Cell Damage Research Articles

Gastroprotective Effect of Isoferulic Acid Derived from Foxtail Millet Bran against Ethanol-Induced Gastric Mucosal Injury by Enhancing GALNT2 Enzyme Activity.

Excessive alcohol consumption has led to the prevalence of gastrointestinal ailments. Alleviating gastric disorders attributed to alcohol-induced thinning of the mucus layer has centered on enhancing mucin secretion as a pivotal approach. In this study, foxtail millet bran polyphenol BPIS was divided into two components with MW < 200 D and MW > 200 D by molecular interception technology. Combined with MTT, cell morphology observation, and trypan blue staining, isoferulic acid (IFA) within the MW < 200 D fraction was determined as the effective constituent to mitigate ethanol-induced damage of gastric epithelial cells. Furthermore, a Wistar rat model with similar clinical features to alcohol-induced gastric mucosal injury was established. Then, gastric morphological observation, H&E staining, and assessments of changes in gastric hexosamine content and gastric wall binding mucus levels were carried out, and the results revealed that IFA (10 mg/Kg) significantly ameliorated alcohol-induced gastric mucosal damage. Finally, we applied techniques including Co-IP, molecular docking, and fluorescence spectroscopy and found that IFA inhibited the alcohol-induced downregulation of N-acetylgalactosamintransferase 2 (GALNT2) activity related to mucus synthesis through direct interaction with GALNT2 in gastric epithelial cells, thus promoting mucin synthesis. Our study lays a foundation for whole grain dietary intervention tailored to individuals suffering from alcoholic gastric mucosal injury.

Weibel-Palade bodies: function and role in thrombotic thrombocytopenic purpura and in diarrhea phase of STEC-hemolytic uremic syndrome.

Vascular endothelial cells are equipped with numerous specialized granules called Weibel-Palade bodies (WPBs). They contain a cocktail of proteins that can be rapidly secreted (3-5min) into the vascular lumen after an appropriate stimulus such as thrombin. These proteins are ready without synthesis. Von Willebrand factor (VWF) and P-selectin are the main constituents of WPBs. Upon stimulation, release of ultralarge VWF multimers occurs and assembles into VWF strings on the apical side of endothelium. The VWF A1 domain becomes exposed in a shear-dependent manner recruiting and activating platelets. VWF is able to recruit leukocytes via direct leukocyte binding or via the activated platelets promoting NETosis. Ultralarge VWF strings are ultimately cleaved into smaller pieces by the protease ADAMTS-13 preventing excessive platelet adhesion. Under carefully performed flowing conditions and adequate dose of Shiga toxins, the toxin induces the release of ultralarge VWF multimers from cultured endothelial cells. This basic information allows insight into the pathogenesis of thrombotic thrombocytopenic purpura (TTP) and of STEC-HUS in the diarrhea phase. In TTP, ADAMTS-13 activity is deficient and systemic aggregation of platelets will occur after a second trigger. In STEC-HUS, stimulated release of WPB components in the diarrhea phase of the disease can be presumed to be the first hit in the damage of Gb3 positive endothelial cells.

LncRNA FGD5-AS1 is required for gastric cancer proliferation by inhibiting cell senescence and ROS production via stabilizing YBX1

BackgroundThe vast majority of lncRNAs have low expression abundance, which greatly limits their functional range and impact. As a high expression abundance lncRNA, FGD5-AS1’s non-ceRNA biological function in cancer is unclear.MethodsRNA-seq studies and chromatin immunoprecipitation (Chip) assays were performed to identify ZEB1-regulated lncRNAs. RNA sequencing, RNA pulldown, RNA Immunoprecipitation assays, and rescue assays were conducted to explore the molecular mechanisms of FGD5-AS1 in GC.ResultsAs one of the most abundant lncRNAs in cells, FGD5-AS1 has been shown to be transcriptionally activated by ZEB1, thus closely related to epithelial-mesenchymal transition (EMT) signaling. Clinical analysis showed that FGD5-AS1 overexpression was clinically associated with lymph node metastasis, and predicted poor survival in GC. Loss-of-function studies confirmed that FGD5-AS1 knockdown inhibited GC proliferation and induced cisplatin chemosensibility, cell senescence, and DNA damage in GC cells. Mechanismically, FGD5-AS1 is a YBX1-binding lncRNA due to its mRNA contains three adjacent structural motifs (UAAUCCCA, ACCAGCCU, and CAGUGAGC) that can be recognized and bound by YBX1. And this RNA-protein interaction prolonged the half-life of the YBX1 protein in GC. Additionally, a rescue assay showed that FGD5-AS1 promotes GC by repressing cell senescence and ROS production via YBX1.ConclusionFGD5-AS1 is a cellular high-abundant lncRNA that is transcriptionally regulated by ZEB1. FGD5-AS1 overexpression promoted GC progression by inhibiting cell senescence and ROS production through binding and stabilizing the YBX1 protein.

