Increased Reactive Oxygen Species Levels Research Articles

Anticancer Effect of Dihydroartemisinin via Dual Control of ROS-induced Apoptosis and Protective Autophagy in Prostate Cancer 22Rv1 Cells.

Dihydroartemisinin (DHA), a natural agent, exhibits potent anticancer activity. However, its biological activity on prostate cancer (PCa) 22Rv1 cells has not been previously investigated. In this study, we demonstrate that DHA induces anticancer effects through the induction of apoptosis and autophagy. Cell viability and proliferation rate were assessed using the CCK-8 assay and cell clone formation assay. The generation of reactive oxygen species (ROS) was detected by flow cytometry. The molecular mechanism of DHA-induced apoptosis and autophagy was examined using Western blot and RT-qPCR. The formation of autophagosomes and the changes in autophagy flux were observed using transmission electron microscopy (TEM) and confocal microscopy. The effect of DHA combined with Chloroquine (CQ) was assessed using the EdU assay and flow cytometry. The expressions of ROS/AMPK/mTOR-related proteins were detected using Western blot. The interaction between Beclin-1 and Bcl-2 was examined using Co-IP. DHA inhibited 22Rv1 cell proliferation and induced apoptosis. DHA exerted its antiprostate cancer effects by increasing ROS levels. DHA promoted autophagy progression in 22Rv1 cells. Inhibition of autophagy enhanced the pro-apoptotic effect of DHA. DHA-induced autophagy initiation depended on the ROS/AMPK/mTOR pathway. After DHA treatment, the impact of Beclin- 1 on Bcl-2 was weakened, and its binding with Vps34 was enhanced. DHA induces apoptosis and autophagy in 22Rv1 cells. The underlying mechanism may involve the regulation of ROS/AMPK/mTOR signaling pathways and the interaction between Beclin-1 and Bcl-2 proteins. Additionally, the combination of DHA and CQ may enhance the efficacy of DHA in inhibiting tumor cell activity.

New CDDO Arylboronate Ester Derivatives with High Selectivity and Low Toxicity.

As an oleanolic acid derivative, CDDO-Me lacks selectivity for tumors. Based on the high reactive oxygen species (ROS) level in cancer cells, compound 4 was selected from 17 new CDDO arylboronate ester derivatives. A preliminary study revealed that 4 displayed the highest selectivity for cancer cells. Furthermore, 4 could be transformed to 4H by ROS to increase its covalent binding ability and antiproliferation effect (IC50 of 2.11 vs 0.37 μM) in BGC-823 cells. Interestingly, 4 increased ROS levels to induce apoptosis in BGC-823 cells. Moreover, the LD50 of 4 (91.2 mg/kg) was much greater than that of CDDO-Me (61.7 mg/kg) in ICR mice. A pharmacokinetic study indicated that 4 could be transformed to 4H in vivo. In addition, 4 exhibited a greater tumor inhibition rate (86.2%) than CDDO-Me (51.7%). Overall, the design of 4 provided an effective modification strategy for CDDO to increase the selectivity for cancer cells.

Differentiated management of ROS level in tumor and kidney to alleviate Cis-platinum induced acute kidney injury with improved efficacy

Cisplatin (DDP) is a prevalent chemotherapeutic agent used in tumor therapy, yet DDP-induced acute kidney injury (AKI) severely limits its clinical application. Antioxidants as reactive oxygen species (ROS) scavengers can circumvent this adverse effect while leading to the decrease of efficacy to tumor. Herein, we report ultrasmall ruthenium nanoparticles (URNPs) as switchable ROS scavengers/generators to alleviate DDP-induced AKI and improve its therapeutic efficacy. In the physiological environment of the kidney, URNPs mimic multi-enzyme activities, such as superoxide dismutase and catalase, effectively protecting the renal cell and tissue by down-regulating the increased ROS level caused by DDP and alleviating AKI. Specifically, URNPs are oxidized by high levels of H2O2 in the tumor microenvironment (TME), resulting in the generation of oxygen vacancies and Ru3+/Ru4+ ions. This unique structure transformation endows URNPs to generate singlet oxygen (1O2) under laser irradiation and hydroxyl radicals (∙OH) through a Fenton-like reaction in tumor cell and tissue. The simultaneous generation of multifarious ROS effectively improves the efficacy of DDP in vitro and in vivo. This TME-responsive ROS scavenger/generator acts as an adjuvant therapeutic agent to minimize side effects and improve the efficacy of chemotherapy drugs, providing a new avenue to chemotherapy and facilitating clinical tumor therapy.Graphical

