Intracellular Glutathione Content Research Articles

Synthesis and characterization of 7-diethylamino-4-Chloromethyl coumarin: Spectroscopic analysis, molecular docking, and anticancer activity on large intestine carcinoma cells

Cancer, characterized by uncontrolled cell growth and metastasis, poses a significant global health burden, ranking as a leading cause of mortality worldwide. Colorectal cancer (CRC) specifically accounts for a substantial portion of cancer cases, with increasing incidence projected over the coming decades. While conventional treatments exist, they often entail adverse effects and limited efficacy, driving interest in natural remedies like coumarin derivatives due to their diverse biological activities and perceived safety profile. This study focuses on the synthesis and characterization of a novel compound, 7-diethylamino-4-chloromethyl coumarin (referred to as 7D4C), derived from coumarin. Structural elucidation employed Fourier transform infrared spectroscopy (FT-IR), proton and carbon-13 nuclear magnetic resonance spectroscopy (1H and 13C NMR), and mass spectrometry (MALDI-TOF-MS). Molecular docking studies were conducted to explore potential biological interactions. Furthermore, the anti-cancer potential of 7D4C was assessed using human epithelial adenocarcinom (LoVo) and healthy fibroblast (CCD-18Co) cell lines. Viability analysis, comet assay for DNA damage, and evaluation of cancer biomarkers including apoptosis, intracellular reactive oxygen species (iROS) levels, mitochondrial membrane potential (MMP), intracellular glutathione (GSH) concentration, and intracellular calcium (iCa2+) levels were performed. The synthesis of 7D4C was successfully completed, and its structure was confirmed. Molecular docking results indicate that 7D4C exhibits strong binding affinity to the p53 protein, highlighting its potential as a novel modulator of p53 activity. Subsequent investigations revealed that the synthesized compound induced apoptosis in cancer cells by reducing MMP and triggering DNA damage through the production of iROS. The promising anti-cancer activity of 7D4C in the LoVo cell line highlights its importance in coumarin-based therapies. Introducing 7D4C could significantly enhance future research in this area, leveraging insights from in vitro coumarin studies.

Paclitaxel Prodrug Enables Glutathione Depletion to Boost Cancer Treatment.

Herein, we constructed a paclitaxel (PTX) prodrug (PA) by conjugating PTX with acrylic acid as a cysteine-depleting agent. The as-synthesized PA can assemble with diacylphosphatidylethanolamine-PEG2000 to form stable nanoparticles (PA NPs). After endocytosis into cells, PA NPs can specifically react with cysteine and trigger release of PTX for chemotherapy. On the other hand, the depletion of cysteine can greatly downregulate the intracellular content of glutathione and lead to oxidative stress outburst-provoking ferroptosis. The released PTX can elicit antitumor immune response by inducing immunogenic cell death, thus promoting dendritic cells maturation and cascaded cytotoxic T lymphocytes activation, which not only produces a robust immunotherapy effect but also synergizes the ferroptosis therapy by inhibiting cysteine transport via the release of interferon-γ in the activated immune system. As a result, PA NPs exhibit favorable in vitro and in vivo antitumor performance with reduced systemic toxicity. Our work highlights the potential of simple molecular design of prodrugs for enhancing the therapeutic efficacy toward malignant cancer.

Trehalose enhances the resistance of Lactiplantibacillus plantarum LIP-1 to spray- and freeze-drying treatments by regulating amino acid metabolism

The effect of adding trehalose in the culture medium on Lactiplantibacillus plantarum LIP-1 was investigated during spray- and freeze-drying. Although previous studies reported that the enhancement of bacterial resistance is due to intracellular trehalose accumulation, our study revealed that it was related to trehalose metabolism by the strain, and not accumulation. The underlying mechanisms were mainly related to the activation of l-glutamic acid and cysteine metabolic pathways during trehalose metabolism. First, murL, murD, and vanY in the l-glutamic acid metabolic pathway were significantly upregulated, increasing peptidoglycan synthesis and thus improving the cell wall integrity during drying process. Second, gshA in the cysteine metabolic pathway was upregulated, increasing intracellular glutathione content, enhancing antioxidant capacity, and thereby reducing the degree of cell membrane damage during drying process. Third, alkaline substances produced during l-glutamic acid and cysteine metabolisms increased the intracellular pH and reduced the degree of DNA damage caused by acid stress.

