Free Radical Research最新文献

Chronic kidney disease (CKD) is a progressive condition marked by persistent kidney damage, leading to high mortality rates and economic burden in advanced stages. Ozone therapy has emerged as a complementary alternative capable of mitigating oxidative stress involved in CKD progression. Ozonated saline solution (OSS) prepared via microbubbling offers enhanced efficacy due to greater ozone dissolution, homogeneity, and stability compared to conventional methods. This study compared the biosafety and efficacy of OSS prepared through bubbling and microbubbling methods in advanced CKD patients. In vitro, hydrogen peroxide (H₂O₂) concentrations were measured at various doses and times for both methods. In healthy volunteer, biosafety was assessed using TMRE and Annexin V in leukocytes. In CKD patients, TMRE, Annexin V, redox markers (catalase, superoxide dismutase, glutathione system, H₂O₂, lipoperoxidation), and renal function markers (urea, creatinine, glomerular filtration rate) were evaluated. Microbubbling produced lower H₂O₂ concentrations in vitro, depending on time and ozone dose. In vivo, both methods increased mitochondrial activity and apoptosis in CKD patient leukocytes. However, microbubbling notably enhanced antioxidant capacity, catalase and superoxide dismutase activity, and redox balance (elevated reduced-to-oxidized glutathione ratio) compared to conventional bubbling. It also showed slight improvements in serum clinical parameters. In conclusion, the microbubbling method demonstrated superior biosafety and therapeutic efficacy in advanced CKD patients, highlighting its potential as a preferred approach in ozone therapy.

Inflammatory bowel diseases (IBD), which include Crohn's disease and ulcerative colitis, represent a global health issue as a prevalence of 1% is expected in the western world by the end of this decade. These diseases are associated with a high oxidative stress that induces inflammatory pathways and severely damages gut tissues. IBD patients suffer from antioxidant defenses weakening, through, for instance, an impaired activity of superoxide dismutases (SOD)-that catalyze the dismutation of superoxide-or other endogenous antioxidant enzymes including catalase and glutathione peroxidase. Manganese complexes mimicking SOD activity have shown beneficial effects on cells and murine models of IBD. However, efficient SOD mimics are often manganese complexes that can suffer from decoordination and thus inactivation in acidic stomachal pH. To improve their delivery in the gut after oral administration, two SOD mimics Mn1 and Mn1C were loaded into lactic acid bacteria that serve as delivery vectors. When orally administrated to mice suffering from a colitis, these chemically modified bacteria (CMB) showed protective effects on the global health status of mice. In addition, they have shown beneficial effects on lipocalin-2 content and intestinal permeability. Interestingly, mRNA SOD2 content in colon homogenates was significantly decreased upon mice feeding with CMB loaded with Mn1C, suggesting that the beneficial effects observed may be due to the release of the SOD mimic in the gut that complement for this enzyme. These CMB represent new efficient chemically modified antioxidant probiotics for IBD treatment.

Methotrexate (MTX) is a well-known anti-metabolite agent recognized for its oxidative effects, particularly in the liver where the enzyme catalase is abundant. This research aimed to clarify the impact of MTX on the behavior of liver catalase. The cytotoxicity of HepG2 cells was assessed across various concentrations of MTX. Following that, the examination focused on the generation of reactive oxygen species (ROS) and the activity of catalase. Furthermore, the kinetic activity of bovine liver catalase (BLC) was examined in the presence of MTX. Finally, the interaction between MTX and the enzyme's protein structure was investigated using docking and dynamic light scattering (DLS) methods. The results indicated a significant decrease in catalase activity and a significant increase in ROS production in HepG2 cells treated with MTX. Although the activity of BLC remained unaffected by MTX directly, molecular docking and DLS techniques revealed MTX binding to BLC, inhibiting its tetramerization. The oxidative effects of MTX were associated with elevated ROS levels in cellular processes, leading to excessive catalase activity and subsequent suicide inactivation. Furthermore, MTX influenced the protein structure of catalase.