Overexpression of EgrZFP6 from Eucalyptus grandis increases ROS levels by downregulating photosynthesis in plants

In plants, abiotic stressors are frequently encountered during growth and development. To counteract these challenges, zinc finger proteins play a critical role as transcriptional regulators. The EgrZFP6 gene, which codes for a zinc finger protein of the C2H2 type, was shown to be considerably elevated in the leaves of Eucalyptus grandis seedlings in the current study when they were subjected to a variety of abiotic stimuli, including heat, salinity, cold, and drought. Analysis conducted later showed that in EgrZFP6 transgenic Arabidopsis thaliana, EgrZFP6 was essential for causing hyponastic leaves and controlling the stress response. Furthermore, the transgenic plants showed elevated levels of reactive oxygen species (ROS), such as superoxide and hydrogen peroxide (H2O2). Additionally, in EgrZFP6-overexpressing plants, transcriptome sequencing analysis demonstrated a considerable downregulation of many genes involved in photosynthesis, decreasing electron transport efficiency and perhaps promoting the buildup of ROS. Auxin levels were higher and auxin signal transduction was compromised in the transgenic plants. Stress-related genes were also upregulated in Arabidopsis as a result of EgrZFP6 overexpression. It is hypothesized that EgrZFP6 can downregulate photosynthesis, which would cause the production of ROS in chloroplasts. As a result, this protein may alter plant stress responses and leaf morphology via a retrograde mechanism driven by ROS. These results highlight the significance of zinc finger proteins in this sophisticated process and advance our understanding of the complex link between gene regulation, ROS signaling, and plant stress responses.

Piceatannol induces caspase-dependent apoptosis by modulating intracellular reactive oxygen species/mitochondrial membrane potential and enhances autophagy in neuroblastoma cells.

The aim of this study was to evaluate the anticancer effects of piceatannol, a natural stilbenoid, on human neuroblastoma cells. In order to accomplish this goal, we performed various cellular assays, including the XTT cell proliferation assay for cell viability, colony formation assay for colony formation capacity, FITC Annexin V and cell death detection kit for apoptosis, matrigel invasion assay for invasion capacity, intracellular reactive oxygen species (ROS) red dye for intracellular ROS levels, TMRM staining method for mitochondrial membrane potential (MMP), and the CYTO-ID autophagy detection kit for autophagy. Furthermore, we analyzed the expression levels of genes associated with apoptosis and autophagy using RT-qPCR. Based on our findings, piceatannol exhibited cytotoxic effects on neuroblastoma cells. Besides, treatment with piceatannol at both 50 and 100 μM concentrations for 72 h decreased colony formation, induced apoptosis and autophagy, inhibited cell invasion, decreased MMP, and increased ROS levels in SH-SY5Y cells. In addition, we observed significant upregulation in the expression levels of CASP8, BECLIN, ATG5, ATG7, and MAPILC3A genes between the two doses. These results suggest that piceatannol enhances autophagic activity and induces caspase-dependent apoptosis, indicating its potential as a therapeutic agent against neuroblastoma cells.

Cantharidin overcomes IL-2Rα signaling-mediated vorinostat resistance in cutaneous T-cell lymphoma through reactive oxygen species

BackgroundVorinostat (SAHA) is a histone deacetylase inhibitor that has shown clinical efficacy against advanced cutaneous T-cell lymphoma (CTCL). However, only a subset of patients with CTCL (30–35%) respond to SAHA...

The Protective Effect of Melatonin on LPS-Induced Myocardial Injury via the Caspase-11/GSDMD Pathway.

