ROS Levels Research Articles

Prenatal exposure to di-n-butyl phthalate promotes RIPK1-regulated necroptosis of uroepithelial cells and induces hypospadias through the epithelial-mesenchymal transition process in newborn male rats

Maternal exposure to di-n-butyl phthalate (DBP) has been linked to the induction of hypospadias; however, the underlying mechanism remains unclear. Necroptosis is reported to be implicated in developmental malformations. This study aimed to investigate the underlying mechanism of necroptosis in the development of hypospadias. DBP was dissolved in corn oil, and pregnant rats were administered a precisely measured dose of DBP (750mg/kg/day) via gastric intubation from gestation day 14 to 18. Control rats received only corn oil. The day of birth was considered postnatal day (PND) 1. Male hypospadias rats were identified on PND 7. Genital tubercle tissues were collected and stored at −80°C for subsequent PCR analysis, cryopreserved in liquid nitrogen for western blot, or fixed in formalin for immunohistochemistry (IHC) staining. IHC staining and western blot analysis revealed increased expression of RIPK1 and necroptosis markers in genital tubercle (GT) tissue compared to the control group. Additionally, higher levels of EMT and impaired androgen receptor expression were observed in GT tissue. Exposure to increased DBP concentrations in rat primary uroepithelial cells (PUCs) led to elevated ROS production. Necroptosis markers and EMT expression levels were upregulated in PUCs following DBP incubation. Notably, treatment with DBP combined with necrostatin-1, a necroptosis inhibitor, reduced the expression of EMT markers and ROS production compared to DBP treatment alone. In vitro studies further revealed that DBP-induced necroptosis promoted the degradation of E-cadherin through the ubiquitin-proteasome pathway in PUCs. Our findings suggest that maternal exposure to DBP promotes necroptosis in uroepithelial cells by elevating ROS level and EMT status. Thus, necroptosis may play an essential role in the development of hypospadias.

Neurotoxic and developmental effects of scented incense stick Smoke: Network toxicology and zebrafish model study

Burning incense sticks is a traditional practice in many cultures, especially in Southeast Asia. While it is often regarded as sacred and beneficial, modern incense sticks contain various chemicals that can pose health risks. A GCMS analysis of the ICS revealed potential compounds. Network toxicology revealed that ICS contains compounds violating Lipinski's rule of five, leading to potential neurotoxic effects. Key pathways affected include neuroactive ligand-receptor interaction and calcium signaling, associated with neurodegenerative diseases like Parkinson's and Alzheimer's. Significant genes involved are STAT3, BCL2, and MTOR, emphasizing the chemical ahzardss of ICS exposure. We investigated the toxicity of ICS using zebrafish (Danio rerio) embryos as a mode. ICS exposure resulted in a dose-dependent increase in toxicity. High concentrations (7 and 14µg/ml) led to immediate mortality, while lower concentrations (0.1, 0.3, 0.5, and 1µg/ml) caused developmental defects such as yolk sac edema, skeletal malformations, and pericardial edema. Mortality rates increased with higher concentrations, confirming dose-dependen ICS exposure caused hypoactive locomotion, with reduced distance traveled and velocityt toxicity. Higher concentrations of ICS led to increased ROS levels and cellular damage, as evidenced by enhanced staining levels. A dose-dependent increase in lipid peroxidation (DPPP assay) and lipid accumulation (Nile red assay) was observed. Higher ICS concentrations led to significant oxidative damage to lipids and increased lipid deposition. Enzymatic assays showed that ICS exposure significantly decreased the activities of antioxidant enzymes SOD and CAT, indicating impaired antioxidant defense, while increasing LDH activity, signaling tissue damage and cytotoxicity. Gene expression analysis revealed downregulation of SOD1 and CAT genes, upregulation of inflammatory genes TNF-α and IL-1β, and increased expression of the apoptotic gene p53 with decreased expression of Bcl-2 and BDNF. These findings highlight ICS's potential to cause oxidative stress, inflammation, apoptosis, and neurodevelopmental impairments.

Impact of Indole Inclusion in the Design of Multi-Tactical Metal-Binding Tetra-Aza Macrocycles that Target the Molecular Features of Neurodegeneration.

