Cell Aggregation Research Articles

Adsorption and uptake of functionalized nanoplastics (NPs) by wetland plant (Sphagnum): A unique pathway for polystyrene-NPs reduction in non-vascular plants

Wetlands are sources and sinks for nanoplastics (NPs), where adsorption and uptake by plants constitute a crucial pathway for NPs accumulation. This study found that Sphagnum exhibited a high potential (~89.75 %) to intercept NPs despite the lack of root systems and stomata. Two pathways for 100nm polystyrene NPs accumulation in Sphagnum were located: (i) Spiral interception and foliar adsorption. Efficient adsorption is credited to the micro/nano-interlocked leaf structure, which is porous, hydrophilic and rough. (ii) Intracellular enrichment through pores. Fluorescence tracking indicates pseudo-leaves (lateral > cephalic branches) as primary organs for internalization. Accumulation of differently functionalized NPs was characterized: PS-Naked-NPs (PS), PS-COOH-NPs (PC) and PS-NH2-NPs (PN) were all largely retained by pathway (i), while pathway (ii) mainly uptake PN and PC. Unlike PS aggregation in transparent cells, PC enrichment in chloroplast cells and PN in intercellular spaces reduced pigment content and fluorescence intensity. Further, the effects of the accumulated NPs on the ecological functions of Sphagnum were evaluated. NPs reduce carbon flux (assimilation rate by 57.78 %, and respiration rate by 33.50%), significantly decreasing biomass (PS = 13.12 %, PC = 26.48 %, PN = 35.23 %). However, toxicity threshold was around 10 μg/mL, environmental levels (≤1 μg/mL) barely affected Sphagnum. This study advances understanding of the behavior and fate of NPs in non-vascular plants, and provides new perspectives for developing Sphagnum substrates for NPs interception.

On-demand bactericidal and self-adaptive antifouling hydrogels for self-healing and lubricant coatings of catheters

Catheter-related infections are one of the most common nosocomial infections with increasing morbidity and mortality, and robust antibacterial or antifouling catheter coatings remain great challenges for long-term implantation. Herein, multifunctional hydrogel coatings were developed to provide persistent and self-adaptive antifouling and antibacterial effects with self-healing and lubricant capabilities. Polyvinyl alcohol (PVA) with β-cyclodextrin (β-CD) grafts (PVA-Cd) and 4-arm polyethylene glycol (PEG) with adamantane and quaternary ammonium compound (QAC) terminals (QA-PEG-Ad) were crosslinked through host-guest recognitions between adamantane and β-CD moieties to acquire PVEQ coatings. In response to bacterial infections, QACs exhibit reversible transformation between zwitterions (pH 7.4) and cationic lactones (pH 5.5) to generate on-demand bactericidal effect. Highly hydrophilic PEG/PVA backbones and zwitterionic QACs build a lubricate surface and decrease the friction coefficient 10 times compared with that of bare catheters. The antifouling hydrated layer significantly inhibits blood protein adsorption and platelet activation and reveals negligible hemolysis and cytotoxicity. The dynamic host-guest crosslinking achieves full self-healing of cracks in PVEQ hydrogels, and the mechanical profiles were recovered to over 90 % after rejuvenating the broken hydrogels, exhibiting a long-term stability after mechanical stretching, twisting, knotting and compression. After subcutaneous implantation and local bacterial infection, the retrieved PVEQ-coated catheters display no tissue adhesion and 3 log folds lower bacterial number than that of bare catheters. PVEQ coatings effectively prevent the repeated bacterial infections and there are few inflammatory reactions in the surrounding tissue, while substantial lymphoid infiltration and inflammatory cell aggregation occur in muscle tissues around the bare catheter. Thus, this study demonstrates a catheter coating strategy by on-demand bactericidal, self-adaptive antifouling, self-healing and lubricant hydrogels to address medical devices-related infections. Statement of significanceIt is estimated over two billion peripheral intravenous catheters are annually used in hospitals around the world, and catheter-associated infection has become a great clinical challenge with rapidly rising morbidity and mortality. Surface coating is considered a promising approach, but substantial challenges remain in the development of coatings that simultaneously satisfy both anti-fouling and antibacterial attributes. Even more, few attempts have been made to design mechanically robust coatings and reversible antibacterial or antifouling capabilities, which are critical for long-term medical implants. To address these challenges, we propose a concise strategy to develop hydrogel coatings from commercially available poly(ethylene glycol) and polyvinyl alcohol. In addition to self-healing and lubricant capabilities, the reversible conversion between zwitterionic and cationic lactones of quaternary ammonium compounds enables on-demand bactericidal and self-adaptive antifouling effects.

