fenton reaction is 53 M⁻¹s⁻¹ up to about pH 4, but above it the rate constant increases linearly with pH.  This acceleration of the Fenton reaction led to the hypothesis that above pH 5 formation of FeO²⁺ predominates.  Thermodynamically, this species is comparable to HO^· as an oxidant.  HCO₃⁻ accelerates the reaction even more, and convincing evidence has been presented that the complex of Fe²⁺ with CO₃²⁻ reacts with H₂O₂ to form CO₃^·− and a Fe(III)-complex, conceivably via FeO²⁺ as an intermediate. The rapid reaction of ONOO⁻ with CO₂ (k > 10⁷ M⁻¹s⁻¹) leads to ONOOCO₂⁻ that, depending on the CO₂ concentration, yields varying amounts of NO₂^· and CO₃^·−.  These two oxidizing radicals together nitrate aromatic residues. Compared to 35 years ago, oxidative biochemistry is no longer concerned with the indiscriminate oxidations and additions of HO^·, but with the more selective reactions of CO₃^·− and NO₂^·.

The human skin, being our outermost protective barrier, sustains continuous contact with the environment. As such, its cells must be kept in a state of constant alert against external increased oxidative stress and massive environmental insults (e.g. sunlight and UV radiation, air pollution, and mechanical stress). All these insults ultimately result in an impaired redox balance and increased cellular oxidation. One of the pivotal oxidation regulation mechanisms in the skin is the Nrf2–Keap1 pathway, and its activity leads to cutaneous redox maintenance which evidently sustains the principle of hormesis. We suggest that moderate environmental stressors and skin microbiome can provide the necessary continuous stimuli for the activation of the Nrf2 pathway. We also suggest that endogenous neurotransmitters play a major role in this activation.

Human extravillous trophoblasts play a key role in implantation, placentation, and successful pregnancy outcomes due to their ability to migrate and invade through the uterine spiral arteries. Abnormalities in the trophoblasts' migratory and invasive abilities may result in insufficient remodeling of the uterine spiral arteries. This leads to the development of preeclampsia, a syndrome in pregnancy. Neuropeptide Y (NPY) is a sympathetic neurotransmitter that plays a significant role in the adaptive stress response as well as in the regulation of body energy balance. This study was designed with the aim of investigating whether preeclampsia is associated with NPY-induced disruption of trophoblast migration and redox balance. For this purpose, the concentration of NPY was determined in the plasma of 20 healthy and 20 preeclamptic pregnant women. The obtained results demonstrated that in preeclampsia, the concentration of NPY is significantly lower (190 pg/mL) than in a healthy pregnancy (260 pg/mL). After that, NPY in concentrations of 190 pg/mL and 260 pg/mL was used to treat the human extravillous trophoblast cell line HTR-8/SVneo for ten weeks. The effect of NPY on trophoblast proliferation was determined by counting cells during each passage. After the end of the treatment, the effect of NPY on migration and intracellular concentrations of superoxide anion radical (O2.-), hydrogen peroxide (H2O2), and nitric oxide (NO) were examined. The obtained results show that NPY induces changes in trophoblast proliferation and reduces their migration at both applied doses. In addition, both doses of NPY induce a decrease in intracellular concentrations of O2.-, H2O2, and NO. The NPY concentration of 190 pg/mL significantly decreased O2.- in trophoblasts in comparison to the concentration of 260 pg/mL. This study demonstrates that NPY affects the migration and redox balance regulation of trophoblasts. It also disrupts the trophoblast redox balance at a level characteristic of preeclamptic pregnancy.