Numerous small molecules have been studied for their ability to counteract oxidative stress, a key contributor to neurodegenerative diseases such as Alzheimer's. Despite these efforts, the pharmacological properties and structure-activity relationships of these compounds remain insufficiently understood, yet they are critical in evaluating a drug molecule's therapeutic potential. A modified tetra-aza macrocycle has demonstrated strong antioxidant activity through various mechanisms; however, its limited permeability presents challenges for advanced formulation studies. To enhance permeability while preserving the beneficial reactivity of the parent molecule, two synthetic modifications involving indole functionality were explored and compared to modifications using methyl groups alone. New synthetic strategies were developed to produce the indole-containing molecules, which were characterized by 1D/2D NMR techniques. Isoelectric points, metal binding, and radical scavenging activity were determined to validate that the reactivity of the parent molecules was retained. The permeability of all molecules explored was improved. Protection against oxidative stress through activation of the Nrf2 pathway was demonstrated for molecules containing indoles in cellular models by measuring ROS levels upon treatment and mRNA levels of HO-1 and Nrf2. In contrast, no protection or Nrf2 activation was observed with the methylation of the O- or N atom. These results suggest that while alkylation improves permeability overall, concomitant antioxidant protection and positive permeability are achieved with the indole congeners alone.

O-GlcNAcylation promotes malignancy and cisplatin resistance of lung cancer by stabilising NRF2.

The transcription factor NRF2 plays a significant role in regulating genes that protect cells from oxidative damage. O-GlcNAc modification, a type of posttranslational modification, is crucial for cellular response to stress. Although the involvement of both NRF2 and O-GlcNAc in maintaining cellular redox balance and promoting cancer malignancy has been demonstrated, the potential mechanisms remain elusive. The immunoblotting, luciferase reporter, ROS assay, co-immunoprecipitation, and immunofluorescence was used to detect the effects of global cellular O-GlcNAcylation on NRF2. Mass spectrometry was utilised to map the O-GlcNAcylation sites on NRF2, which was validated by site-specific mutagenesis and O-GlcNAc enzymatic labelling. Human lung cancer samples were employed to verify the association between O-GlcNAc and NRF2. Subsequently, the impact of NRF2 O-GlcNAcylation in lung cancer malignancy and cisplatin resistance were evaluated in vitro and in vivo. NRF2 is O-GlcNAcylated at Ser103 residue, which hinders its binding to KEAP1 and thus enhances its stability, nuclear localisation, and transcription activity. Oxidative stress and cisplatin can elevate the phosphorylation of OGT at Thr444 through the activation of AMPK kinase, leading to enhanced binding of OGT to NRF2 and subsequent elevation of NRF2 O-GlcNAcylation. Both in cellular and xenograft mouse models, O-GlcNAcylation of NRF2 at Ser103 promotes the malignancy of lung cancer. In human lung cancer tissue samples, there was a significant increase in global O-GlcNAcylation, and elevated levels of NRF2 and its O-GlcNAcylation compared to paired adjacent normal tissues. Chemotherapy promotes NRF2 O-GlcNAcylation, which in turn decreases cellular ROS levels and drives lung cancer cell survival. Our findings indicate that OGT O-GlcNAcylates NRF2 at Ser103, and this modification plays a role in cellular antioxidant, lung cancer malignancy, and cisplatin resistance.

Extreme heat event influences the toxic impacts of nano-TiO2 with different crystal structures in mussel Mytilus coruscus

The wide use of nano‑titanium dioxide (nano-TiO2) and its ubiquitous emission into aquatic environments are threatening environmental health. Ambient temperature can affect the aggregation state of nano-TiO2 in seawater, thus influencing the intake and physiological effects on marine species. We studied the physiological effects of mixed nano-TiO2 (a mixture of anatase crystal and rutile crystal with an average particle size of 25 nm, P25) on mussels. Subsequently, we investigated the oxidative stress, immunotoxicity, neurotoxicity, and detoxification in Mytilus coruscus exposed to two different crystal structures of nano-TiO2 (anatase and rutile) at 100 μg/L concentration under marine heatwaves (MHWs, 28 °C). MHWs and nano-TiO2 exposure induced neurotoxicity and immune damage and caused dysregulation of redox balance in the gills. Moreover, MHWs exposure disturbed the glutathione system and detoxification function of mussels, resulting in enhanced toxicity of nano-TiO2 after co-exposure. Anatase exposure significantly impaired the antioxidative system and downregulated the relative expression of antioxidant-related genes (Nrf2 and Bcl-2), HSP-90, and immune parameters under MHWs, while producing higher ROS levels compared to rutile. Based on integrated biomarker response (IBR), anatase combined with MHWs group showed the highest value (19.29). However, there was no significant difference in bioaccumulation of titanium between anatase (6.07 ± 0.47 μg/g) and rutile (5.3 ± 0.44 μg/g) exposures under MHWs. These results indicate that MHWs would elevate the potential hazard of nanoparticles to marine organisms.