All-aqueous synthesis of alginate complexed with fibrillated protein microcapsules for membrane-bounded culture of tumor spheroids

Water-in-water (W/W) emulsions provide bio-compatible all-aqueous compartments for artificial patterning and assembly of living cells. Successful entrapment of cells within a W/W emulsion via the formation of semipermeable capsules is a prerequisite for regulating on the size, shape, and architecture of cell aggregates. However, the high permeability and instability of the W/W interface, restricting the assembly of stable capsules, pose a fundamental challenge for cell entrapment. The current study addresses this problem by synthesizing multi-armed protein fibrils and controlling their assembly at the W/W interface. The multi-armed protein fibrils, also known as ‘fibril clusters’, were prepared by cross-linking lysozyme fibrils with multi-arm polyethylene glycol (PEG) via click chemistry. Compared to linear-structured fibrils, fibril clusters are strongly adsorbed at the W/W interface, forming an interconnected meshwork that better stabilizes the W/W emulsion. Moreover, when fibril clusters are complexed with alginate, the hybrid microcapsules demonstrate excellent mechanical robustness, semi-permeability, cytocompatibility and biodegradability. These advantages enable the encapsulation, entrapment and long-term culture of tumor spheroids, with great promise for applications for anti-cancer drug screening, tumor disease modeling, and tissue repair engineering.

Novel use of a - 20°C cryoprotectant yields high viability and improved aggregation of marine sponge cells.

Novel use of a - 20°C cryoprotectant yields high viability and improved aggregation of marine sponge cells.

Particle-based modeling and GPU-accelerated simulation of cellular blood flow

Computational modeling and simulation of cellular blood flow is highly desired for understanding blood microcirculation and blood-related diseases such as thrombosis and tumor, but it remains a challenging task primarily because blood in microvessels should be described as a dense suspension of different types of deformable cells. The focus of the present work is on the development of a particle-based and GPU-accelerated numerical method that is able to quickly simulate the various behaviors of deformable cells in three-dimensional arbitrarily complex geometries. We employ a two-fluid model to describe blood flow, incorporating the deformation and aggregation of cells. A smoothed dissipative particle dynamics is used to solve the two-fluid model, and a discrete microstructure model is applied for the cell deformation, as well as a Morse potential model for the cell aggregation. The heterogeneous CPU-GPU environment is established, where each GPU thread is dedicated to a particle, and the CPU is mainly responsible for loading and exporting data. Five test cases are conducted against analytical theory, experimental data, and previous numerical results, for pure fluid, cell deformation, cell aggregation, cell suspension and the cellular flow in a complex network, respectively. It is shown that the methodology can accurately predict various behaviors of cells, and the GPU is well suited for particle-based modeling. Especially for cellular blood flow, where calculating cellular forces is a compute-intensive and time-consuming task, the GPU offers exceptional parallel capabilities, significantly enhancing the simulation efficiency. The speedup is about 3.5 times faster than the CPU parallelization with 96 cores for the pure fluid, and this acceleration nearly reaches 20 times when cells are included in the simulations. Particularly, the calculations for deformation and aggregation forces demonstrate a substantial speedup, achieving the improvements of up to 120 and 640 times, respectively, compared to their serial counterparts. The present methodology can effectively integrate various behaviors of cells, and has the potential in simulating very large microvascular networks at organ levels.