Melatonin (MT) has been demonstrated to have cardioprotective effects. Nevertheless, the precise mechanism through which MT provides protection against the etiology of LPS-induced myocardial injury remains uncertain. In this investigation, our objective was to explore the impact of MT on LPS-induced myocardial injury in an in vitro setting. H9C2 cells were categorized into four groups: a control group (H9C2 group), an MT group, an LPS group, and an MT + LPS group. The H9C2 group received treatment with sterile saline solution, the LPS group was exposed to 5 μg/mL LPS for 24 hours, the MT + LPS group underwent pretreatment with 150 μmol/L MT for 2 hours, followed by exposure to 5 μg/mL LPS for 24 hours, and the MT group received only 150 μmol/L MT for 2 hours. Cell viability and lactate dehydrogenase (LDH) release were assessed using the CCK-8 assay and LDH activity assay, respectively. The levels of reactive oxygen species (ROS) were quantified in each group of cells, and the percentage of propidium iodide (PI)-stained apoptotic cells was determined by flow cytometry. The mRNA levels of caspase11, GSDMD, and IL-18 in each group of cells were quantified. MT treatment significantly protected H9C2 cells from LPS-induced damage, as evidenced by decreased LDH release. LPS treatment markedly increased ROS levels in H9C2 cells, which were subsequently reduced by MT. LPS caused a substantial decrease in superoxide dismutase (SOD) activity and a significant increase in malondialdehyde (MDA) levels, while MT treatment significantly reversed these effects. Additionally, MT markedly enhanced the proportion of viable H9C2 cells compared to LPS-treated controls, as evidenced by the PI staining assay. LPS upregulated both mRNA levels and protein levels of IL-18 in H9C2 cells. However, MT treatment effectively mitigated this LPS-induced increase. Furthermore, MT significantly decreased LPS-induced protein levels of cleaved-caspase 11 and GSDMD-N in H9C2 cells. Overall, our findings suggest that MT inhibits the Caspase11-GSDMD signaling pathway via pyroptosis-related proteins (caspase-11 and GSDMD-N) and reduces the expression of inflammation-related cytokines (IL-18), thereby exerting a protective effect on H9C2 cells after LPS injury.

Targeting ROS in cancer: rationale and strategies.

Reactive oxygen species (ROS) in biological systems are transient but essential molecules that are generated and eliminated by a complex set of delicately balanced molecular machineries. Disruption of redox homeostasis has been associated with various human diseases, especially cancer, in which increased ROS levels are thought to have a major role in tumour development and progression. As such, modulation of cellular redox status by targeting ROS and their regulatory machineries is considered a promising therapeutic strategy for cancer treatment. Recently, there has been major progress in this field, including the discovery of novel redox signalling pathways that affect the metabolism of tumour cells as well as immune cells in the tumour microenvironment, and the intriguing ROS regulation of biomolecular phase separation. Progress has also been made in exploring redox regulation in cancer stem cells, the role of ROS in determining cell fate and new anticancer agents that target ROS. This Review discusses these research developments and their implications for cancer therapy and drug discovery, as well as emerging concepts, paradoxes and future perspectives.

Targeting ROS in cancer: rationale and strategies.

Reactive oxygen species (ROS) in biological systems are transient but essential molecules that are generated and eliminated by a complex set of delicately balanced molecular machineries. Disruption of redox homeostasis has been associated with various human diseases, especially cancer, in which increased ROS levels are thought to have a major role in tumour development and progression. As such, modulation of cellular redox status by targeting ROS and their regulatory machineries is considered a promising therapeutic strategy for cancer treatment. Recently, there has been major progress in this field, including the discovery of novel redox signalling pathways that affect the metabolism of tumour cells as well as immune cells in the tumour microenvironment, and the intriguing ROS regulation of biomolecular phase separation. Progress has also been made in exploring redox regulation in cancer stem cells, the role of ROS in determining cell fate and new anticancer agents that target ROS. This Review discusses these research developments and their implications for cancer therapy and drug discovery, as well as emerging concepts, paradoxes and future perspectives.

