Low Levels Of Reactive Oxygen Species Research Articles

Panax quinquefolius saponins and panax notoginseng saponins attenuate myocardial hypoxia-reoxygenation injury by reducing excessive mitophagy.

Panax quinquefolius saponins (PQS) and Panax notoginseng saponins (PNS) are key bioactive compounds in Panax quinquefolius L. and Panax notoginseng, commonly used in the treatment of clinical ischemic heart disease. However, their potential in mitigating myocardial ischemia-reperfusion injury remains uncertain. This study aims to evaluate the protective effects of combined PQS and PNS administration in myocardial hypoxia/reoxygenation (H/R) injury and explore the underlying mechanisms. To investigate the involvement of HIF-1α/BNIP3 mitophagy pathway in the myocardial protection conferred by PNS and PQS, we employed small interfering BNIP3 (siBNIP3) to silence key proteins of the pathway. H9C2 cells were categorized into four groups: control, H/R, H/R + PQS + PNS, and H/R + PQS + PNS+siBNIP3. Cell viability was assessed by Cell Counting Kit-8, apoptosis rates determined via flow cytometry, mitochondrial membrane potential assessed with the JC-1 fluorescent probes, intracellular reactive oxygen species detected with 2',7'-dichlorodihydrofluorescein diacetate, mitochondrial superoxide production quantified with MitoSOX Red, and autophagic flux monitored with mRFP-GFP-LC3 adenoviral vectors. Autophagosomes and their ultrastructure were visualized through transmission electron microscopy. Moreover, mRNA and protein levels were analyzed via real-time PCR and Western blotting. PQS + PNS administration significantly increased cell viability, reduced apoptosis, lowered reactive oxygen species levels and mitochondrial superoxide production, mitigated mitochondrial dysfunction, and induced autophagic flux. Notably, siBNIP3 intervention did not counteract the cardioprotective effect of PQS + PNS. The PQS + PNS group showed downregulated mRNA expression of HIF-1α and BNIP3, along with reduced HIF-1α protein expression compared to the H/R group. PQS + PNS protects against myocardial H/R injury, potentially by downregulating mitophagy through the HIF-1α/BNIP3 pathway.

Genotyping of rams based on melatonin receptor 1A gene polymorphisms: a tool in sire selection?

Context Several polymorphisms in the melatonin receptor 1A gene (MTNR1A ) have been related to reproductive performance in ovine. Aims To investigate the effect of the Rsa I and Mnl I polymorphisms on ram seminal quality. Methods Eighteen Rasa Aragonesa rams were genotyped for the Rsa I (C/C, C/T, T/T) and Mnl I (G/G, G/A, A/A) allelic variants of the MTNR1A gene. Individual ejaculates were analysed once a month throughout the whole year. Sperm motility, morphology, membrane integrity, levels of reactive oxygen species (ROS), phosphatidylserine (PS) inversion, DNA fragmentation and capacitation status were assessed. The effect of the season and polymorphisms on seminal quality was evaluated by mixed ANOVA. Key results Both polymorphisms had an effect on membrane integrity and viable spermatozoa with low levels of ROS and without PS translocation, and Rsa I also on motile and DNA-intact spermatozoa. An interaction between both polymorphisms was found, pointing to a negative effect on seminal quality of carrying the T or A allele in homozygosity. Differences were higher in the reproductive than in the non-reproductive season. Conclusions Mutations substituting C by T and G by A at Rsa I and Mnl I polymorphic sites, respectively, in the MTNR1A gene in rams could decrease the seminal quality. Implications Genotyping of rams based on melatonin receptor 1A could be a powerful tool in sire selection.

