Intracellular ROS Levels Research Articles

Evaluation of the bactericidal activity of plasma-activated NaCl solution (PAN) against foodborne pathogens: Inactivation mechanism and application to mackerel

The objective of this study was to assess the bactericidal effect of plasma-activated NaCl solution (PAN) against Escherichia coli O157:H7, Salmonella Typhimurium, and Listeria monocytogenes and apply PAN as a brine salting solution for mackerel. To enhance the bactericidal effect of plasma-activated water (PAW), NaCl solutions (0, 3.5, 7, and 10 %) were treated with plasma for 20 and 40 min to generate PAN. Plasma-activated glycerol solution (PAG) was also included to evaluate the influence of water activity on plasma activation and its effect on microbial activity. Physicochemical analysis revealed that elevating the NaCl concentration of PAN led to a decrease in pH, an increase in oxidation–reduction potential, and higher levels of reactive species such as H2O2 and HOCl. PAN showed greater antibacterial activity compared to PAW and PAG, except for L. monocytogenes, where 40 min activation time and treatment time exceeding 20 min was required for significantly higher reduction to occur. PAN exhibited greater antibacterial activity at higher NaCl concentrations, which was attributed to increased ionic strength and reactive chlorine species. Additionally, we evaluated the microbial mechanisms of PAN by assessing cellular damage and alterations. The common observation across the three pathogens was that PAN resulted in increased cell membrane damage, reduced intermembrane enzyme activity, higher intracellular ROS levels, and changes in zeta potential values, while DNA damage was observed only in PAN-treated L. monocytogenes. Furthermore, when PAW and PAN were stored for up to four weeks, PAN showed higher efficacy compared to PAW. 10 % PAN was also effective against foodborne pathogens on mackerel, achieving log reductions of 3.62 for E. coli O157:H7, 4.62 for S. Typhimurium, and 3.18 for L. monocytogenes after a 20 min treatment without adversely affecting quality. Our results demonstrated the antibacterial activity and action mechanism of PAN, presenting its potential application in the seafood industry.

ESTRADIOL IMPACTS MÜLLER GLIA AND ENDOTHELIAL CELL RESPONSES IN HYPERGLYCEMIC MICROENVIRONMENTS WITH ADVANCED GLYCATION END PRODUCTS

Diabetic retinopathy is a leading cause of vision loss in working adults, with disproportionate impact on women with lowered estrogen. Sex hormones and their receptors are significant to neuroprotection of the inner blood-retinal barrier (iBRB), a tissue that regulates transport across the neuroretina and vasculature. Moreover, high glucose levels in diabetes lead to the formation of advanced glycation end products (AGEs), which promote inflammation and iBRB breakdown to result in vision loss. This study examined the effects of supplemental estradiol on cell reactivity and cell barrier resistance within an in vitro model of hyperglycemia. Changes in morphology and expression of reactive oxygen species were examined when cells were exposed to a hyperglycemic medium containing AGEs, with and without supplemental estradiol. Cell morphology was assessed via changes in cell area and cell shape index, while intracellular ROS levels were measured using a ROS-sensitive dye. In addition, trans endothelial resistance (TEER) assays were used to measure changes in cell barrier function in response to hyperglycemic conditions, with and without supplemental estradiol. Results show that ROS levels in Müller glia in hyperglycemic conditions significantly decreased in response to supplemental estradiol. The estradiol further increased the resistivity of Müller glia and endothelial cell barriers cultured in high glucose and AGEs. This project illustrates the restorative effects of estradiol in collective responses of cell barriers formed by endothelial cells and Müller glia.

Design, Synthesis, and Antifungal Evaluation of Novel Cuminic Acid Derivatives as Potential Laccase Inhibitors.