Assessment of objective visual function following idebenone administration in patients with leber hereditary optic neuropathy.

To objectively assess visual function in Leber's Hereditary Optic Neuropathy (LHON) patients; this study evaluated pre- and post-idebenone treatment changes in primary visual cortical (V1) responses using functional magnetic resonance imaging (fMRI), given the challenges in subjective testing due to central retinal ganglion cell damage. A descriptive study involving four confirmed LHON patients. Four patients received 900 mg/day of oral idebenone for 24 weeks. Baseline and post-treatment visual acuity, visual fields, and BOLD fMRI responses while passively viewed drifting contrast pattern visual stimuli were compared with self-reported symptoms. Post-idebenone, one patient showed positive trends across subjective tests, reported symptoms, and fMRI. Two patients had stable symptoms and fMRI responses; one improved on subjective tests, and another worsened slightly. Another patient improved in visual field tests despite worsening symptoms and fMRI trends. fMRI may offer a valuable objective measure of visual functions in LHON and appears to be more relevant in assessing symptoms. Further research with more participants is needed to ascertain fMRI's role in developing objective visual assessments and treatment evaluation.

SGLT2 inhibitor dapagliflozin reduces proximal tubular cell damage biomarkers in patients with acute heart failure

SGLT2 inhibitor dapagliflozin reduces proximal tubular cell damage biomarkers in patients with acute heart failure

Vitamin D alleviation of oxidative stress in human retinal pigment epithelial cells.

Oxidative stress-induced retinal pigment epithelium (RPE) cell damage is a major factor in age-related macular degeneration (AMD). Vitamin D3 (VD3) is a powerful antioxidant and it has been suggested to have anti-aging properties and potential for treating AMD. This study aimed to investigate the effect of VD3 on RPE cell oxidative apoptosis of RPE cells in order to provide experimental evidence for the treatment of AMD. Human retinal pigment epithelial cell 19 (ARPE-19) cells were divided into four groups: blank group (untreated), model group (incubated in medium with 400μmol/L H2O2 for 1h), VD3 group (incubated in medium with 100μmol/L VD3 for 24h), and treatment group (incubated in medium with 400μmol/L H2O2 for 1h and 100μmol/L VD3 for 24h). Cell viability, cell senescence, ROS content, expression levels of vitamin D specific receptors, Akt, Sirt1, NAMPT, and JNK mRNA expression levels, SOD activity, and MDA, GSH, and GPX levels were measured. We first established an ARPE-19 cell stress model with H2O2. Our control experiment showed that VD3 treatment had no significant effect on ARPE-19 cell viability within 6-48h. Treating the stressed ARPE-19 cells with VD3 showed mixed results; caspase-3 expression was decreased, Bcl-2 expression was increased, MDA level of ARPE-19 cells was decreased, GSH-PX, GPX and SOD levels were increased, the relative mRNA expression levels of Akt, Sirt1, NAMPT were increased (P < 0.05), and the relative mRNA expression level of JNK was decreased (P < 0.05). VD3 can potentially slow the development of AMD.

Photoactivated Anticancer Activity of Cobalt(III) Complexes with Naturally Occurring Flavonoids Chrysin and Silibinin.

Photoactive metal complexes of bioessential transition metal ions with natural chelators are gaining interest as photocytotoxic agents for cancer photodynamic therapy (PDT). We report six new cobalt(III) complexes with a mixed-ligand formulation [Co(B)2(L)](ClO4)2 (Co1-Co6), where B represents a N,N-donor α-diimine ligand, namely, phenanthroline (phen; Co1, Co2), dipyrido[3,2-d:2',3'-f]quinoxaline (dpq; Co3, Co4), and dipyrido[3,2-a:2',3'-c]phenazine (dppz; Co5, Co6), and L is the monoanionic form of the naturally occurring flavonoids chrysin (chry; Co1, Co3, Co5) and silibinin (sili; Co2, Co4, Co6). Complexes displayed a d-d absorption band within 500-700 nm and exhibited excellent dark and photostability in solution. Cytotoxicity studies indicated significant activity of Co5 and Co6 against cervical (HeLa) and lung (A549) cancer cells under visible light (400-700 nm) irradiation giving low micromolar IC50 values (2.3-3.4 µM, phototoxicity index ~ 15-30). The complexes demonstrated notably low toxicity against normal HPL1D lung epithelial cells. Flow cytometry assay revealed an apoptotic mode of cell damage triggered by the complexes when irradiated. ROS generation assay indicated the involvement of singlet oxygen species in the cell death mechanism when irradiated with light. Overall, complexes Co5 and Co6 with coordinated dipyridophenazine and flavonoid ligands are potential candidates for cancer PDT applications.