High-throughput fabrication of cell spheroids with 3D acoustic assembly devices.

Acoustic cell assembly devices are applied in cell spheroid fabrication attributed to their rapid, label-free and low-cell damage production of size-uniform spheroids. However, the spheroids yield and production efficiency are still insufficient to meet the requirements of several biomedical applications, especially those that require large quantities of cell spheroids, such as high-throughput screening, macro-scale tissue fabrication, and tissue repair. Here, we developed a novel 3D acoustic cell assembly device combined with a gelatin methacrylamide (GelMA) hydrogels for the high-throughput fabrication of cell spheroids. The acoustic device employs three orthogonal piezoelectric transducers that can generate three orthogonal standing bulk acoustic waves to create a 3D dot-array (25 × 25 × 22) of levitated acoustic nodes, enabling large-scale fabrication of cell aggregates (>13,000 per operation). The GelMA hydrogel serves as a supporting scaffold to preserve the structure of cell aggregates after the withdrawal of acoustic fields. As a result, mostly cell aggregates (>90%) mature into spheroids maintaining good cell viability. We further applied these acoustically assembled spheroids to drug testing to explore their potency in drug response. In conclusion, this 3D acoustic cell assembly device may pave the way for the scale-up fabrication of cell spheroids or even organoids, to enable flexible application in various biomedical applications, such as high-throughput screening, disease modeling, tissue engineering, and regenerative medicine.

Exploring the role of E. faecalis enterococcal polysaccharide antigen (EPA) and lipoproteins in evasion of phagocytosis.

Enterococcus faecalis is an opportunistic pathogen frequently causing nosocomial infections. The virulence of this organism is underpinned by its capacity to evade phagocytosis, allowing dissemination in the host. Immune evasion requires a surface polysaccharide produced by all enterococci, known as the enterococcal polysaccharide antigen (EPA). EPA consists of a cell wall-anchored rhamnose backbone substituted by strain-specific polysaccharides called 'decorations', essential for the biological activity of this polymer. However, the structural determinants required for innate immune evasion remain unknown, partly due to a lack of suitable validated assays. Here, we describe a quantitative, invitro assay to investigate how EPA decorations alter phagocytosis. Using the E. faecalis model strain OG1RF, we demonstrate that a mutant with a deletion of the locus encoding EPA decorations can be used as a platform strain to express heterologous decorations, thereby providing an experimental system to investigate the inhibition of phagocytosis by strain-specific decorations. We show that the aggregation of cells lacking decorations is increasing phagocytosis and that this process does not involve the recognition of lipoproteins by macrophages. Collectively, our work provides novel insights into innate immune evasion by enterococci and paves the way for further studies to explore the structure/function relationship of EPA decorations.

Ultrafiltration of Angelicae Sinensis Radix and Astragali Radix inhibits ferroptosis and improves pulmonary fibrosis in rats by regulating Nrf2/xCT/GPX4 signaling pathway