Exploration of beauvericin's toxic effects and mechanisms in human astrocytes and N-acetylcysteine's protective role

Beauvericin (BEA) is a newly identified mycotoxin produced by various Fusarium species, and its contamination in food and animal feed is widespread globally. This mycotoxin demonstrates cytotoxic effects by inducing oxidative stress in multiple models. Furthermore, evidence indicates that BEA possesses diverse toxic activities, making it a promising candidate for toxicological research. Recent studies have highlighted the ability of BEA to traverse the blood-brain barrier, suggesting its potential neurotoxicity. However, limited information is available regarding the neurotoxic effects of BEA on human astrocytes. Therefore, this study aimed to assess the neurotoxic effects of BEA on the Gibco® Human Astrocyte (GHA) cell line and elucidate the underlying mechanisms. Additionally, the study aimed to investigate the protective effects of the antioxidant N-acetylcysteine (NAC) against BEA-induced toxicity. The data show that exposure to BEA within the 2.5–15 μM concentration range resulted in concentration-dependent cytotoxicity. BEA-treated cells exhibited significantly increased levels of reactive oxygen species (ROS), while intracellular glutathione (GSH) content was significantly reduced. Western blot analysis of cells treated with BEA revealed altered protein levels of Bax, cleaved caspase-9, and caspase-3, along with an increased Bax/Bcl-2 ratio, indicating the induction of apoptosis. Additionally, BEA exposure triggered antioxidant responses, as evidenced by increased protein expression of Nrf2, HO-1, and NQO1. Significantly, pretreatment with NAC partially attenuated the significant toxic effects of BEA. In conclusion, our findings suggest that BEA-induced cytotoxicity in GHA cells involves oxidative stress-associated apoptosis. Furthermore, NAC demonstrates potential as a protective agent against BEA-induced oxidative damage.

Nobiletin as a novel agent to enhance porcine in vitro embryo development and quality

In vitro embryo production (IVP) is of great importance to the porcine industry, as well as for basic research and biomedical applications. Despite the large efforts made in laboratories worldwide to address suboptimal culture conditions, porcine IVP remains inefficient. Nobiletin (Nob, 5,6,7,8,3′,4′ hexamethoxyflavone) supplementation to in vitro culture (IVC) medium, enhances in vitro embryo development in various species. However, its impact on the quality and developmental capacity of in vitro-produced pig embryos is yet to be established. This study evaluated the effects of different concentrations (2.5 and 5 μM) of Nob during the early culture of in vitro-produced pig embryos on embryo developmental competence, mitochondrial activity, lipid content, intracellular Reactive Oxygen Species (ROS) and Glutathione (GSH) content, Total Cell Number (TCN) per blastocyst, and expression of genes related to embryo development, quality and oxidative stress. Embryos cultured in medium without Nob supplementation and in medium supplemented with 0.01 % dimethyl sulfoxide (DMSO-vehicle for Nob) constituted the Control and DMSO groups, respectively. Embryo development rates were evaluated on Days 2, 6 and 7 of IVC. Additionally, a representative group of embryos was selected to assess mitochondrial activity, lipid, ROS and GSH content (on Days 2 and 6 of IVC), TCN assessment and gene expression analyses (on Day 6 of IVC). No significant differences were observed in any of the parameters evaluated on Day 2 of IVC. In contrast, embryos cultured under the presence of Nob 2.5 showed higher developmental rates on Days 6 and 7 of IVC. In addition, Day 6 embryos showed increased mitochondrial activity, with decreased levels of ROS and GSH in the Nob 2.5 group compared to the other groups. Both Nob 2.5 and Nob 5 embryos showed higher TCN compared to the Control and DMSO groups. Furthermore, Nob 2.5 and Nob 5 upregulated the expression of Superoxide dismutase type 1 (SOD1) and Glucose-6-phosphate dehydrogenase (G6PDH) genes, which could help to counteract oxidative stress during IVC. In conclusion, the addition of Nob during the first 48 h of IVC increased porcine embryo development rates and enhanced their quality, including the upregulation of relevant genes that potentially improved the overall efficiency of the IVP system.

