Extent Of Aggregation Research Articles

Role of Peptide Associations in Enhancing the Antimicrobial Activity of Adepantins: Comparative Molecular Dynamics Simulations and Design Assessments

Adepantins are peptides designed to optimize antimicrobial biological activity through the choice of specific amino acid residues, resulting in helical and amphipathic structures. This paper focuses on revealing the atomistic details of the mechanism of action of Adepantins and aligning design concepts with peptide behavior through simulation results. Notably, Adepantin-1a exhibits a broad spectrum of activity against both Gram-positive and Gram-negative bacteria, while Adepantin-1 has a narrow spectrum of activity against Gram-negative bacteria. The simulation results showed that one of the main differences is the extent of aggregation. Both peptides exhibit a strong tendency to cluster due to the amphipathicity embedded during design process. However, the more potent Adepantin-1a forms smaller aggregates than Adepantin-1, confirming the idea that the optimal aggregations, not the strongest aggregations, favor activity. Additionally, we show that incorporation of the cell penetration region affects the mechanisms of action of Adepantin-1a and promotes stronger binding to anionic and neutral membranes.

Neuropathology in an α-synuclein preformed fibril mouse model occurs independent of the Parkinson's disease-linked lysosomal ATP13A2 protein

Loss-of-function mutations in the ATP13A2 (PARK9) gene are implicated in early-onset autosomal recessive Parkinson's disease (PD) and other neurodegenerative disorders. ATP13A2 encodes a lysosomal transmembrane P5B-type ATPase that is highly expressed in brain and specifically within the substantia nigra pars compacta (SNc). Recent studies have revealed its normal role as a lysosomal polyamine transporter, although its contribution to PD-related pathology remains unclear. Cellular studies report that ATP13A2 can regulate α-synuclein (α-syn) secretion via exosomes. However, the relationship between ATP13A2 and α-syn in animal models remains inconclusive. ATP13A2 knockout (KO) mice exhibit lysosomal abnormalities and reactive astrogliosis but do not develop PD-related neuropathology. Studies manipulating α-syn levels in mice lacking ATP13A2 indicate minimal effects on pathology. The delivery of α-syn preformed fibrils (PFFs) into the mouse striatum is a well-defined model to study the development and spread of α-syn pathology. In this study we unilaterally injected wild-type (WT) and homozygous ATP13A2 KO mice with mouse α-syn PFFs in the striatum and evaluated mice for neuropathology after 6 months. The distribution, spread and extent of α-syn aggregation in multiple regions of the mouse brain was largely independent of ATP13A2 expression. The loss of nigrostriatal pathway dopaminergic neurons and their nerve terminals induced by PFFs were equivalent in WT and ATP13A2 KO mice. Reactive astrogliosis was induced equivalently by α-syn PFFs in WT and KO mice but was already significantly higher in ATP13A2 KO mice due to pre-existing reactive gliosis. We did not identify asymmetric motor disturbances, microglial activation, or axonal damage induced by α-syn PFFs in WT or KO mice. Although α-syn PFFs induce an increase in lysosomal number in the SNc in general, TH-positive dopaminergic neurons did not exhibit either increased lysosomal area or intensity, regardless of genotype. Our study evaluating the spread of α-syn pathology reveals no exacerbation of α-syn pathology, neuronal loss, astrogliosis or motor deficits in ATP13A2 KO mice, suggesting that selective lysosomal abnormalities resulting from ATP13A2 loss do not play a major role in α-syn clearance or propagation in vivo.