This study aims to investigate the mechanism of ferroptosis mediated by the nuclear factor-E2-related factor 2(Nrf2)/solute carrier family 7 member 11(SLC7A11, also known as xCT)/glutathione peroxidase 4(GPX4) signaling pathway in radiationinduced pulmonary fibrosis and the intervention effect of Angelicae Sinensis Radix(ASR) and Astragali Radix(AR) ultrafiltration extract. Fifty Wistar rats were randomly divided into five groups, with 10 rats in each group. Except for the blank group without radiation, the rats in each group were anesthetized and subjected to a single local chest irradiation of 40 Gy X-rays once to establish a rat model of radiation-induced pulmonary fibrosis. After radiation, the rats in the intervention groups were orally administered with ASR-AR ultrafiltration extract at doses of 0. 12, 0. 24, and 0. 48 g·kg~(-1), respectively, once a day for 30 days. After 30 days of continuous administration, the levels of oxidative stress indicators superoxide dismutase(SOD) activity, reduced glutathione(GSH),malondialdehyde(MDA), and ferrous ion(Fe~(2+)) in lung tissues of each group were detected by colorimetry. Immunofluorescence was used to detect reactive oxygen species(ROS) fluorescence expression in lung tissues. Hematoxylin-eosin(HE) and Masson staining were performed to observe pathological changes in lung tissues. Immunohistochemistry and Western blot were used to detect the expression levels of Nrf2/xCT/GPX4 signaling pathway and fibrotic proteins in lung tissues. The results showed that compared with the results in the blank group, the levels of Fe~(2+) and MDA in the model group increased, while SOD activity and GSH levels decreased,and ROS levels increased. HE and Masson staining results showed that the structure of lung tissue was seriously damaged, the pulmonary interstitium was significantly proliferated, the alveoli collapsed and consolidated severely, and there were more inflammatory cell aggregates and collagen fiber deposits. Transmission electron microscopy showed that the degree of lung tissue damage in the model group was relatively high, with increased, smaller, and disorganized damaged mitochondria, irregular morphology, shallow matrix,most mitochondria ruptured and shortened, mildly expanded, some mitochondria with increased electron density of the matrix, partial mitochondrial outer membrane rupture, and characteristic changes of ferroptosis-specific mitochondria. Immunohistochemistry showed that the expression of transferrin receptor protein 1(TFR1) in lung tissues was significantly increased, while the expression of GPX4,ferritin heavy chain 1(FTH1), Nrf2, and xCT was significantly decreased. Western blot showed that the expression of α-smooth muscle actin(α-SMA) and collagen Ⅰ protein increased. Compared with the model group, the intervention group with ASR-AR ultrafiltration extract significantly improved lipid peroxidation and antioxidant-related indicators, decreased Fe~(2+) levels, alleviated fibrosis, and decreased the expression of TFR1, α-SMA, and collagen Ⅰ proteins in lung tissues, while increased the expression of GPX4, FTH1, Nrf2, and xCT proteins. In summary, ASR-AR ultrafiltration extract has an ameliorative effect on radiation-induced pulmonary fibrosis, and its mechanism may involve the inhibition of ferroptosis by regulating the Nrf2/xCT/GPX4 signaling pathway.

Evidence that the StlA polyketide synthase is required for the transition of growth to development in Polysphondylium violaceum.

The social amoeba Polysphondylium violaceum uses chemoattractants different from those of Dictyoctelium discoideum for cell aggregation. However, the detailed mechanisms in P. violaceum remain unknown. We have previously reported that the polyketide synthase StlA is involved in inducing aggregation in this species. To elucidate the mechanism of StlA-induced aggregation in P. violaceum, we analyzed the phenotype of P. violaceum stlA- (Pv-stlA-) mutants in more detail. Unlike our previous results, the mutant cells did not exhibit proper chemotaxis toward glorin. Defective aggregation was not restored by glorin pulses, 8Br-cAMP, or deletion of the homologue of PufA that is a translational repressor of protein kinase A, whereas mutant cells grown in the presence of 4-methyl-5-pentylbenzene-1,3-diol (MPBD), the putative Pv-StlA product, aggregated normally without it after starvation. Furthermore, the early developmental marker gene, dscA, was downregulated in the mutant cells. Our data thus suggested that StlA is required for the transition from growth to development in P. violaceum.

Robust bioprocess design and evaluation of commercial media for the serial expansion of human induced pluripotent stem cell aggregate cultures in vertical-wheel bioreactors