Kaempferol Alleviates Injury in Human Retinal Pigment Epithelial Cells via STAT1 Ubiquitination-Mediated Degradation of IRF7.

Retinal pigment epithelial (RPE) cells have a pivotal function in preserving the equilibrium of the retina and moderating the immunological interaction between the choroid and the retina. This study primarily focuses on delineating the protective effect offered by Kaempferol (Kae) against RPE cell damage. Bioinformatics analysis was performed on the GSE30719 dataset to identify hub genes associated with RPE. Subsequently, we analyzed the impact of Kae on RPE apoptosis, cell viability, and inflammatory response through cell experiments, and explored the interaction between hub genes and Kae. Based on the GSE30719 dataset, nine hub genes (ISG15, IFIT1, IFIT3, STAT1, OASL, RSAD2, IRF7, MX2, and MX1) were identified, all of which were highly expressed in the GSE30719 case group. Kae could boost the proliferative activity of RPE cells caused by lipopolysaccharide (LPS), as well as reduce apoptosis and the generation of inflammatory factors (tumor necrosis factor receptor (TNFR), interleukin-1beta (IL-1β)) and cytokines (IL-1, IL-6, IL-12). STAT1 was shown to inhibit cell proliferation, promote apoptosis, and secrete IL-1/IL-6/IL-12 in LPS-induced RPE cells. Moreover, IRF7 was found to interact with STAT1 in LPS-induced RPE cells, and STAT1 could maintain IRF7 levels through deubiquitination. In addition, we also found that the protective effect of Kae on LPS-induced RPE cell injury was mediated through STAT1/IRF7 axis. This study provided evidence that Kae protects RPE cells via regulating the STAT1/IRF7 signaling pathways, indicating its potential therapeutic relevance in the diagnosis and management of retinal disorders linked with RPE cell damage.

Two new cucurbitane-type triterpenoid saponins from the fruit of Citrullus colocynthis

Two new cucurbitane-type triterpenoid saponins, 2,20β,22β-trihydroxy-16α,23(R)-epoxycucurbita-1,5,24-triene-3,11-dione 2-O-β-D-glucopyranoside (1), 2,20β,22α-trihydroxy-16α,23(S)-epoxycucurbita-1,5,11,24-tetraene-3-one 2-O-β-D-glucopyranoside (2) were isolated from the fruit of Citrullus colocynthis (L.) Schrad. Their structures were elucidated by mass spectrometry, IR, 1D, and 2D NMR spectroscopy, etc. Besides, both of the compounds showed significant hepatoprotective activities at 10 μM against paracetamol-induced HepG2 cell damage.

P-156 Ultra-fast warming of blastocysts yields better survival rates for D5 non-PGT and similar survival rates for D5/D6 PGT blastocysts