Rheological and Biochemical Properties of Blood in Runners: A Preliminary Report

Purpose: Physical activity induces numerous modifications in the morphological, rheological, and biochemical properties of blood. The purpose of this study was to evaluate changes in blood rheological and biochemical indicators among runners. Also, we assessed how the rheological and biochemical properties of blood in people who practised running characterised the range and direction of exercise modifications and allowed for the diagnosis of transient adaptive effects. Methods: This study included 12 athletes who regularly trained in middle- and long-distance running (6–8 times a week) and presented a high sports level (national and international class). The athletes performed a 30 min warm-up consisting of 15 min of jogging and exercises. After a 10 min rest, they completed a 3 km run with submaximal effort. Blood samples were collected at baseline and after the effort. Results: No statistically significant changes were revealed in erythrocyte, leukocyte, platelet, iron, ferritin, transferrin, erythropoietin, or C-reactive protein concentrations in the examined runners. The same applied to the elongation index at a shear stress within the range of 0.30–60.00 Pa, amplitude and total extent of aggregation, aggregation half-life, and aggregation index. A significant increase (within standard limits) was only observed in fibrinogen concentration after running. Conclusions: The lack of post-exercise changes in blood rheological and biochemical indicators in the investigated runners points at an efficient haemorheological system. This, in turn, reflects well-executed training and remarkably well-trained adaptive systems responsible for regeneration.

Physicochemical property characterization, amino acid profiling and sensory evaluation of plant-based ice cream incorporated with soy, pea and milk proteins

This study examined the effects of incorporating milk protein concentrate (MPC), pea or soy proteins isolates (PPI and SPI) on the physicochemical, sensorial properties, and amino acid composition of ice creams containing 7% protein, in comparison to dairy ice cream as a reference. As protein ingredients, PPI exhibited higher water and oil holding capacity but lower surface hydrophobicity than SPI and MPC. Viscosity of the mixes were proportional to the firmness of ice cream, and both were highest with use of PPI. MPC ice cream had most similar physical and sensory properties to reference. PPI and SPI ice cream mixes showed higher extent of fat coalescence than MPC and reference. PPI and SPI conferred structural stability to ice cream with lower melting rate and better shape retention, and ability to delay ice recrystallization during temperature flocculation as compared with SMP and MPC. Confocal laser scanning microscope images indicated that higher extent of protein aggregation and more air cells were found in PPI ice cream. Sensory and amino acid profile results revealed that PPI and SPI ice creams were inferior in taste, texture, and essential amino acids like methionine. This study offers insights for the development of high protein frozen desserts.

Facile colorimetric sensor development for sequential detection of coralyne and calf-thymus DNA via tannic acid-assisted silver nanoparticles

Owning to unique size dependent optical properties, plasmonic nanoparticles have been explored as chemical read-outs for assaying various biological and chemical analytes. We propose colorimetric sensing strategy based on tannic acid mediated silver nanoparticles (TA-AgNPs) for the detection of Coralyne (COR) and calf thymus DNA (CT-DNA). Aggregation of TA-AgNPs is observed on addition of COR molecule because of electrostatic interaction between them, which is established by change in color from yellow to brown. TA-AgNPs/COR based colorimetric probe was then employed for CT-DNA detection, utilizing anti-aggregation of nanoparticles. Formation of intercalated complex between CT-DNA and COR prevents the aggregation of nanoparticles and color of TA-AgNPs solution remains yellow. Under optimum sensing conditions, extent of aggregation and anti-aggregation of TA-AgNPs was studied. A linear correlation between absorbance value at 437 nm and COR concentration is obtained in the range of 0 to 1.2 μM with detection limit of 0.44 μM. TA-AgNPs/COR based detection probe exhibited linear response to CT-DNA in concentration range of 0 to 7.62 μM with a limit of detection down to 1.63 μM. The applicability of sensing strategy verified by analysis of COR and CT-DNA in human urine samples, which demonstrated the practical utility of method in biomedical applications.