Lycium Barbarum Polysaccharide-Derived Nanoparticles Protect Visual Function by Inhibiting RGC Ferroptosis and Microglial Activation in Retinal Ischemia‒Reperfusion Mice.

Retinal ischemia‒reperfusion (IR) is a major contributor to vision impairment and irreversible vision loss due to retinal ganglion cell (RGC) injury or loss. Contemporary therapeutic approaches predominantly focus on the amelioration of symptoms rather than addressing the fundamental etiological factors. Oxidative stress is a notable feature and an important mediator of IR damage. Lycium barbarum polysaccharide (LBP), the main active ingredient of Lycium barbarum, has various pharmacological effects, including antioxidation, immunoregulation, and neuroprotective effects. In this study, the ROS-consumable moiety phenylboronic acid pinacol ester (PBA) is introduced to LBP molecules, which can self-assemble into nanoparticles in aqueous solution. This nanoparticle (termed PLBP) can reduce the cellular ROS levels and enhance the antioxidant capability of RGCs by activating the NRF2 pathway, thus protecting RGCs from ferroptosis and preserving visual function in response to IR injury. PLBP also reduces neuroinflammation by inhibiting the ability of microglia to phagocytose, migrate, secrete inflammatory cytokines, and activate the NF-κB pathway. In conclusion, this approach can be used as an inspiration for the future development of neuroprotective drugs.

Leech Granules Inhibit Ferroptosis and Alleviate Renal Injury in Mice with Diabetic Kidney Disease

Ethnopharmacological relevanceThe underlying mechanisms of diabetic kidney disease (DKD) and effective treatment strategies remain unclear. DKD progression is closely associated with abnormal iron metabolism and ferroptosis in vivo. Leeches, used in traditional Chinese medicine for promoting blood circulation and resolving blood stasis, are utilized to treat diabetes and its associated complications. Leeches effectively antagonize oxidative stress injury and exert protective effects on renal function. However, whether leeches can inhibit ferroptosis by modulating oxidative stress and iron metabolism remains unclear. Aim of the studyTo investigate the therapeutic potential of leech granules in DKD and, specifically, their effects on ferroptosis. Materials and methodsWe used a mouse model of DKD. The mice were treated with leech granules via gavage. Component identification and analysis of leech granules were performed using UPLC-MS, and efficacy was assessed by histopathology and analysis of blood glucose, lipids, and renal function. Additionally, the pharmacological mechanisms of leech granules were explored via proteomics, immunohistochemical staining, western blotting, and cell culture. ResultsProteomic analysis showed that iron metabolism was dysregulated and ferroptosis increased in DKD mice. Leech granules significantly reduced iron accumulation and renal pathological damage, decreased ROS levels, upregulated GSH levels, and inhibited ferroptosis in the kidneys of DKD mice. Furthermore, in vitro cellular experiments demonstrated that leech granules could inhibit erastin-induced ferroptosis and protect renal cells. ConclusionsThe regulation of renal iron metabolism and inhibition of ferroptosis mediates the therapeutic effect of leech granules on DKD. Leech granules represent a promising approach for DKD treatment.