Evaluating the phytotoxicological effects of bulk and nano forms of zinc oxide on cellular respiration-related indices and differential gene expression in Hordeum vulgare L.

The increasing use of nanoparticles is driving the growth of research on their effects on living organisms. However, studies on the effects of nanoparticles on cellular respiration are still limited. The remodeling of cellular-respiration-related indices in plants induced by zinc oxide nanoparticles (nnZnO) and its bulk form (blZnO) was investigated for the first time. For this purpose, barley (Hordeum vulgare L.) seedlings were grown hydroponically for one week with the addition of test compounds at concentrations of 0, 0.3, 2, and 10 mg mL−1. The results showed that a low concentration (0.3 mg mL−1) of blZnO did not cause significant changes in the respiration efficiency, ATP content, and total reactive oxygen species (ROS) content in leaf tissues. Moreover, a dose of 0.3 mg mL−1 nnZnO increased respiration efficiency in both leaves (17 %) and roots (38 %). Under the influence of blZnO and nnZnO at medium (2 mg mL−1) and high (10 mg mL−1) concentrations, a dose-dependent decrease in respiration efficiency from 28 % to 87 % was observed. Moreover, the negative effect was greater under the influence of nnZnO. The gene transcription of the subunits of the mitochondria electron transport chain (ETC) changed mainly only under the influence of nnZnO in high concentration. Expression of the ATPase subunit gene, atp1, increased slightly (by 36 %) in leaf tissue under the influence of medium and high concentrations of test compounds, whereas in the root tissues, the atp1 mRNA level decreased significantly (1.6–2.9 times) in all treatments. A dramatic decrease (1.5–2.4 times) in ATP content was also detected in the roots. Against the background of overexpression of the AOX1d1 gene, an isoform of alternative oxidase (AOX), the total ROS content in leaves decreased (with the exception of 10 mg mL−1 nnZnO). However, in the roots, where the pressure of the stress factor is higher, there was a significant increase in ROS levels, with a maximum six-fold increase under 10 mg mL−1 nnZnO. A significant decrease in transcript levels of the pentose phosphate pathway and glycolytic enzymes was also shown in the root tissues compared to leaves. Thus, the disruption of oxidative phosphorylation leads to a decrease in ATP synthesis and an increase in ROS production; concomitantly reducing the efficiency of cellular respiration.

Luteolin and Quercetin combination therapy: Enhanced inhibition of H157 human lung cancer cells

IntroductionTraditional Chinese Medicine (TCM) is renowned for its holistic treatment approach and minimal side effects. Luteolin (LUT) and Quercetin (QUE) each possess notable anticancer properties. This study is the first to elucidate the synergistic anticancer activity of the combination of LUT and QUE (LUT+QUE) against lung cancer. MethodsThe synergistic anticancer activity of QUE+LUT was evaluated through the combination index (CI) value by varying the concentrations of QUE and LUT. Network pharmacology analysis was then employed to identify the key targets and crucial signaling pathways for QUE and LUT. Finally, the synergistic anticancer mechanisms of QUE+LUT were elucidated by verifying these core targets and pathways through various experiments, including apoptosis assays, migration assays, ELISA, Western blot and reactive oxygen species (ROS) assays. ResultsThe key targets identify through network pharmacology analysis include BCL2L1, MMP9, JUN, PTGS2, TP53, AKT1 and CASP3. The crucial pathways are the PI3K-Akt signaling pathway and chemically-induce carcinogenic ROS pathways. Cellular experiments demonstrate that QUE+LUT synergistically induce apoptosis, inhibit cell proliferation, migration and invasion, increase ROS levels and regulate the PI3K-Akt signaling pathway, effectively targeting lung cancer cells. DiscussionThe combined therapy of QUE and LUT holds potential as an effective treatment for cancer patients, providing valuable insights for the selection and mechanistic exploration of important anticancer drugs.

