Cell System Research Articles

Mutant p53 reactivators protect breast cancer cells from ferroptosis.

Ferroptosis is a novel nonapoptotic form of cell death characterized by iron-dependent reactive oxygen species-mediated lipid peroxidation. In several different cell systems, the tumor suppressor p53 can enhance sensitivity to ferroptotic inducers. At least half of all human cancers show loss of function of p53. Furthermore, many of those tumors express mutant forms of p53 that has lost its wild-type function. Several groups have designed small molecules that can reactivate the wild-type function of these missense p53 mutants. We reasoned that p53 reactivators may also enhance sensitivity of certain cancer cells to ferroptosis stimuli. To test this idea we combined a number of different p53 reactivators with small molecule inducers of ferroptosis. In contrast, we observed that several p53 reactivators protected cells from cell death induced by ferroptotic inducers. Surprisingly, this protection still occurred in p53-null cell lines. We observed that these reactivators were neither free radical scavengers nor ion chelators. One of these p53 reactivator molecules, NSC 59984, reduced expression of GPX4, which is unlikely to explain its ability to reduce sensitivity to ferroptosis. We suggest that these p53 reactivators function via an unknown, p53-independent manner to suppress ferroptosis.

Biodegradable pyroptosis inducer with multienzyme-mimic activity kicks up reactive oxygen species storm for sensitizing immunotherapy

Triggering pyroptosis is a major new weathervane for activating tumor immune response. However, biodegradable pyroptosis inducers for the safe and efficient treatment of tumors are still scarce. Herein, a novel tumor microenvironment (TME)-responsive activation nanoneedle for pyroptosis induction, copper-tannic acid (CuTA), was synthesized and combined with the sonosensitizer Chlorin e6 (Ce6) to form a pyroptosis amplifier (CuTA-Ce6) for dual activation and amplification of pyroptosis by exogenous ultrasound (US) and TME. It was demonstrated that Ce6-triggered sonodynamic therapy (SDT) further enhanced the cellular pyroptosis caused by CuTA, activating the body to develop a powerful anti-tumor immune response. Concretely, CuTA nanoneedles with quadruple mimetic enzyme activity could be activated to an “active” state in the TME, destroying the antioxidant defense system of the tumor cells through self-destructive degradation, breaking the “immunosilent” TME, and thus realizing the pyroptosis-mediated immunotherapy with fewer systemic side effects. Considering the outstanding oxygen-producing capacity of CuTA and the distinctive advantages of US, the sonosensitizer Ce6 was attached to CuTA via an amide reaction, which further amplified the pyroptosis and sensitized pyroptosis-induced immunotherapy with the two-pronged strategy of CuTA enzyme-catalyzed cascade and US-driven SDT pathway to generate a “reactive oxygen species (ROS) storm”. Conclusively, this work provided a representative paradigm for achieving safe, reliable and efficient pyroptosis, which was further enhanced by SDT for more robust immunotherapy.

Fuel Cell Electric Vehicle Characterisation under Laboratory and In-use Operation

Fuel Cell Electric Vehicles (FCEVs) are one of the main options considered as possible future alternatives to conventional vehicles. The paper evaluates a light-duty FCEV under both laboratory and on-road conditions. Hydrogen consumption and vehicle efficiency were measured over various driving and temperature conditions. Results show hydrogen consumption between 0.975kg/100km and 1.5kg/100km, close to the officially declared value. A new extended formulation of vehicle efficiency is proposed to consider different energy streams in the vehicle. Vehicle indicative efficiency remained between 50 and 60% on the majority of cases. The paper also investigates the feasibility of applying a fuel cell system of similar characteristics for commercial urban and rural applications, the Vehicle Energy Consumption Calculation TOol (VECTO), as a possible alternative solution to a battery electric equivalent vehicle.