Cerium oxide nanoparticles formation and aggregation dynamics

The conventional synthesis of cerium oxide nanoparticles (CeNPs) relies on the use of trivalent cerium and an oxidizer to ensure a thorough hydrolysis. This study explores CeNPs formation via the hydrolysis of tetravalent cerium, specifically through the spontaneous reaction of ceric ammonium nitrate (Ce(NH4)2(NO3)6) under varied aging conditions, including adjustments in pH, concentration, and heating strategy. This research adopts a thermo-hydrolysis method to synthesize CeNPs with small primary particles (2–3 nm) to loose dendritic clusters (5–8 nm). Additionally, when the starting pH of the solution decreases below 0.8, it favors the formation of clusters with larger diameters (15–20 nm). Analyses using dynamic light scattering (DLS), high-resolution transmission electron microscopy (HRTEM), and X-ray photoelectron spectroscopy (XPS) indicate that CeNPs particle size in acidic environments is predominantly determined by the extent of primary particle aggregation, influenced by nitrate adsorption. Overall, our comprehensive analysis of the CeNPs evolution provides fundamental insights for the development of a size/structure controllable synthetic strategy for nanoparticle manufacturing.

Porphyrin Aggregation under Homogeneous Conditions Inhibits Electrocatalysis: A Case Study on CO2 Reduction.

Metalloporphyrins are widely used as homogeneous electrocatalysts for transformations relevant to clean energy and sustainable organic synthesis. Metalloporphyrins are well-known to aggregate due to π-π stacking, but surprisingly, the influence of aggregation on homogeneous electrocatalytic performance has not been investigated previously. Herein, we present three structurally related iron meso-phenylporphyrins whose aggregation properties are different in commonly used N,N-dimethylformamide (DMF) electrolyte. Both spectroscopy and light scattering provide evidence of extensive porphyrin aggregation under conventional electrocatalytic conditions. Using the electrocatalytic reduction of CO2 to CO as a test reaction, cyclic voltammetry reveals an inverse dependence of the kinetics on the catalyst concentration. The inhibition extends to bulk performance, where up to 75% of the catalyst at 1 mM is inactive compared to at 0.25 mM. We additionally report how aggregation is perturbed by organic additives, axial ligands, and redox state. Periodic boundary calculations provide additional insights into aggregate stability as a function of metalloporphyrin structure. Finally, we generalize the aggregation phenomenon by surveying metalloporphyrins with different metals and substituents. This study demonstrates that homogeneous metalloporphyrins can aggregate severely in well-solubilizing organic electrolytes, that aggregation can be easily modulated through experimental conditions, and that the extent of aggregation must be considered for accurate catalytic benchmarking.

Online monitoring of protein refolding in inclusion body processing using intrinsic fluorescence

Inclusion bodies (IBs) are protein aggregates formed as a result of overexpression of recombinant protein in E. coli. The formation of IBs is a valuable strategy of recombinant protein production despite the need for additional processing steps, i.e., isolation, solubilization and refolding. Industrial process development of protein refolding is a labor-intensive task based largely on empirical approaches rather than knowledge-driven strategies. A prerequisite for knowledge-driven process development is a reliable monitoring strategy. This work explores the potential of intrinsic tryptophan and tyrosine fluorescence for real-time and in situ monitoring of protein refolding. In contrast to commonly established process analytical technology (PAT), this technique showed high sensitivity with reproducible measurements for protein concentrations down to 0.01 g L-1\\documentclass[12pt]{minimal} \\usepackage{amsmath} \\usepackage{wasysym} \\usepackage{amsfonts} \\usepackage{amssymb} \\usepackage{amsbsy} \\usepackage{mathrsfs} \\usepackage{upgreek} \\setlength{\\oddsidemargin}{-69pt} \\begin{document}$$^{-1}$$\\end{document}. The change of protein conformation during refolding is reflected as a shift in the position of the maxima of the tryptophan and tyrosine fluorescence spectra as well as change in the signal intensity. The shift in the peak position, expressed as average emission wavelength of a spectrum, was correlated to the amount of folding intermediates whereas the intensity integral correlates to the extent of aggregation. These correlations were implemented as an observation function into a mechanistic model. The versatility and transferability of the technique were demonstrated on the refolding of three different proteins with varying structural complexity. The technique was also successfully applied to detect the effect of additives and process mode on the refolding process efficiency. Thus, the methodology presented poses a generic and reliable PAT tool enabling real-time process monitoring of protein refolding.Graphical abstractReal-time intrinsic fluorescence monitoring in protein refolding