Neuropathological implication of high blood bilirubin in patients and model rats with depression

PurposeElevated bilirubin levels have been associated with major depressive disorder (MDD); however, the exact impact of bilirubin on MDD and the underlying molecular mechanisms remain unclear. Here, we explored the influence of bilirubin on MDD and sought to identify the mechanisms via which bilirubin induces depressive-like behavior. Patients and methodsForty patients who were diagnosed with MDD and received treatment with selective serotonin reuptake inhibitors (SSRIs) were included, with 43 healthy volunteers serving as controls. Clinical symptoms were evaluated using Hamilton depression rating scale-24 (HAMD-24) and the Hamilton anxiety rating scale. Serum concentrations of total bilirubin (TBIL) and indirect bilirubin (IBIL) were measured at baseline and after treatment using an automated biochemical analyzer. The connection between clinical symptoms and TBIL or IBIL was examined using Pearson correlation. Chronic restraint stress (CRS) was employed to generate a rat model of depression. TBIL, IBIL in rat serum were measured by ELISA. Reactive oxygen species (ROS) contents in rat hippocampal tissues were quantified by flow cytometry. The levels of microglial markers and the extent of neuronal damage in the rat hippocampus were assessed by immunofluorescence and transmission electron microscopy, respectively. ResultsSerum TBIL and IBIL levels were higher in patients with MDD than in the healthy controls. After treatment with SSRIs, the serum levels of TBIL and IBIL in MDD patients were significantly reduced. The levels of TBIL and IBIL were associated with HAMD-24 in MDD patients. Compared with the controls, the serum levels of TBIL, IBIL and the hippocampal ROS contents were elevated in CRS-exposed rats. Fluoxetine lowered inflammatory factor levels, mitigated oxidative stress. ConclusionOur findings indicate a possible correlation between elevated serum bilirubin and depressive symptoms. Increases in ROS levels, along with neuronal damage, may represent pathological mechanisms underlying MDD.

Mechanism of ergosterol peroxide on MCF-7 breast cancer cells based on network pharmacology and in vitro experiments

This study investigated the effects of ergosterol peroxide(EP) on the proliferation and apoptosis of MCF-7 breast cancer cells, explored its possible mechanisms of action, and verified the effects and mechanisms by in vitro experiments. Network pharmaco-logy was used to screen the target proteins of EP and construct target networks and protein-protein interaction(PPI) networks to predict the potential target proteins and related pathways involved in EP anti-breast cancer effects. The MTT assay was performed to measure the inhibitory effect of EP on MCF-7 cell proliferation, and the colony formation assay was used to assess the cell cloning ability. Flow cytometry and laser confocal microscopy were employed to evaluate cell apoptosis, mitochondrial membrane potential and reactive oxygen species(ROS) levels. Western blot analysis was conducted to examine the expression levels of B-cell lymphoma 2(Bcl-2), Bcl-2-associated X protein(Bax), cytochrome C(Cyt C), caspase-7, cleaved caspase-7, phosphatidylinositol 3-kinase(PI3K), and se-rine/threonine kinase B(AKT) in MCF-7 cells treated with EP. The results of network pharmacology prediction yielded 173 common targets between EP and breast cancer; the results of Kyoto Encyclopedia of Genes and Genomes(KEGG) enrichment analysis showed that EP treatment for breast cancer mainly affected the signaling pathways such as cancer pathway, PI3K-AKT signaling pathway, cellular senescence signaling pathway, and viral carcinogenesis pathway; and the MTT assay results showed that the viability of MCF-7 cells in the EP group was significantly lower than that in the control group, exhibiting a time-and concentration-dependent trend, and EP can inhibit colony formation of MCF-7 breast cancer cells. Treatment with 10, 20, and 40 μmol·L~(-1) EP for 24 h resulted in a significant increase in the total apoptosis rate of MCF-7 cells, a significant decrease in mitochondrial membrane potential, and a significant increase in ROS levels. In addition, treatment with EP led to an upregulation of Cyt C, Bax, and cleaved caspase-7 protein expression, and a downregulation of p-PI3K, p-AKT, and Bcl-2 protein expression in MCF-7 cells. Studies have shown that EP inhibits MCF-7 breast cancer cell proliferation and reduces colony formation by a mechanism that may be related to the PI3K-AKT pathway mediating the mitochondrial apoptotic pathway.