Identification of a set of genes potentially responsible for resistance to ferroptosis in lung adenocarcinoma cancer stem cells

Scientific literature supports the evidence that cancer stem cells (CSCs) retain inside low reactive oxygen species (ROS) levels and are, therefore, less susceptible to cell death, including ferroptosis, a type of cell death dependent on iron-driven lipid peroxidation. A collection of lung adenocarcinoma (LUAD) primary cell lines derived from malignant pleural effusions (MPEs) of patients was used to obtain 3D spheroids enriched for stem‐like properties. We observed that the ferroptosis inducer RSL3 triggered lipid peroxidation and cell death in LUAD cells when grown in 2D conditions; however, when grown in 3D conditions, all cell lines underwent a phenotypic switch, exhibiting substantial resistance to RSL3 and, therefore, protection against ferroptotic cell death. Interestingly, this phenomenon was reversed by disrupting 3D cells and growing them back in adherence, supporting the idea of CSCs plasticity, which holds that cancer cells have the dynamic ability to transition between a CSC state and a non-CSC state. Molecular analyses showed that ferroptosis resistance in 3D spheroids correlated with an increased expression of antioxidant genes and high levels of proteins involved in iron storage and export, indicating protection against oxidative stress and low availability of iron for the initiation of ferroptosis. Moreover, transcriptomic analyses highlighted a novel subset of genes commonly modulated in 3D spheroids and potentially capable of driving ferroptosis protection in LUAD-CSCs, thus allowing to better understand the mechanisms of CSC-mediated drug resistance in tumors.

Targeting ferroptosis as a potential strategy to overcome the resistance of cisplatin in oral squamous cell carcinoma.

Oral squamous cell carcinoma (OSCC) is a crucial public health problem, accounting for approximately 2% of all cancers globally and 90% of oral malignancies over the world. Unfortunately, despite the achievements in surgery, radiotherapy, and chemotherapy techniques over the past decades, OSCC patients still low 5-year survival rate. Cisplatin, a platinum-containing drug, serves as one of the first-line chemotherapeutic agents of OSCC. However, the resistance to cisplatin significantly limits the clinical practice and is a crucial factor in tumor recurrence and metastasis after conventional treatments. Ferroptosis is an iron-based form of cell death, which is initiated by the intracellular accumulation of lipid peroxidation and reactive oxygen species (ROS). Interestingly, cisplatin-resistant OSCC cells exhibit lower level of ROS and lipid peroxidation compared to sensitive cells. The reduced ferroptosis in cisplatin resistance cells indicates the potential relationship between cisplatin resistance and ferroptosis, which is proved by recent studies showing that in colorectal cancer cells. However, the modulation pathway of ferroptosis reversing cisplatin resistance in OSCC cells still remains unclear. This article aims to concisely summarize the molecular mechanisms and evaluate the relationship between ferroptosis and cisplatin resistance OSCC cells, thereby providing novel strategies for overcoming cisplatin resistance and developing new therapeutic approaches.

Synthetic modulation of ROS scavenging during host—Sclerotinia sclerotiorum interaction: a new strategy for the development of highly resistant plants

Sclerotinia sclerotiorum is a widespread fungal pathogen responsible for significant crop losses across the globe. The challenge of breeding resistant varieties is exacerbated by the fungus's sophisticated pathogenic mechanisms. A pivotal factor in the host-pathogen interaction is the regulation of reactive oxygen species (ROS) within both the fungi and the host plants. However, there is currently no efficient strategy to leverage this interaction mechanism for developing disease-resistant crop varieties. Here, we introduce an engineered ROS scavenging system designated as syn-ROS for impairing ROS neutralization in S. sclerotiorum while concurrently fortifying it in the host. The syn-ROS system comprises gene silencing constructs targeting the S. sclerotiorum Cu/Zn superoxide dismutase (SsSOD) and its copper chaperone (SsCCS), alongside overexpression constructs for the Arabidopsis thaliana AtSOD1 and AtCCS. Transgenic plants carrying the syn-ROS system demonstrated a marked enhancement in resistance to S. sclerotiorum. Upon infection, the expression of SsSOD and SsCCS was reduced, while the expression of AtSOD1 and AtCCS was enhanced in syn-ROS transgenic plants. Moreover, the infected syn-ROS plants showed decreased Cu/Zn SOD enzyme activity and elevated ROS concentrations within the fungal cells. In contrast, the cells of A. thaliana manifested increased Cu/Zn SOD enzyme activity and lowered ROS levels. Collectively, these findings suggest a novel and promising approach for contriving plants with robust resistance by synthetically manipulating ROS scavenging activities in the interaction between the host and S. sclerotiorum.