Urban heat island analysis based on high resolution measurement data: A case study in Beijing

Urban heat island (UHI) is a widely concerned urban climate which impacts billions of residents worldwide. Nevertheless, inadequate rigorous air temperature (Tair) measurement has become an obstacle to UHI research and mitigation, especially in megacities like Beijing. In this study, we applied a 1-km hourly Tair dataset interpolated from records of 521 stations in Beijing. The dataset enables us to analyze UHI patterns on a block scale and the impacts of landscapes combined with the local climate zones (LCZ) scheme. Our results indicate that: (1) Tair and its temporal variation in each LCZ vary significantly, which are highly relevant to their morphology and thermal-radiative properties. (2) LCZ 1 and LCZ 2 have the highest nocturnal UHI intensities (UHII). Discrepancies among LCZs also cause various temporal variations in their UHII. (3) The spatial pattern of LCZs affects regional temperature. Area ratio and aggregation extent of LCZ 1&2 show significant positive correlations with regional temperature, while LCZ A&D&G have the opposite regularity. (4) Distinct mitigation strategies are put forward for each subarea in Beijing with various endowments of population and the LCZ structure. The study provides insight into Beijing UHI and plausible mitigation advice in megacities like Beijing.

Brazilin-7-2-butenoate inhibits amyloid β-protein aggregation, alleviates cytotoxicity, and protects Caenorhabditis elegans

The fibrillogenesis of amyloid β-protein (Aβ) gradually accumulates to form neurotoxic Aβ aggregates in the human brain, which is the direct cause of Alzheimer's disease (AD) related symptoms. There are currently no effective therapies for AD. Brazilin, a natural polyphenol, inhibits Aβ fibrillogenesis, disrupts the mature fibrils and alleviates the corresponding cytotoxicity, but it also has the high toxic. Therefore, brazilin-7-2-butenoate (B-7-2-B), a brazilin derivative, was designed and synthesized. B-7-2-B exhibited lower toxicity and stronger inhibitory effect on Aβ aggregation than brazilin. B-7-2-B could prevent the formation of Aβ fibrils and oligomers, and depolymerize pre-formed aggregates in a dose-dependent manner. Furthermore, B-7-2-B prominently alleviated the cytotoxicity and the oxidative stress induced by Aβ aggregates in PC12 cells. The protective impacts of B-7-2-B were further demonstrated by using the Caenorhabditis elegans model, including decreasing the extent of Aβ aggregation, improving the motility and sensation disorders. Eventually, B-7-2-B was proven to be no apparent damage to worms. In summarize, it can be concluded that B-7-2-B has the potential as a drug for treating AD.

Effect of a typical anticonvulsant drug carbamazepine on the aggregation of zinc oxide nanoparticles and an insight into their co-transport in porous media

The emergence of new contaminants like engineered nanoparticles (NPs), pharmaceuticals (PhACs), etc., causes a detrimental effect on the ecosystem and the study needs to be done to understand its behavior in the natural environment. In the present study aggregation and transport behavior of zinc oxide nanoparticles (nZnO) was evaluated in the presence of one of the PhACs, carbamazepine (CBZ). At first, a series of batch experiments were performed with 50 mg/L of nZnO and 1 mg/L of CBZ at different pH (6 and 8) and electrolyte concentration (EC: 1 mmol/L and 20 mmol/L), to study the change in the hydrodynamic diameter of nZnO. The result indicates that the extent of aggregation of nZnO increases with an increase in EC from 1 mmol/L to 20 mmol/L of NaCl or CaCl2. However, in the presence of CBZ, shows greater colloidal stability at lower EC, and its effect was found to be nominal at higher EC. The transport behavior of nZnO and CBZ was assessed by column experiment, it was found that more than 97 % of nZnO and 99 % of CBZ are retained in the porous media during the injection for different test conditions. Moreover, during transport experiments of longer duration, less than 10 % of nZnO and about 72 µg/L of CBZ were released in the effluent. Therefore, the risk of the release of nZnO is reduced in the presence of CBZ.

