Cell Model Research Articles

Exploring the Mechanisms of Iron Overload-Induced Liver Injury in Rats Based on Transcriptomics and Proteomics

Iron is a trace element that is indispensable for the growth and development of animals. Excessive iron supplementation may lead to iron overload and elevated reactive oxygen species (ROS) production in animals, causing cellular damage. Nevertheless, the precise mechanism by which iron overload causes cell injury remains to be fully elucidated. In this study, 16 male SD rats aged 6 to 7 weeks were randomly assigned to either a control group (CON) or an iron overload group (IO). Rats in the iron overload group received 150 mg/kg iron dextran injections every three days for a duration of four weeks. The results indicated that iron treatment with iron dextran significantly increased the scores of steatosis (p < 0.05) and inflammation (p < 0.05) in the NAS score. The integrated transcriptomic and proteomic analysis suggests that HO-1 and Lnc286.2 are potentially significant in iron overload-induced liver injury in rats. In vitro experiments utilizing ferric ammonium citrate (FAC) were conducted to establish an iron overload model in rat liver-derived BRL-3A cells. The result found that FAC treatment can significantly increase the BRL-3A cell’s Fe2+ content (p < 0.05), ROS (p < 0.01), lipid ROS (p < 0.01) levels, and the expression of the HO-1 gene and protein (p < 0.01), aligning with proteomic and transcriptomic findings. HO-1 inhibition can significantly decrease BRL-3A cell vitality (p < 0.01) and promote ROS (p < 0.05) and lipid ROS (p < 0.01), thus aggravating FAC-induced BRL-3A cell iron overload damage. Using the agonist of HO-1 agonist cobalt protoporphyrin (CoPP) to induce HO-1 overexpression can significantly alleviate the decrease in FAC-induced BRL-3A cell viability (p < 0.01), ROS (p < 0.01), and lipid ROS (p < 0.01). In addition, siLnc286.2 treatment can increase HO-1 expression, alleviate the decline of FAC-induced BRL-3A cell activity, and increase lipid ROS (p < 0.05) content. In conclusion, the findings of this study suggest that by suppressing the expression of Lnc286.2, we can enhance the expression of HO-1, which in turn alleviates lipid peroxidation in cells and increases their antioxidant capacity, thereby exerting a protective effect against liver cell injury induced by iron overload.

Highly Deformable Phototunable Viscoelastic Fluid Interface Modulates Cellular Adaptive Wetting Behavior

AbstractEmerging evidence shows that the viscoelastic cues of the e Ixtracellular matrix (ECM) regulate cellular functions and fates. However, as cells are viscoelastic, force dissipation occurs within themselves as well as the ECM side, implying the existence of reciprocal viscous regulation between the two. Here, a fluid‐based scaffold with tunable viscoelasticity has been developed to investigate its impact on the cell adhesion process. The platform is based on the water–perfluorocarbon interface decorated with diacetylene‐based phospholipid membranes (IPLMs), whose viscoelasticity can be systematically manipulated by photocrosslinking. Further introduction of a cell‐adhesive peptide and fluorescent tag allows cell adhesion at the highly deformable fluid interface and confocal observation of dynamic cell–model ECM interactions. The viscoelasticity‐tunability is confirmed by fluorescence recovery after photobleaching, interfacial rheology, and atomic force microscopy nanoindentation. Cells seeded at the IPLM exhibit so‐called adaptive wetting, where the interface first deforms toward the out‐of‐plane direction before cellular dimensional changes, followed by cellular flattening and interfacial restoration. Furthermore, the quantification of these parameters reveals a biphasic response against the crosslinking levels, which indicates that the cell‐ECM viscosity balance determines adaptive wetting phenotypes. The platform may enable the prediction of dynamic adhesion responses in physiological and pathological processes.