Abstract Study question Can an ultra-fast single-step warming of vitrified blastocysts yield similar survival rates and post-warming development compared to the standard multi-step rehydration protocol? Summary answer Ultra-fast warming, shows significantly better rates of survival and post-warming development in vitro for non-PGT blastocysts and similar rates for PGT blastocysts. What is known already Excellent survival rates have been reported since the introduction of human blastocyst vitrification. Typically, most warming procedures involve three steps with decreasing concentrations of non-penetrating cryoprotectants. Recently, few studies have evaluated the effectiveness of simplifying the procedure to a single step for rehydration, while maintaining comparable survival and pregnancy rates. Adopting simpler procedures can optimize laboratory protocols, workflow, and safety, supporting further advancements in Assisted Reproduction Technologies (ART). Study design, size, duration This validation study ultra-fast warmed 246 vitrified blastocysts [129 non-PGT day5 (cryopreservation period 23/01/2017-24/04/2018 VIT-kit, Fujifilm), 57 PGT day5 (25/01/2023-31/03/2023 VIT-KitNX, Fujifilm), 60 PGTday6 (03/02/2023-22/08/2023 Vit-KitNX, Fujifilm)] and compared survival rates with 379 blastocysts [240 non-PGT day5, 97 PGT day5, 42 PGT day6] vitrified in the same time-periods and warmed for clinical application using the standard multi-step protocol. Ultra-fast warmed blastocysts were cultured for 24h in a time-lapse incubator and subjected to a viability staining. Participants/materials, setting, methods Blastocysts, originating from 111 patients that were donated to validation studies after the legal storage period(n = 129) or deemed non-transferable after PGT(n = 117), were ultra-fast warmed by immersing the straw in a 37 °C thawing solution (1M trehalose; Vit-kitwarmNX; Fujifilm) for 1 minute. Survival rates [total (≥50% intact) and fully (100% intact) ], transfer rates (2h post-warming, ≥50% intact), re-expansion time and the number of damaged cells (Live/Dead kit L-3224, Invitrogen) were analysed. Fisher’s Exact Test (p &amp;lt; 0.05). Main results and the role of chance Overall, the immediate post-warming survival rates (≥50% intact) were comparable between ultra-fast and multi-step warming, 98.4% vs 96.8% respectively (p = 0.3041). However, the fully intact survival rate immediately post-warming was significantly higher with ultra-fast warming (85.4% vs 76.3%; p = 0.0058) as well as the percentage of blastocysts considered transferable (96.7% vs 91.2%; p = 0.0076). Sub-group analysis demonstrated significantly better results for non-PGT blastocysts following ultra-fast warming. Higher immediate survival post-warming (100% vs 96.6%; p = 0.036) and transfer rates (100% vs 90%; p = 0.0002) were achieved. Similar results were observed for PGT blastocysts on both day 5 and day 6, with no significant differences in total, fully intact, and transfer rates observed between the two warming procedures [PGT Day5: 96.5%, 80.7%, and 93.0% vs. 97.9%(p = 0.6270), 69.1%(p = 0.1334), and 93.8%(p = 1.0000)] [PGT Day6: 96.7%, 68.3%, and 93.3% vs. 95.2%(p = 1.0000), 64.3%(p = 0.6761), and 92.8%(p = 1.0000)]. Ultra-fast warmed blastocysts showed a total re-expansion rate of 93.3% in 3.3h±2.7h on average. Viability staining demonstrated very few cells damaged. Zero cell damage was observed in 31.5% of the blastocysts, 59.0% had 1-10 cells damaged, 9.5% &amp;gt;10 cells damaged, and only 4 blastocysts were completely degenerated. Limitations, reasons for caution In order to limit the amount of embryos needed in this validation study, ultra-fast warming outcome parameters were compared with retrospective clinical results of blastocysts vitrified in the same time periods. Ultra-fast warming is a single-step process where precise temperature control(37 °C) of the warming medium is crucial for success. Wider implications of the findings Minimizing handling steps leads to an overall risk reduction in potential exposure to suboptimal warming conditions and operator/handling-related stress. This optimization brings huge efficiency to the busy work schedule that is characteristic for an IVF lab without dropping cryopreservation quality indicators. Trial registration number Not applicable

3’-Methyl-4-thio-1H-tetrahydropyranspiro-5’-hydantoin platinum complex as a novel deoxyribonuclease I inhibitor

Deoxyribonuclease I (DNase I) is one of the main nucleases involved in deoxyribonucleic acid (DNA) degradation during apoptosis. It catalyzes the hydrolytic cleavage of DNA, producing 5‘-oligonucleotides. The inhibition of DNase I may serve as an important mechanism for protecting DNA against premature degradation during cell damage. Fourteen hydantoin-containing compounds, including two newly synthesized and seven previously synthesized metal complexes, along with five previously synthesized hydantoin ligands, were evaluated in vitro for their inhibitory properties against bovine pancreatic DNase I. As a result, the 3’-methyl-4-thio-1H-tetrahydropyranspiro-5’-hydantoin platinum complex (8) inhibited the enzyme with an IC50 value of 110.20 ± 24.20 µM, a potency 3-fold greater than that of the reference crystal violet (IC50 = 378.27 ± 47.75 µM). To understand the binding mode and mechanism of inhibition of compound 8 with DNase I, molecular docking calculations were performed. The analysis revealed that compound 8 interacts with the most important catalytic residues of DNase I. To the best of our knowledge, this is the first report of a platinum complex inhibiting DNase I.

Assessing the antioxidant properties of Naringin and Rutin and investigating their oxidative DNA damage effects in breast cancer

This work examines the capacity of Naringin and Rutin to influence the DNA damage response (DDR) pathway by investigating their interactions with key DDR proteins, including PARP-1, ATM, ATR, CHK1, and WEE1. Through a combination of in silico molecular docking and in vitro evaluations, we investigated the cytotoxic and genotoxic effects of these compounds on MDA-MB-231 cells, comparing them to normal human fibroblast cells (2DD) and quiescent fibroblast cells (QFC). The research found that Naringin and Rutin had strong affinities for DDR pathway proteins, indicating their capacity to specifically regulate DDR pathways in cancer cells. Both compounds exhibited preferential cytotoxicity towards cancer cells while preserving the vitality of normal 2DD fibroblast cells, as demonstrated by cytotoxicity experiments conducted at a dose of 10 µM. The comet experiments performed particularly on QFC cells provide valuable information on the genotoxic impact of Naringin and Rutin, highlighting the targeted initiation of DNA damage in cancer cells. The need to use precise cell models to appropriately evaluate toxicity and genotoxicity is emphasized by this discrepancy. In addition, ADMET and drug-likeness investigations have emphasized the pharmacological potential of these compounds; however, they have also pointed out the necessity for optimization to improve their therapeutic profiles. The antioxidant capabilities of Naringin and Rutin were assessed using DPPH and free radical scavenging assays at a concentration of 10 µM. The results confirmed that both compounds have a role in reducing oxidative stress, hence enhancing their anticancer effects. Overall, Naringin and Rutin show potential as medicines for modulating the DDR in cancer treatment. They exhibit selective toxicity towards cancer cells while sparing normal cells and possess strong antioxidant properties. This analysis enhances our understanding of the therapeutic uses of natural chemicals in cancer treatment, supporting the need for more research on their mechanisms of action and clinical effectiveness.