Expression levels of the selenium-uptake receptor LRP8, the antioxidant selenoprotein GPX1 and steroidogenic enzymes correlate in granulosa cells.

Reactive oxygen species (ROS) are a by-product of the activity of cytochrome P450 steroidogenic enzymes. Antioxidant enzymes protect against ROS damage. To identify if any particular antioxidant enzyme is used to protect against ROS produced by granulosa cells as follicles enlarge and produce oestradiol, we measured in the bovine granulosa cells the expression of two steroidogenic enzymes (CYP11A1, CYP19A1), important for progesterone and oestradiol production. We also measured expression of the members (FDXR, FDX, POR) of their electron transport chains (ETC). We measured antioxidant enzymes (GPXs 1-8, CAT, SODs 1 and 2, PRDXs 1-6, GSR, TXN, TXNRDs 1-3). Since selenium is an active component of GPXs, the selenium-uptake receptors (LRPs 2 and 8) were measured. Only the selenium-dependent GPX1 showed the same increase in expression as the steroidogenic enzymes did with increasing follicle size. GPX4 and PRDX2/6 decreased with follicle size, whereas SOD1/2, CAT, GSR, and TXNRD3 were lowest at the intermediate sizes. The other antioxidant enzymes were unchanged or expressed at low levels. The expression of the selenium-uptake receptor LRP8 also increased significantly with follicle size. Correlation analysis revealed statistically significant and strongly positive correlations of the steroidogenic enzymes and their ETCs with both GPX1 and LRP8. These results demonstrate a relationship between expression of genes involved in steroidogenesis and selenium-containing antioxidant defence mechanisms. They suggest that during the late stages of folliculogenesis, granulosa cells are dependent on sufficient expression of GPX1 and the selenium transporter LRP8 to counteract increasing ROS levels caused by the production of steroid hormones.

Protective Effects of Allium senescens Extract against 6-Hydroxydopamine in Neurons

Oxidative neurodegeneration causes various neuronal diseases, including Alzheimer’s, Parkinson’s, and Huntington’s diseases. This study aimed to demonstrate the protective effect of Allium senescens leaf extracts on 6-hydroxydopamine (6-OHDA)-stressed SH-SY5Y cells, which are known to be optimal for neurotoxic research. The levels of apoptotic markers were evaluated using quantitative polymerase chain reaction (qPCR) and flow cytometry. The localization of apoptotic cells in vivo was analyzed using whole-mount immunochemistry and terminal deoxynucleotidyl transferase dUTP nick end labeling assay. Additionally, the production of reactive oxygen species (ROS) was estimated using flow cytometry. 6-OHDA induced ROS production in neuroblasts and in vivo, but treatment with the extract protected against the 6-OHDA-induced increase in ROS levels. Under oxidative stress, the extract performs three protective functions: decreasing ROS production, preventing mitochondrial apoptosis, and protecting the central and ventral nervous systems. These results also suggest that the extract can be useful in the development of functional foods for the prevention of neural damage caused by oxidative stress.

Heat stress in symbiotic dinoflagellates: Implications on oxidative stress and cellular changes