Rheological properties of blood in multiple myeloma patients.

Multiple myeloma (MM) is considered to be one of the hematological malignancies formed by excessive and abnormal proliferation of plasmocytes. Among other parameters, several blood tests are used to diagnose multiple myeloma. The hemorheological profile in multiple myeloma is not widely studied. Hemorheology includes the study of measuring the deformability and aggregation of erythrocytes, blood viscosity, and sedimentation rate. The degree of deformability of blood cells is necessary to maintain proper vital functions. Proper deformability of red blood cells ensures proper blood circulation, tissue oxidation and carbon dioxide uptake. The aim of the study was to compare morphology and blood rheology parameters in patients with MM and healthy individuals. The study included 33 patients with MM, and 33 healthy subjects of the same age. The hematological blood parameters were evaluated using ABX MICROS 60 hematology analyzer. The LORCA Analyzer to study erythrocyte aggregation and deformability. Patients with MM had lower red blood cells count (RBC) (9.11%) (p < 0.001) and half time of total aggregation (T1/2) (94.29%) (p < 0.001) values and higher mean corpuscular volume (MCV) (5.50%) (p < 0.001), aggregation index (AI) (68.60%) (p < 0.001), total extent of aggregation (AMP) (87.92%) (p < 0.001) values than the healthy control group. Aggregation in patients with MM is different compared to healthy individuals. It was observed that the percentage of cell aggregation is almost 50% higher than in the control group. The study of morphology, aggregation and deformability of erythrocytes in patients with suspected MM may be helpful in making clinical decisions.

Holistic Prediction of AuNP Aggregation in Diverse Aqueous Suspensions Based on Machine Vision and Dark-Field Scattering Imaging.

The localized surface-plasmon resonance of the AuNP in aqueous media is extremely sensitive to environmental changes. By measuring the signal of plasmon scattering light, the dark-field microscopic (DFM) imaging technique has been used to monitor the aggregation of AuNPs, which has attracted great attention because of its simplicity, low cost, high sensitivity, and universal applicability. However, it is still challenging to interpret DFM images of AuNP aggregation due to the heterogeneous characteristics of the isolated and discontinuous color distribution. Herein, we introduce machine vision algorithms for the training of DFM images of AuNPs in different saline aqueous media. A visual deep learning framework based on AlexNet is constructed for studying the aggregation patterns of AuNPs in aqueous suspensions, which allows for rapid and accurate identification of the aggregation extent of AuNPs, with a prediction accuracy higher than 0.96. With the aid of machine learning analysis, we further demonstrate the prediction ability of various aggregation phenomena induced by both cation species and the concentration of the external saline solution. Our results suggest the great potential of machine vision frameworks in the accurate recognition of subtle pattern changes in DFM images, which can help researchers build predictive analytics based on DFM imaging data.

Adaptable strategy for reactivation and recycling of spent S-Zorb adsorbents at the laboratory and pilot scale