In search of novel natural product-based fungicides, 49 cuminic acid derivatives were designed, synthesized, and screened for their in vitro antifungal effects toward seven phytopathogenic fungi and oomycetes. Consequently, several derivatives exhibited strong antifungal activities toward Fusarium graminearum, Botryosphaeria dothidea, and Valsa mali. Among them, compound 2b exhibited the most potent antifungal activity toward B. dothidea (EC50 = 0.96 mg/L), more powerful than chlorothalonil. The in vivo assay against B. dothidea found that the protective and curative effects of 2b were comparable to chlorothalonil. Meanwhile, SEM and TEM observations indicated that 2b could ruin the integrity of mycelial morphology and organelles of B. dothidea. Preliminary mechanism research showed that 2b increased the cell membrane permeability and intracellular ROS level, as well as conspicuously decreased the mycelial dry weight and cell wall chitin contents of B. dothidea. The phytotoxicity test revealed that 2b showed good safety on seeds of mung bean and radish. The in vitro laccase inhibitory activity assay and molecular docking study demonstrated that 2b could be a promising laccase inhibitor. This type of cuminic acid hydrazide derivative would provide valuable inspiration for developing novel fungicides against B. dothidea.

An amphipathic peptide combats multidrug-resistant Staphylococcus aureus and biofilms

The emergence of drug-resistant Staphylococcus aureus (S. aureus) has resulted in infections in humans and animals that may lead to a crisis in the absence of highly effective drugs. Consequently, the development of alternative or complementary antimicrobial agents is urgently needed. Here, a series of peptides derived from AP138 were designed with high expression, antimicrobial activity, and antibiofilm properties via bioinformatics. Among them, the best derived peptide, A24 (S9A), demonstrated the greatest stability and bactericidal efficiency against multidrug-resistant S. aureus in a physiological environment, with a high hydrophobicity of 35%. This peptide exhibited superior performance compared to the preclinical or clinical antimicrobial peptides (AMPs). A24 displayed increased biocompatibility in vitro and in vivo, exhibiting a low hemolysis rate (less than 3%), minimal cytotoxicity (survival rate exceeding 85%), and no histotoxicity. A24 had the capacity to destroy cell walls, increase cell membrane permeability, and induce increases in intracellular ATP and ROS levels, which resulted in the rapid death of S. aureus. A24 inhibited the formation of early biofilms and eliminated both mature biofilms (40–50%) and persisters (99.9%). Therapeutic doses of A24 were shown to exhibit favorable safety profiles and bactericidal efficacy in vivo and could reduce bacterial loads of multidrug-resistant S. aureus by 4–5 log10 CFU/0.1g levels in mouse peritonitis and endometritis models. Furthermore, A24 increased the survival rate to 100% and exhibited anti-inflammatory properties in a mouse model. The aforementioned data illustrate the potential of A24 as a pharmaceutical agent for the treatment of bacterial infections, including peritonitis and endometritis, in animal husbandry with multidrug-resistant S. aureus infections.

Design, Synthesis and Bioactivity of Benzyl Propiolates with Broad-Spectrum Inhibition Activity on Phytopathogenic Fungi.

As part of our continuing research on propiolic acid derivatives, a series of benzyl propiolate derivatives and analogues were designed, synthesized, and investigated for inhibition activity in vitro and in vivo on phytopathogenic fungi, structure-activity relationship (SAR) and action mechanism. The results showed that most of the compounds had potent and broad-spectrum antifungal activity in vitro at 50 μg/mL. Among the compounds, 2-bromobenzyl propiolate (3h) displayed the highest comprehensive activity with total activity index (TAI) of 4.57 against all the eight test fungi followed by 3-fluorobenzyl propiolate (3c) (TAI = 4.53), superior to positive fungicides thiabendazole (TAI < 4.12) and/or azoxystrobin (TAI < 2.90). Relative to the other fungi, Fusarium solani and Fusarium graminearum revealed higher total susceptibility indexes (TSI) of 11.1 and 8.98, respectively, for all compounds. 3h and 3g (4-chlorobenzyl propiolate) gave the smallest EC50 values of 0.86 and 0.67 μg/mL against F. solani and F. graminearum, respectively, superior or comparable with thiabendazole and azoxystrobin. As a representative, 3h at 200 μg/mL displayed 100% protection on F. solani infections on potatoes over 7 days and high safety for plant growth. In antifungal mechanism, 3h was able to change mycelial morphology, destroy the structures of both hypha and cell membrane, increase the intracellular ROS level, decrease the intracellular mitochondrial membrane potential level, and change the permeability of the cell membrane. The SAR showed that ethynyl is a key group for the activity, and the presence of halogen atoms at the 2-site of the benzene ring can significantly improve the activity in most cases. Thus, benzyl propiolate can be considered a novel antifungal lead compound, while 3h can be considered a promising fungicide candidate to develop new agricultural fungicides.