A Gene-Switch Platform Interfacing with Reactive Oxygen Species Enables Transcription Fine-Tuning by Soluble and Volatile Pharmacologics and Food Additives.

Synthetic biology aims to engineer transgene switches for precise therapeutic protein control in cell-based gene therapies. However, off-the-shelf trigger-inducible gene circuits are usually switched on by single or structurally similar molecules. This study presents a mammalian gene-switch platform that controls therapeutic gene expression by a wide range of molecules generating low, non-toxic levels of reactive oxygen species (ROS). In this system, KEAP1 (Kelch-like ECH-associated protein 1) serves as ROS sensor, regulating the translocation of NRF2 (nuclear factor erythroid 2-related factor 2) to the nucleus, where NRF2 binds to antioxidant response elements (ARE) to activate the expression of a gene of interest. It is found that a promoter containing eight-tandem ARE repeats is highly sensitive to the low ROS levels generated by the soluble and volatile molecules, which include food preservatives, food additives, pharmaceuticals, and signal transduction inducers. In a proof-of-concept study, it is shown that many of these compounds can independently trigger microencapsulated engineered cells to produce sufficient insulin to restore normoglycemia in experimental type-1 diabetic mice. It is believed that this system greatly extends the variety of small-molecule inducers available to drive therapeutic gene switches.

Proline-rich protein PRPL1 enhances Panax notoginseng defence against Fusarium solani by regulating reactive oxygen species balance and strengthening the cell wall barrier.

The root rot mainly caused by Fusarium solaniis a bottleneck in the cultivation of Panax notoginseng. In this study, we reported a gene encoding a plant cell wall structural protein, P. notoginseng proline-rich protein (PnPRPL1), whose transcription was upregulated by F. solani and induced by some hormone signals. The PnPRPL1 recombinant protein significantly inhibited the growth and conidial germination of the root rot pathogens. Downregulation of PnPRPL1 byRNA interference(RNAi) in P. notoginseng leaves increased the susceptibility to F. solani, whereas overexpression of PnPRPL1 in tobacco (Nicotiana tabacum) enhanced the resistance to F. solani. Compared with wild-type tobacco, the PnPRPL1-overexpressing transgenic tobacco had higher reactive oxygen species (ROS)-scavenging enzyme activities, lower ROS levels, and more lignin and callose deposition. The opposite results were obtained for the P. notoginseng expressing PnPRPL1 RNAi fragments. Furthermore, the PnPRPL1 promoter transcription activity was induced by several plant hormones and multiple stress stimuli. In addition, the transcription factor PnWRKY27 activated the expression of PnPRPL1 by directly binding to the promoter region. Thus, PnPRPL1, which is positively regulated by a WRKY transcription factor, encodes an antimicrobial protein that also mediates ROS homoeostasis and callose/lignin deposition during the response to F. solani infection.

Abstract 2960: Deciphering the role of miRNA146-a and its crosstalk with cellular redox state in senescence escape in breast cancer cells