P-606 Maintenance of telomere length through autophagy and androgen receptor inhibition in infertile individuals with differences of sex development (DSD)

Abstract Study question What are the molecular mechanisms underlying genome instability concomitant with increased telomere length (TL) in individuals with DSD? Summary answer Low autophagy activity and androgen receptor signaling stabilize GAR1, a telomerase-specific protein, leading to an increase in TL in DSD cells. What is known already Increased genome instability is linked to infertility, and compromised telomere maintenance is associated with genome instability. Individuals with DSD, characterized by dysgenic gonads and a high risk of Germ Cell Tumor (GCT) development, exhibit concurrent genome instability, increased TL, and deregulated Androgen Receptor (AR) signaling. Telomeric DNA damage promotes autophagy, which induces cell death and prevents propagation of replication crisis and tumorigenesis. We previously demonstrated that inhibition of autophagy, a conservative process of protein degradation, led to TL increase in cell lines derived from individuals with DSD, but the underlying mechanisms remain unclear. Study design, size, duration 112 individuals with DSD, including Swyer syndrome (46,XY females), Complete Androgen Insensitivity Syndrome (CAIS 46,XY females), Turner (45,X0 females), and Mixed Gonadal Dysgenesis (MGD 45,X0/46,XY females and males) were enrolled into study during routine praxis in our clinic. The samples were collected with written patients consent. A control group included fertile women and men, when appropriate, of comparable age. Samples underwent standard genetic and histological characterization. Lymphoblastoid cell lines were used for functional studies. Participants/materials, setting, methods Study groups comprised individuals with CAIS (n = 19); Swyer (n = 8); MGD (n = 12); Turner (n = 23) with mosaic (n = 12), with chr. X/Y deletions (n = 8), with X-isochr./idiocentric/marker chr. (n = 23) syndromes. Blood samples were collected for DNA, RNA and protein isolation from leukocytes. Additionally, individuals with DSD who underwent gonadectomy formed an additional group with histologically proven GCT (DSD-GCT, n = 7). Gonadal tissue was provided by the tissue bank. Samples were analyzed using quantitative real-time PCR (qRT-PCR), immunoblotting, and mass spectrometry. Main results and the role of chance Leukocytes from Swyer (p = 0,0069), Turner (p = 0,0064), and MGD without GCT (p = 0,05) groups exhibited a 1,5-folds increase in TL, diminished with GCT (p = 0,003). The RNA component of telomerase, TERC, was inhibited two times in Swyer (p = 0,0417). Proteomic analysis identified a telomerase protein GAR1 accumulation in Swyer, but downregulated in GCT group together with AR upregulation (p = 0,0379). Immunoblotting confirmed a two-fold increase of GAR1 in Swyer (p = 0,0022) and Turner (p = 0,0401), while GAR1 decreased in all DSD samples (p = 0,0455). GAR1 protein increased stability, not due to upregulated transcription, and promoted TL since we illustrated GAR1 accumulation within 72h of BafilomycinA1 treatment inhibiting autophagy. This treatment induced telomerase activity within 24h. Further autophagy inhibition caused accumulation of DNA damage in DSD cells, measured by γH2AX levels labeling double strand brakes (p = 0,0286). Dysgenic gonadal tissue analysis also showed a negative correlation between autophagy and TL-specific gene expression, with LC3 (p = 0,0341) and P62 (p = 0,0342) increased in all DSD samples, and proteins LC3 (p = 0,0298) and P62 (p = 0,0237) upregulated, and GAR1 (p = 0,0432) inhibited exclusively in Swyer group. Inhibition of AR increased TL (p = 0,0357) together with GAR1 (p = 0,0125) in DSD group at 19h, which was associated with a consecutive γH2AX upregulation (p = 0,0489) at 24h with Enzalutamide. Limitations, reasons for caution This study is based on a limited number of samples from the individuals with rare genetic syndromes, some observed only in 1 out of 10,000 clinical cases. Therefore, in vitro study models are recommended for further research. Wider implications of the findings The study reveals gene expression patterns and molecular regulatory mechanisms associated with DSD and GCT risk. The data suggest possibilities for prophylactic treatment of DSD individuals and new diagnostic approaches of GCT using peripheral blood. The study also emphasizes the importance of addressing genome integrity in infertility research. Trial registration number NA

Exploring the preventive effects of Jie Geng Tang on pulmonary fibrosis induced in vitro and in vivo: a network pharmacology approach.