Methylglyoxal impairs human dermal fibroblast survival and migration by altering RAGE-hTERT mRNA expression in vitro

Fibroblasts are native residents in dermal layer of human skin which are important for dermal regeneration and essential during cutaneous wound healing by releasing inflammatory markers and actively migrate to close an open wound. Premature skin ageing due to methylglyoxal (MGO) has recently caught the attention considering its potential to accelerate the emergence of skin ageing signs, however previous studies were only focused in primary neonatal dermal fibroblast and NIH3t3 fibroblast cell line. Therefore, thorough investigation is required to study the impact of MGO on primary human dermal fibroblast isolated from adult subject (HDFa). In our experiments, short exposure of MGO was observed to induced significant reductions in cell viability at concentrations of 7.5, 10, 12.5, 15, and 17.5 mM (p < 0.005) after 3 hours of treatment. The cellular death of HDFa at 10, 12.5 and 15 mM of MGO were also marked by increased in intracellular ROS level, indicating the involvement of oxidative stress-induced death in these cells. We also observed enlarge scratch areas of cells exposed with 7.5 and 10 mM MGO compared to control after 26 hours, thereby suggesting a decline in cell migration and viability in this group. We propose the increased ROS as the consequence of AGE-RAGE activation which was marked by significant elevation of RAGE mRNA on cells exposed to 10 mM MGO. Our data also suggest the occurrence of DNA damage events via ROS-induced oxidation or mediated by decline in hTERT mRNA expression.

Efficacy of Ergosterol and Ergosterol peroxide in different anti-inflammatory models – a comparison study

ABSTRACT Ergosterol and its peroxide with important regulatory roles in a variety of intracellular biological processes. In this study, both of the LPS- and UVA-induced human keratinocyte (HaCaT) inflammatory models were established to investigate the anti-inflammatory effects of ES and EP. The protective effects of EP and ES on UVA and LPS-induced cell damage were discussed by cell viability assays, reactive oxygen species levels, skin barrier-related proteins, anti-inflammatory factors, and expression of genes involved in related signaling pathways were analyzed. The results showed that both of ES and EP could reduce the intracellular ROS levels; inhibit IL-8 and IL-1β secretion; and had significant promotion effects on AQP3 and FLG, inhibited gene expressions of PI3K, AKt, IL-1β and IL-8 and ES and EP were not irritating according to the safety evaluation. In summary, both of ES and EP showed excellent anti-inflammatory effects and provided a degree of protection against UVA/LPS-induced inflammatory damage.

Chemical Composition of Extracts from Various Parts of Feverfew (Tanacetum parthenium L.) and Their Antioxidant, Protective, and Antimicrobial Activities.

Tanacetum parthenium is a medicinal plant from the Asteraceae family that can be applied externally in the case of various skin diseases. The aim of the study was to perform a phytochemical analysis of hydroethanolic extracts from the aerial parts (herb), flower heads, and leaves of feverfew and to assess their biological properties. Hydrodistilled oils were analyzed using GC-MS. The chemical composition of the extracts was estimated using spectrophotometry and the HPLC method. Moreover, the extracts were evaluated to determine their antioxidant potential using DPPH and FRAP and measuring the intracellular level of ROS. The cytotoxicity of extracts toward keratinocytes and fibroblasts was also analyzed, as well as their antimicrobial properties against 12 microorganisms. The results of the research revealed that chrysanthenone and α-thujone were the dominant volatile compounds in the essential oil from the flowers, while camphor, trans-chrysanthenyl acetate, and camphene were predominant in the essential oil from the leaves and herb. The results of HPLC showed that the major polyphenol compounds present in the hydroethanolic extracts from various parts of T. parthenium were 3,5-dicaffeoyl-quinic acid, chlorogenic acid, and 3,4-dicaffeoyl-quinic acid. The extract from feverfew flowers was shown to have the highest content of total polyphenols, flavonoids, and phenolic acids, as well as the highest antioxidant potential. In turn, the herb extract had the highest content of condensed tannins and terpenoids and exhibited the most effective antimicrobial properties against the 12 bacterial and fungal strains. Moreover, the hydroethanolic extracts from different parts of T. parthenium plants were shown to have a potent protective effect on skin cells. The present study supports the potential applications of Tanacetum parthenium in the cosmetic and pharmaceutical industries.