The purpose of this study is to investigate FABP3's biological function and potential mechanism in cataract. Treatment of H₂O₂ raised FABP3 expression. H₂O₂ decreased cell viability, enhanced apoptosis, promoted Bax and cleaved caspase-3 expression, inhibited Bcl-2 expression, enhanced the levels of IL-6, IL-1β, and TNF-α, raised MDA level, and decreased SOD and GSH levels in HLE-B3 cells. However, the effects of H₂O₂ on cell viability, apoptosis, inflammatory cytokines, and oxidative stress were reversed by FABP3 knockdown and aggravated by FABP3 overexpression. H₂O₂ increased the levels of lipid hydroperoxides and Fe²⁺, but reduced the expression of GPX4, SLC7A11, and Ferritin protein. Nevertheless, knockdown of FABP3 reversed the changes of lipid hydroperoxides, Fe²⁺, GPX4, SLC7A11, and Ferritin protein, and FABP3 overexpression caused the opposite results. In addition, the inhibition of FABP3 knockdown on cell apoptosis, inflammation, and oxidative stress was reversed by ferroptosis inducer (erastin), and the promotion of FABP3 overexpression on cell apoptosis, inflammation, and oxidative stress was reversed by ferroptosis inhibitor (Fer-1). Taken together, knockdown of FABP3 in lens epithelial cells treated with H₂O₂ restrained apoptosis, inflammation, and oxidative stress through regulating ferroptosis, suggesting that FABP3 might be a potential target for cataract treatment.

Background: Acute myocardial infarction (AMI) is a deadly cardiovascular disease with no effective solution except for percutaneous coronary intervention and coronary artery bypass grafting. Inflammation and apoptosis of the injured myocardium after revascularization seriously affect the prognosis. Hydrogen possesses anti-inflammatory, anti-oxidative, and anti-apoptotic effects and may become a new treatment for AMI. This study explored the specific mechanism by which hydrogen operates during AMI treatment.

Methods: Thirty Sprague-Dawley rats were randomly divided into three groups: control, myocardial infarction (MI), and myocardial infarction + hydrogen (MI+H₂), each containing 10 rats. The MI rat model was established by ligation of the left anterior descending branch. The MI+H₂ group received 2% hydrogen inhalation treatment for 3 h/Bid.

Results: Myocardial infarct size was evaluated using triphenyl tetrazolium chloride staining. Transmission electron microscopy showed reduced mitochondrial damage compared with the MI group. JC-1 staining, which indicates mitochondrial membrane potential, showed a low red/green fluorescence intensity ratio in the MI group compared to that in the control group, indicating mitochondrial membrane potential loss. After hydrogen inhalation, this ratio increased, suggesting partial recovery of membrane potential. In addition, mitochondrial ATP content, mitochondrial complex I, and mitochondrial complex III activity were significantly decreased in the MI group, which was improved after hydrogen administration. Western blotting analysis showed decreased Cyt-c protein levels in the myocardial mitochondria and increased levels in the cytoplasm of MI rats. Following hydrogen inhalation, the levels of ROS, 8-OHdG, and MDA that could represent oxidative stress injury significantly decreased. Besides, the expression of Cyt-C, Bax, cleaved-caspase-9, and cleaved-caspase-3 in MI group significantly increased, while the Bcl-2, TRX2, SOD2 expression decreased. The expression of these proteins in MI+H2 group was improved compared with the MI group.

Conclusion: Overall, hydrogen inhalation reduces myocardial infarct size, improves mitochondrial dysfunction, and modulates the levels of apoptosis-related substances. Importantly, Hydrogen reduces acute myocardial infarction damage by downregulating ROS and upregulating antioxidant proteins.