A Smooth Muscle Cell-Based Ferroptosis Model to Evaluate Iron-Chelating Molecules for Cardiovascular Disease Treatment.

Dysregulation of iron homeostasis causes iron-mediated cell death, recently described as ferroptosis. Ferroptosis is reported in many chronic diseases, such as hepatic cancer, renal, and cardiovascular diseases (heart failure, atherosclerosis). However, there is a notable scarcity of research studies in the existing literature that explore treatments capable of preventing ferroptosis. Additionally, as far as the author is aware, there is currently no established model for studying ferroptosis within cardiovascular cells, which would be essential for assessing metal-chelating molecules with the potential ability to inhibit ferroptosis and their application in the treatment of cardiovascular diseases. In this study, a smooth muscle cell-based ferroptosis model is developed upon the inhibition of the system Xc- transporter by erastin associated or not with Fe(III) overload, and its rescue upon the introduction of well-known iron chelators, deferoxamine and deferiprone. We showed that erastin alone decreased the intracellular concentration of glutathione (GSH) without affecting peroxidized lipid concentrations. Erastin with ferric citrate was able to decrease intracellular GSH and induce lipid peroxidation after overnight incubation. Only deferiprone was able to rescue the cells from ferroptosis by decreasing lipid peroxidation via iron ion chelation in a 3:1 molar ratio.

Effect of glutathione-transport-related gene gsiD on desiccation tolerance of Cronobacter sakazakii and its related regulatory mechanism.

The outstanding desiccation tolerance of Cronobacter sakazakii (C. sakazakii) enables long-term persistence in food products with low-water activity to increase the infection risk, especially in low-birth-weight, immuno-compromised neonates, and infants less than 4 weeks of age. In our previous study, the disruption of glutathione transport-related gene gsiD by transposon was found to significantly increase its inactivation rate under drying stress challenges. However, the mechanism underlying the association between glutathione transport and desiccation tolerance of C. sakazakii remains to be clarified. In this study, the mechanism underlying their association was investigated in detail by constructing the gsiD gene deletion mutant. gsiD gene deletion was found to cause the dysfunction of the glutathione transport system GsiABCD and the limitation of glutathione import. The resulting decrease in intracellular glutathione caused the decreased potassium ions uptake and increased potassium ions efflux, inhibited the proline synthesis process, limited extracellular glutathione utilization, increased oxidant stress, reduced biofilm formation, and increased outer membrane permeability, which may be the main reasons for the significant reduction of the desiccation tolerance of C. sakazakii.IMPORTANCEContributing to its superior environmental adaptability, Cronobacter sakazakii can survive under many abiotic stress conditions. The outstanding desiccation tolerance makes this species persist in low-water activity foods, which increases harm to humans. For decades, many studies have focused on the desiccation tolerance of C. sakazakii, but the existing research is still insufficient. Our study found that gsiD gene deletion inhibited glutathione uptake and further decreased intracellular glutathione content, causing a decrease in desiccation tolerance and biofilm formation and an increase in outer membrane permeability. Moreover, the expression level of relative genes verified that gsiD gene deletion made the mutant not conducive to surviving in dry conditions due to restricting potassium ions uptake and efflux, inhibiting the conversion of glutamate to compatible solute proline, and increasing the oxidative stress of C. sakazakii. The above results enrich our knowledge of the desiccation tolerance mechanism of C. sakazakii.

Multienzyme Active Manganese Oxide Alleviates Acute Liver Injury by Mimicking Redox Regulatory System and Inhibiting Ferroptosis.