Activation of Hedgehog pathway by circEEF2/miR-625-5p/TRPM2 axis promotes prostate cancer cell proliferation through mitochondrial stress.

The purpose of this study was to identify the role played by circEEF2 (has-circ-0048559) in prostate cancer (PCa) development and to determine the potential mechanism involved. circEEF2, miR-625-5p, and the transient receptor potential M2 channel protein (TRPM2) were determined using RT-qPCR in PCa. Cell proliferation was determined by CCK-8 assay and colony formation assay, whereas migration and invasion were assessed by Transwell assay, and apoptosis was evaluated by flow cytometry after annexin V-FITC and propidium iodide staining. The interactions between circEEF2 and miRNAs were investigated through the Circular RNA Interactome database, and the downstream targets of miR-625-5p were forecasted using TargetScan. The interaction was confirmed using both the dual luciferase reporter gene assay and RNA pull-down assay. TRPM2, Hedgehog signaling pathway proteins (GLI1 and GLI2), ubiquinone oxidase subunit B8, and cytochrome C oxidase subunit IV (COX4) were analyzed by protein blotting. JC-1 fluorescence detection was applied for mitochondrial membrane potential changes, fluorescent probe assay for intracellular ROS levels, and immunofluorescence staining for γ-H2AX expression. The role of circEEF2 in PCa tumor growth was tested by xenograft experiments. CircEEF2 expression was upregulated in PCa (p<0.05). Cells of PCa were inhibited in proliferation, migration, invasion, and enhanced in apoptosis by depleting circEEF2 (p<0.05). circEEF2 directly targeted adsorbed miR-625-5p. TRPM2 bound to miR-625-5p. Upregulating TRPM2 likewise reversed the therapeutic effect of depleting circEEF2 on cancer development in PCa cells. circEEF2 activated the Hedgehog pathway through the miR-625-5p/TRPM2 axis, promotes mitochondrial stress, and promotes PCa development in vivo. circEEF2 upregulates mitochondrial stress to promote PCa by activating the Hedgehog pathway through the miR-625-5p/TRPM2 axis.

Beta-cypermethrin-induced stress response and ABC transporter-mediated detoxification in Tetrahymena thermophila

β-Cypermethrin (β-CYP), a synthetic pyrethroid pesticide, is widely used for insect management. However, it also affects non-target organisms and pollutes aquatic ecosystems. Tetrahymena thermophila, a unicellular ciliated protist found in fresh water, is in direct contact with aquatic environments and sensitive to environmental changes. The proliferation of T. thermophila was inhibited and the cellular morphology changed under β-CYP stress. The intracellular ROS level significantly increased, and SOD activity gradually rose with increasing β-CYP concentrations. Under 25 mg/L β-CYP stress, 687 genes were up-regulated, primarily enriched in the organic cyclic compound binding and heterocyclic compound binding pathways. These include 8 ATP-binding cassette transporters (ABC) family genes, 2 cytochrome P450 monooxygenase genes, and 2 glutathione peroxidase related genes. Among of them, ABCG14 knockdown affected cellular proliferation under β-CYP stress. In contrast, overexpression of ABCG14 enhanced cellular tolerance to β-CYP. The results demonstrated that Tetrahymena tolerates high β-CYP concentration stress through various detoxification mechanisms, with ABCG14 playing a crucial role in detoxification of β-CYP.