BackgroundWhile pluripotent stem cell (PSC) therapies move toward clinical and commercial applications at a rapid rate, manufacturing reproducibility and robustness are notable bottlenecks in regulatory approval. Therapeutic applications of PSCs require large cell quantities to be generated under highly robust, well-defined, and economically viable conditions. Small-scale and short-term process optimization, however, is often performed in a linear fashion that does not account for time needed to verify the bioprocess protocols and analysis methods used. Design of a reproducible and robust bioprocess should be dynamic and include a continuous effort to understand how the process will respond over time and to different stresses before transitioning into large-scale production where stresses will be amplified.MethodsThis study utilizes a baseline protocol, developed for the short-term culture of PSC aggregates in Vertical-Wheel® bioreactors, to evaluate key process attributes through long-term (serial passage) suspension culture. This was done to access overall process robustness when performed with various commercially available media and cell lines. Process output variables including growth kinetics, aggregate morphology, harvest efficiency, genomic stability, and functional pluripotency were assessed through short and long-term culture.ResultsThe robust nature of the expansion protocol was demonstrated over a six-day culture period where spherical aggregate formation and expansion were observed with high-fold expansions for all five commercial media tested. Profound differences in cell growth and quality were revealed only through long-term serial expansion and in-vessel dissociation operations. Some commercial media formulations tested demonstrated maintenance of cell growth rates, aggregate morphology, and high harvest recovery efficiencies through three bioreactor serial passages using multiple PSC lines. Exceptional bioprocess robustness was even demonstrated with sustained growth and quality maintenance over 10 serial bioreactor passages. However, some commercial media tested proved less equipped for serial passage cultures in bioreactors as cultures led to cell lysis during dissociation, reduction in growth rates, and a loss of aggregate morphology.ConclusionsThis study demonstrates the importance of systematic selection and testing of bioprocess input variables, with multiple bioprocess output variables through serial passages to create a truly reproducible and robust protocol for clinical and commercial PSC production using scalable bioreactor systems.

Dissociation of red blood cell aggregates in extensional flow

Dissociation of red blood cell aggregates in extensional flow

The Buerger’s rabbit model: a closer step to unravelling thromboangiitis obliterans?

ObjectiveThromboangiitis obliterans (TAO) remains clinical challenging due to its rarity and underwhelming management outcomes. This study aimed to describe a novel TAO rabbit model that demonstrates a closer resemblance to TAO.MethodsThirty-six New Zealand rabbits underwent the surgical implantation of calibrated gelatin sponge particles (CGSPs) into their right femoral artery. The CGSPs were soaked in different solutions to simulate different types of thrombi: normal (NT; normal saline); inflammatory TAO thrombus (TAO; dimethylsulfoxide [DMSO]), and DMSO with methotrexate (MTX). All groups underwent clinical assessment, digital subtraction angiography (DSA) and histopathological analysis at time points day 0 (immediate), week 1 (acute), week 2 (subacute), and week 4 (chronic).ResultsThe TAO rabbit presented with signs of ischemia of the right digit at week 4. On DSA, the TAO rabbits exhibited formation of corkscrew collaterals starting week 1. On H&E staining, gradual CGSP degradation was observed along with increased red blood cell aggregation and inflammatory cells migration in week 1. On week 2, disorganization of the tunica media layer and vascular smooth muscle cell (VSMC) proliferation was observed. In the TAO rabbit, migrated VSMCs, inflammatory cells, and extracellular matrix with collagen-like substances gradually occluded the lumen. On week 4, the arterial lumen of the TAO rabbit was filled with relatively-organized VSMC and endothelial cell clusters with less inflammatory cells. Neorevascularization was found in the MTX-treated group.ConclusionThe novel TAO rabbit model shows a closer resemblance to human TAO clinically, radiographically, and histopathologically. Histological analysis of the IT progression in the TAO model suggests that it is of VSMC origin.

Human amniotic epithelial cells improve uterine spiral artery remodeling to ameliorate preeclampsia in a rat model†.