Drug-induced liver injury (DILI) is a severe condition characterized by impaired liver function and the excessive activation of ferroptosis. Unfortunately, there are limited options currently available for preventing or treating DILI. In this study, MnO2 nanoflowers (MnO2Nfs) with remarkable capabilities of mimicking essential antioxidant enzymes, including catalase, superoxide dismutase (SOD), and glutathione peroxidase are successfully synthesized, and SOD is the dominant enzyme among them by density functional theory. Notably, MnO2Nfs demonstrate high efficiency in effectively eliminating diverse reactive oxygen species (ROS) such as hydrogen peroxide (H2O2), superoxide anion (O2 •-), and hydroxyl radical (•OH). Through in vitro experiments, it is demonstrated that MnO2Nfs significantly enhance the recovery of intracellular glutathione content, acting as a potent inhibitor of ferroptosis even in the presence of ferroptosis activators. Moreover, MnO2Nfs exhibit excellent liver accumulation properties, providing robust protection against oxidative damage. Specifically, they attenuate acetaminophen-induced ferroptosis by inhibiting ferritinophagy and activating the P62-NRF2-GPX4 antioxidation signaling pathways. These findings highlight the remarkable ROS scavenging ability of MnO2Nfs and hold great promise as an innovative and potential clinical therapy for DILI and other ROS-related liver diseases.

Bmo-miR-3351 modulates glutathione content and inhibits BmNPV proliferation by targeting BmGSTe6 in Bombyx mori.

MicroRNAs (miRNAs) are small non-coding RNAs that play pivotal roles in the host response to invading pathogens. Among these pathogens, Bombyx mori nucleopolyhedrovirus (BmNPV) is one of the main causes of substantial economic losses in sericulture, and there are relatively few studies on the specific functions of miRNAs in the B. mori-BmNPV interaction. Therefore, we conducted transcriptome sequencing to identify differentially expressed (DE) messenger RNAs (mRNAs) and miRNAs in the midgut of 2 B. mori strains (BmNPV-susceptible strain P50 and BmNPV-resistant strain A35) after BmNPV infection. Through correlation analysis of the miRNA and mRNA data, we identified a comprehensive set of 21 miRNAs and 37 predicted target mRNAs. Notably, miR-3351, which has high expression in A35, exhibited remarkable efficacy in suppressing BmNPV proliferation. Additionally, we confirmed that miR-3351 binds to the 3' untranslated region (3' UTR) of B. mori glutathione S-transferase epsilon 6 (BmGSTe6), resulting in its downregulation. Conversely, BmGSTe6 displayed an opposite expression pattern to miR-3351, effectively promoting BmNPV proliferation. Notably, BmGSTe6 levels were positively correlated with glutathione S-transferase activity, consequently influencing intracellular glutathione content in the infected samples. Furthermore, our investigation revealed the protective role of glutathione against BmNPV infection in BmN cells. In summary, miR-3351 modulates glutathione content by downregulating BmGSTe6 to inhibit BmNPV proliferation in B. mori. Our findings enriched the research on the role of B. mori miRNAs in the defense against BmNPV infection, and suggests that the antiviral molecule, glutathione, offers a novel perspective on preventing viral infection in sericulture.

Effect of in vitro supplementation of branched-chain amino acids on glutathione peroxidase enzyme activity and intracellular glutathione concentration in CACO-2 cells

Effect of in vitro supplementation of branched-chain amino acids on glutathione peroxidase enzyme activity and intracellular glutathione concentration in CACO-2 cells

Autonomous metabolic reprogramming and oxidative stress characterize endothelial dysfunction in acute myocardial infarction.

Compelling evidence has accumulated on the role of oxidative stress on the endothelial cell (EC) dysfunction in acute coronary syndrome. Unveiling the underlying metabolic determinants has been hampered by the scarcity of appropriate cell models to address cell-autonomous mechanisms of EC dysfunction. We have generated endothelial cells derived from thrombectomy specimens from patients affected with acute myocardial infarction (AMI) and conducted phenotypical and metabolic characterizations. AMI-derived endothelial cells (AMIECs) display impaired growth, migration, and tubulogenesis. Metabolically, AMIECs displayed augmented ROS and glutathione intracellular content, with a diminished glucose consumption coupled to high lactate production. In AMIECs, while PFKFB3 protein levels of were downregulated, PFKFB4 levels were upregulated, suggesting a shunting of glycolysis towards the pentose phosphate pathway, supported by upregulation of G6PD. Furthermore, the glutaminolytic enzyme GLS was upregulated in AMIECs, providing an explanation for the increase in glutathione content. Finally, AMIECs displayed a significantly higher mitochondrial membrane potential than control ECs, which, together with high ROS levels, suggests a coupled mitochondrial activity. We suggest that high mitochondrial proton coupling underlies the high production of ROS, balanced by PPP- and glutaminolysis-driven synthesis of glutathione, as a primary, cell-autonomous abnormality driving EC dysfunction in AMI.