Engineered nanoplatform mediated gas therapy enhanced ferroptosis for tumor therapy in vivo

The high glutathione (GSH) environment poses a significant challenge for inducing ferroptosis in tumor cells, necessitating the development of nanoplatforms that can deplete intracellular GSH. In this study, we developed an engineered nanoplatform (MIL-100@Era/L-Arg-HA) that enhances ferroptosis through gas therapy. First, we confirmed that the Fe element in the nanoplatform undergoes valence changes under the influence of high GSH and H2O2 in tumor cells. Meanwhile, L-Arg generates NO gas in the presence of intracellular H2O2, which reacts with GSH. Additionally, Erastin depletes GSH by inhibiting the cystine/glutamate antiporter system, reducing cystine uptake and impairing GPX4, while also increasing intracellular H2O2 levels by activating NOX4 protein expression. Through these combined GSH-depletion mechanisms, we demonstrated that MIL-100@Era/L-Arg-HA effectively depletes GSH levels, disrupts GPX4 function, and increases intracellular lipid ROS levels in vitro. Furthermore, this nanoplatform significantly inhibited tumor cell growth and extended the survival time of tumor-bearing mice in vivo. This engineered nanoplatform, which enhances ferroptosis through gas therapy, shows significant promise for ferroptosis-based cancer therapy and offers potential strategies for clinical tumor treatment.

Damascenone inhibits osteoclastogenesis by epigenetically modulating Nrf2-mediated ROS scavenge and counteracts OVX-induced osteoporosis

BackgroundBone formation and resorption regulate bone homeostasis. Excessive osteoclastogenesis enhances bone resorption and causes osteoporosis. Although medicines targeting osteoclast have been developed, these drugs have several side effects. Natural compounds have advantages in safety and efficiency, making them potential candidates for osteoporosis treatment. PurposeThis study aims to elucidate the role of damascenone (Dama) in osteoclastogenesis and osteoporosis. Study design and methodsTo demonstrate the effect of Dama on osteoclast differentiation and function, we performed multiple in vitro experiments including TRAP staining, F-actin staining, bone slice resorption assay, real-time PCR, and western bolt. Further, ROS detection, network pharmacology, microscale thermophoresis assay, and ChIP assay were conducted to elucidate the underlying molecular mechanism. Finally, the in vivo effects of Dama were verified using an ovariectomy induced osteoporosis mice model. ResultsDama inhibited RANKL-induced osteoclast differentiation and bone resorptive function in vitro. The expression of osteoclast-related genes and activation of MAPKs and NF-κB signaling in osteoclast were also attenuated by Dama. Meanwhile, Dama reduced intracellular ROS level via up-regulating Nrf2 expression. Network pharmacology demonstrated that HDAC2 is the potential direct target of Dama. Dama inhibited HDAC2 function and increased H3K27ac level of Nrf2, which induced Nrf2 expression and activated ROS scavenging enzymes. Inhibiting NRF2 or activating HDAC2 attenuated the effect of Dama on osteoclastogenesis. Finally, Dama injection suppressed in vivo osteoclastogenesis and ameliorated bone loss induced by OVX. ConclusionDama attenuates osteoclastogenesis by epigenetically modulating Nrf2 expression and ROS scavenge. This study provides evidence for Dama being a potential treatment for osteoporosis.

KLF2 Inhibits Ferroptosis and Improves Mitochondrial Dysfunction in Chondrocyte Through SIRT1/GPX4 Signaling to Improve Osteoarthritis.

Osteoarthritis (OA), a disease of articular joints, is the leading cause of disability in the elderly. Repressing ferroptosis and improving mitochondrial function can delay the progression of OA. Kruppel-like factor 2 (KLF2) exerts a protective effect on OA. However, whether KLF2 affects ferroptosis and mitochondrial function during OA remains unknown. The OA in vivo and in vitro models were constructed in this work. The structural damage of knee joint in OA mice was evaluated through Micro-CT scanning. H&E, SOFG, TB, and TUNEL staining were applied for pathological examination of cartilage tissues. ELISA was employed to examine the contents of inflammatory factors. Additionally, iron deposition in cartilage tissues was measured by Prussian blue staining, and the levels of proteins related to ferroptosis were assessed by immunoblotting. Besides, mitochondrial morphology and function were estimated using a transmission electron microscope and JC-1 staining. In interleukin (IL)-1β-treated C28/I2 cells, the levels of inflammatory factors, intracellular ROS, mitochondrial ROS, lipid ROS, and Fe2+ were measured. Mitochondrial function was evaluated by detecting the levels of mitochondrial membrane potential (MMP), ATP, mPTP, and OCR. KLF2 overexpression ameliorated the structural damage of knee cartilage in OA mice. KLF2 upregulation inhibited ferroptosis and alleviated mitochondrial damage in knee cartilage of OA mice and IL-1β-treated C28/I2 cells. Moreover, KLF2 overexpression activated SIRT1/GPX4 signaling in vivo and in vitro. EX527 addition blocked the influences of KLF2 upregulation on ferroptosis and mitochondrial dysfunction in IL-1β-treated C28/I2 cells. Altogether KLF2 inhibits ferroptosis and improves mitochondrial dysfunction in chondrocytes through SIRT1/GPX4 signaling to improve OA.