Targeting Fibroblast-Derived Interleukin 6: A Strategy to Overcome Epithelial-Mesenchymal Transition and Radioresistance in Head and Neck Cancer

Background: Cancer-associated fibroblasts have been reported to play a central role in driving cancer progression, promoting metastasis, and conferring resistance to therapy in HNSCC. Methods: Indirect and direct co-culture models of HPV-positive and HPV-negative HNSCC cells with fibroblasts were developed to study the effect of fibroblasts on cancer cells. ELISA was used to measure IL-6 secretion in these models. To dissect the underlying signalling mechanisms, the effects of IL-6, an IL-6 receptor (IL-6R) inhibitor, a MAPK/ERK inhibitor, and a JAK/STAT inhibitor were evaluated. Epithelial-to-mesenchymal transition (EMT) was assessed by measuring EMT markers and conducting scratch assays and spheroid assays. Radioresistance was evaluated using clonogenic assays. Additionally, radioresistant (RR) cell lines were established from parental cells to examine the correlation between radioresistance and EMT. Results: Fibroblasts were found to drive EMT-like changes and heightened radioresistance in HNSCC cells through IL-6 secretion. Remarkably, these Fb-driven effects were robustly reversed using IL-6R and MAPK/ERK inhibitors in both HPV-positive and HPV-negative cell lines, whereas JAK/STAT inhibitors proved effective only in HPV-negative cells. RR cell lines exhibit a more aggressive phenotype than their parental counterparts, marked by pronounced EMT features and heightened resistance to radiotherapy. Importantly, these aggressive characteristics were substantially attenuated by targeting IL-6R or MAPK/ERK pathways. Conclusions: This study highlights the critical role of fibroblast-secreted IL-6 in driving and maintaining EMT and radioresistance in HNSCC, resulting in a more aggressive tumour phenotype. Targeting the IL-6/IL-6R/ERK pathway emerges as a promising therapeutic approach for combating CAF-driven tumour progression and improving clinical outcomes in patients with aggressive, therapy-resistant HNSCC.

Aging-enhanced autophagy activity promotes fibrotic progression via the TGF-β2/Smad signaling pathway in trabecular meshwork cells—a new insight from POAG

IntroductionGlaucoma, a leading cause of irreversible blindness, is characterized by optic neuropathy and retinopathy, with primary open-angle glaucoma (POAG) being the most prevalent form. The primary pathogenic mechanism of POAG involves elevated intraocular pressure caused by chronic fibrosis of the trabecular meshwork (TM). Autophagy, a critical process for maintaining cellular homeostasis, has been implicated in fibrosis across various organs. However, its precise role in the fibrosis associated with POAG pathogenesis remains unclear. This study investigates the involvement of autophagy in TM fibrosis and explores its potential impact on POAG development, aiming to provide insights into new therapeutic targets.MethodsTo assess autophagy activity and its relationship with fibrosis, we analyzed TM tissues from POAG patients and healthy donors. Autophagic activity in human TM tissues was measured through immunohistochemical analyses. An in vitro aging model using chronic H2O2 treatment was established to investigate the change of fibrosis in TM cells. Additionally, we used dexamethasone-treated TM cells as a POAG model to explore the role of autophagy in fibrotic progression. The involvement of the TGF-β2/Smad signaling pathway was investigated through western blot analysis and quantitative real-time PCR.ResultsThis study reveals increased autophagic activity in tissues from POAG patients and an age-related upregulation of autophagy in healthy human TM tissues. In the H2O2-induced aging model, TM cells displayed both elevated autophagic activity and fibrosis. Further investigation showed that enhanced autophagy activity promoted fibrotic progression via activation of the TGF-β2/Smad signaling pathway. Similarly, in the dexamethasone-treated TM cell model, autophagy was found to exacerbate fibrosis, aligning with observations in the aging model.DiscussionIn this study, we uncover the interplay between autophagy and the TGF-β2/Smad pathway in the pathogenesis of POAG. We observed increased autophagic activity in TM tissues from POAG patients and in TM tissues of aging healthy individuals. In human primary TM cells, we confirmed that autophagy becomes activated in the context of cellular senescence and the development of POAG, which further facilitates fibrotic progression via the TGF-β2/Smad signaling pathway. These findings underscore the important role of autophagy in POAG pathogenesis and confirm senescence as a pivotal risk factor.