Global warming has been shown to harmfully affect symbiosis between Symbiodiniaceae and other marine invertebrates. When symbiotic dinoflagellates (the genus Breviolum) were in vitro exposed to acute heat stress of +7 °C for a period of 5 days, the results revealed the negative impact on all physiological and other cellular parameters measured. Elevated temperatures resulted in a severe reduction in algal density of up to 9.5-fold, as well as pigment concentrations, indicating the status of the physiological stress and early signs of photo-bleaching. Reactive oxygen species (ROS) were increased in all heated dinoflagellate cells, while the antioxidant-reduced glutathione levels initially dropped on day one but increased under prolonged temperature stress. The cell viability parameters were reduced by 97 % over the heating period, with an increased proportion of apoptotic and necrotic cells. Autofluorescence (AF) for Cy5-PE 660-20 was reduced from 1.7-fold at day 1 to up to 50-fold drop at the end of heating time, indicating that the AF changes were highly sensitive to heat stress and that it could be an extremely sensitive tool for assessing the functionality of algal photosynthetic machinery. The addition of the drug 5-AZA-2′-deoxycytidine (5-AZA), which inhibits DNA methylation processes, was assessed in parallel and contributed to some alterations in algal cellular stress response. The presence of drug 5-AZA combined with the temperature stress had an additional impact on Symbiodiniaceae density and cell complexity, including the AF levels. These variations in cellular stress response under heat stress and compromised DNA methylation conditions may indicate the importance of this epigenetic mechanism for symbiotic dinoflagellate thermal tolerance adaptability over a longer period, which needs further exploration. Consequently, the increased ROS levels and changes in AF signals reported during ongoing heat stress in dinoflagellate cells could be used as early stress biomarkers in these microalgae and potentially other photosynthetic species.

Impaired cardiac mitochondrial bioenergetics contribute to loss of cardiac function, in a porcine multiple risk factor model

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Dutch CardioVascular Alliance (an initiative with support of the Dutch Heart Foundation) Grant 2020B008 RECONNEXT, and the German Center for Cardiovascular Research (DZHK; 81Z0600207) Background Heart failure with preserved ejection fraction lacks targeted therapies, due to insufficient understanding of pathogenesis. A multiple comorbidity swine model (MCS) was developed by exposure to three risk factors for six months including diabetes (streptozotocin), chronic kidney disease (renal embolization), and an unhealthy diet (high-fat, high salt diet). An additional risk factor ovariectomy was added on top of the MCS model (MCS+). MCS and MCS+ presented with left ventricular diastolic dysfunction, oxidative stress, coronary microvascular dysfunction and inflammation. Furthermore, myocardial oxygen consumption was increased given the same level of cardiac work (Fig. A), suggesting an impairment in myocardial mitochondrial function. Using a combined-omics approach, the purpose of this study is to unravel alterations in gene expression and proteome abundance regarding mitochondrial function in MCS swine. Methods 15 MCS, 4 MCS+ and 10 healthy control female swine were included in the study. Proteome analysis and single nuclei RNA sequencing, were performed on frozen MCS left ventricle myocardial samples. Cardiac mitochondrial function was measured by an O2k-FlouRespirometer ex vivo in MCS+ swine versus control. Results Proteome analysis showed a reduced abundance of proteins involved in branched-chain amino acid (BCAA) catabolism (Fig. B). Single nuclei RNA sequencing of a subgroup of the same animals demonstrated a downregulation of these genes specifically in cardiomyocyte subpopulations. BCAA can act as a source of ATP via oxidative phosphorylation and can modulate mitochondrial substrate utilisation, and impair mitochondrial function, resulting in increased mitochondrial production of reactive oxygen species (ROS). Mitochondrial function was measured in fresh myocardial tissue, using pyruvate/malate/glutamate as substrate. Low mitochondrial respiratory sensitivity to ADP was observed 26±2.5 in MCS+ vs 43.6±1.5 pmol/(s*mg tissue) in healthy control, reflecting low oxidative capacity. Moreover, the respiratory rate after uncoupling by FCCP was reduced in MCS+ versus healthy control (73.6±11.7 vs 118.4±15.6 pmol/(s*mg tissue). These findings are consistent with our data in the MCS swine and indicate impaired BCAA in the myocardium of animals with comorbidities, which is associated with increased ROS levels (8-isoprostane 12.9±0.8 pg/mg protein in MCS vs 10.3±0.5 in healthy animals) and impaired myocardial efficiency during exercise (Fig. A). Conclusion A combined omics approach suggested an alteration in cardiac mitochondria. Respiratory analyses showed impaired mitochondrial bioenergetics and increased oxidative stress, which can contribute to diastolic dysfunction.

Regulation of Betaine Homocysteine Methyltransferase by Liver Receptor Homolog-1 in the Methionine Cycle.