Preeclampsia (PE) is a multisystem pregnancy disorder characterized by impaired remodeling of placental spiral arteries, which leads to the release of pro-inflammatory cytokines and anti-angiogenic agents. However, treatment options for PE are limited, with termination of pregnancy being the only curative option. In this work, we investigated the effects of human amniotic epithelial cells (hAECs) in PE rat model. The rats were induced with lipopolysaccharide (LPS) on gestational day 14.5 followed by injection of hAECs and human umbilical cord mesenchymal stem cells 24h later. The hAECs treatment resulted in a reduction in blood pressure and proteinuria in the PE rat model. Furthermore, hAECs treatment decreased levels of pro-inflammatory cytokines, reduced inflammatory cells aggregation, and alleviated the damage to placental spiral arteries by downregulating the expression of anti-angiogenic factor and upregulating proangiogenic factor. In vitro experiments confirmed that hAECs treatment restored the proliferation, migration, and angiogenesis of LPS-damaged human umbilical vein endothelial cells. Additionally, hAECs treatment had positive effects on fetal weight and neurological development in the PE group, with no negative effects on the physical development or fertility of offspring rats. These results suggested that hAECs transplantation may be a novel adjuvant therapeutic strategy for PE by reducing the inflammatory and enhancing placental spiral artery angiogenesis.

Intelligent and biocompatible cellulose aerogels featured with high-elastic and fast-hemostatic for epistaxis and wound healing

Nasal tamponade is a commonly employed and highly effective treatment method for preventing nasal bleeding. However, the current nasal packing hemostatic materials exhibit some limitations, such as low hemostatic efficiency, the potential for causing secondary injury when removed from the nasal cavity, limited intelligence in their design, and an inability to promote the healing of nasal mucosa wounds. Herein, we report the fabrication of a smart cellulose aerogel through the covalent cross-linking of carboxymethyl cellulose (CMC) macromolecules, while incorporating one-dimensional cellulose nanofibers (CNF) and two-dimensional MXene as reinforcing network scaffolds and conductive fillers. The abundant hydrogen and ether bonds in aerogels make them possess high elasticity in both dry and wet states, which can be compressed 100 times at 90 % deformation with a stress loss of <10 % under water. The highly elastic aerogels can be filled into the narrow nasal passages, pressuring the capillaries and reducing the amount of bleeding. Moreover, the strong interface between aerogels and blood can promote red blood cell aggregation, platelet adhesion and activation, activate intrinsic coagulation pathway and accelerate blood coagulation, resulting in excellent hemostatic ability. Furthermore, the aerogels exhibit excellent hemocompatibility and cytocompatibility, making them suitable for wound healing and capable of fully healing wounds within 15 days. Notably, the presence of MXene causes the aerogels to form a conductive network when exposed to blood, enabling them to perform real-time hemostatic monitoring without removing the dressing. This innovative biomedical aerogel, prepared from natural materials, shows excellent potential for applications in rapid nasal hemostasis.

Antifungal and antibiofilm activities of flavonoids against Candida albicans: Focus on 3,2′-dihydroxyflavone as a potential therapeutic agent

Effective management of microbial biofilms holds significance within food and medical environments. Candida albicans, an opportunistic fungus, forms mucosal biofilms closely linked to candidiasis and drug-resistant infections due to their drug tolerance. Morphologic change from yeast to filamentous cells is a key virulence factor and a prerequisite for biofilm development. This study investigated the anti-fungal and antibiofilm activities of 20 flavonoids against C. albicans. With their known antioxidant capabilities, flavonoids hold promise in combating infections associated with biofilms. Among them, flavone and its derivatives exhibited moderate antifungal activity, 3,2′-dihydroxyflavone (3,2′-DHF) at 1 μg/mL exhibited strong antibiofilm activity (MIC 50 μg/mL). In addition, 3,2′-DHF dramatically inhibited cell aggregation and germ tube/hyphae formation. Transcriptomic analyses revealed that flavone and 3,2′-DHF behaved differently, as 3,2′-DHF downregulated the expressions of germ tube/hyphae-forming and biofilm-related genes (ECE1, HWP1, TEC1, and UME6) but upregulated the biofilm/hyphal regulators (CHK1, IFD6, UCF1, and YWP1). Tests evaluating toxicity with plant and nematode models revealed that flavone and 3,2′-DHF exhibited mild toxicity. Current results indicate that hydroxylated flavone derivatives can enhance anti-fungal and antibiofilm activities and provide a source of potential anti-fungal agents against drug-resistant C. albicans.