The spent S-Zorb adsorbents containing Ni and Zn elements are hazardous wastes. It would generate significant economic and environmental benefits to reactivate and recycle these solid wastes through a reactivation strategy. Furthermore, adaptability investigation of this strategy is also indispensable before its industrial application. Herein, the spent S-Zorb adsorbents (Spent-TJ/MM/QD) from different plants were reactivated at laboratory and pilot scale in 3 m3 reactor via an acid-base coupling reactivation strategy. The spent adsorbents exhibit distinct phase compositions and microstructures of active components. Formation of ZnSi2O4 and ZnS is the primary reason for abandonment of the Spent-TJ (Spent-MM) and Spent-QD, respectively. The nickel species also exhibit different aggregation extent. Fortunately, the inert zinc and nickel species are respectively converted into ZnO and NiO during the reactivation process. Higher surface area (1.7–4.0 times that of the spent adsorbents) and more acid sites are generated over the reactivated adsorbents. Besides, all the reactivated adsorbents possess similar phase compositions and microstructures. Both the adsorbents reactivated at pilot and laboratory scale exhibit comparable desulfurization activity to fresh ones. The sulfur content of the gasoline desulfurized by the reactivated adsorbents is below 10 μg g−1, meeting the Euro V legislations. All the results indicate the excellent adaptability and commercial potential of the reactivation strategy. The possible mechanism for the excellent adaptability of the reactivation method was proposed.

An experimental study of stress-dependent water retention behaviour of two lateritic clays with different minerals

Lateritic soils are associated with a high content of sesquioxides, which enhance the aggregation of fine particles. The formation of aggregates can induce significant changes in the pore size distribution (PSD) and consequently affect the hydro-mechanical behaviour of lateritic soils. The influence of mineral types on the extent of aggregation and the stress-dependent water retention behaviour of lateritic clays is not well understood. In this study, the stress-dependent water retention curves (SDSWRCs) of a compacted lateritic sandy fat clay with halloysite (i.e., GLH) obtained from Ghana were measured at different net vertical stresses using an improved stress-controlled pressure plate apparatus. The results were then compared with those of another lateritic sandy lean clay with goethite (NLG) obtained from Nigeria. Additionally, scanning electron microscopy (SEM) was employed to examine the microstructure associated with these soils. The experimental results show that GLH has a higher ability to retain water than NLG at any given suction, even though the former has a higher void ratio than the latter prior to the drying-wetting cycle. This is because of the presence of relatively small aggregates and small-size macropores in GLH, as revealed by SEM images. On the other hand, the water retention capacity of both soils reduces with elevated stress levels, except at net vertical stress of 120 kPa for GLH, with a corresponding decrease in the rate of desorption. Furthermore, the overall increase in the total degree of hysteresis measured is almost 2 times higher for NLG (i.e., 116%) than GLH (i.e., 72%) in the stress and suction range considered, contrary to the decreasing trends commonly reported in the literature. It was found that the sizes of aggregates in GLH reduced (‘finer soil’) as stress level increased and caused more water to be retained. Conversely to GLH, the aggregate size remained unchanged for NLG, as revealed by aggregate size analysis. This is mainly because of the differences in the strengths provided by the unique minerals in the two soils (i.e., halloysite for GLH and goethite for NLG) to the aggregates, due to the different parent rocks from which the soils were formed.

Thin-Film Electrodes Based on Conjugated Polyelectrolytes for High-Performance Pseudocapacitance in Aqueous Media

Conjugated polyelectrolytes (CPEs) are an emerging class of materials that exhibits competitive pseudocapacitive performances in their hydrogel form due to their mixed electronic/ionic conductive nature. Interest in developing solid-state electrodes based on CPEs for practical applications is needed. However, the water-soluble nature of the CPEs hinders their thin-film stability in aqueous media. Herein, we report that the slow-dried drop-casted CPE-K thin films formed a solid-state electrode insoluble in aqueous electrolyte solutions when the salt concentration is higher than 0.1 M. XRD measurements revealed that slow-dried, drop-casted CPE-K film possesses a more ordered macromolecular structure when compared to its pre-deposited powder form. DLS measurements revealed that the extent of CPE-K aggregation increases as a function of salt concentration, which enhances the stability of the films in the electrolyte solutions. In addition, AFM imaging suggests well-ordered and uniform nanostructures. As a result, it is possible to use CPE-K thin-film as a pseudocapacitive electrodes capable to maintain 85% (71 F g-1 ) of its original capacitance (84 F g-1) at a specific current of 500 A g-1. In addition, CPE-K film electrodes exhibit good cycling stability where a capacitance retention of 93% after 100,000 cycles at a current density of 35 A g-1 was obtained. To our knowledge, such high charge/discharge rates and stabilities are greater than those reported for state-of-the-art CP in any form. Further electrochemical characterization revealed that surface-controlled pseudocapacitive behavior dominates the charge-storing mechanism of the CPE-K electrodes. In addition, our electrochemical analysis suggests that facile ionic diffusion attributed to the presence of an ionic lattice in the ordered structure of the CPE-K films is responsible for the unprecedented charging and cycling performances. These findings provide a new strategy to obtain solid-state electrodes based on CPEs without the need for binders or additives. Additional advantages such as water processability, low cost, and bio-compatibility may allow CPE-K to be a sustainable solution for the future of electrochemical energy storage.