Using cortical organoids to understand the pathogenesis of malformations of cortical development

Malformations of cortical development encompass a broad range of disorders associated with abnormalities in corticogenesis. Widespread abnormalities in neuronal formation or migration can lead to small head size or microcephaly with disorganized placement of cell types. Specific, localized malformations are termed focal cortical dysplasias (FCD). Neurodevelopmental disorders are common in all types of malformations of cortical development with the most prominent being refractory epilepsy, behavioral disorders such as autism spectrum disorder (ASD), and learning disorders. Several genetic pathways have been associated with these disorders from control of cell cycle and cytoskeletal dynamics in global malformations to variants in growth factor signaling pathways, especially those interacting with the mechanistic target of rapamycin (mTOR), in FCDs. Despite advances in understanding these disorders, the underlying developmental pathways that lead to lesion formation and mechanisms through which defects in cortical development cause specific neurological symptoms often remains unclear. One limitation is the difficulty in modeling these disorders, as animal models frequently do not faithfully mirror the human phenotype. To circumvent this obstacle, many investigators have turned to three-dimensional human stem cell models of the brain, known as organoids, because they recapitulate early neurodevelopmental processes. High throughput analysis of these organoids presents a promising opportunity to model pathophysiological processes across the breadth of malformations of cortical development. In this review, we highlight advances in understanding the pathophysiology of brain malformations using organoid models.

A review of 3D bioprinting for organoids

Abstract Current two-dimensional (2D) cell models for effective drug screening suffer from significant limitations imposed by the lack of realism in the physiological environment. Three-dimensional (3D) organoids models hold immense potential in mimicking the key functions of human organs by overcoming the limitations of traditional 2D cell models. However, current techniques for preparation of 3D organoids models had limitations in reproducibility, scalability, and the ability to closely replicate the complex microenvironment found in vivo. Additionally, traditional 3D cell culture systems often involve lengthy and labor-intensive processes that hinder high-throughput applications necessary for a large-scale drug screening. Advancements in 3D bioprinting technologies offer promising solutions to these challenges by enabling precise spatial control over cell placement and material composition, thereby facilitating the creation of more physiologically relevant organoids than current techniques. This review provides a comprehensive summary of recent advances in 3D bioprinting technologies for creating organoids models, which begins with an introduction to different types of 3D bioprinting techniques (especially focus on volumetric bioprinting (VBP) technique), followed by an overview of bioinks utilized for organoids bioprinting. Moreover, we also introduce the applications of 3D bioprinting organoids in disease models, drug efficiency evaluation and regenerative medicine. Finally, the challenges and possible strategies for the development and clinical translation of 3D bioprinting organoids are concluded.

Irisin and Metastatic Melanoma: Selective Anti-Invasiveness Activity in BRAF Wild-Type Cells

Irisin is a newly discovered 12 kDa messenger protein involved in energy metabolism. Irisin affects signaling pathways in several types of cancer; however, the role of irisin in metastatic melanoma (MM) has not been described yet. We explored the biological effects of irisin in in vitro models of MM cells (HBLwt/wt, LND1wt/wt, Hmel1V600K/wt and M3V600E/V600E) capable of the oncogenic activation of BRAF. We treated MM cells with different concentrations of r-irisin (10 nM, 25 nM, 50 nM, 100 nM) for 24 h–48 h. An MTT assay highlighted that r-irisin did not affect the proliferation of MM cells. We subsequently treated MM cells with 10 nM r-irisin, corresponding to the dose exhibiting biological activity in vitro. Irisin reduced the invasive ability of only LND1wt/wt (p < 0.05), which highly expressed αv gene levels, but did not affect the invasion of BRAFmut cells. Gelatin zymography analysis showed a reduction in the enzymatic activity of MMP-2 and MMP-9 in BRAFwt/wt cells treated with 10 nM r-irisin. Moreover, gene expression analysis (qPCR) of MMP-2 and MMP-9 and of the fibrinolytic system (uPAR, uPA and PAI-1) highlighted a crucial role of 10 nM r-irisin treatment in the inhibition of pro-invasive systems in BRAFwt/wt. In conclusion, our results may suggest a possible differential role of irisin in melanoma cells.