Abstract Senescence serves as a double-edged sword in cancer as studies revealed that senescence exhibits both tumor-suppressing and tumor-promoting effects. A subpopulation of cancer cells that enters senescence can evade senescence to proliferate, thereby resulting in tumor regrowth, which has been associated with chemoresistance and tumor aggressiveness. Unlike senescent cells, the senescence-escaped cells harbor low levels of reactive oxygen species (ROS), however, the mechanism(s) underlying this redox dichotomy in senescence and senescence evasion remains elusive. In this regard, recent reports have highlighted the pivotal role of miRNAs in regulating cellular senescence. To that end, miR-146a has been reported as a biomarker of senescence probably via modulating pro-inflammatory and antioxidant pathways. We set up a model of doxorubicin-induced senescence in MDA-MB-231 breast carcinoma cells to study the role and involvement of miRNA146a in senescence induction and its regulation. Senescence induction was confirmed by upregulation of p21, increase in senescence-associated beta galactosidase and C12FDG staining, as well as propidium iodide positivity. We show that, following the initial phase of apoptotic cell death, as evidenced by increased yH2AX and cleaved caspase 3, a population of senescent cells escaped growth arrest to re-enter cell cycle and proliferate. Senescence-escaped cells harbored lower ROS levels compared to senescent cells, which correlated with upregulation of antioxidant enzymes such as MnSOD and CuZnSOD as well as increased transcription of NRF2 and NQO1. Our results also provide evidence for a novel senescence associated redox state, which implicates intracellular peroxynitrite as a driver of cellular senescence. Interestingly, in our model system, p21 upregulation was not associated with intracellular ROS accumulation, which was reported in earlier findings. Interestingly, these changes coincided with a significant increase in miR-146 expression (during senescence), which was previously shown to downregulate ROS in other model systems. While modulating miR-146a did not seem to change ROS levels upon doxorubicin-induced senescence in MDA-MB231 cells, the precise crosstalk between cellular ROS, miR146-a and senescence evasion is currently being pursued. Citation Format: Shazib Pervaiz, Angeline Q. Liew, Marie-Veronique Clement. Deciphering the role of miRNA146-a and its crosstalk with cellular redox state in senescence escape in breast cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2960.

Unveiling the Impact of Light-Induced Acceptor-Generated ROS on Device Stability in Organic Photovoltaics.

The intrinsic stability of the acceptor is a crucial component of the photovoltaic device stability. In this study, we investigated the efficiency and stability of the nonfused-ring acceptors LC8 and BC8 under indoor light conditions. Interestingly, we found that devices based on BC8 with terminal side chains exhibited a higher indoor efficiency and stability. Through accelerated aging experiments, we discovered that the acceptors generate singlet oxygen under light exposure with BC8 demonstrating lower levels of ROS compared to LC8. We attribute this difference to the modulation of the acceptor aggregation orientation. Furthermore, the generated reactive oxygen species (ROS) further deteriorate the acceptor structure, and this phenomenon is also observed in high-efficiency acceptor structures, such as Y6. Our research reveals important mechanisms of acceptor photo-oxidation processes, providing a theoretical basis for enhancing the intrinsic stability of acceptors.

MiR-375 impairs breast cancer cell stemness by targeting the KLF5/G6PD signaling axis.

Recurrence of breast cancer may be due to the presence of breast cancer stem cells (BCSC). Abnormal tumor cell growth is closely associated with increased reactive oxygen species (ROS) and disruption of redox homeostasis, and BCSCs exhibit low levels of ROS. The detailed mechanism between the low levels of ROS in BCSCs and their maintenance of stemness characteristics has not been reported. A growing number of studies have shown that tumor development is often accompanied by metabolic reprogramming, which is an important hallmark of tumor cells. As the first rate-limiting enzyme of pentose phosphate pathway (PPP), the expression of G6PD is precisely regulated in tumor cells, and there is a certain correlation between PPP and BCSCs. MiR-375 has been shown to inhibit stem cell-like properties in breast cancer, but the exact mechanism is not clear. Here, KLF5, as a transcription factor, was identified to bind to the promoter of G6PD to promote its expression, whereas miR-375 inhibited the expression of KLF5 by binding to the 3'UTR region of KLF5 mRNA and thus reduced the expression of G6PD expression, inhibits PPP to reduce NADPH, and increases ROS levels in breast cancer cells, thereby weakening breast cancer cell stemness. Our study reveals the specific mechanism by which miR-375 targets the KLF5/G6PD signaling axis to diminish the stemness of breast cancer cells, providing a therapeutic strategy against BCSCs.

Manipulating Redox Homeostasis of Cancer Stem Cells Overcome Chemotherapeutic Resistance through Photoactivatable Biomimetic Nanodiscs.