Establishment of a high-content imaging assay for tau aggregation in hiPSC-derived neurons differentiated from two protocols to routinely evaluate compounds and genetic perturbations

Aberrant protein aggregation is a pathological cellular hallmark of many neurodegenerative diseases, such as Alzheimer's disease (AD) and frontotemporal dementia (FTD), where the tau protein is aggregating, forming neurofibrillary tangles (NFTs), and propagating from neuron to neuron. These processes have been linked to disease progression and a decline in cognitive function. Various therapeutic approaches aim at the prevention or reduction of tau aggregates in neurons. Human induced pluripotent stem cells (hiPSCs) are a very valuable tool in neuroscience discovery, as they offer access to potentially unlimited amounts of cell types that are affected in disease, including cortical neurons, for in vitro studies. We have generated an in vitro model for tau aggregation that uses hiPSC – derived neurons expressing an aggregation prone, fluorescently tagged version of the human tau protein after lentiviral transduction. Upon addition of tau seeds in the form of recombinant sonicated paired helical filaments (sPHFs), the neurons show robust, disease-like aggregation of the tau protein. The model was developed as a plate-based high content screening assay coupled with an image analysis algorithm to evaluate the impact of small molecules or genetic perturbations on tau. We show that the assay can be used to evaluate small molecules or screen targeted compound libraries. Using siRNA-based gene knockdown, genes of interest can be evaluated, and we could show that a targeted gene library can be screened, by screening nearly 100 deubiquitinating enzymes (DUBs) in that assay. The assay uses an imaging-based readout, a relatively short timeline, quantifies the extent of tau aggregation, and also allows the assessment of cell viability. Furthermore, it can be easily adapted to different hiPSC lines or neuronal subtypes. Taken together, this complex and highly relevant approach can be routinely applied on a weekly basis in the screening funnels of several projects and generates data with a turnaround time of approximately five weeks.

Aggregation of human plasma and of human blood induced in vitro by filgrastim originator product; effect of PEGylation

We herein report that filgrastim product Neupogen® and the filgrastim formulation buffer induced aggregate formation when mixed in vitro with human plasma, and formation of large membranous erythrocyte aggregates when mixed with human blood, similar to the aggregation induced by pegfilgrastim and by pegfilgrastim buffer [T. Arvinte, E. Poirier, N. Ersayin, G. Darpin, A. Cudd, J. Dowd, S. Brokx, Aggregation of human plasma and of human blood induced in vitro by pegfilgrastim originator formulation buffer and pegfilgrastim products, Eur. J. Pharmaceut. Biopharmaceut. (2023), doi: 10.1016/j.ejpb.2023.10.019]. The data identify the filgrastim buffer (which is practically the same in filgrastim and pegfilgrastim products) as the main driver of human plasma and blood aggregation. Kinetic experiments showed differences in the extent of plasma aggregation induced by a filgrastim product manufactured in EU and one manufactured in USA. Human donor variability in the plasma aggregation induced by filgrastim was observed. To study the effect of PEGylation of the filgrastim protein on plasma aggregation we compared filgrastim (Neupogen®) with pegfilgrastim (Neulasta®) solutions at the same protein concentration. These data show that PEGylation has a beneficial effect in inhibiting to an extent plasma aggregation. Interestingly, 20 kDa polyethylene glycol in the filgrastim buffer induced more plasma aggregation compared to the buffer, similar to the aggregation induced by pegfilgrastim. For intravenous infusion filgrastim solutions (300 µg/ml, vials only) may be diluted in 5 % dextrose from a concentration of 300 µg/ml to 5 µg/ml. Aggregation of human plasma was also induced by filgrastim solutions diluted in 5 % dextrose to 50 µg/ml, 15 µg/ml and 5 µg/ml filgrastim, as well as by the filgrastim buffer similarly diluted in 5 % dextrose (1:6, 1:20 and 1:60 dilution). These data show that filgrastim solutions used for intravenous administration in patients induce human plasma aggregation in vitro. Such aggregation phenomena may be related to known infusion side effects of filgrastim therapy.