Structural characterization, antioxidant activities and physicochemical properties analysis of a galactose-rich exopolysaccharide produced by Limosilactobacillus fermentum YL-11

Lactobacilli-derived exopolysaccharides (EPSs) are widely regarded as safe and have been employed as potential natural, non-toxic additives and functional agents in food industry. The structural properties of a galactose-rich exopolysaccharide produced by Limosilactobacillus fermentum YL-11 (YL-11 EPS) were investigated with chemical and instrumental analysis. The YL-11 EPS was determined to have a molecular weight (Mw) of 13.2 kDa. Nine different types of glycosidic bonds were identified in YL-11 EPS. The inferred structure of the YL-11 EPS contained the backbone of →2)-β-D-Galp-(1→[3)-β-D-Galp-(1]4 → 3)-β-D-Galp-(1 → 3)-β-D-Galp-(1→, with two side chains, β-D-Galp-(1 → 6)-α-D-Manp-(1 → 6)-[α-D-Glcp-1→2]-α-D-Manp-(1 → 3)-[α-D-Glcp-1→6]-α-D-Glcp-(1 → 3)-[α-D-Glcp-1→6]-α-D-Glcp-(1→ with an acetyl group at the O-3 position of →2,6-α-D-Manp-1→ and α-L-Araf-(1→, which were present at the C6 location of the 1,3-β-D-Galp. The YL-11 EPS showed concentration-dependent antioxidant activities and the DPPH, ABTS, hydroxyl groups and superoxide radicals scavenging rates of 10 mg/mL YL-11 EPS reached 61%, 74%, 35.4% and 40%, respectively. The YL-11 EPS also has a beneficial protective impact on H2O2-injured cells, which pretreatment with YL-11 EPS can substantially decrease ROS levels and enhance SOD enzyme activity. Additionally, the YL-11 EPS exhibited good thickening properties and thermal stability. The EPS's structural information and physicochemical properties provide a basis for studying their structure-function relationships.

Hypoxia-inducible factor-1α promotes ferroptosis by inducing ferritinophagy and promoting lactate production in yak longissimus thoracis et lumborum postmortem

Ferroptosis has emerged as a novel, crucial regulator of meat quality in the postmortem hypoxia environment, with its role in mediating protein oxidation and cell death. However, the interaction between ferroptosis and the hypoxia response, especially the involvement of hypoxia-inducible factor-1α (HIF-1α), remains poorly studied. This study aimed to characterize whether HIF-1α influences ferroptosis, and, if so, explore the underlying mechanisms involved. The results showed that ferroptosis mediated by HIF-1α negatively impacts meat color and water holding capacity (WHC) but improving tenderness. Inhibition of HIF-1α by 3-(5′-hydroxymethyl-2′-furyl)-1-benzyl indazole (YC-1) reduced ferroptosis, as evidenced by lower lipid ROS levels, malondialdehyde (MDA), along with higher glutathione (GSH) levels compared to the control (P < 0.05). Additionally, inhibition of HIF-1α shifted iron homeostasis towards decreased uptake via downregulation of transferrin receptor 1 (TfR1) and induced export/storage via upregulation of ferroportin (FPN) and ferritin heavy chain (FTH) (P < 0.05). The relative expression of the ferritinophagy mediator nuclear receptor coactivator 4 (NCOA4), LC3-II/LC3-I ratio, and ATG were inhibited by YC-1 (P < 0.05), these findings suggest a general decrease in ferritinophagy associated with HIF-1α inhibition. YC-1-treated samples exhibited significantly diminished lactate accumulation and lactate dehydrogenase (LDH) activity compared to the control (P < 0.05). Unexpectedly, the inhibition of ferroptosis caused by YC-1 was further amplified by lactate enhancement, suggesting that lactate can exert its suppressive effects on ferroptosis independently of HIF-1α. Collectively, these findings demonstrate that HIF-1α drives ferroptosis by regulating iron metabolism, while lactate inhibits ferroptosis in a HIF-1α-independent manner. Overall, the HIF-1α mediated ferroptosis of postmortem yak muscle had a negative impact on WHC and color, while as a contributing factor of tenderness.

Low fluid shear stress stimulates the uptake of noxious endothelial extracellular vesicles via MCAM and PECAM-1 cell adhesion molecules.