Prospects of elafibranor in treating alcohol-associated liver diseases

Alcohol-related liver disease (ALD), which is induced by excessive alcohol consumption, is a leading cause of liver-related morbidity and mortality. ALD patients exhibit a spectrum of liver injuries, including hepatic steatosis, inflammation, and fibrosis, similar to symptoms of nonalcohol-associated liver diseases such as primary biliary cholangitis, metabolic dysfunction-associated steatotic liver disease, and nonalcoholic steatohepatitis. Elafibranor has been approved for the treatment of primary biliary cholangitis and has been shown to improve symptoms in both animal models and in vitro cell models of metabolic dysfunction-associated steatotic liver disease and nonalcoholic steatohepatitis. However, the efficacy of elafibranor in treating ALD remains unclear. In this article, we comment on the recent publication by Koizumi et al that evaluated the effects of elafibranor on liver fibrosis and gut barrier function in an ALD mouse model. Their findings indicate the potential of elafibranor for ALD treatment, but further experimental investigations and clinical trials are warranted.

Study on the Synergistic Effect of Klotho and KRAS on Reducing Ferroptosis After Myocardial Infarction by Regulating RAP1/ERK Signaling Pathway.

Myocardial infarction (MI) is a coronary artery-related disease that seriously threatens human life and is the leading cause of sudden death worldwide, where a lack of nutrients and oxygen leads to an inflammatory response and death of cardiomyocytes. Ferroptosis is a form of non-apoptotic cell death associated with metabolic dysfunction, resulting in abnormal breakdown of glutamine and iron-dependent accumulation of reactive oxygen species (ROS) during metabolism. However, the molecular mechanism of ferroptosis in the pathogenesis of MI and the function of Klotho and KRAS on ferroptosis during MI remain unclear. The MI rat model was established by LAD ligation with a 6-0 suture. H9c2 cells were placed in glucose-deficient DMEM (Thermo) and cultured in an anaerobic environment (1% CO2 and 5% CO) to establish an in vitro OGD cell model. The damage to rat heart tissue was detected by HE staining, and Klotho and KRAS were determined by RT-qPCR, Western Blot, and IHC. TUNEL staining was used to determine apoptosis in rat heart tissue samples. The interaction between Klotho and KRAS was verified by co-immunoprecipitation and Western Blot. The cardiomyocyte activity was measured by CCK-8 assay. LDH, CK-MB, cTnT, and Fe2+ markers were detected by the kits. For the assessment of ferroptosis, GSH and ROS in cardiomyocytes and serum were detected by kits, and PTSG was detected by Western Blot. IL-1β and IL-6 in cardiomyocytes and serum were determined by ELISA. Klotho was downregulated in MI. Downregulation of Klotho promoted myocardial injury; increased apoptosis of cardiomyocytes; promoted LDH, CK-MB, and cTnT concentrations; inhibited GSH; and promoted ROS levels, PTGS2 expression, and ferroptosis in rats. The same results were obtained in vitro. Klotho and KRAS had endogenous interactions. KRAS knockdown can reverse Klotho knockdown-mediated MI and ferroptosis. RAP1/ERK pathway was highly expressed in MI, and inhibiting RAP1/ERK pathway activation can reverse the promoting effect of overexpressed KRAS on MI progression and ferroptosis. Klotho interacts with KRAS and inhibits ferroptosis after MI by regulating the RAP1/ERK pathway.

Identification, cellular localization, and function analysis of a novel toll‐like receptor (PtToll4) in Portunus trituberculatus