The Antifungal Effects of Berberine and Its Proposed Mechanism of Action Through CYP51 Inhibition, as Predicted by Molecular Docking and Binding Analysis.

Fungal infections present a significant health risk, particularly in immunocompromised individuals. Berberine, a natural isoquinoline alkaloid, has demonstrated broad-spectrum antimicrobial activity, though its antifungal potential and underlying mechanisms against both yeast-like and filamentous fungi are not fully understood. This study investigates the antifungal efficacy of berberine against Candida albicans, Cryptococcus neoformans, Trichophyton rubrum, and Trichophyton mentagrophytes in vitro, as well as its therapeutic potential in a murine model of cryptococcal infection. Berberine showed strong antifungal activity, with MIC values ranging from 64 to 128 µg/mL. SEM and TEM analyses revealed that berberine induced notable disruptions to the cell wall and membrane in C. neoformans. No signs of cell necrosis or apoptosis were observed in fungal cells treated with 2 × MIC berberine, and it did not increase intracellular ROS levels or affect mitochondrial membrane potential. Molecular docking and binding affinity assays demonstrated a strong interaction between berberine and the fungal enzyme CYP51, with a dissociation constant (KD) of less than 1 × 10-12 M, suggesting potent inhibition of ergosterol biosynthesis. In vivo studies further showed that berberine promoted healing in guinea pigs infected with T. mentagrophytes, and in a murine cryptococcal infection model, it prolonged survival and reduced lung inflammation, showing comparable efficacy to fluconazole. These findings indicate that berberine exerts broad-spectrum antifungal effects through membrane disruption and CYP51 inhibition, highlighting its potential as a promising therapeutic option for fungal infections.

Plasma activated solution synergistic with exogenous RONS enhances cancer cell apoptosis for improved tumor therapy

Abstract The role of reactive oxygen and nitrogen species (RONS) in plasma-activated solution (PAS) for inducing cancer cell apoptosis is well-established, but suboptimal concentrations and rapid degradation often limit their efficacy. This study addresses a significant gap in current research by exploring how the incorporation of exogenous RONS into PAS can enhance apoptosis in cancer cells, potentially overcoming the limitations of traditional PAS treatments. In this study, the effect of treating saline with a plasma jet for different durations was investigated to analyze how its chemical composition triggers apoptosis in cancer cells in vitro. It was observed that longer plasma treatment times resulted in lower cell viability, the reactive species of H2O2, NO2- and ONOO- were found to play a crucial role in inducing apoptosis in cancer cells. Based on these findings, the anticancer efficacy of PAS was further evaluated after the addition of specific exogenous RONS, using measures such as cell viability, intracellular ROS levels, and microscopic imaging. Biological experiments demonstrated that the anticancer efficacy followed the order: addition of RONS to saline before plasma discharge treatment &gt; addition of RONS to saline after plasma discharge treatment &gt; exogenous RONS solution without plasma treatment. This suggests that the external introduction of RONS can modify the chemical composition of PAS to effectively enhance its anticancer properties, with the addition of H2O2 before plasma discharge treatment proving to be the most effective strategy. These results provide a novel perspective on harnessing the anticancer potential of PAS and expanding its potential applications in tumor therapy.