Tumor heterogeneity remains a significant obstacle in cancer therapy due to diverse cells with varying treatment responses. Cancer stem-like cells (CSCs) contribute significantly to intratumor heterogeneity, characterized by high tumorigenicity and chemoresistance. CSCs reside in the depth of the tumor, possessing low reactive oxygen species (ROS) levels and robust antioxidant defense systems to maintain self-renewal and stemness. A nanotherapeutic strategy is developed using tumor-penetrating peptide iRGD-modified high-density lipoprotein (HDL)-mimetic nanodiscs (IPCND) that ingeniously loaded with pyropheophorbide-a (Ppa), bis (2-hydroxyethyl) disulfide (S-S), and camptothecin (CPT) by synthesizing two amphiphilic drug-conjugated sphingomyelin derivatives. Photoactivatable Ppa can generate massive ROS which as intracellular signaling molecules effectively shut down self-renewal and trigger differentiation of the CSCs, while S-S is utilized to deplete GSH and sustainably imbalance redox homeostasis by reducing ROS clearance. Simultaneously, the depletion of GSH is accompanied by the release of CPT, which leads to subsequent cell death. This dual strategy successfully disturbed the redox equilibrium of CSCs, prompting their differentiation and boosting the ability of CPT to kill CSCs upon laser irradiation. Additionally, it demonstrated a synergistic anti-cancer effect by concurrently eliminating therapeutically resistant CSCs and bulk tumor cells, effectively suppressing tumor growth in CSC-enriched heterogeneous colon tumor mouse models.

PH and ROS Responsiveness of Polymersome Nanovaccines for Antigen and Adjuvant Codelivery: An In Vitro and In Vivo Comparison.

The antitumor immunity can be enhanced through the synchronized codelivery of antigens and immunostimulatory adjuvants to antigen-presenting cells, particularly dendritic cells (DCs), using nanovaccines (NVs). To study the influence of intracellular vaccine cargo release kinetics on the T cell activating capacities of DCs, we compared stimuli-responsive to nonresponsive polymersome NVs. To do so, we employed "AND gate" multiresponsive (MR) amphiphilic block copolymers that decompose only in response to the combination of chemical cues present in the environment of the intracellular compartments in antigen cross-presenting DCs: low pH and high reactive oxygen species (ROS) levels. After being unmasked by ROS, pH-responsive side chains are exposed and can undergo a charge shift within a relevant pH window of the intracellular compartments in antigen cross-presenting DCs. NVs containing the model antigen Ovalbumin (OVA) and the iNKT cell activating adjuvant α-Galactosylceramide (α-Galcer) were fabricated using microfluidics self-assembly. The MR NVs outperformed the nonresponsive NV in vitro, inducing enhanced classical- and cross-presentation of the OVA by DCs, effectively activating CD8+, CD4+ T cells, and iNKT cells. Interestingly, in vivo, the nonresponsive NVs outperformed the responsive vaccines. These differences in polymersome vaccine performance are likely linked to the kinetics of cargo release, highlighting the crucial chemical requirements for successful cancer nanovaccines.

Apigenin-7-glucoside-loaded nanoparticle alleviates intestinal ischemia-reperfusion by ATF3/SLC7A11-mediated ferroptosis

Intestinal ischemia reperfusion injury (II/R injury) is a common and intractable pathophysiological process in critical patients, for which exploring new treatments and mechanisms is of great importance to improve treatment outcomes. Apigenin-7-O-Glucoside (AGL) is a sugar derivative of apigenin natural product with various pharmacological activities to protect against intestinal diseases. In this study, we synthesized two amphiphilic molecules, namely DTPA-N10–10 and mPEG-TK-DA, which can scavenge free radicals and reactive oxygen species (ROS). They were successfully encapsulated AGL through self-assembly, resulting in the formation of multi-site ROS scavenging nanoparticles called PDN@AGL. In vitro and in vivo experiments demonstrated that PDN@AGL could protect intestinal tissues by reducing lipid peroxidation, lowering ROS levels and inhibiting ferroptosis during II/R injury. Furthermore, our study revealed, for the first time, that the regulation of the ATF3/SLC7A11 pathway by PDN@AGL may play a crucial role in mitigating II/R injury. In conclusion, our study confirmed the beneficial effects of PDN@AGL in combating II/R injury through the ATF3/SLC7A11-mediated regulation of ferroptosis and oxidative stress. These findings lay the groundwork for the potential application of PDN@AGL in the treatment of II/R injury.