Paracetamol degradation in a dual-chamber rectangular membrane bioreactor using microbial fuel cell system with a microbial consortium from sewage sludge

Paracetamol degradation in a dual-chamber rectangular membrane bioreactor using microbial fuel cell system with a microbial consortium from sewage sludge

Fault diagnosis method of PEMFC system based on ensemble learning

Due to the coupling of multiple physical fields inside proton exchange membrane fuel cell (PEMFC) and the complex and changeable external application scenarios, the faults of PEMFC system greatly affects its engineering application. The paper proposes an ensemble learning-based fault diagnosis method for PEMFC systems, utilizing five distinct algorithms to develop an integrated ensemble classifier for fault detection. The original sample dataset collected from fuel cells undergoes a process of deep feature extraction to obtain a composite feature sample dataset, which is then inputted into an ensemble classifier to establish a fault diagnosis model. The experimental data of four kinds of faults of 80 W fuel cell test bench, EC fuel cell system and 50 W water-cooled fuel cell test bench are analyzed by using this classification model. The results show that the accuracy and generalization of the ensemble learning algorithm for different fault diagnosis of PEMFC system in different application environments are verified.

A Mouse Model of X-Linked Chronic Granulomatous Disease for the Development of CRISPR/Cas9 Gene Therapy

Chronic granulomatous disease (CGD) is an inherited immunodeficiency disease mainly caused by mutations in the X-linked CYBB gene that abrogate reactive oxygen species (ROS) production in phagocytes and microbial defense. Gene repair using the CRISPR/Cas9 system in hematopoietic stem and progenitor cells (HSPCs) is a promising technology for therapy for CGD. To support the establishment of efficient and safe gene therapies for CGD, we generated a mouse model harboring a patient-derived mutation in the CYBB gene. Our CybbC517del mouse line shows the hallmarks of CGD and provides a source for Cybb-deficient HSPCs that can be used to evaluate gene-therapy approaches in vitro and in vivo. In a setup using Cas9 RNPs and an AAV repair vector in HSPCs, we show that the mutation can be repaired in 19% of treated cells and that treatment restores ROS production by macrophages. In conclusion, our CybbC517del mouse line provides a new platform for refining and evaluating novel gene therapies and studying X-CGD pathophysiology.

A real-time hydrogen consumption estimation method for fuel cell vehicles

ABSTRACT As an economic evaluation indicator for fuel cell vehicles (FCVs), real-time hydrogen consumption’s accuracy and immediateness are important. Real-time hydrogen consumption obtained through the fuel cell system (FC Power method) has strong immediateness but poor accuracy, while through the hydrogen mass (Mass method) has good accuracy but poor immediateness. Given the difficulty in obtaining sufficient training data on actual vehicles, this paper establishes a hydrogen storage system model whose input and output can be used to obtain data. Then, a new estimation method based on the prediction of the mass’s difference (PMD method) has been constructed based on the Neural basis expansion analysis with exogenous variables (NBEATSx) algorithm. After the NBEATSx algorithm is trained, the prediction accuracy of the PMD method is verified under urban and high-speed conditions. The result shows that the new method proposed in this article improves Root Mean Square Error (RMSE) and Mean Absolute Error (MAE) by nearly 45% compared to the FC Power method, and the hydrogen consumption’s sampling point is advanced compared to the Mass method. The trained model can be input into Hydrogen Management System (HMS) which is installed on the hydrogen storage system and output data in real-time.

Treatment of nitrogenous wastewater by a constructed wetland-coupled microbial fuel cell system: a review

Treatment of nitrogenous wastewater by a constructed wetland-coupled microbial fuel cell system: a review

An in-situ view cell system for investigating swelling behavior of elastomers upon high-pressure hydrogen exposure

The transition to hydrogen as a clean and efficient energy carrier is impeded by challenges in the compatibility of hydrogen with materials used within hydrogen infrastructure. Elastomers, crucial in sealing components, often exhibit premature failures in high-pressure hydrogen environments due to excessive swelling. This study employs an innovative in-situ view cell system to assess the swelling behavior of hydrogenated nitrile butadiene rubber (HNBR) under various hydrogen conditions. The system, designed to withstand pressures up to 96.5 MPa, incorporates Digital Image Correlation (DIC) for strain measurements and volume estimation. Results reveal non-linear volume increases during depressurization, challenging conventional assumptions. Furthermore, investigations into peak hydrogen pressures and pressure-holding scenarios during decompression highlight complex swelling trends. The introduction of a novel computer vision (CV) method enhances precision in volume estimation, overcoming DIC limitations. The study provides insights into mitigating elastomer swelling, crucial for developing robust materials to support future hydrogen-driven energy systems.