Transplantation of human umbilical cord-derived mesenchymal stem cells improves age-related ovarian functional decline via regulating the local renin–angiotensin system on inflammation and oxidative stress

BackgroundAge-related reproductive aging is a natural and irreversible physiological process, and delaying childbearing is increasingly common all over the world. Transplantation of mesenchymal stem cells (MSCs) is considered a new and effective therapy to restore ovarian function, but the relevant mechanisms remain unclear. Recently, it has been found that there is a local Renin–angiotensin system (RAS) in human ovary and it plays a key role.MethodsAfter collecting follicular fluid from women who received oocyte retrieval for pure male factor infertility, the level of RAS components in it were detected, and the correlation analysis by linear regression. Then, the in vivo experiments on female C57BL/6 mice were designed to measure ovarian function, and the transcription and translation levels of RAS pathway were detected by molecular biology methods. Moreover, the role of RAS in regulating inflammation and oxidative stress in the co-culture system were explored in in vitro experiments on KGN cells.ResultsFirst, a total of 139 samples of analyzable follicular fluid were obtained. The local RAS of ovary, which is independent of systemic RAS (P > 0.05), is affected by age (Pearson r < 0, P < 0.05) and related to ovarian function, inflammation, oxidative stress indexes and assisted reproduction laboratory outcomes (P < 0.05). Next, the ovary/body weight of aging mice decreased significantly and serum sex hormones levels changed significantly (P < 0.01). The number of functional follicles decreased, while the atresia follicles increased (P < 0.05). After MSCs transplantation, all the above measures have been partially recovered (P < 0.05). Although several RAS components in aging ovary changed, MSCs only improved the expression level of AT1R (P < 0.05). Furthermore, the secretion ability and mitochondrial membrane potential of aging KGN cells decreased, while the intracellular ROS level and the aging cells ratio increased (P < 0.01). All the above measures have been partially recovered when co-cultured with MSCs (P < 0.05). After Ang(1–7) were added into the co-culture system, the above have been more significantly restored compared with Ang II (P < 0.05). Nevertheless, there was no statistical difference in estradiol level no matter which one was added (P > 0.05).ConclusionsTogether, our findings indicate that a novel possible mechanism to explain how stem cells restore age-related ovarian functional decline.

Green synthesis of Au nanoparticles by Scutellaria barbata extract for chemo-photothermal anticancer therapy

IntroductionThe combination of photothermal therapy and chemotherapy has emerged as a promising strategy for cancer treatment. The green synthesis of gold nanoparticles (Au NPs) using extracts from traditional Chinese medicine Scutellaria barbata D. Don (Scu) has revealed an economical, sustainable, and eco-friendly approach for the outstanding anticancer activities. MethodsScu extract was mixed with HAuCl4 solution to fabricate Scu-Au NPs. The synthesis process was optimized by varying the reaction temperature, reaction time, and HAuCl4 concentration. The changes in the components of Scu before and after synthesis were analyzed using the 3,5-dinitrosalicylic acid (DNS) reduction test, the rutin colorimetric method, and the Folin–Ciocalteu method. The morphology, size, and structure of Scu-Au NPs were characterized by TEM, DLS, XRD, and XPS. The photothermal characteristics induced by near-infrared (NIR) irradiation were assessed by continuously monitoring the temperature elevation. The anticancer activity and mechanisms of Scu-Au NPs were investigated using MTT assay, cell uptake studies, ROS level detection, cell apoptosis and necrosis assays, flow cytometry analysis, intracellular Caspase-3 protein activity fluorescence detection, and Western blot analysis on A549, 4T1, and RAW264.7 cells. ResultsScu-Au NPs were successfully fabricated using Scu extract as a reducing agent. The Scu-Au NPs were spherical with a size of 50 nm. During the synthesis process, the levels of reducing sugars, flavonoids, and polyphenols in the Scu extract decreased by 80.8%, 54.3%, and 70.1%, respectively. Scu-Au NPs demonstrated excellent photothermal responsiveness, thermostability, and biocompatibility. For chemo-photothermal anticancer therapy, the cytotoxicity of Scu-Au NPs on 4T1 cells reached approximately 85% upon exposure to 808 nm NIR irradiation. The anticancer activity was attributed to the induction of apoptosis and necrosis, achieved by increasing intracellular ROS levels, causing cell cycle arrest at the S phase, and modulating the expression of apoptosis-related proteins. DiscussionOur synthesized Scu-Au NPs exhibit favorable photothermal conversion characteristics, demonstrating excellent therapeutic efficacy and safety for cancer treatment.