Macrophage-hitchhiking interleukin-10 plasmid DNA delivery system modulates rheumatoid arthritis microenvironment via the re-polarization of macrophages

Rheumatoid arthritis (RA) is a common chronic inflammatory disease and the current treatments for RA are unsatisfactory due to the buffer barrier from the RA microenvironment. Interleukin-10 (IL-10) was found to be able to modulate rheumatoid arthritis microenvironment (RAM) by the metabolic reprogramming of macrophages. However, its clinical application has proven challenging mainly due to its short half-life, non-ideal targeting efficiency via systemic administration and poor tissue penetration. Taking advantage of the recruitment of macrophages, we constructed an innovative macrophage-hitchhiking IL-10 pDNA delivery system using the peptides bearing nuclear localization signal (NLS) and tuftsin-modified disulfide-crosslinked polymers (bPEI-SS-PEG-T) which was responsive to the intracellular glutathione concentration. Firstly, we formed NLS/DNA complexes via electrostatic interactions to enhance the nuclear entry efficiency. Then NLS/DNA complexes were combined with bPEI-SS-PEG-T to construct bPEI-SS-PEG-T/NLS/DNA NPs with sizes of 168.4 nm and zeta potentials of + 10.9 mV. As the bPEI-SS-PEG-T was biodegradable and the tuftsin peptides enhanced macrophage-mediated phagocytosis, the nanoparticles exhibited lower cytotoxicity and better cellular uptake in addition to higher nuclear entry efficiency, leading to better IL-10 plasmid transfection. In addition, these macrophage-hitchhiking nanoparticles could effectively accumulate at inflammatory sites of RA via intra-peritoneal injection due to the macrophage migration to the inflammatory joints and alleviate symptoms of inflammation in vivo by inducing the re-polarization of macrophages, which might be mediated by the inhibition of mammalian target of rapamycin (mTOR) and the induction of arginase-2 (Arg2). As a result, this macrophage-hitchhiking gene delivery system demonstrated excellent inflammation targeting ability, good nuclear entry ability, high IL-10 plasmid transfection efficiency and great therapeutic efficacy in vivo, which is promising for the treatment of rheumatoid arthritis. Besides, this delivery system also provides a novel strategy for the gene therapy and the gene delivery system design for rheumatoid arthritis and other similar inflammatory diseases from the perspective of the inflammatory microenvironment.

Role of Glutathione in Neutrophil Chemotaxis in Periodontitis

Periodontitis is a common non-communicable inflammatory disease that leads to the destruction of periodontal tissues and tooth loss. Initiated by the plaque biofilm, there is a strong innate immune response with an abundance of neutrophils in the periodontium of affected individuals. Previous reports have shown that the intracellular concentration of glutathione in peripheral blood neutrophils from periodontitis patients and the chemotactic ability of these cells are compromised. Furthermore, other studies have described that in oxidative stress conditions neutrophil chemotaxis is aberrant and causes the glutathionylation of F-actin, a key player in chemotaxis. In this study, the effects of glutathione-modulating compounds were assessed in neutrophils isolated from healthy donors, showing that the perturbation of glutathione homeostasis decreases the chemotaxis of neutrophils. Following this, the intracellular glutathione status and chemotactic ability of neutrophils isolated from periodontitis patients was compared to that of age- and sex-matched controls. A decrease in glutathione and chemotactic ability were confirmed. Finally, the proteome of these neutrophils was explored, demonstrating a change in the abundance of proteins involved in glutathione homeostasis. Together these data suggest that peripheral blood neutrophils from periodontitis patients are compromised in their ability to cope with oxidative stress and move.