Techno-economic and environmental analysis for the application of renewable energy sources in seaports.

Ports have an indisputable effect on the decarbonization of urban areas, helping to minimize air and environmental pollution and achieve sustainable development. In this instance, it is crucial to do research that can advance our understanding of how to increase ports' energy independence by utilizing renewable energy sources. The current study aims to study the environmental benefits and techno-economic challenges of converting three Egyptian ports to eco-friendly green ports by using solar panels, offshore wind turbines, and hydrogen fuel cells. The study shows that from a technical point of view, the required green power to be installed at Alexandria, Port Said, and Suez ports is around 13 MW, 5 MW, and 1.5 MW, respectively. Furthermore, the environmental analysis findings demonstrate that integrating green energy will significantly lower emissions in seaports. It is anticipated that the ports of Alexandria, Port Said, and Suez will achieve annual reductions in carbon dioxide emissions of roughly 68,7 k-tons, 25,8 k-tons, and 6,4 k-tons, respectively. From an economic point of view, the ports could be supplied with green energy from wind turbines for a cost of between 0.115 and 0.125 USD/kWh, while solar panels have a cost range of 0.098 to 0.129 USD/kWh. Additionally, hydrogen fuel cell systems cost about 0.102 USD/kWh.

Chondrocyte-targeted exosome-mediated delivery of Nrf2 alleviates cartilaginous endplate degeneration by modulating mitochondrial fission

BackgroundCartilaginous endplate (CEP) degeneration, which is an important contributor to intervertebral disc degeneration (IVDD), is characterized by chondrocyte death. Accumulating evidence has revealed that dynamin-related protein 1 (Drp1)-mediated mitochondrial fission and dysfunction lead to apoptosis during CEP degeneration and IVDD. Exosomes are promising agents for the treatment of many diseases, including osteoporosis, osteosarcoma, osteoarthritis and IVDD. Despite their major success in drug delivery, the full potential of exosomes remains untapped.Materials and methodsIn vitro and in vivo models of CEP degeneration were established by using lipopolysaccharide (LPS). We designed genetically engineered exosomes (CAP-Nrf2-Exos) expressing chondrocyte-affinity peptide (CAP) on the surface and carrying the antioxidant transcription factor nuclear factor E2-related factor 2 (Nrf2). The affinity between CAP-Nrf2-Exos and CEP was evaluated by in vitro internalization assays and in vivo imaging assays. qRT‒PCR, Western blotting and immunofluorescence assays were performed to examine the expression level of Nrf2 and the subcellular localization of Nrf2 and Drp1. Mitochondrial function was measured by the JC-1 probe and MitoSOX Red. Mitochondrial morphology was visualized by MitoTracker staining and transmission electron microscopy (TEM). After subendplate injection of the engineered exosomes, the degree of CEP degeneration and IVDD was validated radiologically and histologically.ResultsWe found that the cargo delivery efficiency of exosomes after cargo packaging was increased by surface modification. CAP-Nrf2-Exos facilitated chondrocyte-targeted delivery of Nrf2 and activated the endogenous antioxidant defence system in CEP cells. The engineered exosomes inhibited Drp1 S616 phosphorylation and mitochondrial translocation, thereby preventing mitochondrial fragmentation and dysfunction. LPS-induced CEP cell apoptosis was alleviated by CAP-Nrf2-Exo treatment. In a rat model of CEP degeneration, the engineered exosomes successfully attenuated CEP degeneration and IVDD and exhibited better repair capacity than natural exosomes.ConclusionCollectively, our findings showed that exosome-mediated chondrocyte-targeted delivery of Nrf2 was an effective strategy for treating CEP degeneration.Graphic abstract CAP-Nrf2-Exos delivered Nrf2 into CEP cells and alleviated LPS-induced apoptosis by inhibiting Drp1-mediated mitochondrial fission