Exploration of glutaredoxin-1 oxidative modification in carbon nanomaterial-induced hepatotoxicity.

Herein, we present toxicological assessments of carbon nanomaterials in HL-7702 cells, and it was found that reactive oxygen species (ROS) levels were elevated. Mass spectrometry results indicated that cysteine sulfhydryl of glutaredoxin-1 (GLRX1) was oxidized to sulfenic acids and sulfonic acids by excessive ROS, which broke the binding of GLRX1 to apoptosis signal-regulating kinase 1, causing the activation of the JNK/p38 signaling pathway and ultimately hepatocyte apoptosis. However, a lower level of ROS upregulated GLRX1 instead of sulfonation modification of its active sites. Highly expressed GLRX1 in turn enabled the removal of intracellular ROS, thereby exerting inconspicuous toxic effects on cells. Taken together, these findings emphasized that CNM-induced hepatotoxicity is attributable to oxidative modifications of GLRX1 arising from redox imbalance.

M6A demethylase ALKBH5 regulates FOXO1 mRNA stability and chemoresistance in triple-negative breast cancer

Resistance to chemotherapy is the main reason for treatment failure and poor prognosis in patients with triple-negative breast cancer (TNBC). Although the association of RNA N6-methyladenosine (m6A) modifications with therapy resistance is noticed, its role in the development of therapeutic resistance in TNBC is not well documented. This study aimed to investigate the potential mechanisms underlying reactive oxygen species (ROS) regulation in doxorubicin (DOX)-resistant TNBC. Here, we found that DOX-resistant TNBC cells displayed low ROS levels because of increased expression of superoxide dismutase (SOD2), thus maintaining cancer stem cells (CSCs) characteristics and DOX resistance. FOXO1 is a master regulator that reduces cellular ROS in DOX-resistant TNBC cells, and knockdown of FOXO1 significantly increased ROS levels by inhibiting SOD2 expression. Moreover, the m6A demethylase ALKBH5 promoted m6A demethylation of FOXO1 mRNA and increased FOXO1 mRNA stability in DOX-resistant TNBC cells. The analysis of clinical samples revealed that the increased expression levels of ALKBH5, FOXO1, and SOD2 were significantly positively correlated with chemoresistance and poor prognosis in patients with TNBC. To our knowledge, this is the first study to highlight that ALKBH5-mediated FOXO1 mRNA demethylation contributes to CSCs characteristics and DOX resistance in TNBC cells. Furthermore, pharmacological targeting of FOXO1 profoundly restored the response of DOX-resistant TNBC cells, both in vitro and in vivo. In conclusion, we demonstrated a critical function of ALKBH5-mediated m6A demethylation of FOXO1 mRNA in restoring redox balance, which in turn promoting CSCs characteristics and DOX resistance in TNBC, and suggested that targeting the ALKBH5/FOXO1 axis has therapeutic potential for patients with TNBC refractory to chemotherapy.