Paper-Based Microfluidic Analytical Device Patterned by Label Printer for Point-of-Care Blood Glucose and Hematocrit Detection Using 3D-Printed Smartphone Cassette.

This study presents a portable, low-cost, point-of-care (POC) system for the simultaneous detection of blood glucose and hematocrit. The system consists of a disposable origami microfluidic paper-based analytical device (μPAD) for plasma separation, filtration, and reaction functions and a 3D-printed cassette for hematocrit and blood glucose detection using a smartphone. The origami μPAD is patterned using a cost-effective label printing technique instead of the conventional wax printing method. The 3D-printed cassette incorporates an array of LED lights, which mitigates the effects of intensity variations in the ambient light and hence improves the accuracy of the blood glucose and hematocrit concentration measurements. The hematocrit concentration is determined quantitatively by measuring the distance of plasma wicking along the upper layer of the origami μPAD, which is pretreated with sodium chloride and Tween 20 to induce dehydration and aggregation of the red blood cells. The filtered plasma also penetrates to the lower layer of the origami μPAD, where it reacts with embedded colorimetric assay reagents to produce a yellowish-brown complex. A color image of the reaction complex is captured using a smartphone inserted into the 3D-printed cassette. The image is analyzed using self-written RGB software to quantify the blood glucose concentration. The calibration results indicate that the proposed detection platform provides an accurate assessment of the blood glucose level over the range of 45-630 mg/dL (R2 = 0.9958). The practical feasibility of the proposed platform is demonstrated by measuring the blood glucose and hematocrit concentrations in 13 human whole blood samples. Taking the measurements obtained from commercial glucose and hematocrit meters as a benchmark, the proposed system has a differential of no more than 6.4% for blood glucose detection and 9.1% for hematocrit detection. Overall, the results confirm that the proposed μPAD is a promising solution for cost-effective and reliable POC health monitoring.

Structural Mapping of Protein Aggregates in Live Cells Modeling Huntington's Disease

AbstractWhile protein aggregation is a hallmark of many neurodegenerative diseases, acquiring structural information on protein aggregates inside live cells remains challenging. Traditional microscopy does not provide structural information on protein systems. Routinely used fluorescent protein tags, such as Green Fluorescent Protein (GFP), might perturb native structures. Here, we report a counter‐propagating mid‐infrared photothermal imaging approach enabling mapping of secondary structure of protein aggregates in live cells modeling Huntington's disease. By comparing mid‐infrared photothermal spectra of label‐free and GFP‐tagged huntingtin inclusions, we demonstrate that GFP fusions indeed perturb the secondary structure of aggregates. By implementing spectra with small spatial step for dissecting spectral features within sub‐micrometer distances, we reveal that huntingtin inclusions partition into a β‐sheet‐rich core and a ɑ‐helix‐rich shell. We further demonstrate that this structural partition exists only in cells with the [RNQ+] prion state, while [rnq−] cells only carry smaller β‐rich non‐toxic aggregates. Collectively, our methodology has the potential to unveil detailed structural information on protein assemblies in live cells, enabling high‐throughput structural screenings of macromolecular assemblies.

Structural Mapping of Protein Aggregates in Live Cells Modeling Huntington's Disease.

While protein aggregation is a hallmark of many neurodegenerative diseases, acquiring structural information on protein aggregates inside live cells remains challenging. Traditional microscopy does not provide structural information on protein systems. Routinely used fluorescent protein tags, such as Green Fluorescent Protein (GFP), might perturb native structures. Here, we report a counter-propagating mid-infrared photothermal imaging approach enabling mapping of secondary structure of protein aggregates in live cells modeling Huntington's disease. By comparing mid-infrared photothermal spectra of label-free and GFP-tagged huntingtin inclusions, we demonstrate that GFP fusions indeed perturb the secondary structure of aggregates. By implementing spectra with small spatial step for dissecting spectral features within sub-micrometer distances, we reveal that huntingtin inclusions partition into a β-sheet-rich core and a ɑ-helix-rich shell. We further demonstrate that this structural partition exists only in cells with the [RNQ+] prion state, while [rnq-] cells only carry smaller β-rich non-toxic aggregates. Collectively, our methodology has the potential to unveil detailed structural information on protein assemblies in live cells, enabling high-throughput structural screenings of macromolecular assemblies.