MRP1-Dependent Extracellular Release of Glutathione Induces Cardiomyocyte Ferroptosis After Ischemia-Reperfusion.

The membrane components of cardiomyocytes are rich in polyunsaturated fatty acids, which are easily oxidized. Thus, an efficient glutathione-based lipid redox system is essential for maintaining cellular functions. However, the relationship between disruption of the redox system during ischemia-reperfusion (IR), oxidized lipid production, and consequent cell death (ferroptosis) remains unclear. We investigated the mechanisms underlying the disruption of the glutathione-mediated reduction system related to ferroptosis during IR and developed intervention strategies to suppress ferroptosis. In vivo fluctuations of both intra- and extracellular metabolite levels during IR were explored via microdialysis and tissue metabolome analysis. Oxidized phosphatidylcholines were assessed using liquid chromatography high-resolution mass spectrometry. The areas at risk following IR were assessed using triphenyl-tetrazolium chloride/Evans blue stain. Metabolomic analysis combined with microdialysis revealed a significant release of glutathione from the ischemic region into extracellular spaces during ischemia and after reperfusion. The release of glutathione into extracellular spaces and a concomitant decrease in intracellular glutathione concentrations were also observed during anoxia-reperfusion in an in vitro cardiomyocyte model. This extracellular glutathione release was prevented by chemical inhibition or genetic suppression of glutathione transporters, mainly MRP1 (multidrug resistance protein 1). Treatment with MRP1 inhibitor reduced the intracellular reactive oxygen species levels and lipid peroxidation, thereby inhibiting cell death. Subsequent in vivo evaluation of endogenously oxidized phospholipids following IR demonstrated the involvement of ferroptosis, as levels of multiple oxidized phosphatidylcholines were significantly elevated in the ischemic region 12 hours after reperfusion. Inhibition of the MRP1 transporter also alleviated intracellular glutathione depletion in vivo and significantly reduced the generation of oxidized phosphatidylcholines. Administration of MRP1 inhibitors significantly attenuated infarct size after IR injury. Glutathione was released continuously during IR, primarily in an MRP1-dependent manner, and induced ferroptosis. Suppression of glutathione release attenuated ferroptosis and reduced myocardial infarct size following IR.

A redox-responsive nanosystem to suppress chemoresistant lung cancer through targeting STAT3

Cancer stem cells (CSCs) have been demonstrated to be involved in tumor initiation and relapse, and the presence of CSCs in the tumor tissue often leads to therapeutic failure. BBI608 has been identified to eliminate CSCs by inhibiting signal transducer and activator of transcription 3 (STAT3). In this study, we confirm that BBI608 can efficiently suppress the proliferation and migration of non-small cell lung cancer (NSCLC) cells, and specifically kill the stemness-high population in chemoresistant NSCLC cells. To improve its bioavailability and tumor accumulation, BBI608 is successfully encapsulated into redox-responsive PEGylated branched N-(2-hydroxypropyl) methacrylamide (HPMA)-deoxy cholic acid (DA) polymeric nanoparticles (BBI608-SS-NPs). The BBI608-SS-NPs can release the drug in response to high concentrations of intracellular glutathione, and exhibit cytotoxicity against lung cancer cells and CSCs comparable to the free drug BBI608. Furthermore, the BBI608-SS-NPs preferentially accumulate in tumor sites, resulting in a superior anti-tumor efficacy in both cisplatin-resistant cell line-derived xenograft (CDX) and patient-derived xenograft (PDX) models of NSCLC. Mechanistic studies demonstrate that BBI608-SS-NPs not only directly inhibit the downstream genes of the STAT3 pathway, but also indirectly inhibit the Wnt pathway. Overall, this stimuli-responsive polymeric nanoformulation of BBI608 shows great potential in the treatment of chemoresistant NSCLC by targeting CSCs.

Signaling pathways upregulating glutathione‑specific γ‑glutamylcyclotransferase 1 by 3‑(5'‑hydroxymethyl‑2'‑furyl)‑1‑benzylindazole.