Atherosclerotic lesions mainly form in arterial areas exposed to low shear stress (LSS), where endothelial cells express a senescent and inflammatory phenotype. Conversely, areas exposed to high shear stress (HSS) are protected from plaque development. Endothelial extracellular vesicles (EVs) have been shown to regulate inflammation and senescence, and therefore play a crucial role in vascular homeostasis. Whilst previous studies have shown links between hemodynamic forces and EV release, the effects of shear stress on the release and uptake of endothelial EVs remains elusive. We aim to decipher the interplay between these processes in endothelial cells exposed to atheroprone or atheroprotective shear stress. Confluent HUVECs were exposed to LSS or HSS for 24h. Large and small EVs were isolated from conditioned medium by centrifugation and size exclusion chromatography. They were characterised by TEM, Western blot, tunable resistive pulse sensing, flow cytometry and proteomics. Uptake experiments were performed using fluorescently-labelled EVs and differences between groups were assessed by flow cytometry and confocal microscopy. We found that levels of large and small EVs in conditioned media were fifty and five times higher in HSS than in LSS conditions, respectively. In vivo and in vitro uptake experiments revealed greater EV incorporation by cells exposed to LSS conditions. Additionally, endothelial LSS-EVs have a greater affinity for HUVECs than HSS-EVs or EVs derived from platelets, erythrocytes and leukocytes. Proteomic analysis revealed that LSS-EVs were enriched in adhesion proteins (PECAM1, MCAM), participating in EV uptake by endothelial cells. LSS-EVs also carried mitochondrial material, which may be implicated in elevating ROS levels in recipient cells. These findings suggest that shear stress influences EV biogenesis and uptake. Given the major role of EVs and shear stress in vascular health, deciphering the relation between these processes may yield innovative strategies for the early detection and treatment of endothelial dysfunction.

Edible thermosensitive chitosan/hydroxypropyl β-cyclodextrin hydrogel with natural licoricidin for enhancing oral health: Biofilm disruption and demineralization prevention

Dental caries, a widespread and significantly detrimental health condition, is characterized by demineralization, pain, compromised tooth functionality, and various other adverse effects. Licoricidin (LC), a natural isoflavonoid, demonstrates potent antimicrobial properties for maintaining oral health. However, its practical application is significantly hindered by its limited water solubility and susceptibility to removal within the oral environment. To tackle this issue, we developed a delivery oral system by an edible thermosensitive chitosan- disodium beta-glycerol phosphate pentahydrate (CS/β-GP) hydrogel to load LC/Hydroxypropyl beta-cyclodextrin (HP-β-CD) inclusion complexes. These hydrogels (LC/HP-β-CD/CS/β-GP) could solidify rapidly at oral temperature and sustainably release LC, thereby preventing its rapid clearance from the oral cavity. We confirmed the significant antibacterial activity of this hydrogel against Streptococcus mutans and Staphylococcus aureus. Additionally, the HP-β-CD combination enhanced LC to penetrate bacterial biofilms and inhibit biofilm growth, leading to leakage of cellular proteins and DNA. Additionally, we studied the effect of LC/HP-β-CD/CS/β-GP on intracellular ROS levels and MMP, comprehensively exploring its antimicrobial mechanism. Furthermore, LC/HP-β-CD/CS/β-GP exhibited the ability to inhibit demineralization and demonstrated excellent biocompatibility. In summary, this study presented a safer approach to oral delivering bioactive substances, offering a promising strategy for enhanced oral health and safety.

MiR-224-5p alleviates preeclampsia-like mouse symptoms by targeting PANX1 to inhibit ferroptosis in trophoblast cells

Preeclampsia (PE) is a high morbidity and lethality disease specific to pregnancy, and insufficient placental trophoblast invasion acts as a crucial factor contributing to PE development. The present study investigated the function and potential mechanism of microRNA (miR)-224-5p within PE. In the study, miR-224-5p expression was reduced within placental tissue samples of the PE mouse model and PE cell model. Restoration of miR-224-5p expression markedly inhibited ROS levels and ferroptosis, lowered blood pressure in pregnant mice, increased the live birth rate, and enhanced trophoblast cell proliferation and invasion as well as suppressed their apoptosis. miR-224-5p could target and suppress PANX1, and overexpression of PANX1 could significantly advance ferroptosis and cause trophoblast dysfunction, a process that might be relieved via restoring miR-224-5p expression. In conclusion, miR-224-5p/PANX1 ameliorates trophoblast dysfunction by inhibiting ferroptosis, which provides a potential new option for clinical treatment of PE.