Betaine-homocysteine S-methyltransferase (BHMT) is one of the most abundant proteins in the liver and regulates homocysteine metabolism. However, the molecular mechanisms underlying Bhmt transcription have not yet been elucidated. This study aimed to assess the molecular mechanisms underlying Bhmt transcription and the effect of BHMT deficiency on metabolic functions in the liver mediated by liver receptor homolog-1 (LRH-1). During fasting, both Bhmt and Lrh-1 expression increased in the liver of Lrh-1f/f mice; however, Bhmt expression was decreased in LRH-1 liver specific knockout mice. Promoter activity analysis confirmed that LRH-1 binds to a specific site in the Bhmt promoter region. LRH-1 deficiency was associated with elevated production of reactive oxygen species (ROS), lipid peroxidation, and mitochondrial stress in hepatocytes, contributing to hepatic triglyceride (TG) accumulation. In conclusion, this study suggests that the absence of an LRH-1-mediated decrease in Bhmt expression promotes TG accumulation by increasing ROS levels and inducing mitochondrial stress. Therefore, LRH-1 deficiency not only leads to excess ROS production and mitochondrial stress in hepatocytes, but also disrupts the methionine cycle. Understanding these regulatory pathways may pave the way for novel therapeutic interventions against metabolic disorders associated with hepatic lipid accumulation.

Oxidative stress plays an important role in the central regulatory mechanism of orofacial hyperalgesia under low estrogen conditions

Hyperalgesia occurs in the orofacial region of rats when estrogen levels are low, although the specific mechanism needs to be investigated further. Furthermore, oxidative stress plays an important role in the transmission of pain signals. This study aimed to explore the role of oxidative stress in orofacial hyperalgesia under low estrogen conditions. We firstly found an imbalance between oxidative and antioxidant capacity within the spinal trigeminal subnucleus caudalis (SP5C) of rats after ovariectomy (OVX), resulting in oxidative stress and then a decrease in the orofacial pain threshold. To investigate the mechanism by which oxidative stress occurs, we used virus as a tool to silence or overexpress the excitatory amino acid transporter 3 (EAAT3) gene. Further investigation revealed that the regulation of glutathione (GSH) and reactive oxygen species (ROS) can be achieved by regulating EAAT3, which in turn impacts the occurrence of oxidative stress. In summary, our findings suggest that reduced expression of EAAT3 within the SP5C of rats in the low estrogen state may decrease GSH content and increase ROS levels, resulting in oxidative stress and ultimately lead to orofacial hyperalgesia. This suggests that antioxidants could be a potential therapeutic direction for orofacial hyperalgesia under low estrogen conditions, though more research is needed to understand its mechanism.

Inhibiting AGS Cancer Cell Proliferation through the Combined Application of Aucklandiae Radix and Hyperthermia: Investigating the Roles of Heat Shock Proteins and Reactive Oxygen Species.

Cancer is a major global health concern. To address this, the combination of traditional medicine and newly appreciated therapeutic modalities has been gaining considerable attention. This study explores the combined effects of Aucklandiae Radix (AR) and 43 °C hyperthermia (HT) on human gastric adenocarcinoma (AGS) cell proliferation and apoptosis. We investigated the synergistic effects of AR and HT on cell viability, apoptosis, cell cycle progression, and reactive oxygen species (ROS)-dependent mechanisms. Our findings suggest that the combined treatment led to a notable decrease in AGS cell viability and increased apoptosis. Furthermore, cell cycle arrest at the G2/M phase contributed to the inhibition of cancer cell proliferation. Notably, the roles of heat shock proteins (HSPs) were highlighted, particularly in the context of ROS regulation and the induction of apoptosis. Overexpression of HSPs was observed in cells subjected to HT, whereas their levels were markedly reduced following AR treatment. The suppression of HSPs and the subsequent increase in ROS levels appeared to contribute to the activation of apoptosis, suggesting a potential role for HSPs in the combined therapy's anti-cancer mechanisms. These findings provide valuable insights into the potential of integrating AR and HT in cancer and HSPs.