Effects of hyperglycemia on the TGF-β pathway in trabecular meshwork cells

BackgroundHyperglycemia, which can lead to apoptosis, hypertrophy, fibrosis, and induces hyperinflammation in diabetic vascular complications due to oxidative stress. In order to elucidate the potential dual roles and regulatory signal transduction of TGF-β1 and TGF-β2 in human trabecular meshwork cells (HTMCs), we established an oxidative cell model in HTMCs using 5.5, 25, 50, and 100 mM d-glucose-supplemented media and characterized the TGF-β-related oxidative stress pathway. MethodsFurther analysis was conducted to investigate oxidative damage and protein alterations in the HTMC caused by the signal transduction. This was done through a series of qualitative cell function studies, such as cell viability/apoptosis analysis, intracellular reactive oxygen species (ROS) detection, analysis of calcium release concentration, immunoblot analysis to detect the related protein expression alteration, and analysis of cell fibrosis to study the effect of different severities of hyperglycemia. Also, we illustrated the role of TGF-β1/2 in oxidative stress-induced injury by shRNA-mediated knockdown or stimulation with recombinant human TGF-β1 protein (rhTGF-β1). ResultsResults from the protein expression analysis showed that p-JNK, p-p38, p-AKT, and related SMAD family members were upregulated in HTMCs under hyperglycemia. In the cell functional assays, HTMCs treated with rhTGFβ-1 (1 ng/mL) under hyperglycemic conditions showed higher proliferation rates and lower ROS and calcium levels. ConclusionsTo summarize, mechanistic analyses in HTMCs showed that hyperglycemia-induced oxidative stress activated TGF-β1 along with its associated pathway. General Significance: While at low concentrations, TGF-β1 protects cells from antioxidation, whereas at high concentrations, it accumulates in the extracellular matrix, causing further HTMC dysfunction.

ROS regulation in gliomas: implications for treatment strategies.

Gliomas are one of the most common primary malignant tumours of the central nervous system (CNS), of which glioblastomas (GBMs) are the most common and destructive type. The glioma tumour microenvironment (TME) has unique characteristics, such as hypoxia, the blood-brain barrier (BBB), reactive oxygen species (ROS) and tumour neovascularization. Therefore, the traditional treatment effect is limited. As cellular oxidative metabolites, ROS not only promote the occurrence and development of gliomas but also affect immune cells in the immune microenvironment. In contrast, either too high or too low ROS levels are detrimental to the survival of glioma cells, which indicates the threshold of ROS. Therefore, an in-depth understanding of the mechanisms of ROS production and scavenging, the threshold of ROS, and the role of ROS in the glioma TME can provide new methods and strategies for glioma treatment. Current methods to increase ROS include photodynamic therapy (PDT), sonodynamic therapy (SDT), and chemodynamic therapy (CDT), etc., and methods to eliminate ROS include the ingestion of antioxidants. Increasing/scavenging ROS is potentially applicable treatment, and further studies will help to provide more effective strategies for glioma treatment.

Aerobic exercise improves BKCa channel-mediated vasodilation in diabetic vascular smooth muscle via AMPK/Nrf2/HO-1 pathway.

This study aims to investigate the effect of aerobic exercise training on BKCa channel in diabetic vascular smooth muscle and explore the underlying mechanism. Control m/m mice and diabetic db/db mice were randomly assigned to sedentary groups (W and D) and exercise training groups (WE and DE). Mice in exercise groups underwent training sessions lasting for 12weeks, with a speed of 12m/min for 60min, five times per week. The thoracic aorta was extracted isolated and examined for measurement of vascular structure, global levels of reactive oxygen species (ROS), vasodilation, and protein expression. Rat thoracic aorta vascular smooth muscle cells (USMCs) were cultured, and siRNA transfection was conducted to detect whether AMPK contributed to the regulation. ROS level and protein expression were measured. Compared with control mice, diabetic mice had a larger vascular medium thickness, impaired BKCa-mediated vasodilation, a higher level of ROS, and a lower expression of BKCa α, BKCa β1, Nrf2, p-Nrf2, p-Nrf2/Nrf2, HO-1, and p-AMPK/AMPK. Exercise training increased the expression of BKCa α, Nrf2, p-Nrf2, p-Nrf2/Nrf2, HO-1, and p-AMPK/AMPK. AMPK deletion led to lower ROS levels and expression of BKCa α, β1, Nrf2, and HO-1 in USMCs cultured in high glucose conditions. BKCa channel protein expression reduction in VSMCs contributes to vasodilation and vascular remodeling dysfunction in diabetes mellitus. Aerobic exercise can promote the expression of BKCa channel and improve BKCa-mediated vascular dysfunction in diabetic VSMCs through AMPK/Nrf2/HO-1 signaling pathway.