Molnupiravir is a prodrug of the antiviral ribonucleoside analogue N⁴-hydroxycytidine (NHC), for use in the treatment of coronavirus disease 2019 (COVID-19). However, it is generally considered that NHC-triphosphate is incorporated into the host genome to induce mutations. In our previous preliminary report, we proposed oxidative DNA damage by NHC via cytidine deaminase (CDA)-mediated ROS formation. In the present study, we investigated cell viability using the HL-60 human leukemia cell line and its H₂O₂-resistant clone, HP100 cells. The survival rate was significantly reduced in HL-60 cells treated with NHC, but not in HP100 cells. LC-MS analysis revealed that uridine formation occurred from CDA-treated NHC, suggesting that CDA metabolizes NHC to uridine and hydroxylamine. We clarified mechanisms of CDA-mediated reactive oxygen species (ROS) generation and DNA damage by NHC using isolated DNA. CDA-treated NHC induced DNA damage in the presence of Cu(II). The DNA damage was enhanced by NADH addition and piperidine treatment. CDA-treated NHC and Cu(II) caused piperidine-labile sites at thymine, cytosine, and guanine, and the DNA cleavage pattern was similar to that of hydroxylamine. Catalase and bathocuproine inhibited the DNA damage, indicating the involvement of H₂O₂ and Cu(I). An indicator of oxidative DNA damage, 8-oxo-7,8-dihydro-2'-deoxyguanosine formation by CDA-treated NHC, was lower under hypoxic conditions than under normal conditions. Therefore, hydroxylamine, possibly produced from NHC treated with CDA, could induce metal-dependent H₂O₂ generation during the redox reactions, suggesting that oxidative DNA damage induced by ROS plays an important role in molnupiravir-related cytotoxicity and mutagenicity.

Elevated levels of the enzyme GPX4 have been detected in tumor tissues, which may play a role in cancer progression. We did a meta-analysis of eight studies encompassing 1180 individuals to evaluate the importance of GPX4 in cancer, particularly in terms of prognosis and clinicopathological characteristics. Research results indicate that higher levels of GPX4 were linked to worse overall survival (OS) (HR = 1.47 [95%CI = 1.18-1.76], p < .001). Elevated levels of GPX4 were linked to lymph node invasion (OR.69 [95% CI.44-1.10], p =.12), metastasis (OR 1.58 [95% CI.97-2.55], p =.06, p <.0001), and advanced clinical stage III-IV (OR.82 [95% CI.70-.96], p =.001). A sensitivity study revealed that the general findings were constant across all levels of impact intensity. The findings of this meta-analysis suggest that increased GPX4 levels are not only correlated with reduced overall survival rates for patients with tumors but it also offers valuable insights regarding the clinical traits of tumor malignancy and metastasis. Based on these connections, GPX4 has the potential to serve as a biomarker for tumor detection, prognosis, and targeted therapy.

Skeletal muscle satellite cells (SMSCs) are pivotal for skeletal muscle regeneration post-injury, and their development is intricately influenced by regulatory factors. Selenoprotein K (SELENOK), an endoplasmic reticulum resident selenoprotein, is known for its crucial role in maintaining skeletal muscle redox sensing. However, the specific molecular mechanism of SELENOK in SMSCs remains unclear. In this study, a SELENOK knockdown model was established to delve into its role in SMSCs. The results revealed that SELENOK knockdown hindered SMSCs proliferation and differentiation, as evidenced by the regulation of key proteins such as Pax7, Myf5, CyclinD1, MyoD, and Myf6, and the inhibitory effects were mitigated by N-Acetyl-l-cysteine (NAC). SELENOK knockdown induced oxidative stress, further analyses uncovered that SELENOK knockdown downregulated nuclear transcription factor nuclear erythroid factor 2-like 2 (Nrf2) protein expression while upregulating cytoplasmic kelch-like ECH-associated protein 1 (Keap1) protein expression. SELENOK knockdown impeded Nestin and sequestosome 1/p62 (p62) interaction with Keap1, leading to increased Nrf2 ubiquitination. This prevented Nrf2 transportation from cytoplasm to nucleus mediated by Keap1, ultimately resulting in the downregulation of catalase (CAT), heme oxygenase-1 (HO-1), and glutathione peroxidase 4 (GPX4) protein expression. Notably, SELENOK knockdown-induced inhibition of SMSCs proliferation and differentiation was alleviated by Oltipraz, an activator of the Nrf2 pathway. This study provided novel insights, demonstrating that SELENOK is a key player in SMSCs proliferation and differentiation by influencing the Nrf2 antioxidant signaling pathway.