AbstractToll‐like receptor (TLR) is one of the most important pattern recognition receptors in both invertebrate and vertebrate animals. In previous study, three different kinds of TLR cDNAs (PtToll1‐3) have been cloned from Portunus trituberculatus. In this study, a novel PtToll4 cDNA sequence was identified in this crab. The PtToll4 ORF is predicted to encode 894 peptides with an initiating signal peptide, an extracellular LRRs domain, a transmembrane region, and an intracellular toll‐interleukin‐1 receptor domain. Based on the sequence and phylogenetic analysis, PtToll4 distinctly clustered with almost all crustacean tolls, which mostly clustered with some shrimp TLRs and Eriocheir sinensis Toll 2 (EsToll 2). The real‐time quantitative polymerase chain reaction analysis showed that the transcript of PtToll4 was constitutively expressed in tissues. In hemocytes, the transcript of PtToll4 in large granulocytes is approximately threefold higher than in small granulocytes and hyalocytes. Moreover, the expression of PtToll4 could be moderately and shortly upregulated by Vibrio alginolyticus or lipopolysaccharide challenge in the primary cultured hemocytes. In HEK 293T cell model, over‐expressed PtToll4 was not able to activate the mammal NF‐κB, compared with a positive plasmid. In the Drosophila S2 cell model, over‐expressed PtToll4 distributed mainly in cell membrane and could affect the activation of some Drosophila antimicrobial peptides promoters. The RNA interference experiment showed that the expressions of PtToll4 was significantly inhibited by its specific double‐stranded RNA in the crab hemocytes and accompanied by the suppressed expressions of the anti‐lipopolysaccharide factor 4 (ALF4), ALF7, hyastatin3, crustin1, and crustin3.

Anti-atherosclerotic effects of LncRNA NEXN-AS1 by regulation of the canonical inflammasome pathway of pyroptosis via NEXN.

. Pyroptosis is closely related to many chronic diseases including atherosclerosis, but the potential pathomechanisms are still unclear. This research aimed to explore how lncRNAs may contribute to pyroptosis and the potential mechanisms. . We performed in vitro assays to investigate the effects of a relatively newly discovered lncRNA, NEXN-AS1, on pyroptosis. Two types of vascular wall cells, namely, human vascular smooth muscle cells (VSMCs) and endothelial cells (ECs), were used as cell models in this study. A previously constructed lentivirus vector was used to overexpress lncRNA NEXN-AS1 and a small interfering RNA (siRNA) mimic against NEXN to knockdown NEXN. The mRNA and protein levels of molecules of pyroptosis in the canonical inflammasome pathway, including nucleotide-binding oligomerization segment-like receptor family 3 (NLRP3), caspase-1, gasdermin D (GSDMD), interleukin-1β (IL-1β), and interleukin-18 (IL-18) were detected using quantitative real-time PCR and western blot analysis, respectively. . We observed that lentivirus-mediated overexpression of lncRNA NEXN-AS1 increased the expression levels of NEXN and markedly reduced the expression of critical molecules involved in pyroptosis, including NLRP3, caspase-1, GSDMD, IL-1β, and IL-18, in both VSMCs and ECs. Furthermore, NEXN knockdown could reverse the effects of lncRNA NEXN-AS1 overexpression on pyroptosis. lncRNA NEXN-AS1 could act as an target for maintaining endothelium homeostasis, plaque stability, and delaying the progression of atherosclerosis.

Nitrogen doped carbon dots for in vitro intracellular redox modulation via optical stimulation.

Carbon dots (CDs) are promising candidates as oxygen photosensitizers, in cancer therapeutic applications due to their high quantum yield, superior chemical and photostability, low cytotoxicity and ease of chemical functionalization/tuning. Nitrogen doping can further improve oxygen photosensitization performance. Besides photodynamic therapy, however, the possibility to finely and remotely regulate the intracellular redox balance by using physical stimuli has been attracting more and more interest not only for nanotheranostic application, but also as a novel, fully biocompatible therapeutic tool. Here, we report on the synthesis of nitrogen-doped CDs by solvothermal methods starting from abundant, bioderived, low-cost precursors, and we characterize their interface with in vitro cultures of human embryonic kidney (HEK-293) cells, a widely accepted model of non-tumoral cells. While not affecting cell proliferation, synthesized CDs efficiently modulate, under visible light and physiological eustress conditions, intracellular calcium ion dynamics and reactive oxygen species concentration, resulting in a 4-fold increase. The reported results may broaden the application of CDs beyond photodynamic therapy, unveiling new opportunities in the field of redox medicine assisted by carbon-based nanomaterials and optical stimulation.