Enhancement of porcine in vitro embryonic development through luteolin-mediated activation of the Nrf2/Keap1 signaling pathway

BackgroundOxidative stress, caused by an imbalance in the production and elimination of intracellular reactive oxygen species (ROS), has been recognized for its detrimental effects on mammalian embryonic development. Luteolin (Lut) has been documented for its protective effects against oxidative stress in various studies. However, its specific role in embryonic development remains unexplored. This study aims to investigate the influence of Lut on porcine embryonic development and to elucidate the underlying mechanism.ResultsAfter undergoing parthenogenetic activation (PA) or in vitro fertilization, embryos supplemented with 0.5 µmol/L Lut displayed a significant enhancement in cleavage and blastocyst formation rates, with an increase in total cell numbers and a decrease in the apoptosis rate compared to the control. Measurements on D2 and D6 revealed that embryos with Lut supplementation had lower ROS levels and higher glutathione levels compared to the control. Moreover, Lut supplementation significantly augmented mitochondrial content and membrane potential. Intriguingly, activation of the Nrf2/Keap1 signaling pathway was observed in embryos supplemented with Lut, leading to the upregulation of antioxidant-related gene transcription levels. To further validate the relationship between the Nrf2/Keap1 signaling pathway and effects of Lut in porcine embryonic development, we cultured PA embryos in a medium supplemented with brusatol, with or without the inclusion of Lut. The positive effects of Lut on developmental competence were negated by brusatol treatment.ConclusionsOur findings indicate that Lut-mediated activation of the Nrf2/Keap1 signaling pathway contributes to the enhanced production of porcine embryos with high developmental competence, and offers insight into the mechanisms regulating early embryonic development.

Sphingomyelinase Regulates the Release of Protumoral Mitochondria Containing-Vesicles from AML Cells

Mitochondrial transfer from bone marrow stromal cells to acute myeloid leukaemia (AML) blasts is protumoral (Marlein et al., 2017; Moschoi et al., 2016) . We have previously shown that this was through a NADPH oxidase-2 (Nox2) dependent mechanism which increases AML fitness both in vitro and in vivo. Furthermore, we recently have shown that AML cells export mitochondria in vesicles which are subsequently phagocytosed by bone marrow macrophages (Moore et al., 2022) . Next, we have investigated how the release of mitochondria-containing vesicles is regulated and how this process contributes to the fitness of AML blasts. Sphingolipids are a component of the cell membrane which impact the stability and curvature of the membrane. They have been shown to play a role in the biogenesis of vesicles and a number of the enzymes involved in sphingolipid metabolism are dysregulated in AML, therefore we investigated the role of sphingolipids in mitochondrial exocytosis via vesicle formation. To understand the role of vesicles in regulating AML fitness, we used primary human AML (collected with approval on the East of England - Cambridgeshire and Hertfordshire Research Ethics Committee and the UK Health Research Authority, IRAS Project ID - 33753) and mouse syngeneic AML models (MN1 and HOXA9/Meis). We find that the myosin inhibitor blebbistatin inhibits vesicle release and results in an increased mitochondrial mass and increased apoptosis. In order to investigate the involvement of one of the major sphingolipids, ceramide, we targeted neutral sphingomyelinase (GW4869) which is responsible for ceramide synthesis. We treated the AML cells in vitro with GW4869 for 24 h which induced a large increase in vesicle release. Using the mitochondrial stain Mitotracker Green (MTG) we confirm that these vesicles contain mitochondria. The vesicles were isolated, DNA was extracted, and RT-qPCR performed which confirms increased mitochondrial DNA content. Others have previously shown that low levels of cellular reactive oxygen species (ROS) are associated with leukaemic stem cells (LSCs). DCF staining of AML treated with GW4869 shows low levels of cellular ROS. To assess how this process might impact the stemness of AML, we performed in vivo experiments. Pretreatment of MN1 cells with GW4869 accelerated engraftment in C57Bl/6 animals which corresponded with a decrease in cellular ROS. In summary, we have demonstrated the mechanism of mitochondrial exocytosis is important for AML fitness and that by disrupting sphingolipid metabolism, dysregulation of this process leads to increased tumour burden.