A new hiPSC-based viral expression system to study virus host-cell interaction in hiPSC-derived cardiomyocytes

Abstract Coxsackievirus B3 (CVB3) belongs to the Picornaviridae family and is a member of the enterovirus genus. CVB3 is the most frequently detected pathogen in viral myocarditis, which leads to acute heart infections in 20-40% of cases and to dilated cardiomyopathy (DCM) in cases of prolonged viral persistence. Complex arrhythmias and sudden deaths due to CVB infections have also been reported, particularly in newborns and children. Observed arrhythmias include bradycardia, AV block and junctional ectopic tachycardia, suggesting that in addition to the functioning myocardium, the pacemaker cells and cardiac conduction system may also be impaired by CVB3 infection. The symptomatic phenotype is highly variable and depends on unclear host factors. It ranges from inconspicuous inflammation and rapid healing to acute congestive heart failure or viral persistence with progressive chronic disease. Despite the increasing and profound knowledge about viral myocarditis, it is still a challenge to identify further host factors that contribute to acute and chronic myocarditis, as science lacks sensitive and controllable disease models. The presented project shows the generation of a controlled, inducible virus-expressing hiPSC line that provides a solution for the beforementioned problems. The new human induced pluripotent stem cell line (hiPSC) i can be used to carry out virologic experiments in a controlled manner and without risk to the user. For this purpose, the modified and non-infectious genome of the enterovirus Coxsackievirus B3 (CVB3) is stably integrated into the genome of a wild-type hiPSC line. The CVB3 genome to be stably integrated was modified to prevent the formation of functional capsids by a DVP0 mutation. In addition, the 3'UTR and 5'UTR regions of the CVB3 genome were removed, which prevents uncontrolled self-replication of viral RNA. As a result, work with this CVB3DVP0-UTR genome is classified as S1 compliant. The expression of the CVB3DP0-UTR virus genome can be controlled by adding doxycycline to the cell culture medium by a TeT-On System. The transfected cells are then cultivated as clones and tested for virus expression. The dose-dependent CVB3DVP0 expression is verified using qPCR and FACS after 1 day of doxycycline application in the cell culture medium. Clones isolated and positively tested for CVB3 expression are used to carry out initial experiments on the interaction of the expressed viral proteins with the cell. Particular focus is placed on the induction of cell death, remodelling of the cytoskeleton and increase of reactive oxygen species (ROS) in mitochondria after CVB3DVP0-UTR induction in order to validate the cell system for application. Differentiation of the CVB3-expressing hiPSC-line into cardiomyocytes with subsequent virus induction results in the elevation of mitochondrial ROS, degradation of the cytoskeleton and arrhythmic behaviour of the cells.

#1949 Comparison of two commercial available primary human mesangial cell clones reveal differences