Glutathione‑specific γ‑glutamylcyclotransferase1 (CHAC1), is an unfolded protein response‑induced gene. Although it has been previously reported that CHAC1 transcription is regulated by activating transcription factor (ATF)4, ATF3 and CCAAT/enhancer‑binding protein β (C/EBPβ), the signaling pathways that regulate CHAC1 are largely unknown. It was revealed that 3‑(5'‑hydroxymethyl‑2'‑furyl)‑1‑benzylindazole (YC‑1; PubChem ID: 5712), a nitric oxide‑independent activator ofsoluble guanylyl cyclase (sGC), increases CHAC1 levels in cultured human kidney proximal tubular cells (HK‑2). Therefore, in the present study, the signaling pathways that induce CHAC1 by YC‑1 were investigated in HK‑2 cells. YC‑1 induced CHAC1 expression in a dose‑ and time‑dependent manner. KT5823, an inhibitor of cGMP‑dependent protein kinase (PKG), partially inhibited CHAC1 upregulation, indicating that the sGC‑cGMP‑PKG pathway participates in CHAC1 regulation. These results also suggested that other signaling pathways are involved in the regulation of CHAC1. Since antibody array analysis showed the activation of p38, mTOR and Akt, the involvement of these factors was further investigated. Although LY294002 and KU0063794 (inhibitors of Akt and mTOR, respectively) inhibited YC‑1‑induced CHAC1 expression, SB203580 (an inhibitor of p38) did not. These results indicated that CHAC1 is regulated by the Akt‑mTOR pathway. In addition, YC‑1 induced endoplasmic reticulum (ER) stress, a regulator of CHAC1 induction. These findings suggested that CHAC1 is regulated by YC‑1 through the sGC‑cGMP‑PKG, Akt‑mTOR and ER stress pathways. The present study demonstrated that CHAC1 induction reduced the intracellular glutathione concentration, indicating that CHAC1 plays an important role in intracellular redox homeostasis in tubular cells.

Intracellular delivery of anticancer agents using dual responsive nanomicelles synthesized via RAFT polymerization

In this study, diselenide core crosslinked nanomicelles were developed for the delivery of docetaxel (DTX) to tumor cells, aiming to achieve synergistic antitumor chemotherapy in combination with radiotherapy.The nanomicelles were composed of a random triblock copolymer, Poly(poly(ethylene glycol) methyl ether methacrylate-co-N-isopropylacrylamide-co-Acrylic acid N-hydroxysuccinimide ester), synthesized via reversible addition-fragmentation chain transfer (RAFT) polymerization, and crosslinked using selenocystamine dihydrochloride. Physicochemical experiments were conducted to characterize the polymer synthesis and nanoparticles. The in vitro release profile was studied in response to redox conditions. In vitro cytotoxicity studies were performed on the breast cancer (MCF-7) cell line.The results showed the formation of 81 ± 5.2 nm spherical nanomicelles with a drug loading and encapsulation efficiency (EE) of 8.09% (w/w) and 97% (w/w), respectively. The nanomicelles exhibited dual responsive controlled release profiles, with approximately 80% of the drug being released within the first 2 h in response to intracellular glutathione (GSH) concentration and X-ray exposure. Fluorescent microscopy confirmed the effective internalization of the nanomicelles. Furthermore, the nanomicelles demonstrated increased cytotoxicity and apoptosis rates. The DTX-loaded nanomicelles exhibited a lower IC50 value (0.037 µg/ml) compared to free DTX (0.092 µg/ml) (P < 0.05). The combination of X-ray irradiation (2 Gy) and DTX release exhibited the highest cytotoxicity on MCF-7 cells (IC50 = 0.019 µg/ml).Overall, the nanomicelles prepared in this study, in combination with X-ray irradiation, demonstrated enhanced antitumor efficacy of the system.

Glutathione-Responsive Methotrexate Polymersomes for Potential Management of Ectopic Pregnancy.