Impact of secondary ice in a frozen NaCl freeze-concentrated solution on the extent of methylene blue aggregation

Freezing and lyophilization have been utilized for decades to stabilize pharmaceutical and food products. Freezing a solution that contains dissolved salt and/or organic matter produces pure primary ice crystal grains separated by freeze-concentrated solutions (FCS). The microscopic size of the primary ice crystals depends on the cooling conditions and the concentration of the solutes. It is generally accepted that primary ice crystals size influences the rate of sublimation and also can impact physico-chemical behaviour of the species in the FCS. This article, however, presents a case where the secondary ice formed inside the FCS plays a critical role. We microscoped the structures of ice-cast FCS with an environmental scanning electron microscope and applied the aggregation-sensitive spectroscopic probe methylene blue to determine how the microstructure affects the molecular arrangement. We show that slow cooling at −50 °C produces large salt crystals with a small specific surface, resulting in a high degree of molecular aggregation within the FCS. In contrast, fast liquid nitrogen cooling yields an ultrafine structure of salt crystals having a large specific surface area and, therefore, inducing smaller aggregation. The study highlights a critical role of secondary ice in solute aggregation and introduces methylene blue as a molecular probe to investigate freezing behaviour of aqueous systems with crystalline solute.

Abstract 16734: Patterns of Inflammatory Activation in Cardiovascular Disease: Insights From the Randomized Colchicine PCI Trial

INTRODUCTION: Recent studies demonstrate that neutrophils accumulate in atherosclerotic lesions and that neutrophil counts correlate with disease severity. The randomized double-blind Colchicine-PCI study demonstrated that colchicine attenuated systemic inflammation (IL-6, CRP) after percutaneous coronary intervention compared with placebo. GOALS/AIMS: We characterized neutrophil activity in patients across a spectrum of cardiovascular phenotypes, including severity of coronary artery disease (CAD) and presentation of acute myocardial infarct (AMI). METHODS: The Colchicine-PCI biomarker sub-study enrolled 478 participants with pretreatment assessment of inflammatory biomarkers. White blood cell count and differential were performed using a hematology analyzer. Soluble markers of neutrophil activity, including neutrophil extracellular traps (NETs), and systemic inflammation were assessed using multiplex assays. Neutrophil surface markers of activity and neutrophil platelet aggregates were assessed using an Accuri C6 flow cytometer. RESULTS: We observed neutrophil concentration, adhesion, transmigration, and activation, as well as increased NETs complexes; but no difference in extent of neutrophil platelet aggregation or systemic inflammation in patients with versus without obstructive CAD. Conversely, in patients with versus without AMI, neutrophil markers of adhesion and transmigration did not differ, but systemic markers of inflammation and neutrophil activation were elevated. CONCLUSION: This single-site, prospective study is the largest neutrophil biorepository in patients with cardiovascular risk factors. Inflammatory activation, and particularly neutrophil profiles, differ in patients with obstructive CAD and in the setting of AMI.