Abstract Background and Aims Mesangial cells and their matrix form the central stalk of the glomerulus but is often overlooked when studying glomerular function. The cells are however highly involved in glomerular function and crosstalk between the different cell types of the glomerulus. They are important in onset and progression of several glomerular diseases, such as IgA nephropathy and diabetic kidney disease. As mesangial cells have several functions and are involved in glomerular disease they are often used for in vitro work studying response to various stimuli and downstream signaling. This includes changes in the production and regulation or their matrix, proliferation, contractility, protein expression and release of different inflammatory mediators such as cytokines and growth factors [1]. Using primary cells when doing in vitro work is usually considered the best option, but there might be differences between origin of cells in their response and physiology. For this study, our aim was to compare two types of commercially available primary human mesangial cells from Cell systems (CS) and Novabiosis (NBS). Method For characterization of the CS and NBS mesangial cells, gene expression of commonly used mesangial cell markers, including KRT19, ACTA2, PDGFRA and PDGFRB, was analyzed with real time qPCR. Staining of smooth muscle actin was done in order to compare the phenotype of CS and NBS. To compare the cells, differences in protein expression were determined using proteomics. CS and NBS Cells were cultured in the same way and were also treated with PDGF-BB for 24 h to compare with untreated control. For further comparison, proliferation assay was used to compare proliferation after treating cells with PDGF-BB for 24 h. Results Both sources of cells expressed all markers, but there was a difference in level of gene expression. CS cells showed higher expression than NBS cells for all markers, apart from ACTA2. NBS cells had a more stellate phenotype whereas the CS cells were more elongated. There was a difference in proteins expressed by CS and NBS mesangial cells from our proteomics. The level of upregulation of proteins seen with proteomics was stronger for CS than NBS cells when treated with PDGF-BB. The number of proteins upregulated was also higher for CS than NBS cells. Both mesangial cell types had increased proliferation when treated with PDGF-BB, however CS cells were seen to grow faster in cell culture and seemed to be more responsive to the treatment with PDGF-BB. Conclusion In conclusion, this study compares two commercially available primary human mesangial cells and show that there is a difference between the two. The cell types react differently when treated with PDGF-BB and has some difference in what proteins they express. This needs to be considered when conducting cell culture experiments with mesangial cells, depending on what the aim and focus for the study is.

Ebselen improves fungal keratitis through exerting anti-inflammation, anti-oxidative stress, and antifungal effects

Fungal keratitis is a severely vision-threatening corneal infection, where the prognosis depends on both fungal virulence and host immune defense. Inappropriate host responses can induce substantial inflammatory damage to the cornea. Therefore, in the treatment of fungal keratitis, it is important to concurrently regulate the immune response while efforts are made to eliminate the pathogen. Ebselen is a widely studied organo-selenium compound and has been demonstrated to have antifungal, antibacterial, anti-inflammatory, and oxidative stress-regulatory properties. The effectiveness of ebselen for the treatment of fungal keratitis remains unknown. In this study, ebselen was demonstrated to produce a marked inhibitory effect on Aspergillus fumigatus (A. fumigatus), including spore germination inhibition, mycelial growth reduction, and fungal biofilm disruption. The antifungal activity of ebselen was related to the cell membrane damage caused by thioredoxin (Trx) system inhibition-mediated oxidative stress. On the contrary, ebselen enhanced the antioxidation of Trx system in mammalian cells. Further, ebselen was proven to suppress the expressions of inflammatory mediators (IL-1β, IL-6, TNF-α, COX-2, iNOS, and CCL2) and reduce the production of oxidative stress-associated indicators (ROS, NO, and MDA) in fungi-stimulated RAW264.7 cells. In addition, ebselen regulated PI3K/Akt/Nrf2 and p38 MAPK signaling pathways, which contributed to the improvement of inflammation and oxidative stress. Finally, we verified the therapeutic effect of ebselen on mouse fungal keratitis. Ebselen improved the prognosis and reduced the fungal burden in mouse corneas. Expressions of inflammatory mediators, as well as the infiltration of macrophages and neutrophils in the cornea were also obviously decreased by ebselen. In summary, ebselen exerted therapeutic effects by reducing fungal load and protecting host tissues in fungal keratitis, making it a promising treatment for fungal infections.

The role of elasticity on adhesion and clustering of neurons on soft surfaces

The question of whether material stiffness enhances cell adhesion and clustering is still open to debate. Results from the literature are seemingly contradictory, with some reports illustrating that adhesion increases with surface stiffness and others suggesting that the performance of a system of cells is curbed by high values of elasticity. To address the role of elasticity as a regulator in neuronal cell adhesion and clustering, we investigated the topological characteristics of networks of neurons on polydimethylsiloxane (PDMS) surfaces - with values of elasticity (E) varying in the 0.55–2.65 MPa range. Results illustrate that, as elasticity increases, the number of neurons adhering on the surface decreases. Notably, the small-world coefficient – a topological measure of networks – also decreases. Numerical simulations and functional multi-calcium imaging experiments further indicated that the activity of neuronal cells on soft surfaces improves for decreasing E. Experimental findings are supported by a mathematical model, that explains adhesion and clustering of cells on soft materials as a function of few parameters - including the Young’s modulus and roughness of the material. Overall, results indicate that – in the considered elasticity interval – increasing the compliance of a material improves adhesion, improves clustering, and enhances communication of neurons.