Inhibition of sulfur assimilation by S-benzyl-L-cysteine: Impacts on growth, photosynthesis, and leaf proteome of maize plants

Sulfur is an essential nutrient for various physiological processes, including protein synthesis and enzyme activation. We aimed to evaluate how S-benzyl-L-cysteine (SBC), an inhibitor of the sulfur assimilation pathway, affects maize plants' growth, photosynthesis, and leaf proteomic profile. Thus, maize plants were grown for 14 days in vermiculite supplemented with SBC. Photosynthesis was assessed using light and CO2 response curves and chlorophyll a fluorescence. Leaf proteome analysis was conducted to evaluate photosynthetic protein biosynthesis, and ROS content was quantified to assess oxidative stress. Applying SBC resulted in a significant decrease in the growth of maize plants. The gas exchange analysis revealed that maize plants exhibited a diminished rate of CO2 assimilation attributable to both stomatal and non-stomatal limitations. Furthermore, SBC suppressed the activity of important elements involved in the photosynthetic electron transport chain (including photosystems I and II, cytochrome b6f, and ATP synthase) and enzymes responsible for the Calvin cycle, some of which have sulfur-containing prosthetic groups. Consequently, the diminished electron flow rate resulted in a substantial increase in the levels of ROS within the leaves. Our research highlights the crucial role of SBC in disrupting maize photosynthesis by limiting L-cysteine and assimilated sulfur availability, which are essential for the synthesis of protein and prosthetic groups and photosynthetic processes, emphasizing the potential of OAS-TL as a new herbicide site of action.

RNA m6A methylation regulatory mechanism of resveratrol in premature senescence cells

AbstractResveratrol, a natural compound found in various plants, is known for its anti‐inflammatory, antioxidant, and senescence‐delaying properties. RNA N6‐methyladenosine (m6A) methylation plays a crucial role in oxidative stress and premature cellular senescence processes and is closely associated with age‐related disorders. However, the anti‐premature senescence via RNA m6A methylation mechanism of resveratrol is still not fully understood. In this study, based on premature senescence model of human embryonic lung fibroblasts (HEFs) induced by hydrogen peroxide (H2O2), a widely accepted model of premature senescence caused by oxidative stress, we explored the anti‐aging regulatory effects of resveratrol at the RNA m6A methylation level. Our data suggested that resveratrol significantly delayed premature senescence by increasing cell viability, reducing SA‐β‐gal blue staining rate, ROS levels, and senescence‐associated secretory phenotypes (SASP) expression in HEFs. Meanwhile, resveratrol increased the whole RNA methyltransferases activity and the overall m6A level during senescence. Furthermore, three genes CCND2, E2F1, and GADD45B have been identified as the main ones regulating premature by resveratrol. Specifically, it decreased E2F1, GADD45B RNA m6A methylation level, and increased CCND2 level in premature cells. Our study provided new clues for exploring the mechanism and application of resveratrol in the field of premature aging.

Protective Effects of Wild Sulla coronaria (Fabaceae) Flowers Phytocomplex in Human Dermal Fibroblasts Stimulated with Interleukin-1β

Sulla coronaria is indigenous to the Mediterranean region. It is grown as fodder in southern Italy because it contains various secondary metabolites with beneficial activities on animals. Recently, its potential use in cosmeceutical treatments for skin problems was reported. In this scenario, to contribute to a possible cosmeceutical application, we characterized the phytochemical profile of Sulla coronaria flowers’ hydroalcoholic extract by HPLC-DAD, Folin-Ciocalteu, Aluminum Chloride methods, DPPH assay, and, for the first time, we evaluated the antioxidant and anti-inflammatory activities on dermal fibroblasts. The phytochemical analysis confirmed the significant content of phenolic compounds (TPC 69.8 ± 0.6 mg GAE/g extract, TFC 15.07 mg CE/g extract) and the remarkable presence of rutin, quercetin, and isorhamnetin derivatives that give to the phytocomplex a good antioxidant activity as highlighted by the DPPH assay (IC50 of 8.04 ± 0.5 µg/mL). Through the reduction in NO• and ROS levels in human dermal fibroblasts, the biological tests demonstrated both the safety of the extract and its ability to counteract the inflammatory state generated by Interleukin-1β exposure. Our findings indicate that the antioxidant activities of the phytocomplex are strictly related to the anti-inflammatory action of the Sulla coronaria flowers extract, confirming that this plant could be a valuable source of bioactive molecules for cosmeceutical and nutraceutical applications.