The Role of RyR2 Mutations in Congenital Heart Diseases: Insights Into Cardiac Electrophysiological Mechanisms.

Ryanodine receptor 2 (RyR2) protein, a calcium ion release channel in the sarcoplasmic reticulum (SR) of myocardial cells, plays a crucial role in regulating cardiac systolic and diastolic functions. Mutations in RyR2 and its dysfunction are implicated in various congenital heart diseases (CHDs). Studies have shown that mutations in the RYR2 gene, which encodes the RyR2 protein, are linked to several cardiac arrhythmias, including catecholaminergic polymorphic ventricular tachycardia (CPVT), long QT syndrome (LQTS), calcium release deficiency syndrome (CRDS), and atrial fibrillation (AF). Additionally, RyR2 mutations have been associated with multiple genetic cardiomyopathies, such as left ventricular non-compaction cardiomyopathy (LVNC), arrhythmogenic right ventricular cardiomyopathy (ARVC), hypertrophic cardiomyopathy (HCM) and dilated cardiomyopathy (DCM). Through various cell and animal models, researchers have developed mutant RyR2 models demonstrated that these mutations often lead to calcium dysregulation, typically resulting in either a gain or loss of function. This comprehensive review delves into the current understanding of RyR2 mutations and their impact on cardiac electrophysiology, focusing on the molecular mechanisms linking these mutations to arrhythmias and cardiomyopathies-an essential step in advancing diagnostic and therapeutic strategies.

Predicting the Anti-SARS-CoV-2 Potential of Isoquinoline Alkaloids from Brazilian Siparunaceae Species Using Chemometric Tools

The COVID-19 pandemic has caused over 7 million deaths globally in the past four years. Siparuna spp. (Siparunaceae), which is used in Brazilian folk medicine, is considered a genus with potential antiviral alternatives. This study explored the correlation between phytochemicals in Siparuna leaf extracts (S. ficoides, S. decipiens, S. glycycarpa, S. reginae, and S. cymosa) and their potential against various SARS-CoV-2 targets. In vitro assays examined interactions between the spike protein and the ACE2 receptor, protease activity, and viral replication inhibition in Calu-3 cell models. UHPLC-MS/MS analysis, processed with MZmine and evaluated chemometrically, revealed isoquinoline alkaloids with bulbocapnine, showing promising therapeutic potential. Predictions regarding absorption, distribution, metabolism, excretion, and toxicity were conducted, along with molecular docking and dynamics simulations, to evaluate protein−ligand interaction stability. The results confirmed the antiviral activity of the Siparuna genus against SARS-CoV-2 targets, with 92% of the extracts maintaining over 70% cellular viability at 200 μg·mL−1 and 80% achieving more than 50% viral activity suppression at 50 μg·mL−1. These findings highlight the potential of isoquinoline alkaloids as novel anti-coronavirus agents and support the need for further exploration, isolation, and testing of Siparuna compounds in the fight against COVID-19.

Morphology remodelling and membrane channel formation in synthetic cells via reconfigurable DNA nanorafts.

The shape of biological matter is central to cell function at different length scales and determines how cellular components recognize, interact and respond to one another. However, their shapes are often transient and hard to reprogramme. Here we construct a synthetic cell model composed of signal-responsive DNA nanorafts, biogenic pores and giant unilamellar vesicles (GUVs). We demonstrate that reshaping of DNA rafts at the nanoscale can be coupled to reshaping of GUVs at the microscale. The nanorafts collectively undergo reversible transitions between isotropic and short-range local order on the lipid membrane, programmably remodelling the GUV shape. Assisted by the biogenic pores, during GUV shape recovery the locally ordered DNA rafts perforate the membrane, forming sealable synthetic channels for large cargo transport. Our work outlines a versatile platform for interfacing reconfigurable DNA nanostructures with synthetic cells, expanding the potential of DNA nanotechnology in synthetic biology.

Naringenin Inhibits Ferroptosis in Renal Tubular Epithelial Cells of Diabetic Nephropathy Through SIRT1/FOXO3a Signaling Pathway.