Pulmonary fibrosis is a debilitating lung disease marked by excessive fibrotic tissue accumulation, which significantly impairs respiratory function. Given the limitations of current therapies, there is an increasing interest in exploring traditional herbal formulations like Jie Geng Tang (JGT) for treatment. This study examines the potential of JGT and its bioactive component, quercetin, in reversing bleomycin (BLM)-induced pulmonary fibrosis in mice. We employed a BLM-induced MLE-12 cell damage model for in vitro studies and a bleomycin-induced fibrosis model in C57BL/6 mice for in vivo experiments. In vitro assessments showed that JGT significantly enhanced cell viability and reduced apoptosis in MLE-12 cells treated with BLM. These findings underscore JGT's potential for cytoprotection against fibrotic agents. In vivo, JGT was effective in modulating the expression of E-cadherin and vimentin, key markers of the epithelial-mesenchymal transition (EMT) pathway, indicating its role in mitigating EMT-associated fibrotic changes in lung tissue. Quercetin, identified through network pharmacology analysis as a potential key bioactive component of JGT, was highlighted for its role in the regulatory mechanisms underlying fibrosis progression, particularly through the modulation of the IL-17 pathway and Il6 expression. By targeting inflammatory pathways and key processes like EMT, JGT and quercetin offer a potent alternative to conventional therapies, meriting further clinical exploration to harness their full therapeutic potential in fibrotic diseases.

In Vitro Protective Effect of Pea-Derived Peptides (PPs) via the Keap1/Nrf2 Signaling Pathway on Alpha-Gliadin-Sensitizing Peptide Induced Cacao-2 Cells.

Celiac disease (CD) is an allergic intestinal disease caused mainly by gliadin in wheat, which is widespread in the population and currently lacks effective treatment. α-Gliadin peptides cause cellular damage by substantially increasing cellular reactive oxygen species (ROS) levels. This study investigates the protective effect of 11 pea-derived peptides (PPs) on ɑ-gliadin peptide (P31-43) treated Caco-2 cells. Results show that cells treated with PP2, PP5, and PP6 peptides significantly reduce the cell mortality caused by P31-43. Three PPs significantly reduce the P31-43-induced decrease in ROS levels to control levels, and there is no difference between them and the vitamin C (Vc) group. The results in terms of antioxidant-related enzymes show that PPs significantly decrease superoxide dismutase activity (SOD), glutathione reductases (GR), and glutathione (GSH)/oxidized glutathione (GSSG) levels, thus significantly enhancing the antioxidant level of cells. By studying the key proteins of the Kelch-like ECH-associated protein 1 (Keap1)/NF-E2-related factor 2 (Nrf2) pathway, it is found that PPs activate the Keap1/Nrf2 signaling pathway. The study finds that peptides from peas can effectively alleviate ɑ-gliadin peptide-induced cell damage. The discovery of these food-derived peptides provides novel potential solutions for the prevention and treatment of CD.

P-544 Viability assessment using fluorescent markers, ultrastructure and implantation of human embryos following blastocyst biopsy using three different laser systems