Ceftriaxone and MC-100093 mitigate fentanyl-induced cardiac injury in mice: Preclinical investigation of its underlying molecular mechanisms

Drug addiction is considered a worldwide concern and one of the most prevailing causes of death globally. Opioids are highly addictive drugs, and one of the most common opioids that is frequently used clinically is fentanyl. The potential harmful effects of chronic exposure to opioids on the heart are still to be elucidated. Although β-lactam antibiotics are well recognized for their ability to fight bacteria, its protective effect in the brain and liver has been reported. In this study, we hypothesize that β-lactam antibiotic, ceftriaxone, and the novel synthetic non-antibiotic β-lactam, MC-100093, are cardioprotective against fentanyl induced-cardiac injury by upregulating xCT expression. Mice were exposed to repeated low dose (0.05 mg/kg, i.p.) of fentanyl for one week and then challenged on day 9 with higher dose of fentanyl (1 mg/kg, i.p.). This study investigated cardiac histopathology and target genes and proteins in serum and cardiac tissues in mice exposed to fentanyl overdose and β-lactams. We revealed that fentanyl treatment induced cardiac damage as evidenced by elevated cardiac enzymes (troponin I). Furthermore, fentanyl treatment caused large aggregations of inflammatory cells and elevation in the areas and volumes of myocardial fibers, indicating hypertrophy and severe cardiac damage. Ceftriaxone and MC-100093 treatment, However, induced cardioprotective effects as evidenced by marked reduction in cardiac enzymes (troponin I) and changes in histopathology. Furthermore, ceftriaxone and MC-100093 treatment decreased the levels of hypertrophic genes (α-MHC & β-MHC), apoptotic (caspase-3), and inflammatory markers (IL-6 & NF-κB). This study reports for the first time the cardioprotective effect of β-lactams against fentanyl-induced cardiac injury. Further studies are greatly encouraged to completely identify the cardioprotective properties of ceftriaxone and MC-100093.

Cyanobacterial Bloom Formation by Enhanced Ecological Adaptability and Competitive Advantage of Microcystis-Non-Negligible Role of Quorum Sensing.

Microcystis-dominated cyanobacterial blooms (MCBs) frequently occur in freshwaters worldwide due to massive Microcystis colony formation and severely threaten human and ecosystem health. Quorum sensing (QS) is a direct cause of Microcystis colony formation that drives MCBs outbreak by regulating Microcystis population characteristics and behaviors. Many novel findings regarding the fundamental knowledge of the Microcystis QS phenomenon and the signaling molecules have been documented. However, little effort has been devoted to comprehensively summarizing and discussing the research progress and exploration directions of QS signaling molecules-mediated QS system in Microcystis. This review summarizes the action process of N-acyl homoserine lactones (AHLs) as major signaling molecules in Microcystis and discusses the detailed roles of AHL-mediated QS system in cellular morphology, physiological adaptability, and cell aggregation for colony formation to strengthen ecological adaptability and competitive advantage of Microcystis. The research progress on QS mechanisms in Microcystis are also summarized. Compared to other QS systems, the LuxI/LuxR-type QS system is more likely to be found in Microcystis. Also, we introduce quorum quenching (QQ), a QS-blocking process in Microcystis, to emphasize its potential as QS inhibitors in MCBs control. Finally, in response to the research deficiencies and gaps in Microcystis QS, we propose several future research directions in this field. This review deepens the understanding on Microcystis QS knowledge and provide theoretical guidance in developing strategies to monitor, control, and harness MCBs.