The first-line treatment for ectopic pregnancy (EP), the chemotherapeutic methotrexate (MTX), has a failure rate of more than 10%, which can lead to severe complications or death. Inadequate accumulation of administered MTX at the ectopic implantation site significantly contributes to therapeutic failure. This study reports the first glutathione-responsive polymersomes for efficient delivery of MTX to the implantation site and its triggered release in placental cells. Fluorescence and photoacoustic imaging have confirmed that the developed polymersomes preferentially accumulate after systemic administration in the implantation site of pregnant mice at early gestational stages. The high concentrations of intracellular glutathione (GSH) reduce an incorporated disulfide bond within polymersomes upon internalization into placental cells, resulting in their disintegration and efficient drug release. Consequently, MTX delivered by polymersomes induces pregnancy demise in mice, as opposed to free MTX at the same dose regimen. To achieve the same therapeutic efficacy with free MTX, a sixfold increase in dosage is required. In addition, mice successfully conceive and birth healthy pups following a prior complete pregnancy demise induced by methotrexate polymersomes. Therefore, the developed MTX nanomedicine can potentially improve EP management and reduce associated mortality rates and related cost.

Exosomes derived from M2-type microglia ameliorate oxygen-glucose deprivation/reoxygenation-induced HT22 cell injury by regulating miR-124-3p/NCOA4-mediated ferroptosis

BackgroundAlthough it has been reported that miRNA carried by M2 microglial exosomes protects neurons from ischemia–reperfusion brain injury, the mechanism of action remains poorly understood. This study aimed to explore the miRNA signaling pathway by which M2-type microglia-derived exosomes (M2-exosomes) ameliorate oxygen–glucose deprivation/reoxygenation (OGD/R)-induced cytotoxicity in HT22 cells. MethodsBV2 microglia were induced by M2 polarization. Then, M2-exosomes were identified via transmission electron microscopy and special biomarker detection and co-cultured with HT22 cells. Cell proliferation was evaluated using the Cell Counting Kit-8 (CCK-8) assay. Intracellular concentrations of reactive oxygen species (ROS), Fe2+, glutathione (GSH), and malondialdehyde (MDA) were determined using dichlorofluorescein fluorescence and biochemical determination. miR-124-3p levels were determined using qRT-PCR, and protein expressions were examined via western blotting. ResultsOGD/R suppressed the proliferation and induced the accumulation of Fe2+, ROS, and MDA and reduction of GSH in mouse HT22 cells, suggesting ferroptosis of HT22 cells. OGD/R-induced changes in the above mentioned indexes was ameliorated by M2-exosomes but restored by the exosome inhibitor GW4869. M2-exosomes with (mimic-exo) or without miR-124-3p (inhibitor-exo) promoted and suppressed proliferation and ferroptosis-associated indexes of HT22 cells, respectively. Moreover, mimic-exo and inhibitor-exo inhibited and enhanced NCOA4 expression in HT22 cells, respectively. NCOA4 overexpression reversed the protective effects of miR-124-3p mimic-exo in OGD/R-conditioned cells. NCOA4 was targeted and regulated by miR-124-3p. ConclusionsM2-exosome protects HT22 cells against OGD/R-induced ferroptosis injury by transferring miR-124-3p and NCOA4 into HT22 cells, with the latter being a target gene for miR-124-3p.

Перспективы применения N-ацетилцистеина в качестве нефропротектора у пациентов с сочетанной травмой и другие фармакологические аспекты клинического использования данного вещества

Summary. Objectives of the study were to analyze a wide range of published data; to study biological activity and bioavailability of N-acetylcysteine (N-AC) as well as a possibility of using the drug in clinical practice to eliminate oxidative stress and inflammatory reactions in pathological conditions and nosologies; to study its therapeutic effectiveness in renal dysfunction in patients with concomitant trauma. Study materials and methods. Materials of research: scientific publications on the specified problems. Research method: analytical. Results of the study and their analysis. A review of domestic and foreign literature on the role of N-acetylcysteine in correction of oxidative stress and protection against free radicals is presented. The great role of N-acetylcysteine for the replenishment of the intracellular content of glutathione, the main cellular antioxidant, as well as the potential possibility of using N-acetylcysteine in clinical practice for various pathologies and diseases is investigated. Review of the results of a large number of studies leads the authors to the conclusion that this pharmacological substance is a very promising means for replenishing the intracellular content of glutathione and can be used in the treatment of a number of diseases.