Abstract Study question Are there any differences in viability, ultrastructure and implantation of human blastocysts biopsied using three different laser systems? Summary answer No statistically significant differences are observed in viability, ultrastructure and implantation of human blastocysts biopsied using three different laser systems What is known already Most studies in human embryos concentrate on assessing the success of biopsy/vitrification on survival rates and clinical outcomes following transfer. Our previous published studies examined in addition to the survival rates, the cytoskeleton, ultrastructure and viability of cleavage and blastocyst stage biopsied embryos, using a specific laser system, emphasizing differences between the two methods and alerting on effective strategies to minimise potential laser induced damage. This is the first study to investigate the safety of 3 different laser systems for blastocyst biopsy by comparing implantation rates and by analysing cell viability using Fluorescence Microscopy and ultrastructure by Transmission Electron Microscopy(TEM). Study design, size, duration 405 human blastocysts were biopsied on day 5 for PGT-A or PGT-SR or PGT-M using the Saturn laser (n = 145-58 cycles) or the Octax laser (n = 132-55 cycles) or the Lykos laser (n = 128-46 cycles). Following transfer of normal embryos, spare embryos, rejected for transfer due to being diagnosed as abnormal were processed following vitrification/warming for viability analysis (n = 30 Saturn laser, n = 30 Octax laser, n = 30 Lykos laser) or TEM (n = 20 Saturn laser, n = 20 Octax laser, n = 20 Lykos laser). Participants/materials, setting, methods The study was conducted in an academic hospital with an IVF/PGT laboratory, 2 private IVF Units and an IVF Unit of a Naval hospital. Blastocyst viability after biopsy/vitrification/warming was assessed using two fluorescent markers: carboxyfluorescein-diacetate succinimidylester (CFSE-viable cells) and propidium iodide (PI–damaged cells) in combination with DAPI to visualise DNA. TEM analysis was carried out following embryo fixation in gluteraldehyde, incubation in osmium, aqueous uranyl acetate, dehydration through ethanol series, and immersion in Epon. Main results and the role of chance The implantation rates (+ve hCG / ET) were not statistically different among the 3 laser systems (64.4% -Saturn, 63.4%- Octax, 64.7%- Lykos p &amp;gt; 0.05). Viability staining with CFSE/PI following vitrification/warming revealed that all biopsied blastocysts had damaged cells at the position of cutting (range 4-8cells) and additional PI stained cells in other parts (range 2-15), possibly due to damage sustained during vitrification/warming. The majority of embryos across the 3 groups had ≤7% PI stained damaged cells (73.3% Saturn, 73.3% Octax, 76.7% Lykos), indicating no statistically significant difference in blastocyst viability among the 3 laser systems (p &amp;gt; 0.05). No difference was also observed in the incidence of lysed biopsied TE cells or the PGT results. TEM analysis revealed similar ultrastracture among the 3 groups, as documented by the presence of normal organelles’ structures including smooth/rough endoplasmic reticulum, Golgi apparatus, mature mitochondria with cisternae, zona and microvilli. Multivesicular bodies, residual bodies, lipofuscins and autophagic vacuoles were also evident and indicative of elimination and rescue process. Excessive accumulation of glycogen granules and distension of mitochondria were more prominent in Grade B blastocysts compared to Grade A blastocysts, which may indicate a ‘block’ of glycogenolysis secondary to the mitochondrial alterations, suggesting changes in normal embryo metabolism. Limitations, reasons for caution The biopsied blastocysts used for viability and ultrastructural analysis in this study were all diagnosed with either chromosomal abnormalities or single gene defects following PGT-A or PGT-SR or PGT-M. Wider implications of the findings The similarities observed in implantation rates, the minimal levels of cell damage and the embryo quality dependent limited ultrastuctural anomalies identified confirm the safety of all three laser systems and emphasize the blastocysts’ ability to initiate elimination/rescue processes in their attempt to fully recover from ‘traumas’ sustained during biopsy/vitrification. Trial registration number not applicable

Mitochondrial Quality Control Processes at the Crossroads of Cell Death and Survival: Mechanisms and Signaling Pathways.

Biological aging results from an accumulation of damage in the face of reduced resilience. One major driver of aging is cell senescence, a state in which cells remain viable but lose their proliferative capacity, undergo metabolic alterations, and become resistant to apoptosis. This is accompanied by complex cellular changes that enable the development of a senescence-associated secretory phenotype (SASP). Mitochondria, organelles involved in energy provision and activities essential for regulating cell survival and death, are negatively impacted by aging. The age-associated decline in mitochondrial function is also accompanied by the development of chronic low-grade sterile inflammation. The latter shares some features and mediators with the SASP. Indeed, the unloading of damage-associated molecular patterns (DAMPs) at the extracellular level can trigger sterile inflammatory responses and mitochondria can contribute to the generation of DAMPs with pro-inflammatory properties. The extrusion of mitochondrial DNA (mtDNA) via mitochondrial outer membrane permeabilization under an apoptotic stress triggers senescence programs. Additional pathways can contribute to sterile inflammation. For instance, pyroptosis is a caspase-dependent inducer of systemic inflammation, which is also elicited by mtDNA release and contributes to aging. Herein, we overview the molecular mechanisms that may link mitochondrial dyshomeostasis, pyroptosis, sterile inflammation, and senescence and discuss how these contribute to aging and could be exploited as molecular targets for alleviating the cell damage burden and achieving healthy longevity.

Mitochondrial-targeted cyclometalated Ir(III)-5,7-dibromo/dichloro-2-methyl-8-hydroxyquinoline complexes and their anticancer efficacy evaluation in Hep-G2 cells.