Parkinson's disease (PD) is a neurodegenerative disorder characterized by the loss of dopaminergic neurons in the substantia nigra. Recently, disorders in metabolism of metals, including copper (Cu) and iron (Fe), have been reported to be linked to the pathogenesis of PD. We previously demonstrated that 6-hydoroxydopamine (6-OHDA), a neurotoxin used for the production of PD model animals, decreases Atox1, a Cu chaperone, and ATP7A, a Cu transporter, and disrupts intracellular Cu metabolism in human neuroblastoma SH-SY5Y cells. However, the exact mechanisms remain unclear. Meanwhile, intracellular Fe modulates 6-OHDA-induced cellular responses. In this study, we investigated whether Fe participates in 6-OHDA-induced abnormality in Cu metabolism. 6-OHDA-induced reactive oxygen species (ROS) production and cellular injury were suppressed by Fe chelators, deferoxamine and 2,2'-bipyridyl (BIP). These chelators also restored 6-OHDA-induced degradation of Atox1 and ATP7A proteins and subsequent Cu accumulation, indicating that intracellular Fe is involved in the disruption of Cu homeostasis associated with 6-OHDA. Atox1 has redox-sensitive cysteine (Cys) residues in its Cu-binding site. The Cys residues of Atox1 were oxidized by 6-OHDA, and BIP suppressed their oxidation. Moreover, the replacement of Cys with histidine in the Cu-binding site conferred resistance to 6-OHDA-induced Atox1 degradation. These results suggest that oxidized modification of Atox1 by 6-OHDA is likely to accelerate its degradation. Thus, we conclude that Fe and Cu metabolisms are closely related to each other in the pathogenesis of PD.

The concept of dual-state hyper-energy metabolism characterized by elevated glycolysis and OxPhos has gained considerable attention during tumor growth and metastasis in different malignancies. However, it is largely unknown how such metabolic phenotypes influence the radiation response in aggressive cancers. Therefore, the present study aimed to investigate the impact of hyper-energy metabolism (increased glycolysis and OxPhos) on the radiation response of a human glioma cell line. Modulation of the mitochondrial electron transport chain was carried out using a 2,4-dinitrophenol (DNP). Metabolic characterization was carried out by assessing glucose uptake, lactate production, mitochondrial mass, membrane potential, and ATP production. The radiation response was examined by cell growth, clonogenic survival, and cell death assays. Macromolecular oxidation was assessed by DNA damage, lipid peroxidation, and protein carbonylation assay. Hypermetabolic OPM-BMG cells exhibited a significant increase in glycolysis and OxPhos following irradiation as compared to the parental BMG-1 cells. Enhanced radioresistance of OPM-BMG cells was evidenced by the increase in α/β ratio (9.58) and D1 dose (4.18 Gy) as compared to 4.36 and 2.19 Gy in BMG-1 cells respectively. Moreover, OPM-BMG cells were found to exhibit increased resistance against radiation-induced cell death, and macromolecular oxidation as compared to BMG-1 cells. Inhibition of glycolysis and mitochondrial complex-II significantly enhanced the radiosensitivity of OPM-BMG cells compared to BMG-1 cells. Our results demonstrate that the hyper-energy metabolism of increased glycolysis and OxPhos confer radioresistance. Consequently targeting glycolysis and OxPhos in combination with radiation may overcome therapeutic resistance in aggressive cancers like glioma.