HECTD2 as a target for veratric acid in the regulation of ferroptosis in renal cell carcinoma

Function of HECTD2 in renal cell carcinoma malignant progression is undefined. Molecular mechanism behind anti-cancer effects of veratric acid (VA) from traditional Chinese medicine (TCM) is underexplored. The Cancer Genome Atlas was leveraged to study HECTD2 expression in renal cell carcinoma and its relationship with histological grading. Kaplan–Meier survival analysis was performed. HECTD2 expression was detected in cancer cells and tissues via qRT-PCR and immunohistochemistry. GPX4 and SLC7A11 expression in tumor samples with high or low HECTD2 expression was examined by immunohistochemistry, cell viability by CCK8, cell proliferation by colony formation assay, lipid ROS and mitochondrial superoxide levels by flow cytometry, Fe2+ and MDA content by assay kits, and GPX4 and SLC7A11 proteins by western blot. SeeSAR software screened TCM small molecule compounds with highest affinity to HECTD2, confirmed with cellular thermal shift assay. VA IC50 was measured by CCK8. Xenograft model was developed and treated with VA. Tumor size and weight were monitored, with immunohistochemistry to detect HECTD2 expression in tumors and assess ferroptosis-related markers. HECTD2 was overexpressed in tumor tissues and cells, which positively correlated with histological grading. HECTD2 depletion inhibited cell vitality and proliferation, raised intracellular lipid ROS, mitochondrial superoxide, Fe2+, and MDA. HECTD2 was a target with highest VA affinity. In vitro and vivo experiments concurred that VA treatment hindered malignancy of renal cell carcinoma and enhanced its susceptibility to ferroptosis. HECTD2 supports ferroptosis resistance in renal cell carcinoma, but VA, through its targeting of HECTD2, initiates ferroptosis, showcasing its anti-cancer efficacy.

PRKN-mediated the ubiquitination of IQGAP3 regulates cell growth, metastasis and ferroptosis in early-onset colorectal cancer.

High IQ motif-containing GTPase activating protein 3 (IQGAP3) expression is considered to be associated with poor prognosis of colorectal cancer (CRC). However, its role in early-onset CRC (EOCRC) progress is unclear. The mRNA and protein levels of IQGAP3 and Parkin (PRKN) were examined by qRT-PCR and western blot. Cell proliferation, apoptosis and metastasis were determined by CCK8 assay, EdU assay, flow cytometry and transwell assay. ROS, MDA, GSH, Fe2+, ACSL4 and SLC7A11 levels were detected to assess cell ferroptosis. The interaction between PRKN and IQGAP3 was assessed by Co-IP assay and ubiquitination assay. Xenograft tumor models were constructed to explore the effect of PRKN and IQGAP3 on the tumorigenesis in vivo. IQGAP3 was upregulated, while PRKN was downregulated in EOCRC tissues and cells. IQGAP3 knockdown inhibited CRC cell proliferation, migration and invasion, while enhanced apoptosis and ferroptosis. PRKN ubiquitinated IQGAP3 to promote its degradation. PRKN overexpression suppressed CRC cell growth, metastasis and promoted ferroptosis, while these effects were reversed by upregulating IQGAP3. In animal study, upregulation of PRKN reduced CRC tumorigenesis by decreasing IQGAP3 expression in vivo. IQGAP3, ubiquitinated by PRKN, promoted EOCRC progression by enhancing cell proliferation, metastasis, repressing apoptosis and ferroptosis, which provided a novel target for EOCRC treatment.

Epigenetic mechanism of EZH2-mediated histone methylation modification in regulating ferroptosis of alveolar epithelial cells in sepsis-induced acute lung injury.