Integrative Human Genetic and Cellular Analysis of the Pathophysiological Roles of AnxA2 in Alzheimer's Disease.

Alzheimer's disease (AD) is an intractable and progressive neurodegenerative disease. Amyloid beta (Aβ) aggregation is the hallmark of AD. Aβ induces neurotoxicity through a variety of mechanisms, including interacting with membrane receptors to alter downstream signaling, damaging cellular or organelle membranes, interfering with protein degradation and synthesis, and inducing an excessive immune-inflammatory response, all of which lead to neuronal death and other pathological changes associated with AD. In this study, we extracted gene expression profiles from the GSE5281 and GSE97760 microarray datasets in the GEO (Gene Expression Omnibus) database, as well as from the Human Gene Database. We identified differentially expressed genes in the brain tissues of AD patients and healthy persons. Through GO, KEGG, and ROC analyses, annexin A2 (AnxA2) was identified as a putative target gene. Notably, accumulating evidence suggests that intracellular AnxA2 is a key regulator in various biological processes, including endocytosis, transmembrane transport, neuroinflammation, and apoptosis. Thus, we conducted a series of cell biology experiments to explore the biological function of AnxA2 in AD. The results indicate that AnxA2 gene knockdown primarily affects oxidative phosphorylation, cell cycle, AD, protein processing in the endoplasmic reticulum, SNARE interactions in vesicular transport, and autophagy. In SH-SY5Y cells secreting Aβ42, AnxA2 gene knockdown exacerbated Aβ42-induced cytotoxicity, including cell death, intracellular ROS levels, and neuronal senescence, altered cell cycle, and reduced ATP levels, suggesting its critical role in mitochondrial function maintenance. AnxA2 gene knockdown also exacerbated the inhibitory effect of Aβ42 on cell migration. AnxA2 overexpression reduced the inflammatory response induced by Aβ42, while its absence increased pro-inflammatory and decreased anti-inflammatory responses. Furthermore, AnxA2 gene knockdown facilitated apoptosis and decreased autophagy. These results indicated potential pathophysiological roles of AnxA2 in AD.

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

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

Elevated circulating LncRNA NORAD fosters endothelial cell growth and averts ferroptosis by modulating the miR-106a/CCND1 axis in CAD patients

Atherosclerosis is a leading cause of cardiovascular diseases, characterized by endothelial dysfunction and lipid accumulation. Long non-coding RNAs (lncRNAs) are emerging as key regulators of endothelial cell behavior. This study aimed to investigate the role of lncRNA NORAD in endothelial cell proliferation and as a potential therapeutic target for atherosclerosis. A total of 75 CAD patients and 76 controls were recruited, and plasma NORAD levels were measured using qRT-PCR. HUVECs were transfected with si-NORAD to evaluate its effects on cell cycle, proliferation, migration, and apoptosis. Plasma NORAD levels were significantly elevated in CAD patients. The NORAD-miRNA-mRNA ceRNA regulatory network was constructed based on GEO database, and G1/S-specific cyclin-D1 (CCND1) was identified as one of the hub factors. NORAD deficiency suppressed cell migration and induced G1 cell cycle arrest in HUVECs by downregulating CCND1 in vitro. NORAD upregulated CCND1 in HUVECs via sponging miR-106a that inhibited cell migration. The dual-luciferase assay confirmed the direct targeting of miR-106a by NORAD, and overexpression of miR-106a inhibited HUVEC proliferation and migration. Si-NORAD transfection resulted in induced early apoptosis, increased intracellular ROS levels, decreased GSH levels, and reduced mitochondrial membrane potential. Additionally, si-NORAD decreased the expression of GPX4, FTH1, KEAP1, NCOA4, and Nrf2, while increasing Xct levels, confirming the involvement of ferroptosis. Our findings reveal that NORAD plays a critical role in endothelial cell proliferation, migration, and apoptosis, and its silencing induces ferroptosis. The regulatory network involving NORAD, miR-106a, and their target genes provides a potential therapeutic avenue for atherosclerosis.