The global pandemic crisis affected almost every society and economy, challenged almost every health system worldwide. Above all, governments and non-governmental organizations had to fight the misinformation and conspiracy theories placed by the social and mass media. All of this had a profound impact on the public in terms of vaccine safety and the advantages of vitamin use in fighting the virus. This fear has opened doors to alternative medicines such as supplements (vitamins, minerals, herbal products, oils) that may have profound effects on the immune system. To determine the pattern of use of supplements during the pandemic in healthy individuals who tested negative for SARS-CoV-2. The 33 healthy individuals tested negative for SARS-CoV-2 in the pandemic period were included (Group 1). Total antioxidant power, iron-reducing (PAT), and plasma peroxides (d-ROMs) were measured using FRAS5 analytical photometric system and are reported in equivalents of ascorbic acid and H2O2, respectively. The oxidative stress index (OSI) was automatically calculated by the software. The obtained values were compared with 30 healthy individuals analyzed prior to the pandemic (Group 2). The mean values for oxidative stress parameters in Group 1 vs Group 2 were: d-ROMs 418 vs 266 U. Carr, PAT 3862 vs 2554 U. Carr, and OSI 111 vs 36. In all comparisons, a statistically significant difference was obtained (p<0.05, t-test). Individuals belonging to Group 1 had reported that they have consumed daily doses of Zinc (30 mg), Vitamin C (at least 1000 mg) and Vitamin D (at least 2000 IU) in a period of >1 month. Several of them have also used Isoprinosine, magnesium, and selenium. Uncontrolled intake of supplements can have a profound effect on the pro- and antioxidant balance resulting in interruption of the phycological balance and leading to increased oxidative stress index in otherwise healthy individuals. 

Canine hyperadrenocorticism (HAC) or Cushing’s syndrome is a multisystemic clinical condition caused by chronic exposure to elevated concentrations of glucocorticoids. It has been considered that oxidative stress is implicated in pathophysiology of HAC. The exact impact of uric acid (UA) on oxidative stress in hyperadrenocorticism remains unclear, given its ability to act as both an antioxidant and a pro-oxidant. In addition, increased UA levels are related to the development of hypertension, dyslipidemia, and type II diabetes in humans with HAC. For this purpose, we aimed to investigate the association of UA with the components of oxidative stress in dogs with HAC. This study included 12 dogs with newly diagnosed HAC and 12 healthy controls. The oxidative stress in serum samples was assessed by advanced oxidation protein product (AOPP) and thiobarbituric acid–reactive substances (TBARS), and antioxidative status by total antioxidant capacity (TAC), reduced glutathione (GSH) and paraoxonase-1 (PON-1). Uric acid was compared between two groups and correlated with oxidative stress parameters. The results showed that dogs with HAC exerted markedly higher level of UA compared to healthy controls (p<0.001). Additionally, higher levels of AOPP and TBARS (p=0.001; p =0.043) were observed in the HAC group, indicating oxidative damage compared to the controls. Among antioxidants, only GSH exhibited a difference between groups (p=0.001). Correlation analysis of UA revealed strong association with TBARS (r=0.615; p=0.037), which implies that UA is linked to an increase of oxidative stress in canine Cushing’s syndrome. The results of this study indicate a possible pro-oxidant role of UA in dogs with HAC. 

A 60-day experiment was conducted to investigate the impact of dietary sulfur amino acids (AA) on altered haemato-biochemical and redox parameters of Inshas cockerel chicks raised under restricted feeding. Male Inshas strain chickens were divided into five groups, each with five replicates of eight birds. The control group received the full National Research Council (NRC) requirements (100%). The other groups received diets meeting 90% of NRC requirements: 90% NRC; 90% NRC+Methionine; 90% NRC+Cysteine; and 90% NRC+both Methionine and Cysteine (AA-mix). AA supplementation improved growth performance compared to the control group. The birds that were given a combination of AA supplementation exhibited the highest body weight and carcass weight compared to other groups. AA supplementation improves blood physiological characteristics by reducing damage caused by feed restriction conditions. Serum parameters, including aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase, alkaline phosphatase, gamma-glutamyl transpeptidase, and total protein concentration, showed decreased levels. Treatment with mixed AA maintained urea and uric acid concentrations at a level similar to the control group. Remarkably, a combination of AA treatments reduced the negative effects of feed restriction on young male chickens by enhancing the overall antioxidant capacity and activity of antioxidant enzymes in liver tissue (glutathione S-transferase, total superoxide dismutase, glutathione peroxidase, and amount of total glutathione), and decreased the malondialdehyde concentration. Feed restriction impacted liver histological structure, where hepatocytes were susceptible to feed restriction and included numerous cytoplasmic vacuoles, congested blood vesicles, lymphocytic infiltration, and pyknotic nuclei in treated cockerels. AA therapy restored most hepatic histological abnormalities. The findings suggest that AA supplementation significantly mitigated the adverse effects of feed restriction by improving haemato-biochemical parameters and hepatic redox status.

Acknowledgement: The authors extend their appreciation to the Department of Animal and Poultry Production, Faculty of Agriculture, Minia University, Egypt.