Naringenin has the potential to regulate ferroptosis and mitigate renal damage in diabetic nephropathy (DN). However, it remains unclear whether the naringenin's effects in DN are linked to its ability to regulate ferroptosis. This study investigated the potential anti-ferroptosis properties of naringenin in high glucose (HG)-induced renal tubular epithelial cell models. HK-2 cells were cultured in HG medium to establish the DN cell model. HK-2 cells were treated with different doses of naringenin to explore the effect of naringenin. The CCK-8 results show that 50 μM ~ 200 μM of naringenin do not affect the viability of HK-2 cells and the viability of HG-induced HK-2 cells increase in a dose-dependent manner with naringenin treatment. Additionally, naringenin increased the levels of IL-10 while decreasing the levels of IL-1β, TNF-α, IL-6, and ROS in HG-induced HK-2 cells. Naringenin also reduced the levels of Fe2+, oxidized lipid ROS, MDA, 4-HNE, ACSL4, and TFR1 in HG-induced HK-2 cells, while increasing the levels of non-oxidized lipid ROS, SOD, GSH-Px, SLC7A11, and GPX4. Meanwhile, naringenin restored the levels of MMP, ATP and MPTP opening, reduced OCR in HG-induced HK-2 cells. Furthermore, naringenin reversed the decreased expression of SIRT1, p-FOXO3a, Nrf2 and Nuclear Nrf2 caused by HG. SIRT1 inhibitor EX527 and Nrf2 inhibitor ML385 attenuated the effects of naringenin on ferroptosis in HG-induced HK-2 cells, with EX527 demonstrating a stronger reversal effect on ferroptosis than ML385. These results suggest that naringenin inhibits ferroptosis in HG-induced HK-2 cells mainly through SIRT1/FOXO3a signaling pathway. This finding further enhanced our understanding of the mechanism behind naringenin's protective effect on DN.

Passion fruit seed extract protects hydrogen peroxide-induced cell damage in human retinal pigment epithelium ARPE-19 cells

Age-related macular degeneration (AMD) is a major cause of vision loss among adults. We investigated the protective effects of passion fruit seed extract (PFSE) and its rich polyphenol piceatannol in an AMD cell model in which human retinal pigment epithelial ARPE-19 cells were exposed to hydrogen peroxide (H2O2). Using a cell viability WST-8 assay, we revealed that PFSE and piceatannol increased the cellular viability of ARPE-19 cells by 130% and 133%, respectively. Moreover, PFSE and piceatannol recovered the cell viability of ARPE-19 cells, which had decreased to 60% owing to H2O2-induced damage, to approximately 84% and 89%, respectively. In addition, we found that the treatment of ARPE-19 cells with H2O2 decreased the mitochondrial and glycolytic ATP production rate to approximately 54% that of healthy control ARPE-19 cells using a Seahorse extracellular flux analyzer. Furthermore, pretreatment with PFSE and piceatannol restored the oxidative stress-induced decrease in the mitochondrial and glycolytic ATP production rate to approximately 97% and 82%, respectively. These results indicated the cytoprotective effects of PFSE and piceatannol against oxidative stress in human ARPE-19 cells by resolving the dysfunction of mitochondrial and glycolytic energy metabolism.

Comprehensive Metabolomics in Mouse Mast Cell Model of Allergic Rhinitis for Profiling, Modulation, Semiquantitative Analysis, and Pathway Analysis

Allergic rhinitis affects millions globally, causing significant discomfort and reducing the quality of life. This study investigates the metabolic alterations in murine mast cells (MC/9) under allergic rhinitis conditions induced by lipopolysaccharide (LPS) stimulation, employing UHPLC-QTOF-MS-based untargeted and targeted metabolomics. The analysis identified 44 significantly regulated metabolites, including histamine, leukotrienes, prostaglandins, thromboxanes, and ceramides. Key metabolic pathways such as arachidonic acid, histidine, and sphingolipid metabolisms were notably modulated. The study further examined the therapeutic effects of triprolidine and zileuton, demonstrating their capacity to reverse LPS-induced metabolic shifts. Triprolidine primarily modulated histidine and sphingolipid metabolism, while zileuton targeted arachidonic acid and sphingolipid metabolism. These findings underscore the utility of metabolomics analysis in elucidating the complex biochemical pathways involved in allergic rhinitis and highlight the potential of metabolomics for evaluating therapeutic interventions. This study enhances our understanding of mast cell metabolism in allergic responses and provides a robust model for assessing the efficacy of anti-allergic agents, paving the way for more effective treatments.