Impact of the Transboundary Interference Inhibitor on RNAi and the Baculovirus Expression System in Insect Cells

Impact of the Transboundary Interference Inhibitor on RNAi and the Baculovirus Expression System in Insect Cells

Catalyst Interaction in Unitized Regenerative Fuel Cells

Abstract Unitized regenerative fuel cells have emerged as promising energy conversion and storage systems for various applications. However, in order to optimize their efficiency, it is crucial to enhance the performance of the bifunctional catalyst. This study aims to provide deeper insights into the electrochemical behavior and performance of the bifunctional catalyst. Several electrocatalysts were prepared and evaluated using rotating disc electrode measurements. The primary focus was placed on investigating the interaction between Pt, Ir, and the support material, antimony doped tin oxide (ATO), and their impact on the oxygen evolution reaction and oxygen reduction reaction. Among the analyzed catalysts, Pt black mixed with synthesized IrO2 supported on developed ATO exhibited the highest performance, considering the results from both the fuel cell and electrolyzer systems.

Chicken intestinal organoids: a novel method to measure the mode of action of feed additives

There is a rapidly growing interest in how the avian intestine is affected by dietary components and feed additives. The paucity of physiologically relevant models has limited research in this field of poultry gut health and led to an over-reliance on the use of live birds for experiments. The development of complex 3D intestinal organoids or “mini-guts” has created ample opportunities for poultry research in this field. A major advantage of the floating chicken intestinal organoids is the combination of a complex cell system with an easily accessible apical-out orientation grown in a simple culture medium without an extracellular matrix. The objective was to investigate the impact of a commercial proprietary blend of organic acids and essential oils (OA+EO) on the innate immune responses and kinome of chicken intestinal organoids in a Salmonella challenge model. To mimic the in vivo prolonged exposure of the intestine to the product, the intestinal organoids were treated for 2 days with 0.5 or 0.25 mg/mL OA+EO and either uninfected or infected with Salmonella and bacterial load in the organoids was quantified at 3 hours post infection. The bacteria were also treated with OA+EO for 1 day prior to challenge of the organoids to mimic intestinal exposure. The treatment of the organoids with OA+EO resulted in a significant decrease in the bacterial load compared to untreated infected organoids. The expression of 88 innate immune genes was investigated using a high throughput qPCR array, measuring the expression of 88 innate immune genes. Salmonella invasion of the untreated intestinal organoids resulted in a significant increase in the expression of inflammatory cytokine and chemokines as well as genes involved in intracellular signaling. In contrast, when the organoids were treated with OA+EO and challenged with Salmonella, the inflammatory responses were significantly downregulated. The kinome array data suggested decreased phosphorylation elicited by the OA+EO with Salmonella in agreement with the gene expression data sets. This study demonstrates that the in vitro chicken intestinal organoids are a new tool to measure the effect of the feed additives in a bacterial challenge model by measuring innate immune and protein kinases responses.

Research and Development of Control Strategies for Fuel Cell Systems

For the self-developed kilowatt level proton exchange membrane fuel cell system, the control algorithm of the fuel cell system is built in MATLAB/Simulink, and the control models of each subsystem, the entire system control process, and steady-state calibration process are discussed. Data analysis through experiments shows that the system has good performance.

Modeling and Control of Ejector-Based Hydrogen Circulation System for Proton Exchange Membrane Fuel Cell Systems

Modeling and Control of Ejector-Based Hydrogen Circulation System for Proton Exchange Membrane Fuel Cell Systems