Sepsis-induced acute lung injury (SI-ALI) leads to significant deaths in critically ill patients worldwide. This study explores the mechanism of EZH2 regulating ferroptosis of alveolar epithelial cells (AECs) in SI-ALI. In vitro cell model and in vivo mouse lung injury model of sepsis were established. EZH2 expression in lung tissues was intervened by sh-EZH2, followed by H&E staining observation of lung tissue pathological changes. EZH2, H3K27me3, USP10, GPX4, and ACSL4 expressions were determined by qRT-PCR or Western blot. ROS, GSH, and iron ion levels were detected using fluorescent labeling and reagent kits, respectively. ChIP analyzed the enrichment of EZH2 and H3K27me3 on USP10 promoter. The binding between USP10 and GPX4, and the ubiquitination level of GPX4 were detected using Co-IP. EZH2 was highly expressed in lung tissues of SI-ALI mice. EZH2 silencing alleviated ALI and ferroptosis of AECs; EZH2 increased the H3K27me3 level on USP10 promoter through histone methylation. USP10 stabilized GPX4 protein expression through ubiquitination; inhibition of USP10 partially reversed the inhibitory effect of EZH2 silencing on ferroptosis of AECs. In conclusion, EZH2 depresses USP10 expression by promoting histone H3K27me3 modification on USP10 promoter, thereby enhancing ubiquitination degradation of GPX4 and ultimately facilitating ferroptosis of AECs in sepsis.

CDKN1A promotes Cis-induced AKI by inducing cytoplasmic ROS production and ferroptosis

Background and objectiveThis study focuses on investigating the role of CDKN1A in cisplatin-induced AKI (acute kidney injury, AKI) and its potential as a biomarker for early diagnosis and therapeutic intervention by integrating bioinformatics analysis, machine learning, and experimental validation. MethodsWe analyzed the GSE85957 dataset to find genes that changed between control and cisplatin-treated rats. Using bioinformatics and machine learning, we found 13 important genes related to ferroptosis and the P53 pathway. The key gene, CDKN1A, was identified using various algorithms. We then tested how reducing CDKN1A in human kidney cells affected cell health, ROS, and iron levels. We also checked how CDKN1A changes the levels of proteins linked to ferroptosis using Q-PCR and Western Blot. ResultsCDKN1A was found to negatively regulate the G1/S phase transition and was associated with ferroptosis in p53 signaling. Experiments in human renal tubular epithelial cells (HK-2) and rat NRK-52E cells showed that CDKN1A knockdown mitigated cisplatin-induced cell injury by reducing oxidative stress and ferroptosis. ConclusionOur integrated approach identified CDKN1A as a biomarker for cisplatin-induced AKI. Its regulation could be key in AKI pathogenesis, offering new therapeutic insights and aiding in early diagnosis and intervention.

Self-cascade ROS-trapping bioreaction system reverses stem cell oxidative stress fate for osteogenesis

eactive oxygen species (ROS) scavenging is essential for periodontal regeneration. However, the dynamic change of the applied materials within the ROS-rich environment and the residual oxidation products in the host highly impact periodontal regeneration. This study successfully constructs a bioreaction system via thiol-ene click chemistry, leveraging the high affinity of glutathione (GSH) for ROS to attract excess ROS to the crosslinking points. Two minutes after hydrogen peroxide (H2O2) treatment, the ROS level in the G8-0 hydrogel acutely decreases, reaching a 4.4% reduction within 10minutes, confirming the ROS-trapping efficacy. Through a ‘bait switch-on’ mechanism, hexagonal boron nitride (hBN) takes over the captured ROS and the oxidation products of pectin further drive the reduction reaction, ultimately restoring the extracellular environment. The self-cascade products, oxidized hBN, reshape the intracellular oxidative stress (OS) environment, achieving a synergistic extra- and intra-cellular treatment. The significantly high reduced to oxidized glutathione (GSH/GSSG) ratio in G8-10 hydrogel (~80%) illustrates a reversal of oxidative stress in bone marrow stem cells (BMSCs). On a molecular level, the bioreaction system inhibits the NF-κB pathway, promoting the expression of key antioxidant genes (Nqo1 and Nrf2) and osteogenic molecules (ALP and OCN), thereby reversing the detrimental effects of OS on BMSCs. In vivo application demonstrated the system’s strong redox-balancing and osteogenic capabilities in the periodontal inflammation environment. This novel antioxidant bioreaction system, characterized by self-cascade ROS-trapping and product utilization, offers innovative treatment strategies for tissue regeneration under conditions of excessive OS.