High glucose induces renal tubular epithelial cell senescence by inhibiting autophagic flux.

Autophagy, a cellular degradation process involving the formation and clearance of autophagosomes, is mediated by autophagic proteins, such as microtubule-associated protein 1 light chain 3 (LC3) and sequestosome 1 (p62), and modulated by 3-methyladenine (3-MA) as well as chloroquine (CQ). Senescence, characterised by permanent cell cycle arrest, is marked by proteins such as cyclin-dependent kinase inhibitor 1 (p21) and tumour protein 53 (p53). This study aims to investigate the relationship between cell senescence and renal function in diabetic kidney disease (DKD) and the effect of autophagy on high-glucose-induced cell senescence. We categorised 46 patients with DKD diagnosed by renal biopsy into classes I, IIa, IIb, III and IV and used four normal kidney specimens from patients with renal trauma as controls. We evaluated pathological changes, LC3 and p21. We used streptozotocin-induced DKD models in rats and 35mM glucose-cultured human proximal tubular epithelial cells (HK-2) with or without 3-MA and CQ. We assessed p53, p21, LC3 and p62. We observed autophagosomes and detected senescence-associated galactosidase (SA-β-gal) activity. In patients with DKD, p21 and LC3 expression levels increased over time and correlated positively with blood creatinine and proteinuria. In DKD rats and HK-2 cells, p21, p53, LC3 and p62 expression levels were higher than in the controls, as were SA-β-gal-positive cells, renal tubular autophagosomes and co-expression of p21 and LC3. The 3-MA reduced p16, p21 and p53 expression compared with the high glucose group, whereas CQ had the opposite effect. These results suggest that renal tubular cell senescence is associated with the progression of DKD. Additionally, autophagic flux may play a role in mediating high-glucose-induced senescence in renal tubular cells.

IL-22-mediated microRNA-124-3p/GRB2 axis regulates hyperproliferation and inflammatory response of keratinocytes in psoriasis.

Psoriasis is an inflammatory dermatosis that features overproliferation and inflammatory reaction of keratinocytes. A study reported that IL-22 is involved in the pathogenesis of psoriasis by mediating miR-124 to regulate the expression of fibroblast growth factor receptor 2 in keratinocytes. A microRNA may target multiple target genes. Therefore, we speculate that miR-124-3p may also target other downstream genes to affect IL -22-induced keratinocyte function. A possible target gene of miR-124-3p, growth factor receptor-bound protein 2 (GRB2), was screened by analyzing the target gene databases. GRB2 expression was elevated and miR-124-3p expression was decreased in psoriatic lesions compared to psoriatic adjacent normal skins and healthy controls. We performed the following cell experiments in the IL-22-stimulated HaCaT cell model. In keratinocytes transfected with the miR-124-3p mimics, GRB2 expression was significantly lower. We analyzed the regulation of keratinocyte proliferation by GRB2 and miR-124-3p. High levels of GRB2 promoted keratinocyte proliferation and expression of Ki67, PCNA, and K16, which were inhibited by low expression of GRB2. In addition, we found that the effect of GRB2 inhibitors on the proliferation and inflammatory response of keratinocytes was dose-dependent. Finally, we investigated the influence of GRB2 on inflammatory mediators in keratinocytes with the ELISA. After low expression of GRB2, the mRNA expression and secretion of the pro-inflammatory factor were suppressed. When both GRB2 and miR-124-3p were overexpressed, the cellular overproliferation and inflammation caused by GRB2 overexpression were significantly reversed by miR-124-3p. In summary, IL-22-mediated miR-124-3p regulates keratinocyte hyperproliferation and inflammatory response by suppressing GRB2 expression in psoriasis.