Both 8-hydroxyquinoline compounds and iridium (Ir) complexes have emerged as potential novel agents for tumor therapy. In this study, we synthesized and characterized two new Ir(III) complexes, [Ir(L1)(bppy)2] (Br-Ir) and [Ir(L2)(bppy)2] (Cl-Ir), with 5,7-dibromo-2-methyl-8-hydroxyquinoline (HL-1) or 5,7-dichloro-2-methyl-8-hydroxyquinoline as the primary ligand. Complexes Br-Ir and Cl-Ir successfully inhibited antitumor activity in Hep-G2 cells. In addition, complexes Br-Ir and Cl-Ir were localized in the mitochondrial membrane and caused mitochondrial damage, autophagy, and cellular immunity in Hep-G2 cells. We tested the proteins related to mitochondrial and mitophagy by western blot analysis, which showed that they triggered mitophagy-mediated apoptotic cell death. Remarkably, complex Br-Ir showed high in vivo antitumor activity, and the tumor growth inhibition rate was 63.0% (P<0.05). In summary, our study on complex Br-Ir revealed promising results in in vitro and in vivo antitumor activity assays.

Conversion of notoginsenoside R1 to 3β,12β-dihydroxydammar-(E)-20(22),24-diene-6-O-β-D-xylopyranosyl-(1→2)-β-D-glucopyranoside by Lactiplantibacillus plantarum S165 enhanced protective effects of LPS-induced intestinal epithelial barrier injury in Caco-2 cells.

Microbial transformation to modify saponins and enhance their biological activities has received increasing attention in recent years. This study aimed to screen the strain that can biotransform notoginsenoside R1, identify the product and study its biological activity. A lactic acid bacteria strain S165 with glycosidase-producing activity was isolated from traditional Chinese fermented foods, which was identified and grouped according to API 50 CHL kit and 16S rDNA sequence analysis. Subsequently, notoginsenoside R1 underwent a 30-day fermentation period by the strain S165, and the resulting products were analyzed using High-performance liquid chromatography (HPLC), Ultra-performance liquid chromatography (UPLC)-mass spectrometry (MS)/MS, and 13C-Nuclear magnetic resonance (NMR) techniques. Employing a model of Lipopolysaccharide (LPS)-induced damage to Caco-2 cells, the damage of Caco-2 cells was detected by Hoechst 33 258 staining, and the activity of notoginsenoside R1 biotransformation product was investigated by CCK-8 and western blotting assay. The strain S165 was identified as Lactiplantibacillus plantarum and was used to biotransform notoginsenoside R1. Through a 30-day biotransformation, L. plantarum S165 predominantly converts notoginsenoside R1 into 3β,12β-dihydroxydammar-(E)-20(22),24-diene-6-O-β-D-xylopyranosyl-(1→2)-β-D-glucopyranoside, temporarily named notoginsenoside T6 (NGT6) according to HPLC, UPLC-MS/MS, and 13C-NMR analysis. Results from CCK-8 and Hoechst 33258 staining indicated that the ability notoginsenoside T6 to alleviate the intestinal injury induced by LPS in the Caco-2 cell was stronger than that of notoginsenoside R1. In addition, Western blotting result showed that notoginsenoside T6 could prevent intestinal injury by protecting tight junction proteins (Claudin-1, Occludin, and ZO-1). Notoginsenoside R1 was biotransformed into the notoginsenoside T6 by L. plantarum S165, and the biotransformed product showed an enhanced intestinal protective effect in vitro.

Obesity Induces DNA Damage in Mammary Epithelial Cells Exacerbated by Acrylamide Treatment through CYP2E1-Mediated Oxidative Stress.

Obesity and environmental toxins are risk factors for breast cancer; however, there is limited knowledge on how these risk factors interact to promote breast cancer. Acrylamide, a probable carcinogen and obesogen, is a by-product in foods prevalent in the obesity-inducing Western diet. Acrylamide is metabolized by cytochrome P450 2E1 (CYP2E1) to the genotoxic epoxide, glycidamide, and is associated with an increased risk for breast cancer. To investigate how acrylamide and obesity interact to increase breast cancer risk, female mice were fed a low-fat (LFD) or high-fat diet (HFD) and control water or water supplemented with acrylamide at levels similar to the average daily exposure in humans. While HFD significantly enhanced weight gain in mice, the addition of acrylamide did not significantly alter body weights compared to respective controls. Mammary epithelial cells from obese, acrylamide-treated mice had increased DNA strand breaks and oxidative DNA damage compared to all other groups. In vitro, glycidamide-treated COMMA-D cells showed significantly increased DNA strand breaks, while acrylamide-treated cells demonstrated significantly higher levels of intracellular reactive oxygen species. The knockdown of CYP2E1 rescued the acrylamide-induced oxidative stress. These studies suggest that long-term acrylamide exposure through foods common in the Western diet may enhance DNA damage and the CYP2E1-induced generation of oxidative stress in mammary epithelial cells, potentially enhancing obesity-induced breast cancer risk.